ASBMR 29th Annual Meeting

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Therapeutic Effects of Risedronate for Two Years, on Bone Metabolic Markers, Especially on TRACP-5b. H. Naka*¹, H. Masaki*¹, Y. Imanishi*², T. Miki¹, Y. Nishizawa². ¹Geriatric Medicine, Osaka City University Graduate School of Medicine, Osaka, Japan. ²Department of Metabolism, Endocrinology and Molecular Medicine, Osaka City University Graduate School of Medicine, Osaka, Japan.

The prevention of treatment cessation for osteoporosis patients is important for providing protection against fractures. Assessing bone resorption in the early stage of treatment by evaluating various bone metabolic markers over time in risedronate therapy is considered useful in preventing cessation of treatment. Therefore, we conducted prospective study on the extent to which the therapeutic effect on the patient can be assessed by evaluating bone metabolic markers over time, including the new bone resorption marker TRACP-5b.

Risedronate was administered to 36 postmenopausal osteoporosis patients (mean age 66.3 years) for 2 year and the therapeutic effects on bone density and bone metabolic markers were evaluated. As a control, 8 people received Ca and were evaluated in the same manner. When bone metabolic markers (BAP, uNTX, uCTX, sNTX, TRACP-5b) were measured before treatment and in Months 1, 3 and 6, some of the bone resorption markers were significantly inhibited from Month 1 and by Month 6 mean inhibition was 15–60%. Bone formation markers were suppressed 25–35% in Months 3–6, and lumbar BMD had increased 3.7% in 6 months, 4.2% in 12 months and 4.5% in 24 months. The proportion of patients who showed an inhibition rate (S/N ratio) against the least significant change of 1 or more in Month 1 was 83.3% for TRACP-5b, 52.8% for sNTX, 66.7% for uNTX and 44.4% for uCTX. In Month 3 they were 81.1%, 70.3%, 75.7% and 51.4%, respectively. In the Ca administration controls, no long-term significant changes in TRACP-5b were observed, but fluctuations in the collagen degradation products NTX and CTX were observed. Significant correlation (p < 0.05) between the rate of TRACP-5b inhibition and the rate of increase in lumbar bone density in Month 12 was observed at −0.386 in Month 1 and −0.513 in Month 3. In contrast, correlation between the rate of change in BMD and the rate of uCTX inhibition was −0.335 and -.0425, respectively. There was no significant correlation between the rate of increase in BMD and uNTX, sNTX or DPD. Correlation with the rate of BAP inhibition in Months 3 and 6 was −0.492 and −0.633. Furthermore, about 28 patients who could be measured BMD in 24 months, Significant correlation (p < 0.05) between the rate of TRACP-5b inhibition and the rate of increase in BMD in Month 24 was observed at −0.419 in Month 6, but other bone resorption marker were not. Correlation with the rate of BAP inhibition in Months 6 was −0.536.

Some bone metabolic markers can predict the therapeutic effect for short periods like a year after. However, only fewer markers can predict the therapeutic effect for long periods.

Disclosures: H. Naka, None.

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Three-Year Effects of Bisphosphonates on Treatment of Osteoporosis and on Reducing the Risk for Vertebral Fractures of Rheumatoid Arthritis Patients. (Fracture Intervention Trial). H. Nakayama¹, F. Hagiwara*², K. Shimada*¹, T. Matsui*¹, T. Tohma*². ¹Department of Rheumatology, National Hospital Organization, Sagamihara National Hospital, Sagamihara-City, Kanagawa Pref., Japan. ²Department of Rheumatology, Clinical Reseach Center for Allergy and Rheumatology, National Hospital Organization, Sagamihara National Hospital, Sagamihara-City, Kanagawa Pref., Japan.

[PURPOSE] To evaluate the efficacy of alendronate, risedronate and etidronate in the change of bone mineral density (BMD), bone metabolic markers and, reducing the risk for vertebral fractures of rheumatoid arthritis(RA) patients. [METHODS] We carried out 36-month, randomized, intervention trial of five groups in 400 patients with RA(366 women and 34 men). GroupA (n=77): alendronate (ALN) 5mg, GroupR(n=80): risedronate (RIS) 2.5mg, GroupE4 (n=77): cyclical (every 3 month) etidronate (EHDP)400mg, GroupE2(n=72): cyclical EHDP200mg, and GroupD(n=90): not treated with any bisphosphonate(BP). All patients received supplemental calcium and alfacalcidol(VD) 0.5μg. Every dose of BPs was licensed in Japan. BMD was measured at lumbar spine(LS), femoral neck(FN) total hip(TH) by dual energy X-ray absorptiometry technique, and we checked thoracic and lumbar vertebral fractures by X-ray at baseline, 6, and 12 month later. BAP and urine NTX(uNTX) were measured at baseline, 3, 6, and 12 month. [RESULTS] Mean age of the patients of every group was 61–63 years. The mean percent change from baseline to 36 month in LS/FN/TH BMD was +3.7/+3.5% in GroupA, +3.3/-0.7% in GroupR, +2.0/-2.2% in GroupE4, +0.7/-1.9% in GroupE2, and +0.5/2.4% in GroupD. The mean percent change in BAP/uNTX was −25.7/-37.9% in GroupA, −23.2/-23.8% in GroupR, −8.2/-26.7% in GroupE4, −13.1/-11.2% in GroupE2, and +3.9/-7.7% in GroupD. The frequency of incidental vertebral fracture (per 100 person-year) for latter 30 months was 1.8 in GroupA, 1.9 in GroupR, 7.3 in GroupE4, 8.7 in GroupE2, and 16.8 in GroupD. Reductions in risk of vertebral fractures were 89% in GroupA, 88% in GroupR, 56% in GroupE4, 48% in GroupE2[CONCLUSIONS]ALN increased both LS and FN BMD significantly, and RIS also increased LS BMD. ALN and RIS decreased both BAP and uNTX significantly, cyclic EHDP400mg also decreased uNTX. ALN, RIS and cyclic EHDP400mg reduced the risk of vertebral fractures by less than half. These effects were superior to that with cyclic EHDP200mg or VD alone.

Disclosures: H. Nakayama, None.

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Prevalence of Severe Suppression of Bone Turnover Among Patients with Atypical Osteoporotic Fractures. N. Napoli¹, D. Novack¹, K. M. Diemer*¹, M. Watkins¹, S. L. Teitelbaum², R. C. Armamento-Villareal¹. ¹Div. of Bone and Mineral Diseases, Washington University in St Louis, St Louis, MO, USA. ²Department of Pathology, Washington University in St Louis, St Louis, MO, USA.

Recent reports of patients on long-term bisphosphonates (BPs) developing fragility fractures in sites that are not typical for osteoporosis with bone biopsy findings suggestive of severe suppression of bone turnover (SSBT), have raised important safety concerns on the prolonged use of the drug. This study is a retrospective analysis of the patients (14 women and 6 men) at the Bone Health Program at Washington University School of Medicine who underwent bone biopsy because of unusual osteoporotic fractures (femoral shaft, pelvis, rib, metatarsal, shoulder) from November 2004 to January 2007. Ten of 15 patients were found to have biopsy findings consistent with SSBT, defined as total absence of double tetracycline labels in trabecular bone. There were no significant differences in bone mineral density by DEXA, clinical and biochemical data (age, BMI, serum Ca, alkaline phosphatase, PTH, Vitamin D) between the 2 groups, except for the duration of therapy. On the average, SSBT cases took BPs for a significantly longer period of time than non-SSBT (6.1±0.58 vs 3.9±0.83, P =0.05). Histomorphometric analysis also demonstrated significantly lower Ob.S./BS (0.57±0.28 vs 2.23±0.42, P<0.01), but no difference in Oc.S/BS, in cases of SSBT. Our findings suggest that a majority of patients treated with BPs who develop fractures at sites not typical of osteoporosis have SSBT, and that duration of BP therapy is a risk factor. Further studies are needed to determine whether patients at risk for SSBT and atypical fracture can be identified clinically, without biopsy. Given the large numbers of patients who have been treated with BPs for > 5 years without atypical fractures, and reports of normal turnover in a random sample of treated patients (Black DM et al., JAMA, 2006 Dec 27;296(24):2927–38) the overall risk for SSBT in all patients taking BPs is low. However, it is important to define the subpopulation at risk for these atypical fractures.

Disclosures: N. Napoli, None.

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CFOS Trial: Alendronate Once Weekly for the Prevention and Treatment of Osteoporosis in Canadian Adult Cystic Fibrosis Patients. A. Papaioannou¹, C. C. Kennedy¹, G. Ioannidis¹, J. O'Neill*², C. Webber², M. Pui*², A. Freitag*¹, R. Josse*³, A. Cheung*³, Y. Berthiaume*⁴, H. Rabin*⁵, N. Paterson*⁶, A. Jeanneret*², E. Matouk*⁷, J. Villeneuve*⁸, R. McCallum*¹, M. Nixon*¹, J. D. Adachi¹. ¹Medicine, McMaster University, Hamilton, ON, Canada. ²Radiology, McMaster University, Hamilton, ON, Canada. ³Medicine, University of Toronto, Toronto, ON, Canada. ⁴Medicine, Centre hospitalier de l'Université de Montréal, Montreal, PQ, Canada. ⁵Medicine, University of Calgary, Calgary, AB, Canada. ⁶Medicine, London Health Science Centre, London, ON, Canada. ⁷Medicine, Montreal Chest Institute, Montreal, PQ, Canada. ⁸Medicine, Le Centre hospitalier universitaire de Québec, Quebec City, PQ, Canada.

Introduction: A multicentre, randomized controlled trial (RCT) was conducted in adults with cystic fibrosis to assess the efficacy and safety of oral alendronate (FOSAMAX®). This is the only Canadian RCT to examine a bisphosphonate in this patient population.

Methods: Patients received placebo or alendronate 70 mg once weekly for 12-months. All patients received 800 IU Vitamin D and 1000 mg calcium (500 mg supplementation, 500 mg from diet) daily. Patients age 18 years and older with cystic fibrosis (confirmed by positive sweat test or DNA analysis) with a bone mineral density (BMD) T-score <−1.0 were eligible for inclusion. Patients with a prior organ transplantation; endoscopy-proven esophagitis, gastritis, ulceration; metabolic bone disorders; severe renal disease; or using systemic corticosteroids (dose of 7.5 mg/day or greater) in previous 6-months, were excluded. The primary measure was percent change in lumbar spine (LS) BMD after 12-months. Secondary measures were percent change in total hip BMD, proximal femur (PF) BMD, and N-telopeptide after 12-months. Results: A total of 56 patients (34 males, 22 females) were enrolled in the study. The mean age was 29.1 (SD=8.78) years. Mean baseline 25-OHD was 60.3 (SD=27.9) nmol/L. Five patients (4 Placebo, 1 Alendronate) had a new vertebral fracture (all Grade 1) during the study. The number of withdrawals was equal between the treatment groups (4 Alendronate, 4 Placebo). Over 12-months, the active treatment group had a 4.99% (95% CI: 2.40, 7.59) greater increase in LS BMD and a 3.50% (95% CI: 1.37, 5.63) greater increase in PF BMD compared with the placebo group (adjusted for age, body mass index, baseline BMD, number of baseline vertebral fractures).

Conclusion: Patients with cystic fibrosis are at risk for early bone loss due to several factors including delayed pubertal maturation, glucocorticoid therapy, malabsorption of vitamin D, poor nutritional status, inactivity, and hypogonadism. Alendronate was well tolerated and produced a significantly greater increase in BMD over 12-months compared with placebo.

Disclosures: A. Papaioannou, Merck Frosst 2, 5.

This study received funding from: Merck Frosst Canada.

W361

The Bisphosphonates Cause Inflammatory Response In Vivo and in Human Peripheral Blood Mononuclear and Polymorphonuclear Cells In Vitro. W. Park¹, M. Lim*¹, S. Kwon*¹, S. Hong¹, S. Lee*². ¹Medicine/Rheumatology, IN-HA University Hospital, Incheon, Republic of Korea. ²Medicine/Rheumatology, Konkuk University Hospital, Seoul, Republic of Korea.

Bisphosphonates (BPs) are currently mainstay therapy for various clinical conditions including osteoporosis, Paget's disease, multiple myeloma, and metastatic bone disease. However BP associated osteonecrosis of jaw (ONJ) and influenza-like illness were reported mainly in patients who received intravenous (IV) BPs.

Therefore we speculated that high blood concentration of BP after IV or intermittent large oral dose would cause inflammatory reaction and the reaction would be related to those side effects.

Five patients with idiopathic osteoporosis without recent bisphosphonate, glucocorticoid, or the immunosuppressant use were selected. They were given 30 mg of IV pamidronate. Acute phase reactants such as erythrocyte sedimentation rate (ESR) and C-reactive protein (CRP) were measured for 3 days. The in vitro effects of BP on the mRNA expression of the pro-inflammatory cytokines was measured using the human peripheral blood mononuclear (PBMC) and polymorphonucelar (PBPMNs) cells isolated by centrifugation over Histopaque-1077 and Histopaque-1119 (Sigma, 1119–1, St. Louis, MO, USA). The isolated PBMCs and PBPMNCs were incubated in RPMI-1640 containing alendronate 0, 1, 5, 25, 125 μM for 18 hours. One million cells were harvested to extract RNA. The real-time RT-PCR quantified the expression of the interferon (IFN)-γ, TGF-β, interleukin (IL)-1β, IL-10, IL-6, RANKL, TNF-α, TRAP, cathepsin-K (CTK).

At baseline, serum CRP level was normal but the level rose 24–72 hours after pamidronate IV infusion (0.08 ± 0.074 vs. 0.23 ± 0.240 mg/dL, p=0.042, paired T-test). The ESR at the concurrent time did not rose significantly (13.4 ± 18.02 vs. 15.6 ± 15.13 mm/hr). Most of the inflammatory cytokines increased after alendronate pulse dose-dependently. The expression of IFN-γ, IL-6, and TNF-α were most prominent in the PBMCs and those of IFN-γ, IL-6, and IL-1β in the PBPMNCs. (Table 1) RANKL and CTK expression were not detectable in PBPMNCs.

Acute inflammation caused by BP may be due to its high blood level after intravenous infusion or large intermittent oral dose. The clinical study concerning whether the very slow IV infusion or small dose daily oral therapy would minimize those inflammatory side effects is warranted 

Table  .  

Disclosures: W. Park, None.

W362

Comparable Adherence and Improvement in Gastrointestinal (GI) Tolerability with Monthly Oral and Quarterly Intravenous Ibandronate in Patients Who Discontinued Previous Bisphosphonates Because of GI Intolerance. V. K. Piziak¹, A. M. Babbitt*², J. D. Kohles³, E. M. Lewiecki⁴. ¹The Children's Hospital at Scott & White, Temple, TX, USA. ¹²Greater Portland Bone & Joint Specialists, South Portland, ME, USA. ³Roche Laboratories, Inc, Nutley, NJ, USA. ⁴NM Clin Res & Osteoporosis Center, Albuquerque, NM, USA.

The PRIOR study evaluated adherence and gastrointestinal (GI) tolerance to monthly oral (PO) ibandronate 150 mg and quarterly intravenous (IV) ibandronate 3 mg in patients who discontinued previous daily or weekly bisphosphonate (BP) therapy because of GI intolerance.

PRIOR was a 12-month, prospective, open-label, multicenter, noninferiority study in postmenopausal women with osteoporosis or osteopenia. Patients chose either PO or IV ibandronate and could switch formulations once because of side effects. Patients were considered adherent to therapy if adherence (ratio of the actual duration of trial medication intake over the maximum duration of the trial medication intake) was ≥75%. In comparing the IV and PO groups, the between-group difference in adherence rates was adjusted by propensity score and a 2-sided 90% confidence interval (CI) was constructed. If the upper limit of the CI was <20%, then noninferiority of the adherence of the PO group compared with the IV group was concluded. Patients reported GI symptoms by completing a self-administered GI Experience Survey at screening, and Months 1, 4, 7, and 10.

Of 543 patients in the intent-to-treat population, 147 (27.1%) chose PO and 396 (72.9%) chose IV as the initial therapy. Twenty-seven patients switched administration route; 11 switched to IV ibandronate because of GI intolerance (all completed the study), and 16 patients switched to PO therapy for various reasons, including bone or joint pain (n=4), influenza-like symptoms (n=3), and injection-site reactions (n=3) (5 withdrew prior to study completion). Within the per-protocol (PP) population, 69.7% of patients in the PO group and 82.9% in the IV group were adherent to their originally chosen therapy at study end. Based on the adjusted difference in adherence rates in the PP population, adherence to PO therapy was noninferior to adherence to IV therapy (12.4% difference; 90% CI, 5.1% to 19.7%). Overall mean GI tolerance scores improved significantly from baseline to all postbaseline assessments for both the PO and IV treatment groups (P<0.0001 for both). Both dosing regimens were generally well tolerated.

In this population of patients who had discontinued daily or weekly BP treatment because of GI symptoms, adherence rates were similar between patients on monthly PO and quarterly IV ibandronate. Patients reported improvement of GI symptoms compared with baseline while receiving either form of ibandronate.

Disclosures: V.K. Piziak, Proctor and Gamble 2, 8; Roche 8; Lilly 2; GlaxoSmithKline 8.

This study received funding from: Roche Laboratories, Inc.

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Quarterly Intravenous Ibandronate Injections Provide Continuing Benefits in Women with Postmenopausal Osteoporosis: DIVA Study Long-Term Extension. C. Recknor¹, M. Lillestol*², R. Grant*³, C. Neate*⁴, R. R. Recker⁵. ¹United Osteoporosis Centers, Gainesville, GA, USA. ²Internal Medicine Associates, Fargo, GA, USA. ³Hoffmann-La Roche Inc., Nutley, NJ, USA. ⁴Roche Products Ltd, Welwyn Garden City, United Kingdom. ⁵Creighton University, Osteoporosis Research Center, Omaha, NE, USA.

Oral and intravenous (i.v.) ibandronate are available for treatment of postmenopausal osteoporosis. The daily oral regimen (2.5mg) has proven antifracture efficacy,¹ and in the DIVA trial, i.v. injections of 2mg every 2 months (q2mo) or 3mg every 3 months (q3mo) were shown to be superior to daily oral.^2,3 After 2 years, patients in the i.v. arms had superior gains in lumbar spine (LS) BMD (q2mo 6.4%, q3mo 6.3%) compared with daily oral (4.8%; p<0.001). Gains in total hip (TH) BMD were also superior (p<0.001) and reductions in serum CTX were comparable (53.4–59.9%).³

After completion of the 2-year, double-blind DIVA study, patients could be enrolled in a long-term extension (LTE). Eligible patients from the i.v. arms continued to receive open-label i.v. ibandronate at the same dose for a further 3 years (q2mo, n= 253; q3mo, n=263), those in the oral arm were re-randomized to receive q2mo (n=128) or q3mo (n=137) i.v. according to whether they had received 2- or 3-monthly i.v. placebo during DIVA. Here we present the LTE first year results.

Further increases were seen after 1 year in LS BMD (q2mo 0.92%; q3mo 0.95%) and TH BMD (q2mo 0.48%; q3mo 0.13%). A pooled analysis evaluating only those patients who remained on the same treatment for 3 years showed 3-year gains in LS BMD of 7.6% (q2mo; p<0.0001) and 7.0% (q3mo; p<0.0001). 3-year increases in TH BMD were 3.7% (q2mo; p<0.0001) and 3.3% (q3mo; p<0.0001). The reduction of serum CTX seen in the first 2 years of DIVA was maintained in the LTE. In the pooled analysis, median reductions in peak (30 months) and trough (36 months) serum CTX were 89.0% (q2mo), 92.4% (q3mo), and 40.2% (q2mo), 41.4% (q3mo), respectively. Both i.v. regimens continued to be well tolerated, with a similar incidence of adverse events (AEs) to those seen during the 2 years of DIVA and no evidence of late or cumulative toxicity. Treatment-related AEs were reported in 14% (q2mo) and 9% (q3mo) of patients, with serious AEs in 8% (q2mo) and 9% (q3mo), of which two were considered treatment related (both GI disorders in the q2mo group). Five patients (all q2mo) were withdrawn for AEs. The continuing efficacy benefits and good tolerability seen here show that i.v. ibandronate is a valuable long-term treatment option for postmenopausal osteoporosis.

• Chesnut CH, et al. J Bone Miner Res 2004;19:1241–9.

• Delmas PD, et al. Arthritis Rheum 2006;54:1838–46.

• Emkey R, et al. Arthritis Rheum 2005;52:4060.

Disclosures: C. Recknor, F. Hoffmann-La Roche Ltd/GlaxoSmithKline 5.

This study received funding from: F. Hoffmann-La Roche Ltd/GlaxoSmithKline.

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The Efficacy of Fosamax 35mg a Week for the Treatment of Osteoporosis in Postmenopausal Women with Subclinical Hyperthyroidism. L. Y. Rozhinskaya*, G. A. Melnichenko*, Z. E. Belaya*, G. S. Kolesnikova*, A. V. Iljin*, N. I. Sasonova*. Neuroendocrinology and Osteoporosis, The National Research Centre for Endocrinology, Moscow, Russian Federation.

The aim was to estimate the effects of treatment with alendronate (Fosomax 35 mg) in postmenopausal women with osteoporosis and subclinical hyperthyroidism. Thirty postmenopausal women (64 (60–69) years old) with osteoporosis and subclinical hyperthyroidism (77% with endogenous subclinical hyperthyroidism and 23% on L-thyroxine suppressive therapy after thyroidectomy due to differentiated thyroid cancer) were randomly assigned into two groups: (1) 14 women received Fosamax 35 mg a week in combination with 500 mg of calcium and 400 UI of Vitamin D3 (VD) daily; (2) 16 women received 1000 mg of calcium and 800 UI of VD daily. Euthyroidism was achieved in all women with endogenous subclinical hyperthyroidism. An increase in physical activity was recommended to all patients and a hypolipidemic diet was given to those who had had high cholesterol level. Calcium (Ca), phosphorous (P), creatinine (Cre), alkaline phosphatase (ALP), cholesterol, low density lipoprotein (LDL), high density lipoprotein (HDL), triglycerides (TG), cholesterol/HDL ratio) in fasting serum as well as calcium/creatinine ratio in fasting urine (U-Ca/U-Cre); osteocalcin (OC) and C-terminal telopeptide of type I collagen (b-CTx) serum, BMD (DXA; Prodigy, Lunar) at the lumbar spine (L1–L4), femoral neck (FN), total hip (TH) and radius total (RT) were measured at the baseline visit and after 1 year of treatment. At the baseline visit there were not found any differences between the (1) and the (2) groups. After 12 months of treatment the markers of bone metabolism as well as ALP decreased significantly in both groups, though the decreases were significantly greater (p<0.001 for both OC and b-CTx) in the (1) versus the (2) group. BMD in the (1) group increased by 7,6% at L1–L4 (p=0,003), 2,8% at FN (p=0,013), 3,3% at TH (p=0,012) and 3,2% at RT (p=0,047). There was not detected any BMD loss in the (2) group. The changes were not significant between the two groups. The levels of Ca, P, Cre, U-Ca/U-Cre did not change in both groups. Significant improvements in lipid levels were found: HDL increased (p=0.035 (1) p=0,034 (2)), cholesterol/HDL ratio decreased (p=0,011 (1); p=0,004 (2)). In addition, cholesterol (p=0,003); TG (p=0,016) and LDL (p=0,006) decreased for patients from the (1) group. Conclusion: The achievement of euthyroidism and Fosamax 35 mg a week increase BMD in all regions of the skeleton in postmenopausal women with osteoporosis and subclinical hyperthyroidism and reduce bone resorption significantly more than Calcium and VD supplementation.

Disclosures: L. Y. Rozhinskaya, None.

W365

Relationship Between Increasing Annual Cumulative Exposure to Ibandronate, Increases in BMD and Reductions in Clinical Fractures. A. Sebba*¹, R. D. Emkey*², W. A. Blumentals*³, P. N. Sambrook*⁴. ¹University of South Florida, Tampa, Palm Harbor, FL, USA. ²Emkey Arthritis and Osteoporosis Clinic, Wyomissing, PA, USA. ³Roche Laboratories, Inc, Nutley, NJ, USA. ⁴Institute of Bone & Joint Research, University of Sydney, Sydney, Australia.

Analyses of the relationship between changes in bone mineral density (BMD) and fracture reduction with bisphosphonate therapy have produced varying results, but only a limited range of doses has been examined. Data from 4 pivotal phase III clinical trials of ibandronate (2 oral and 2 intravenous [IV]) were pooled and analyzed to examine increases in BMD at various annual cumulative exposures (ACEs) in women with postmenopausal osteoporosis.

Data from the intent-to-treat populations, comprising patients who received oral or IV ibandronate or placebo in 4 studies (Study 4380 [IV fracture study], BONE, MOBILE, and DIVA), were pooled. BONE and the IV fracture study were 3-year placebo-controlled fracture trials, and MOBILE and DIVA were 2-year BMD studies that examined fractures as secondary endpoints. Oral doses included 2.5 mg daily, 20 mg intermittent, 100 mg monthly, 2×50 mg monthly, and 150 mg monthly. IV doses included 0.5 mg quarterly, 1 mg quarterly, 2 mg every 2 months, and 3 mg quarterly. ACE was computed by multiplying the drug strength in milligrams by the number of annual doses and by an absorption factor (0.6% for oral and 100% for IV). Rates of BMD increases at the lumbar spine (LS) and total hip (TH) and rates of all clinical fractures over 2 years were calculated. Linear models, weighted by the sample sizes of the trials, were constructed to examine the clinical fracture rate as a function of increases in LS BMD.

This analysis included a total of 8710 patients. Plots constructed to visually observe any potential association between clinical fractures and ACE showed a trend to decreased clinical fractures with increasing ACE. Plots of change in LS BMD vs ACE showed an increase in percentage change in LS BMD with increasing ACE. The same relationship was seen for TH BMD. There was a statistically significant inverse linear relationship between clinical fracture rates and gains in LS BMD (Figure 1: β = −0.397. P = 0.0046; R² = 0.65).

Increasing gains in BMD at the LS correlated with decreasing rate of clinical fractures. Higher ibandronate doses were associated with the highest level of BMD gain and lowest rate of clinical fracture. 

Figure Figure.. Clinical fracture rates vs percent change in lumbar spine BMD at 2 years

Disclosures: A. Sebba, Roche Laboratories, Inc. 2, 8; Eli Lilly 2, 8; Merck 2, 8; Amgen 2.

This study received funding from: Roche Laboratories, Inc.

W366

Women Are More Persistent with Monthly Bisphosphonate Therapy Compared to Weekly Bisphosphonates: 12 Month Results from Two Retrospective Databases. S. L. Silverman¹, J. A. Cramer*², J. A. Sunyecz*³, C. Sarawate*⁴, C. Harley*⁵, W. A. Blumentals*⁶, S. Poston*⁷, E. M. Lewiecki⁸. ¹Cedars-Sinai Medical Center, Beverly Hills, CA, USA. ²Yale University, West Haven, CT, USA. ³Laurel Highlands Ob/Gyn, Hopwood, PA, USA. ⁴HealthCore, Inc., Wilmington, DE, USA. ⁵i3 Innovus, Eden Prairie, MN, USA. ⁶Roche, Nutley, NJ, USA. ⁷US Health Outcomes, GlaxoSmithKline, Research Triangle Park, NC, USA. ⁸NM Clin Res & Osteoporosis Ctr, Albuquerque, NM, USA.

This study evaluated medication persistence among patients receiving monthly ibandronate versus weekly bisphosphonates. The 12-month results of 2 retrospective database analyses are presented here.

Deidentified patient data were obtained from 2 large managed care claims databases (provided by i3 Innovus and HealthCore Integrated Research Network™) in parallel studies. The combined databases represent over 30 million covered lives. In both studies, eligible women were ≥45 years of age and filled ≥1 pharmacy claim(s) for a weekly or monthly bisphosphonate from April 1, 2005 to November 30, 2006 (i3 Innovus) or October 31, 2006 (HealthCore). Baseline data were collected for 6 months prior to the index date (date of first prescription filled). Persistence, defined as continuous use, was evaluated at 12 months based on a refill gap of 30 days for weekly bisphosphonates; a gap of 45 days was used for monthly ibandronate because of its longer dosing window. Persistence was assessed using Cox proportional hazard models to control for baseline characteristics and potential confounders such as patient age, co-pay amount, comorbidities, long-term prescriptions, concomitant medications, fracture history, and prior DXA scans.

The i3 Innovus database provided claims data from 3,512 women prescribed monthly ibandronate and 13,967 prescribed weekly bisphosphonates. The HealthCore database included 1,006 women receiving monthly ibandronate and 10,658 receiving weekly bisphosphonates. The analyses determined that women on monthly therapy demonstrated higher persistence than women receiving weekly therapy (Table). After adjusting for age, co-pay and comorbidities, monthly users were 25.1% and 37.7% less likely to discontinue therapy versus weekly users in the i3 Innovus and HealthCore analyses, respectively (Table). 

Table  .  

The 12 month results of these analyses suggest that women prescribed monthly ibandronate are more persistent than women on weekly bisphosphonates.

Disclosures: S.L. Silverman, Roche Laboratories, Inc. 2, 5, 8; Merck 2, 5, 8; Procter and Gamble 2, 5, 8; Wyeth 2, 5.

This study received funding from: Roche Laboratories, Inc

W367

Oral Monthly Ibandronate Is Associated with Rapid Suppression of Serum CTX Within Three Days of Treatment Initiation. S. L. Silverman¹, E. Barrett-Connor*², C. Simonelli³, J. D. Kohles*⁴, G. Dasic⁵, N. C. Binkley⁶. ¹Cedars-Sinai Medical Center/UCLA, Beverly Hills, CA, USA. ²University of California at San Diego, La Jolla, CA, USA. ³Health East Osteoporosis Care, Woodbury, MN, USA. ⁴Roche Laboratories, Inc, Nutley, NJ, USA. ⁵GlaxoSmithKline, King of Prussia, PA, USA. ⁶University of Wisconsin, Madison, WI, USA.

The purpose of the Rapid Onset study was to examine the speed of onset and pattern of suppression of the bone resorption marker serum C-terminal telopeptide of type 1 collagen (sCTX) in postmenopausal women with osteoporosis treated for 6 months with monthly ibandronate.

Rapid Onset was a randomized, double blind, placebo-controlled study of women diagnosed with postmenopausal osteoporosis within the past 12 months and with no more than 3 months exposure to daily or weekly bisphosphonate therapy for the 5 years before screening. Participants received once monthly oral ibandronate (150 mg) or placebo for 6 months. Levels of sCTX were measured at baseline and Day 3 (first month only), as well as on Day 7, 14, 21, and 28 after the doses were administered. The primary study endpoint was the relative change in median sCTX from baseline to Day 3. Responder analyses (defined as ≥50% and ≥70% sCTX decreases) were also performed.

An Independent Data Monitoring Committee performed the analyses. Sixty-seven women participated in this study; I did not take any study drug, 49 received ibandronate and 17 received placebo. Mean baseline sCTX measurements levels were the same (0.63 ng/mL) for patients receiving ibandronate and placebo. Rapid suppression of sCTX occurred in those receiving ibandronate; median sCTX was reduced by almost 70% within 3 days of ibandronate administration (P < 0.0001 vs placebo) and remained suppressed at day 28 (median decrease of 43% from baseline, [P = 0.0014 vs placebo]). In contrast, in the placebo group, the median sCTX was reduced by almost 6% on Day 3 and the maximum median percent change in sCTX from baseline was 22% on Day 14. A high proportion of women responded to ibandronate; at Day 3, 71% of patients receiving ibandronate had ≥50% decreases in sCTX and 47% had ≥70% decreases in sCTX. No patients receiving placebo were considered to be responders at Day 3.

This study demonstrated that ibandronate treatment rapidly decreases sCTX levels within 3 days of treatment initiation.

Disclosures: S.L. Silverman, Lilly 2, 8: Merck 2, 5, 8; Procter & Gamble 2, 5, 8: Roche 2, 5, 8.

This study received funding from: Roche Laboratories, Inc.

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Daily or Intermittent Oral Ibandronate Preserves Trabecular Microarchitecture: Micro-Computed Tomography Analysis of Patients in the 3-Year BONE Study. L. G. Ste-Marie¹, B. Langdahl*², D. Masanauskaite*³, D. Ethgen*⁴, R. R. Recker⁵. ¹CHUM Université de Montreal, Montreal, PQ, Canada. ²Aarhus University Hospital, Aarhus, Denmark. ³F. Hoffmann-La Roche Ltd, Basel, Switzerland. ⁴GlaxoSmithKline, Collegeville, PA, USA. ⁵Creighton University, Osteoporosis Research Center, Omaha, NE, USA.

In postmenopausal osteoporosis, trabecular bone microarchitecture contributes significantly to overall bone strength. Micro-computed tomography (microCT) is a quantitative 3D scanning procedure used to assess trabecular architecture. In the 3-year BONE study, oral ibandronate (IBN) administered daily (2.5mg) or intermittently (20mg every other day for 12 doses every 3 months) significantly reduced vertebral fracture risk (primary endpoint) by 62% (p=0.001) and 50% (p=0.006) respectively; significantly increased lumbar spine and proximal hip bone mineral density (BMD) and reduced markers of bone turnover, vs placebo (PBO).¹ 2D histomorphometric analysis of biopsies taken at months 22 and 34 of the study indicated that newly formed bone was of normal quality.² In the current analysis, biopsies have been analyzed by microCT to assess 3D trabecular microarchitecture. Biopsies were analyzed at Creighton University with a Scanco microCT 40 scanner (Scanco Medical, Bassersdorf, Switzerland) and rod and plate distribution quantified by differential analysis of the triangulated bone surface. The structural model index (SMI), was then calculated, with a lower SMI indicating an increased ratio of plates to rods and thus improved trabecular microarchitecture.

Biopsies were obtained from 110 patients, with 84 evaluable by microCT (28 PBO, 56 IBN [both arms]). Median SMI was 1.001 with IBN vs 1.365 with PBO (90% CI for difference in medians: −0.626, −0.033) and connectivity density was higher in IBN-treated patients (median 3.904 vs 3.112/mm³, 90% CI for difference in medians: 0.159, 1.517). This indicates that trabecular microarchitecture was better preserved in patients receiving IBN than PBO. Additional measurements showed that IBN is consistently beneficial on multiple parameters of bone structure and architecture vs PBO. Taken together with previous results from the BONE study showing reduced fracture risk, improved BMD and decreased bone turnover as well as histomorphometric and hip structural analyses,³ these findings indicate that treatment with ibandronate preserves bone strength by maintaining good quality trabecular microarchitecture in women with postmenopausal osteoporosis.

• Chesnut CH, et al. J Bone Miner Res 2004;19:1241-9.

• Recker RR, et al. Osteoporos Int 2004;15:231-7.

• Fuerst T, et al. J Bone Miner Res 2006;21(Suppl. 1):S287 (Abst SU323).

Disclosures: L. G. Ste-Marie, F. Hoffmann-La Roche Ltd/GlaxoSmithKline 5.

This study received funding from: F. Hoffmann-La Roche Ltd/GlaxoSmithKline.

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10-Years-Follow-Up of a Pregnancy-associated Osteoporosis (PAO). A Clinical Case Report. U. C. Stumpf¹, J. Windolf*¹, W. J. Fassbender². ¹Department of Traumatology and Handsurgery, University Hospital, Duesseldorf, Germany. ²Department of Internal Medicine, Hospital z. Hl. Geist, Kempen, Germany.

Pregnancy-associated osteoporosis (PAO) is an uncommon condition characterized by the occurrence of painful fractures during late pregnancy or lactation. The pathophysiology of this disease is uncertain, and its therapeutical management is poorly defined. At the moment exist no guidelines for the therapy. Although rare, diagnosis of pregnancy-associated osteoporosis should be suspected when lumbar or thoracic spine pain occur during pregnancy or in the post-partum period as it can lead to multiple vertebral fractures. Case report: A 41-years-old, premenopausal, slim (BMI= 19) woman is accompanied by pain of the whole spine and the none in general since 10 years. Within her adolescence she got traumatic fractures of the jaw, metatarsus and toes. She had two pregnancies 10 and 8 years ago. Since the first pregnancy a PAO was known, which impaired after the second one. A secondary cause is the treatment for asthma bronchiale over 12 years with inhalative, intermittent oral, corticosteroids (CS) in the range between 5-16 mg /die. Seven years ago, the first DXA-scan was performed and showed a t-score (L2-L4): −2,1 (z-score:-2,0); medication started with etidronate, calcitonin and supplementation of calcium and vitamin D3. In the DXA-scan after 1 year, a consolidation of the PAO could be observed (t-score L2-L4: −1,7; z-score-1,6). With this medication back pain got under control. 5 years ago, the patient had a horse-riding accident with an instable burstfracture of L 1, which was stabilised by internal fixator from Th12 to L2. In the actual DXA-scan an increase of bone mineral density up to a t-score L2-L4 of −1,3 (z-score:-1,3) was observed. In the first laboratory investigation an overall low turnover of bone was seen, due to CS therapy without accelerated bone resorption, PTH and vitamin D3 in normal range. Futhermore a low estradiol (24 pg/ml range: 0.0–270.00 female) despite regular menstrual cycle was noticed. We started a combinated osteoporosis (ibandronate 3 mg iv /12 weeks) and pain treatment (level I WHO) with supplementation of 1000 mg calcium and 800 IU vitamin D3 per die.

Conclusion: As we can observe in this long time follow up the PAO accompanies the patient their whole life and should not be underestimated. We conclude that it is necessary to avoid a long course of pain and fractures in these young women in order to treat the PAO initially adequate with antiresoptive or even osteoanbolic substances to obtain a fast consolidation of BMD and to lower the risk of further fractures and chronification of pain.

Disclosures: U.C. Stumpf, None.

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The Effects of Risedronate and Raloxifene on Serum Lipid Levels in Postmenopausal Women. H. Suh*, K. Lee*. Family Medicine, Gachon Medical School, Gil Medical Center, Incheon, Republic of Korea.

Purpose: Some studies in other countries have reported that the intravenous injection of bisphosphonate and the oral administration of raloxifene, which are used in the prevention and treatment of osteoporosis, have the effect of lowering lipid levels. However, there have been no studies on the effect of the oral administration of bisphosphonate. Furthermore, there have been no studies on the effect of raloxifene in korea. Thus, we studied the effect of typical bisphosphonate medication, risedronate and raloxifene, classified as SERM(Selective Estrogen Receptor Modulator) on the serum lipid levels of post-menopausal korean women.

Methods: At a certain university hospital, each 199 postmenopausal women who had either taken 35mg of risedronate per week or 60mg of raloxifene daily for at least 1 year were selected. We followed them up and compared their baseline serum lipid levels with that of 1 year after.

Results: The risedronate group showed a valid decrease in the level of total cholesterol, LDL cholesterol, non-HDL cholesterol(P<0.05). The raloxifene group showed a valid decrease in the level of total cholesterol, LDL cholesterol, non-HDL cholesterol(P<0.05) and showed a valid increase in HDL cholesterol levels(P<0.05). The raloxifene group showed a valid decrease in the level of total cholesterol, LDL cholesterol, non-HDL cholesterol compared to the risedronate group.(P<0.05)

Conclusions: In post-menopausal korean women, the oral administration of risedronate and raloxifene reduced total cholesterol, LDL cholesterol and non-HDL cholesterol levels. As well, raloxifene showed superior effects to risedronate.

Disclosures: H. Suh, None.

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Efficacy and Safety of Alendronate for the Treatment of Glucocorticoid-induced Osteoporosis in Children: A Prospective Multicenter Study in Japan. H. Tanaka¹, Y. Seino². ¹Pediatrics, Okayama University, Okayama, Japan. ²Pediatrics, Osaka Koseinenkin Hospital, Osaka, Japan.

OBJECTIVE: Glucocorticoid-induced Osteoporosis (GIOP) is being increasingly reported even in children. And efficacy and safety of the use of oral bisphosphonate treatment have been established in adult area. But few randomized control trials have been published and its efficacy and safety are still controversial in growing children. In this preliminary study, we evaluated the effect of an oral bisphosphonate (alendronate) on bone mass in children under long-term glucocorticoid treatment. METHODS: Fifty six children under long-term (over 6months) glucocorticoid treatment were randomly assigned into 2 groups; 30 children (Group A) were treated with alfacalcidol and alendronate, and 26 (Group B) were treated with only alfacalcidol as a control for 2 years. We measured lumbar bone mineral density (LBMD) and metabolic bone markers (NTx and Bone specific ALP) every 6months. RESULTS: Mean LBMD at the start of the trial was 88.7% and 86% of the healthy Japanese children in group A and B respectively. In group A, Mean LBMD increased to 93.6% (6 months), 98.5% (12 months) and maintained until the end of the trial, while it decreased to 83.3% (6 months) and 80.5% (12 months) and then continued to decreased slowly in group B. Urinary excretion of NTx decreased by 33.7% at 6 month while it increased by 9.6% in group B. Considering their condition, increases in the height of all patients but one enrolled in this trial was satisfactory. No new fractures were observed in group A, but multiple vertebral fractures were observed in one case of group B. CONCLUSION: Bisphosphonates can be considered essential components of the treatment of GIOP, not only in adults, but also in pediatric patients. Alendronate has a positive effect on GIOP in children.

Disclosures: H. Tanaka, None.

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Identifying the Treatment Care Gap in Postmenopausal Women at the Southlake Regional Health Centre: The Role of a “Shared Care” Model. C. Thome¹, G. Ioannidis², L. Fraser*¹, E. Ng*¹, J. D. Adachi². ¹Southlake Regional Health Centre, Newmarket, ON, Canada. ²McMaster University, Hamilton, ON, Canada.

Inadequate osteoporosis management in patients with fragility fractures has been well documented in Canada. We performed a patient chart audit to examine the treatment care gap in postmenopausal women 50 years of age and older with at least one fragility fracture and who were admitted to the acute musculoskeletal inpatient nursing unit in the Southlake Regional Health Centre (Newmarket, ON, CAN) in 1999 (T1), 2003 (T2) and 2005 (T3). During this time period, a “Shared Care” program was developed where by all patients with fragility fractures admitted by orthopedic surgeons were seen by one of two rheumatologists. The program began in 1999 and was fully implemented in 2005. The treatment care gap was defined as any fracture patient who was not on therapy at the time of discharge (potential therapies included bisphosphonates, calcitonin, raloxifene and parathyroid hormone). Exact multivariable logistic regression analysis was performed to determine how the treatment care gap changed from T1 to T3. Covariates included in the analysis were patients' history of osteoporosis (yes/no), past osteoporosis medications (yes/no), age of patient (years), length of hospital stay (days), and most responsible physician (orthopedic surgeon, rheumatologist, or general internal medicine physicians alone). Odds ratios (OR) and 95% confidence intervals (95% CI) were calculated. A total of 27, 43 and 44 patients with fragility fractures were evaluated at T1, T2, and T3, respectively. Of these patients, 90, 86 and 96% had a hip fracture. The mean (standard deviation) age, and length of hospital stay of the patients were 81.1 (7.7) years and 8.2 (5.8) days at T1, 79.0 (9.6) years and 10.5 (14.0) days at T2, and 84.8 (7.9) years and 9.6 (7.9) days at T3. The number (%) of patients with a history of osteoporosis and past osteoporosis drug use were 2 (7.4) and 0 (0.0) at T1, 20 (46.5) and 10 (23.3) at T2, and 26 (59.1) and 25 (56.8) at T3. A large number (%) of responsible physicians were orthopedic surgeons (T1=27 (100%), T2=36 (83.7%), T3=34 (77.3%)). The number (%) of patients on therapy at the time of discharge was 0 (0.0), 13 (30.2) and 35 (79.6) at T1, T2, and T3 respectively. As compared with treated patients at T1, the OR of patients treated in T2 and T3 was 1.6 (95% CI: 0.166,∞), and 20.3 95% CI: (2.6, ∞). A patient's history of osteoporosis (OR= 12.5, 95% CI: 2.4, 128.2) and past osteoporosis medications (OR= 6.2, 95% CI: 1.2, 45.5) was associated with higher treatment rates. In conclusion, the treatment gap at the health centre has decrease from 1999 to 2005. The development of the “Shared Care” program has resulted in improved management of patients with fragility fractures.

Disclosures: C. Thorne, Alliance of Better Bone Health 2.

This study received funding from: Alliance for Better Bone Health.

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Two-Consecutive Days-a-Month Risedronate 75 Mg (2Cd) Reduces Vertebral Fracture Risk at One Year. N. Watts¹, J. Brown*², G. Cline*¹, P. D. Delmas⁴. ¹University of Cincinnati, Cincinnati, OH, USA. ²Laval University, Quebec City, PQ, Canada. ³Procter & Gamble Pharmaceuticals, Mason, OH, USA. ⁴INSERM Research Unit 403, Lyon, France.

The antifracture efficacy of new osteoporosis therapies is usually based on placebo-controlled trials, but inclusion of a placebo arm in subsequent clinical trials may be limited by practical or ethical considerations. In these cases, use of historical controls can be explored as an interesting alternative. A recent active-controlled study of risedronate 75 mg for 2 consecutive days per month (2CD) demonstrated that this regimen increases bone mineral density (BMD) comparable to what is seen with risedronate 5 mg daily, which has proven anti-fracture efficacy. To assess the anti-fracture efficacy of this new regimen, we analyzed fracture data in an active-controlled study of risedronate dosing regimens (the 2CD study) using matched historical controls from previous placebo-controlled trials. Women in the 2CD study were matched for age, years since menopause, BMD and prevalent vertebral fractures, with placebo patients in the Vertebral Efficacy of Risedronate Therapy (VERT) trials, forming an historical placebo group. We also constructed an historical active-treatment group from the risedronate 5 mg daily arm of the VERT trials for comparison with the 5 mg daily (n=613) and 2CD (n=616) treatment groups in the 2CD study; historical control groups consisted of daily placebo patients (n=99, matched from 993) and risedronate 5 mg daily patients (n=96; matched from 990) in the VERT studies. Over 1 year of treatment, risk of new vertebral fractures in the 2CD group (1.1%) was reduced by 79% relative to the historical placebo group (5.1%) (OR 0.21; 95% CI, 0.05 to 0.88, P=0.016), similar to the 1-year risk reduction observed in the VERT trials of risedronate 5 mg daily (61-65%). The incidence of new vertebral fractures, assessed for internal validation, was similar in the three treatment groups: 1.0% in the historical risedronate 5 mg group, 1.5% in the risedronate 5 mg daily group from the 2CD study, and 1.1% in the 2CD group. The use of appropriate historical control data provides an approach to the assessment of fracture effects in osteoporosis trials for which placebo-controlled data are not available. Risedronate 75 mg for 2 consecutive days each month appears as effective as the 5 mg daily dose in reducing the risk of new vertebral fractures in the first year of treatment.

Disclosures: N. Watts, Procter & Gamble Pharmaceuticals 5.

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Vertebral Fracture Efficacy with Risedronate Is Apparent Early and Is Sustained. N. B. Watts¹, C. Roux*², X. Zhou*³, A. Grauer¹. ¹University of Cincinnati, Cincinnati, OH, USA. ²Cochin Hospital, Rene Descartes University, Paris, France. ³Procter & Gamble Pharmaceuticals, Mason, OH, USA.

Treatment with an agent that provides both early and sustained fracture protection can benefit patients. Superficial examination of published studies suggests that the early effect of treatment on vertebral fracture (as reflected in relative risk reduction) may lessen over time. In this study, absolute vertebral fracture risk reduction was examined in the Vertebral Efficacy with Risedronate Trials (VERT).

The population included 2442 patients from VERT-NA and VERT-MN who were treated with at least 1 dose of either placebo or risedronate (RIS) 5 mg daily. The mean age was 69 and the mean spine T-score was −2.5. Treatment groups were balanced with respect to key baseline characteristics. Fracture endpoints included clinical vertebral fracture and radiographic vertebral fractures. Difference in risk of vertebral fractures between placebo and RIS 5 mg groups was estimated using the difference of the Kaplan-Meier (KM) estimators at months 3,6,… and 36.

The findings for clinical vertebral fracture were consistent with the rapid onset of risk reduction with risedronate treatment; relative to placebo, RIS significantly reduced the risk for clinical vertebral fracture starting from Month 6, with significant absolute risk reduction of 1.0% (95% CI: 0.3%, 1.6%). The magnitude of the difference in fracture risk between the placebo and risedronate 5 mg groups increased over time (Figure). Relative to placebo, RIS significantly reduced the risk for radiographic vertebral fracture starting from the first scheduled radiographic visit at one year. In general, the mean magnitude of the difference in the fracture risk between the placebo and risedronate 5 mg groups increased over time (from 5.3% at 12 months up to 8.6%, (95% CI: 5.5%, 11.6%).

In summary, the results confirmed the risedronate rapid protection against clinical vertebral fracture as early as Month 6. Further, risedronate anti-vertebral fracture efficacy as reflected in absolute fracture risk reduction increased over 3 years for both clinical and radiographic vertebral fractures

Disclosures: N.B. Watts, Procter & Gamble Pharmaceuticals 5.

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Long-term Bisphosphonates Therapy and Increase Fracture Risks. S. J. Wimalawansa. Medicine, Div. Endocrinology, UMDNJ-RWJMS, New Brunswick, NJ, USA.

Bisphosphonate have been in the market for over 2 decades. However, no credible evidence available on fracture benefits of continuation of bisphosphonate therapy > 5 years. The terminal half-life of alendronate is similar to bone mineral, ∼10 years, while risedronate is ∼2 years. The skeletal effects of these two bisphosphonates last for years after stopping therapy. Studies with alendronate show 5 years after discontinuation of therapy, patients bone marker levels remain below the pretreatment levels.

The patients on long-term bisphosphonate therapy continue to maintain their BMD with reduced bone remodeling markers. Once bisphosphonate therapy is discontinued, BMD is stable for about 1 to 2 years depending on bisphosphonate used. Increase in bone remodeling, as indicated by markers of bone turnover will occur before reduction of BMD begins. Therefore, bone markers are much sensitive indicators than BMD to consider starting bisphosphonate or another therapy following discontinuation of bisphosphonate.

Current data suggest that discontinuation of bisphosphonate after 4 to 5 years of therapy does not increase fracture risks, but may in fact be beneficial to patients. Whereas, patients who continue to have fractures while on therapy are non-compliant, or likely to have an underlying secondary cause of bone loss. After several years' therapy with bisphosphonates, stable BMD (with suppressed bone turnover markers) or demonstration of normal histology by biopsy does not signify that the drug protects against fractures. Not only biopsies can miss generalize trends, but also the very small sample sizes used in these studies give false sense of security of the safety of long-term bisphosphonate use. Although the optimal duration of treatment with bisphosphonate (or any therapy) for osteoporosis is uncertain. Data accumulating to suggest that there are no benefits to patients by continuation of therapy beyond five years. The only useful therapy to be continued in the long-term is calcium and vitamin D supplementation. To-date, no independent data available on fracture efficacy of long term anti-osteoporosis medications; this need to be critically evaluated. Prospective clinical trial addressing this issue is underway at this institute. It is now a decade since the first FDA approval of bisphosphonates for osteoporosis. It is time now to examine this issue seriously and to develop clinical practice guidelines (and cost-benefit analysis) for minimizing complications of long-term use of bisphosphonates. This pro-active measure may save millions of tax payer dollars and other health costs (i.e., cost of unnecessary drug therapy, management of complications, and the potential for increased fractures incidence).

Disclosures: S.J. Wimalawansa, None.

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Bringing Bone Health to Seniors' Communities: A Unique Approach to Help Manage Osteoporosis. M. Kloseck*¹, M. van Zandvoort*², R. Crilly³, M. Speechley*⁴. ¹Faculty of Health Sciences, University of Western Ontario, London, ON, Canada. ²Department of Health and Rehabilitation Sciences, Faculty of Health Sciences, University of Western Ontario, London, ON, Canada. ³Department of Medicine, University of Western Ontario, London, ON, Canada. ⁴Department of Epidemiology and Biostatistics, University of Western Ontario, London, ON, Canada.

Osteoporosis is preventable and treatable, yet there is a significant care gap in Canada. We hypothesized that engaging seniors in raising awareness, identifying neighbours and others in their community at risk and a peer-led senior-friendly education program would increase diagnosis and treatment of osteoporosis, and that ongoing peer support would encourage compliance and persistence with medication and lifestyle change. This hypothesis is being tested in a randomized controlled trial (RCT). In Phase 1 (completed) community capacity building and participatory action research methodology was used to train seniors in a local community to act as peer educators and mentors within their community. Ten seniors (mean age=82 yrs. ± 6.74 yrs. SD, range=71-90) participated in a 2-week training program containing 5 osteoporosis education modules and 1 session on public presentation skills delivered by local experts. Once trained, these seniors formed the Community Osteoporosis Advisory Committee which collaboratively developed a peer advisor training manual, designed a community information and recruiting program, and developed a senior-friendly education and support program which they practiced until all were comfortable. Pre- and post-knowledge surveys showed a statistically significant change (p=.04) in knowledge from project start to end of Phase 1. Confidence by seniors in their role as osteoporosis advisors increased significantly (p=.02), as did confidence in their ability to deliver information to their neighbours through community presentations (p=.02). Overall confidence in their role as peer educators and mentors increased significantly from project start to end of Phase 1 (p=.01). In Phase 2 (in progress) the senior-led education and support program is being tested in a RCT (n=100) in a local community of seniors (n=2500, mean age=79 ± 9.53 SD). Outcomes include BMDs performed, treatments begun, appropriateness of treatment and persistence. Preliminary baseline knowledge surveys (n=33, mean age=81 ± 7.18 yrs. SD, range 67-91 yrs., 97% female, 3% male) show a substantial gap in knowledge. On the 19-item osteoporosis questionnaire the mean correct minus incorrect score=6.15 ± 4.79 SD. Scores ranged from −4 to 16 out of 19. Forty-one percent reported having fallen in the past year. This unique approach to diagnosing and treating osteoporosis may be a useful model for other communities of seniors.

Disclosures: R. Crilly, Alliance for Better Bone Health 2.

This study received funding from: Alliance for Better Bone Health.

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Compliance with Osteoporosis Treatment and Incidence of Hip and Wrist Fracture after Forearm BMD Screening. M. W. J. Davie, T. Jones*, N. Dugard*. Charles Salt Research Centre, Robert Jones & Agnes Hunt Orthopaedic Hospital NHS Trust, Oswestry, United Kingdom.

Treatment compliance in osteoporosis may affect fracture outcome. To investigate compliance and fracture after a screening programme, we revisited 1299 women aged 50-97y screened by forearm scanning after a minimum of 3 years. Women were scanned at the request of their doctor (GP) and treatment advised if BMD values were <0.34g/cm² (osteoporosis (OP)). Treatment was decided by the woman's GP. No follow up was arranged. Women completed a detailed questionnaire at the scanning visit and again when revisited. Women lost to follow up were traced through health records and treatment details between scanning and revisit obtained from the women's own GP. 74% of women returned their questionnaires with no difference between those osteoporotic and those not. Fractures were validated with a 10% sample (and were correct within 5%).

All but 98.5% were traced of whom 14.4% had died. 254 women had osteoporosis at the first visit (aged 50-9, 2.4% had OP, 60-9 15.8%, and > 69 54.3%). Of these 37.4% had never started any treatment (excluding Ca and Vit D only) — with no difference across ages. 1.3% had HRT and 98.7% Bisphosphonate (BP) of which 38% used Didronel, 43% Fosamax and 19% had had > 1 BP. Compliance with BP dropped to 73.8% after 1yr (no difference across ages) with 22.9% only collecting 1 script. Fewer women with OP in institutions had ever had treatment (28%, p<0.01) and were also more likely to stop treatment (28.6% had only 1 script). Excluding women who were no longer available compliance at 2yr was 68.3%, 3yr 58.6%, 4yr 50% and 49.6% at 5yr. Hip fracture occurred in 1 woman with OP out of 38 (age 76.6±8.1y) who were never treated and in 7 of 55 women (73.2±7.3y (age p<0.05)) fully compliant for 5yr. The excess in women fully compliant occurred after > 3y treatment (2 had used Didronel, 2 Fosamax, 3 both). Wrist fractures occurred in 3 women in each group.

Compliance declined early suggesting early follow up is indicated. Treatment in institutions may be overlooked. Compliance is not necessarily associated with hip fracture reduction. However the age of patients with hip fracture (75% were > 80y) is consistent with BP efficacy being less in this age group. Moreover most of the hip fractures occurred > 3y after treatment started when BP may be less effective at preventing hip fracture.

Disclosures: M.W.J. Davie, None.

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An Outreach Program Improved Osteoporosis Management after a Fracture. A. C. Feldstein¹, W. M. Vollmer*¹, D. H. Smith*¹, A. Petrick*¹, J. Schneider*¹, H. Glauber*², M. Herson*². ¹Center for Health Research, Kaiser Permanente Northwest, Portland, OR, USA. ²Northwest Permanente, Portland, OR, USA.

This longitudinal retrospective cohort study evaluated implementation of a 2-phase intervention to improve management of osteoporosis after a fracture. Stakeholder barriers and facilitators of osteoporosis management were also determined. The study setting was a nonprofit group model HMO in the U.S. Pacific Northwest with 15 clinics, 480,000 members, and comprehensive electronic medical record data. Study participants were women members aged 67 or older who sustained a qualifying clinical fracture(s) and who had not received a bone mineral density (BMD) measurement or osteoporosis treatment in the 12 months prior to the fracture (n=3588) and their 255 primary care providers (PCPs). Interviews/focus groups were conducted with 58 patients, health care managers, PCPs, and orthopedic clinicians. Phase I included outreach to clinicians and patients; Phase 2 added clinician/staff incentives. The primary outcome was “osteoporosis management” _receipt of a BMD measurement or osteoporosis medication in the 6 months after an index fracture. Prior to the intervention, 13.4% (95% confidence interval [CI] = 12.0%-14.8%) of fracture patients had received osteoporosis management; the pre-intervention time trend was not significant. After the intervention, the unadjusted probability of osteoporosis management increased on average by 3.1% (95% CI = 2.6%-3.5%) every 2 months throughout both study phases. There was no significant added improvement in Phase 2. Overall, the probability of osteoporosis management increased from the baseline level to 44.0% (95% CI = 40.0%-48.0%) by the end of the study period (20 months post intervention). Adjusted models revealed that osteoporosis management was less likely in older patients and in those with dementia and was more likely in those with fractures more highly associated with osteoporosis. Improvement varied by clinic and was less likely for patients with dementia. Patient knowledge gaps and fatalism were common, especially among older patients. Common clinician barriers were lack of time, difficulty interpreting all relevant results, patient management concerns in the transition from specialty to primary care, and frustration with the side effects of and poor patient adherence to osteoporosis medications. This study found that an outreach program to clinicians and patients improved the management of osteoporosis after a fracture. More-tailored interventions may be necessary for high-risk subgroups. There is also a need for more effective patient and clinician education and a stronger role for specialists.

Disclosures: A.C. Feldstein, None.

This study received funding from: Merck & Co., Inc.

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Adherence to Raloxifene Therapy: Assessment Methods and Relationship with Efficacy. J. Finigan¹, J. A. Clowes², M. A. Paggiosi*¹, D. K. Swindell*¹, K. E. Naylor*¹, N. F. A. Peel¹, R. Eastell¹. ¹Academic Unit of Bone Metabolism, University of Sheffield, Sheffield, United Kingdom. ²Mayo Clinic School of Medicine, Rochester, MN, USA.

Adherence to study medication is usually estimated by counting returned tablets. This method relies on subjects' honesty and may be inaccurate.

We aimed to assess adherence more accurately, and examine its effect on measures of bone response, by using monitoring caps on the medication bottles in two 2-year studies of osteopenic women, ages 50 to 80. Study 1: 75 women took 60 mg Raloxifene. Study 2: 50 were randomised to take 60 mg Raloxifene (n=25) or no treatment (n=25). Electronic caps (MEMS, Aardex) recorded the date and time whenever the bottle was opened. Returned tablets were also counted. We measured bone mineral density (BMD) by DXA at the lumbar spine (LS) and total hip (TH) twice, a week apart, at baseline and 2 years. Percentage changes from mean baseline to mean 2-year BMD were calculated. We measured urinary NTX from samples taken while fasting on 4 consecutive mornings at baseline, 1 and 2 years. We calculated the mean NTX for baseline and the mean for years 1 and 2 combined, and hence the mean percentage change since baseline.

Combining data from both studies, 100 women took Raloxifene, of whom 29 withdrew during the 2 years. Adherence was assessed by both methods for 71 subjects as a percentage of the total study days. The two methods correlated significantly (p<0.001, Spearman's rho=0.73) but the tablet count showed a higher mean adherence than the MEMS caps (91.4% v 79.8%, p<0.001), with greater divergence at lower adherence levels. Using the MEMS results for 65 subjects with complete data, adherence correlated with NTX response (p<0.01, rho= −0.33) but not BMD response (LS p=0.087, rho=0.21, TH p=0.25, rho=0.15). However there was a threshold effect, with adherence in the lowest quartile associated with poorer BMD response at LS (+0.61% v +2.10%, p=0.040) and at TH (-0.66% v +0.91%, p=0.012).

Monitoring caps may assess adherence more accurately than tablet counts. The degree of adherence is associated with both bone turnover and BMD responses.

Disclosures: J. Finigan, None.

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Adherence and Persistence with Teriparatide Therapy in Commercial Insurance Plans and Managed Medicare. S. A. Foster¹, K. A. Foley*², E. S. Meadows*¹, J. A. Johnston*¹, S. Wang*², G. M. Pohl*¹, S. R. Long*². ¹Eli Lilly and Company, Indianapolis, IN, USA. ²Thomson Medstat, Ann Arbor, MI, USA.

The purpose of this study was to evaluate adherence to teriparatide (TPTD) therapy and to identify the factors that impact persistence. Beneficiaries (45 years and older) with at least one claim for teriparatide in 2003 or 2004 and continuous enrollment in the previous 12 months and subsequent 6 months were identified in a large (7.6 million covered lives) national commercial and Medicare administrative claims database (MarketScan). Adherence was assessed through the calculation of the medication possession ratio (MPR) at 6 months and 12 months. The MPR was calculated as the sum of days supply dispensed between the start of the observation period and the end of the observation period and divides that number by the total number of days being examined. Persistence was calculated as the time (days) to discontinuation (meaning the last prescription for the index drug regardless of gaps in fills — with at least 2 months remaining enrollment without the script). In addition, factors associated with time to discontinuation in treatment were assessed using Cox proportional hazards models. The average age of patients in this analysis was 70.3 and 89.7% were female. In the 12 months prior to initiating TPTD, 38% of patients had experienced a fracture. The average MPR at 6 months was 0.74 (N=2218) and 12 months was 0.66 (N=1303). At 6 months (N=2218), 64.6% of patients remained on TPTD therapy and at 12 months (N=1303), 56.7% of patients remained on TPTD. Prior use of an antiresorptive therapy, having BMD screening within the 12 months prior to starting TPTD, and lower patient copayments were associated with longer times until discontinuation. Overall, despite being an injectable, TPTD adherence was comparable with adherence rates found with other osteoporosis medications in the literature. Understanding the factors that impact persistence with therapy may allow clinicians to make informed decisions about patient selection that may improve persistence.

Disclosures: S.A. Foster, Eli Lilly and Company 3.

This study received funding from: Eli Lilly and Company.

W381

Patient Perceptions and Reasons for Discontinuing Teriparatide Treatment: Initial Results from the DANCE Observational Study Addendum. D. T. Gold¹, D. L. Weinstein*², G. Pohl*³, K. D. Krohn³, E. Meadows*³. ¹Psychiatry & Behaviorial Sciences, Sociology, and Psychology, Duke University, Durham, NC, USA. ²Medicine, Washington University, St. Louis, MO, USA. ³Outcomes Research, Eli Lilly, Indianapolis, IN, USA.

Purpose: To determine the perceptions of patients who initiate and persist on treatment with teriparatide (Forteo) for osteoporosis.

Methods: The Direct Assessment of Non-vertebral Fractures in the Community Experience (DANCE) is a multi-center, prospective, observational study. The DANCE addendum includes questionnaires (Q1 and Q2) completed by the teriparatide patients. Administered at initiation of therapy, Q1 includes items related to the patient's experience with self-injection drugs, perceptions about the severity of their osteoporosis, relationships with their prescribing physician and staff, and concerns about starting treatment. The Q2 instrument (completed 2 to 6 months after initiating therapy) includes items regarding training or other support offered at therapy initiation, the patient's early experience with teriparatide treatment (including problems, adverse events, and concerns), and whether the patient is still taking treatment. If the patient has stopped teriparatide treatment when Q2 is completed, the patient is asked to choose one of 6 pre-specified reasons why they stopped treatment.

Results: As of January 2007, 700 patients had completed both Q1 and Q2. The DANCE Addendum patients were relatively homogeneous in their perceptions at initiation of teriparatide treatment. Fewer than 1 out of 4 (n=150, 22%) patients reported that they had previous experience with self-injectable medications. Many (n=409, 59%) patients believed their osteoporosis was severe or very severe. Nearly all (n=662, 95%) believed that it was extremely or very important to treat their osteoporosis. Additionally, some (n=167, 24%) patients were extremely or very concerned about their ability to pay for teriparatide. Few patients (n=50, 7%) reported discontinuing teriparatide treatment by the time Q2 was administered. The reasons for discontinuation are shown below. 

Table  .  

Conclusions:: Few patients have discontinued teriparatide treatment in the DANCE Addendum before the first follow-up visit 2-6 months after initiation. The 2 most common reasons for discontinuation were (1) that treatment concerns outweighed the benefits of teriparatide and (2) concern about paying for teriparatide.

Disclosures: D. T. Gold, Eli Lilly and Company 5; Procter and Gamble, sanofi-aventis, GlaxoSmithKline, Roche, and Amgen 5, 8.

This study received funding from: Eli Lilly & Company.

W382

A Group-based Patient-Education Program Increases Knowledge of Osteoporosis and Adherence to Pharmacological Treatment — A Randomised Trial. D. Nielsen¹, J. Ryg*¹, W. Nielsen*², B. Knold*¹, N. Nissen*¹, L. Stilgren¹, A. Riis Madsen*¹, K. Brixen¹. ¹Endocrinology, Odense University Hospital, Odense, Denmark. ²Physiotherapy, Odense University Hospital, Odense, Denmark.

Purpose: A large number of studies have demonstrated that pharmacological therapy of osteoporosis is effective; however non-adherence to such therapy is a well recognized problem. Few studies, have examined the effect of particular patient education programs on knowledge and adherence to therapy. We hypothesised that group-based, multi-disciplinary, education program increases the patients' knowledge of osteoporosis and adherence with pharmacological therapy. Methods: A total of 300 patients (32 men aged 65 ± 9 yrs and 268 women aged 63 ± 8 yrs), recently diagnosed with osteoporosis and started on specific treatment, were randomised to either the “school” (n=150) or “control” (n=150) group. In the school-group, patients attended four classes with 8-12 participants during four weeks (a total of 12 hours). Teaching was performed by nurses, physiotherapists, dieticians, and doctors and was based on dialogs and situated learning. The classes covered “facts on osteoporosis”, “fractures and pain”, “diet”, “preventive measures”, “balance and exercise”, and “medical treatment”. Teaching was designed to increase empowerment. We assessed the patients' knowledge about osteoporosis and compliance to pharmacological treatment, using validated questionnaires. Results: At baseline, no significant difference was seen between the groups regarding knowledge score (20.1 ± 5.6 versus 21.0 ± 4.7). The change in knowledge at 3 months was significantly higher in the school group (3.43 ± 4.0) compared with the control group (0.33 ± 2.48) (p<0.001). Adherence with pharmacological therapy at 2 years was significantly higher in the school group (99%) compared to controls (84%) (p<0.001, Log Rank test). Neither baseline knowledge score, nor change in knowledge score, however, was significantly associated with compliance. Conclusion: A multidisciplinary patient education program leads to increased knowledge of osteoporosis and increased adherence to treatment. Knowledge, however, did not predict adherence. There are more issues important when it comes to adherence to treatment.

Disclosures: D. Nielsen, Eli Lilly 2, 8; Nycomed 2; Servier 2, 8; Pharma Vinci 2, 8.

This study received funding from: Danish Osteoporosis Society, Augustinus Foundation, Frimondt-Heineke Foundation, Sygekassernes Helse Foundation, Medical Industries: Novo Nordisk, Aventis, Eli Lilly, MSD and Pharma Vinci, Nycomed and Servier.

W383

Convenience and Preference Perceptions After 1 Year of Oral Bisphosphonate Treatment in a Group of Brazilian Women. S. Ragi-Eis¹, I. F. Francischetto*², B. Albergaria*², A. A. Oliveira*², F. V. Simøes*², K. Milagre*², C. Bonomo*². ¹Clinical Research Department, CEDOES, Vitoria — ES, Brazil. ²Clinical Research, CEDOES, Vitoria — ES, Brazil.

Once monthly dosing (OM) of ibandronate has been shown to provide similar efficacy in increasing BMD to once weekly dosing (OW) of alendronate for treatment of osteoporosis in postmenopausal women. Whether patients will prefer monthly dosing to weekly dosing for a chronic condition such as osteoporosis has not been well documented in a Latin American population. The aim of this study was to assess the preference as well as their perception of convenience for monthly versus weekly dosing regimens among sample of Brazilian osteoporotic postmenopausal women.

One hundred and eighty one postmenopausal women with osteoporosis were originally randomized in the Motion Study, to receive treatment with either (1) alendronate 70 mg once weekly plus once monthly placebo tablet or (2) ibandronate 150 mg once monthly plus once weekly placebo for 12 months. From this subset, 145 (One hundred and forty five) women (80.1%) were interviewed during the follow-up visits, after completion of the study. The patients were asked about the convenience as well as their preference about treatment regimens to continue their treatment.

Among those expressing preference (n=115), 69 % (79) pointed the once monthly dosing regimen as the preferred regimen to continue their treatment from that moment on. The main reasons to justify their option were “better to remember” (30 subject for the OM vs 22 for the OW) and “more favorable time interval” (22 subjects for the OM vs 0). In addition, from the patients expressing opinion about convenience (n=81) the once monthly regimen was considered by 73% (59) to be the more convenient one. The key factor to justify their opinion was “fits better in my lifestyle” (56 subjects for the OM vs 16).

After having the opportunity to try both weekly and monthly regimens for a 12 month period, the majority of the patients preferred the OM one to continue their treatment. They also considered this option the more convenient one. In conclusion, patient preference and convenience perceptions about dosing regimen should also be taken into consideration when taking the decision to prescribe an oral bisphosphonate for Brazilian Women. This could have a favorable impact on treatment compliance.

Disclosures: S. Ragi-Eis, Sanofi-Aventis, Eli Lilly, GE 2; Merck, Roche, Sanofi-Aventis, Eli Lilly 8.

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Persistence with Teriparatide Therapy Among Participants in the DANCE Trial. K. Taylor*¹, D. Gold*², S. Silverman*³, P. Chen*¹, M. Wong¹, K. Krohn*¹. ¹Eli Lilly and Company, Indianapolis, IN, USA. ²Duke University, Durham, NC, USA. ³OMC Research, Beverly Hills, CA, USA.

Despite the benefits of osteoporosis (OP) therapies shown in clinical trials, poor persistence is a common cause of reduced patient benefit in a “real world” setting [Mayo Clin Proc 2006;81:1013-22]. Overall persistence with OP therapies has been reported to be between 25-45% at 1 yr [Maturitas 2004;48:271-87; Arch Int Med 2005;165:2414-9]. Few, if any, US studies have assessed persistency with teriparatide (TPTD) in community settings. A prescription tracking database [IMS Health LRx™] estimated persistency with TPTD (20 μg/d) to be about 30% at 12 months (Lilly data on file). The ongoing, observational DANCE study captures data on persistence with TPTD therapy. A survival model (Figure) assesses the rate of patients persisting with therapy at defined time points after their start date. This analysis included 2474 patients who were eligible to have had 12 months of TPTD therapy. The model estimated ∼70% of patients persisted with therapy at 12 months.

Persistence was not significantly affected by age, co-morbidity, or baseline severity of OP, as assessed by BMD T-score, prevalent fracture status, or clinical risk factors for fracture. Of these 2474 patients, 419 completed 18-24 months of therapy, 852 discontinued therapy before completing the 18-24 month course, and 1203 remained in the study. Reasons for discontinuation were categorized as patient decision (422,49.5%), physician decision (136 patients, 16%), serious adverse event (57, 6.7%), sponsor decision (2, 0.2%) and other unstated reasons (172, 20.2%), and 63 patients (7.4%) had no data available. For the 422 patients who listed “patient decision” as the reason for discontinuation, additional comments about their discontinuation were collected, and included insurance reimbursement, inconvenience, and concerns about injection. Persistence with TPTD in DANCE subjects at 12 months may be higher than that measured using a prescription tracking database. Different methodologies used to measure persistence may reflect some of this disparity. Further analysis of the DANCE cohort may elucidate factors affecting persistence to TPTD therapy and offer new approaches to optimize TPTD use in the community.

Disclosures: K. Taylor, K. Taylor, P. Chen, M. Wong, K. Krohn 1, 3; D. Gold 2; S. Silverman 2.

This study received funding from: Eli Lilly and Company.

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Safety and Tolerability of Bazedoxifene in Postmenopausal Women with Osteoporosis: Results from a 3-Year, Randomized, Placebo- and Active-Controlled Clinical Trial. J. D. Adachi¹, C. H. Chesnut², J. P. Brown³, C. Christiansen⁴, L. A. Russo*⁵, C. E. Fernandes*⁶, J. C. Menegoci*⁷, A. Kung⁸, A. A. Chines⁹, L. Bessac*⁹, D. Chakrabarti*⁹. ¹St. Joseph's Hospital — McMaster University, Hamilton, ON, Canada. ²University of Washington, Seattle, WA, USA. ³CHUL Research Center, Laval University, Quebec, PQ, Canada. ⁴Center for Clinical and Basic Research (CCBR), Ballerup, Denmark. ⁵CCBR, Rio de Janeiro, Brazil. ⁶Women's Health and Wellness Institute (ISBEM), São Paulo, Brazil. ⁷San Francisco Clinic, São Paulo, Brazil. ⁸The University of Hong Kong, Hong Kong, China. ⁹Wyeth Pharmaceuticals, Collegeville, PA, USA.

Bazedoxifene (BZA) is a novel selective estrogen receptor modulator selected for clinical development based on preclinical evidence of tissue-selective estrogen agonist activity on the skeletal system and lipid metabolism and estrogen antagonist activity on breast and uterine tissues. A total of 7492 postmenopausal women with osteoporosis (mean age ± SD, 66.4 ± 6.7 years) were randomized to receive BZA 20 mg, BZA 40 mg, raloxifene (RLX) 60 mg, or placebo (PBO) daily and received ≥1 dose of study medication. At 36 months, BZA exhibited a statistically significant decrease in new vertebral fracture compared with placebo (primary efficacy endpoint to be reported elsewhere). We report here the 3-year safety and tolerability data from this study.

After 3 years of treatment, BZA was overall well tolerated and exhibited a favorable safety profile. Findings of selected safety/tolerability analyses are shown below. 

Table  .  

No safety concerns related to the cardiovascular and reproductive systems, including breast, were observed in the BZA treatment groups. There was a higher incidence of DVT in the BZA groups compared with the PBO group. The incidence of breast cancer and cystic/fibrocystic breast disease was lower in women receiving BZA than in those receiving PBO. Transvaginal ultrasonography examinations revealed that BZA had no significant effects on the endometrium or ovaries.

In conclusion, BZA has a favorable safety and tolerability profile in postmenopausal women with osteoporosis. The safety and tolerability profile was similar between BZA 20 and 40 mg groups. BZA is a promising new SERM for the prevention and treatment of postmenopausal osteoporosis.

Disclosures: J.D. Adachi, Amgen, Astra Zeneca, Eli Lilly, GlaxoSmithKline, Merck, Novartis, Pfizer, Procter & Gamble, Roche, Servier 5, 8; Amgen, Astra Zeneca, Eli Lilly, GlaxoSmithKline, Merck, Novartis, Pfizer, Procter & Gamble, Roche 2.

This study received funding from: Wyeth Phamaceuticals.

W386

Postmenopausal Osteoporosis and Weight Gain. A. Bazarra-Fernandez*. ObGyn, Juan Canalejo University Hospital Trust. La Coruña. Spain, La Corunna, Spain.

Background: Recent studies have found weight gained during menopause increase the risk of high blood pressure, the diabetes, heart disease, and has been strongly linked to increased incidence of breast and other hormone-related postmenopausal malignancies. These healthcare concerns have led to the conception of specific products that target menopausal weight gain.

Aim: Looking over weight gain and osteoporosis treatment in climacteric. Material and methods: 20 women who were 44 to 58 years old have been recruited. BMI was increased to age. Those with an intact uterus have moderate to severe vasomotor symptoms associated with the menopause, moderate to severe symptoms of vulvar and vaginal atrophy and risk of postmenopausal osteoporosis. They were ascribed to equal two 10 women groups. One group was assigned to 2 mg drospirenone /1 mg 17 beta- estradiol hemihydrate. The other group treated with 40 mg soy bean.

Results: in the women on 2 mg drospirenone /1 mg 17 beta- estradiol hemihydrate medication decreased, moderate to severe symptoms of vulvar and vaginal atrophy vasomotor symptoms associated with the menopause in regard to the other group treated with 40 mg soy bean. They had weight main loss of 3 kg in one year, (P < 0.05).

Conclusions: Human HRT is in relation to decrease osteoporosis. Nobody noted 17 beta-estradiol is in relation to breast cancer. Estradiol is the same oestrogen produced by the ovaries before menopause. Drospirenone has the unique property of reducing water retention often associated with the use of oestrogen and other synthetic progestin hormones. The impact of obesity on hormone replacement therapy is due to many women associate hormones with weight gain. This late medication formula can be beneficial in minimizing uncomfortable symptoms, such as weight gain, hot flashes, night sweats, and mood swings, associated with the natural progression of a woman's life cycle. So, it is due to conduct one great try to make clear and more comprehensible these points.

Disclosures: A. Bazarra-Fernandez, None.

W387

Safety and Tolerability of Bazedoxifene for the Prevention of Postmenopausal Osteoporosis. C. H. Chesnut¹, C. Christiansen², H. C. Hoeck*², H. Genant³, D. van Duren*⁴, A. B. Levine*⁵, A. A. Chines⁵, G. Constantine*⁵. ¹University of Washington, Seattle, WA, USA. ²Center for Clinical and Basic Research, Ballerup, Denmark. ³University of California San Francisco, San Francisco, CA, USA. ⁴Menox BV, Nijmegen, The Netherlands. ⁵Wyeth Pharmaceuticals, Collegeville, PA, USA.

Bazedoxifene (BZA) is a novel selective estrogen receptor modulator (SERM) selected for its antagonist activity on endometrial and breast tissues. Here we report on the safety and tolerability of BZA for the prevention of postmenopausal osteoporosis in a randomized, double-blind, placebo- and raloxifene-controlled phase III trial. Healthy postmenopausal women (aged ≥45 years) with lumbar spine or femoral neck bone mineral density T-scores no less than −2.5 (mean, −1.2) were enrolled if they did not have vasomotor symptoms requiring treatment, bone diseases (other than osteoporosis), previous vertebral fractures, or endometrial hyperplasia at baseline. Subjects were randomized to take BZA 10, 20, or 40 mg; raloxifene 60 mg; or placebo daily for 2 years. Efficacy assessments (bone mineral density, bone markers, and lipids) are presented in detail elsewhere. Safety was evaluated based on adverse event (AE) reporting, laboratory analyses, and physical examination. Endometrial and ovarian safety were assessed by periodic transvaginal ultrasound and endometrial biopsy. Of 1,583 women (mean age ± standard deviation, 57.6 ± 6.5 years) included in the safety population; 1,113 (70.3%) completed the 2-year study. The rates of treatment-emergent AEs, serious AEs, and discontinuations due to AEs were similar among treatment groups. Vasodilatation was more common with BZA 20 and 40 mg (19.9% and 22.6%, respectively) than with placebo (13.2%; P <0.05 for both), but similar to raloxifene (18.3%). The incidence of leg cramps was similar across treatment groups (range, 9.3%-11.6%). The incidence of venous thrombotic events with BZA was low (<1%) and similar to that with RLX or placebo. No cases of endometrial hyperplasia or malignancy were diagnosed in women treated with BZA. In conclusion, BZA was well tolerated and had a safety profile similar to placebo in a population of relatively young postmenopausal women. BZA is a new promising SERM that could become an important addition to the available therapies for the prevention of postmenopausal osteoporosis.

Disclosures: C.H. Chesnut, Roche 8.

This study received funding from: Wyeth Research.

W388

Methods of Transition from Hormone Therapy to Raloxifene or Other Regimens Administered to Postmenopausal Women for the Prevention or Treatment of Osteoporosis: The HORTHON Study. G. Christodoulakos¹, V. Drossinos*², C. Barker*³, E. Korelis*², G. Kreatsas*¹. the Hellenic HORTHON Study Group*¹. ¹2nd Dpt of Obstetrics & Gynecology, University of Athens, Aretaeion Athens Hospital, Greece. ²Dpt of Medical Research, Pharmaserve-Lilly, Athens, Greece. ³EuMIS Statistics, Eli Lilly & Company, Erl Wood, United Kingdom.

HORTHON is a Hellenic prospective observational study designed to investigate the preferred method of transition from hormone therapy (HT) to Raloxifene or other pharmaceutical agents for the prevention or treatment of osteoporosis.

The study design was non-interventional. Patients were asked to sign an informed consent document before entering the study. Inclusion criteria were a postmenopausal status for at least 2 years and continuous treatment with HT for at least 6 months prior to inclusion. 494 patients were included in the study, enrolled by 39 investigative sites. Mean age (SD) was 52.2 (5.12) years, mean (SD) age at start of menopause was 47.1 (4.94), mean (SD) time since menopause was 5.2 (4.13) and mean (SD) duration of HT was 4.0 (2.85) years. 49.0% had a history of osteoporosis in mother and 7.7% had a family history of breast cancer. 62.2% of enrolled patients were diagnosed with Osteopenia and 14.8% with Osteoporosis.

83.0% of patients received Raloxifene either as monotherapy (Raloxifene only) or combined mainly with calcium and vitamin D (Raloxifene plus Other), 11.6% received other treatment (Other Therapy) while 5.3% were not administered any therapy, as chosen in the course of normal clinical practice. The primary reason of choice was safety in the “Raloxifene only” (52.8%) and “Raloxifene plus other” (41.0%), while in the “Other therapies” was efficacy (34.5%).

Transition methods included “no wash out and no taper”, “wash out only”, “wash out and taper” and “taper only” (45.2%, 45.2%, 7.6% and 1.9% of patients respectively). For “Raloxifene only” group, “no wash out and no taper” method applied to 55.9% of patients. “For Raloxifene plus Other” and “Other” new therapies “wash out only” method was chosen for the majority of patients (46.0% and 49.1% respectively). Mean (SD) satisfaction from transition method (taper and/or wash out), estimated by a visual analogue scale, was 32.9 (19.30).

The main reason for discontinuation of HT was prolonged use (42.3% of patients). Mean (SD) satisfaction to previous HT treatment was 30.8 (20.16) while for patients who received no new therapy, satisfaction (SD) was 22.7 (15.48). In this study, patients who discontinue HT, mainly for prolonged use, were more likely to transition to Raloxifene rather than another pharmaceutical agent. Safety is the major reason for choosing the new therapy. The most frequent transition methods were “no wash out and no taper” and “wash out only”.

Disclosures: G. Christodoulakos, None.

This study received funding from: Pharmaserve — Lilly S.A.C.I.

W389

Possible Role of Equol Status in the Effects of Isoflavone on Bone and Fat Mass in Postmenopausal Japanese Women: A Double-Blind Randomized Controlled Trial. Y. Ishimi¹, J. Oka*², S. Uchiyama*³, T. Ueno*³, T. Toda*⁴, M. Uehara*⁵, J. Ezaki*¹, J. Wu*⁶. ¹Nutritional Epidemiology Program, National Institute of Health and Nutrition, Tokyo, Japan. ²Home Economy, Tokyo Kasei University, Tokyo, Japan. ³Saga Nutraceuticals Research Institute, Otsuka Pharmaceutidcal Co. Ltd., Saga, Japan. ⁴Fuijcco Co. Ltd., Kobe, Japan. ⁵Nutritional Science, Tokyo University of Agriculture, Tokyo, Japan. ⁶Central Research Laboratory, Nisshin OilliO Group, Ltd., Yokosuka, Japan.

Introduction: Equol is more biologically active than its precursor daidzein, which is the principal isoflavone found in soybean. There are inter-individual differences in the ability to produce equol; this may lead to differences in the effects of isoflavone intervention on human health. This study aims to investigate whether the effects of soy isoflavones on bone and fat mass are related to an individual's equol status.

Methods: We performed a 1 -year double-blind randomized trial to compare the effects of isoflavone (75 mg of isoflavone conjugates/day; equivalent to 47 mg/day of aglycone form) with those of placebo on bone mineral density (BMD), fat mass, and serum isoflavone concentrations in early postmenopausal Japanese women who were classified based on their equol-producer phenotype.

Results: After 1-year, the isoflavone intervention significantly increased serum equol concentration in the equol producers but not in the nonproducers. In the isoflavone group, the annualized changes in the BMD of the total hip and inter-trochanteric regions were −0.46% and −0.04%, respectively, in the equol producers and −2.28% and −2.61%, respectively, in the nonproducers; these values were significantly different (p < 0.05 for both the regions). Significant differences were observed between the equol producers and nonproducers in the isoflavone group with regard to the annualized changes in the fat mass. No significant difference in the annualized changes in the BMD and fat mass was observed between the equol producers and nonproducers in the placebo group.

Conclusions: Our data suggest that the preventive effects of isoflavones on bone loss and fat accumulation in early postmenopausal women depend on an individual's equol-producing capacity.

Disclosures: Y. Ishimi, None.

This study received funding from: Japan Health Science Foundation.

W390

Success and Complication Rate of Percutaneous Vertebroplasty (PVP) in Patients with Osteoporosis (OP). M. Gaugg*¹, F. Karnel*², P. Breitner*², W. Kumpan*², B. Kutil*¹, L. Erlacher*¹. ¹Internal Medicine, Division of Rheumatology and Osteology, SMZ Süd, Vienna, Austria. ²Radiology, SMZ Süd, Vienna, Austria.

Background: Percutaneous vertebroplasty (PVP) is an evidence-based, minimally-invasive procedure to manage pain and vertebral spine deformation after vertebral compression fracture (VCF) in patients with OP^1,2 Objective: To describe success and complication rates in patients affected by VCF and treated by PVP in specialized clinical care, to evaluate their clinical and treatment characteristics. Materials and Methods: Data were collected retrospectively by review of examination reports. PVP was performed by two interventionists. CT fluoroscopy guided PVP with additional guidance by conventional C-arm fluoroscopy was performed as described previously^3,4, between October 2003 and February 2007. Results: A total of n= 133 procedures were available for analysis. PVP was performed in n=85 patients [(71 females/14 males, mean age 75,5 +/- 9.5 a; (53-93a)]. Post-interventional cement leakage was found in 44/85 patients (51.7%) or 50/133 vertebrae (37,6%). The location of the leakage was distributed as follows: 9,7% (n=13) to basivertebral veins, 5 % (n=7) to paravertebral structures including paravertebral veins, 15 % (n=20) to adjacent vertebral disc (upper or lower end plate), and 7% (n=10) to epidural venous system. All PVP's were completed successfully with conscious sedation and local anesthesia. During treatment of 133 vertebrae (TH4-L5), no clinically evident neurologic or embolic events occurred. There was no procedure related mortality. Conclusion: PVP can be performed safely and effectively in patients with OP and VCF. Post-interventional cement leakage is found frequently by combination of CT and conventional flouroscopy. However, clinically relevant neurologic and/or thromboembolic complications may be rare with careful patient selection by clinicians and appropriate radiologic imaging.

References:

1) Interventional Techniques: Evidence-based Practice Guidelines in the Management of Chronic Spinal Pain. Mark V. Boswell, et al. Pain Physician 2007 Jan;10(1):7-111.

2) Balloon kyphoplasty and vertebroplasty for vertebral compression fractures: a comparative systematic review of efficacy and safety. RS Taylor et al. Spine 2006 Nov 1;31(23):2747-55

3) Percutaneous vertebroplasty guided by a combination of CT and fluoroscopy. A Gangi, et al. AJNR Am J Neuroradiol. 1994 Jan; 15(1): 83-6

4) Evaluation of percutaneous vertebroplasty in osteoporotic vertebral fractures using a combination of CT fluoroscopy and conventional lateral fluoroscopy. MB Pitton et al. Rofo. 2004 Jul; 176(7): 1005-12

Disclosures: M. Gaugg, None.

W391

Health-related Quality Of Life 7 Years After Hip Or Vertebral Fractures. I. Hallberg¹, M. Bachrach-Lindström*¹, G. Toss², A. C. Ek*¹. ¹Dept of Medicine and Care, Div of Nursing Science, Faculty of Health Sciences, Linköping University, SE-581 85 Linköping, Sweden. ²Dept of Medicine and Care, Div of Internal medicine, Faculty of Health Sciences, Linköping University, SE-581 85 Linköping, Sweden.

In a previous paper we reported that women with vertebral fracture had severe reduction of health-related quality of life two years after a fracture. In order to estimate the long-term impact of osteoporotic fractures on HRQOL we examined 67 women, 7 years after a hip or vertebral fracture. Patients were otherwise attending routine primary health care. HRQOL was evaluated by the SF-36 questionnaire and compared with a local age-matched reference material (n=804). High scores in SF-36 indicate better HRQOL. Bone mineral density (BMD) was measured with Hologic 4500A. Physical function was assessed by handgrip strength (Jamar dynamometer) and static balance on dominant leg with eyes open. The number of the vertebral fractures was evaluated by lateral radiographs of the spine and calculation of spinal deformity index (SDI, range 0-39), according to the Genant visual semiquantitative criteria.

At the 7 year investigation 51 women had one or more vertebral fractures (9 of which also had had a hip fracture). Mean SDI was 5.7 (range 1-25). Of the women with a hip fracture 16 were without vertebral fractures. Mean age (SD) was in vertebral fracture group 76.0 (4.7) and in hip fracture group 73.6 (4.2) years (p=0.07). Bisphosphonate treatment was used by 23 of 67 women (17/51 and 6/16 respectively, n.s.).

HRQOL (SF-36) was reduced in the vertebral fracture group regarding physical function (p=0.005), role-physical (p=<0.001), bodily pain (p=<0.001), role-emotional (p=0.046), vitality and social function (p=0.06). Women with hip fracture (n=16) but no vertebral fracture did not differ from the reference material.

In the total fracture group (n=67) static balance showed a significant positive correlation (p<0.05) to SF-36 regarding all 8 domains (range r_s=0.268-0.549). Also handgrip strength correlated in all domains except role-emotional (range r_s=0.261-0.387). SDI showed negative correlation to all domains except physical function, general health and mental health (range r_s=-0.269-0.346). Age showed negative correlation to vitality and mental health (range r_s=0.246-0.292). BMD in hip or lumbar spine did not correlate with any domain in SF-36.

In conclusion women with vertebral fractures still had pronounced reduction of most HRQOL domains 7 years after fracture and treatment in routine health care. The long-term reduction of HRQOL and its correlation to static balance and handgrip strength raise questions for further investigation.

Disclosures: I. Hallberg, None.

W392

Importance of Balloon Kyphoplastiy in the Management of Vertebral Fractures. J. Hamon*, P. Longis*. Orthopedie, Hospital, Saint-Nazaire, France.

Objectives: To show the importance of balloon kyphoplasty, in the management of vertebral compression fractures of the elderly and vertebral fractures secondary to cancer. This minimally invasive procedure is designed to reduce the fracture, correct the deformity, stabilize the lesion by injection of cement, decrease the pain, limit the hospitalization time and improve the patients' quality of life.

Methods: Retrospective study from Dec 2005 to Apr 2007. All patients older than 40 years and presenting with a vertebral compression fracture or a metastatic vertebral fracture, localized between T6 and S4 without neurological complication, were included. The procedure is performed under general anesthesia and is characterized by controlled inflation of 2 small balloons under fluoroscopic control in order to correct the vertebral deformity before injection of the cement. Results: 44 patients, 26 women and 18 men were included in the study, 51 vertebrae were treated. The average age is 64 years. 35 vertebral compression fractures (from T7 to L4) and 16 metastatic vertebral lesions (from T6 to S4) were treated (2 prostate, 3 pulmonary, 3 hematologic, 5 gastro-intestinal, 1 gynecologic and 2 neuro-endocrine primary cancer). For traumatic fractures, mean fracture age is 67 days. Pre-operative use of analgesics, according to the classification of the W.H.O, is of level 1 in 8 patients, level 2 in 11 patients and level 3 in 25 patients. The mean cement volume injected per vertebra is 5 cc. For the trauma lesions, height restoration of the anterior vertebral body wall is on average 15% lost height restored and the correction of the local kyphosis is 5°. Cement leakages in the adjacent discs are found in 9 patients without any clinical consequences. No post-operative complications were seen. Post-operative mobilization is on average on day 1. Hospital discharge is on average on day 2. Post-operatively prescribed analgesics, according to the classification of the W.H.O, are of level 1 in 29 patients, level 2 in 9 patients and level 3 in 6 patients. 38 patients returned home, 6 patients needed revalidation (1 because of associated traumatic lesions and 5 patients needed specialized care for their primary tumor).

Discussion: The usual treatments of this type of vertebral lesions as well as their frequent adverse events are known. The interest of Kyphoplasty is thus highlighted. The review of the literature also highlights the advantages of this technique compared to vertebroplasty Conclusion: Balloon kyphoplasty is an alternative with reduced surgery time, good safety profile, early mobilization and without need for bracing. Balloon kyphoplasty allows reduction in analgesics consumption, an early hospital discharge and improvement of the patients' quality of life

Disclosures: J. Hamon, None.

W393

Significant Reduction of Vertebral Fractures: Comparison of Rehabilitation of Osteoporosis Program-Exercise (ROPE) versus No-ROPE, With or Without Pharmacotherapy. D. A. Kurmen Figueroa*, M. Sinaki. Physical Medicne and Rehabilitation, Mayo Clinic, Rochester, MN, USA.

The objective of this study was to determine whether patients with osteoporosis who received the Rehabilitation of Osteoporosis Program-Exercise (ROPE) had reduced risk of vertebral fracture as compared with those who did not (No-ROPE). To avoid bias in choice of treatment, none of the subjects or evaluators had knowledge of group assignment.

We reviewed the medical records of 200 patients. One hundred and twelve patients with osteoporosis met the inclusion criteria, i.e. women older that age 40 with the diagnosis of osteoporosis, baseline and follow-up radiographs of spine. Patients with steroid-induced bone loss and active malignancy were excluded. Two groups were formed: 71 subjects in the ROPE group and 41 subjects in the No-ROPE group met the enrollment criteria. Follow up was the date of follow up x-rays or recurrence of fracture/back pain. The duration of follow up ranged from 3 months to 118 months with an average of 45.5 months. The results showed that fifty-six of the 71 ROPE subjects (78.9%) and 20 of the 41 No-ROPE subjects (48.8%) had no new fracture at the time of final follow up. Interestingly, 56 subjects (78.9%) of the 71 ROPE subjects had received pharmacotherapy whereas all 41 (100%) of the No-ROPE subjects had received pharmacotherapy for bone loss.

The 79% of the ROPE population without new fracture were distributed as: 29.6% combination therapy, 26.8% bisphosphonates only, 9.9% calcium only, and 12.7 % no pharmacotherapy. The 49% of the No- ROPE population without new fracture were distributed as: 26.8% combination therapy, 17.1% bisphosphonates only, and 4.9% calcium only.

In the population taking some kind of pharmacotherapy, 21.1% had new fractures during the time to follow up in the ROPE group versus 51.2% in the No-ROPE group.

The No-ROPE group was four times more susceptible to fracture during the time to follow up (P = 0.001), Pearson Chi Sq test. The group with the lowest incidence of fractures was the ROPE group with combination pharmacotherapy.

To prevent vertebral osteoporosis fractures, we enthusiastically suggest that ROPE be mandatory for management of every woman with osteoporosis, regardless of choice of pharmacotherapy.

Disclosures: D.A. Kurmen Figueroa, None.

W394

Relationship Between Serum 25-hydroxyvitamin D₃ Concentration and Walking Ability, Leg Strength, or Balance in Community-Dwelling Japanese Frail Elderlies. J. Okuno*, S. Tomura*, H. Yanagi*, N. Yabushita*, T. Okura*, K. Tanaka*. Graduate school of Comprehensive Human Sciences, University of Tsukuba, Tsukuba City, Japan.

The aim of this study was to evaluate the relationship between serum 25-hydroxyvitamin D₃ (25OHD) concentration and walking ability, muscle strength, or balance in community-dwelling Japanese frail elderlies. The study was a longitudinal study conducted in a town near Tsukuba city (latitude 36°north) from June to September, in 2005 and 2006. Forty-five participants were community dwelling elderlies aged 65 years and over who required minimum support or nursing care to maintain their ADL. Their mean age was 76.5±5.9 years (mean±SD, range: 65-90). Of the participants, 32 attended a 3-month exercise program for nursing care prevention once per week (exercise group). The remaining 13 received only advice for nursing care prevention (control group). The Ethics Committee of University of Tsukuba approved the study. An interview was conducted based on a questionnaire including score of functional capacity of ADL (Tokyo Metropolitan Institute of Gerontology (TMIG) score), experiences of fall, stumbling and body sway during the past one year, walking ability, and the frequency of going outside of the home. The serum levels of 25OHD, intact parathyroid hormone (iPTH) and calcium were measured. The following physical tests were performed at baseline and 3 months: Timed Up & Go (TUG) and a 5-meter walk for walking ability, functional reach and foot balance with open eyes for balance, trunk flexion for flexibility, and ankle strength and grip strength for muscle strength. Among 45 patients who underwent the tests, 66.7% and 40% had difficulties in standing up and in walking, respectively. 54.3% experienced falls, 73.8% experienced stumbling and body sway more than once during the past one year. The mean level of 25OHD was 62.1±13.7 nmol/L (mean±SD, range: 27.5-87.5). At baseline, the rate of 25OHD levels less than 50 nmol/L that is the cut-off point of 25OHD resulting from the significant difference of intact PTH level was 18.0%. At baseline and 3 months there were no significant differences in physical tests between the control group and the exercise group. The subjects with 25OHD levels more than 50 nmol/L at baseline improved their walking ability and ankle strength for a period of three months in both groups. On the other hand, the subjects with 25OHD levels less than 50 nmol/L did not improve their physical performance even in the exercise group. It is suggested that serum 25OHD level is related to walking ability and muscle strength in Japanese frail elderlies, and that serum 25OHD level more than 50 nmol/L may be needed for maintaining their mobility or balance.

Disclosures: J. Okuno, None.

W395

Treatment of Osteoporotic Vertebral Body Fractures by Means of Percutaneous Balloon Kyphoplasty. Long Term Results of a Prospective, Clinical Trial. T. R. Blattert*. Trauma & Reconstructive Surgery, Leipzig University, Leipzig, Germany.

Introduction: Balloon kyphoplasty is a minimally-invasive, percutaneous surgical technique for reduction and stabilization of osteoporotic vertebral body fractures. However, there is no prospective, clinical trial on long term results concerning the safety and efficacy of the method so far.

Patients and Methods: This prospective, clinical trial investigated both safety and efficacy of percutaneous Balloon kyphoplasty. All vertebrae were stabilized with Polymethylmethacrylate (PMMA). Pre- and postoperatively, the following data were acquired: subjective rating of pain (Visual Analog Scale, VAS), bisegmental endplate-angle (EA2), anterior and posterior height of vertebra. Inclusion criteria were osteoporotic fractures of vertebral bodies in the thoracolumbar spine, patient age ≥ 65 years, fracture age ≤ 4 months, and t-score ≤ −2.5 (DEXA). Exclusion criteria were tumor lesions and additional posterior instrumentation.

Results: 352 vertebrae of 314 patients suffering from acute pain could be included with a minimum-follow up of two years. The average patient age was 74 years (57-92). Average t-score was −2.7. (-3.1 to −2.5). 262 patients suffered from pain for three weeks on average, whereas 52 patients were not able to recall the onset of pain. Fractures were only localized within the thoracolumbar spine with only A type of injuries occuring. 309 of 314 patients experienced marked pain-relief as expressed on the VAS (2.1 ± 1.9 to 8.2 ± 1.5; 0 “worst” to 10 “best”). Average correction of EA2 was 6.2°. The anterior vertebral height could be restored by 7.4 mm on average, posterior height by 3.0 mm. There were no neurological complications. In 32 (9.1%) vertebrae, we saw intraoperative leakage of cement (6 (1.7%) out of these with epidural leakage), however, no clinical consequences had to be noted. There were 6 (1.7%) cases of intraoperative ballon-perforation, and 11 (3.1%) cases of subsequent vertebral body fractures in the adjacent level.

Conclusion: Balloon-kyphoplasty is an efficient, and minimally-invasive therapeutic option for the treatment of painful vertebral body fractures having its focus on cases with underlying osteoporosis. Depending on the age and type of fracture, this treatment can obtain a significant rate of fracture reduction that is being maintained over the course of two years minimum. Compared to current literature on vertebroplasty, this technique presents fewer leakage complications at equal long term success in reducing pain.

Disclosures: T.R. Blattert, Kyphon Europe 5.

W396

Life Quality in Patients with Surgically Treated Vertebral Stenosis and the Relation with Bone Quality. I. D. A. Nemes*¹, D. V. Poenaru*², H. Vermesan*², R. Preibeanu*², D. Vermesan*², I. Branea*², M. Dragoi*¹, O. Bereteu*¹, R. Onofrei*¹, E. Amaricai*¹, D. Popa*¹, I. Ilia*¹, C. Nemes*³. ¹Timisoara City Universitary and Emergency Hospital — Rehabilitation and Rheumatology Department, Timisoara “Victor Babes” University of Medicine and Pharmacology, Timisoara, Romania. ²Timis County Universitary and Emergency Hospital — Orthopaedic and Traumatology Department, Timisoara “Victor Babes” University of Medicine and Pharmacology, Timisoara, Romania. ³Medical Lab, Timisoara City Universitary and Emergency Hospital, Timisoara, Romania.

Purpose: To establish the ratio of real vertebral stenosis versus those suppose to be and to assess life quality of patients with surgically treated vertebral stenosis and to the relation with bone quality.

Methods: In 3 years period from 380 patients considered to have vertebral stenosis, only 130 had the real disease. They were divided in 3 therapy groups: 25 patients with mild to moderate symptoms and a conservative treatment; 46 patients with more severe symptoms and surgically treated with decompressive laminectomy ± chyphovertebroplasty; 59 patients with severe symptoms and surgically treated with decompressive laminectomy + instrumented fusion.

All patients benefit from diet, pain management, antiosteoporotic drugs in group II and III and physical therapy (local analgetic electrotherapy, biofeed-back training of the local muscle and Williams programme in group I/ biofeed-back training of the local muscle and Florida Spine Institute post-operative protocol in group II and III). All patients were evaluated, regarding their life quality, according to WOMAC scale at baseline, day 21 and at 3 month and DXA bone densitometry and lab parameters of bone metabolism were performed at 40 patients of group III.

Results: In group I and II the three months evaluation showed a clinical and functional improvement. The clinical and functional status of the third group patients was significantly improved at 3 month assessment. A “slow” evolution was seen at 40 patients after six months, despite the fact that DXA scores and lab parameters had an improvement.

Conclusion: The ratio of vertebral stenosis/ vertebral hernia is almost 1/3 in our area. If the conservative therapy applied in moderate forms has no significant result in 3 month we should consider the surgical therapy as the appropriate therapy. As a result of the protocol's applying a better life quality, related with DXA scores and lab parameters, was seen. The difference in outcomes between the post surgical evaluations in group III may be due to the type or intensity of the rehabilitation programme followed after the hospitalization period or cultural issues, patients having greater expectation from the surgical treatment than this is meant to realise.

Disclosures: I.D.A. Nemes, None.

W397

Adjacent Fractures in the Thoracic and Lumbar Spine after the Treatment with Balloon-Kyphoplasty −2 Years Prospective Follow-up. R. Pflugmacher*. Centrum für Muskuloskeletale Chirurgie, Charité — Universitätsmedizin Berlin, Berlin, Germany.

Purpose: To evaluate the long-term outcomes of 57 patients with 87 osteoporotic vertebral fractures, located in the thoracic and lumbar spine, treated with Balloon Kyphoplasty.

Material and Methods: 63 patients (20 males and 43 females) with 96 osteoporotic vertebral fractures were treated with Balloon Kyphoplasty. We were able to have a 2 year follow up in 57 patients with 87 vertebrae treated. Symptomatic levels were identified by correlating the clinical presentation with conventional radiographs, CT and / or MRI. During the 2 year follow-up reduction in pain was determined. The effects on pain symptoms were measured on a self-reported Visual analog Scale (VAS) and the Oswestry score was documented to assess disability. Radiographic scans were performed pre- and postoperatively and after 3,6, 12 and 24 months. The vertebral height and kyphosis angle were measured to assess the restoration of the sagittal alignment.

Results: The median pain scores (VAS) improved significantly from pre- to post-treatment as did the Oswestry Disability Score (p<0.001). This improvement was maintained at 2 year follow up. In 9 patients (15.8%) (6 female, 3 male) an adjacent fracture occurred in 12 vertebrae (13.8 %) within 2 weeks to 22 months follow-up. In four patients the adjacent fractures were asymptomatic. Five patients with symptomatic adjacent fractures were treated again with Balloon Kyphoplasty. Clinically asymptomatic cement leakage occurred in 12 of 96 vertebral bodies (12.5 %). During 2 year follow-up this surgical technique demonstrated restoration and stabilization of the height of the vertebral body.

Conclusion: Balloon Kyphoplasty is an effective minimal invasive procedure for the stabilization of osteoporotic vertebral fractures leading to a statistically significant reduction of pain status.

Disclosures: R. Pflugmacher, None.

W398

Improving Trunk Strength and Endurance in Older Women with Vertebral Fractures. K. M. Shipp, D. T. Gold, C. F. Pieper*, K. W. Lyles. Duke University, Durham, NC, USA.

The trunk musculature is important for people with osteoporosis. First, strong, fatigue-resistant trunk muscles are necessary for erect postural alignment. Second, strong trunk extensors may prevent vertebral fractures. An observational study associated trunk extension strengthening over 2 years with decreased incidence of vertebral fractures in the next 8 years [Sinaki M et al. Bone 2002;836-841]. The relationships between trunk alignment, strength, and endurance; functional status; and fracture risk are complex and not yet fully understood.

Results will be presented from a randomized clinical trial of a group exercise intervention (3X/week for 6 months) in older women (n=122, mean age 81 years) with vertebral fractures (mean # 2.3) [Gold DT et al. J Am Geriatr Soc 2004;52:1471-1478]. Randomization was by site: 3 trunk exercise intervention sites (n=53) and 3 control sites (n=69). The trial was a modified cross-over design: the control sites received the exercise intervention after 6 months of health education sessions. The exercises included 1) stretches to increase trunk extension flexibility and 2) progressive resistive strengthening exercises for trunk extensors, abdominals, and posterior scapular groups.

Using mixed-model ANCOVA (intention-to-treat), we found a significant difference between groups for trunk extension strength at 6 months (p=0.0001; +6.6 ft-lbs. [up 24%] exercisers, −4.6 ft-lbs. controls). When the control subjects received the exercise intervention, there was a mean increase of 15.0 ft-lbs [up 45%].

An ancillary study examined the effect of the intervention on Timed Loaded Standing (TLS), a measure of combined trunk and arm endurance [Shipp KM et al. Osteoporos Int 2000;11:914-922]. Using mixed-model ANCOVA (intention-to-treat), we found a trend for difference between groups for TLS time at 6 months (p=0.0542; +8.8 sec exercisers, −8.9 sec controls). Among compliers (those attending 2/3s of sessions), group was significant (p=0.035; +14.9 sec exercisers, −10.3 sec controls). Change in TLS time, but not change in trunk extensor strength, over 6 months was significantly correlated (p<0.05) with change in psychological symptoms and functional status.

These results demonstrated that trunk extension strength and combined trunk and arm endurance could be increased, in older women with vertebral fractures, by 6 months of group, physical therapist-led, trunk-specific exercise classes. Improved trunk endurance was associated with fewer psychological symptoms and better functional status.

Disclosures: K.M. Shipp. None.

This study received funding from: NIH (NIA and NICHD).

W399

Peak Vertical Impact Forces During Backward Falls onto the Buttocks and the Influence of Compliant Flooring. M. M. Sran, S. N. Robinovitch*. Simon Fraser University, Burnaby, BC, Canada.

Fall-related vertebral fractures are common and backward falls result in impact to the buttocks and pelvis. Compliant flooring is a promising technique for reducing impact force and risk for vertebral fracture during a fall. However, we have little knowledge of the peak forces applied to the body during a backward fall, or how floor stiffness affects this force. Our goal was to measure the peak vertical force applied to the buttocks in a backward fall from standing, and to determine whether this force is lowered by reductions in floor stiffness. Participants included 11 males mean age 25 ± 5(SD) yrs and body mass mean 81 ± 16 (SD) kg. A tether and electromagnet suddenly released the participant from a backward lean of 15°, causing him to fall backward onto the ground, which was covered with a layer of ethylene-vinyl acetate (EVA) foam rubber. We conducted 5 trials for each of 3 foam thicknesses [4.5 cm (Firm), 7.5 cm (Medium), and 10.5 cm (Soft)]. Participants were instructed to avoid contacting the ground with their hands until their buttocks had contacted the ground, not to squat or step. We measured peak vertical impact forces applied to the buttocks at 960 Hz with a force plate (model 4060H, Bertec Corp.). An 8-camera, 240 Hz motion measurement system (EVaRT 4.6, Motion Analysis Corp.) was used to track peak velocity (m/s) of a skin surface marker on the sacrum. We also modeled peak vertical force for falls onto a rigid (bare) floor. We used repeated measures ANOVA and post-hoc t-tests to compare peak forces between the 3 conditions (p = 0.016). There was a significant difference in peak normalized vertical force (N/kg) between falls onto the Soft foam condition compared with the Medium (p= 0.002) and Firm (p< 0.001) conditions. Peak normalized vertical force (N/kg) was (mean ± SD) 63.6 ± 6.2, 59.9 ± 6.2, and 56.9 ± 5.9 for Firm, Medium, and Soft respectively. Estimated peak normalized vertical force (N/ kg) for the rigid (bare floor) condition was 75.3 ± 7.6. In comparison to the rigid floor, falling onto the Firm, Medium and Soft floors provided, on average, 15, 20 and 24% force attenuation. This novel data improves our understanding of this mechanism of vertebral injury and is essential if we are to design techniques for the prevention of vertebral fractures, such as protective equipment and safe movement environments. Peak vertical forces were 5099 N ± 868(SD) for the thinnest foam condition. Studies report the compressive force required to cause fracture of elderly lumbar vertebrae to be between (mean ± SD) 3009 ±1505¹ and 6910 ± 2480 N², similar to the peak loads measured in this study. A thin (4.5cm) layer of foam overlying the floor can provide 15% force attenuation during a fall onto the buttocks. ¹Burklein D et al. J Biomech 2001 ²Cheng XG et al. J Bone Miner Res 1998.

Disclosures: M.M. Sran, None.

W400

A Community-Based Program to Minimize Osteoporosis and Fractures Risks. S. J. Wimalawansa¹, M. Bartello*², K. Morgan*³, M. Wagner*⁴, K. Hodapp*⁵, S. Lachenmayr*⁶. ¹Medicine, UMDNJ-RWJMS, New Brunswick, NJ, USA. ²Memorial Hospital, Morristown, NJ, USA. ³Rutgers University, New Brunswick, NJ, USA. ⁴Rutgers University, Milltown, NJ, USA. ⁵Morristown Memorial Hospital, Morristown, NJ, USA. ⁶NJ Dept. of Health, Trenton, NJ, USA.

In New Jersey (NJ), ∼1 million residents have low bone mass and ∼8,000 individuals over 65 fractures a hip annually. The NJ Interagency Council on Osteoporosis (ICO) oversees osteoporosis (OP) education & prevention initiatives in the state. Since 1997, Project Healthy Bones, a 24-week, comprehensive, peer-led OP exercise & education program provides a public health approach to healthy behaviors improving BMD, dietary calcium intake, balance & strength, and minimize falls and associated fracture risk.

In 2004, reduced funding resulted in creating new training at local level for community class leaders, establish program sites for local program demand, provision of resources / technical assistance and provide program oversight. This model utilizes regional, trainers-training program with community-based partnerships. ICO provides 2-day training program in volunteer Peer Leader training, evaluation protocols and oversight accountability.

Our data demonstrated improved balance, increased strength & higher bone density through this program. The impact of Project Healthy Bones was evaluated through exercise tracking forms, pre-post- knowledge tests and food diaries. With the new program, 90% of participants increased weight lifting and exercise tolerance, and 68% increased their calcium intake (∼500 mg per day).

From March 05 to December 06, a network of 82 agency staff and 318 volunteer peer leaders provided > two-hundred 24-week programs to nearly 2,000 participants in every county in NJ (over 15,000 volunteer hour contributions). This is in addition to continuing of 80% regular classes beyond the initial 24-weeks. A statewide ICO listserv also provides a forum answers questions/challenges, disseminates current research and OP news, and coordinates local program offerings. Many physicians and HMOs are referring patients directly to the program based on improved DXA results (pre- and post-class) and decrease falls.

The Project Healthy Bones maintain existing community programs; create new programs based on local demand; and oversight to assure program integrity. The program is on CD-ROM, which is designed to use locally by peer leaders. Program demonstrated improve balance and nutrition; prevent falls, increased strength, morale and social well-being among participants. Despite lack of state funding, the modeled program has been strengthened and expanded with extensive community partnerships, fostered volunteerism, and has shown a positive impact in the health and quality of individuals' lives.

Disclosures: S.J. Wimalawansa, None.

W401

Overexpression of Osteoblast IGF-I Blunts the Deleterious Effects of lOw Protein Intake on Bone Strength. P. Ammann¹, B. Kream², C. Rosen³, R. Rizzoli¹. ¹Department of Rehabilitation and Geriatrics, Division of Bone Diseases, Geneva, Switzerland. ²University of Connecticut Health Center, Farmington, CT, USA. ³St Josephs Hospital Maine Center for Osteoporosis Rsearch, Bangor, ME, USA.

Isocaloric low protein intake decreases bone mass and bone strength. These alterations are associated with decreased circulating IGF-I levels. Osteoblast IGF-I expression is decreased by low amino-acids concentration. Whether circulating and/or bone locally produced IGF-I are responsible for low protein diet-induced bone damages is not established yet. We investigated 6-month adult transgenic male mice overexpressing IGF-I in osteoblasts under the control of collagene type-1 promoter (TG-IGF) and wild type mice (WT), fed a normal or an isocaloric low protein diet, for 8 weeks. After sacrifice, tibia were tested for biomechanics, size, microstructure using microcomputerized tomography, cortical histomorphometry and bony tissue nano-indentation. Blood was also collected. In WT on low protein diet, compression strength was significantly decreased and resistance to bending displays similar trend, whereas they were not changed in TG-IGF. Data in the table show a cortex thinning, and also alterations of intrinsic bone tissue quality in WT but not in TG-IGF. Outer bone diameter was higher in TG-IGF-I, irrespective of the protein intake. Endosteal BFR was reduced in WT on a low protein diet, but not in TG-IGF. Trabecular bone mass was significantly decreased in WT only. Plasma IGF-I was similar in WT and TG-IGF, and equally decreased by the low protein diet. These results in adult male mice indicate that bone local overexpression of IGF-I blunts the deleterious effects of a low protein diet, even in the presence of lower IGF-I circulating levels. 

Table  .  

Disclosures: P. Ammann, None.

This study received funding from: Swiss National Research Fondation.

W402

Sub-Optimal 25(OH)D₃ Levels Causes Increased Osteoclastogenesis and Bone Loss. P. H. Anderson*¹, R. K. Sawyer*¹, A. J. More*¹, A. Muliabrahimovic*¹, B. K. May*¹, P. D. O'Loughlin*¹, H. A. Morris². ¹Endocrine Bone Research, Institute of Medical and Veterinary Science, Adelaide, Australia. ²Hanson Institute, Institute of Medical and Veterinary Science, Adelaide, Australia.

The association between increased risk of hip fracture and a low vitamin D status has long been recognised. However, the level of circulating vitamin D required to maintain bone strength is controversial. To determine the circulating 25-hydroxyvitamin D (25D) level required for the maintenance of bone, we generated 6 groups of Sprague-Dawley rats with stable, mean 25D level ranging from 11 nmol/L to 118 nmol/L. Vitamin D depleted animals were fed various levels of vitamin D (Ong-500ng/d) with 0.4% Ca for 4 months before being killed at 7 months of age for serum biochemistry and bone histomorphometry analyses. Proximal femora structure was analysed by μCT and distal femora histomorphometry was analysed using sagittal 5 μm undecalcified sections. Fasting serum calcium, PTH and phosphate levels were normal in animals with low 25D levels. Evidence of osteomalacia was seen only in animals with 11(±0.7) nmol/L 25D, with elevated serum ALP and cross-laps, increased osteoid surface and delayed osteoid maturation time (OMT) compared to animals fed higher levels of vitamin D. OMT was constant in animals fed between 50 and 500ng/d vitamin D and trabecular bone volume (BV/TV) was positively related to circulating 25D (R²=0.51, P<0.001), but not to 1,25(OH)₂D₃ levels, nor PTH levels. Animals fed between 50 and 100ng/d vitamin D (mean 25D <80nmol/L) had lower BV/TV (Fig 1) and both osteoid surface and osteoclast surface (Fig 2) were elevated compared to animals fed higher vitamin D. In summary, while osteomalacia occurs at extremely low 25D levels, less severe depletion of vitamin D was associated with increased osteoclast numbers, and significant bone loss. The molecular mechanism for the changes in cellular activity due to vitamin D insufficiency are currently under investigation. In conclusion, for animals fed adequate dietary Ca, a circulating 25D above 80nmol/L is required to prevent bone loss. ,  

Figure Figure 1. Trabecular bone volume with changes in vitamin D intake

Figure Figure 2. Osteoclast Numbers with changes in vitamin D intake

Disclosures: P.H. Anderson, None.

This study received funding from: NH&MRC of Australia.

W403

A More Alkaline Diet May Enhance the Favorable Impact of Dietary Protein on Lean Tissue Mass in Older Adults. B. Dawson-Hughes, S. S. Harris*, L. Ceglia*. Bone Metabolism Laboratory, The Jean Mayer Human Nutrition Research Center on Aging at Tufts University, Boston, MA, USA.

Maintaining muscle mass in aging is important to prevent falls and fractures. Dietary protein is required to preserve muscle mass, however the acid load from diets rich in acidogenic protein foods and cereal grains relative to alkalinogenic fruits and vegetables may contribute to loss of lean tissue mass in older adults.

This analysis was conducted to determine whether for a given protein intake, a more alkaline diet has a more favorable effect on lean body mass (LBM). Subjects were 384 men and women age 65+ who participated in a 3-yr calcium and vitamin D versus placebo trial. All subjects gave written informed consent.

Urinary nitrogen (N), an indicator of protein intake, and potassium (K) were measured at baseline in 24-hr collections. N was measured by the Dumas combustion method and K by direct-current emission spectroscopy. The N:K ratio, reflecting the ratio of acidic protein foods to alkaline fruit/vegetable intake, was calculated. LBM, defined as total body non-fat, non-bone tissue mass, was measured by DXA at baseline and 3 years. Physical activity, measured by Physical Activity Score for the Elderly (PASE) questionnaire, height, and weight were assessed at baseline and 3 years.

At baseline, the mean N:K was 5.98 ± 1.70 (SD) mmol/mmol. The N:K was inversely associated with baseline LBM (regression coefficient = −0.245, P = 0.011, adjusted for sex, age, weight, physical activity score, and nitrogen excretion) and with 3-yr change in LBM (regression coefficient −0.105, P = 0.046, adjusted in addition for treatment group and baseline LBM).

These results suggest that the effect of protein on muscle may be enhanced when the overall diet is less acidic, that is, higher in fruit and vegetable content.

Disclosures: B. Dawson-Hughes, None.

This study received funding from: NIH.

W404

Propensity to Accumulate Bone Microdamages Is Increased in Adult Female Rats Fed an Isocaloric Low Protein Diet. V. Dubois-Ferrière*, R. Rizzoli, P. Ammann. Division of Bone Diseases, Geneva, Switzerland.

Low protein intake decreases bone strength through a change in bone mass, bone microstructure and intrinsic bony tissue quality. Whether the low protein diet-induced alterations of bony tissue could favor the accumulation of bone microdamages is not known. We investigated the effects of repeated loading on humerus bone strength in 6-month-old female rats pair-fed a control (15% casein, n=10) or an isocaloric low-protein (2.5%, n=10) diet for 10 weeks. The humeri were cyclically loaded in three-point bending under load control for 2000 cycles. The peak load selected corresponded to 60% of the maximal load of the contralateral humerus. The humeri were then loaded to failure. We compared the load/displacement curve of the cyclically loaded humerus to the contralateral non-cyclically loaded humerus.

Whereas cyclic loading did not induce any deterioration in rats fed a normal protein diet, this cyclic loading regimen negatively influenced the post-yield behaviour of humerus in rats fed a low protein diet, as indicated by the significant decrease of post-yield load and plastic deflection. This suggests that bone microdamages could be more prominent in rats fed a low protein diet than in control bones submitted to the same loading regimen. 

Table  .  

Disclosures: V. Dubois-Ferrière, None.

W405

Thin Healthy Women Have a Similar Low Bone Mass as Women with Anorexia Nervosa. D. Fernandez-Garcia*¹, J. Garcia-Aleman*¹, A. Sebastian Ochoa*², M. Muñoz-Torres², F. Tinahones*¹. ¹Department of Endocrinology, University Hospital Virgen de la Victoria, Malaga, Spain. ²Department of Endocrinology, University Hospital San Cecilio, Granada, Spain.

Association between anorexia nerviosa (AN) and low bone mass has been demonstrated. Bone loss associated with AN involve hormonal and nutritional impairments, though their exact contribution is not clearly established.

The aim: of our study was to compare bone mass in AN patients with women of similar weight with no criteria for anorexia nervosa, and a third group of healthy, normal-weight, age-matched women.

We studied 48 patients with AN (DSM-IV criteria), 22 healthy eumenorrheic women with low weight (LW Group; BMI <18.5 kg/m²) and 20 healthy women with BMI > 18.5 kg/m² (Control Group), all of similar age. We measured by DEXA lean body mass, percentage of fat mass, total bone mineral content (t BMC) and bone mineral density in lumbar spine (BMD LS) and total (BMD T). We measured anthropometric parameters, leptin and GH.

The control group had greater BMD T and BMD LS than the other groups, with no differences between the AN and LW groups. No differences were found in BMD T, BMD LS and t BMC between the restrictive (n=25) and binge-purge type (n=23) in AN patients. In AN, minimum weight (p=0.002) and percent of fat mass (p=0.02) explained BMD LS variation (r²:0,48) and minimum weight (r²:0,42; p=0.002) for BMD T in stepwise regression analyses. In LW group, BMI explained BMD LS (r²:0,72; p=0.01) and BMD T (r²:0,57; p=0.04).

Conclusions: Patients with AN had similar BMD as healthy thin women. Anthropometric parameters could contribute more significantly than estrogen deficiency in achievement of peak bone mass in AN patients.

Disclosures: D. Fernandez-Garcia, None.

W406

Vitamin D Insufficiency and Normal Parathyroid Function: Disease or No Disease?, K. E. Hansen¹, J. Riggert*¹, A. N. Jones*¹, J. Engelke*¹, M. Shafer*². ¹Medicine, University of Wisconsin, Madison, WI, USA. ²Wisconsin State Lab of Hygeine, Madison, WI, USA.

Vitamin D insufficiency (VDI) is allegedly widespread. In patients with normal parathyroid hormone (PTH), the diagnosis rests upon an increase in calcium absorption (CA) following increments in serum 25(OH)D to levels > 30 ng/ml. However, estimates of increased CA with correction of VDI vary dramatically, depending on methods used to measure CA. Using the increment for serum calcium as an index of CA, correction of VDI reportedly increases CA by 65% (J Am Coll Nutr 2003;22(2):142). Of concern, estimates of CA using serum calcium levels correlate poorly with estimates using calcium isotopes (r = 0.42, Calcif Tissue Int, 1983;35(6):819). We initiated a dual stable calcium isotope study to clarify the impact of vitamin D repletion on CA in postmenopausal women with VDI and normal PTH.

Eligible women were ≥ 5 years postmenopausal with a serum 25(OH)D of 16-24 ng/ml by HPLC assay, normal PTH and an estimated calcium intake ≤ 1,100 mg daily. Exclusion criteria included hypercalcemia, hypercalciuria, chronic intestinal disorders, use of medications known to interfere with vitamin D or calcium metabolism, osteomalacia, prior adult fragility fracture or a T-scores below −3.0 (spine or femur).

Each woman underwent inpatient dual calcium isotope studies when vitamin D insufficient and subsequently when vitamin D replete (25(OH)D > 35 ng/ml after ergocalciferol 50,000 IU/daily for 15 days). During each CA study, women consumed 18 mg of ⁴⁴Ca orally with breakfast and received 3 mg of ⁴²Ca intravenously. Each woman's typical diet was replicated during each CA study using 7-day diet records and Food Processor software. Urine was collected for 24 hours and analyzed for its calcium isotope content by mass spectrometry. We calculated CA using the formula by Eastell (JBMR 1989;4(4):463-468). Thus far, 6 of 22 women have completed the study. All had normal PTH at baseline (46 (9) pg/ml). Basal serum 25(OH)D was 20 (4) ng/ml, increasing to 73 (22) ng/ml with ergocalciferol. CA was 23% (8%) initially and 20% (8%) following vitamin D repletion (p>0.05, Wilcoxon signed-rank test).

Our observations suggest that calcium absorption does not increase in women with VDI who achieve unequivocal vitamin D repletion with ergocalciferol. Although preliminary, our data raise significant questions about whether VDI is a true disorder in patients with normal parathyroid function.

Disclosures: K.E. Hansen, None.

This study received funding from: CReFF, WI Partnership Program. Jackson Foundation.

W407

Effect of Citrus Flavanones on Bone Metabolism in Old Male Rats. V. Habauzit*¹, A. Trzeciakiewicz*¹, J. Mardon*¹, J. Mardon*¹, P. Lebecque*¹, M. Davicco*¹, C. Morand*¹, E. Offord*², V. Coxam¹, M. Horcajada*¹. ¹Human Nutrition, INRA, Saint Genès Champanelle, France. ²Nutrition and Health, NESTLE, Lausanne, Switzerland.

Currently, there is an increasing interest in fruit and vegetables as their consumption appears to be related to lower risk of metabolic disorders occurring during ageing. Polyphenols could be involved in these benefits. In this study we investigated the potency of hesperidin and naringin, their two major citrus flavonoids (flavanones), in regulating bone metabolism in senescent male rats.

Forty-eight 20-m-old male Wistar rats were divided into 4 groups. Control group received a casein-based diet. The others were fed, for 90 days, the same diet but added with either 0.5 % hesperidin (Hp), 0.5 % naringin (Nar) or a mix of both flavanones (0.25%Hp + 0.25%Nar). Femoral mineral density and strength, bone biomarkers (deoxypyridinoline and osteocalcin) were measured. Plasma and serum analyses (leptin, cholesterol, triglycerides, insulin, glucose, IGF-1, IL-6, FRAP, NO) were performed.

During the experiment, body weights were steady in each group and flavanones intake did not affect this parameter. Besides, plasma leptin was significantly decreased in Hp and Hp+Nar groups, suggesting an effect on body composition. At necropsy, daily Hp intake improved bone mineral density compared with control group, whatever bone sites considered (p<0.05). While the same pattern for the animals fed with the mix of flavanones was observed, only metaphyseal density was affected by Nar consumption (p<0.05). However bone strength was not modified by flavanones. Bone resorption was slowed down by Hp and Nar whereas accretion remained unchanged. Flavanones induced a decrease in plasma triglycerides and total cholesterol and no changes in blood glucose and plasma insulin were detected.

The consumption of flavanones appeared to prevent serum IL-6 from increasing during aging-related inflammation. Oxidative status seemed also to be improved in animals consuming flavanones as shown by FRAP assay and decreased NO production. Finally, circulating flavanones were only detected in rat plasma receiving Hp and/or Nar.

These results suggest that both hesperidin and naringin were able to affect bone metabolism in senescent male rats, hesperidin being apparently the more efficient. This effect could be partly due to the ability of flavanones in slowing down bone resorption rate. Bone sparing effect of hesperidin and naringin observed in senescent intact male rats, a model of ageing, should also be linked with anti-inflammatory, antioxidant and also hypolipidemic capacities of these compounds.

Disclosures: M. Horcajada. None.

W408

Influence of Moderate Energy Restriction and Seafood Consumption on Bone Turnover in Overweight Young Adults. A. J. Lucey*¹, G. Paschos*¹, K. D. Cashman², J. A. Martinéz*³, N. Bandarra*⁴, I. Thorsdottir*⁵, M. Kiely*¹. ¹Department of Food & Nutritional Sciences, University College Cork, Cork, Ireland. ²Department of Food & Nutritional Sciences & Department of Food & Medicine, University College Cork, Cork, Ireland. ³Department of Physiology & Nutrition, University of Navarra, Pamplona, Spain. ⁴National Institute on Agriculture and Fisheries Research, Lisbon, Portugal. ⁵Unit for Nutrition Research, Landspitali University Hospital, Reykjavik, Iceland.

While weight loss may increase bone turnover, increased intakes of n-3 fatty acids (FA) may maintain skeletal health by reducing bone resorption. This study examined relationships between weight loss, changes in FA in red blood cell (RBC) membrane phospholipids (%) and bone turnover biomarkers, induced by an 8-week dietary intervention, which included fish or fish oil as part of an energy restricted diet (-30%) in 20-40 year old adults (BMI 27.5-32.5 kg/m²). Participants (n 276) were randomised to 1 of 4 groups: placebo (sunflower oil capsules; 3g/day); cod (3 × 150g/week); salmon (3 × 150g/week); fish oil capsules (3g/day). At baseline and endpoint serum levels of the biomarkers of bone formation (osteocalcin (OC) and bone specific alkaline phosphatase (BAP)) and resorption (serum crosslaps (CTx) and N-telopeptide of Type 1 collagen (NTx)) were measured by enzyme linked immunosorbent assay (ELISA) and the FA composition of RBC membrane phospholipids (%) was measured by gas chromatography. Using repeated measures analysis of variance, before adjusting for confounding factors, serum levels of CTx (P < 0.001) and NTx (P < 0.05) increased, BAP showed little response, while levels of OC (P < 0.05) decreased from baseline to end point, however, these changes were no longer present after adjusting for country, gender, sampling month, age, smoking status and % weight loss. Mean weight loss was 5.8% (range 0 to 15) over the 8-week intervention and was similar across the 4 study groups. Weight loss and changes in body composition were the main factors influencing the increases in CTx (P < 0.001) and NTx (P < 0.05), while OC was unrelated to weight loss. Increased n-3 FA consumption in the salmon and fish oil groups resulted in increases in n-3 FA in RBC membranes and decreases in n-6 FA (P < 0.05). After adjusting for confounding, OC was positively correlated with the % RBC n-3 FA at baseline and endpoint. However, the overall inclusion of fish or fish oil in the diet had no significant effect on bone turnover. We conclude that a nutritionally adequate energy-restricted diet over 8 weeks, which resulted in modest weight loss and changes in body composition influenced biomarkers of bone resorption in young overweight adults.

Disclosures: A.J. Lucey, None.

This study received funding from: The YOUNG study (co-ordinator Prof. Inga Thorsdottir) is part of the SEAFOODplus Integrated Project (co-ordinator Prof. Torger Børresen), which is funded by the EC through the 6th Framework Programme Contract No FOOD-CT-2004-506359.

W409

Effects of Calcium Sources and Soluble Silicate on Bone Metabolism and the Related Gene Expression in Mice. F. Maehira¹, I. Miyagi*¹, Y. Eguchi*². ¹Lab. Metabolic Chemistry, Faculty of Medicine, University of the Ryukyus, Okinawa, Japan. ²Research Laboratory Center, Faculty of Medicine, University of the Ryukyus, Okinawa, Japan.

Silicon has been known as an essential element for the formation of collagen and glycosaminoglycans in bone and cartilage. The effects of five calcium (Ca) sources were compared on bone biochemical and mechanical properties and the related gene expression using the control strain of senescence-accelerated mice (SAMR1), from a viewpoint of their soluble silicon (Si) content. The experimental protocols of this study were approved by our Institutional Animal Studies Committee. Weanling male SAMR1 were fed the diets containing 1% Ca supplemented with CaCO₃ as the control (CT), coral sand (CS), fossil stony coral (FSC), fish (FC), egg-shell (EC) powders, and 50 ppm Si in the CT diet for 6 months. The mRNA expression of Runx2, BMP-2, IL-11, COL1A1, PPAR-γ, OPG, RANKL, TGF-β was quantified by a real-time PCR. Soluble Si content was 9.83, 7.17, 2.48, 0.29, and 0.20 ppm for CS, FC, FSC, EC, and Ca-deficient basal diet, respectively. In bone biochemical properties, Si, CS, and FSC in order increased (P < 0.05) dry and ash weights, Ca and hydroxyproline (OHPro) contents, and alkaline phosphatase and decreased (P < 0.05) tartrate resistant acid phosphatase, urinary excretion of OHPro comparing with the CT group. In bone mechanical properties, Si increased (P < 0.05) and FC decreased (P < 0.05) strength and stiffness. In the mRNA expression related to osteoblastogenesis, Si increased (P < 0.05) Runx2. Si, CS, and FSC in order decreased (P < 0.05) and FC, EC increased (P < 0.05) PPAR-γ. In the mRNA expression related to osteoclastogenesis. Si, CS increased (P < 0.05) and FC, EC decreased (P < 0.05) OPG/RANKL ratio whereas Si, CS decreased (P < 0.05) TGF-β. FC in spite of its high Si content showed worst effects on bone, probably due to its high phosphorus content. The results indicated that soluble silicate and coral sand with highest soluble Si content among Ca sources improved bone biochemical and mechanical properties through stimulation of gene expression related to osteoblastogenesis and suppression of that related to osteoclastogenesis.

Disclosures: F. Maehira, None.

W410

The Effect of a High Protein Diet on Bone and Body Composition in Young Female Rats. K. M. Pye*¹, A. Wakefield*², H. Aukema*², J. House*², M. Ogborn*², H. Weiler¹. ¹School of Dietetics and Human Nutrition, McGill University, Ste-Anne de Bellevue, PQ, Canada. ²Department of Human Nutritional Sciences, University of Manitoba, Winnipeg, MB, Canada.

Long-term, high protein diets at 35% of energy may have implications in bone biology. Increased consumption of protein lowers blood pH levels which may interfere with blood calcium homeostasis leading to an increased stimulation of osteoclasts and elevated calcium losses. However, inconsistencies within the literature make it impossible to state whether this level of bone resorption, over long periods, has a negative impact on bone health. No one study within the eighty years of high protein diet research has examined all the biochemical, physical, and largely the biomechanical markers of bone health. This study will be the first to give a complete analysis and examine whether a high protein diet at the 35% energy level can be deemed safe with respect to long-term bone health.

The objective of this research is to examine the effect a high protein diet at 35% of energy has on bone mass and strength in the rat model.

Eighty female Sprague-Dawley rats were randomized to receive 4, 8, 12, or 17 months of a control diet (15% of energy as protein) or the high protein diet (35% energy). Weekly food intake and weight was documented. Whole body composition and regional bone mass were measured in vivo using dual-energy x-ray absorptiometry (DXA; Hologic QDR version 11.2, Hologic, Inc.). Following termination, femurs and tibias were excised. Ex vivo, femur and tibia bone mass were measured using DXA and right femurs were tested for mechanical strength using designed 3-point bending tests (INSTRON 5544, Instron Inc.). Main effects of diet and time were detected using factorial ANOVA (p < 0.05).

The high protein diet increased BMC (g/kg) in femur, tibia, and vertebrae as illustrated in the table below. High protein consumption also decreased whole body fat (normal diet: 165.17 ± 87.92 g, high diet: 110.60 ± 70.92 g, p < 0.0001), increased lean mass (normal diet: 634.68 ± 99.95 g/kg, high diet: 717.11 ± 92.75 g/kg, p<0.0001), and decreased abdominal fat (normal diet: 44.56 ± 22.32 g, high diet: 31.55 ± 19.18 g, p=0.0006). Diet had no effect on the biomechanical strength of femurs. Thus preliminary results suggest that high protein diets at the current upper limit of 35% of energy increase mineral content, decrease body fat content, but does not hinder the mechanical and weight bearing abilities of bone. 

Table  .  

Disclosures: K.M. Pye, None.

This study received funding from: Canadian Institutes for Health Research.

W411

Low Calcium Diet Accelerates Alveolar Bone Resorption in Rat Experimental Periodontitis. K. Tokunaga¹, C. Wada¹, H. Ohba¹, H. Seto¹, E. Takeda², T. Nagata¹. ¹Periodontology and Endodontology, Health Bioscience, Tokushima, Japan. ²Clinical Nutrition, Health Bioscience, Tokushima, Japan.

Periodontitis is characterized by the inflammatory destruction of periodontal tissues including alveolar bone, periodontal ligament, and gingiva. Alveolar bone resorption is a major cause of tooth loss. It is possible that a decrease of general bone density may affect the pathogenesis of periodontitis. Estrogen deficiency by menopause and lack of Ca ingestion is known to be a risk factor for osteoporosis. However, it is not clear whether the lack of Ca ingestion influences alveolar bone resorption in periodontitis. In this study, we examined the effect of low Ca diet on alveolar bone resorption in rat experimental periodontitis. Male Wistar rats, 8 weeks old, were firstly divided into 2 groups: rats with low Ca diet (Ca: 0.18%), and the control (Ca: 1.08%). The diets were given for 5-20 days. Nylon ligature was placed around the cervix of maxillary lateral second molars to induce periodontitis with bone resorption. Then ligature-treated group and ligature-treated low Ca group were prepared. On days 5 and 20, maxillae was extracted and analysed using micro CT. Samples were decalcified and the thin sections were stained by tartrate-resistant acid phosphatase (TRAP). Multinuclear osteoclast-like cells were visualized under light microscopy. Micro CT images of maxillae showed that there was no change in the non-ligature group and that alveolar bone resorption was strongly induced around the second molar from days 5 to 20 in the ligature-teated groups. The ligature-treated sites in low Ca group showed a greater bone resorption than the ligature-treated control. Histrogical examination showed that TRAP positive cells appeared at the top of alveolar bone on day 5 in the ligature-treated groups. The number of osteoclasts in the ligature-treated low Ca group was more than that in the ligature-treated control. These osteoclast-like cells disappeared on day 20 in both groups. Taken together, low Ca diet may increase the number of osteoclasts at the top of alveolar bone and induce the greater resorption of alveolar bone in the experimental periodontitis. These findings suggest that the lack of Ca ingestion in daily life may accelerate alveolar bone resorption in periodontitis.

Disclosures: K. Tokunaga, None.

W412

A Marked Abnormality in Myeloid and Mesenchymal Somatic Stem Cells Explains the Skeletal Defect in Glucagon-Like Polypeptide-2 Receptor KO Mice. J. Xiao¹, D. J. Drucker*², J. A. Clowes¹. ¹Mayo Clinic, Rochester, MN, USA. ²Department of Medicine, University of Toronto, Toronto, MN, USA.

Clinical studies have established nutritional intake as a key factor regulating the marked circadian rhythm in bone turnover. In addition, at pharmacological concentrations the intestinal hormone, glucagon like peptide 2 (GLP2) which is released in response to nutrient intake acutely decreases bone resorption. Furthermore, we have established that there is a marked skeletal deficit during growth in GLP2 receptor knockout (GLP2R -/-) mice.

In addition, previous studies have established that CDllb+ myeloid precursors differentiate into mature osteoclasts (OCs) and express key surface receptors, including c-Fms (the receptor for MCSF) and RANK (the receptor for RANKL). Furthermore, cells expressing strol a mesenchymal stem cell (MSC) marker differentiate into osteoblast (OB) precursors following co-expression with RANKL. Finally CD133 is a marker of the most primitive somatic stem cell (SSC).

Although the importance of nutrition in bone biology has been clearly established the mechanism(s) for how GLP2 regulates bone metabolism in vivo at physiological concentrations has not yet been determined. We determined this in 6 week old GLP2R -/- mice (n = 4) compared to their wild type (WT) C57BL/6 littermates (n = 5) by examining the percentage of cells (as a fraction of bone marrow MNCs) expressing the various receptors using flow cytometry.

As shown in the Table, a defect in the GLP2R resulted in a significant increase in the % of total cells expressing RANK+ and the % of RANK+/CD34- and RANK+/cFms- cells. Furthermore, there was a significant increase in the % of total cells expressing the MSC marker strol, the % of strol+/CD133- and strol+/RANKL+ cells. 

Table  .  

Data are mean ± SD; *P < 0.05

These studies thus represent the first systematic examination of the effects of a nutrient-regulated hormone at physiological concentrations on OC and OB precursor populations in vivo. Collectively, these data indicate that the absence of GLP2 signaling increases the differentiation of myeloid cells towards mature osteoclasts. Since there is a profound defect in OB differentiation and mineralization in vitro it suggests the increase in MSC differentiation towards mature OBs may either represent a compensatory mechanism or an increase in OB precursors due to a distal block in OB differentiation.

Disclosures: J. Xiao, None.

This study received funding from: Arthritis Research Campaign. IUK.

W413

Comprehensive Analysis of Bone-related Genes Expression Induced by Acute Oxidative Stress. K. Mori¹, R. Kitazawa¹, T. Kondo¹, Y. Hamada*², S. Kitazawa¹. ¹Division of Pathology, Kobe University Graduate School of Medicne, Kobe, Japan. ²Division of Nephrology and Kidney Center, Kobe University Hospital, Kobe, Japan.

In the diabetic condition, accumulation of methylglyoxal (MG), an intermediate metabolite of glucose formed through glycolysis, precedes formation of advanced glycation end products (AGE), which then cause oxidative stress in various cells and organs. This oxidative stress is now regarded as a major cause of diabetic complications such as diabetic nephropathy, retinopathy, angiopathy and bone loss. In this study, we comprehensively analyzed gene expression by the DNA microarray system to identify genes in osteoblastic/ stromal cells modulated by oxidative stress. Mouse bone marrow stromal cells, ST2, were treated with MG (100μM) for 48hr, and RNA extracted from ST2 cells cultured with or without MG was examined. DNA microarray analysis revealed that the expression of osteoprotegerin (OPG), a decoy receptor of receptor activator of nuclear factor-kB ligand (RANKL), was markedly reduced by MG treatment. In contrast, the expression of soluble Frizzled-related protein 4 (sFRP-4), a decoy receptor for Wnt proteins, increased by MG treatment. This reciprocal gene expression between OPG and sFRP-4 was also reproduced and confirmed by quantitative real-time RT-PCR. Since the canonical Wnt/β-catenin signaling pathway contributes to OPG gene expression in osteoblastic cells through the transactivation of the gene, the rapid and marked decrease of OPG gene expression after MG-induced acute oxidative stress may be due to reduced Wnt/β-catenin signaling by the upregulated expression of its antagonist, sFRP-4. Furthermore, the expression of several target genes of the Wnt/β-catenin signaling pathway (e.g. cyclin D1, Wnt1 inducible signaling pathway protein 1 (WISP1), WISP2) also decreased in MG-treated ST2 cells. On the other hand, the expression of mesenchymal cell differentiation regulating transcription factors, such as Runx2 for osteoblasts and peroxisome proliferators- activated receptor γ (PPAR-γ) for adipocytes, did not change significantly by short-term MG treatment. In conclusion, oxidative stress induces a rapid reduction of Wnt/β-catenin signaling and expression of its target genes. We speculate therefore that diabetic bone loss is ultimately caused by decreased Wnt/β-catenin signaling, and that the early bone-resorption-dominant phase attributed to rapid and marked decrease of OPG gene expression may precede final low turnover osteoporosis as a late complication of diabetes.

Disclosures: K. Mori, None.

W414

Poor Long Term Glycemic Control Is Not Associated with Decreased Bone Mineral Density in Typ 1 Diabetes. T. Neumann*¹, A. Saemann*², S. Lodes*¹, B. Kaestner*¹, S. Franke*¹, C. Hemmelmann*³, U. A. Mueller*², G. Hein*¹, G. Wolf*⁴. ¹Department of Medicine III, Rheumatology/ Osteology, Friedrich-Schiller-University of Jena, Jena, Germany. ²Department of Medicine III, Endocrinology, Friedrich-Schiller-University of Jena, Jena, Germany. ³Institute of Medical Statistics, Computer Sciences and Documentation, Friedrich-Schiller-University of Jena, Jena, Germany. ⁴Department of Medicine III, Nephrology, Friedrich-Schiller-University of Jena, Jena, Germany.

It is assumed that type 1 diabetes is associated with lower bone mineral density (BMD) and increased fractures risk. The impact of duration of diabetes on BMD is controversial. In a cross-sectional study design we investigated the influence of long term glycemic control on BMD in type 1 diabetes. We studied 126 unselected patients with diabetes mellitus type 1 from our outpatient clinic, 63 premenopausal women (41 +/- 8 years) and 63 men (45 +/- 10 years), duration of diabetes was 22 +/- 10,5 years. All patients received standard questionnaire on diabetic history and risk factors for osteoporosis. History of HbAlc was obtained from medical records (mean HbAlc for each year of diabetes duration). 15 patients (10 women and 5 men) were excluded from analysis of long term glycemic control because of missing HbAlc data. BMD was measured using DXA (Lunar Prodigy Advance) at lumbar spine (LS), right femoral neck (FN) and total hip (TH).

Osteoporosis (T-score <−2,5 SD) at the LS (FN; TH) was found in 5 (4; 2) %. Osteopenia (T-score between −1 and −2,5 SD) at the LS (FN; TH) was found in 33 (34; 26) %. BMD was not associated with cigarette smoking or a familial history of osteoporotic fracture. Neither diabetes duration nor quartile of duration (1^st, 2 to <14 years; 2^nd, 14 to <21 years; 3^rd, 21 to <31 years; 4^th, 31 −48 years) were associated with BMD. Quartiles of median HbAlc (1^st, 5,4 to <6,8 %; 2^nd, 6,8 to <7,4 %; 3^rd, 7,4 to <8,1 %; 4^th, 8,1-10,8 %) over the time of diabetes were not associated with BMD. No association was found for markers of bone turnover and median HbA lc by measuring serum C-terminal telopeptide of type 1 collagen (CTX).

Our data suggest that osteoporosis is a clinical significant and commonly underestimated problem in type 1 diabetes. However, the quality of long term glycemic control as well as the duration of diabetes type 1 has no impact on BMD of lumbar spine, femoral neck and total hip.

Disclosures: T. Neumann, None.

W415

Antiepileptic Drugs Affect Bone Loss Via Reproductive Hormones. A. Pack¹, E. Shane², D. McMahon², A. Randall*¹, M. Morrell*³. ¹Neurology, Columbia University, New York, NY, USA. ²Medicine, Columbia University, New York, NY, USA. ³Neurology, Stanford University, Palo Alto, CA, USA.

The antiepileptic drugs (AEDs), phenytoin (PHT) and carbamazepine (CBZ), are commonly associated with low bone density (BMD) and high bone turnover markers (BTM). Traditionally they are thought to affect bone metabolism by inducing the hepatic cytochrome P450 enzyme CYP3A4 which in turn increases catabolism of vitamin D to inactive metabolites, causing lower serum calcium (SCa) and higher parathyroid hormone (PTH) secretion. Notably, however, CYP3A4 also increases metabolism of estradiol (E2) to inactive 2-OH-estradiol. Moreover, CYP enzyme inducing AEDs (EIAEDs) are associated with increased oral contraceptive failure rates, decreased serum E2 and androgens, and increased serum sex hormone binding globulin (SHBG), thus decreasing free, biologically active hormone. These effects of EIAEDs on E metabolism may also compromise bone health. We hypothesized that women with epilepsy (WWE) treated with EIAEDs have lower E levels, higher bone turnover and lower BMD than WWE treated with lamotrigine (LTG), an AED with no effect on CYP450.

WWE aged 18-40 on monotherapy with EIAEDs CBZ or PHT (n=18) or LTG (n=13) were studied during the follicular phase of the menstrual cycle. E2, estrone (E1), testosterone (T), and SHBG were measured. Free E2 and T were calculated. BMD was measured by DXA. SCa, 25-OHD, 1,25(OH)₂D, PTH, osteocalcin (OC), bone alkaline phosphatase (BSAP) and urine N-telopeptide (UNTX) were assessed.

El, % free E2, and % free T were lower and SHBG was higher in women on EIAEDs compared to LTG. SCa was lower and UNTX was higher in women on EIAEDs, although there was no correlation between BTMs and reproductive hormones. In contrast, vitamin D metabolites and PTH did not differ. BMD did not differ between groups at any site. In the EIAED group, spine BMD correlated positively with E2 (r=0.58; p =0.02) but femoral neck BMD correlated negatively with % free E2 (r=-0.54; p=0.03). There was no relationship between BMD and calciotropic hormones or SCa in either group.

In summary, women on PHT or CBZ had lower levels of gonadal hormones and higher SHBG than those on LTG, while calciotropic hormones did not differ. Although BMD was not lower in the EIAED group, lower E exposure may lead to higher rates of bone loss over time. We conclude that adverse effects of EIAEDS on bone health may be mediated in part through effects on gonadal hormone status. 

Table  .  

Disclosures: A. Pack, GlaxoSmithKline 2, 5, 8; Novartis 8; Abbott 8.

This study received funding from: GlaxoSmithKline.

W416

Hepatic Osteodystrophy: Expression of IGF-1 and GH Receptor (GHR) and Biomechanical Studies in Experimental Model of Cholestatic Liver. F. A. Pereira¹, I. Facincani*¹, L. N. Z. Ramalho*², J. B. Volpon*³, F. J. A. Paula¹. ¹Internal Medicine, School of Medicine of Ribeirão Preto, University of São Paulo, Ribeirão Preto, Brazil. ²Pathology, School of Medicine of Ribeirão Preto, University of São Paulo, Ribeirão Preto, Brazil. ³Biomechanical and Rehabilitation of the Locomotor System, School of Medicine of Ribeirão Preto, University of São Paulo, Ribeirão Preto, Brazil.

Hepatic osteodystrophy is an important co-morbidity of chronic liver disease. In the present study we assessed the impact of chronic cholestasis on the expression of IGF-1 and GHR in liver and in growth plate and bone resistance in rats submitted to bile-duct obstruction (BDO). Male Wistar rats were divided into two groups: Sham (n=15) and Cirrhotic (n=15). On day 30 post-surgery, the rats were sacrificed and blood was withdrawn to determine serum liver enzymes and bilirubin. Liver and left tibia were used for immunohistochemistry and right femur for biomechanical study. Liver histology was evaluated. Immunohistochemistry semiquantitative expression was performed according to the following criteria: (-) no stained cells; (+) less than 10% positive cells; (++) 10% −50% positive cells; (+++) more than 50% positive cells. Bilirubin (Sham =0.08 ± 0.01 mg/dl vs Cirrhotic 6.6 ± 0.1 mg/dl p<0.001), alkaline phosphatase (Sham =68.3 ± 0.3 U/L vs Cirrhotic 1864 ± 0.6 U/L p<0.001), γGT (Sham =5.8 ± 0.2 U/L vs Cirrhotic 66.5 ± 0.2 U/L p<0.001) levels were significantly higher in Cirrhotic group. In this group, all animals showed histological evidence of cholestatic cirrhosis. Also, in liver tissue, the levels of IGF-1 expression were significantly higher in Cirrhotic group in comparison with Sham group (p<0.01), and the level of GHR expression were significantly lower in cirrhotic group (p<0.01). In growth plate cartilage the levels of IGF-1 expression was significantly lower in cirrhotic group (all animal showed no stained cells or less than 10% of positive cells) in comparison with Sham group (the majority of animals showed more than 50% IGF-1 positive cells) (p<0.01). The same pattern was observed in the growth plate cartilage in relation to GHR expression (p<0.01). The applied forces at the fracture moment were: Sham = 82.5 ± 8.6 N vs Cirrhotic = 57.1 ± 12.1 N 9 p<0.001). Our results indicate that the bone quality is significantly impaired on chronic liver cholestasis in rats and probably the reduction of expression of growth factors on bone microenvironment is involved in this process.

Disclosures: F.A. Pereira, None.

W417

An Integrated Model for Evaluation and Simulation of Therapeutic Responses to Bone-Related Therapies. M. C. Peterson¹, M. M. Riggs*². ¹Amgen Inc., Thousand Oaks, CA, USA. ²Metrum Research Group, LLC, Tariffville, CT, USA.

Calcium (Ca) homeostasis and bone remodeling are complexly integrated, highly regulated processes involving multiple organs and endocrine pathways. Published generalized models have failed to provide adequate platforms for describing multiple therapeutic class effects on the system. Our aim was to develop a model that predicts longitudinal Ca homeostasis and co-factor levels (including phosphate, PTH, and calcitriol), as well as expected rates of bone turnover due to natural progression, therapeutic intervention, or disease states.

Our approach is based on physiologic requirements to maintain Ca balance, with the general model predicated on 3 manuscripts [Raposo et. al, J Clin Endocrin Metab, 2002; Lemaire et. al, J Theor Biol, 2004; Bellido et. al, J Biological Chem, 2003]. Based on literature and Amgen clinical data, significant modifications and additional feedback mechanisms have been added to appropriately integrate the model. The model includes the RANK-RANKL-OPG system to enable characterization of osteoclast (OC) and osteoblast (OB) genesis and survival, PTH-mediated anti-apoptotic signaling in OB to allow description of anabolic and catabolic effects of PTH on bone remodeling, and TGFbeta-mediated feedback mechanisms to capture homeostatic observations.

The model has enabled evaluation of various affecting mechanisms, ramifications of therapeutic interventions, and disease state sequelae, including the following: (1) administration of the RANKL inhibitor, denosumab, (2) intermittent- and (3) constant-PTH infusion, and (4) progressive renal insufficiency. The integrated model appears to provide a generalizable structure with known or plausible cell signaling concepts and physiologically consistent parameters. Model predictions under conditions (1)–(4) provide results consistent with clinical observations.

In conclusion, a robust physiologic model that provides continuous descriptions of biomarkers and electrolytes including calcium, phosphate, PTH, and markers of bone turnover consistent with literature and Amgen internal data has been developed. This model may facilitate characterization of therapeutics under development, enable predictions of cytokine mediators of bone homeostasis, and provide a platform for hypothesis testing prior to clinical investigations.

Disclosures: M.C. Peterson, Amgen Inc. 1, 3.

This study received funding from: Amgen, Inc.

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Effects of Estrogen on Circulating dlk1/pref-1/FA1 Levels in Postmenopausal Women and Correlations with Bone Turnover and Volumetric BMD. B. Srinivasan¹, B. M. Abdallah*², S. Khosla¹, M. Kassem². ¹Mayo Clinic, Rochester, MN, USA. ²Odense University Hospital, Odense, Denmark.

dlk1/FA1 (Delta-like 1/fetal antigen1), also known as Pref-1, (Pre-adipocyte factor 1), is a member of the epidermal growth factor-like protein family, known to modulate differentiation of mesenchymal and hematopoietic stem cells in bone marrow in a paracrine/endocrine fashion. Studies in mice have shown that dlk1/FA1 inhibits adipogenesis and is a marker for pre-adipocytes. Recently, Dlk1/FA1 was identified as a novel paracrine/endocrine inhibitor of human bone marrow stromal (hMSC) cell differentiation. Since estrogen (E) is known to enhance osteoblast differentiation and inhibit adipocytic differentiation, we measured circulating FA1 levels by immunoassay in untreated and E-treated postmenopausal women (n = 83 per group, mean age, 63 yrs) who were matched for age and BMI. Circulating FA1 levels were significantly higher in the untreated women (mean ± SEM, 41.5 ± 1.8 ng/ml) as compared to E-treated women (29.9 ± 1.4 ng/ml, P<0.001). Serum FA1 levels were correlated with serum osteocalcin levels in untreated (age and BMI adjusted correlation coefficient, R^age,BMI = 0.26. P = 0.02) and E-treated women (R^age,BMI = 0.27, P = 0.02), with femur neck cortical volumetric BMD (vBMD) measured by QCT in both groups (R^age,BMI = 0.24 and 0.26, respectively, P < 0.05), and with trabecular vBMD at the distal radius in the E-treated women (R^age,BMI = 0.24, P = 0.03). No significant associations with other bone turnover markers (PINP or CTx) or vertebral vBMD were noted. Collectively, these findings indicate that E suppresses circulating FA1 levels in postmenopausal women, perhaps reflecting the known effects of E in inhibiting differentiation of bone marrow stromal cells to pre-adipocytes. The positive associations with serum osteocalcin levels and with vBMD that were noted raise the possibility that higher dlk1/FA1 levels, whether in untreated or E-treated women, may suppress adipocyte differentiation of bone marrow stromal cells to a greater extent than osteoblastic differentiation, leading to a net positive effect of dlk1/FA1 on bone. Further studies are needed to test this hypothesis and to better understand the endocrine role of dlk1/FA1 in regulating bone turnover and bone mass.

Disclosures: B. Srinivasan, None.

This study received funding from: NIH/NIA — AR27065.

W419

To Detamine Serum Fluoride Levels in Patients with Osteoporosis Is Quite Useful for the Judgment of Effectiveness of Osteoporosis Treatment. K. Tanno*¹, K. Itai*¹, M. Ohsawa*¹, T. Onoda*¹, Y. Yaegashi*¹, T. Sato*¹, K. Sakata*¹, G. Murcoka*², T. Shimamura*², A. Okayama*³. ¹Department of Hygiene and Preventive Medicine, Iwate Medical University, Morioka, Japan. ²Department of Orthopedic Surgery, Iwate Medical University, Morioka, Japan. ³Department of Preventive Cardiology, National Cardiovascular Center, Suita, Japan.

Endogenous fluoride is mostly stored within the bone and vividly released into blood circulation along with bone resorption, therefore, serum fluoride levels may faithfully reflect resorption rates of bone metabolism. Purpose of this study is to determine serum fluoride levels in postmenopausal women with osteoporosis before and after treatment of antiresorptive agents and to examine whether serum fluoride levels faithfully reflect resorption rates represented as changes in bone mineral density (BMD) and changes in biochemical markers of bone turn over. A total of 55 postmenopausal women with osteoporosis were enrolled (mean age 69.4 years). BMD (measured by Dual energy X-ray absorptiometry), serum levels of bone alkaline phosphatase (BAP), serum levels of type 1 collagen cross-linked N-telopeptide (NTX), urinary levels of deoxypyridinoline (DPD), and serum fluoride levels (measured by flow injection method with ion selective electrode) were determined and compared before and after treatment with antiresorptive agents (Alendronate, Risedronate). Mean treatment period was 6.2 months (3.8–7.6 months). Serum fluoride levels, as well as serum BAP levels, serum NTX levels, and urinary DPD levels significantly decreased along with increases in BMD in patients with osteoporosis after antiresorptive treatment (see table). In conclusion, markedly decreased serum fluoride levels indicated attenuated resorption rates of bone metabolism, and serum fluoride should be a useful marker that faithfully reflects bone metabolism. To determine serum fluoride levels in patients with osteoporosis should become increasingly essential for the judgment of disease severity and effectiveness of osteoporosis treatment. 

Table  .  

Disclosures: K. Tanno, None.

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Case Series: Osteomalacia Following Gastric Bypass and Biliopancreatic Diversion Surgery. A. Al-Shoha¹, R. Lakhdar*¹, D. S. Rao². ¹Internal Medicine, Henry Ford Hospital, Detroit, MI, USA. ²Bone & Mineral Metabolism, Henry Ford Hospital, Detroit, MI, USA.

Gastric bypass surgery (GBS) and Biliopancreatic diversion (BPD) are the most common procedures for management of morbid obesity, a disorder that is increasing in epidemic proportions in the US and world wide. Metabolic bone disease after GBS has been reported. However, to our knowledge, only two cases of post-GBS osteomalacia (OM) have been reported, but the diagnosis of OM in both cases was based solely on biochemical findings. We report 4 patients with bone biopsy confirmed OM after GBS.

Three women and one man (age range 29-53y) were seen for evaluation of metabolic bone disease not responding to “usual” therapy. Two of the 4 patients had BPD; one of those 2 had gastric stapling (GS) about 9y before BPD (Case 2). The other 2 patients had Roux en-Y GBS; one of those had GS at the time of GBS (Case 3) and the other had jejunoileal bypass 26y before GBS (Case 4). Clinical features included generalized bone pain and tenderness, muscle weakness, difficulty walking, and waddling gait, stooping posture, and fractures. Two patients (Cases 2 and 4) had history of kidney stones at presentation. Diagnoses prior to referral were varied including arthritis, gout, vitamin D deficiency and osteoporosis. Serum chemistry (Table) and radiological findings were suggestive of OM in all the 4 cases, and the diagnosis was confirmed by bone biopsies. Following high doses of Vitamin D and calcium therapy there was significant improvement in the symptoms and functional status and biochemical variables.

Our experience suggests that GBS may predispose to severe vitamin D depletion and OM in the absence of high dose vitamin D and calcium supplements. The current “usual” supplements are grossly inadequate in this population and the presentation may be nonspecific and misleading. Prospective long-term studies are needed to determine the appropriate vitamin D dose to prevent metabolic bone disease in this uniquely susceptible population. 

Table  .  

Disclosures: A. Al-Shoha, None.

W421

Rickets Is Associated with Failure of Epiphyseal/Metaphyseal Bone Tether Formation. J. Chen*¹, C. H. Lohmann*², R. Coleman*¹, R. E. Guldberg*¹, Z. Schwartz¹, B. D. Boyan¹. ¹Petit Institute of Bioengineering and Bioscience, Georgia Institute of Technology, Atlanta, GA, USA. ²University of Hamburg-Eppendorf, Hamburg, Germany.

Recently, we showed that bone bridges (tethers) connect the metaphyseal and epiphyseal trabecular bone through the rat tibial growth plate (GP); altered tether formation was highly correlated to growth disruption. We hypothesized that altered chondro-osseous morphology typical of rickets might involve changes in tether number, distribution, or structure. To test this, we imaged the GP via micro-CT and histology to assess if 8-week old VDR-/- mice have changes in tether parameters compared to VDR+/+ littermates. GPs were isolated from micro-CT scans (resolution = 16 μm) of right distal femurs from 15 VDR-/- and 15 VDR+/+ mice and average thickness and volume were calculated. To define the tether area, GPs were marked with ellipses using Adobe Photoshop. The “tether probability index” was set as the fraction of samples with tethers in each area. Randomly selected GPs were processed for decalcified (8) and undecalcified (1) histomorphometry; serial sagittal sections were stained with H&E or silver nitrate, respectively. GP and tether vertical height (GP width), horizontal width and area were measured (Image-Pro Plus) and GP volume, GP projection area, tether volume and tether occupancy rate calculated; tether number was determined by reconstruction the consecutive histological images. For reconstructed 3D micro-CT images, tether number was counted directly, GP and tether projection area were measured with Image-Pro Plus software, tether volume was calculated from tether area and average GP thickness. Tethers occurred in all VDR+/+ femurs; in contrast, only 20% of VDR-/- femurs had tethers, and their area and distribution were different. Although VDR-/- GPs had greater area and thickness, tether number, area, occupancy rate and volume fraction were dramatically reduced. This was confirmed by histology of non-decalcified femurs. Statistical analysis showed that the histological results correlated with micro-CT for all tether parameters except tether volume. This study demonstrates the utility of the micro-CT method for assessing GP morphology and pathology. Tethers did form at specific sites in some VDR-/- animals, suggesting that rickets does not affect all regions of the GP equally. The results indicate that failure of the GP to develop properly has consequences for establishing the appropriate interface between two mechanically dissimilar tissues.

Disclosures: B.D. Boyan, None.

This study received funding from: Children's Healthcare of Atlanta, Price Gilbert. Jr.

Foundation. NSF 9731643.

W422

Minimally Invasive Treatment of Oncogenic Osteomalacia by Radiofrequency Ablation. E. Hesse¹, H. Rosenthal*², G. Brabant*³, E. Länger*⁴, K. F. Gratz*⁵, L. Bastian*⁶. ¹Orthopaedics and Cell Biology, Yale University School of Medicine, New Haven, CT, USA. ²Diagnostic Radiology, Hannover Medical School, Hannover, Germany. ³Endocrinology, Christie's Hospital, Manchester, United Kingdom. ⁴Pathology, Hannover Medical School, Hannover, Germany. ⁵Nuclear Medicine, Hannover Medical School, Hannover, Germany. ⁶Trauma Surgery, Klinikum Leverkusen, Leverkusen, Germany.

Oncogenic osteomalacia (OOM) is a rare disorder characterized by hyperphosphaturia, hypophosphatemia, and osteomalacia. OOM is treated by surgical excision, which rapidly and permanently abrogates all symptoms. However, tumor removal can be complicated because the lesion is usually small and difficult to distinguish from the surrounding tissue. Complete excision is necessary, however, because the syndrome typically persists if any tumor tissue remains. To ensure complete excision, surgery may require wide resection margins, causing iatrogenic tissue damage that is disproportional to tumor size and to the almost always benign nature of the tumor. We report for the first time the use of minimally invasive radiofrequency ablation (RFA) to completely destroy a tumor causing OOM and thereby avoid arthroplasty. A 40-year-old woman was diagnosed for an acquired hypophosphatemic vitamin D-resistant osteomalacia due to a tumor in her right femoral head that was detected as recently described (JBMR, 2007 Jan;22(1): 158-62). Laboratory analyses revealed hypophosphatemia and a diminished TmP/GFR. A complete tumor removal by an open surgical procedure would have required total hip arthroplasty. Because of her young age, we attempted to preserve the hip joint, which was healthy and not affected by the tumor. Histology revealed a benign mesenchymal tumor. We performed CT-guided RFA to destroy the lesion, thereby avoiding arthroplasty. Complete tumor ablation was achieved six days later after a second round of RFA and confirmed on the following day by magnetic resonance imaging (MRI). A few weeks after RFA, biochemical markers returned to normal and all symptoms resolved. A one year clinical follow-up was unremarkable. RFA is a well-established procedure for selectively removing small tissue volumes. In bone, RFA is an effective palliative method for treating skeletal metastases. RFA of benign primary bone tumors has been primarily used to treat osteoid osteoma. This case demonstrates that CT-guided RFA can successfully treat a tumor causing OOM. We believe that RFA may broaden the scope of treatment options in OOM and may offer an effective, less invasive technique that would be a very attractive alternative to classical surgery.

Disclosures: E. Hesse, None.

W423

Severe Vitamin D Depletion in Arab-American Women Living in Dearborn, MI, USA. R. D. Hobbs*¹, Z. Habib*¹, D. Alromaihi*¹, L. Ida*², N. Parikh*², J. Schmidt*³, J. Kordosky*³, D. S. Rao². ¹Internal Medicine, Henry Ford Hospital, Detroit, MI, USA. ²Bone & Mineral Research Laboratory, Henry Ford Hospital, Detroit, MI, USA. ³DiaSorin, Inc, Stillwater, MN, USA.

Although vitamin supplements and dairy intake are important determinants of vitamin D nutrition, most individuals can maintain sufficient 25-OHD levels with adequate sun exposure. Secular habits, geographic location (latitudes <40°) or diminished sun exposure may lead to vitamin D depletion. European studies have documented vitamin D depletion in veiled Muslim women living in areas with decreased ambient sunlight, but no such data exist for this ethnic group living in the US. Accordingly we assessed vitamin D nutritional status by serum 25-OHD in Arab-American women living in Dearborn, MI, the largest, most concentrated Arab settlement in the US.

Serum 25-OHD and PTH were measured in 87 Arab-American women who attended an ethnic market on either April 7 or 14, 2007. Demographic variables, dress, medication use, medical history, vitamin supplementation and a food analysis were performed on each subject. A cross sectional approach with analysis using MANOVA or Fischer's exact test was used.

Three groups were formed as follows: unveiled (UV; n=22), veiled with supplementation (VS; n=45), and veiled non-supplemented (VNS; n=20).

Mean 25-OHD levels were lowest and mean PTH levels highest in the veiled non-supplemented group, and differed significantly from the other two groups and from each other (p=0.004, p=0.003 respectively). Although mean age and location did not differ, the mean time in the US was significantly different between the groups (p=0.002). Vitamin D supplementation between the veiled/supplemented (1250 ± 1243 IU/week)) and unveiled groups (1000 ± 1276 IU/week) did not differ (p=.45).

Vitamin D depletion, as assessed by 25-OHD, is highly prevalent in a sample of Muslim women living in Dearborn, MI and probably the highest among any demographic groups studied thus far. Similar to European studies the findings are at least in part attributable to conservative dress in an area with decreased sun exposure. With increasing time spent in the United States vitamin D levels increase across all tested groups although most (61/87) had levels ≤ 10 ng/ml. These findings potentially identify a problem in the largest, most concentrated Arab-American population in the United States. 

Table  .  

Disclosures: R.D. Hobbs, None.

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Lack of Efficacy of Octreotide in the Treatment of Tumor-induced Osteomalacia. M. H. Kelly*¹, B. Brillante*¹, A. Khosravi*², M. T. Collins¹. ¹National Institute of Dental and Craniofacial Research, National Institutes of Health, Bethesda, MD, USA. ²Neurorehabilitation Department, Kennedy Krieger Institute, Johns Hopkins University, Baltimore, MD, USA.

The purpose of this study was to test the efficacy of the somatostatin analogue, octreotide, in the treatment of tumor-induced osteomalacia (TIO).

TIO is a rare, acquired disease characterized by low serum phosphate and 1,25-dihydroxyvitamin D₃ (1,25-D), and osteomalacia. It is due to mesenchymal tumors that produce the phosphate and vitamin D-regulating hormone, FGF-23. Removal of the tumor is curative, but the tumors sometimes cannot be located and medical treatment is required. Octreotide has been reported to be an effective medical therapy. We sought to confirm the efficacy of octreotide by treating six patients with TIO.

Six patients with TIO who were off phosphate and calcitriol replacement were studied. Five of the six subjects had FGF-23 secreting tumors that were localized on Indium-111-pentetreotide scans (a radiolabeled analogue of octreotide). Subjects were treated with octreotide (Sandostatin, Novartis) 100 mcg by subcutaneous injection every 8 hours for three days. Serum phosphorus, 1,25-D, and FGF-23 were measured serially for 3 days. Repeated measures ANOVA was performed to test for significant changes in serum measurements.

None of the subjects responded to the octreotide. The mean changes in serum values over the 3 day period were: phosphorus 4% (range −3 to 13), 1,25-D 10% (-5 to 14), and FGF-23 −1% (-11 to 8). There was no significant change in serum phosphorus, 1,25-D, or FGF-23 in response to treatment with octreotide.

During 3 days of treatment with relatively high doses of octreotide, there was no effect on serum phosphorus, 1,25-D, or serum FGF-23 in patients with FGF-23 secreting tumors, in spite of the fact that 5/6 tumors took up pentetreotide.

Disclosures: M.H. Kelly, None.

W425

Skeletal Fluorosis from Instant Tea: A Second Case. M. P. Whyte¹, W. G. Totty*², G. M. Whitford*³. ¹Division of Bone and Mineral Diseases, Washington University School of Medicine, St. Louis, MO, USA. ²Mallinckrodt Institute of Radiology, Washington University School of Medicine, St. Louis, MO, USA. ³School of Dentistry, Medical College of Georgia, Augusta, GA, USA.

Skeletal fluorosis (SF) usually results from prolonged consumption of well water with > 4 parts per million (ppm) fluoride ion (F); i.e., > 4 mg/L. Black and green teas can contain significant amounts of F. In 2005, SF due to drinking 1–2 gallons of double-strength instant tea daily throughout adult life was reported in a 52-year-old woman (Am J Med 118:78).

A 49-year-old woman developed widespread musculoskeletal pains, considered fibromyalgia, in her mid-30's. Additionally, there was unexplained, increasing, axial osteosclerosis. She reported drinking 2 gallons (7.56 liters) of instant tea each day since age 12 years, and took fluoxetine intermittently for 5 years. Ion-selective electrode methodology quantitated F in her blood, urine, municipal tap water, beverage, and extracts of fingernail and toenail clippings.

Radiographs showed uniform osteosclerosis involving the axial skeleton, but enthesopathy was consistent with her age. DXA Z-scores were + 10.3 in the lumbar spine, and + 2.8 in the total hip. Serum F concentration was 120 μg/L (20–80, reference range), and a 24-hour urine specimen contained 18 mg F/gm creatinine (< 3, reference range). Mean F levels were 2.2 and 2.8 times greater than for 3 age-matched controls in fingernails and toenails, respectively (p < 0.001). Her instant tea beverage, prepared extra-strength as usual and using tap water with ∼ 1.14 ppm F, contained 5.8 ppm F. In the U.S.A., this F level exceeds the Environmental Protection Agency limit of 4.0 ppm for primary drinking water, the Food and Drug Administration limit of 1.4–2.4 ppm for bottled water or beverages, and the Public Health Service optimum fluoridation level in community water ranging from 0.7–1.2 ppm. The tea product contributed ∼35 of the 44 mg daily F exposure from her beverage. Fluoxetine could have added as much as 3.3 mg F daily.

A second case of SF from habitual consumption of large volumes of extra-strength instant tea calls for recognition and better understanding of a skeletal safety limit for this modern preparation of the world's most popular beverage.

Legend to Figure:

Anteroposterior radiograph of the lower thoracic and upper lumbar spine at age 48 years shows uniformly dense, but properly shaped, vertebrae.

Disclosures: M.P. Whyte, None.

This study received funding from: Shriners Hospitals for Children, The Clark and Mildred Cox Inherited Metabolic Bone Disease Research Fund, and the Barnes-Jewish Hospital Foundation.

W426

The Relationship between Bone Formation Rate and Bone Marrow Adipocytes in Pediatric Metabolic Bone Diseases. M. S. Cheung*, F. H. Glorieux, L. M. Ward, R. Travers*, F. Rauch. Genetics Unit, Shriners Hospital for Children, Montreal, PQ, Canada.

Adipocytes are derived from the same mesenchymal stem cells as osteoblasts. If mesenchymal stem cells differentiate into either adipocytes or osteoblasts depending on functional requirements, there should be an inverse relationship between the number of adipocytes in the bone marrow and bone formation rate on adjoining bone surfaces. Indeed, histomorphometric studies in adults seem to confirm this hypothesis. In the present study we examined whether a similar relationship between adipocyte numbers and bone formation rate is present in a range of pediatric metabolic bone disorders. Iliac bone samples from 71 children and adolescents with osteogenesis imperfecta (OI) -before medical treatment and after two years of pamidronate therapy (OI-PAM) — idiopathic juvenile osteoporosis (IJO), Crohn's disease and pseudohypoparathyroidism (PHP) were assessed for cancellous bone formation rate per bone surface (BFR/BS) and the presence of adipocytes. Results were compared to those of 58 controls who were individually age-matched to each disease cohort. 

Table  .  

*p<0.05; **p<0.01;***p<0.001 compared to controls

In untreated OI, both BFR/BS and the proportion of samples containing adipocytes were higher than in controls. Pamidronate treatment led to lower BFR/BS, but had no effect on the number of samples with adipocytes. In Crohn's disease and PHP, adipocytes were more prevalent than in controls, even though average BFR/BS was similar or higher. When all biopsy samples were examined, there was no difference in BFR/BS between samples with adipocytes present compared to those without adipocytes (p=0.40). In conclusion, no evidence for an inverse relationship between bone formation and the presence of adipocytes was found in the disease groups studied here. Factors other than bone formation rate seem to be more important determinants of adipogenesis in these conditions.

Disclosures: M.S. Cheung, None.

W427

Exostoses After Pediatric Stem Cell Transplant Are Associated with Growth Hormone Treatment. K. J. Dilley¹, K. Danner-Koptik*². ¹Pediatric Hematology-Oncology-Transplant, Children's Memorial Hospital, Feinberg School of Medicine, Chicago, IL, USA. ²Pediatric Hematology-Oncology-Transplant, Children's Memorial Hospital, Chicago, IL, USA.

Allogeneic hematopoeitic stem cell transplant (SCT) is a curative therapy for pediatric patients with both malignant and non-malignant diseases. Single or multiple benign exostoses or osteochondromas have been reported following total body irradiation (TBI) as well as focal irradiation. Patients exposed to TBI at a young age are at highest risk of developing exostoses. The objective of this study was to look at potential factors besides radiation that may play a role in development of exostoses. All patients who have undergone SCT at a single institution between March 1992 and October 1998 and who have had an exostosis identified by clinical findings or as an incidental finding on a radiologic study were included. A case-control design matched patients with controls who had the same stem cell source. 11/13 cases were matched to controls within the same age group at SCT of <4 yrs or ≥4 yrs but 2 others were not able to be so closely matched to unique controls. Treatment-related variables as well as physical and laboratory findings were collected from medical record reviews. Of 110 eligible patients, 13 (12%) were found to have exostoses at the time of data collection, 7 male and 6 female. All but two were transplanted for malignancy. All but one received TBI, that case did receive radiation but to a site remote from the exostosis. Multiple exostoses were noted in 62% (8/13). Treatment with growth hormone was more likely in cases (n=6) than in controls (n=2), p=0.046. Also, cases had a nonsignificant trend toward more negatively skewed height Z scores (mean −2.1 vs −1.5, p=0.056) and weight Z scores (mean −1.4 vs −0.2, p=0.097) at most recent follow-up than did controls. Patients available for use as controls tended to be older than cases, anecdotally supporting other studies that have reported younger age to be a risk factor for exostoses. No other SCT treatment factors such as HLA-mismatch or the occurrence of GVHD were associated with exostosis risk. Calcium (9.8 vs 9.9, p=0.75) and IGF-1 (382 vs 331, p=0.60) values were not significantly different at latest follow-up (cases at mean age 14.6 yrs vs controls at mean age 13.5 yrs), but IGF-BP3 approached significance with a mean of 5.0 in cases compared to 3.7 in controls (p=0.066). In conclusion, allogeneic SCT patients with the most extreme growth disruption and subsequent treatment with growth hormone appear to be at highest risk for development of exostoses. All reported samples to date have been small, use of multi-institution data to further examine risk factors is warranted.

Disclosures: K.J. Dilley, None.

W428

Postnatal Vitamin D Supplementation Normalized Neonatal Bone and Lean Mass Following Maternal Dietary Vitamin D Deficiency in the Guinea Pig. S. L. Finch, H. A. Weiler. School of Dietetics and Human Nutrition, McGill, Ste. Anne-de-Bellevue, PQ, Canada.

While vitamin D deficiency is common at birth, the consequences to growth and bone mass by weaning are unclear. This study was designed to determine if maternal dietary vitamin D deficiency in pregnancy has a negative impact on mineralization of full term infants and if postnatal supplementation would recover the limited bone mass using a 2-by-2 design. Forty pregnant guinea pigs were randomized to receive a diet with or without 1 IU vitamin D3/g diet. Blood was sampled in sows before mating at the start of the 3^rd trimester (d42) to coincide with fetal mineralization and postpartum to measure 25(OH)D as the marker for vitamin D status. At birth pups were randomized to receive 10 IU of cholecalciferol or a placebo of equal volume until weaning at d28. Diets and supplements were blinded using letter coding. Dual energy x-ray absorptiometry (DXA) was used to measure body composition, as well as bone mineral density (BMD), bone area and bone mineral content (BMC) of the whole body, tibia, femur and lumbar spine at birth and d28 of life. Main and interaction effects were detected using factorial ANOVA and post-hoc testing using Bonferroni tests. Weight of the pups was lower in the vitamin D deficient group at birth and d28 (Birth: 169.59± 97.94 vs 155.34± 97.89 g, P=0.042; d28: 279.13± 47.43 vs 255.91± 44.19 g, P=0.024). Main effects of the maternal diet showed vitamin D deficient offspring had lower whole body BMC (5.58±2.69 vs 5.24±2.52 g, P=0.012) and lower tibia BMC (0.15±0.07 vs 0.14±0.07 g, P=0.012) than the control group. Interaction effects showed that the deficient group were lower than the control group in whole body BMD (P=0.045) and lean mass (P=0.024) unless they received supplemental vitamin D. Additionally, femur BMC was lower at d 28 in the deficient males unless they received the supplement (P=0.012). Weight and whole body BMC increased 2.5 to 3 times from birth to weaning demonstrating the magnitude of growth that this novel model is capable of in a short period of time. In summary, postnatal vitamin D supplementation normalized neonatal bone and lean mass following maternal dietary vitamin D deficiency in the guinea pig.

Disclosures: S.L. Finch, None.

This study received funding from: Dairy Farmers of Canada and The Natural Sciences and Engineering Council of Canada.

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Hyperparathyroid Bone Disease in Two Sisters with Parathyroid Hormone Resistance (pseudohypopoarathyroidism 1b). R. C. Gensure¹, M. Bastepe². ¹Pediatrics, Ochsner Clinic Foundation, New Orleans, LA, USA. ²Endocrine Unit, Massachusetts General Hospital and Harvard Medical School, Boston, MA, USA.

Pseudohypoparathyroidism (PHP) is a disorder of hypocalcemia and elevated serum parathyroid hormone (PTH) levels caused by defective signaling via the stimulatory G protein α subunit (Gsα). PHP type-la results from mutations in Gsα-coding GNAS exons, and is characterized by multi-hormone resistance and a bone dysplasia syndrome called Albright's hereditary osteodystrophy (AHO). PHP-lb is caused by mutations that disrupt the imprinting of GNAS and is characterized by resistance to PTH (and occasionally to TSH) with no bone dysplasia. In each subtype, hormone resistance develops only after maternal inheritance of the genetic defect, consistent with the predominantly maternal expression of Gsα in some tissues, including the renal proximal tubule. We report on two sisters with PHP-lb, demonstrating both PTH- and TSH-resistance in the absence of AHO, seen in consultation because of increased fractures. The genetic defect in this family, a 4.4-kb deletion in STX16, has been reported previously (Am J Hum Genet. 2005 May;76(5):804-14). The elder sister (patient A, 15 9/12 y.o.) had one pathological fracture and was found to have a low lumbar bone mineral density (BMD) (Z-score −1.7). Importantly, the BMD did not show the expected increase with age, but rather decreased 12.6% over the past year. The younger sister (patient B, 14 0/12 y.o.) had two recent fractures and had a BMD closer to the normal range (Z-score −0.7), with a slight increase (2.6%) over the past year. Each had been managed with calcium and calcitriol according to the standard practice to maintain low-normal serum calcium levels to reduce risk of kidney stones; the PTH levels were elevated in both patients (272 pg/ml for patient A, 320 pg/ml for patient B). Gsα is not imprinted in bone; hence, while patients with PHP-la have a 50 percent reduction in Gsα signaling leading to partial PTH resistance in bone, patients with PHP-lb are predicted to have normal Gsα signaling in bone and could therefore develop hyperparathyroid bone disease. The patients' medications were adjusted to lower PTH levels to the normal range (57 pg/ml and 35 pg/ml, respectively). After 8 months, their bone mineral densities improved to Z=-0.9 and Z=-0.3, respectively (increased by ∼0.5 S.D. in each case), and they did not have any additional fractures. Neither patient developed hypercalcuria, likely because of the lack of imprinting of GNAS in the renal distal tubule. Given that bone is not resistant to PTH in PHP-lb, we recommend that these patients' serum PTH levels be maintained in the normal range to prevent hyperparathyroid bone disease.

Disclosures: R.C. Gensure, None.

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Physeal Closure from Chronic Vitamin A Intoxication. D. Wenkert¹, W. H. McAlister*², M. P. Whyte³. ¹Res Ctr, Shriners Hospt Children, St. Louis, MO, USA. ²Mallinckrodt Inst Radiol, Wash Univ Sch Med, St. Louis, MO, USA. ³Div Bone & Miner Dis, Wash Univ Sch Med, St. Louis, MO, USA.

Vitamin A excess causes periosteal calcification, diffuse idiopathic skeletal hyperostosis, and fractures. In pediatric patients, skeletal growth, modeling, and remodeling can be disrupted. Premature growth plate fusion from vitamin A and D toxicity was first reported in animal hind limbs (“hyena disease”). Subsequently, in 1962, 3 of 7 children with growth retardation from vitamin A toxicity had early lower limb physeal closure (JAMA; 182:980). Further reports concerned vitamin A derivatives used in severe pediatric dermatoses or cancers.

A 6-yr-old boy with a history of hypervitaminosis A, associated with liver fibrosis, was referred for “epimetaphyseal disease”. Birth weight was 10 lbs, and he walked at 10 mo. At that time, gastroenteritis was reportedly treated by a chiropractor with a dose of 3 million units (U) of vitamin A over 16 days, followed by 60,000 U/d except when intercurrent illness prompted ∼20 additional 3 million U, 16-day courses.

By age 2 1/2 yrs, fingernails reportedly “bubbled up and fell off”, speech regressed, and gross motor skills were lost. Hospitalization elsewhere revealed a critically ill, anorexic child with shiny, hairless, jaundiced skin, icterus, mouth sores, mucosal bleeding, pseudotumor cerebri, ataxia, and marked hepatosplenomegaly. Laboratory studies revealed a coagulopathy, hepatitis, anemia, and thrombocytopenia, but normal serum Ca, P, and Mg levels. Alkaline phosphatase was elevated at 827 IU/l. Hypervitaminosis A was diagnosed. After vitamins were stopped, he resumed walking and eating within 9 mo. Nevertheless, leg bowing, noted at age 3, worsened. Radiographs at age 5 showed physeal closure within distal femurs and proximal and distal tibias.

Upon referral, he had dysphagia, dyspnea, anemia, bleeding tendency, morning stiffness, knee and ankle pain, but took only vitamin K. Height was 5%, weight 75%, sitting height 50%, arm span 75%, and head circumference > >95%. He had petechia, bruises, telangiectasias, hypertelorism, cupped optic discs, and splenomegaly (17 cm) without hepatomegaly, 30° knee contractures, enlarged warm ankles and knees, and bowed tibias. Radiographs revealed the physeal closure, now with relative overgrowth of fibulae, and 1/2 yr later, closure of the proximal fibular physes. Physes elsewhere remained open.

Long-term follow-up of physeal fusion from vitamin A toxicity (eg. 12 yrs: JBJS[Am] 1974;56:1283) suggests disproportionate, assymetric shortening of lower limbs. Narrowing of physes from isotretinoin can resolve (Am J Dis Child 1988;142:316-8). Experience with our patient suggests that reversal of physeal fusion is not possible if vitamin A toxicity has been severe and prolonged.

Disclosures: D. Wenkert, None.

This study received funding from: Shriners Hospitals for Children, The Clark and Mildred Cox Inherited Metabolic Bone Disease Research Fund, and the Barnes-Jewish Hospital Foundation.

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The Importance of Biochemical Markers of Bone and Cartilage Turnover as the Predictors of Radiographic Progression in Patients with Early Rheumatoid Arthritis. J. Hashimoto¹, P. Garnero², N. Miyasaka*³, K. Yamamoto*⁴, S. Kawai*⁵, T. Takeuchi*⁶, N. Murata*⁷, H. Yoshikawa¹, N. Nishimoto*⁸. ¹Orthopaedic Surgery, Osaka University Graduate School of Medicine, Suita, Japan. ²INSERM Research Unit 664 and Synarc, Lyon, France. ³Tokyo Medical and Dental University, Tokyo, Japan. ⁴The University of Tokyo, Tokyo, Japan. ⁵Toho University Omori Medical Center, Tokyo, Japan. ⁶Saitama Medical Center/School, Saitama, Japan. ⁷Orthopaedic Surgery, Kyowakai Hospital, Suita, Japan. ⁸Osaka University Graduate School of Medicine, Suita, Japan.

Background: Rheumatoid arthritis (RA) is characterized by chronic arthritis leading to joint damage due to bone and cartilage destruction. Although early intervention of RA could prevent the progression of joint damages, it is difficult to predict the radiological progression in patients with early RA.

Objectives: To evaluate the predictive value of biological, radiological and clinical parameters for the prognosis of radiographic joint damage in RA patients treated with conventional DMARDs.

Methods: Among 302 patients with active RA of less than five years duration who participated in the prospective 1-year randomized controlled trial of tocilizumab monotherapy, termed SAMURAI study, the control arms (n=145), in which the dose regimens of the conventional DMARDs therapy excluding any biologics, were analyzed. Biochemical markers (bone turnover, cartilage turnover and inflammation), clinical score (number of swollen joint, tender joint, DAS score) and body mass index were measured at baseline and radiographic joint damage were scored at baseline, week 28 and 52 by two independent blinded readers according to the van der Heijde modified Sharp method.

Results: At baseline, mean age, disease duration, DAS28 score, total Sharp score and CRP were 53 years, 2.3 years, 6.5, 29.4 and 4.8 mg/dl, respectively. Increased urinary C-terminal crosslinking telopeptide (CTX-II) levels, higher urinary pyridinoline/deoxypyridinoline ratio (PYD/DPD), higher JSN score and low BMI at baseline were associated with increased radiological progression at 1 year both for erosion and the JSN score. When categorized into two or three groups of baseline risk factors (cut-off values: 500 and 1000 ng/mmol/creatinine for CTX-II, median for PYD/DPD, 0 unit or more for JSN, 18.5 and 25 for BMI), logistic regression analysis showed that these four variables were significant independent predictors of radiological progression (+0.5 unit increase of erosion or JSN score at week 52).

Conclusion: High baseline levels of biomarkers of bone and cartilage turnover including CTX-II and PYD/DPD, JSN score and low BMI are significantly and independently associated with 1-year progression of joint destruction in early RA.

Disclosures: J. Hashimoto, None.

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Tartrate-Resistant Acid Phosphatase Isoform 5a as a Marker of Disease Severity in Rheumatoid Arthritis. A. J. Janckila¹, D. H. Neustadt*², L. T. Yam*¹. ¹Medicine, VA Medical Center, Louisville, KY, USA. ²Medicine, University of Louisville, Louisville, KY, USA.

Objective: This study was undertaken to determine the clinical significance of tartrate-resistant acid phosphatase (TRACP) isoforms 5a and 5b as markers of chronic inflammation and cartilage/bone destruction in rheumatoid arthritis (RA).

Methods: 118 patients were recruited including 50 with RA (25 with subcutaneous nodules), 26 with osteoarthritis (OA) and 42 with other rheumatic diseases. 26 healthy adults served as controls. Serum TRACP-5a activity, TRACP-5a protein and TRACP-5b activity were determined by in-house immunoassays. C-reactive protein (CRP) was determined by in-house immunoassay using commercial antibodies and CRP. Other commercial markers included bone-specific alkaline phosphatase (bALP), C-telopeptides of type-I collagen (ICTP), cartilage glycoprotein-39 (YKL-40), and IgM rheumatoid factors (IgM-RF).

Results: TRACP-5a protein was elevated in RA and correlated only with IgM-RF. Amongst RA patients, TRACP-5a protein and IgM-RF were higher in those with nodules. TRACP-5a protein and IgM-RF tended to decline with disease duration, but not age in RA patients with nodules. In contrast, TRACP-5b activity was slightly elevated in RA and correlated with bALP and ICTP, but not with IgM-RF or CRP. TRACP-5b activity was not related to presence of nodules, however it increased with age and disease duration in RA.

Conclusion, TRACP isoforms may be useful disease markers in RA. TRACP-5a protein may be a measure of systemic inflammatory macrophage burden and a marker for disease severity. TRACP-5b activity is a marker for osteoclast number and may be useful to monitor local or systemic bone destruction in early RA.

Disclosures: A.J. Janckila, None.

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Association of tri-nucleotide (CAG and GGC) Repeat Polymorphism of androgen Receptor Gene in Taiwanese Women with Refractory or Remission Rheumatoid Arthritis. H. Kang¹, S. Yu*¹, Y. Hsu², C. Chang*³. ¹Graduate Institute of Clinical Medical Sciences, Chang Gung University, Kaohsiung, Taiwan. ²Program for Population Genetics, Harvard School of Public Health, Boston, MA, USA. ³Departments of Pathology, Urology, Radiation Oncology, Rochester University, Rochester, NY, USA.

To investigate the relationship between CAG and GGC repeat polymorphism of the androgen receptor (AR) gene and rheumatoid arthritis (RA) in female patients with different disease subtypes. This case-control study enrolled 215 females in three groups: RA patients refractory to standardized therapy (n=51); RA patients at complete remission phase (n=60); and healthy controls (n=104). CAG and GGC repeat lengths were determined by automated fluorescence-based DNA fragment-sizing method. Demographic data, allele lengths, allele distribution, and zygosity status of CAG/GGC repeats were assessed for the three group. Refractory RA patients tend to have a significantly younger onset age of RA and more elevated erythrocyte sedimentation rates than do remission RA patients. Mean and median values of CAG and GGC repeat lengths are similar in both RA and control patients. However, RA patients harboring any long CAG alleles with more than 23 repeats had an increased risk of a refractory course, whereas differences in risk were not observed between these patients and RA subtypes harboring any long GGC alleles with more than 16 repeats. In addition, the homozygous frequency of CAG but not GGC alleles was lower in refractory RA than in remission RA patients or in controls (p=0.042). Neither CAG nor GGC repeat lengths had a significant relationship with rheumatoid factor reactivity. Our observations indicate that short CAG repeats of the AR gene with higher transactivation activity may have protective effects against refractory course of RA development and that homozygous frequency of CAG alleles may be involved in the disease remission subtype. In contrast, lack of association of GGC polymorphism and RA was also observed. Together, these data imply that CAG but not GGC alleles in the AR polymorphism may play an important role in modulating the disease pattern of RA among Taiwanese women.

Disclosures: H. Kang, None.

This study received funding from: Chang Gung Memorial Hospital and University.

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The Effect of Etanercept on Bone Metabolism in Patients with Rheumatoid Arthritis. D. Kida*, Y. Eto*, M. Tsukamoto*, T. Sato*, A. Kaneko*, G. Ishihara*, H. Sugishita*, K. Saito*. Department of Orthopedic surgery and Rheumatology, National Hospital Organization Nagoya Medical Center, Nagoya, Japan.

Background: Recently, the discovery of the RANKL-RANK-OPG pathway contributed to a better understanding of bone diseases such as RA, osteoporosis, and bone metastasis. On the other hand, TNF has been shown to increase the expression of RANKL and act directly on osteoclasts by enhancing their resorpting activity. Furthermore, TNF enhances bone loss in RA by inducing osteoblast apoptosis. Therefore, the imbalance between bone resorption and bone formation leads to focal and systemic bone loss in RA. Although the most recent data suggest that infliximab therapy may inhibit generalized bone loss in patients with RA, it has not been studied in etanercept (ETN).

Purpose: The aim of this study was to evaluate urinary N-telopeptide of type-I collagen / creatinine (NTx) and serum bone specific alkaline phosphatase (BAP) in RA before and after treated with ETN.

Methods: Sixty postmenoposal females (mean age 62 yr; mean disease duration 14 yr) with RA who had not been treated for osteoporosis were divided into three groups: Twenty treated with bisphosphonate (BP), twenty treated without BP and twenty treated with ETN alone. To evaluate disease activity, ESR, CRP, DAS28ESR-4, modified Stanford Health Assessment Questionnaire (mHAQ) score, NTx, and BAP were measured in all patients at baseline, and after 6 months.

Results. However, ESR (p<0.01), CRP (p<0.001), DAS28ESR-4 (p<0.001), and mHAQ score (p<0.05) had decreased significantly, NTx levels did not differ significantly in 6 months after initial treatment with ETN. BAP levels (p<0.01) were significantly increased, and higher (p<0.01) in patients treated with ETN compared to others.

Conclusion. ETN therapy may not inhibit bone resorption but stimulate bone formation in patients with RA. This effect may be due to a beneficial roll of TNF-alpha blockade on bone turnover. Studies of longer duration are needed to assess the effect of ETN on bone metabolism.

Disclosures: D. Kida, None.

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Human Rheumatoid Synovial Fibroblasts Promote Osteoclastogenic Activity by Activating RANKL via TLR-2 and TLR-4 Activation. H. Kim*¹, S. Lee*¹, K. Kim*², M. Cho*², W. Park*³, H. Kim*². ¹Internal Medicine, Konkuk University Hospital, Seoul, Republic of Korea. ²Internal Medicine, Catholic University of Korea, Seoul, Republic of Korea. ³Internal Medicine, INHA University Hospital, Seoul, Republic of Korea.

Objective: The interplay between the innate immune system and inflammatory bone destruction in the joints of individuals with rheumatoid arthritis (RA) remains unclear. This study was undertaken to explore the effect of toll-like receptor (TLR) signaling in fibroblast-like synoviocytes (FLS) on the expression of RANKL and induction of osteoclastogenic activity.

Methods: The levels of RANKL mRNA and protein were measured using RT-PCR, realtime PCR, and immunostaining. Monocytes were cocultured with RA -FLS that had been stimulated with TLR ligands in fresh media and subsequently stained for tartrate-resistant acid phosphatase (TRAP) activity. Osteoclast molecule markers were measured using realtime PCR.

Results: Expression of TLR-2 and TLR-4 was higher in RA-FLS than in OA-FLS and normal skin fibroblasts. TLR-2 and TLR-4 ligands induced RANKL expression in RA-FLS. TLR stimulation of RA-FLS also induced the production of IL-1β and TNF-α to a lesser extent; however, it had no effect on IL-17 production. Inhibition of TLR induced IL-1β production, which partially reversed the upregulation of RANKL induced by TLR ligands. RA-FLS stimulated by TLR-2 and TLR-4 ligands and cocultured with human monocytes induced high levels of expression of TRAP, RANK, cathepsin K, calcitonin receptor, and matrix metalloproteinase-9, suggesting that RA-FLS promote osteoclast differentiation. Our results suggest that the TLR signaling pathway, through TLR-2 and TLR-4, induces RANKL expression in RA-FLS and the expression of RANKL promotes the differentiation of osteoclasts in RA synovium.

Conclusion: Targeting specific TLRs may be a promising approach to prevent inflammatory bone destruction in the pathogenesis of RA.

Disclosures: S. Lee, None.

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Alpha Lipoic Acid Attenuates Vascular Smooth Muscle Calcification via the Expressional Regulation of Genes Related to Apoptosis and Oxidative Stress. E. Kim*¹, I. Lee*¹, H. Kim*², W. Jang*¹, I. Park¹. ¹Kyungbook National University, Daegu, Republic of Korea. ²Keimyung University, Daegu, Republic of Korea.

Vascular smooth muscle cell (VSMC) calcification is an important risk factor for cardiovascular diseases. It is reported that alpha lipoic acid (ALA) has many beneficial effects on vascular cells in atherosclerosis. The present study was designed to investigate whether ALA can prevent VSMC calcification. We explored the effects of ALA on human VSMC calcification in vitro and found that ALA attenuates VSMC calcification induced by high phosphate conditions as quantified by the o-cresolphthalein complexone method. ALA also inhibits vitamin D3-induced aortic calcification in vivo. Using DNA microarray analysis and secretome analysis using LC/MS/MS, we identified genes and proteins whose expression was changed by high phosphate conditions and ALA. Proteins up-regulated by high phosphate condition that were subsequently down-regulated in the presence of ALA were related to apoptosis and oxidative stress. Gene expression analysis using DNA microarray tools confirmed the up-regulation or down-regulation of some, but not all, of the proteins observed in ALA challenged, in high phosphate condition. The present data suggest that vascular calcification is inhibited by ALA through modulation of apoptosis and oxidative stress pathway and provide valuable information about the underlying mechanisms of atherosclerosis.

Disclosures: I. Lee, None.

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Survey on the Effect of Immunomodulatory Therapy on Bone in Multiple Sclerosis. M. J. McKenna¹, M. Shuhaiber*², K. Foy*², R. Lonergan*², M. AuYeong*¹, J. Brady*³, B. Murray*³, J. M. T. Redmond*². ¹DXA Unit, St. Vincent's University Hospital, Dublin 4, Ireland. ²Neurology Department, St. James' Hospital, Dublin 8, Ireland. ³Metabolism Laboratory, St. Vincent's University Hospital, Dublin 4, Ireland.

Osteoporosis is a complication of multiple sclerosis (MS) especially if corticosteroid therapy is given. Little is known about the effect on bone of immunomodulator therapy for MS. We sought to evaluate bone mass in patients with MS on immunomodulators. We performed a cross-sectional survey and a follow-up survey.

At baseline, we studied bone mineral density (BMD) by dual energy X-ray absorptiometry (DXA) in 37 patients with MS who received immunomodulators. The BMD was measured at two sites: the lumbar spine (L1 to L4) and the left total femur site, using a Hologic QDR4000 Elite Densitometer (Bedford, MA). In our DXA unit, the least significant change at the 95% level is 3.2% at the spine and 2.5% at the femur.

Different immunomodulators were administered: Interferon beta — la in 70%, Interferon beta-lb in 27% and Glatiramer in 3%. High-dose pulse steroid therapy (intravenous methylprednisolone 500 mg) was given to 81% ranging from one course to 17 courses. The mean age was age 38.8 ± 8.9 years, the mean duration of MS was 5.8 ± 3.7 years; the mean number of relapses was 4.0 ± 0.7; and the mean disability score (EDSS) was 3.1 ± 1.9. Both mean BMD Z-score at spine of 0.53 (CI: 0.15-0.92; p<0.001) and mean BMD Z-score at femur of 0.72 (CI: 0.42-1.01; p<0.0001) were significantly greater than zero.

A follow-up survey after nearly 4 years of continuous immunomodulatory therapy is now being conducted. BMD is being remeasured. In addition, biochemical studies are being performed including: vitamin D status; parathyroid status; and bone turnover markers (serum bone specific alkaline phosphatase, intact osteocalcin, procollagen type I aminopropeptide, beta-carboxyterminal telopeptide of type I collagen; and, urine aminoterminal telopeptide of type I collagen, and deoxypyridinoline crosslinks). Preliminary analysis of DXA in 7 subjects shows no change in BMD at the spine (1.046±0.16 g/cm² vs 1.051±g/cm²; p =0.58) and at the femur (1.024±0.20 g/cm² vs 1.02±0.20 g/cm²; p=0.46).

In conclusion, immunomodulator therapy in MS does not appear to have an adverse effect on bone.

Disclosures: M.J. McKenna, None.

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Prevalence of Radiographic Osteoarthritis of Knee and Lumbar Spine and Its Association with Pain: The Research on Osteoarthritis Against Disability (ROAD) Study. S. Muraki¹, N. Yoshimura¹, H. Oka*¹, A. Mabuchi*¹, Y. En-yo*², M. Yoshida*², T. Suzuki³, H. Yoshida³, H. Ishibashi*³, S. Yamamoto³, H. Kawaguchi¹, K. Nakamura¹, ¹22nd Century Medical Ctr., Univ. of Tokyo, Tokyo, Japan. ²Wakayama Medical Univ., Wakayama, Japan. ³Tokyo Metropolitan Institute of Gerontology, Tokyo, Japan.

Although osteoarthritis (OA) of knee and lumbar spine is a major cause of disability in the elderly, few epidemiologic studies have been performed. We established a large-scale nationwide clinical study called ROAD (Research on Osteoarthritis Against Disability) in 2005, and created a comprehensive and systemic database including clinical and genetic information in three cohorts of urban, mountainous and seacoast areas. We recruited 3,040 participants in total, from which 2,288 subjects older than 60 years (818 men & 1,470 women; mean age=74.3 yrs.) were enrolled for investigation of the prevalence of radiographic OA of knee and lumbar spine, as well as its association with the respective local pain. The radiographic severity of OA was determined according to the Kellgren/Lawrence (KL) grade (0-4) at femoral-tibial joints of bilateral knees and at intervertebral spaces from L1/2 to L5/S1 of the lumbar spine. Prevalence of radiographic OA (KL> or = 2) in either knee joint was 47.0% in men and 70.2% in women, while that in either intervertebral space was 84.1% in men and 70.7% in women. Prevalence of radiographic knee OA was higher in the female sex (odds ratio [OR]=3.28, 95% confidential interval [CI]=2.71-3.97) and mountainous residents (OR=1.62, 95%CI=1.44-1.83 compared to urban residents), whereas that of lumbar OA was higher in the male sex (OR=2.06, 95%CI=1.63-2.61) and urban residents (OR=1.43, 95%CI=1.13-1.83 compared to mountainous residents). We next compared the association between radiographic severity and local pain by logistic regression analysis after adjustment for age and BMI. The radiographic severity of knee OA was positively associated with knee pain (OR=6.7, 95%CI=4.10-11.2 in men; OR=4.3, 95%CI=3.07-6.02 in women; for KL3/4 compared to KL0/1), while that of lumbar OA was weakly associated with low back pain at all spaces in women (OR=1.6-2.1 for KL3/4 compared to KL0/1), but not in men. The initial investigation in the ROAD study using the baseline data revealed a high prevalence of radiographic OA in knee and lumbar spine among elderly people, and found that gender and community differences were distinctly associated with the knee and lumbar OA. Furthermore, the radiographic knee OA showed a strong association with knee pain in both genders. Contrarily, the radiographic lumbar OA had a moderate association with low back pain only in women. With the progress of the ROAD study, the underlying environmental and genetic backgrounds will be elucidated.

Disclosures: S. Muraki, None.

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Treatment with Anti-TNF-α Antibody Recovers Osteoclast Maturation Deviated from Bone Remodeling Cycle in RA Joint Destruction. M. Nawata*, Y. Okada*, K. Saito*, Y. Tanaka*. The First Department of Internal Medicine, School of Medicine, University of Occupational and Environmental Health, Japan, Kitakyusyu, Japan.

Rheumatoid arthritis (RA) is characterized by inflammatory synovitis, erosive arthritis and joint destruction. Recent evidence indicates that TNF-α plays a pathological role in joint destruction and that infliximab (anti-TNF-α antibody) provides significant protection against the destructive process, even in patients who show no clinical improvement, suggesting the dissociation of these two disease processes. However, the mechanism of bone destruction in RA remains unknown. Here we assessed bone metabolism in RA patients before and after infliximab treatment. RA patients (n=100, mean age, 53.6±12.1 years, ±SD) were treated with either methotrexate (MTX) or infliximab+MTX. We evaluated DAS (Disease Activity Score)-28, bone metabolism markers and bone mineral density (BMD) at 0, 6, 12 and 24 months after treatment. At enrollment, BMD was within the normal aged-adjusted level (L2-4; 0.89±0.18 g/cm², left hip; 0.67±0.14 g/cm²), as was serum bone alkaline phosphatase (BAP; 26.9±10.5 U/I), a bone formation marker, but urinary crosslinked N-telopeptide of type I collagen (u-NTx), a bone resorption marker, was notably high (76.4±37.7 nmol/mmol CRE). Treatment with infliximab resulted in the following: i) No change in BMD for 2 years (L2-4 BMD 0.90±0.22, left hip 0.68±0.11 g/cm²). ii) Decrease in u-NTx to 61.3±32.1 (p<0.05), 48.8±27.0 (p<0.01) and 42.3±18.3 nmol/mmol CRE (p<0.01) at 6, 12 and 24 months, respectively. iii) Increase in BAP to 31.6±11.5 (p<0.05), 31.9±11.7 (p<0.05) and 29.2±11.2 U/I at 6, 12 and 24 months, respectively, iv) These changes were independent of the dose of steroids or disease activity. vi) Improvement of bone erosion was noticeable on the joint X-p after 24 months. BAP and u-NTx did not change in patients treated with MTX alone. Bone metabolism is normally maintained by bone remodeling, bone resorption by osteoclast and formation by osteoblasts, respectively. In contrast, we here propose that joint destruction in RA is induced by osteoclast maturation in “deviation” from the remodeling cycle in a TNF-dependent manner, based on the followings: i) increase in u-NTx with normal BMD and BAP levels, ii) TNF-α and/or sRANKL induce osteoclast maturation in the absence of osteoblasts in RA synovium, iii) joint erosion is observed within 10 weeks in TNF-α-transgenic mice. Furthermore, infliximab treatment allowed recovery from the “deviation” into the remodeling cycle as evident by the decrease in u-NTx and increase in BAP. These results imply the dissociated efficacy of infliximab for joint destruction from clinical improvement, which leads to repair of RA bone erosion.

Disclosures: M. Nawata, None.

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Evaluation of the Bone Mineral Density and Body Composition in Early Rheumatoid Arthritis — The OsteoPorosis and Early Rheumatoid Arthritis (OPERA) Study. A. C. Azevedo*¹, M. M. Pinheiro¹, K. S. Sarkis*², E. T. Reis Neto*¹, L. A. Martini², V. L. Szejnfeld¹. ¹Rheumatology, Unifesp, Escola Paulista de Medicina, Sao Paulo, Brazil. ²Nutrition, Faculdade de Saude Publica, Universidade de Sao Paulo, Sao Paulo, Brazil.

BACKGROUND: Traditional and not traditional risk factors for osteoporotic fractures have been described in patients with rheumatoid arthritis (RA), like menopause, smoking, glucocorticoids, immobilization and bone mineral density (BMD). Recently, some authors have showed that aspects related to body composition could also be associated with bone loss. OBJECTIVES: To evaluate bone mineral density (BMD) and body composition in patients with early rheumatoid arthritis (ERA) before and after the treatment. PATIENTS AND METHODS: Thirty-two RA patients (ACR, 1987) with time of symptoms less 15 months and without previous treatment with DMARDs (disease-modifying anti-rheumatic drugs) were enrolled in this study. Patients using medications that interfered on bone mass were excluded. Details on lifestyle, demographic data, fractures, dietary intake, physical activity, quality of life were evaluated by a specific questionnaire. Disease's characteristics and activity (DAS28) and current treatment of RA were also evaluated. Moreover, it was applied SF-36 and HAQ (Health Assessment Questionnaire). All the patients underwent bone densitometry (spine and femur BMD) by DXA (GE-Lunar, DPX MD+). Body composition was measured by DXA (lean, fat and bone mass). Descriptive statistical analysis, chi-squared test and logistic regression were used. P<0.05 was adopted. After 12 months, the questionnaires and DXA measurements were repeated. RESULTS: Mean age and BMI were 43.4±11.8 years and 24.7±6.5 kg/m², respectively. Women were 26 (83.9%) and mixed race were 13 (41.9%). At baseline, disease duration, DAS28 and HAQ was 7.97±4.5 months, 5.41±1.45 and 0.896±0.605, respectively. Corticosteroids were referred by 21 patients (67.7%) with mean dosage 7±8,9 mg/ day. Spine and total femur BMD was 1.068±0.15 and 0.987±0.12 g/ cm², respectively. Total and skeletal lean mass was 37.180±6.4 kg and 6.34±0.9 kg/ m², respectively. Fat mass was 36.8±6.9%. After 12 months of follow-up, we did not verify significant bone or lean loss. On the other hand, there were significant reduction of the disease's activity and significant increase of functional ability and fat mass. CONCLUSIONS: We observed low prevalence of osteopenia/ osteoporosis and sarcopenia in patients with ERA at baseline. Although the patients were considered as euthrophic by BMI, there was higher percentage of fat mass than lean mass between 2 visits. Thus, our data suggest that health interventions may emphasize more prevention of fat mass increase than lean or bone mass loss in ERA patients.

Disclosures: M.M. Pinheiro, None.

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Bone Metabolism in an Animal Model of Ankylosing Spondylitis. M. Rauner*¹, D. Stupphann*¹, J. Patsch*¹, M. Haas*², M. Breban*³, I. Fert*³, S. Glatigny*³, P. Pietschmann¹. ¹Institute of Pathophysiology, Medical University of Vienna, Vienna, Austria. ²Department of Internal Medicine III, Medical University of Vienna, Vienna, Austria. ³Institut Cochin, Hǒpital Cochin, Paris, France.

Ankylosing Spondylitis (AS) is a chronic inflammatory disease characterized by the main involvement of axial joints and bilateral sacroiliitis. Besides the formation of new bone leading to syndesmophytes and ankylosis, osteoporosis is a reported feature as well. Thus, AS raises the paradox of new bone formation at sites of inflammation and the association of reduced bone mass and high fracture risk. The major histocompatibility complex class I allele HLA-B27 has been found to be strongly associated with AS. HLA-B27 transgenic (tg) rats spontaneously develop a multisystemic disease with the appearance of spondylitis and syndesmophytes. However, there is no information about bone metabolism in this animal model. Hence, the aim of our study was to investigate the suitability of the HLA-B27 tg rat as a model for the bone-related alteration in AS. Therefore, bone histomorphometry was conducted for the femur, tibia and fourth vertebral body from six control rats, six healthy HLA-B27 tg rats and six HLA-B27 tg rats exhibiting the phenotype of spondyloathropathies. Each group consisted of two male and four female rats aged between seven and ten months. Sections, stained with Goldner's trichrome, were analyzed using the Osteomeasure Software (Osteometrics, USA). Femur, tibia and the fourth vertebral body of the diseased HLA-B27 tg rats exhibited a significant reduction of bone volume over total volume compared with control rats and healthy tg rats. Moreover, trabecular thickness and trabecular number of the diseased rats were significantly reduced. The trabecular separation was significantly increased. The number of osteoblasts and osteoid volume did not show any alterations, however, the number of osteoblasts as well as osteoid volume were higher in femur and tibia than in the vertebral body. Osteoclasts tended to be more abundant in the diseased HLA-B27 tg rats. The decrease in bone volume, trabecular number and thickness as well as the increase of trabecular separation in the diseased HLA-B27 tg rats indicate an enhanced bone resorption, possibly mediated by the increased number of osteoclasts. Moreover, the indices of osteoid volume show, that the mineralization process in the diseased rats was not disturbed. Our results demonstrate that HLA-B27 tg rats is an appropriate model for the study of bone loss in AS. In further experiments, we will analyze markers of bone formation and resorption in the serum and bone of HLA-B27 tg rats to assess bone turnover on a systemic level.

Disclosures: M. Rauner, None.

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Osteoporosis and Rheumatoid Arthritis, Which Nutritional Factors Are Associated?, K. S. Sarkis*¹, R. G. Silva*², C. F. Zerbini*², M. M. Pinheiro³, L. A. Martini¹, ¹Nutrition, Universidade de Sao Paulo, Sao Paulo, Brazil, ²School of Medicine, Universidade de Sao Paulo, Sao Paulo, Brazil, ³Reumathology, UNIFESP, Sao Paulo, Brazil.

The aim of this study was investigate the association between diet, body composition and Bone Mineral Density (BMD) in women with Rheumatoid Arthritis (RA). This is a cross-sectional study performed at RA outpatient clinic and included forty-five (45) patients who fulfilled the ACR criteria for RA and agreed to participate. Mean age was 55 ± 10 y and the mean duration of disease was 7.5 ± 5.6y. The patients were divided in two groups according to BMD status: group 1 — normal BMD (n=24); group 2 — patients with osteoporosis (n=21) (WHO 1994). Standardized questionnaires were used to ascertain demographic characteristics, medical history and use of oral glucocorticosteroids. Functional status was assessed using the Health Assessment Questionnaire (HAQ) score and the disease activity by the Disease Activity Score (DAS28). A validated food frequency questionnaire was used to assess the dietary intake. Mean daily intake, including total energy (kcal/day) and all others macronutrients, calcium, phosphorus and vitamin D was calculated and adjusted for energy. Percentage Body Fat (%BF), Total Fat Free Mass (TFFM), Appendicular Lean Mass (ALM), Relative Skeletal Muscle Index (REM1) and BMD was measured at the total femoral, femoral neck and lumbar spine (L1-L4) in g/cm2 by using DXA (GE Lunar Radiation Corporation, DPX IQ, Madison, WI, USA). Statistical analysis included Student T test, Pearson correlation and multiple linear regression. BMD was considered dependent variable. Significance was adopted as p<0.05. There were no significant difference in HAQ, DAS 28 and use of oral glucocorticosteroids between groups. Also no differences were observed in total energy, protein, carbohydrate, calcium, phosphorus and vitamin D intake. However both groups presented mean calcium and vitamin D intake lower than recommended values (597.3 ± 281.7mg/day vs 692.2 ± 217.1 mg/day and 60.7 ± 56.9 vs 88.1 ± 89.6 UI/day, group 1 and 2 for calcium and vitamin D, respectively). Patients with lumbar or femoral osteoporosis were significantly older (p<0.01), had a lower %BF (p<0.05), FFM (p<0.01), ALM (p<0.01), RSMI (p<0.01), longer disease duration (p<0.05) and a higher lipid intake (p<0.05). In the regression linear model, being older with more than 7.5 years with AR, lower %BF, fat free mass, appendicular lean mass together with higher lipid intake, were negatively associated with BMD [Femural total = 0.637–0.008 (age) −0.009 (disease duration) + 0,073 (RSMI) + 0,006 (%BF), with R²=66.8;p<0.001] The present study indicated two important modifiable factors that could improve BMD in patients with RA; the body composition and a reduction in fat intake.

Disclosures: K.S. Sarkis, None.

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Vitamin D Receptor Deficiency Aggravates TNF-Mediated Arthritis. K. Polzer*¹, J. Zwerina*¹, K. Redlich*², J. Smolen*², G. Schett*¹. ¹University of Erlangen-Nuremberg, Erlangen, Germany. ²Medical University of Vienna, Vienna, Austria.

Background: Vitamin D is an important hormone for calcium homeostasis and bone metabolism and may have immunomodulatory effects. The aim of this study was to investigate the role of the Vitamin D Receptors (VDR) in inflammatoryarthritis.

Methods: To determine the role of the Vitamin D in TNF-mediated joint disease, we crossed mice deficient for the VDR with arthritic human TNF transgenic (hTNFtg) mice. Offsprings were analysed for clinical signs of arthritis as well as the histological degree of synovitis, cartilage damage and bone erosion. Systemic mineral bone density was evaluated by bone histomorphometry. Furthermore, serum levels of cytokines and other inflammatory mediators were determined by ELISA.

Results: hTNFtg mice with VDR deficiency showed pronounced weight loss, increased paw swelling and decreased grip strength compared to hTNFtg mice. In addition, VDR-/-hTNFtg mice showed lower bone mineral density compared to hTNFtg mice. In line, serum levels of TNF were significantly increased in VDR-/-hTNFtg mice compared to hTNFtg mice. Consequently, histological analysis showed an increase in synovial inflammation in VDR-/-hTNFtg mice compared to hTNFtg mice. The VDR-/-hTNFtg mice showed a dramatic increase in osteoclast number compared to hTNFtg mice, accompanied with strengthened bone erosions in the joints. These data were verified in osteoclast assays of the four genotypes in vitro. Cartilage proteoglycan loss was also enhanced in VDR-/-hTNFtg mice compared to hTNFtg mice.

Conclusion: These findings suggest that deficiency of the VDR leads to an increase in inflammation, cartilage destruction and bone erosion with enhanced recruitment of osteoclasts. This leads to the assumption that additional treatment with active metabolites of Vitamin D may have potentially positive effects as an additive therapy in RA.

Disclosures: G. Schett, None.

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Bone Microarchitecture as Assessed by High Resolution Peripheral Quantitative CT (HRpQCT) in HIV+ Postmenopausal Women. A. D. Shu*¹, M. T. Yin¹, D. J. McMahon*¹, S. Cabral*¹, H. Eisenberg*¹, D. Ferris*², E. Shane¹. ¹Columbia University, NY, NY, USA. ²Bronx-Lebanon, Bronx, NY, USA.

As a result of effective antiretroviral therapy (ART), more HIV+ women are surviving beyond menopause and are at increased risk for osteoporosis (OP). We previously reported lower BMD but similar rates of bone loss in HIV+ PM Hispanic women compared to HIV-controls. There are no studies on bone microarchitecture and limited fracture data in HIV+ individuals. We hypothesized that lower BMD in HIV+ women is associated with microarchitectural deterioration. Xtreme CT (Scanco Medical AG) is a new HRpQCT system with sufficiently high resolution (<100μm) to permit in vivo assessment of trabecular (Tb) architecture at the radius (RAD) and tibia (TIB). DXA and HRpQCT were performed in 32 HIV+ (75% on ART) and 38 HIV- Hispanic and African American PM women from outpatient clinics, excluding women with known OP, metabolic bone disease and serum Cr > 2.5 mg/dL. HIV+ and HIV- groups were similar with regard to age (mean 60 y), prevalence of hypertension, diabetes, thyroid disease, and use of HRT, glucocorticoids and antiepileptics. HIV+ women were more likely to use tobacco. BMI was higher in HIV- women (31 v 28, p = 0.04). In contrast to our previous findings in Hispanics, mean BMD did not differ at any site. Similarly, HRpQCT measurements of 13 cortical (Cort) or Tb microstructural parameters did not differ between groups (Table). Analyses stratified by HIV status and race and adjusted for covariates of age, BMI and tobacco use were compared by repeated measures ANCOVA. No differences between HIV and race-defined groups were observed at the TIB. At the RAD, Hispanics had higher D100 (average bone density) (p<0.003) and Cort thickness (p<0.03); however, HIV status was not associated with any differences. In summary, DXA and HRpQCT parameters did not differ between HIV+ and HIV- PM women. The groups may be similar because the controls are drawn from a clinic population and many have chronic illnesses common in older minority women. The similarities in microarchitecure may also explain why retrospective studies of HIV+ patients on ART have not found that fracture rates are higher than expected.

Table Table. Mean HRpQCT and DXA values (± SD) with reference values*

* DXA reference NHANES/Hologic. HRpQCT reference in Caucasian women from Boutroy et. al. JCEM 90:6508–6515. ^†PreM, premenopausal or T ≥ 1.0; LBM, low bone mass or T-score < 1.0; OP, T-score ≤ 2.5.

Disclosures: A.D. Shu, None.

This study received funding from: R0IAI065200.

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OPG Prevents Osteopenia in Arthritic Rats Via Different Mechanisms than Bisphosphonate Therapy. M. Stolina, D. Dwyer*, S. Middleton*, S. Adamu*, M. S. Ominsky, D. Zack*, P. J. Kostenuik. Amgen Inc., Thousand Oaks, CA, USA.

RANKL was recently suggested to be a mediator of systemic bone loss in arthritic rats (Stolina et al, JBMR 2005). The catabolic effects of RANKL are inhibited by osteoprotegerin (OPG), and the RANKL:OPG ratio may be an important determinant of bone resorption. Skeletal catabolism can also be prevented by bisphosphonates, which can exert modest changes in OPG levels in patients and in osteoblast cultures. However, local and systemic changes to the RANKL:OPG ratio have not been described with BP treatment in an osteopenia model. We therefore compared the effects of OPG versus zoledronic acid (ZA) on serum and bone proteins and BMD in a rat arthritis model.

Adjuvant arthritis (AIA) was induced in rats by heat-killed mycobacteria. AIA rats were treated with human OPG-Fc (1 mg/kg), ZA (0.1 mg/kg), or PBS (Veh) at the clinical onset of arthritis (day 0), day 3, and day 6. Non-arthritic (NA) controls were treated with Veh. BMD was assessed at the lumbar vertebrae (LV) by DXA on day 9 when serum was collected and total protein was extracted from LV for evaluation of OPG, RANKL, and the resorption marker TRAP-5b.

AIA-Veh rats had lower LV BMD compared to NA controls (P < 0.05). Histology showed no evidence of inflammation in the LV of AIA-Veh rats, so osteopenia may have been related to an 80% increase in serum RANKL (P < 0.05 vs NA controls). Osteopenia was fully prevented by OPG or ZA treatment. Whereas OPG treatment prevented the AIA-related increase in serum RANKL, ZA treatment led to a further non-significant RANKL increase (vs AIA-Veh) and had no effect on serum OPG levels. The serum RANKL:OPG ratio was markedly reduced by OPG treatment (P < 0.05, vs AIA-Veh), and non-significantly increased by ZA (vs AIA-Veh). Serum TRAP-5b was significantly reduced by both treatments (P < 0.05 vs. AIA-Veh), more so with OPG (P < 0.05 vs ZA).

The RANKL:OPG ratio in LV protein extracts was significantly increased in AIA-Veh rats (P < 0.05 vs NA controls). This increase was primarily related to local suppression of OPG. OPG treatment significantly reduced LV RANKL, whereas ZA caused a 2-fold increase in LV RANKL (P < 0.05 vs AIA-Veh). OPG treatment markedly increased LV OPG levels, while ZA caused a modest reduction in LV OPG (vs AIA-Veh). The RANKL:OPG ratio was thereby reduced with OPG treatment, while ZA led to a 5-fold increase (both P < 0.05 vs AIA-Veh). LV TRAP-5b was significantly reduced by both treatments, to a greater extent with OPG (P < 0.05 vs ZA).

In summary, two potent agents of different therapeutic classes fully prevented systemic bone loss in arthritic rats. RANKL inhibition by OPG reduced the RANKL:OPG ratio in bone whereas ZA increased this ratio, highlighting the different mechanisms of action of these therapeutics.

Disclosures: M. Stolina, Amgen Inc. 1, 3.

This study received funding from: Amgen, Inc.

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Increased Expression of Receptor Activator of NF-kappaB Ligand (RANKL) in Stimulated T-cells from Patients with Ankylosing Spondylitis. D. Stupphann*¹, M. Rauner*¹, D. Krenbek*¹, J. Patsch*¹, T. Pirker*², C. Muschitz*², H. Resch*², P. Pietschmann¹. ¹Medical University of Vienna, Vienna, Austria. ²St. Vincent Hospital, Medical Department II, Vienna, Austria.

Ankylosing spondylitis (AS) is a chronic, disabling rheumatic disease, which initially mainly involves the sacroiliac joints and in later stages affects the vertebral column. Besides new bone formation leading to syndesmophytes and ankylosis, osteoporosis is a reported feature as well. Based on the hypothesis, that activated T cells are the major source of RANKL in inflammatory bone-resorptive diseases, we investigated the presence of intracellular (ic) and membrane bound (mb) RANKL on activated T cells from patients with ankylosing spondylitis (AS). Further we determined their relationship to bone mineral density (BMD), disease activity and bone metabolism markers. T-cell surface and intracellular expression of RANKL was analyzed by flow cytometry. BMD at the lumbar spine and total hip was measured by dual-energy x-ray absorptiometry (DXA) and quantitative computed tomography (QCT). Serum OPG levels were determined by ELISA. Expression of icRANKL was statistically significantly increased in patients with AS compared to the healthy volunteers (p < 0.05). In contrast expression of mbRANKL was significantly lower in the group of patients compared to the controls (p < 0.01). The cytokine expression was not associated with bone loss that was found with DXA at the total hop in 45% of all cases, and with QCT at the lumbar spine in 48% of all cases. Spinal BMD values measured by QCT were positively correlated with age (r = −0.630; p = 0.003) and negatively correlated with serum OPG levels (r = −0.571; p = 0.025). We conclude that intracellular RANKL in T cells is overexpressed in patients with AS, indicating that these cells may act as a storage for soluble RANKL which rapidly can be mobilized and secreted into the circulatory system after an inflammatory stimulus. In contrast, the expression of mbRANKL is significantly decreased in the AS patients, that might be due to a different activity or prevalence of the enzymes, responsible for the ectodomain shedding of RANKL.

Disclosures: D. Stupphann, None.

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Advanced Glycation-end-products Induce Vascular Calcification Through the RAGE/p38 MAPK Signaling Pathway. T. Tanikawa, R. Tanikawa, Y. Okada, Y. Tanaka. First Department of Internal Medicine, University of Occupational and Environmental Health, Japan, Kitakyushu, Japan.

Mönckeberg's calcification in type 2 diabetes, known as medial artery calcification is a strong independent predictor of cardiovascular mortality. However, the mechanism of vascular calcification in diabetes remains to be elucidated. Advanced glycation-end-products (AGEs), long-term products of the Maillard reaction, are formed at an accelerated rate in diabetes. They are thought to contribute to the development of diabetic complications, including atherosclerosis and microvascular diseases. In our in vitro studies, AGEs induced calcification of cultured vascular smooth muscle cells by von kossa staining and also induced the expression of Runx2 mRNA by RT-PCR, known as a transcription factor in osteoblast differentiation. Furthermore, AGEs increased alkaline phosphatase (ALP) activity and osteocalcin secretion, two osteoblast differentiation markers. Finally, AGEs induced phosphorylation of p38 MAPK and the phosphorylation was inhibited by anti-RAGE blocking antibody, which was assayed by western blotting. Increased ALP activity was inhibited by a p38 MAPK inhibitor. Our findings indicate that AGEs induce vascular calcification by osteoblast-like differentiation of smooth muscle cells and through the RAGE/p38 MAPK signaling pathway and suggest that inhibition of the AGE/RAGE pathway could be potentially effective therapeutically for prevention of vascular calcification in diabetic patients.

Disclosures: T. Tanikawa, None.

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Type 2 Diabetic Patients with Charcot Arthropathy Display Reduced Calcaneal Stiffness Despite Normal Foot BMD. K. A. Witzke¹, H. K. Parson*², A. I. Vinik*². ¹Kinesiology, California State University, San Marcos, San Marcos, CA, USA. ²Internal Medicine, Eastern Virginia Medical School, Norfolk, VA, USA.

Because bone is innervated it is subject to degradation when the nerves that innervate it lose their function. Charcot neuroarthropathy is a complication of diabetes associated with severe nerve dysfunction that often leads to unusual fractures that collapse the ankle and/or midfoot. These fractures may be caused by declines in bone mineral density (BMD), although BMD of the feet is not a common evaluation site. We believe that Charcot probably compromises the quality of bone, specifically its microarchitecture, crystalinity, and collagen properties, and these changes may not be completely explained by BMD alone. Calcaneal stiffness, measured by Quantitative Ultrasound (QUS), is a simple method of determining the stiffness of bone, an important biomechanical property related to bone quality and fracture. The purpose of this preliminary study was to determine if poor nerve function is related to reduced calcaneal stiffness and/or reduced BMD of the feet, and whether type 2 diabetics with Charcot display significantly different bone stiffness and BMD than either controls or type 2 diabetics without Charcot. We measured BMD of the whole body, hips, and feet (BMD, GE Lunar Prodigy), Calcaneal stiffness (QUS, GE Achilles Express), peroneal nerve conduction velocity (NCV), vibration detection threshold (VDT), and pressure perception (PRES), in 56 male subjects (n=22 controls; n=31 type 2 diabetic; n=3 diabetic patients with Charcot) aged 56.1 +/- 8.4 y. Diabetics averaged 127.4 +/- 80.4 months of diabetes. As expected, all nerve function measures were significantly worse in Charcot patients than the other two groups. For bone measures, Charcot patients displayed significantly lower calcaneal stiffness but similar BMD measures at all sites including the feet, compared to the diabetic and control groups. This may suggest a direct relationship between motor nerve dysfunction and the structural properties of bone in Charcot neuroarthropathy. Although the Charcot patients in this study were fairly homogeneous, these assumptions need to be tested in more patients, and a more direct measure of bone collagen would also be useful. 

Table  .  

* p < 0.05 for Charcot compared to Control and Diabetic

Disclosures: K.A. Witzke, None.

This study received funding from: Commonwealth Health Research Board.

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Morphological Texture Analysis of Radiographs of the Proximal Femur: In Vitro Study Using Biomechanical Strength as a Standard of Reference. M. B. Huber*¹, J. Carballido-Gamio*¹, K. Fritscher*², R. Schubert*², M. Haenni*³, C. Hengg*⁴, S. Majumdar*¹, T. M. Link*¹. ¹University of California, San Francisco, San Francisco, CA, USA. ²University of Health Sciences, Medical Informatics and Technology, Innsbruck, Austria. ³AO Development Institute, Davos Platz, Switzerland. ⁴Medical University Innsbruck, Innsbruck, Austria.

Texture analysis of femur radiographs may serve as a potential low cost technique to predict osteoporotic fracture risk and has received considerable attention in the last years. The goal of this in vitro study was to use automatically placed regions of interest (ROI) and morphological texture analysis methods in radiographs of proximal femur specimens to investigate their biomechanical strength.

Radiographs were obtained from 14 femoral specimens and bone mineral density (BMD) was measured in the femoral neck. Biomechanical testing in terms of failure load (FL), breakaway torque (BT) and number of cycles (NC) was performed. Five ROIs were placed automatically in the head, upper and lower neck, trochanteric and intertrochanteric compartment. In each ROI, we obtained isotropic and anisotropic morphological gradients (MG) and Minkowski Dimensions (MD), and three Minkowski Functionals [Area (MFA), Perimeter (MFP), Euler number (MFE)] and calculated coefficient of determination with parameters of biomechanical strength. In a stepwise multiregression analysis, we identified the most predictive parameters in different models.

A moderate correlation between neck BMD and FL of R²=0.40' (p<0.05) was found and a high correlation R²=0.78” (p<0.01) between BMD and BT. For isotropic MG-filters, correlations with FL up to R²=0.64* (p<0.01) and with BMD, BT and NC up to R²=0.62* were found. Using anisotropic MGs even higher correlations were obtained (R²=0.70*) for FL and NC, but no significant correlations were found with BMD. Correlations between MDs and FL were in the range R²=0.37″-0.61*. MFA and MFP correlated significantly with FL and NC (R²=0.55”), and were independent of BMD. All these correlations were based on femoral head and lower neck ROIs. Our multiregression analysis showed, that BMD was not able to improve the prediction of FL using a model with three MGs (R²=0.85'). The model testing all parameters resulted in a high prediction of R²=0.97' and included MGs, MD and MFP, however, not BMD. In conclusion morphological texture features were highly correlated with biomechanical strength and were partially independent from BMD. We showed that femoral head and lower neck ROI were best suited to estimate bone strength. Morphological information contained in trabecular bone structure visualized on radiographs may provide osteoporotic fracture risk prediction as a low-cost modality.

Disclosures: M.B. Huber, None.

This study received funding from: AO Foundation.

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Micro-architectural Changes of Developing Bone in Male and Female Rats. R. J. Arends*¹, M. V. Beuningen — Vaan*¹, M. V. Boven — Nunen*¹, M. T. Brok — Bardoel*¹, D. V. Dobbelsteen*², G. B. Janssen*², T. V. Klundert*¹, P. E. Langerwerf*¹, T. J. Robben*¹, M. G. Scheepers*¹, J. V. Vliet*², A. G. Ederveen¹. ¹Pharmacology, N.V. Organon, Oss, The Netherlands. ²Toxicology & Drug Disposition, N.V. Organon, Schaijk, The Netherlands.

MicroCT measurements are extensively applied in pre-clinical research on bone and are well validated to study micro architecture in adult and aged animals. However, the number of studies on the growing skeleton is limited. In this report, we have combined the metaphyseal trabecular bone and mid-diaphyseal bone μCT data of a number control groups of male and female Wistar rats at the age of 5, 8, 11 and 14 weeks to obtain a reference dataset of μCT parameters in growing animals.

From week 5 to 14 bodyweight increased from 107 to 424 gram in male and from 97–228 in female rats. Cortical BMD decreased rapidly between 5 and 8 weeks in males (1441 to 1148 mg/cm³) and in females (1459 to 1187) and thereafter remained constant until week 14. Moreover, between week 5 and 14 cortical thickness increased gradually; more in males (0.30 to 0.6 mm) than in females (0.31–0.51 mm). As a result of this, polar moment of inertia increased; more pronounced in males (4.2 to 19.6 mm⁴) than in females (3.6 to 8.4).

Trabecular BMD gradually decreased between week 5 and 14 in males (283–239 mg/cm³), however it increased in females (321–378). BV/TV increased steadily in females from 26.7 to 52.8% between 5 and 14 weeks and only slightly in males (24.8–31.8). Trabecular number decreased rapidly between 5 and 8 weeks in both males and females (3.1 to 1.8 l/mm and 3.3 to 2.7 l/mm, resp.) and this was accompanied by an increase in trabecular thickness (0.079 to 0.170 mm in males, 0.082 to 0.190 in females). As a net effect of these changes, trabecular separation strongly increased between 5 and 14 weeks in males (0.19–0.54 mm) and only slightly in female rats (0.18–0.23). Between 5 and 14 weeks, the degree of anisotropy decreased gradually in both males (3.29 to 2.15) and females (3.04 to 1.80), furthermore a modest decrease in structure model index was detected in males (1.49–1.12), while in females a strong decrease was measured (1.38–0.31).

In conclusion, the cortical femur is still growing in both sexes up to 14 weeks of age; males grow more than females. In femur of growing male rats, cortical thickness and diameter are the major parameters that determine bone strength as is predicted by a high polar moment of inertia. However, the cancellous compartment showed a different profile. In female rats, during skeletal growth trabecular bone volume and trabecular thickness increase in the femur, resulting in a gain of trabecular bone and an architectural change from a rod like structure to a more plate like structure, whereas in males trabecular bone is partially lost before it reaches a plateau phase.

Disclosures: R.J. Arends, N.V. Organon 3.

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Correlation Between the Mechanical Properties of Trabecular Bone Assessed by Finite Element Analysis and Experimental Test. Y. Won¹, B. Min*², Y. Chung³, M. Baek*¹, W. Cui*¹, M. Hahn*¹. ¹Orthopaedics, Ajou University School of Medicine, Suwon, Republic of Korea. ²Orthopaedics, Keimyung University School of Medicine, Daegu, Republic of Korea. ³Endocrinology and Metabolism, Ajou University School of Medicine, Suwon, Republic of Korea. ⁴Orthopaedics, Kwandong University School of Medicine, Gangneung, Republic of Korea.

Introduction: Following development of imaging technology, image-based finite element analysis has been commonly used to assess the mechanical properties of bone as a surrogate of experimental test. The aim of this study was to investigate the correlation between the mechanical properties of trabecular bone from the primary compressive trabecular system of human femoral head assessed by micro- finite element analysis and the experimental test.

Materials and Methods: Twenty-one human femoral heads were obtained from 11 patients undergoing total hip arthroplasty and 10 cadavers. Cylindrical trabecular bone was cored from the primary compressive trabecular system of the femoral head. The sample size was diameter of 19 mm, height of 15 mm. All samples were scanned with a high-resolution micro-computed tomography system at a spatial resolution of 21.31 μm. Based on the serial micro-CT images, a volume of interest (VOI) was selected to be 9.50 mm of diameter and 7.50 mm of side length. Based on these micro-images of trabecular bone, the finite element model was created, and the finite element analysis and the compressive test were performed sequentially. The yield stress and Young's modulus were calculated from two methods.

Results: The average yield stress and Young's modulus from FEA and Instron system were 13.04 MPa and 1.30 GPa, 13.06 MPa and 0.11 GPa, respectively. The correlations between the yield calculated from FEA and Instron were significant (p = 0.001), but not for the Young's modulus (p=0.055).

Conclusion: The mechanical properties of trabecular bone calculated from FEA are well correlated with those from the mechanical test, suggesting that the micro-image-based finite element analysis as a substitute of the mechanical test can be a useful tool to evaluate the mechanical properties of trabecular bone, further to predict the fracture risk.

Disclosures: M. Baek, None.

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A Methodology for Bone Strength Prediction using Parametric Statistical Shape and Density Finite Element Modeling. T. L. Bredbenner*, D. P. Nicolella. Materials Engineering, Southwest Research Institute, San Antonio, TX, USA.

Bone strength is the result of the complex combination of traits related to bone shape and bone density distribution. The use of finite element (FE) modeling methods improve the estimation of bone strength over predictions based on clinical image data alone since FE models can explicitly incorporate detailed descriptions of bone geometry and bone density distribution. However, the efficient construction of finite element models that accurately represent the complex morphology of skeletal structures is a major challenge thereby limiting their clinical use in diagnosing fracture risk. In the present study, methodology based on statistical shape modeling was developed and implemented to parametrically describe the complex geometry and bone density distribution using clinical imaging data for a set of 7 human female proximal femurs. The femurs were scanned using a clinical CT scanner (Lightspeed Ultra, GE Healthcare) along with a density calibration phantom (Mindways Software). Triangulated surfaces were created to describe the outer geometry for each femur (Amira, Mercury Computing) and a volumetric finite element mesh was mapped onto each femur using optimal point-to-point correspondence methods. Apparent ash density was estimated at each node for all femur models using the calibrated CT image data. Statistical shape and density modeling methods were then used to develop a parametric finite element model describing the set of cadaver femurs. Principal Component Analysis of the statistical shape and density FE model showed that 78% of the variability in bone geometry and density can be described using only 3 independent components. Reconstructing the FE model using each component independently and predicting bone strength under simulated fall conditions estimated the contribution of each component to proximal femur strength. Finally, individual finite element models of femurs within the training set could be accurately reconstructed, and a leave one out analysis demonstrated that a volumetric finite element model of a previously unseen femur could be reconstructed with a maximum geometry error of 1.25 mm and an average error of 0.04 mm.

Disclosures: T.L. Bredbenner, None.

This study received funding from: Southwest Research Institute Advisory Committee for Research.

W453

Assessment of Vertebral Fracture Threshold and Alendronate Therapeutic Effects by a QCT-based Nonlinear Finite Element Model in Postmenopausal Women. K. Imai¹, I. Ohnishi*², S. Yamamoto¹, K. Nakamura². ¹Orthopaedic Surgery, Tokyo Metropolitan Geriatric Medical Center, Tokyo, Japan. ²Orthopaedic Surgery, Tokyo University, Tokyo, Japan.

A QCT-based nonlinear finite element model (CT/FEM) accurately predicts vertebral strength, fracture sites and distribution of minimum principal strain (Spine 2006; 31:1789–94). This study aimed to assess the risk of vertebral fracture and the therapeutic effects of alendronate on osteoporosis using a CT/FEM. We performed a cross-sectional analysis of L2 vertebral strength for 104 postmenopausal Japanese women (mean age, 71.3 yrs; range, 49–85; mean height, 149.7 cm; mean weight, 50.5 kg) with no history of previous L2 fracture, traumatic vertebral fracture or osteoporosis medication. Subjects comprised 75 women without vertebral fracture and 29 women with non-traumatic vertebral fracture. The optimal point on the ROC curve that was used as vertebral fracture threshold was 1950 N with 75.9% sensitivity and 73.3% specificity. We also prospectively assessed the biomechanical effects of alendronate in 33 patients (mean age, 76.5 yrs) who were treated with alendronate at a dose of 5 mg/day for 1 year. QCT scans of the L2 were obtained at baseline and 3, 6 and 12 months, and the vertebral strength and mechanical parameters for each patient were analyzed using the CT/FEM. BMD of the L2–4 was measured by DXA at baseline and 6 and 12 months. BMD tended to increase from baseline at 6 months (3.7%; p=0.0451), and was significantly increased at 12 months (7.5%; p<0.0001). Vertebral strength from the CT/FEM significantly increased by 10.2% from baseline at 3 months (p<0.0001), 16.7% at 6 months, and 26.9% at 12 months. The CT/FEM also revealed that the area with large negative value of minimum principal strain before medication, where a high risk of fracture exists, decreased after medication. After 1 year of medication, the density of the inner cancellous bone with a distance of more than 2 mm from the outer surface increased by 8.3% (p=0.0013), while the density of the peri-cortical bone with a thickness of 2mm increased by 13.6% (p=0.0004). The CT/FEM could assess the vertebral fracture threshold with higher sensitivity and specificity, and also assess the therapeutic effects of alendronate with higher sensitivity and earlier (at 3 months) than BMD. The increase in the vertebral strength might be attributed to the alterations in the distribution of bone density, stress, and strain within the vertebra. The alendronate treatment was more effective in the peri-cortical and cortical areas compared to the inner trabecular compartment. The CT/FEM allows us to analyze quantitatively vertebral strength and biomechanical characteristics and is useful to assess the risk of vertebral fracture and therapeutic effects on osteoporosis.

Disclosures: K. Imai, None.

W454

Radiographic Analysis of Trabecular Bone Structure and Bone Geometry in the Prediction of Hip Fracture Load. T. J. Jämsä*¹, P. Pulkkinen*¹, E. Lochmüller*², V. Kuhn*³, M. Nieminen*⁴, F. Eckstein*⁵, ¹Department of Medical Technology, University of Oulu, Oulu, Finland, ²1st Gynecology Hospital, Ludwig-Maximilians-University, Munich, Germany, ³Department of Trauma Surgery, Medical University Innsbruck, Innsbruck, Germany, ⁴Department of Radiology, Oulu University Hospital, Oulu, Finland, ⁵Institute of Anatomy and Musculoskeletal Research, Paracelsus Medical Private University, Salzburg, Germany.

The clinical diagnosis of osteoporosis is typically based on measuring bone mineral density (BMD) with dual-energy X-ray absorptiometry (DXA). However, access to DXA is limited in many regions, while radiography is widely available. Here, we used an image processing method for assessment of trabecular structure, density and geometry from radiographs, to estimate the mechanical strength of femur.

The sample consisted of 62 femoral cadaver specimens (34 females, 81.6±10.7 yrs, and 28 males, 77.8±11.1 yrs). After radiography and DXA, the femora were mechanically tested in a side impact configuration. The fracture patterns were classified as being cervical or trochanteric. Computerized image analysis was applied to obtain several structure-related trabecular parameters, and a set of geometrical variables was also defined. Multiple linear regression analysis was performed to identify the variables that best explain variation in BMD and failure load between subjects.

In the cervical fracture cases, trabecular bone area (TBA) and calcar femoral cortex thickness (CFC) explained 68% of the variability in DXA-based femoral neck BMD (p < 0.001). TBA and femoral neck axis length (FNAL) explained 64% of the variability in failure loads (p < 0.001), while femoral neck BMD also explained 64%. In the trochanteric fracture cases, trabecular homogeneity index (HI) and femoral shaft cortex thickness explained as much as 87% of the variation in trochanteric BMD (p < 0.001). Euler number (EN) and calcar femoral cortex thickness (CFC) explained 66% of the variability in failure load (p = 0.001), while trochanteric BMD explained 72% (p < 0.001).

In conclusion, trabecular bone structure, as determined from conventional radiographs, reflects a substantial proportion of the variability in DXA-based BMD. Structural parameters of trabecular bone and bone geometry predict in vitro failure loads of the proximal femur with quite similar accuracy as DXA, when using appropriate image analysis technology.

Disclosures: T.J. Jämsä, None.

This study received funding from: Finnish Funding Agency for Technology and Innovation.

W455

Evolution of the Crack-Growth Resistance and Toughening Mechanisms of Human Cortical Bone via In-Situ Electron Microscopy. K. J. Koester*¹, J. W. Ager*², R. O. Ritchie*¹. ¹Materials Science and Engineering, University of California, Berkeley, Berkeley, CA, USA. ²Lawrence Berkeley National Laboratory, Berkeley, CA, USA.

Mammalian bone has evolved to support a variety of physiologically relevant loads and also to resist fracture, which, in many cases, can have life endangering consequences. It is imperative to understand the toughening mechanisms in human bone to characterize influence of age, disease, and drug treatments on the fracture properties of bone. Previous works have established that bone “toughens” to resist crack growth. However, without looking at the fracture of human bone in situ, these studies lack the potency to elucidate the phenomenon taking place during the fracture process. Here, in-situ crack propagation measurements performed in an environmental scanning electron microscope have allowed simultaneous measurement of the crack-growth resistance curve and real-time visualization of the toughening mechanisms. This work is the first study of the evolution of toughness in the transverse and longitudinal orientations in human cortical bone (humerus). The dominant toughening mechanisms were found to be crack deflection in the transverse direction and uncracked ligament bridging in the longitudinal direction.

Disclosures: K.J. Koester, None.

W456

Compromised Fracture Healing in Lrp5 Null Mice. D. E. Komatsu¹, A. G. Robling², C. H. Turner³, S. J. Warden¹. ¹Physical Therapy, Indiana University, Indianapolis, IN, USA. ²Anatomy & Cell Biology, Indiana University, Indianapolis, IN, USA. ³Biomedical Engineering and Orthopaedic Surgery, Indiana University, Indianapolis, IN, USA.

The importance of Wnt signaling in skeletal development and regulation has been unequivocally demonstrated in recent years. However, the role of this pathway in fracture repair has been the subject to far less attention. At the cellular level, Wnt pathway activation is initiated by the binding of Wnt ligands to the Wnt co-receptor Low-Density Lipoprotein Receptor-Related Protein 5 (Lrp5). Mice with homozygous null Lrp5 mutations (Lrp5-/-) demonstrate a low bone mass phenotype, concomitant with decreased mechanical properties. As Lrp5 is also known to be up-regulated during fracture repair, we hypothesized that fracture repair in Lrp5-/- mice would be compromised compared to wild-type littermates (Lrp5+/+). To test this hypothesis, closed fixed femoral fractures were generated in the left femurs of 4 month old male Lrp5-/- (N=7) and Lrp5+/+ (N=6) mice. Fracture repair was qualitatively assessed by radiography, performed post-surgery and weekly until sacrifice at day 28 post-fracture. The fractured and contralateral intact femurs were harvested and quantitatively analyzed by peripheral quantitative computed tomography (pQCT), dual energy X-ray absorptiometry (DXA) and biomechanical testing (4-point bending). To correct for baseline phenotypic differences, the results obtained for the fractured femurs were normalized to those of their contralateral intact femurs. No apparent differences in callus development were seen in the weekly radiographs. Similarly, analysis by DXA showed no differences in area or BMD, though a 15% reduction in BMC was observed in fractured femurs from Lrp5-/- mice, compared to Lrp5+/+ mice (p < 0.02). Likewise, the pQCT results also failed to identify substantial changes in callus morphology, with the sole significant difference being a 32% reduction in cross-sectional mature bone area in Lrp5-/- fracture calluses, as compared to Lrp5+/+ calluses (p < 0.05). However, biomechanical testing revealed a gross hindrance in the mechanical properties of fractured femurs from Lrp5-/- mice, compared to Lrp5+/+ mice, with reductions of 64-76% seen for ultimate load, stiffness, yield force, energy to yield and energy to ultimate force (all p < 0.05). The striking mechanical impairment in Lrp5-/- calluses, combined with minimal radiographic differences, indicates that the newly formed bone is biomechanically unsound despite the fact that these calluses appear to undergo almost normal development and mineralization. We conclude that signaling through Lrp5 is critical for the establishment of biomechanical integrity in healing bone.

Disclosures: D.E. Komatsu, None.

W457

Biomechanical Testing in Experimental Bone Interventions — May the Power Be with You. O. Leppänen¹, H. Sievänen², T. L. N. Järvinen¹, ¹Department of Trauma, Musculoskeletal Surgery and Rehabilitation, University of Tampere, Tampere, Finland, ²Bone Research Group, UKK Institute, Tampere, Finland.

Total variation in any measured variable, in conjunction with the expected treatment effect, dictate the minimum sample size (minSS) required to detect the expected effect with statistical confidence should the effect truly exist. The objective of this study was to introduce a practical algorithm for calculating the minimum sample size needed to detect a statistically significant treatment effect in biomechanical parameters.

A comprehensive literature survey comprising all 3472 original studies published in four major bone journals during 1999–2003 was carried out to identify studies in which biomechanical testing of whole bones was applied. The total variation in most commonly used biomechanical tests of various bones was statistically estimated. In addition, the average intervention effects were calculated for most common settings (ovariectomy, increased and decreased loading) in rats. Employing this data, the adequately-powered sample size for common biomechanical testing procedures was determined.

According to our literature survey, the total variation in different biomechanical tests of bones from different species was similar, justifying the use of rat femur as a model for detailed calculations. Due to poorer repeatability in terms of total variation, minSS in the femoral neck compression test is considerably larger compared to the midshaft bending test. Similarly, the use of stiffness and energy absorption required substantially larger sample size than the use of breaking load. For example, minSS to show a 10% treatment effect in the femoral shaft breaking load with 80% power was 11 rats/group, while the respective minSSs for the stiffness and energy absorption were 23 and 53, respectively. For the femoral neck compression, these numbers were 16, 51, and 134, respectively. Of note, a typical ovariectomy experiment in rats resulted in ∼5% decrease in femoral shaft breaking load, thus requiring a sample size of 42 rats.

The present analysis underscores the need for considerably larger sample sizes in experimental bone interventions using mechanical parameters as primary outcome variables. In particular, the poor repeatability in terms of total variation and the generally small expected treatment effects appear to compromise the general utility of stiffness and energy absorption results in typically small experimental bone interventions.

Disclosures: O. Leppänen, None.

W458

Prednisolone Induces Osteocyte's Death and Erosion of Osteocytic Lacunae Walls. M. Li¹, K. Hara², Y. Akiyama², N. Amizuka¹. ¹Center for Transdisciplinary Research, Niigata University, Niigata, Japan. ²Pharmacological Evaluation Section, Eisai Co., Ltd., Tokyo, Japan.

We have examined the abnormalities in the bones of Wistar rats that received prednisolone (20mg/kg) three times per week. Four, 8 and 12 weeks after beginning the steroid administration, rats were perfused with 4% paraformaldehyde in 0.1 M phosphate buffer (pH 7.4), and their femora and tibiae were processed for paraffin and epoxy-resin embedding. Bone abnormalities in prednisolone-administered rats were assessed by performing alkaline phosphatase (ALP), acid phosphatase (ACP), tartrate resistant acid phosphate (TRAP) histochemistry, TUNEL assay and transmission electron microscope (TEM) imaging. Peripheral quantitative computed tomography (pQCT) was performed for evaluating bone mineral density (BMD) and bone mineral content at the metaphyseal and diaphyseal regions of prednisolone-administered and control rats. Three-point bending test was employed for estimating bone strength. BMD and mineral contents at metaphyseal and diaphyseal regions, as well as values of maximum loading were reduced as early as 4 weeks in prednisolone-administered rats. Ratio of maximum load divided by bone mineral content was much lower in prednisolone-treated rats, so that the reduced maximum load-bearing values appear to relate not only with a decreased BMD, but also with diminished bone quality in prednisolone-administered bones. ALP and TRAP assessments did not reveal obvious difference between control and prednisolone-treated bones. When examining cortical bones, however, several empty osteocytic lacunae were seen in prednisolone-administered rats, and a statistical analysis on the number of empty lacunae showed a significant increase in the prednisolone group. TUNEL assay unveiled increased osteocytic apoptosis in prednisolone bones. However, empty lacunae seem not to be a result of apoptosis only, as the number of apoptotic cells was lower than the number of such lacunae. TEM images demonstrated disrupted osteocytes in relatively enlarged osteocytic lacunae, as well as irregularly shaped osteocytes that were embedded in lacunae featuring eroded peripheral walls. In addition, osteocytes distant from bone-resorbing osteoclasts tended to show ACP-positive granules, indicative of activated lysosomal enzymes in prednisolone-treated rats. Thus, in our study, predonisolone appears to target osteocytes, inducing their cell death and erosion of the surrounding walls of their lacunae, which might influence overall bone quality.

Disclosures: M. Li. None.

W459

New Method to Determine Variation of the Elasticity of Single Trabeculae. S. Lorenzetti*, E. Stüssi. Institute for Biomechanics, ETHZ, Zürich, Switzerland.

By comparing the deflection of an experimental 3 point bending test with FE calculation, the Young's modulus of single trabeculae can be determined.

With DXA and QCT the bone mineral density (BMD) is measured. Due to this technique, variations of the calcium apatite are detectable, but changes of the collagen mesh are not visible. With the presented new method, variations of the elastic behavior of single trabeculae are accessible.

A special micro-positioning device enables sample rotation around the trabecular axis. The force was applied normal to the optical axis and the trabecular with a nylon thread, and the deflection could be measured with the microscope. Based on the image stack from microscopy, a CAD body of the trabecula can be built using Amira^TM and Geomagic^TM. FE calculations were performed with ANSYS^TM. By comparing the FE modeling with the deflection of the experimental 3 point bending test, the Young's modulus can be determined.

To obtain the accuracy of the method, two types of error calculations were performed: First, a straight forward estimation according to Gauss which includes the length, the force and the deflection, and second, FE calculations with variation of the volume. The expected resulting error for the Young's modulus is <10%.

To test the new method the collagen was removed from a set of samples. Samples from the femur of an adult sheep were treated with sodium hypochlorite (NaOCl). The treatment with NaOCl is a standard method to deproteinize bone, and therefore, to alter the Young's modulus without changing the geometric properties or the inorganic part of the bone. The analysis of a native (untreated) femural sample resulted in a Young's modulus of 15.0 ± 1.4 GPa. This is in the upper range of the published data in the literature of 1 up to 19 GPa. The treated samples showed a reduction of the Young's modulus by a factor of 15. Broz et al (1997) observed a decrease at macroscopic samples by a factor of 2.7.

Using DXA, no variation of the BMD would have been observed in this type of samples. With the new method, changes of the organic part (collagen) of the bone become visible.

It is concluded that the new method including sample preparation, experimental 3 point bending test with optical control, determination of the volume and FE modeling is suitable to determine the Young's modulus of single trabeculae in the natural boundary conditions of the sponiosa.

Broz et al. (1997) J. mat. Sci. 8:395–401

The technical support of C. Hauser is gratefully appreciated.

Disclosures: S. Lorenzetti, None.

W460

Fracture Strength of the Distal Radius Can Be Estimated by Non-linear Finite Element Analysis Using 3D High Resolution Peripheral Computed Tomography. J. A. M. MacNeil*, S. K. Boyd. Mechanical Engineering, University of Calgary, Calgary, AB, Canada.

Finite element (FE) modeling based on micro-computed tomography data has the potential to provide significant insight into disease- and treatment-related bone quality changes through estimation of fracture strength. The goals of this study were to determine if linear finite element models of bone stiffness can predict bone strength, and determine if non-linear finite element models provide an improvement over linear finite element models, through a direct assessment of strength. It was also investigated whether a non-homogeneous tissue Young's modulus scaled according to attenuation values can improve bone stiffness and strength measurements.

Fresh human cadaver forearms (5 male and 5 female) with ages ranging from 55 to 93 were obtained from the Gross Anatomy Lab at the University of Calgary. The distal radius was dissected free from each forearm and four 9.1 mm sections were cut beginning at the subchondral plate. The sections were scanned with high resolution peripheral quantitative computed tomography (HR-pQCT) (XtremeCT, Scanco Medical, Switzerland; voxel size 82 μm) in saline solution. These forty samples were mechanically tested and used to determine appropriate homogeneous and non-homogeneous tissue Young's moduli for finite element analyses. Ten samples corresponding to the standard clinical measurement location for in vivo HR-pQCT measurements were used for non-linear finite element evaluation.

Bone stiffness measurements by linear FE analysis using a non-homogeneous tissue Young's modulus had a significantly higher correlation to bone ultimate strength than the homogeneous tissue Young's modulus models, likely due to implicit correction for partial volume effects (R²=0.983 vs. R²=0.960; p<0.05). For the clinical measurement samples, the non-linear FE analysis predicted the experimentally determined ultimate strength better than correlations based on apparent stiffness estimates from linear FE. This trend was true for homogeneous tissue Young's modulus models (R²=0.937 vs. R²=0.868) and was statistically significant for non-homogeneous tissue Young's modulus models (R²=0.951 vs. R²=0.928; p>0.05).

It was concluded that for estimation of bone strength using in vivo HR-pQCT measurements, non-homogeneous tissue Young's modulus improves estimations, and that linear finite element measurements were highly predictive of bone ultimate strength, albeit based on correlations rather than direct measurements. A non-linear finite element measurement directly estimated bone strength, and can provide improved accuracy when the increased computation is justified.

Disclosures: J.A.M. MacNeil, None.

W461

Posterior Cortical BMC in the Femoral Neck Predicts Proximal Femur Failure Load. S. L. Manske*¹, D. M. L. Cooper*², T. Liu-Ambrose*², P. Guy*², H. A. McKay². ¹University of Calgary, Calgary, AB, Canada. ²University of British Columbia, Vancouver, BC, Canada.

Region-specific deterioration of the femoral neck (e.g. superoposterior) may play a key role in hip fractures. We aimed to determine the region of the femoral neck where cortical bone mineral content (CoBMC) best predicted failure load. We obtained 36 unembalmed, previously frozen human cadaveric femora (36 – 92 years), and scanned each with a 16-slice helical QCT scanner (GE LightSpeed Ultra). QCT scans were acquired axially and images (0.5 × 0.5 × 0.63 mm) were reconstructed perpendicular to the femoral neck axis (0.5 × 0.5 × 2.5 mm). We assessed total area (mm²) and CoBMC (g) in the femoral neck cross-section with the lowest total area (i.e., narrowest-neck). We also measured cortical BMC regionally within 8 sectors (octants) radially defined about the centroid of the cross-section (Figure). The femora were then loaded to failure in an apparatus that simulated a sideways fall at a rate of 100 mm/s. Femora that fractured outside of the neck or intertrochanteric regions (n = 7) were excluded from analyses. We performed hierarchical linear regression with failure load (N) as the dependent variable. In the regression model, we entered total area at the first level to account for differences in bone size and then utilized stepwise selection of the sector-based CoBMC values at the second level. Two additional specimens were excluded as outliers based on Cook's distance. The highest correlations between CoBMC and failure load were along the anteroinferior to superoposterior axis (R ranged from 0.63 to 0.71, Figure). The stepwise regression selected CoBMC from the posterior sector based on its association with failure load. CoBMC in the posterior region explained 65% of the variance (R²) in failure load after accounting for total area. Our results agree with the predictions of Mayhew et al. (2005) and Lotz et al. (1995) who suggest that the superoposterior region may be critical to the strength of the femoral neck. We extend their findings by providing measurements of proximal femur strength in ex vivo specimens. Our quantitative results further correspond with our observation that most of the fracture lines propagated through the superoposterior region. We conclude that the superoposterior region requires further investigation as a key site of fracture initiation and propagation in sideways falls.

Disclosures: S.L. Manske, None.

W462

Evaluation of Vertebrae Fragility by Osteoporosis. N. Nango¹, N. Endo*², T. Yamamoto*³, H. Takahashi⁴, M. Machida*⁵, Y. Suga*⁶, T. Asai*⁶. ¹Ratoc System Engineering Co., Ltd., Tokyo, Japan. ²Niigata University, Niigata, Japan. ³Niigata Bone Science Institute, Niiigata, Japan. ⁴Niigata University of Health and Welfare, Niigata, Japan. ⁵Murayama Medecal Center, Tokyo, Japan. ⁶Kawaguchi Municipal Medical Center, Saitama, Japan.

We developed a method to quantify the trabecula path that connects between the two endplates of the vertebrae. We consider this connecting path primarily prevent the vertebral deformation. We used in-vivo Multi Detector CT images of females aged 70 with lumbar stress fracture (a), females aged 86 without fracture(b), and males aged 57 without fracture (c). Based on the CT values of these bone images, we determined the trabecula bone (TRI/3D-BON software by RATOC), and measured the length (L) and cross section (S) of each trabecula. We define the trabecula elasticity coefficient (k) as k ∝ S/L, and the resistance force (f) as f=k*d for deformation distance (d). We define the simulation model as follows: loaded surface, fixed surface and trabecula network connecting both surfaces. Using given external deformation (D), deformation (d) for each trabecula was obtained: each trabecula path connecting the two surfaces satisfy D=Σd; we also calculated the resistance force(fe) of each trabecula composing the network. The resistance force of vertebrae (Fe) was calculated as the sum of (fe) of the trabeculae contacting loaded surface Fe=Σfe.

This calculated Fe is the simulation of resistance force generated in vertebrae when stress deformation is applied to the loaded surface. The figure shows the results, of the connecting trabecula paths between upper and lower endplates which resistance force greater than the specified threshold. The value of the resistance force decreased in the order of young male(c), old female(b), female with stress fracture(a). The resistance force in females with stress fracture was half of that in the male.

As the images show, in the female with stress fracture, the density of trabecula bone connecting the endplates is low at the front of the vertebrae and there are few straight connecting paths, instead the paths are connected by many detours. The paths of the old female without fracture have many fragmented disconnections. The trabecula bone network of the female with stress fracture has disappeared locally and is regarded as the cause of the fragility.

This model shows the mechanical strength of vertebrae trabecula bones but also demonstrates the fragility.

Disclosures: N. Nango, None.

W463

Improved Analysis of Bone Nano-mechanical Properties Using a Novel Nanoindentation Technique. S. Pathak*¹, S. R. Kalidindi*¹, H. W. Courtland², K. J. Jepsen², H. M. Goldman³. ¹Materials Science and Engineering, Drexel University, Philadelphia, PA, USA. ²Orthopedics, Mount Sinai School of Medicine, New York, NY, USA. ³Neurobiology and Anatomy, Drexel University College of Medicine, Philadelphia, PA, USA.

Currently, there exists a critical need to develop better techniques to understand the relationship between bone matrix composition and organization and tissue-level mechanical properties, thereby improving our ability to predict fracture risk. To address this gap, we have developed a new technique for extracting mechanical property information at the nano-scale in bone, by translating nanoindentation generated load-displacement curves into indentation stress-strain (ISS) curves for a better representation of material deformation at the lamellar level. We applied this method to a sample of femora obtained from strains of inbred mice known to have varying whole-bone mechanical properties.

Femora from A/J, C3H/HeJ (C3H), and C57BL/6J (B6) mice (16 wks of age, n=3 each) were embedded in PMMA, sectioned transversely below the third-trochanter and surface polished to 0.05μm. Backscattered electron microscopy (BSE-SEM) was used to assess mineralization density across the postero-lateral (PL) cortex of each sample. Nanoindentations (20μm apart) were carried out across this same cortex using a nanoindenter (MTS XP*) with a 13.5 μm radius spherical diamond tip, and corresponding ISS curves were computed. Examination of the ISS curves taken at a 40μm distance from the actively forming periosteal surface of each strain demonstrate that all three have similar slopes during (elastic) loading, but vary in post yield behavior; A/J demonstrates the highest hardening slope, while B6 demonstrates the lowest. Obtaining the elastic modulus values from the loading portion of these ISS curves rather than the unloading portion of the load-displacement curve also helps us more readily discern the differences in modulus between bone of lower and higher mineralization density.

Our results demonstrate that this method can be used to elucidate trends in yield and post-yield behavior at the lamellar level that are consistent with findings found in macro-mechanical testing of these mouse strains. This represents a significant advancement in the characterization of mechanical behavior of bone under indentation pressure beyond the regimes of elastic limit to which it has been restricted to thus far. The additional local property information provided by our nanoindentation technique, in conjunction with compositional analyses, will help us tremendously in developing an understanding of the complex interactions between bone quality, composition and mechanical properties.

Disclosures: S. Pathak, None.

This study received funding from: NIH AR44927.

W464

Osteoblast-specific Knockout of IGF Receptor Gene in Mice Alters Mechanical Properties of Cortical Bone. G. Ramaswamy*¹, X. Cao*², S. Chowdhury*³, C. Wan², G. McGwin*⁴, J. E. Lemons*⁵, T. L. Clemens², A. W. Eberhardt*¹. ¹Department of Biomedical Engineering, University of Alabama at Birmingham, Birmingham, AL, USA. ²Department of Pathology, University of Alabama at Birmingham, Birmingham, AL, USA. ³Department of Physics, University of Alabama at Birmingham, Birmingham, AL, USA. ⁴Department of Epidemiology, University of Alabama at Birmingham, Birmingham, AL, USA. ⁵Department of Prosthodontics and Biomaterials, University of Alabama at Birmingham, Birmingham, AL, USA.

Insulin-like growth factor-1 is essential for normal embryonic and postnatal growth. Previous studies have shown that selective disruption of the IGF receptor gene (Igflr) in mouse osteoblasts decreased bone formation and mineral apposition rates over the pubertal period.¹ The present study compared the geometric and mechanical properties of cortical bone at the mid-diaphysis of tibiae in 3 week old Igflr knockout and control mice (n = 14 per group) at the whole bone level by microCT and three-point bending and at the microstructural level using nanoindentation. MicroCT data revealed a decrease in cortical thickness (p < 0.05) which translated into a dramatic reduction in bone area (p < 0.05) and bone volume fraction (p < 0.0001) in the knockouts. Elastic modulus and toughness obtained from bending showed significant reductions (p < 0.05) in the knockouts as compared to controls while overall bending stiffness was only mildly affected. These bones showed geometric adaptation with increased mediolateral radii (p < 0.05) and moments of inertia similar to controls, which partially counteracted for the material degradation to maintain their structural integrity. Microstructural properties from nanoindentation did not show any genotype effect, which was consistent with bone mineral density measurements from microCT that were similar between controls and knockouts. Gender specific comparison demonstrated females to be smaller in size with structural properties more severely reduced following ablation of IGF signaling. In conclusion, the current study suggests that local IGF signaling in murine osteoblasts is essential for normal appositional growth of cortical bone during diaphyseal modeling and its disruption affects the mechanical properties, more severely in females, at the prepubertal stage in mice.

1. Zhang M, et al. Endocrinology 2002;277:44005–44012.

Disclosures: G. Ramaswamy, None.

W465

Txnip Deficient HcB/19 Mice Have Decreased BMD and Femoral Strength. N. Saless¹, J. Chen*², A. Shalev*², R. D. Blank². ¹Cellular and Molecular Biology, University of Wisconsin, Madison, WI, USA. ²Medicine, University of Wisconsin, Madison, WI, USA.

Thioredoxin interacting protein (Txnip) mediates the oxidative stress response. Txnip-thioredoxin interaction reduces thioredoxin activity, so that in vitro Txnip overexpression leads to increased apoptosis and decreased proliferation following exposure to oxidative stress. HcB/19 recombinant congenic mice harbor a nonsense mutation of Txnip, resulting in a metabolic profile similar to human familial combined hyperlipidemia. Mice harboring the Txnip mutation also display an increase in fat mass relative to wild-type. Hypothesizing that the observed increase in fat mass is accompanied by a concomitant decrease in bone mass, we compared bone density and biomechanical performance in HcB/19 mice and C3H/DiSn mice, their recipient progenitor, which harbor the normal Txnip allele.

Mice were studied at 4 months of age, examining BMD by DXA (PIXIMus, GE-Lunar) and femoral biomechanics by 3-point bend testing. The latter was performed under displacement control at a rate of 0.3 mm/second and post spacing of 3.75 mm. Data were analyzed for total displacement, structural stiffness, maximum load, and energy to failure. Data from the right and left femora were averaged for each individual. Data were analyzed by 2 way ANOVA with α = 0.05.

We found that HcB/19 mice display decreased total body BMD, and decreased maximum load and stiffness, in spite of similar body mass (see table). Although energy to failure tended to be lower in HcB/19, the difference did not reach significance. There were no interactions between strain and sex. Biomechanical performance was greater in females than males.

Our data show that Txnip deficiency reduces peak bone mass and bone strength. Txnip is known to be induced by 1,25 di-OH D and repressed by mechanical loading. Our data do not address the question of whether either of these mechanisms contributes to the observed biomechanical phenotype. Our data also do not allow us to determine whether the Txnip mutation alters the balance between the osteoblastic and adipocytic lineages during development. Our experiment is limited in that other loci aalso segregate between the strains. Nevertheless, the epidemiological association between atherosclerosis and osteoporosis and our biomechanical data justify further investigation of Txnip 

Table  .  

Data shown as mean ± SEM, NS = not significant, * = significant strain difference, † = significant sex difference

Disclosures: N. Saless, None.

W466

Biomechanics and Body Size Quantitative Trait Loci in HcB/8 × HcB/23 F2 Mice. N. Saless¹, G. E. Lopez Franco*², S. J. Litscher*², K. Abdul Raheem*³, K. Kozarek*³, S. Sudhakaran*³, M. J. Houlihan*³, A. S. Ahmed*³, T. K. O'Neil*³, S. J. Han*³, R. D. Blank². ¹Cellular and Molecular Biology, University of Wisconsin, Madison, WI, USA. ²Medicine, University of Wisconsin, Madison, WI, USA. ³University of Wisconsin, Madison, WI, USA.

Recombinant congenic mouse strains HcB/8 and HcB/23 differ in biomechanical performance in spite of similar ash percentages. To identify quantitative trait loci contributing to the differences in biomechanical performance between these strains, we performed a reciprocal F2 intercross to map the responsible QTLs. Below, the female progenitor is listed first, in accordance with convention.

F2 mice were kept to 17 ± 1 weeks under standard husbandry conditions. Following sacrifice, we performed microsatellite genotyping and measured femoral whole bone biomechanics and BMD by DXA. From 3-point bend testing performed under displacement control, post separation of 3.5 mm, and a deflection rate of 0.3 mm/sec, we extracted total and post-yield displacement, stiffness, maximum load, and energy to failure. We analyzed the data with QTL Cartographer and established experiment-wide significance levels by permutation testing. We studied 124 8×23 females, 163 23×8 females, 169 8×23 males and 147 23×8 males.

Significance thresholds ranged between LOD scores of 1.9 for stiffness to 3.7 for post-yield displacement, reflecting the reproducibility of the trait assays. In the entire F2 population, the only significant QTL is for body mass at 27 cM on chromosome 2. In the 8 × 23 arm, there is a significant QTL at 68 cM on chromosome 3 for total displacement and post-yield deflection (LOD = 4.9 and 5.7, respectively). On chromosome 10, there is a stiffness QTL (LOD = 2.6) at 28 cM. In the 23×8 arm, there are a pair of chromosome 3 QTLs for length centered at 12 cM (LOD = 3.0) and 68 cM (LOD = 3.2). Among males of both arms, there is a broad body mass QTL (LOD = 3.3) located between 18 and 72 cM on chromosome 3. Among females of both arms, there is a body mass QTL (LOD = 3.2) at 79 cM on chromosome 3. Arm and sex specific subgroups display some of these QTLs and also display additional significant QTLs not listed above. Comparison of the overall phenotypic distributions also revealed evidence of one or more X-linked QTLs.

The data show that there are both cross and sex effects, as others have found in previous bone linkage studies. Epistatic interactions between pairs of genes, genomic imprinting, and mitochondrial inheritance are all possible mechanisms underlying the observations. Future work to isolate the genes responsible for the QTLs found in this cross will be facilitated by the fact that the donor segments in the parental strains have already undergone multiple recombination events, limiting their length and providing physical landmarks for fine structure genetic mapping.

Disclosures: N. Saless, None.

This study received funding from: Veterans Administration Biomedical Research and Development Service.

W467

Interrelation Between Dietary Fatty Acids and Fibers in Modulating Bone Quantity and Quality During Skeletal Growth. A. Schirmer*¹, S. Ferreri*¹, L. M. Miller*², Y. Qin*¹, N. Turner*³, J. Lupton*³, S. Judex¹. ¹Biomedical Engineering, Stony Brook University, Stony Brook, NY, USA. ²Biomedical Engineering, Brookhaven National Laboratory, Upton, NY, USA. ³Texas A&M University, College Station, TX, USA.

Dietary factors can influence the levels of bone formation, bone resorption and impact bone quantity and quality. Increased quantities of dietary lipids may have detrimental effects on bone mass but emphasizing specific lipid sources, while maintaining the total amount of dietary fat, can be beneficial. Little is known about the interaction between dietary lipid and fiber source. The objective of this study two different lipid sources were factorially combined with two different fiber sources to investigate their skeletal impact in rapidly growing rats. Forty-nine 4wk old male rats were randomly divided and fed one of four dietary combinations for 4wk (fish oil/cellulose, fish oil/pectin, corn oil/cellulose, corn oil/pectin). At completion of the protocol, micro-computed tomography determined differences in trabecular and cortical bone morphology of the distal and diaphyseal femur. Mechanical and chemical properties of the femoral diaphysis were determined by 3-point bending and Fourier transform infrared spectroscopy. There were no significant differences between the four dietary groups for any morphological or micro-architectural parameter in the distal femoral metaphysis. In contrast, middiaphyseal cortical bone was sensitive to differences in dietary components. Rats fed a fish oil rather than a corn oil diet had a 5% (p<0.02) greater cortical bone area, a 5% (p<0.005) greater periosteal area, and a 6% (p<0.02) greater endocortical envelope area. Conversely, bones from rats on the corn oil diet showed a 26% (p<0.002) higher energy to failure. There were no significant differences in collagen structure between groups but rats fed the corn oil diet had a 16% (p<0.02) higher level of mineralization compared to rats on the fish oil diet. When comparing rats on cellulose versus pectin diets, no differences were found for any morphological, mechanical, or chemical properties. These data indicate that dietary fish oil, compared to corn oil, can moderately promote cortical, but not trabecular, bone growth but, at the same time, may decrease measures of bone quality. In the absence of effects, or interactions, of pectin and cellulose, any skeletal differences observed in this study could be attributed to differences in fatty acid content. Whether the only modest differences in bone quantity and quality are related to the relatively short experimental protocol or the genetically modulated high levels of cellular activity in the rapidly growing rat remains to be determined.

Disclosures: A. Schirmer, None.

W468

Vitamin D Deficiency: A Common Occurrence in Both High and Low Energy Fractures. B. Schreck*¹, A. Serota*¹, D. Helfet*¹, M. G. E. Peterson*¹, N. Sinha², J. M. Lane¹. ¹Hospital for Special Surgery, New York, NY, USA. ²New York Presbyterian Hospital, New York, NY, USA.

Vitamin D insufficiency is an under-recognized problem in adults in the United States. Vitamin D influences Ca homeostasis, bone mineral density, muscle strength, and bone strength and healing. This retrospective study tests the hypothesis that vitamin D deficiency is prevalent among fracture patients, regardless of high vs. low energy fracture etiology.

A retrospective analysis of the medical records for 44 in-patients with non-vertebral fractures on the orthopedic trauma service at the Hospital for Special Surgery was conducted from June 1, 2006 to February 1, 2007. The obtained data included a 25-hydroxyvitamin D level, age, gender, and reason for admission; high energy fracture vs. low energy fracture (a fall from standing height or less).

Vitamin D deficiency, 25(OH)D<32 ng/ml, was found in 59.1% of the patients and the mean 25(OH)D was 30 ng/ml ± 13. Significantly more women (75%) than men (40%) were vitamin D deficient. Significantly more women than men with high energy fractures were vitamin D deficient (p=0.01). In women, both high and low energy fractures present with vitamin D deficiency (80% of high energy fractures and 71.4% of low energy fractures). In men, the mean vitamin D level was lower for low energy fractures (16 ng/ml) compared to high energy fractures (32 ng/ml), p=0.007. In addition, men with low energy fractures were significantly older than men with high energy fractures, p=0.02, but this age difference was not significant among women, p=0.053. The mean age for women (66 years) was higher than the mean age for men (51 years), p=0.02. Women showed a significant correlation of age vs. 25(OH)D as older women actually had higher values of 25(OH)D than younger women, rho=0.46 (p=0.02). For men, there was no significant correlation of age vs. 25(OH)D, rho= −0.2 (p= 0.2).

A high prevalence (59%) of vitamin D deficiency is seen among in-patients on the orthopedic trauma service. Statistically more vitamin D deficiency is seen in women and our results are consistent with the gender difference seen in the general population. The higher values of 25(OH)D in older women compared to younger women may be due to the fact that older women are more likely to be treated with calcium and vitamin D. Even among younger men who sustain a high energy fracture, 25% are vitamin D deficient. Women regardless of age or energy level causing fracture have low vitamin D levels. 25(OH)D levels should be measured in all orthopedic trauma patients and deficient patients should be treated.

Disclosures: B. Schreck, None.

W469

Variation of the Young's Modulus of Single Trabeculae in a Sheep Femur. E. Stüssi, S. Lorenzetti*. Institute for Biomechanics, Ethz, Zürich, Switzerland.

In this work, first results of a study of the variation of the Young's modulus of single trabeculae within a femur are presented.

The fracture risk of bone is influenced by several parameters. The important factors are age, gender, mobility, type of bone, the bone mineral density (BMD) and the material properties. The mechanical properties are influenced by the architecture, the structure and the quality of the bone forming material. One of the major parameters of the material properties is the Young's (elastic) modulus E. This value describes the linear deformation under loading.

Various standard methods are known to measure the stiffness of the entire spongiosa or bone at macroscopic scale. Furthermore, indentation tests are available to determine the elastic behavior of microscopic spongiosa samples. In this work, the first results of a new method to determine the Young's modulus of single trabecula using experimental and FE simulation of a 3 point bending test are presented.

This method takes the exact 3D geometry and the behavior under loading into account. Stacks of images were taken using a laser scanning microscope (LSM) and a near infrared laser. Based on the image stack from microscopy, a CAD body of the trabecula can be built using Amira^TM and Geomagic^TM. FE calculations were performed with ANSYS^TM. The experimental 3 point bending test was performed with the boundary conditions of the natural trabecular network. The force was applied normal to the optical axis with a thread, and the deflection could be measured with the microscope. By fitting the FE modeling to the deflection of the experimental 3 point bending test, the Young's modulus can be determined. The experimental error estimated by Gaussian error calculation and variation of the input parameters of the FE calculations is < 10%.

The spongiosa samples were taken from the femur of a healthy 2.5 y old sheep. The bone was cut into cubic samples using a band saw. The bone marrow was extracted using ultrasound wave bath and pressure air.

The presented data shows variation of the Young's modulus within one femur by a factor of about 4. This is a similar range as summarized by Bayraktar et al. 2004.

Bayraktar et al. J. of Biomech. 37(11):27–35

The technical support of C. Hauser is gratefully appreciated. 

Table  .  

Disclosures: E. Stüssi, None.

W470

Bone Fragility May Be Caused by Mechanostress Inequality in Transitional Period from a Steady-state Bone Metabolism to Another State. Y. Tamura, K. Nakayama, S. Katayama*. Saitama Medical University, Saitama Pref., Japan.

We have presented a computer simulation program of bone metabolism in a past ASBMR meeting. In this meeting, we want to show unloading model of bone on this program.

At first, we made a two-dimensional square bone model in computer and loaded 10 units of mechanical force to the initial bone. After dozens of simulation time, this bone became a structure like trabecular bone and reached steady-state bone metabolism. Then we reduced this force from 10 to 5 units to simulate mechanical unloading. BV/TV decreased promptly, but Tb.N decreased slowly. Dozens of simulation time later, this bone reached new steady state.

We postulated that the phase between fast and slow convergence of BV/TV and Tb.N was transient period for bone to adapt to change of external load. So we examined local mechanical stress, bone formation and resorption in this period.

Before and after transient period, local mechanical stresses were uniform and steady, and bone formation and resorption were also stable. However, during transient period, local stresses varied from place to place, causing increased and fluctuated bone metabolism. Trabecular perforation caused stress concentration to parts of residual trabecula and the places became very weak transiently.

These results suggest that bone fragility with increased bone metabolic marker may occur in transitional period from a steady state bone metabolism to another one, as shown in this unloading simulation model. If so, fractures in osteoporosis may be caused by mechanostress inequality in transient and unstable structures during adaptation processes to another environment of external load and hormones. And it may be very important that this unstable state continued for a long time after bone mass had changed. This condition may mean poor bone quality in osteoporosis.

Disclosures: Y. Tamura, None.

W471

Lean and Not Fat Mass Is Associated with Male Proximal Femur Strength. T. G. Travison¹, A. B. Araujo¹, G. R. Esche*¹, T. J. Beck², J. B. McKinlay¹. ¹New England Research Institutes, Watertown, MA, USA. ²Johns Hopkins University School of Medicine, Baltimore, MD, USA.

Obesity is thought to confer protection against fracture, but the mechanism driving this association is poorly understood. We hypothesized that the effect of increased weight or BMI on bone structure would be accounted for by total and/or appendicular lean mass (LM), and that once these trends were removed, fat mass (FM) would demonstrate no protective influence. To test this hypothesis, we examined body composition and geometric indices of proximal femur strength in a diverse sample of randomly-selected men, ages 30–79 y.

Data were obtained from N=1,171 community-dwelling men randomly selected for inclusion in the cross-sectional Boston Area Community Health / Bone study. Body composition was obtained by dual X-ray absorptiometry (DXA). Hip geometry parameters at the narrow neck (NN), intertrochanter, and shaft regions of the proximal femur were obtained via Hip Structural Analysis of DXA images. These measures included bone mineral density (BMD), bone material in cross sections (cross-sectional area, CSA), bending strength (section modulus, Z) and propensity to buckle under compression (average buckling ratio, ABR). Analyses controlled for age, race/ethnicity (black, Hispanic, white), height, and physical activity.

When considered individually, BMI, LM and FM were each positively associated with hip strength. However, controlling for LM removed the positive, and induced a negative, association for BMI or FM (the Figure depicts results for BMI and NN CSA). In joint models including LM, FM and covariates, NN CSA and Z increased by an estimated 12.4 percent (95% CI: 10.2, 14.6) and 15.1 percent (95% CI: 12.2, 17.9), respectively, per 10 kg increase in LM, while FM had no positive effect. Finally, LM alone was sufficient to account for a substantial proportion of racial difference in hip strength measures (≥ 50% for the majority of comparisons), while FM exhibited no explanatory power. Findings were similar whether total or appendicular mass was considered.

These results indicate that the positive association between relative weight and proximal femur strength is accounted for by lean mass, and suggest that the protective effects of BMI in preventing fracture risk is mediated not by adipose tissue but by the influence of increased muscle mass accompanying elevated BMI.

Disclosures: T.G. Travison, None.

W472

Development of a High-Resolution 3D Micro-CT Based Model to Predict Fracture Callus Histological Architecture. J. A. Weis*¹, F. Granero-Molto¹, L. D. O'Rear*¹, M. I. Miga*², A. Spagnoli¹. ¹Pediatrics, Vanderbilt University, Nashville, TN, USA. ²Biomedical Engineering, Vanderbilt University, Nashville, TN, USA.

Annually ∼600,000 people in the US suffer from fracture healing failure. There is a compelling need for studies that define the dynamic mechanisms underlying the fracture healing process and predict when it fails. The purpose of this study is to determine appropriate microCT thresholds that can predict callus bone/cartilage architecture. A stabilized tibia fracture was created in FVB mice by three-point bending. The tibia was harvested after 14 days and subjected to microCT scanning at 6 micron voxel resolution for 7.4 mm length around the callus (Scanco μCT 40). The tibia was sectioned at 6 micron for 5.2 mm length around the callus. Sections were placed on 288 slides with 3 sections per slide. In-situ hybridization for collagen 1 and collagen 10 was performed for 210 sections for each probe. The collagen 1 and 10 stained sections were quantified and labeled with a defined color value and spatially reconstructed by a custom built image registration code (MATLAB). Each section was centered and rotated to align with the previous section. The reconstructions were then overlayed to produce a bone/cartilage 3-D model based on collagen 1 and 10 expressions. The 3-D model was then compared to the 3-D reconstruction from microCT. As shown in Figure 1, callus volumetric analysis demonstrated that the CT voxel based 3-D reconstruction well predicted the 3-D bone/cartilage model based of collagen 1 (A) and 10 (B) expressions. Figure 1 depicts representative sections of collagen 1 (A), collagen 10 (B), voxel based CT image (C), and predicted 3-D bone/cartilage model (D) clearly indicating the high resolution predictability of the CT model. This study provides basic information for developing a CT voxel based 3-D model that correlates with histological structure of a fractured callus. This model can potentially overcome the limitation of histological studies that can only be based on a limited number of tissue sectional analysis. ,  

Figure Figure 1.. Volumetric analysis of bone/cartilage model vs the predicted voxel-based CT reconstruction.

Figure Figure 2.. Representative callus sections of Col 1 (A) Col 10 (B) in situ hybridization, CT imaging (C), and predicted model (D)

Disclosures: J.A. Weis, None.

This study received funding from: NIH-NIDDK.

W473

Unilateral Osteoarthritic Hips Have Lower Trochanteric BMD and Higher Femoral Neck BMD Compared with the Unaffected Side — An Explanation Why Trochanteric Fractures Are More Frequent Than Femoral Neck Fractures in Osteoarthritic Hips. O. Wolf*, J. Milbrink*, S. Larsson*, H. Mallmin. Surgical Sciences, Uppsala, Sweden.

Patients with osteoarthritis of the hip, OAH, are subject to trochanteric hip fractures but very seldom to femoral neck fractures. One reason might be differences in bone strength due to altered bone composition in the proximal femora. In OAH there might be a problem using DXA, as external contracture is common and this might affect BMD compared to the recommended slight inward rotation. Our aim was to compare BMD at the proximal femur and heel in OAH-affected and healthy legs as well as body composition in the lower extremities. In addition we measured the effect of differences in rotation on DXA values in patients with no hip disease.

41 patients (20 women, 54 ± 8,8 years) with unilateral OAH scheduled for arthroplasty were included. Before surgery DXA was done bilaterally at the proximal femora and heels, as well as total body. Three regions of interest (ROI) for the proximal femur (femoral neck-FN, trochanter-TR and total hip-TH) were investigated. Preoperative weight bearing (WB) was measured with the F Scan system and hip abductor strength with a dynamometer (MS). Differences in rotation on DXA images between OAH and control side were thoroughly assessed. In 21 hip-healthy patients (64.1 ± 11.5years; 42 hips) DXA was done with the foot in a vertical position as well as the recommended inward rotation to assess the effect of rotation. OAH had 5% higher BMD_FN (p<0.007), 8% lower BMD_TR (p<0.000002) and 5% lower BMD_TH (p<0.001) compared with the control side. 21/38 had no differences in rotation whereas 16 had an increased outward rotation of the OAH side. Regional analyses of the total body measurement showed significantly lower BMC and fat mass, 5% and 2%, respectively, in OAH legs but no difference in lean mass (p<0.67). However, when calculated as ratios these differences did not remain significant. We found no significant differences in BMD_HEE1, WB or MS between OAH and controls. For the hip-healthy patients there was a significant increase in BMD_TR (2.5%, p<0.000001) but no significant influence on BMD_FN and BMD_TH with the foot in vertical position compared with slight inward rotation.

Higher BMD_FN and lower BMD_TR in OAH compared with controls, indicates that the trochanteric region is the weakest part of the proximal femur in patients with OAH. This finding can not be explained as false low values due to external contracture. On the contrary, neutral position seems to provide higher BMD_TR values. In conclusion, low BMD_TR offers a potential explanation why OAH-patients are far more likely to suffer a trochanteric rather than a femoral neck fracture.

Disclosures: O. Wolf, None.

W474

The Effect of Collagen Degradation on the Stiffness of Bone in an Emu Model. C. Wynnyckyj*¹, S. Omelon*², M. D. Grynpas². ¹Materials Science Engineering, University of Toronto, Toronto, ON, Canada. ²Samuel Lunenfeld Research Institute, Mount Sinai Hospital, Toronto, ON, Canada.

Current methods of bone quality assessment, such as Dual energy X-ray Absorptiometry (DXA) and Quantitative Ultrasound (QUS) measure bone mineral content and density, while less emphasis has been devoted to collagen's role in fracture risk assessment. Neither DXA nor QUS provide a direct measurement of the mechanical properties of bone. DXA measures bone mineral density (BMD) and QUS measures speed of sound (SOS) transmitted through bone. Although these surrogate measurements are related to fracture risk, they are not sufficient to adequately predict bone fragility. Consequently, there is a need for a technique that can provide more information related to fracture than the data from DXA or QUS. A promising candidate is the Mechanical Response Tissue Analyzer (MRTA), a radiation free, non-invasive instrument developed by NASA to investigate the effect of space travel on bones. The MRTA directly measures a mechanical property of bone, the cross-sectional bending stiffness, which describes the ability of bone to resist deformation.

In order to assess the contribution of collagen on the mechanical properties of bone, the emu tibia was selected as a model due to its size and approximate cylindrical shape, making it ideal for devices designed to accommodate human long bones. The effect of deproteinization in ex-vivo emu bone was explored using DXA, QUS and MRTA. Four groups (n=7–10) of female emu tibiae were endocortically deproteinized in 1M KOH solution for 1, 3, 7 and 14 days, resulting in negligible mass loss (0.5%). Hydroxyproline assays were used to confirm minimal loss in collagen content. Statistical analysis of results over time and between techniques was performed. The negligible mass loss resulted in significant changes in modulus of elasticity between all time points (except between 3 and 7 days), as determined by 3-point bending and MRTA. The change in SOS and BMD varied by less than 2%. This is not surprising as both techniques measure changes in bone mineral content, which is not affected by KOH. In contrast, the MRTA reflected changes in the modulus caused by deproteinization of bone more significantly. No significant differences were seen between 3-point bending and MRTA. The significant reduction in modulus caused by the small amount of bone mass removal may be due to in situ collagen degradation rather than collagen loss. 

Table  .  

Disclosures: C. Wynnyckyj, None.

W475

A Novel Patient-Specific Finite Element Model to Predict Damage Accumulation in Vertebral Bodies under Axial Compression. Y. Chevalier*, M. Charlebois*, P. Varga*, D. Pahr*, P. K. Zysset. Institute of Lightweight Design and Structural Biomechanics, Vienna University of Technology, Vienna, Austria.

In the past years, QCT-based finite element (FE) approaches were applied to monitor the effect of anti-resorbtive or anabolic treatments on vertebral body stiffness under axial compression or bending.

However, these FE analyses were either elastic, that is they excluded any representation of damage accumulation, or utilized constitutive laws based on metal plasticity which have limited accuracy in simulating irreversible strain and damage in bone tissue.

In this work, a novel constitutive law specifically designed for trabecular bone elasticity, plasticity and damage was applied to a collection of 12 cadaveric human vertebral bodies (L1-L5, age 47 to 83) in the framework of an automated finite element model generation software.

The vertebral bodies were scanned by pQCT at 80 microns, segmented and coarsened into 1.3mm isotropic voxels. Mineral density was converted into bone volume fraction (BV/TV) on which the elastic, yield and ultimate properties of the developed transverse isotropic constitutive law were based. Superior and inferior endplates in PMMA were added to transmit load to the vertebral bodies. Axial compression was applied to the digital meshes in the form of axial displacements on the superior endplate. Generation of the mesh and application of the boundary conditions were fully automated and a typical FE simulation took approximately 240 minutes of CPU time on 3GHz Xeon processors with 16GB RAM.

Beyond the usual compression stiffness, the yield and ultimate loads as well as the irreversible deformation of the vertebral bodies were computed. A typical force versus displacement curve is shown in Fig. 1A. while a map of the induced damage is visualized in Fig. 1B. Both vertebral stiffness and ultimate force predicted by the finite element models were linearly related to the experimental values (r²=0.64 and r²=0.76 respectively). The slopes and intercepts of these regressions were not significantly different from 1 and 0 respectively.

This novel finite element model of the human vertebral body was therefore properly validated by experimental data, computationally efficient and could be applied to investigate the effect of 2 years teriparatide treatment within the Eurofors study (see abstract this meeting).

Disclosures: P.K. Zysset, None.

This study received funding from: Lilly Deutschland GmbH.

W476

A Combined Axial Compression and Flexion Loading Mode to Evaluate Strength of Vertebral Bodies using a Voxel-Based Finite Element Method. M. Charlebois*, Y. Chevalier*, P. Varga*, D. Pahr*, P. K. Zysset. Institute of Lightweight Design and Structural Biomechanics, Vienna University of Technology, Vienna, Austria.

The effect of pharmacological treatment on the mechanical properties of bone relies increasingly on finite element models. For human vertebral bodies, axial compression is the loading mode of choice to investigate stiffness or nonlinear characteristics such as yield or ultimate force. However, in daily activities, the human lumbar spine is subjected to a variety of loading modes and vertebral fractures also exhibit different morphologies. In this context, the goal of this study was to compare stiffness and ultimate mechanical properties of human vertebral bodies under axial compression, antero-posterior flexion and a combination of these two loading modes.

A group of 12 human lumbar vertebrae (age 47–83) were scanned with high-resolution pQCT and digitized to 1.3mm voxels. A novel transverse isotropic constitutive law for bone including elasticity, plasticity and damage accumulation was used for all loading cases. Superior and inferior endplates in PMMA were added to the vertebral models to apply loads in the form of axial displacements on the superior endplate and flexion angles about a transverse axis located at the most posterior voxel of the superior endplate. A combined proportional loading mode was then applied using the axial displacement and flexion angle obtained at the yield point of the individual loading modes.

Beyond the independent axial and flexion stiffnesses, the full stiffness matrix of the sagittal motion was computed. Ultimate load and moment were calculated as the maximum force and torque resisted by each vertebrae.

As a first result, the axial and flexion stiffnesses as well as their respective ultimate properties were linearly related (r²=0.87, r²=0.74). In all loading modes, the level of damage was substantially lower in the cortical than in the trabecular compartment where damage localization occurred preferentially (Fig. 1). As expected, the amount of damage was increased for the combined loading mode and was higher than the sum of the damage produced by the individual compression and flexion modes. In conclusion, the combined loading mode applied in voxel-based finite element analyses provides a refined insight into the damage mechanisms leading to failure of vertebral bodies.

Disclosures: P.K. Zysset, None.

This study received funding from: Project 05Z26 of the AO Research Fund, AO Foundation; Lilly Deutschland GmbH.

W477

Effects of Strontium Ranelate Therapy after Long Term Bisphosphonate Treatment. Histomorphometric and μXRF/EDX Analysis of Paired Iliac Crest Bone Biopsies in 15 Patients. B. Busse¹, M. Priemel¹, B. Jobke*², M. Hahn*¹, J. Zustin*³, J. Semler*⁴, M. Amling¹. ¹Center for Biomechanics & Trauma Surgery, Univ. Med. Center Hamburg, Hamburg, Germany. ²MQIR, Radiology, UCSF, San Francisco, CA, USA. ³Bone Pathology, Univ. Med. Center, Hamburg, Germany. ⁴Internal Medicine, Immanuel Hospital, Berlin, Germany.

Several medications have proved effective in reducing the fracture risk in patients with osteoporosis. However, the optimal duration of use of these medications and the effects of changes between treatment strategies remain to be established. The latter are of special clinical interest in patients who have been on bisphosphonate (BP) therapy for years and present with persistent high fracture risk. To gain insides into the effects of strontium ranelate (SR) therapy after long term BP treatment we studied paired transiliac crest biopsies from 15 patients. All patients had previous BP therapy (∅ 32 months) and were subjected to bone biopsy at baseline. Patients were treated with 2g SR/d and subjected to a second biopsy after 6 months (group I, n=5) and 12 months (group II, n=10), respectively. Biopsies were undecalcified histologically processed. The bone mineral density distribution was evaluated by the gray level allocation (mean GL) of backscattered signal intensities. Energy dispersive x-ray analysis (EDX) combined with μ-x-ray fluorescence analysis (μXRF) was used to monitor Sr concentration (Sr/(Sr+Ca), Wt%). Static analysis was performed using the Osteomeasure system according to ASBMR standard and included the quantification of bone volume (BV/TV, %), trabecular interconnection (TBPF, mm-1), trabecular thickness (Tb.Th, μm), osteoid volume (OV/BV, %), osteoid surface (OS/BS, %), as well as osteoclast number (NOc/BPm, mm-1). In group I there was an increase in Sr content while all other indices did not show significant changes as compared to baseline. In sharp contrast in group II there was not only a further increase in Sr content of bone and mean GL but also a significant increase in BV/TV (11.12 ± 3.78 vs. 8.53 ± 2.14) with an improved TBPF (0.69 ± 0.52 vs. 1.35 ± 0.63) and increased Tb.Th (99.39 ± 9.89 vs. 89.99 ± 10.96). These structural changes were explained by activated bone formation indicated by increased OS/BS (12.04 ± 6.81 vs. 7.97 ± 5.6) and OV/BV in face of persistent suppression of bone resorption and unchanged low NOc/BPm (0.011 ± 0.016 vs. 0.035 ± 0.065). These data show that Sr is present in bone in patients that have been on BP treatment previously and that one year after SR therapy active bone formation sites are increased while osteoclasts are still repressed with beneficial effects on bone quality. Therefore SR might be considered as an alternative therapeutic option in patients that have been on long term BP treatment.

Disclosures: B. Busse, None.

W478

Bone Turnover and Quality: How Discrepancies in BMD Changes and Fracture Risk May Be Explained. M. A. Karsdal, I. Byrjalsen, D. Leeming, P. Qvist, C. Christiansen. Nordic Bioscience, Herlev, Denmark.

Introduction: Bone matrix quality may be important for anti-fracture efficacy as the reduction in risk is only partly explained by a concomitant increase in BMD during anti-resorptive therapy. Some of these discrepancies may in part be explained by careful investigation of novel bone turnover markers.

The main protein component of the extracellular bone matrix is collagen type I of which the C-telopeptide motif is released during osteoclastic bone resorption (CTX). Newly synthesized bone matrix contains the α-form of CTX, but with aging the motif spontaneously isomerizes to the β-form. Thereby index of alpha/ beta fragments released during osteoclastic of bone resorption may be a representative for endogenous bone age, which may be related to bone quality and affected differently by different treatments regimes.

The aim of the present study was to evaluate the effect of different anti-resorptive treatments (bisphosphonates, HRT, SERMs and calcitonin) on the age profile of bone collagen degradation measured as the ratio between the degradation products of newly synthesized and mature isomerized C-telopeptides of type I collagen.

Methods: Participants were from cohorts of healthy postmenopausal women participating in double blind, placebo-controlled 2-year studies of alendronate, ibandronate, intranasal hormone replacement therapy (HRT), oral HRT, transdermal HRT, oral calcitonin or raloxifene (n = 487). The non-isomerized ααCTX and isomerized ββCTX were assessed by ALPHA CrossLaps and BETA CrossLaps ELISAs, respectively, in urine samples obtained at baseline, and after 6, 12, and 24 months of therapy. Presently only 3 months data are available for calcitonin.

Results: Bone collagen age measured as the ratio between ααCTX and ββCTX showed that bisphosphonate treatment induced a pronouncedly older age profile with a 52% (alendronate) and 38% (ibandronate) reduction in the ratio between the two CTX isoforms in face of 3% and 15% with HRT or raloxifene. Calcitonin treatment resulted in an increase of 10%, rather than a decrease.

Conclusions: The different responses of ααCTX to ββCTX ratio by the various anti-resorptive therapies reflect a treatment-dependent effect on the endogenous age profile of bone collagen. These data highlight that even though the treatments have comparable effects on BMD, the endogenous bone composition, which may be associated to bone quality, is strongly affected by the type of intervention.

Disclosures: M.A. Karsdal, None.

W479

Quality and Strength of Cortical Bone in Both Ovariectomized and Sham-operated Rats Are Lowered by Limiting Physical Activity. Y. Mikuni-Tkagaki¹, K. Miyagawa*¹, Y. Kozai*², Y. Ito*³, K. Naruse*⁴, Y. Nagai*⁵, H. Yamato⁵, I. Kashima*², K. Ohya⁶, K. Aoki⁷. ¹Functional Biology, Kanagawa Dental College, Yokosuka, Japan. ²Maxillofacial Diagnostic Science, Kanagawa Dental College, Yokosuka, Japan. ³Yokohama Training Center, Kanagawa Dental College, Yokohama, Japan. ⁴Department of Orthopedic Surgery, Kitasato University School of Medicine, Sagamihara, Japan. ⁵Kureha Special Laboratory Co., Ltd, Tokyo, Japan. ⁶Hard Tissue Engineering, Graduate School, Tokyo Medical and Dental University, Tokyo, Japan. ⁷Department of Hard Tissue Engineering, Department of Hard Tissue Engineering, Section of Pharmacology, Tokyo, Japan.

Lack of estrogen after menopause and reduced activity (hypokinesis) are both important factors in the pathogenesis of osteoporosis in the elderly women. To compare the contribution of these two factors independently, we developed an animal model comparing ovariectomized (OVX) or sham-operated rats, which were housed in cages with limited space or in regular institutional cages. The experimental cages restricted standing and walking, while allowing other activities such as eating, drinking and changes in position. Thirty-two Wistar rats, 14 weeks of age, were assigned to four groups of OVX-Restricted, OVX-Walking, Sham-Walking and Sham-Restricted rats. Serum samples were collected during postoperative weeks 1, 4, 8 and 12 to study metabolic bone markers. After sacrificing rats at week 12, femora and tibia were examined with peripheral quantitative computed tomography (pQCT), laser Raman spectroscopy, three point bending, and histopathology. Although significantly elevated levels of metabolic markers, CTx, and osteocalcin were recorded in OVX groups compared to the Sham groups, results of bone strength as well as structural parameters revealed significant differences only between Sham and Restricted groups. In the Restricted rat tibial cortices, both the number of empty osteocytic lacunae and lacuno-canalicular sclerostin staining were significantly increased. Osteoclast activity was not necessarily associated with the osteoporotic pathology in restrained rat bones, suggesting a predominant role of osteocytic control of osteolysis in the bone loss of individuals with restricted activity.

Disclosures: Y. Mikuni-Takagaki, None.

This study received funding from: Ministry of Science, Education, and Culture of Japan to YM-T.

W480

Changes in Dental Enamel Crystals by Bleaching. M. Ogiwara, Y. Miake*, T. Yanagisawa*. Ultrastructural Science, Tokyo Dental College, Masago 1-2-2, Mihama-ku, Chiba -city, Japan.

Recently, the number of requests for dental bleaching has been rapidly increasing for aesthetic reasons. Clinically, dental bleaching is widely performed. Bleaching is considered to occur by the decomposition of organic substances, without affecting inorganic substances. However, there have been only a few number of basic morphological studies, and no study has been conducted on changes in the crystal structure examined by ultrahigh-resolution transmission electron microscopy. We considered that crystal-morphological evaluation of the effects of dental bleaching on enamel crystals was necessary.

We observed the structural changes in the superficial layer of extracted teeth that agreed with patient and approved ethics committee, after trimming and bleaching by scanning electron microscopy (SEM,3D-SEM), and examined the radiolucency by contact microradiography (CMR). Furthermore, trabecular and crystallographic changes in the enamel were analyzed by ultrahigh-resolution transmission electron microscopy (HR-TEM).

SEM and 3D-SEM revealed trabecular capsules, indicating a change caused by bleaching. Furthermore, HR-TEM demonstrated the decalcification of enamel crystals in some samples.

It has been considered that since bleaching occurs through the action of generated reactive oxygen species (ROS) on organic substances, it does not affect inorganic substances. However, the results of this study suggested that bleaching had considerable effects on inorganic substances. The pH is reduced by a bleaching agent, suggesting that the decalcification of enamel crystals caused by bleaching was related to the pH reduction.

Disclosures: M. Ogiwara, None.

W481

Image Processing for Three Dimensional Dynamic Histomorphometry. C. R. Slyfield*¹, R. E. Tomlinson*², E. V. Tkachenko*¹, G. J. Steyer*², C. G. Patthanacharoenphon*¹, G. J. Kazakia*³, D. L. Wilson*², C. J. Hernandez¹, ¹Mechanical and Aerospace Engineering, Case Western Reserve University, Cleveland, OH, USA, ²Biomedical Engineering, Case Western Reserve University, Cleveland, OH, USA, ³Radiology, University of California, San Francisco, San Francisco, CA, USA.

Bone remodeling has been implicated as an indicator of bone quality. The manner in which bone remodeling influences bone strength independent of bone mass is not yet well understood but is believed to be influenced by cavities formed during remodeling. Cavity number and size may therefore influence bone quality and strength. Two dimensional (2D) histomorphometry is currently the only way to examine local bone remodeling. Unfortunately, 2D histomorphometry cannot quantify the number and size of remodeling events. In addition, 2D histomorphometry is very labor intensive and requires special training. Here we present methods for performing automated three dimensional dynamic histomorphometry which enable the measurement of remodeling cavity number and size. 3D images of bone and two different fluorescent formation markers are obtained using a automated serial milling technique at a resolution of five microns (Kazakia et al., 2007). A 3D model is created using an automated image pre-processing technique. Subsurface fluorescence is removed and an adaptive threshold automatically labels mineralizing bone. While 2D histomorphometry requires manual labeling of five or more bone surface traits (labeled surface, bone surface, osteoid surface, etc.), the 3D histomorphometry technique only requires eroded surfaces to be traced manually. Just as micro computed tomography led to improved precision and accuracy in trabecular microarchitecture measures, 3D dynamic histomorphometry is expected to be more precise and accurate than 2D methods. This is the first image processing technique to allow precise 3D measures of bone and fluorescent markers in cancellous bone. This technique will enable investigations of the effect of remodeling size and number on bone biomechanics and help to identify differences among osteoporosis therapies.

Disclosures: C.R. Slyfield, None.

This study received funding from: Case Western Reserve University.

W482

The Role of Dynamic Muscle Contractions on Inhibition of Bone loss and Muscle Atrophy in a Functional Disuse Mouse Model. A. M. Ali*, Y. Qin. Biomedical Engineering, SUNY Stony Brook, Stony Brook, NY, USA.

It is proposed that musculoskeletal disorder may be counterbalanced by understanding the relationship between muscle function and bone adaptation. Skeletal muscle dynamics and its related mechanotransduction function have been suggested as a modulator of intramedullary pressure, and may play a role in bone adaptation. The objective of this study was to evaluate optimized dynamic muscle stimulation in the therapeutic potentials in prevention of bone loss and muscle atrophy. The aims of this study were to test the role of varying the frequency and duration of repetitive muscle contractions at 1 Hz, 20 Hz, and 50 Hz on muscle mass and bone morphology in a functional hind limb disuse mouse model. Forty-four adult female BALB/c mice were randomized into 6 groups: baseline control (n=6), age-matched control (n=6), hind limb suspension non-stimulated (HLS-NS) (n=8), HLS with electrically stimulated muscle contractions at 1 Hz (n=8), 20 Hz (n=8), and 50 Hz (n=8). Pulsatile signal was delivered to the quadriceps muscle with stainless steel needle electrodes for 2 seconds, followed by a 3 second rest period, for total of 10 minutes per day, for 3 weeks. Upon sacrifice, soleus, gastrocnemius, quadriceps, and whole hind limb weights were recorded. Femoral trabecular bone was quantified using micro-CT (SCANCO) at 8 micron resolution for a 1-mm region of interest at the distal metaphysis. HLS-NS showed a decrease in the weight of soleus (-21%), gastrocnemius (-26%, p<0.01), quadriceps (-20%, p<0.01), and the whole hind limb (-15%, p<0.05). Electrically stimulated muscle contractions at all frequencies showed increasing trends in soleus (+6% to +21%), gastrocnemius (+7% to +18%), quadriceps (+3% to +12%), and the whole hind limb (+7 to +13%, p<0.05). Micro-CT revealed that HLS-NS reduced bone volume fraction (-62%, p<0.01), connectivity density (-40%, p<0.01), trabecular number (-19%, p<0.01), and increased trabecular separation (+22%, p<0.01). Muscle contractions at 1 Hz, 20 Hz, and 50 Hz significantly recovered bone volume fraction (+34% p<0.05; +47% p<0.01; +48% p<0.01), connectivity density (+50% p<0.01; +59% p<0.001; +65% p<0.001), trabecular number (+10% p<0.05; 15% p<0.001; 15% p<0.001), and decreased trabecular separation (-10% p<0.01; −14% p<0.001; −14% p<0.001), respectively. The significant recovery of muscle mass and bone structural parameters in the treatment animals suggests that the dynamic muscle contraction is a valid potential therapeutic signal for musculoskeletal disuse. Among all the applied signals, 50 Hz contraction generated the greatest response in bone morphology. These data suggest that an optimized range of physiologic signals can be responsible for prevention of bone loss and muscle atrophy noninvasively and at low magnitude.

Disclosures: A.M. Ali, None.

W483

The Long Term Macro-architectural Benefits of High Impact Exercise During Growth: A Study of Retired Gymnasts. P. Eser*, B. Hill*, S. Bass. Centre for Physical Activity and Nutrition Research, Deakin University, Burwood, Australia.

The greatest skeletal benefits from exercise occur during growth; for these benefits to be important in the primary prevention of osteoporosis they must be maintained into adulthood. Little is known however about the macro-architectural structure of bone changes in response to exercise during growth and how these changes may be maintained when exercise is ceased.

The objective of this study was to determine if retired gymnasts have greater bone strength (i.e. bigger bone size, mass and density) at both cortical and trabecular sites compared to age-matched controls. In this study we compared 30 female gymnasts (age: 23.0 ± 4.4 years, training duration: 10.5 years) who had been retired for 6.1 years, with 20 age, height and weight-matched controls (less than 2 hours of exercise per week). Volumetric BMD and bone cross-sectional area (CSA) were assessed by peripheral quantitative computed tomography (pQCT) at the epiphyses and diaphyses of the radius, humerus, tibia and femur. Polar bone strength strain index (SSlpol) and muscle CSA were assessed at the mid-shaft of each limb.

Upper Limb: At the radial and humeral shafts, cortical CSA, total CSA, BMC and SSlpol were significantly greater (15%–40%) in the retired gymnasts (p<0.01). Muscle CSA at the arm and forearm was also increased by 14% and 16%, respectively (p<0.05). At the distal radius, cortical CSA, BMC and trabecular vBMD was 19%-23% greater in the retired gymnasts (p<0.001). Lower Limb: At the tibial shaft, cortical CSA and BMC was 10% greater (p<0.05). At the femoral shaft BMC was 10% greater (p<0.08). At the distal tibia and femur, trabecular vBMD was greater by 8% and 7%, respectively (p<0.05). Muscle CSA at the upper and lower leg was not different between groups. At the lower extremity, the magnitude of the differences were approximately half that observed in the upper extremity. Between-group differences in bone geometry and vBMD paralleled the differences in muscle CSA. No significant linear relationship was found between years of retirement and any of the bone or muscle parameters.

Despite a greatly reduced level of physical activity (less than 2 hrs/week) and at least 6 years of retirement gymnasts maintained benefits that were site and region specific; at the diaphysis the macro-architectural changes were due to greater bone size and cortical CSA at the upper limb but only a greater cortical CSA at the lower limb. At the epiphysis the benefit was predominantly due to a greater trabecular vBMD at both upper and lower limbs. In conclusion, greater bone strength is maintained in gymnasts retired for up to 6 years. The macro-architectural basis of these benefits varies depending on the skeletal site and region.

Disclosures: S. Bass, None.

This study received funding from: Victorian Golf Association.

W484

The Effect of Vibration Resistance Exercise During 56 Days of Bed Rest on Different Muscle Properties. B. Buehring*¹, U. Gast*², I. Michaelis*², D. Belavy*², D. Felsenberg², J. Rittweger*³. ¹Internal Medicine, University of Wisconsin Hospital and Clinics, Madison, WI, USA. ²Center for Muscle and Bone Research — Charite Berlin, Berlin, Germany. ³Institute for Biophysical and Clinical Research into Human Movement, Manchester Metropolitan University, Alsager, United Kingdom.

Multiple studies have examined the effect of microgravity on muscle function. There is ongoing debate whether the observed loss of muscle function is solely correlated to loss of muscle mass. Finding the causes of muscular function loss in microgravity is important not only for future space flight missions but also for immobilized patients on earth.

Maximal voluntary contraction force (MVC) and dynamic muscle power during countermovement jumps were examined in a 56 day bed rest setting. The effect of vibration resistance exercise (VRE) on the two tasks was also assessed. VRE, consists of a combination low frequency vibration applied to the feet and resistance training. 20 healthy male subjects participated and were divided into an exercise and a control group (CTRL). Mean MVC and surface electromyography (EMG) was measured in the plantarflexors with a dynamometer. Muscle power and jump height were used to evaluate countermovement jumps.

After bed rest, MVC in CTRL was decreased by 17% (p<0.01), but remained unchanged in the VRE group. Maximal EMG amplitude was uneffected in both groups. Neuromuscular drive (ND) did not change in the VRE group but a trend in the decline of ND was found (p=0.052) in CTRL. Countermovement jumps revealed a loss in peak power in both groups. Jumping ability decreased by 13% (p<0.001) in the VRE group whereas the CTRL lost 26% (p<0.001).

This study showed that bed rest without any countermeasures lead to a significant loss of MVC and a decrease in neuromuscular drive of 21% which was on the border of statistical significance. It proved that VRE was able to preserve MVC in the plantarflexors, maintain neuromuscular properties but could not prevent a decrease in the ability to perform countermovement jumps. Mulder et al report a 14.1% loss of total quadriceps cross sectional area (CSA) in the same control group. The VRE group had a significantly smaller loss of 3.5%. We conclude that besides muscle atrophy, other factors like coordination, neuromuscular drive and changes in muscle cell function contribute to loss of certain muscle functions. VRE can preserve MVC, however it can only alleviate the loss of function in more complex tasks. Therefore it is important to add training methods like VRE to astronauts in space or immobilized patients on earth. However more research is needed to examine the effects of microgravity and immobilization on muscular functions besides maximal force.

Disclosures: B. Buehring, None.

W485

Exercise Induced Changes in the Macro-Architectural Parameters of Cortical Bone as Boys Progress Through Puberty: A Large Window of Opportunity?, G. Ducher*¹, J. Black*², L. Saxon*³, R. Daly¹, C. Turner⁴, S. Bass¹. ¹Centre for Physical Activity and Nutrition Research, Deakin University, Burwood, Australia. ²Musculoskeletal Research Centre, La Trobe University, Heidelberg, Australia. ³Basic Sciences, The Royal Veterinary College, London, United Kingdom. ⁴Department of Orthopaedic Surgery, Indiana University School of Medicine, Indianapolis, IN, USA.

Very little is known about the macro-architectural changes in cortical bone in response to exercise during different stages of growth. In this study we compared the side-to-side differences in cortical bone parameters in pre- (n=6), peri- (n=16) and post- (n=16) pubertal boys who played tennis to determine the site- and maturation-specific changes in cortical bone parameters in response to loading. Humeral BMC was assessed by DXA and bone geometry of the mid (40–50%) and distal (60–70%) humeral diaphyses was assessed by magnetic resonance imaging. The pre-, peri, and late or post- pubertal players were aged 11.2±0.5, 12.9±0.3, and 15.3±0.3 yrs and were currently training for 9.1±1.8, 15.4±1.7, 13.0±1.7 hr/wk respectively. Relative side-to-side differences in total and cortical areas ranged from 6.9% to 33.6% at both sites for all pubertal groups (p ranging <0.09 to 0.001). The absolute side-to-side difference in cortical CSA was greater in the peri- and post pubertal boys compared with the pre-pubertal boys (p< 0.05). Medullary contraction was detected at the distal site (ranging from 7.9 to 14%) in all pubertal groups. In contrast, expansion in the medullary cavity was observed at the mid site (∼5%) in pre- and peripubertal boys. 

Table Table 1. Absolute side-to-side differences in humeral bone parameters in the young male tennis players according to their pubertal status (mean ± SE).

* Playing vs non-playing significant at p<0.09-0.001

Between groups comparison: ‡ Pre < Peri at p<0.05; † Pre < Post at p<0.01

Large absolute and relative side-to-side differences in total and cortical cross-sectional areas were detected in prepubertal boys; these differences were greater in the peri and postpubertal boys. These findings are in contrast to our previous work in girls where we report an initial response in the prepubertal players but no further increase in peri and postpubertal girls despite a longer duration of loading. These findings together suggest that males may have a larger window of opportunity to optimize the osteogenic response to loading.

Disclosures: G. Ducher, None.

This study received funding from: National Health and Medical Research Council.

W486

Optimising Bone Health in Pre-Pubertal Children: The Importance of Muscle Strength. R. English*¹, P. Eser*¹, A. Patchett*¹, R. Daly¹, G. Naughton², M. Seibel³, R. Telford*⁴, S. Bass¹. ¹Centre for Physical Activity and Nutrition Research, Deakin University, Burwood, Australia. ²Centre of Physical Activity Across the Lifespan, Australian Catholic University, Sydney, Australia. ³ANZAC Research Institute, University of Sydney, Sydney, Australia. ⁴Australian Institute of Sport, Australian National University, Belconnen, Australia.

Peak muscle force is a determinant of bone strength; however its contribution to sex-specific differences in skeletal development remains uncertain. We investigated the role of muscle cross-sectional area (MCSA); a surrogate measure of peak muscle force on bone mineral content (BMC), geometry and strength at different skeletal sites (weight bearing and non-weight bearing) and bone compartments (trabecular versus cortical). Peripheral QCT was used to assess non-dominant tibia and radius (distal 4% and mid-shaft 66%) BMC, total and cortical CSA and MCSA in 598 pre-pubertal boys (n=302) and girls (n=296) aged 8-9 years.

No sex differences were detected in age, weight or tibial length. Compared to girls, boys were taller (1.4 cm), had greater radial length (0.4 cm) (p<0.001) and greater MCSA at the lower leg (3.5%) and arm (7.9%) (p<0.001). At the tibia and radius metaphyses boys had greater BMC (∼10%) and total CSA (7%) (p<0.0001). These differences remained after adjusting for MCSA. At the diaphysis total and cortical CSA, and polar strength strain indices were 2–10% greater in boys (p<0.05). These differences were not maintained after adjusting for MCSA, which accounted for 10 to 40% of the variance in bone parameters. All results were maintained after adjusting for bone length. In conclusion, MCSA accounted for the greater bone strength at the diaphyses in boys compared to girls. Factors other than MCSA contributed to the sex-specific differences in skeletal parameters at the metaphyses. These findings highlight the importance of developing MCSA for improved bone health at the diaphysis in children.

Disclosures: R. English, None.

This study received funding from: Commonwealth Institute of Australia.

W487

Bone Turnover Response of Men and Women to a Strenuous Gender-Integrated Recruit Training Regimen. R. K. Evans¹, A. J. Antczak*¹, M. E. Lester*¹, R. Yanovich*², E. Israeli*², D. S. Moran*². ¹Military Performance Division, US Army Research Institute of Environmental Medicine, Natick, MA, USA. ²Sheba Medical Center, Heller Institute of Medical Research, Tel Hashomer, Israel.

Rapid onset of strenuous exercise training can lead to stress fracture, a bone overuse injury. Stress fracture susceptibility is hypothesized to result from accelerated bone remodeling and changes in bone turnover that favor resorption. Whether the bone metabolism response is different in women, who present with a five-fold greater risk for stress fracture than men, has not been studied. The purpose of this study was to assess gender disparity and changes in biomarkers of bone metabolism during a four-month recruit training program. Healthy age-matched men (n=58) and women (n=199) (19.0 ± 1.0 yr) entering a four-month gender integrated basic combat training program in the Israeli Defense Forces volunteered for this study. Blood was collected at baseline and at 2 and 4 months. Serum was analyzed for biomarkers reflecting bone formation (BAP and PINP), bone resorption (CTx and TRAP5b), nutrition (PTH, calcium, and 25-OH-D), and the cytokine response (IL-1, IL-6 and TNFalpha). A repeated measures ANOVA was used to analyze changes over time and between genders for each variable. Bone formation and resorption markers were higher in men than women (p<0.001) over the training period, though there were no gender differences in the response to training. For men and women combined, an increase in bone turnover was evident from 0 to 2 months as reflected by significant increases in BAP, PINP, CTx, and TRAP5b (see table), which decreased or stabilized from 2 to 4 months. 

Table  .  

*denotes increase for both genders 0–2 mos (p<0.001); **denotes decrease for both genders 2–4 mos (p<0.01)

Serum calcium and PTH decreased similarly in both genders from 0–2 months, returning to baseline by 4 months. 25(OH)D decreased over the study period in men, but not women. IL-1B, IL-6, and TNFalpha values did not differ between genders, and did not change over the training period. Our results show that a four-month strenuous recruit training regimen increased markers of bone formation and resorption in both men and women during the first two months of training. These data suggest that rapid onset of strenuous training initially results in acceleration of bone turnover. Whether this is related to changes in bone that predispose women to stress fracture warrants further examination.

Disclosures: R.K. Evans, None.

W488

The Functional Muscle-Bone Unit in Young Active Women. P. C. Fehling, L. Ishkanian*, K. Hawkins*, A. Lange*. Exercise Science, Skidmore College, Saratoga Springs, NY, USA.

Several authors have proposed a functional bone to muscle unit (FBMU) defined as the ratio of bone mineral content (BMC) to muscle (BMC/muscle)^1,2,3 to characterize the relationship between bone and muscle. However, which measure of bone and muscle to use in calculating FBMU remains unclear. Therefore, this study investigated FBMU in young, healthy women, using various measures of bone and muscle to calculate FBMU.

Participants in the study were 39 women (20.1 yr ± 1.1); categorized as advanced dancers (DN; n=15), Division III volleyball players (VB; n=9) or non-athletic controls (CN; n=15). A total body (TB) DXA scan was acquired to assess total BMC and leg BMC (Lunar DPX-IQ, V.4.5). A 1 RM leg press (LP; kg) and squat (kg) were used to asses leg strength. The muscle portion of FBMU (denominator) was calculated using TB fat free mass (FFM: kg), TB weight (TBW; kg), leg FFM (kg), leg mass (kg) or strength (leg press & squat; kg). Differences among groups were assessed with a 1-way ANOVA with a Tukey Post Hoc analysis. An ANCOVA was used to adjust for height and weight group differences. Significance level was set at p < .05.

The VB were significantly taller (172.2 vs. 163.3 and 163.1 cm) and heavier (65.8 vs. 59.5 and 59.0 kg) than the DN and CN groups, respectively. After adjusting for height and weight the DN and VB had significantly greater BMC than the CN in the legs; had greater TB and leg FFM; and were stronger (all measures). However, when FBMU was calculated using various measures of bone and muscle (mass or strength), no consistent FBMU pattern emerged (Table 1). For example, when FBMU was expressed as TB BMC/squat, CN had significantly greater FBMU than DN & VB. However, when FBMU was express as leg BMC/squat CN were significantly lower than VB. It appears that depicting the physiological relationship between bone tissue and muscle tissue using FBMU is problematic due to functional, theoretical and practical measurement issues. Several questions about FBMU remain unanswered, including, what does this variable tell us and how should it be calculated?

¹ Rittweger, J et al. Bone. 2000; 27: 319–326

² Sumnik, Z et al. J Musculoskelet Neuronal Interact. 2006; 6: 195–200

³ Dowthwaite, J et al. J Clin Densitom. 2007; 10:65–75 

Table  .  

*FBMU (g/kg), × ± se

^a CN > VB & DN; ^b CN > DN; ^c DN & VB > CN; ^d VB > CN; p < .05

Disclosures: P.C. Fehling, None.

This study received funding from: Skidmore College Opportunity Funds.

W489

Characterization of Muscle Disuse Atrophy and Recovery in Mice Using an In Vivo Casting Model. L. V. Hale*, B. C. Yaden*, J. L. Andrews*, L. M. Helvering*, S. T. Estrem*, F. Lawrence*, S. Li*, S. McAhren*, G. Field*, H. A. Bullock*, V. Krishnan. Musculoskeletal, Lilly Research Laboratories, Indianapolis, IN, USA.

Atrophy of skeletal muscle is characteristic of sarcopenia, cancer cachexia, and disuse in both humans and rodents. Treatment of muscle atrophy is an unmet therapeutic need in these conditions and several preclinical models have been previously described. We have developed and characterized an in vivo model of disuse muscle atrophy in mice by immobilizing one hindlimb in the plantar flexion position by means of a cast. This model is very well-tolerated by the animals and results in no body weight changes with casting. Muscle loss is observed as a decrease in wet weight in both the gastrocnemius and soleus within one day of immobilization with an increasing gradual loss (15–20%) over a 7 day period. This loss is largely reversed after 3 weeks following removal of the cast and reloading of the hindlimb by the animal. Upon reloading, there is edema and some additional loss in gastrocnemius wet weight before noticeable improvement in muscle mass is observed. Atrophied and control gastrocnemius muscles were also subjected to microarray analysis and compared histologically. The beta-adrenergic agonist, Clenbuterol (2mg/kg/day), is able to diminish the extent to which the gastrocnemius atrophies during casting. IGF-1 mRNA is elevated in both the Clenbuterol casted and contralateral legs. We conclude that this in vivo model allows for consistent and characteristic muscle wasting and is useful for the evaluation of pharmacological agents which may prevent atrophy and/or enhance the recovery of muscles which are atrophied from disuse.

Disclosures: L.V. Hale, None.

W490

Electrically Stimulated Eccentric Muscle Contraction Benefits Bone in the Tibia of Hindlimb Unloaded Rats at Both the Mid-Diaphysis and Proximal Methaphysis. L. R. Sumner*¹, M. L. Nilsson*², J. M. Swift*², J. M. Jeffery*¹, C. J. Walthall*¹, J. L. Stallone*², S. A. Bloomfield², H. A. Hogan¹. ¹Mechanical Engineering, Texas A&M University, College Station, TX, USA. ²Health and Kinesiology, Texas A&M University, College Station, TX, USA.

The objective of this study was to investigate controlled electrical muscle stimulation as a countermeasure for bone degradation due to hindlimb unloading in adult male rats. The stimulation protocol uses fine wire electrodes to stimulate the sciatic nerve and induce contractions of the muscles of the lower left hindlimb. The ankle is dorsiflexed during stimulation to create eccentric contractions of the posterior muscles below the knee. In the current study, each session of this simulated resistance training (SRT) consisted of 4 sets of 5 contractions (1000ms) with ankle torques equal to 100% of peak isometric torque. Six-mo. old male Sprague Dawley rats were divided into 3 groups of 12 each: CC (cage control) animals were allowed normal ambulation; HU animals were hindlimb unloaded using standard protocols for 28 days; HU+RT animals were hindlimb unloaded for 28 days with SRT (every other day). The mid-diaphysis and proximal metaphysis of the tibia were assessed by peripheral computed tomography (pQCT). In addition, reduced platen compression (RPC) testing was used to evaluate the mechanical properties of the cancellous bone of the proximal metaphysis. On a separate group of animals, strain gages were applied to the anteromedial surface of the tibial metaphysis. Values ranged from 800 to 1200 microstrain, which are levels generally expected to be osteogenic. The SRT protocol was highly effective as a countermeasure at both sites in the tibia. At mid-diphysis, total BMD was significantly higher for HU+RT than both CC (7.2%) and HU (7.0%). Total BMC was similarly higher in HU+RT (7.8% vs. CC; 14.2% vs. HU). Mid-diaphysis cross-sectional area was also higher in HU+RT (6.6% vs. CC; 13.2% vs. HU). Beneficial effects of SRT were even more dramatic at the proximal metaphysis. Total BMD was higher in HU+RT (13.4% vs. CC; 16.8% vs. HU), as was total BMC (2.4% vs. CC; 21.9% vs. HU). Ultimate stress of the cancellous bone of the metaphysis was 175% higher for HU+RT compared to CC and 268% higher compared to HU. To our knowledge, these data (along with those first presented by our laboratory last year; JBMR 2006 21 (SI): S171) are the first to demonstrate not only effective mitigation of bone loss but absolute gains in bone mass during a simulated microgravity exposure with a physiological exercise intervention. In the earlier study, each exercise session consisted of 4 sets of 10 contractions (500ms) with ankle torques of 120% of peak isometric torque

Disclosures: H.A. Hogan, None.

This study received funding from: NASA Cooperative Agreement NCC-948 with the National Space Biomedical Research Institute.

W491

The Effects of Endurance Running Training on Bone Quality in Growing Rats. T. Huang¹, R. Yang², F. Chang*³, S. Lin*², S. Liu*⁴, S. S. Hsieh*⁵. ¹Institute of Physical Education, Health and Leisure Studies, National Chengkung University, Tainan, Taiwan. ²Department of Orthopaedics, National Taiwan University, Taipei, Taiwan. ³Department of Physical Education, National Taiwan Normal University, Taipei, Taiwan. ⁴Institute of Toxicology, National Taiwan University, Taipei, Taiwan. ⁵Institute of Sports Science, National Taiwan Normal University, Taipei, Taiwan.

To investigate the effects of two different endurance-running modes on bone quality in the growing rats. Thirty-one male Wistar rats were randomly assigned into three groups: the END group (n=10), undergoing continuous endurance treadmill running; the INT group (n=11), undergoing intermittent endurance treadmill running; the CON group, served as sedentary control group (n=10). For equalizing the training volume, the daily training distance was set equal for both exercise groups through out the eight-week training period. Intensity and distance of the two training programs were progressively increased. Both exercise groups showed decreased body weight gain after 8 weeks of running training. Regarding the effects of training on aerobic capacity, both exercise groups showed significantly higher levels in citrate synthase of soleus and heart muscle (p< .05) after eight-week training. Densitometric analysis of femur showed a significantly lower areal bone mineral density (aBMD) in both exercise groups as compared with CON group (p< .05). A three-point bending test was used to measure the biomaterial properties of bone. In extrinsic biomechanical analysis, both the END and INT groups showed significantly higher energy to maximal load and fracture load (p< .05), whereas significantly lower in bending stiffness as compared to the CON group. In intrinsic biomechanical analysis, both exercise groups showed a significantly higher bending stress, bending toughness and Young's modulus (p< .05). Additionally, the INT group was even better than the END group in some of the intrinsic biomechanical parameters. In conclusion, either continuous or intermittent endurance training did benefit the biomaterial bone properties but seemed not to be necessarily associated with aBMD. In addition to conventional geometric and densitometric measurements, further study would be important to investigate the contribution of exercise-enhanced micro-architecture on bone quality.

Disclosures: T. Huang, None.

This study received funding from: National Science Council.

W492

Hypergravity Prevents Bone Loss in Rats Following Ovariectomy. A. Kawaguchi*¹, T. Ninomiya*², Y. Nakamichi*², M. Nakamura*³, N. Udagawa³, N. Takahashi², H. Kato*¹, K. Takaoka⁴, S. Wakitani*⁴. ¹Department of Orthopedic Surgery, Shinsyu University of Medicine, Matsumoto, Japan. ²Division of Hard Tissue Research, Institute for Oral Science, Matsumoto Dental University, Shiojiri, Japan. ³Department of Biochemistry, Matsumoto Dental University, Shiojiri, Japan. ⁴Department of Orthopedic Surgery, Osaka City University Graduate School of Medicine, Osaka, Japan.

There are a lot of reports that characterize the negative effects of microgravity environment on the bone. However, little is known about the effects of hypergravity on bone loss, such as after ovariectomy in rats. Therefore, the purposes of this study were to examine whether hypergravity prevents bone loss after ovariectomy, and to elucidate the mechanisms of hypergravity effects on bone formation and bone resorption. Adult sixteen female Sprague-Dawley rats were randomized to four groups of four rats (sham or ovariectomized, and control or hypergravity exposed). Rats were performed ovariectomy or sham operation at 20 weeks of age, and the gravity treatment was started at the next day of operation. Hypergravity group were subjected to about 3G by centrifugation in accelerated gravity loading machine. The control group at 1G in an identical compartment on the floor of the centrifuge room experienced the same noise, temperature, and light environment as the hypergravity groups. The centrifuge was run continuously for 28 days, except for one hour stops each day for rat care. All animals were triple labeled with tetracycline hydrochloride and calcein injection. After 28 day of centrifugation, femur bones were excised and used for structural analysis by micro computed tomography, measurement of bone mineral density (BMD) by PQCT, and histomorphometric analysis. Micro computed tomography images showed that cancellous bone was greatly decreased in OVX control group, but the bone loss was prevented in OVX with hypergravity group. As a result of PQCT analysis, sham and OVX treated with hypergravity had significantly higher BMD of trabecular bone in distal femur metaphysis compared to control groups. In histomorphometric analysis, OVX decreased BV/TV as a result of increased bone turnover, but OVX with hypergravity group had significantly higher BV/TV, and significantly lower Ob.S/BS, BFR/BS, MAR, N.Oc/BS, OcS/BS. Sham groups had same tendency as OVX groups. These results demonstrate for the first time that hypergravity can suppress high bone turnover following Ovariectomy, and resulting in prevention of bone loss in aged OVX rats.

Disclosures: A. Kawaguchi, None.

W493

Association of Body Weight and Daily Walking Steps with QUS Parameters and Urinary DPD in Elderly Japanese Women. J. Kitagawa¹, Y. Nakahara*². ¹Human System Science, Tokyo Institute of Technology, Tokyo, Japan. ²Health and Nutrition, Wayo Women's University, Chiba, Japan.

High levels of physical activity (PA) have positive effects on quantitative ultrasound (QUS) parameters of the calcaneus and bone resorption markers. However, it remains unclear whether the effects of PA are similar between elderly women with a high body weight and a low body weight. The aim of this study was to investigate the association of body weight and daily walking steps, as an outcome measure of PA, with QUS parameters and urinary DPD in elderly Japanese women.

The subjects were 114 elderly women aged 60 to 85 years (71.4±;4.8 years). They were members of a senior citizen's club. Subjects with a history of disease known to affect bone metabolism were excluded. QUS parameters (SOS, BUA and Stiffness) in the right calcaneus were measured with an A-1000 (Lunar, USA). A pedometer (HJ-005, Omron, Japan) was given to each subject. The subjects were instructed to wear the pedometer during waking hours for 7 consecutive days.

There were significant decreases in body weight and daily walking steps with age in the simple regression analysis. Then, the subjects plotted on the upper part and lower part of the regression line were defined as a high body weight group (BW⁺) and a low body weight group (BW⁻) respectively. Similarly, the subjects were divided into a high walking steps group (WS⁺) and a low walking steps group (WS⁻) according to the regression line for daily walking steps plotted against age. Finally, the subjects were assigned to one of four groups (BW⁺WS⁺; n=17, BW⁺WS; n=35, BW⁻WS⁺; n=33, BW⁻WS⁻; n=29). The associations of body weight and walking steps with QUS parameters and DPD were analyzed by two-way ANOVA in which body weight and walking steps were used as factor variables. There were no significant differences in age among the groups.

The main effects of body weight and walking steps on Stiffness were significant (p<0.001, p<0.05, respectively). Although there was no significant body weight × walking steps interaction for Stiffness (p=0.16), BW⁺WS⁺ had a significantly (p<0.05) greater Stiffness compared with BW⁺WS⁻, whereas there was no significant difference in Stiffness between BW⁻WS⁺ and BW⁻WS⁻. The main effect of walking steps on DPD was significant (p<0.05), whereas that of body weight was not significant. The body weight × walking steps interaction for DPD was not significant. BW⁻WS⁺ showed significantly (p<0.05) lower DPD compared with BW⁺WS⁻. BW⁺WS⁺ showed non-significant 16% lower DPD compared with BW⁺WS⁻. These results indicate that there may be additive effects of walking steps on QUS parameters for elderly women with a high body weight. On the other hand, higher levels of walking steps may be effective in reducing DPD irrespective of the body weight levels.

Disclosures: J. Kitagawa, None.

W494

Role of the L-Type Voltage-gated Calcium Channel Cav1.3 in Bone Mechanotransduction. J. Li¹, C. H. Turner². ¹Department of Biology, Indiana University Purdue University Indianapolis, Indianapolis, IN, USA. ²Department of Orthopedic Surgery, Indiana University School of Medicine, Indianapolis, IN, USA.

Osteoblasts respond to mechanical stimulation with a rapid increase in intracellular calcium ion that is dependent on both extracellular calcium entry and release of calcium ion from intracellular stores. Previous studies have demonstrated that extracellular calcium entry via L-type voltage sensitive calcium channels (L-VSCC) is important in osteoblast proliferation. An alpha 1 subunit forms the pore of the channel and contains the sequences for the dihydropyridine receptor and the voltage sensor. CaV1.3 (alpha 1D) is one of the four different genes that code for the L-VSCC alpha 1 subunit and has been found in osteoblastic cells. We hypothesized that CaV1.3 plays a role in bone growth and mediates mechanically induced bone formation. In order to test this hypothesis, we measured the bone length and bone mineral content in CaV1.3 knockout (KO) mice (obtained from D James Surmeier, Northwestern University, Chicago, USA). We also investigated the skeletal response to loading in CaV1.3 KO mice. The body weight of male KO mice was significant lower than the wildtype male mice (p < 0.05). There was no difference in femur length and bone mineral content of femur and tibia between KO and WT control mice. Axial loading of the ulna with a peak force of 2.5 N (2 Hz for 120 cycles) for 3 days induced new lamellar bone formation on the periosteal surface at midshaft in both KO and normal control mice. There was no difference in mechanically induced mineralizing surface and bone formation rate between KO and WT mice (p = 0.12 and p = 0.17). Others have shown that L-VSCCs are important for osteoblast signaling after mechanical loading. However, our data suggest that CaV1.3 is not necessary for bone growth and skeletal response to mechanical loading. We conclude that the CaV1.3 subtype of L-VSCC is not primarily responsible for the role of L-VSCC in bone development and mechanotransduction in osteoblasts.

Disclosures: J. Li, None.

This study received funding from: NASA.

W495

Mechanical Enhancement of Bone Quality Is Paralleled by Suppression of Fat Production: Can Osteoporosis and Obesity be Countered Through the Same Mechanism?, Y. K. Luu*¹, E. Capilla*², B. J. Lee*¹, J. E. Pessin*², S. Judex¹, C. T. Rubin¹. ¹Biomedical Engineering, Stony Brook University, Stony Brook, NY, USA. ²Pharmacology, Stony Brook University, Stony Brook, NY, USA.

An inverse yet interdependent relationship between osteoblasts and adipocytes in the marrow cavity has been identified, emphasizing that both bone and fat stem from a common precursor — the mesenchymal stem cell. We posit that the introduction of a low magnitude mechanical signal that is anabolic to bone can in effect alter the balance of bone and fat development at the tissue level, and while improving bone quality could also serve as an effective preventative measure towards the onset of obesity. Twenty four C57/BL6 male mice, 7wks of age, were randomly split into two groups, VIB: 12 wks of whole body vibration (0.2g, 90Hz, 15min/d, 5d/wk), and PL: an equal number of placebo sham controls. To promote adiposity, all animals were fed a high fat diet (45 kcal % fat) through the study duration. Body mass gains and the average food intake/wk were similar for both vibrated and control groups. In vivo microCT scans at 12 wks were used to quantify bone and fat (subcutaneous and visceral) of the entire torso of each animal. Normalized to body mass, VIB showed a 6.6% increase (p = 0.015) in lean volume and 5.3% increase (p = 0.098) in bone volume over PL. Post-sacrifice, high resolution (12 μm) microCT scans of the left tibia of VIB indicted a 13.8% (p = 0.058) increase in trabecular bone volume over PL. Additionally, connectivity density (+27.1%, p = 0.058) and trabecular number (+12.1%, p = 0.060) in VIB were increased over PL, while trabecular spacing (-12.5%, p = 0.051) decreased.

In contrast to the increase in bone quantity and quality measured in VIB, these animals also had 28.5% less fat volume (p=0.030) as compared to PL. Weights of epididymal fat pads harvested at sacrifice correlated strongly with CT data (R² = 0.80), and also demonstrated a reduction in VIB animals. That the anabolic effect to bone is achieved at the “cost” of a significant reduction in adiposity, achieved without a metabolic challenge (i.e., the signal is low and the duration is short), suggests that these mechanical signals influence the skeletal and fat phenotype by preferentially driving the lineage commitment and subsequent differentiation of the mesenchymal stem cells towards bone.

Disclosures: Y.K. Luu, None.

This study received funding from: NIH (AR 43498), NASA (NAG 9-1499), and a Wallace H. Coulter Early Career Translational Research Award.

W496

Using pQCT to Assess Regional Bone Changes Resulting from Short-Term Exercise Interventions. C. H. Negus¹, R. K. Evans*², W. Shen*¹, H. M. Isome*², M. L. Lester*², D. E. Catrambone*², B. A. Spiering*³, D. L. Hatfield*³, W. J. Kraemer*³, B. C. Nindl*². ¹L-3 Jaycor, San Diego, CA, USA. ²U.S. Army Research Institute of Environmental Medicine, Natick, MA, USA. ³University of Connecticut, Storrs, CT, USA.

The purpose of this randomized controlled trial was to evaluate the effect of several short term exercise interventions over 13 weeks on regional bone strength parameters of the tibia using peripheral quantitative computed tomography (pQCT). Fifty-seven female volunteers (age 20.4 ± 1.8) were assigned to one of four groups: a sedentary control group, a resistance training group, an aerobic group, or a combined aerobic-resistance group. pQCT scans were taken at sites 4%, 38% and 66% from the distal end plate of the tibia at baseline and at the end of the 13-week training period. Cross-sectional images were divided into six 60° sectors centered on the intramedullary canal. Matlab was used to register and align images and to assess geometry (4%: trabecular area; 38%, 66%: cortical area, periosteal diameter, cortical thickness, & cross-sectional moment of inertia), density (4%: trabecular density; 38%, 66%: cortical density) and overall bone strength (38%, 66%: bone strength index, Slenderness Index). A repeated-measures ANOVA compared the four training groups over time; Fisher's LSD was used to analyze significant interactions. Trabecular density increased significantly at the 4% site with exercise intervention, while the control group exhibited no change. Changes were most pronounced in the groups incorporating impact exercise, and were evident on the medial aspect of the tibia. Measures derived from the 38% and 66% site did not change significantly. 

Table  .  

Trabecular density (mg/cm³) at the 4% site. Values expressed as mean ± SD. ± significance at p<0.01

These results indicate that a subtle regional increase in trabecular density may be the earliest manifestation of improvements in bone strength resulting from exercise intervention.

Disclosures: C.H. Negus, None.

W497

Load-specific Differences in Femoral Neck Cortical Geometry: Preliminary data from 3-D MRI Study of Proximal Femur of Athletes. R. Nikander¹, H. Sievänen¹, P. Dastidar*², A. Heinonen³, P. Kannus*⁴. ¹Bone Research, UKK Institute for Health Promotion Research, Tampere, Finland. ²Department of Radiology, University Hospital, Tampere, Finland. ³Department of Health Science, University of Jyväskylä, Jyväskylä, Finland. ⁴Department of Trauma, Musculoskeletal Surgery and Rehabilitation, University and University Hospital, Tampere, Finland.

During normal walking, femoral neck (FN) is subjected to a large number of consecutive, low magnitude impacts from typical directions. In contrast, during sport performance, power-lifters' femoral neck experiences very high magnitude, low rate loading from unusual direction, while swimmers' femoral neck is subjected to high rate muscle activity lacking virtually the weight-bearing component. We aimed to investigate the differences in the femoral neck cortical geometry among world-glass power-lifters, national level swimmers and normal exercisers serving as referents.

In this preliminary analysis of accumulating bone data from five power-lifters, five swimmers and five referents were compared. In all groups, the femoral neck cortical structure was assessed with DXA and 3-D MRI of the proximal femur. ANCOVA (age, weight and height as covariates) were used for statistical analysis.

The femoral neck BMC and BMD were 35% and 31% higher in power-lifters, but 11% and 13% lower in swimmers compared with controls (0.02<p<0.07). Interestingly, the shape of power-lifters' femoral neck appeared to be round at the narrowest section, while swimmers' and controls' FN seemed to be oval being wider in the superior-inferior direction.

Although the inter-group differences in the femoral neck cortical thickness did not reach statistical significance, a clear trend suggesting substantially thicker cortices in power-lifters in all anatomic directions (anterior 22%, posterior 52%, superior 12% and inferior 45%) was found, while the cortices between swimmers and referents were similar.

The strong femoral neck structure (more mass, denser bone, round shape and thick cortices) seemed to be characteristic of power-lifting performance. High magnitude loading produced at low rate seemed to account for strong femoral neck, while non-weight bearing, low-magnitude loading, despite large number of repetitions at relatively high rate, seems not benefit to bone structure at all.

Disclosures: R. Nikander, None.

W498

The Effect of Resistance Exercise on In Vivo-Derived Tomography Measures and Mechanical Properties of Rodent Bone: Is More Load Always Better?, M. Nilsson*¹, J. M. Swift*¹, C. J. Walthall*², J. L. Stallone*¹, H. A. Hogan², J. D. Fluckey*¹, S. A. Bloomfield¹. ¹Department of Health and Kinesiology, Texas A&M University, College Station, TX, USA. ²Department of Mechanical Engineering, Texas A&M University, College Station, TX, USA.

Weight-bearing resistance exercise and high-impact jump training have previously been shown to be particularly beneficial to skeletal growth compared to other modes of exercise. Exercising against an increased resistance, such as traditional strength training, would theoretically yield higher bone strains compared to exercise with just your own bodyweight (BW), and is expected to produce a greater anabolic response in bone. The purpose of this study was to compare the osteogenic effects of a 5-week progressive resistance exercise paradigm with a BW exercise protocol in rats. Mature male Sprague-Dawleys (mean BW of 407 ± 26.5 g) were operantly conditioned to perform a full leg extension to reach an illuminated lever (mimicking a traditional squat), and thereafter assigned to a resistance exercise group (RT; n=16), a BW exercise group (BW-EX; n=15), or a cage-control group (CC; n=7). Three training sessions were performed per week and resistance was progressively increased from 80 g during session 1 to 410 g during session 15 in the RE group. The BW-EX completed the same number squats per session as the RT group, but without added weights, resulting in many rats being able to jump off the cage floor. In vivo peripheral quantitative computed tomography scans (pQCT) of midshaft and proximal tibiae were done at baseline and after 5 weeks of training. Strength, stiffness, and toughness of mid-diaphyseal tibial bone were assessed with 3-point bending tests to failure post sacrifice. Ex vivo pQCT scans and bending/compression tests of the femoral neck were also performed. RE and BW-EX resulted in similar increases in tibial total bone mineral content (BMC) at the metaphysis (+10% and +11%, respectively) and midshaft (+19% and +18%, respectively). Diaphyseal bone area, cortical shell thickness, and polar moment of inertia increased significantly in RT (+14%, +9%, and 29%, respectively) and BW-EX (+13%, +9%, and +26%, respectively), and both groups had significantly stronger, stiffer, and tougher bone compared to CC (p < 0.05). Femoral neck total BMC (p<0.001) and max force (p<0.01) were significantly higher in exercise groups compared to CC, but there were no differences between RT and BW-EX. In summary, increasing load magnitude by up to ∼100% of the animal's bodyweight in this voluntary resistance exercise paradigm did not produce additional gains in BMC, bone geometry, or mechanical properties. We speculate that the additional impact provided by jumping in the BW-EX group may account for the similar benefits.

Disclosures: M. Nilsson, None.

This study received funding from: Huffine's Institute, Texas A&M University.

W499

Musculoskeletal Response to Altered Mechanical Demand Is Tissue Specific and Influenced by Genetic Make-up. E. Ozcjvici¹, S. Lublinsky*¹, C. Rubin¹, L. Donahue², S. Judex¹. ¹Biomedical Engineering, SUNY Stony Brook, Stony Brook, NY, USA. ²Bone Biology Group, Jackson Labs, Bar Harbor, ME, USA.

The degree of musculoskeletal devastation by the loss of functional weightbearing is heavily influenced by genetic variations. Here, we subjected genetically heterogeneous mice to 3wk of disuse (hindlimb unloading) and 3wk of subsequent reambulation to investigate: (a) if the degree of mechanosensitivity of cortical bone, trabecular bone and muscle are similar, (b) whether the amount of tissue at baseline is associated with disuse induced changes in morphology (c) to what extent the loss in tissue morphology can be recovered upon reambulation. In vivo μCT quantified longitudinal changes in metaphyseal trabecular bone, middiaphyseal cortical bone, and muscle surrounding cortex in the femur of adult (4mo) F2 female progeny of BALB and C3H (high and low mechanosensitivity) mice (n=450). During disuse and reambulation, changes in cortical area amounted to −1(±4)% and 4(±4)%, respectively. Age-matched controls (n=29) added 3(±1)% and 4(±1)% of cortical bone in an identical time frame indicating that appositional growth was retarded with disuse. Muscle loss during disuse was 9(±11)%, 82% of which was recovered upon reambulation. Trabecular bone was devastated during disuse as characterized by a 44(±17)% smaller volume fraction and a 18(±8)% drop in both trabecular number and thickness. Recovery of trabecular volume fraction upon reambulation reached only to 10% of the loss during disuse. Even worse, trabecular number was reduced by an additional 8(±9)% upon reambulation while 71% of the loss was recovered in trabecular thickness. Baseline trabecular volume fraction and muscle area were inversely related to the loss during disuse (R2=34% and 16%, respectively), while changes in cortical area were not associated with baseline morphology (R2<1%). The relation between the loss of skeletal tissue during disuse and gain upon reambulation was weak (R2<10%) for both trabecular and cortical bone, while in muscle, a moderate correlation (R2=32%) was observed. In summary, changes in mechanical demand resulted in a very tissue- and compartment-specific response with large differences between muscle, cortical bone, and trabecular bone. The data also suggest a genetically regulated protective effect of the quantity of trabecular and muscle tissue at baseline on the preservation of tissue during disuse. In contrast to muscle that recovered most of its losses upon reambulation, the recovery of trabecular bone was minimal with some architectural aspects even continuing to deteriorate. The large variations between individuals emphasizes that the identification of the underlying molecular basis will be invaluable for the future development of tailored treatments.

Disclosures: E. Ozcivici, None.

This study received funding from: NASA.

W500

Males Have an Enhanced Osteogenic Responsiveness to Loading vs. Females — A Consequence of Higher Mechanical Competence of Female Bones Relative to Locomotive Loading. I. Pajamäki*¹, O. Leppänen*¹, L. Jokihaara*¹, H. Sievänen*², P. Kannus*², T. L. N. Järvinen¹, ¹Orthopaedics and Traumatology, University of Tampere, Tampere, Finland, ²UKK Institute, Tampere, Finland.

Sex is considered one of the primary factors modulating the responsiveness of bone to mechanical loading. The purpose of this study was to assess whether the loading-induced bone gain (mechanoresponsiveness) differs between young male and female rats.

A total of 40 three-week old male (M) and female (F) rats were used. At entry, unilateral sciatic neurectomy was performed to remove loading from left limb (L-) while the contralateral right limb remained intact (subjected to normal physiological loading, L+). After a 6-month study period, a comprehensive structural analysis of the femoral midshaft was carried out utilizing peripheral quantitative computed tomography (pQCT) and mechanical testing. Total cross-sectional area (tCSA), cortical cross-sectional area (cCSA), cortical bone mineral content (cBMC) and fracture load (Fmax) were determined. To eliminate the inherent bias arising from comparisons between animals differing in body weight and size, all data pertaining to bone mechanical competence was equalized in terms of the animal's apparent loading environment by using the body weight and femoral length as covariates

A clear sex-difference was observed in the mechanoresponsiveness, males exhibiting significantly higher loading-induced responses than females (Figure). However, it should be noted that the non-loaded (L-) bones of females displayed clearly higher mechanical competence relative to the animal's apparent loading. Thus, it is quite plausible that there is no need for these already “overstrengthened” female bones to adapt to mechanical loading with the same magnitude as the male bones.

In conjunction with our previous findings (Järvinen et al. Bone 2003 and JBMR 2003), the reduced mechanoresponsiveness of female bones seems to be explained by the difference in the mechanical (locomotive) competence of bones between sexes. Thus, our results emphasize the importance of acknowledging the locomotive context (existing bone characteristics) in studies evaluating the mechanoresponsiveness of bones 

Figure Figure.. The mechanoresponsiveness of female and male bones. Bars represent mean ± standard error of mean (SEM). Significant differences between L+ vs. L- are indicated as follows: *** p < 0.001. The p-value for interaction (sex and loading) is presented on top of each figure.⊐ denote the difference in the normal (control) bones between females and males.

Disclosures: I. Pajamäki, None.

W501

Effects of Vibration Plus Resistance Training on Bone Metabolism in Postmenopausal Women. I. J. Palmer*, M. G. Bemben*, D. A. Bemben. Health and Exercise Science, University of Oklahoma, Norman, OK, USA.

Whole body vibration has been shown to be osteogenic in animal models, however the efficacy of vibration as an intervention for the prevention of osteoporosis in humans is not clear. The purpose of this study was to examine the effect a vibration stimulus preceding resistance exercise on bone mineral density (BMD) and bone biomarkers in healthy estrogen-deficient postmenopausal women, 55 to 75 years of age. Subjects were assigned to a control group (C, n=12), resistance training group (R, n=22), or a vibration plus resistance training group (VR, n=21). R and VR trained 3 days/week for 8 months, with 3 sets of 10 reps (80% of IRM) on 8 exercises. VR received vibration in three different positions beginning with 1 set of 15 sec (30Hz, 2.16g) and progressing to 2 sets of 60 seconds (40Hz, 2.8g). Training loads were adjusted every 5 weeks. Daily calcium intake was estimated by a food frequency questionnaire. Bone-specific alkaline phosphatase (BAP) and C-terminal telopeptides of Type-1 collagen (CTX) were measured as indicators of bone turnover. BMD and bone mineral content (BMC) of the AP lumbar spine, dual proximal femur, radius 33%, and total body were measured by DXA (GE Lunar Prodigy, enCORE version 8.80.001). At baseline, there were no significant (p>.05) group differences in age, height, weight, strength, calcium intake, BMD, or BMC. Two-way repeated measures ANOVA (group x time) did not detect significant group or time effects for BAP, CTX, or BMD or BMC at any site. VR had significantly (p<0.05) higher relative strength increases in 5 of the eight exercises, compared with R. Multiple regression analyses were used to determine significant (p<0.05) predictors of absolute changes in BMD and BMC in response to training for VR and R groups. Total BMD was predicted by upper body strength change (R²=0.089 p=0.031). Right and left trochanter BMD was predicted by fat mass change (R²=0.083 and 0.127, p<0.05). Right femoral neck BMD was predicted by change in BAP (R²=0.113, p=0.019), by fat mass change (R²=0.108, p=0.021), and by weight change and calcium intake change (R²=0.203, p=0.005). Right total hip BMD was predicted by change in calcium intake (R²=0.12, p=0.015). Spine (L2-L4) BMD was predicted by change in lean mass (R²=0.079, p=0.042). Radius 33% BMD was predicted by change in fat mass (R²=0.091, p=0.028) and by group (R²=0.074, p=0.045). In conclusion, the vibration plus resistance training protocol did not result in significant alterations in bone biomarkers, BMD, or BMC. Although the addition of the vibration stimulus to a traditional resistance training program significantly enhanced muscle strength, no benefits for skeletal health were evident.

Disclosures: I.J. Palmer, None.

This study received funding from: Graduate College, University of Oklahoma.

W502

Flexible Multibody Simulation Approach in The Analysis of Tibial Strains During Walking. T. Rantalainen*¹, R. Al Nazer*², A. Heinonen³, H. Sievänen⁴, A. Mikkola*². ¹Department of Biology of Physical Activity, University of Jyväskylä, Jyväskylä, Finland. ²Department of Mechanical engineering, University of Lappeenranta, Laappeenranta, Finland. ³Department of Health sciences, University of Jyväskylä, Jyväskylä, Finland. ⁴UKK-Institute, Tampere, Finland.

Strains within the bone play a major role in bone remodeling. These small deformations can be determined using experimental strain gage measurements which are challenging and invasive tasks. Further, the strain measurements are, in practise, limited to certain regions of superficial bones only, e.g. the anterior surface of tibia. In this study, tibial strains occurring during walking were estimated using a numerical approach based on the flexible multibody dynamics. A written informed consent was obtained from healthy male volunteer (25 years, 184 cm, 89 kg), who served as a subject. The study was approved by the local ethical committee. The subject was asked to walk on a level surface at a natural cadence and at a constant velocity (1.47 m/s). The walking performance was recorded using four digital video cameras (COHU High Performance CCD Camera, San Diego CA, USA). A detailed lower body musculoskeletal model was developed by employing motion analysis data obtained from the walking test. The motion analysis data was used in inverse dynamic simulation to train the muscles to replicate the motion in forward dynamics simulation using a commercial human movement modeling software (BRG.LifeMOD Biomechanics Modeler®, Biomechanics Research Group, Inc, USA). The tibial strains were then estimated by applying the forward dynamic simulation to a lower body model with a flexible tibia. The maximum and minimum principal strains predicted by the model were 376 and −472 microstrains, respectively, which are in line with literature values from in vivo measurements. It is concluded that the non-invasive flexible multibody simulation approach may be used as a surrogate for experimental bone strain measurements and be of use in the detailed strain analyses of bones.

Disclosures: T. Rantalainen, None.

This study received funding from: European Regional Development Fund.

W503

The Relationship Between Serum 25-hydroxyvitamin D and Bone Markers in the Professional Football Players. Y. Saita, H. Nakajima*, H. Ikeda*, H. Kurosawa*. Department of Orthopaedic Surgery and Sports medicine, Juntendo University, Tokyo, Japan.

Stress fractures are one of the most common types of overuse injuries in top athletes. A recent report suggested a relationship between the incidence of stress fractures and low 25-hydroxyvitamin D (25OH(D)) concentration in military recruits. However, relationships between 25OH(D) and bone markers in top athletes are not well known. Therefore, serum levels of 25OH(D) and bone markers were measured in the professional football (soccer) players in Japan (n = 32, mean age 24 years). The average values of 25OH(D) and bone markers were as follows; 25OH(D): 25.9 ± 2.8 ng/ml (mean • SD), bone-specific alkaline phosphatase (BAP): 31.3 ± 9.1 U/L, cross-linked N-telopeptides of type I collagen (NTX): 18.8 ± 5.2 nmolBCE/l. The levels of BAP and NTX were elevated above normal range in 28.1% and 62.5% in all players, respectively. There are significant positive correlations between BAP and NTX values (r = 0.37, p<0.05). Then, we assessed the correlations between 25OH(D) and bone markers. The players were divided into two groups according to their 25OH(D) levels (low-vitD group: less than mean value of 25OH(D), n=15; high-vitD group; n=17), and bone markers were compared. Interestingly, low-vitD group showed higher levels of BAP compared to high-vitD group (p<0.05). Similarly, low-vitD group showed relatively higher levels of NTX, though it was not statistically significant. The levels of 25OH(D) and BAP showed negative correlations (r = −0.25), yet it was not statistically significant. These findings indicate that football (soccer) players tend to be in a high turnover state in bone remodeling and the serum 25OH(D) concentration would negatively correlate with it. Further study may reveal whether 25OH(D) level could be a biomarker to predict the occurrence of stress fractures in athletes.

Disclosures: Y. Saita, None.

W504

Bone Mineral Density and Bone Quality in Unilateral Lower Limb Amputees. V. D. Sherk*, M. G. Bemben*, J. T. Cramer*, D. A. Bemben. Health and Exercise Science, University of Oklahoma, Norman, OK, USA.

Bone(s) in a residual limb undergo prolonged unloading after amputation, potentially causing significant bone loss in the hip and distal bony end of the residual limb. The purpose of this study was to examine the effect of amputation on bone geometry, volumetric bone mineral density (vBMD) and areal bone mineral density (aBMD) by comparing the intact and residual limbs in unilateral transfemoral and transtibial amputees. Five above-knee and four below-knee amputee subjects (23–50 years of age) who had been ambulatory with a prosthesis for at least 6 months gave informed consent to participate in this study. Bone geometry and vBMD were assessed at the distal end of the residual limb and comparable cross-sectional slice of the intact limb using pQCT (Stratec XCT 3000). aBMD of the proximal femurs was assessed using DXA (GE Lunar Prodigy, enCORE version 8.80.001). Paired t-tests were used to determine differences in bone variables between the amputated and intact limbs. There were significant differences in the hip BMD sites (p<.05) with total hip aBMD for the amputated side being 32% lower than that of the intact side. Femoral neck and trochanter aBMD were 27% and 37% lower in the amputated hip, respectively. Forty-four percent of amputees had hip T-scores < −2.5. As shown in the table below, total and cortical areas and densities were significantly lower at the end of the residual limb compared to the midshaft of the intact limb (p<0.05), but there were no significant differences in the trabecular area and density. In conclusion, amputees had significant decrements in total bone area and density in the residual limb compared to the intact limb, which was attributed to concomitant changes in the cortical bone variables. Based on the aBMD for the hip sites, these individuals are at increased risk for osteoporosis. 

Table Table 1. Bone Area (A) and Density (D) pQCT Variables (Means ± SE)

*p<0.01

Disclosures: V.D. Sherk, None.

This study received funding from: College of Arts and Sciences, University of Oklahoma.

W505

Effect of Combined Strength and Speed Training on Cortical Bone Geometry and Mass Distribution in Master Athletes. H. Suominen¹, M. Korhonen*¹, J. Hautakangas*¹, T. Suominen*¹, M. Alen*¹, A. Mero*². ¹Department of Health Sciences, University of Jyväskylä, Jyväskylä, Finland. ²Department of Biology of Physical Activity, University of Jyväskylä, Jyväskylä, Finland.

Cross-sectional and longitudinal observational studies have shown that athletes competing in strength and power events have superior bone mineral mass and cross-sectional geometry of long bones compared with their non-athletic counterparts. The purpose of the present study was to investigate the effects of intensive physical training on cortical bone geometry and mass distribution in master sprinters. Seventy-three male athletes aged 40–85 years with long-term experience and success in sprint events were randomly assigned into two groups. The men in the experimental group (n=40) participated in a periodized strength and speed training program for 6 months while the controls (n=32) continued their accustomed training routines. Volumetric bone mineral density (vBMD), cortical thickness (CTh), and polar mass distribution were assessed in the tibia shaft by pQCT (XCT 2000) using a Geanie 2.1. software. No significant interaction of group by time was observed in vBMD, while the mean change in CTh was 2–3% greater in the experimental group compared to controls, particularly among the older sprinters (Fig.). According to polar mass distribution, the changes were more evident in the medioanterior region of the tibia shaft. The results indicate that high-load and high-speed exercise training may improve cortical bone geometry even in elderly athletes with long-term training background.

Disclosures: H. Suominen, None.

W506

Association of Serum Osteocalcin with Bone Gain in Pubertal Girls and Bone Loss in Postmenopausal Women. S. M. Cheng*¹, K. M. Fagerlund*², P. Rahkila*¹, E. Helkala*¹, F. Tylavsky³, F. Tylavsky³, H. K. Väänänen⁴, S. Cheng¹. ¹Department of Health Sciences, University of Jyväskylä, Jyväskylä, Finland. ²Department of Medicine, University of Turku, Turku, Finland. ³Department of Preventive Medicine, University of Tennessee, Memphis, TN, USA. ⁴Department of Preventive Medicine, University of Turku, Turku, Finland.

Serum osteocalcin (OC) has been proposed as a specific and sensitive marker for bone formation because it is present exclusively in bone tissue and increases significantly when skeletal growth becomes boosted. Furthermore, since OC is incorporated into the bone matrix it may be released even during bone resorption. In this study we evaluate whether OC reflects bone modeling in pubertal girls, and bone loss in a group of their mothers and grandmothers. The study subjects consisted of 258 girls (mean age at baseline 11.2 yrs, menarche age 13 yrs), and 80 pre-menopausal mothers (PreM, mean age 43yrs), and 100 post-menopausal mothers and grandmothers (PostM, mean age 63yrs). Serum total OC was assessed by an in-house immunofluorometric assay. Bone mineral content (BMC) and density (BMD) of whole body (WB), total femur (TF) and lumbar spine (L2–4) were assessed by DXA (Prodigy, GE Lunar). Data for the girls were collected at baseline and 12, 24, 36, 48 and 84 months intervals. Mother and grandmother participated in the study twice at 36 and 84 months time points. The correlation between OC (not measured at 84 months) and BMC, BMD and the annual changes of BMC and BMD were calculated for successive measurement interval. In girls the mean OC concentration decreased from 45.5 ± 12.5 ng/ml at baseline to 44.4 ± 11.9 (12m), 42.5 ± 12.9 (24m), 35.1 ± 12.6 (36m), and 25.6 ± 10.4 ng/ml (48m). The decrease was significant from 36 to 48 months. In PreM the OC concentration was lower than in PostM (6.9 ± 2.3 vs 8.5 ± 4.4 ng/ml p=0.023). In girls the OC concentrations between different time points were highly correlated (r=0.387–0.827). In girls a significant positive correlation was found between OC and BMC or BMD of WB, TF, and L2–4 at baseline, but significant negative correlations at 24 months and 36 months. In PostM the OC concentration was negatively correlated with BMC or BMD of the WB. Further, we found that in girls the initial OC value is associated with a positive annual BMC or BMD change during the following measurement interval. For all measured bone sites this positive relationship was strongest during 24–36 months (r=0.447–0.595) and 36–84 months (r=0.701–0.771) intervals. In PostM, the association of OC and the annual change of BMC was negative (TF r=-0.427, p=0.037 and WB r=-0.551, p=0.005). Our results indicate that in pubertal girls serum OC concentration reflects the bone gain during the bone modeling period, and bone loss by resorption in postmenopausal women.

Disclosures: S.M. Cheng, None.

This study received funding from: ASBMR Bridge Funding, Academy of Finland and Finnish Ministry of Educztion.

W507

Sexual Dimorphism in the Relationship Between Serum Leptin and Skeletal Health in Healthy Adolescents. J. Maalouf¹, N. Hwalla*², Z. Mahfoud*³, O. Obeid*², G. El-Hajj Fuleihan¹. ¹Calcium Metabolism and Osteoporosis Program, American University of Beirut-Medical Center, Internal Medicine, Beirut, Lebanon. ²American University of Beirut, Nutrition & Food Sciences, Beirut, Lebanon. ³American University of Beirut, Epidemiology & Population Health, Beirut, Lebanon.

Serum leptin and bone mass are concordant in several respects and both increase at the initiation of puberty. Few studies have investigated the relationship between serum leptin concentrations and bone mineral in both genders in the context of other mediators of bone mineral accretion.

The purpose of this cross-sectional study is to determine the relationship between leptin and bone mass in 363 healthy adolescents (184 boys and 179 girls), 10–17 years old.

Height (Ht), weight, Tanner stages (TS), dietary calcium and exercise were evaluated. Bone mineral density (BMD), content (BMC) and body composition were measured by DEXA. Serum leptin, 25-hydroxyvitamin and IGF1 were also measured.

Girls had higher leptin levels than boys at all TS and reached significance at TS3 (p=0.025) and TS4 (p= 0.012). In girls, leptin levels increased substantially across TS and were two-fold higher in post-pubertal girls than pre-pubertal, while in boys the increments were negligible (p=0.87) (Figure). This significant sexual dimorphism persisted (ß = 4.75 ng/ml, p<0.001) after controlling for fat mass (FM) and TS.

There was a correlation between leptin and BMD at the lumbar spine, total hip (TH), femoral neck (FN), radius and subtotal body (TB) and at FNBMC in boys (r= 0.17–0.26, p<0.05) and at all skeletal sites for BMD and BMC in girls (r=0.28–0.38, p<0.001). Serum leptin was correlated with lean mass (LM) in boys (r= 0.31 p<0.001) and girls (r=0.48, p<0.001). It was highly correlated with FM (r= 0.78, p<0.001) in boys and (r=0.67, p<0.001) in girls.

There was no independent effect of serum leptin on BMD or BMC in boys after adjusting for TS, LM, FM, Ht, exercise, dietary calcium, IGF1 and serum vitamin D. Conversely, in girls, after adjustment, leptin was a consistent predictor of BMD but not BMC at cortical sites i.e. at the TH, FN, trochanter, and TB explaining 4–4.5% of variability in BMD.

Leptin correlates with BMD in girls independent of fat and other classical bone mass predictors. The basis for the sexual dimorphism in the relationship between leptin and bone mass is unclear and deserves further investigation

Disclosures: G. El-Hajj Fuleihan, Nestle 2; Merck KGaA 2.

This study received funding from: Nestle/KGaA.

W508

Analysis and Quantification of Spontaneous Bone Regeneration. T. P. Fairbanks*¹, T. M. Link*², D. Ellis*¹, J. S. Lee¹. ¹Oral & Maxillofacial Surgery, UCSF, San Francisco, CA, USA. ²Radiology, UCSF, San Francisco, CA, USA.

Rib grafts are commonly used for reconstruction in the craniofacial region. Despite the size of the defect at the donor site, the rib will regenerate completely and can be harvested a second time. Little is known about this spontaneous regeneration. This unique ability provides an opportunity to study bone regeneration in the only known site to spontaneously and completely form bone in the postnatal setting. The purpose of this study is to use quantitative CT to analyze in vivo rib regeneration at the costochondral bone graft site. This data will provide further understanding of spontaneous bone regeneration and the effects of age on bone formation.

8 Subjects (3 Female, 5 Male) were enrolled at the UCSF Oral and Maxillofacial Surgery Clinic. IRB approved consent was obtained from each participant for CT imaging at 3 intervals: immediately post-operative (T0), 3- and 6- months follow-up (T1, T2 respectively). A standard imaging protocol based on height/weight was used for each patient in a 16 row multi-slice CT scanner (Lightspeed, GE, Milwaukee, Wisconsin). The CT scans were analyzed at PACS workstations (AGFA, Somerville, New Jersey). Using image analysis software, the CT images were analyzed and quantified for densitometric and volumetric data of the regenerated osseous tissue. Using known values of bone mineral density from the calibration phantom, the computed density was converted from Hounsfield Units to mg/mL hydroxy-apatite. The area of the outlined form was given in units of cm2; volume was calculated by multiplying the computed area in the region of interest by the thickness of the axial CT slice and represented in units of mm3.

To compare regenerative capacity as a function of age, the subjects were assigned to the adult (>16 years) or pediatric (≤ 16 years) group. The adult group: 3 subjects (2 Female, 1 Male); each subject demonstrated incomplete regeneration at T2. The average volume of regenerated bone was 2049.45 mm³, with an average density of 323.76 mg/ml hydroxy-apatite. The pediatric group: 5 subjects (1 Female, 4 Male) exhibited complete rib regeneration, typically by Tl. The average final volume of the young group was 4511.45 mm³, and a density of 305.45 mg/ml hydroxy-apatite.

Despite evidence of bone regeneration in all of the subjects enrolled in the study, the pediatric group exhibited substantial and complete regeneration of osseous tissue compared to the adult group.

Disclosures: T.P. Fairbanks, None.

This study received funding from: UCSF School of Dentistry, Dean's Creativity Fund.

W509

Tempo and Timing of Bone Mineral Accrual During the Pre, Peri and Post Adolescent Growth Periods. M. R. Forwood¹, A. D. G. Baxter-Jones*², R. A. Faulkner², R. L. Mirwald*², D. A. Bailey². ¹School of Biomedical Sciences, The University of Queensland, Brisbane, Australia. ²College of Kinesiology, University of Saskatchewan, Saskatoon, SK, Canada.

Existing data on bone mineral acquisition during growth is limited because most studies have been cross-sectional in design or have failed to adequately control for maturational differences in growing children. Starting in 1991 we have been monitoring bone mineral accrual and bone strength in a sample of children who are now in their mid-twenties. From our original sample we present data on 52 boys and 52 girls who have been measured annually for seven consecutive years across the adolescent growth spurt years and again for up to three years as adults. Bone mineral content of the lumbar spine, femoral neck and total body was measured by DXA (Hologic 2000 QDR in the array mode). To control for maturational differences in children of the same chronological age a cubic spline curve was applied to each subject's longitudinal height data and an age of peak linear growth (PHV) for each subject was determined. Pre-adolescence was then defined as the stage of growth up to 2 years before the age of PHV; Peri-adolescence from 2 years before the age of PHV until two years after PHV. Post adolescence was defined as the stage of growth from PHV +2 until adult status (PHV + 10 years). The purpose of the study was to describe bone mineral accrual at the three bone sites during these three developmental time periods; pre-adolescence, peri-adolescence and post- adolescence. The following graph shows the percent of adult accrual during the four year growth period surrounding PHV (Defined as Peri-adolescence = PHV ±2yr). At all sites, for both sexes, over 98% of bone mineral has been laid down by 4 years beyond the age of PHV.

Disclosures: M.R. Forwood, None.

This study received funding from: National Health Researcfh Development Program, Health Canada.

W510

A Model of Secondary Amenorrhea in the Adolescent Female Rat. R. Joshi¹, V. R. Yingling². ¹Kinesiology, Temple University, Philadelphia, PA, USA. ²Kinesiology & Anatomy and Cell Biology, Temple University, Philadelphia, PA, USA.

Optimizing peak bone mass during late adolescence may be effective in reducing the effects of osteoporosis. Osteopenia, osteoporosis, stress fracture as well as a failure to achieve peak bone strength are all observed in young women with secondary amenorrhea. Multiple factors contribute to bone deficits in secondary amenorrhea including increased bone resorption through suppressed estrogen levels and suppressed bone formation resulting from decreased insulin-like growth factor-1 (IGF-1) levels. Therefore it is important to develop an animal model to reproduce a milieu similar to that of secondary amenorrhea in humans to study the effects of this devastating condition on bone structure. The purpose of this study was to access the effects of secondary amenorrhea induced by gonadotropin releasing hormone antagonist (GnRH-a) injections on serum levels of estradiol and IGF-1. At 23 days of age, 40 female Sprague Dawley rats were randomly assigned into a baseline group (BL) (n=6) sacrificed on day 25, a baseline 65 group (BL65) (n=10) sacrificed on day 65, an aged-match control group (C) (n=15) sacrificed on day 90, and an experimental group (Ex) (n=9) that received daily injections (2.5 mg/kg/dose) intraperitoneally for a 25 day period from day 65 to day 90. The study was approved by the IACUC. Differences in body weights and serum levels of estradiol and IGF-1 were detected using a One-way ANOVA (p < 0.05). Body weights were similar on day 65 prior to the injection protocol however on day 90, Ex group weights were significantly greater than C. Estradiol levels were suppressed by 36% in Ex compared to C and 26% compared to BL65. IGF-1 levels were 19% higher compared to C and similar to BL65 levels, IGF-1 values typically decrease as animals mature. The decrease in estradiol levels following GnRH suppression resulted from decreased pulsatile secretion of GnRH from the hypothalamus which then resulted in decreased LH and FSH secretion from the pituitary. The increase in IGF-1 levels is not similar to the clinical condition of secondary amenorrhea which results in suppressed IGF-1 levels. These data suggest that secondary amenorrhea is not only a suppression of the reproductive axis (Hypothalamic-Pituitary-Gonadal) but also results from a suppressed somatic axis (GH-IGF-1). In conclusion, modeling secondary amenorrhea using a GnRH-a may model a subset of the clinical condition. 

Table  .  

Disclosures: R. Joshi, None.

This study received funding from: NIH Grant AG19654, PSC-CUNY Grant 64293-00-33.

W511

Is Development of Ovulatory Cycles in Adolescence Important for Peak Bone Mass?, S. Kalyan*, S. I. Barr*, R. Alamoudi*, J. C. Prior. Medicine, University of British Columbia, Vancouver, BC, Canada.

The purpose of this study was to investigate the potential importance of establishing ovulatory cycles following menarche in bone mineral accretion in peripubescent girls. Healthy premenarcheal Caucasian girls were recruited to this 3-year prospective study of bone development (SI Barr, JBMR, 2001). The mean enrollment age of the 38 girls who completed the study was 10.6 ± 0.6 years. Body mass index (BMI, kg²/m) and bone mineral density (BMD) by DXA of the total body (TB) and lumbar spine (LS) were measured every 12 months for 36 months. Participants reported when and if menarche occurred. 33 experienced menarche and 5 did not. Progesterone was measured in saliva 3-weeks after flow and weekly thereafter until the next flow by 13 of the menstruating girls; levels > 40 nmol indicated ovulation. Nine of the 13 girls had at least one ovulatory cycle. The earliest following menarche that ovulation occurred was 10 ± 5 months. The 4 girls who did not ovulate were only sampled in cycles occurring < 8 months post-menarche. BMI at baseline (18.31 ± 2.6) or 36 months (21.21 ± 3.5) did not relate to change in BMD (r = 0.017 and 0.163, respectively). However, there was a significant positive correlation between the change in both LS and TB BMD and time since menarche (r² = 0.14, p<0.05; r² = 0.40, p<0.0001, respectively). Regression analysis (Figure 1) with the 95% confidence interval suggests that greater gains in BMD are noted at the time following menarche when cycles are more likely to became ovulatory. Although the effect of menarche on bone accrual is already appreciated, the influence of ovulatory cycles in peripubertal girls as a determinant of bone gain has not been investigated. Results of this study provide preliminary evidence to suggest that attaining ovulatory cycles contributes to optimal peak bone mass. This hypothesis is line with previous work showing that ovulatory disturbances accounted for 24% of the variance seen in changes of cancellous-bone density in regularly cycling premenopausal women (JC Prior, NEJM, 1990). 

Figure Figure 1. Regression analysis of the change in BMD over 3 years in relation to time since menarche.

Disclosures: S. Kalyan, None.

This study received funding from: British Columbia Health Care Research Foundation and the British Columbia Children's Hospital Foundation.

W512

5-Year Follow-up of Bone Mineral Distribution and Bending Strength Changes Across Puberty: Comparison Between Biological Age-Aligned Boys and Girls. S. Kontulainen¹, H. Macdonald*², J. Johnston*², H. McKay². ¹University of Saskatchewan, Saskatoon, SK, Canada. ²University of British Columbia, Vancouver, BC, Canada.

The greater incidence of fragility fractures in older women compared to men may be related to sex-specific patterns of bone strength development during growth [1]. Thus, the aim of this 5-year follow-up was to assess changes in bone bending strength to the smallest (Imin) and greatest (Imax) direction of rigidity, and polar distribution of bone mineral content (PDBMC) at the tibial diaphysis in boys and girls at the same maturational stage. Biological age was defined as years from age at peak height velocity for 98 participants (52 girls) in the Healthy Bones II Study. A single slice at the 50% site of the tibial diaphysis was acquired using the Norland/Stratec XCT 2000. Bonalyse 2.1 was used to assess cortical bone Imax and Imin, the polar distribution of bone mineral content (PDBMC) and muscle cross-sectional area (MCSA). K-mode with an outer threshold of 200 mg/cm³ was used to define the periosteal bone surface and an inner threshold of 480 mg/cm³ was used to separate cortical bone from trabecular bone. PDBMC defines the angular distribution of bone around the centre of mass. The distance from the centre represents the mass (in grams) in a 1 cm thick section. Data were analyzed using a hierarchical (random effects) modelling approach. During and after rapid skeletal growth boys distributed their bone mass further from the neutral axis, especially towards the anterior and posterior directions of the tibial diaphysis, when compared with girls at the same biological age (Fig I). This resulted in a greater magnitude of the sex-difference in the direction of greatest bending rigidity (Imax). When biological age, tibial length and MCSA were controlled for, boys had a statistically significant greater increase in bending strength in both directions. These results are in accordance with evidence from anthropological samples suggesting that lower limb bones of adult males are more adapted for anterior-posterior (A-P) bending and common loading direction compared with females [2]. However, this phenomenon may be site-specific and further research at other bone sites is warranted.

References:

1. Seeman E, Delmas PD. N Engl J Med. 2006;354:2250-612.

2. Ruff CB, Hayes WC. Am J Phys Anthropol. 1983;60:383–400. 

Figure Figure 1.. Change in PD_BMC in both sexes aligned with biological age

Disclosures: S. Kontulainen, None.

W513

Age-Related Distribution of Bone and Skeletal Parameters in 1,322 Young Women. T. Kuroda*¹, Y. Onoe¹, S. Orito*¹, M. Ohara*¹, M. Sakai*¹, K. Ishitani*¹, H. Okano¹, A. Uyama*², M. Kume*², H. Ohta¹. ¹Dept. of Obstetrics and Gynecology, Tokyo Women's Medical University, Tokyo, Japan. ²School of Nursing, Tokyo Women's Medical University, Tokyo, Japan.

Objective: The age-related distribution of bone and skeletal parameters was examined in young female subjects to establish methodologies for prophylaxis of osteoporosis through acquisition of high bone mineral density (BMD) in younger years as well as to identify interventional measures aimed at acquisition of high BMD in these women.

Subjects and Methods: All students attending School of Nursing at Tokyo Women's Medical University, and Toho Girls Junior and Senior High Schools during August 2004 and August 2006 were enrolled in the study with the exclusion of those who were receiving treatments that could affect bone. The subjects were inquired about their date of birth, birth weight, weeks at birth, and age at menarche using a questionnaire, and their body height and weight were also measured. As relevant bone-related parameters, BMD, bone mineral content (BMC), and bone area (BA) in the lumbar spine (L2-4) as well as in the total proximal femur (hip) were measured in these subjects using QDR-4500; (Hologic Inc., USA). In addition, blood samples were drawn from the subjects to measure their calcium (Ca), phosphorus (P), bone-specific alkaline phosphatase (BAP), and urinary N telopeptide of type I collagen (NTX) levels.

Results: 1,322 students comprised our study subjects who ranged in age from 12 to 30 years of age. Their mean birth weight was 3,101 ± 431.3 g, and their mean age at menarche was 12.0 ± 1.2 years old. Their body height peaked at the age of 14 years, and their body weight peaked at the age of 21 years of age. Their BMD peaked at the age of 18 years of age but their L2-4 BMD gradually increased thereafter. In contrast, their total hip BMD peaked at the age of 18 years but decreased thereafter. Their L2-4 BA showed a rapid increase until the age of 14 years, while their total hip BA continued to increase gradually beyond the age of 14 years. Their L2-4 BMC exhibited a rapid increase until the age of 18 years and continued to increase gradually with increasing weight, while their total hip BMC remained constant between the ages of 18 and 23 years old and then decreased thereafter. Additionally, their BAP and NTX values were shown to decrease by about 80% between the ages of 12 and 18 years. Conclusions: Of all constituents of BMD examined, BMC was associated with increases in body weight and changes in bone metabolic markers. In contrast, BA was associated with increases in body height until the age of 14 years, although it appeared to be influenced by body weight beyond that age. Study results pointed to a need for lifestyle intervention by the age of 18 and increasing body weight beyond that age to ensure acquisition of high BMD in young women.

Disclosures: T. Kuroda, None.

W514

Correcting DXA Pediatric Bone Mineral Density Measurements to Account for Fat Inhomogeneity. D. C. Lee, T. A. L. Wren, V. Gilsanz. Departments of Orthopaedic Surgery & Radiology, Childrens Hospital Los Angeles, Los Angeles, CA, USA.

Dual energy x-ray absorptiometry (DXA) assumes that the soft-tissue anterior/posterior (AP) to the vertebrae (bone region) has the same attenuation properties as soft tissue adjacent/lateral to the bone (soft tissue region). DXA areal bone mineral density (aBMD) measurements are therefore subject to error when there is an inhomogeneous distribution of soft-tissue. The purpose of this study was to use computed tomography (CT) to calculate the error in vertebral aBMD due to inhomogeneous fat distribution and to develop an equation to correct for this error using easily obtained clinical parameters.

CT (General Electric CT-T 9800) scans were performed on the lumbar vertebrae of 700 children (350 boys, 350 girls), age 5-24 yr. Based on CT axial slices through L3, fat percent was determined in the bone and soft tissue regions. The error in vertebral aBMD was then calculated and correlated with anthropometric parameters including age, height, weight, body mass index (BMI), and trunk width (laterally), thickness (AP), and circumference. A correction equation was generated using stepwise regression. This equation was tested on a separate validation group of 101 subjects (46 boys, 55 girls) to determine the extent to which the soft tissue error was reduced.

Fat percentage was higher in the bone region than in the soft tissue region in 599 out of 700 subjects (85.5%), with an average difference of 6.0±5.7% (p<0.001). This inhomogeneity resulted in an average error of −7.2±7.4% in aBMD measurements (range: −21.6% to 5.5%). Stepwise regression analysis indicated that weight and trunk width had the strongest influence on the measured error (p<0.001). The correction equation derived from the stepwise regression was (Corrected aBMD [g/cm²]) = aBMD [g/cm²] −0.1144 + 0.0011 *Weight [kg] + 0.0004*(Body Width [mm]). When this equation was applied to the validation group, the range of error decreased by 25.7% and the average was reduced from −7.7±8.3% to −0.9±6.3% (Figure).

Our findings show that soft-tissue AP to the vertebrae contains more fat than soft-tissue lateral to the vertebrae, resulting in erroneously low aBMD values. Using a correction equation based on weight and trunk width, this error can be reduced significantly. Further correction may be obtained by developing equations tailored for specific subgroups such as by age, gender, sexual or skeletal maturity, or measures of body composition.

Disclosures: D.C. Lee, None.

This study received funding from: NIH & Army.

W515

Body Composition — Bone Relationship in Females Is Hormonal Stage Dependent: A Longitudinal Study. A. E. Lyytikäinen¹, F. Tylavsky², Q. Wang¹, H. Suominen¹, M. Alén*¹, V. Ristimaa*¹, E. Völgyi¹, S. M. Cheng¹, P. Rahkila*¹, U. M. Kujala*¹, E. Seeman³, S. Cheng¹. ¹Department of Health Sciences, University of Jyväskylä, Jyväskylä, Finland. ²Department of Preventive Medicine, University of Tennessee, Memphis, TN, USA. ³Department of Medicine, University of Melbourne, Melbourne, Australia.

The relationships of bone mass to lean and fat mass are controversial. We hypothesized that 1) the associations of bone mass (BM) with lean tissue mass (LM) and fat mass (FM) will change during puberty and 2) the loss of BM in premenopausal and postmenopausal females is companied by the loss of LM. We followed the growth of 10-13-year (y) old girls (n=258) 84 months (mo). In addition 80 pre-menopause (PreM, mean age 43y) and 28 peri-menopause (PeriM, mean age 49y) mothers, and 72 post-menopause grandmothers (PostGM, mean age 67y) of these girls were measured twice at a 42 mo interval. Whole body BM, LM, and FM were assessed by DXA (Prodigy, GE Lunar). Hierarchical linear model was used to evaluate the changes in BM, LM, and FM relative to menarche for the girls and annual change in the mothers and grandmothers. The mean menarche age for the girls was 13.0±0.9 y. The girls' BM accrual was accompanied by LM accrual until 3y after menarche. Thereafter, growth of LM ceased, but FM continued to increase (0.96 kg/y) and was the main contributor of body weight change (1.12 kg/y) near peak HT [164.5 cm (48mo), 164.8 cm (84mo)]. The annual changes of BM were associated with those of LM (r²=0.327 for 0-24mo, r²=0,277 for 24-36mo and r²0,404 for 36-84mo) and FM (r²=0.165 for 0-24mo; r²=0.185 for 24-36mo; and r²=0.313 for 36-84mo). In PreM, no changes in BM and FM were observed, but LM decreased 0.4 kg, 0.94%/y. LM decreased both in PeriM (0.53kg, 1.3%/y) and PostGM (0.63kg, 1.5%/y. This decrease was the factor that accounted for the loss of BM (PeriM=33g, 1.3%/y, and PostGM=20g, 0.9%/y). In summary, the changes in BM and LM are synchronized during rapid growth period in early puberty. In late puberty LM ceases to increase, and the fat accumulation is associated with bone accrual. The small increase in LM has higher impact than that of fat during postpuberty to early adulthood. In premenopausal women, although LM decreases, its effect on BM is minor. However, in both peri- and postmenopausal females, the loss of LM is of great importance in the loss of BM. In conclusion, the influence of body composition on bone mass gain or loss is stage-dependent from prepuberty to late postmenopausal years.

Disclosures: A.E. Lyytikäinen, None.

This study received funding from: Finnish Ministry of Education, Academy of Finland, Juho Vainio Foundation Finland, ASBMR Bridge Fuding.

W516

Tibial Bone Mineral Density and Geometric Properties in Females of Three Generations. H. O. Ma, E. Völgyi, H. Suominen, O. Wang, P. Rahkila*, S. M. Cheng, A. Lyytikäinen, M. Alén*, S. Cheng. Department of Health Sciences, University of Jyväskylä, Jyväskylä, Finland.

The evolution of bone mineral density (BMD) and geometric properties throughout life is not well understood. In this study we investigated the volumetric BMD and geometric properties of the tibial shaft in female of three generations. Altogether 82 girls (G, mean age 17.9±1.0 yrs), 76 mothers (M, 47±4.8 yrs) and 47 grandmothers (GM, 71±5.0 yrs) were measured using peripheral quantitative computed tomography. Total tibial cross-section area (tCSA), total bone mineral density (tBMD), cortical cross-section area (cCSA), cortical bone mineral density (cBMD), cortical thickness (cTh), bone marrow cross-section area (bmCSA), and muscle cross-section area (mCSA) were determined. Analysis of variance was used to compare the differences between generation groups. The girls had smaller tCSA than mothers and grandmothers showing the unaccomplished bone periosteal expansion. However, the tBMD of the GM group was 10% lower than either in the G or M groups due to the age related bone loss. The cCSA and cBMD were highest in the M group. The G group also had significantly higher cBMD than the GM group. The GM group had the thinnest cTh and largest bmCSA. Our results showed that the periosteal apposition may continue throughout life and endocortical resorption occurs in adult, but to a similar degree to the periosteal appostion in young adults keeping their cortical thickness unchanged. In older female, the periosteal apposition is less than the endocortical resorption thus cortex become thinner and bones become fragile.

Disclosures: H.Q. Ma, None.

This study received funding from: ASBMR Bridge Funding, Academy of Finland and Finnish Ministry of Educztion.

W517

Effect of the Dietary Habit of Skipping Breakfast on Bone Metabolism, Skeleton and Nutritional Status in Young Japanese Women. Y. Onoe¹, T. Kuroda*¹, Y. Miyabara*¹, M. Sakai*¹, R. Yoshikata*¹, M. Ohara*¹, S. Orito*¹, H. Okano¹, M. Kume*², S. Sasaki*³, H. Ohta¹. ¹Dpt. of Obstetrics and Gynecology, Tokyo Women's Medical University, Tokyo, Japan. ²Faculty of Nursing, Tokyo Women's Medical University, Tokyo, Japan. ³School of Public Health, the University of Tokyo, Tokyo, Japan.

The dietary habit of skipping meals not only affects systemic nutritional status but also influences intake of nutrients that are related to bone metabolism. However, it is unclear what effect such dietary habit may have on bone metabolism and skeleton. In our previous study, we have reported that ensuring appropriate physical activity, BMI, and serum 25(OH) D levels are crucial to acquisition of high bone volume during younger years. In this study, we examined the effect of skipping breakfast on nutritional status, intake of nutrients related to bone, and bone metabolism. Two hundred seventy-five healthy female volunteers aged 19 to 25 years old were enrolled in our study. All subjects were interviewed about their background information, and their height and body weight were measured. Their sera were tested for bone metabolic markers, such as BAP and NTX. Bone mineral density (BMD) was measured in these subjects using DXA (QDR 4500). The subjects were also asked about their current physical activity using a questionnaire as well as about their current diet using the self-administered Diet History Questionnaire (DHQ). The number of meals each subject skipped for breakfast, lunch and dinner during 1 week was obtained from the DHQ. Eighteen subjects (6.5%) had no breakfast during the week, while 131 subjects (45%) had breakfast every day of the week. The subjects were divided into two groups: those who had breakfast every day (group I) and those who skipped breakfast (group II). Significant differences were noted between groups in total energy intake and intake of three major nutrients (carbohydrate, protein and fat), with total energy intake and intake of the major nutrients becoming significantly lower with the increasing frequency of breakfast skipped. Oral intake of bone metabolism-related nutrients (Vit D, Vit K, Ca, P, Mg and K) was highest in group I with intake of each nutrient becoming significantly lower with the increasing number of meals skipped. Group I showed significantly higher BMD than group II in both the hip and L2-4. However, the habit of skipping breakfast had no significant effect on BMI. The habit of skipping meals had no significant effect on BAP, while it was significantly negatively correlated with NTX. Our data demonstrates that the habit of skipping breakfast leads to a decrease in systemic nutritional status, intake of nutrients, and bone metabolic and skeletal indices, thus suggesting that the habit of skipping breakfast hinders acquisition of high bone volume.

Disclosures: Y. Onoe, None.

W518

Correlation of Isokinetic Knee and Trunk Muscular Strength with Bone Mineral Content in Female Soccer Players. M. F. Saccol*¹, N. C. Oliveira*¹, A. S. Pinto*¹, R. M. Pereira¹, L. Takayama*¹, J. D. Greve*², F. R. Lima*¹. ¹Rheumatology Division (Bone Metabolism Laboratory), University of São Paulo, São Paulo, Brazil. ²Orthopedic Institute, University of São Paulo, São Paulo, Brazil.

The purpose of this study was to asses the correlation between knee and trunk muscle strength and bone mineral values in female junior soccer players. Twenty-two consecutive females soccer players with at least 15 h.wk⁻¹ of training during the previous year were compared with 20 females not engaged in regular exercise. Bone mineral density (BMD) and bone mineral content (BMC) were measured in lumbar spine (L1-L4), femoral neck and total femur of dominant hip, dominant leg and whole body by dual-energy X-ray absorptiometry (DXA). Fat mass and lean mass were also determined using whole body scan by DXA. Isokinetic muscle measurements were performed using the Biodex System 3 dynamometer with knee or trunk attachment on. Knee and trunk flexors and extensors strength were evaluated with 5 repetitions (concentric and eccentric modes) for the following variables: peak torque (PT), total work (TW) and set total work (STW). Statistical analysis was performed using Student t-test and Pearson's correlation. The two groups were alike regarding age, height, weight and calcium intake (p>0.05). Female soccer players had higher percentage of lean body mass (72.45±3.52 vs. 66.75±5.45g, p<0.001) and lower percentage of body fat (23.7±3.67 vs. 29.94±5.75%, p<0.001) compared to controls. For bone mineral values, the athletes demonstrated significant higher BMD and BMC compared to control group, in all sites evaluated (p<0.05).

Dominant knee strength were also significantly higher for soccer group (concentric extension PT 150.46±31.08 vs. 129.47±21.72Nm, p=0.016; eccentric extension PT 127.21±27.15 vs. 104.85±25.46Nm, p=0.009). Related to trunk strength, only concentric mode had higher performance in the soccer group (concentric extension PT 214.23±52.24 vs. 146.12±49.72Nm, p<0.001). All knee strength extension variables demonstrated positive correlation to BMC of leg, femoral neck and total femur of dominant hip (0.46<r<0.70). This correlation was also found for flexion knee strength, but only with PT variable (0.42< r< 0.64). On trunk evaluation, only flexors strength performance were correlated with BMC of total trunk at variables TW (r=0.525) e STW (r=0.513). These findings of positive correlation between muscle strength and bone mineral values in a specific site reinforces that there is a direct role of muscle function on the accretion of the regional bone mineral mass.

Disclosures: R.M. Pereira, None.

W519

Weight Bearing Bones Are More Sensitive to Physical Exercise in Boys than in Girls During Pre- and Early Puberty. S. Kriemler*¹, L. Zahner*¹, J. Puder*², C. Braun-Fahrlaender*³, C. Schindler*³, M. Kraenzlin⁴, R. E. Rizzoli⁵. ¹Institute Sport & Health, Basel, Switzerland. ²Division Endocrinology, Diabetes & Clinical Nutrition, Basel, Switzerland. ³Institute Social & Preventive Medicine, Basel, Switzerland. ⁴Endocrinology, Diabetes and Clinical Nutrition, Basel, Switzerland. ⁵Rehabilitation & Geriatrics, University Hospital, Geneva, Switzerland.

Background: Physical activity (PA) positively influences bone mineral accrual. However, little is known whether there are gender differences in bone sensitivity to loading and to what extent spontaneous objectively recorded PA influences bone mineral mass independently from muscle mass. We investigated gender differences in the association between bone mineral density/content (BMD/BMC) and measures of PA, during pre- and early puberty.

Methods: We measured BMD/BMC and fat-free mass in 374 6-13-year-old children from randomly selected schools at the hip, lumbar spine and total body by DXA. PA was evaluated by accelerometers, and lower extremity strength by a jump & reach test.

Results: Boys had higher hip and total body BMD than girls at all ages. Boys had higher fat-free mass, greater lower extremity muscle strength and were more physically active than girls at all ages. Total hip BMD was positively associated with time spent in total and vigorous PA in boys (r=0.33 and 0.27, respectively, p<0.01), but not in girls (r=0.02 and 0.04, p=ns), even after adjusting for fat-free mass and lower extremity strength. While boys and girls in the lowest tertile of vigorous PA (22 min per day) had similar hip BMD (0.668 vs 0.679 g*cm-2), those boys in the highest tertile (72 min per day) had significantly higher values than the corresponding girls (0.730 vs. 0.692 g*cm-2, p<0.05). In multiple logistic regression analyses, a low hip BMD in boys was best predicted by fat-free mass (OR 0.55 [95%CI 0.38, 0.78] per 1 kg) and total PA (OR 0.69 [95%CI 0.49, 0.98] per 106 counts), while in girls predictors included fat-free mass (OR 0.63 [95%CI 0.50, 0.79] per 1 kg) and jump & reach (OR 0.90 [95%CI 0.84, 0.98] per 1 cm). Similar data were obtained for femoral neck BMD/BMC. There was no influence of PA on lumbar spine BMD in either gender.

Conclusion: Though at low physical activity during pre- and early puberty, hip BMD is similar in both genders, it increases proportionally to PA, and differences in bone mineral mass between genders appears, likely related to a different sensitivity of bone to physical loading, independently from muscle mass.

Disclosures: R.E. Rizzoli, None.

W520

Accrual of Lean Mass Has More Impact Than the Accrual of Fat Mass on BMC During Growth. F. A. Tylavsky¹, G. Somes*¹, M. Alen², A. Lyytikäinen³, V. Ristimaa*³, S. M. Cheng³, H. Suominen³, U. M. Kujala*³, S. Cheng³. ¹Preventive Medicine, University of Tennessee, Memphis, TN, USA. ²Health Sciences, University of Jyvaskyla, Jyvaskyla, Finland. ³University of Jyvaskyla, Jyvaskyla, Finland.

Many reports have focused on the role lean (LM) and fat mass (FM) plays in the accrual of bone mass. The objective of this study was to examine the relationship between FM and LM and bone mineral content (BMC), bone mineral density (BMD) as girls transition through puberty.

The study subjects consisted of 258 10-13-year old girls at the baseline. They were followed on average of 6.5 years and their results reflect the entire follow-up period (n=217). Whole body BMC, LM and FM were assessed by DXA (Prodigy, GE Lunar). Height, weight, Tanner stage and menarche status was obtained at baseline, 6, 12, 24, 36, and 84 months. 218 participants provided 1160 observations with a minimum of 2 and a maximum of 8 observations. 206 participants provided observations before menarche with a range of 1 to 6 observations from 51.6 months before menarche. 193 subjects provided observations after menarche with a range of 1 to 7 observations to 95.6 months after menarche. 

Table  .  

Using random coefficients modeling in the MIXED procedure of SAS we found that subjects gained about 0.49 kg/month before menarche (BM) and 0.17 kg/month after menarche.

The weight gain was respectively portioned into 124 g/mo of fat and 355 g/mo of lean mass before menarche; and 108 g/mo of fat mass and 59 g/mo of lean mass after menarche. Regardless of menarche status, the effect of gains in lean and fat on BMC is that for every kg of fat mass there is a 16 g gain of BMC during puberty; for every kg of lean mass there is a gain of 47 grams of BMC; indicating the important effect of lean mass on the bone mass. We also found that the relative position within a cohort, LM, and BMC is already established by early puberty with fat mass being subject to environmental and pubertal changes. These findings provide evidence and implications for prevention of fat mass gain in those children who are at risk of overweight or obesity.

Disclosures: F.A. Tylavsky, None.

This study received funding from: ASBMR Bridge Funding. Academy of Finland and Finnish Ministry of Educztion.

W521

Determinants of Change in Bone Volumetric Density and Geometry Differ Between Boys and Girls in a Multiethnic Population. R. J. Wetzsteon, B. C. Kaufman*, J. M. Hughes*, S. Stovitz*, M. A. Petit. Kinesiology, University of Minnesota, Minneapolis, MN, USA.

The purpose of this study was to explore determinants of change in bone volumetric density (vBMD, mg/mm3), bone area and estimated bone strength in a multi-ethnic population of boys and girls. A total of 116 (60 boys, 56 girls) children aged 10.9 ± 0.8 years at baseline participated. Ethnicity was based on CDC categories (17% Caucasian, 27% African American, 24% Hispanic, 10% Asian, 22% Other). Measurements of height, weight, and tibia length were taken by standard techniques; daily calcium intake and physical activity (PA score) were assessed by questionnaire; and dynamic power estimated by a vertical jump test. Peripheral quantitative computed tomography (pQCT) was used to assess the 50% site of the tibia for cortical volumetric density (CoD), area (CoA), polar strength-strain index (SSIp), muscle cross-sectional area (MCSA), and total fat area (FatA). All measurements were taken at baseline and after 5-months. At baseline, boys and girls were similar in age, weight, calcium intake, PAscore, and vertical jump (p < 0.10 for all). Girls were significantly taller (p = 0.04). Baseline CoA and SSIp did not differ between the boys and girls, but girls had greater CoD (+ 2.2%, p = .001) which remained significant after adjusting for height. All bone values increased significantly over 5 months, but the magnitude of the change did not differ between boys and girls. Stepwise regression analyses were run to determine predictors of change with change in tibia length entered in the first step, and other predictors (change in MCSA, change in FatA, age, ethnicity, PAscore, average daily calcium, and change in vertical jump) entered in a stepwise manner if p < 0.05. Separate models were run for boys and girls. For both boys and girls, change in tibia length explained ∼20% of the variance for change in SSIp. For girls, change in MCSA explained an additional 31%, with daily calcium and change in vertical jump explaining and additional 10% of the variance. For boys, only change in MCSA entered the model explaining an additional 23% of the variance for change in SSIp. These data are consistent with recently published data showing higher bone vBMD in girls. They also suggest that MCSA may be a reasonable surrogate of muscle force in boys, but for girls other factors (muscle power and calcium) may be important predictors of change in bone strength.

Disclosures: R.J. Wetzsteon, None.

VITAMIN D WORKING GROUP

WG1

Deletion of the 25 Hydroxy Vitamin D 1α Hydroxylase from Mice Expressing the Null Mutation for the Calcium Sensing Receptor Converts a Hypercalcemic to a Hypocalcemic Hyperparathyroid Phenotype. C. Richard*¹, D. Miao², R. Samadfam*¹, Y. Wang*¹, G. N. Hendy¹, D. Goltzman¹. ¹Dept of Medicine, Calcium Laboratory, McGill University, Royal Victoria Hospital, Montreal, PQ, Canada. ²Department of Human Anatomy, Nanjing Medical University, Nanjing, China, China.

The calcium sensing receptor (CaSR) is functional mainly in the parathyroid gland and kidney, but likely as well in other tissues such as the gastrointestinal tract and the skeleton. These tissues are also target sites for the action of 1,25- dihydroxyvitamin D [1,25(OH)₂D]. Mice which are heterozygous or homozygous for the calcium sensing receptor null mutation (CaSR^+/− and CaSR^−/− respectively) develop hypercalcemia and hyperparathyroidism to a moderate or severe degree, respectively. Mice which are homozygous for targeted deletion of the 25 hydroxyvitamin-D 1α-hydroxylase [1α(OH)ase^−/−] which synthesizes 1,25(OH)₂D develop hypocalcemia, hyperparathyroidism and rickets but survive long term. To assess the interactions between the important calcium regulating molecules, CaSR and 1,25(OH)₂D, we created compound mutants of the CaSR and 1α (OH)ase mutants and assessed their growth, biochemistry and skeletal morphology. CaSR +/-;1α (OH)ase+/+ mice grew normally and exhibited normal longevity. CaSR +/+; 1α (OH)ase-/- mice developed severe rickets and osteomalacia as assessed by faxitron x-ray analysis and von Kossa staining of the skeleton but survived at least 2 months. CaSR +/-; 1α (OH)ase-/- mice developed even more severe rickets, osteomalacia and growth retardation but also survived long term. BMD, assessed by piximus, was reduced by 68% in the lumbar spine and by 49% in the femur compared to wild type mice, but only by 58% and 38% in the lumbar spine and femur respectively of the CaSR +/+; 1α (OH)ase-/- mice. Although CaSR-/-; 1α (OH)ase+/+ mice develop severe hypercalcemia and most die before or shortly after weaning, deletion of the 1α (OH)ase in CaSR-/-; 1α (OH)ase-/- mice facilitated survival for at least 2 months although these animals exhibited the most severe growth retardation, rickets and osteomalacia of all mutants. All mutant animals showed evidence of hyperparathyroidism with CaSR+/-; 1α (OH)ase+/+ and CaSR-/-; 1α (OH)ase+/+ mice displaying hypercalcemia. In contrast, CaSR+/-; 1α (OH)ase-/- mice and CaSR-/-; 1α (OH)ase-/- mice were severely hypocalcemic (serum calcium 1.28 and 1.27mM respectively). The results indicate that the phenotypic manifestations of hyperparathyroidism may be markedly influenced by the vitamin D status of animals with CaSR mutations, confirm that elimination of hypercalcemia sustains longevity in mice homozygous for the CaSR null mutation, and suggest that mutations in the CaSR contribute to worsening of skeletal mineralization.

Disclosures: C. Richard. None.

WG2

Targeted Inactivation of the Vitamin D Receptor by Caspase-3. P. J. Malloy*, D. Feldman. Department of Medicine, Stanford University, Stanford, CA, USA.

Calcitriol inhibits the growth of many cells including breast, colon, ovarian, pancreatic, and prostate cancer cells by causing cell cycle arrest and in some cells by inducing apoptosis. In breast cancer cells, calcitriol induces apoptosis by a caspase independent mechanism. On the other hand, in prostate cancer cells, calcitriol induction of apoptosis involves activation of caspase activity. Calcitriol actions are mediated by the vitamin D receptor (VDR) a member of the steroid-thyroid-retinoid receptor superfamily of nuclear transcription factors. In LNCaP prostate cancer cells induction of apoptosis by staurosporine abolished [³H]1,25(OH)₂D₃-binding and VDR protein suggesting to us that the VDR might be targeted for inactivation by caspases, a family of cysteine proteases that are activated during apoptosis. We identified a potential caspase-3 cleavage site (DxxDS) in helix H2n (D₁₉₅MMDS₁₉₉) in the VDR ligand-binding domain. Mutations (D195A, D198A and S199A) were generated in the putative caspase-3 cleavage site and their affects on caspase cleavage of VDR examined. In reporter gene assays, all of the VDR mutants exhibited transactivation activity similar to the WT VDR. COS-7 cells transfected with WT and mutant VDR cDNA expression vectors were then treated with staurosporine and VDR examined by western blot. A 22 kDa cleavage fragment was detected on western blot from cleavage of the WT VDR and the S199A mutant VDR but not the D195A or D198A mutant VDRs. Addition of the caspase-3 specific inhibitor z-DEVD-FMK prevented cleavage of the WT VDR in vivo. Treatment of transfected COS-7 cells with calcitriol also resulted in cleavage of the WT VDR but not the D198A mutant VDR. Calcitriol-induced cleavage of the WT VDR was blocked with z-DEVD-FMK demonstrating that calcitriol activates caspase-3 in these cells. In vitro treatment with caspase-3 resulted in the cleavage of WT VDR and S199A but not D195A or D198A mutant VDRs. WT VDR was also cleaved by caspase-6, and −7 but not caspase-8 in vitro. In conclusion, our results demonstrate that the VDR is cleaved by caspase-3 in vivo. We further demonstrate that calcitriol induces caspase-3 activity that results in the cleavage of the VDR. Our results suggest that activation of caspase-3 by calcitriol, while contributing to apoptosis may also may limit the apoptotic effects of calcitriol in cells that express caspase-3. Since caspases also regulate the activity of many proteins under non-apoptotic conditions, the inactivation of VDR by caspases may be of general importance as a mechanism to limit calcitriol activity.

Disclosures: P.J. Malloy. None.

WG3

The Vitamin D Receptor, Hedgehog Signaling and Epidermal Carcinogenesis. A. E. Teichert*¹, J. J. Welsh*², D. D. Bikle¹. ¹Endocrine Unit, UCSF-NCIRE, San Francisco, CA, USA. ²Department of Biological Sciences, University of Notre Dame, Notre Dame, IN, USA.

Over 1 million skin cancers occur annually in the United States, 80% of which are basal cell carcinomas (BCC) (16% squamous cell carcinomas, 4% melanomas), making it by far the most common cancer. Extensive epidemiologic data support the concept that the incidence of a number of epithelial malignancies (e.g. breast, prostate, colon) is reduced by increasing 25OHD levels. 1,25(OH)₂D₃, produced by the kidney from 25OHD or locally produced, reduces proliferation and enhances differentiation and thus has been investigated for a role in preventing or treating cancer. However, UVB exposure, which increases 1,25(OH)₂D₃ production in the keratinocyte, causes skin cancer. Conceivably, the 1,25(OH)₂D₃ produced by the skin provides at least partial protection from UVB induced skin cancer such that lack of 1,25(OH)₂D₃ or its receptor would make the skin even more sensitive to UVB induced malignancy.

We have chosen UVB epidermal carcinogenesis as a model to directly address the issue of the balance between the increased risk of UVB exposure as an inducer of carcinogenesis with the benefit of promoting vitamin D and thus 1,25(OH)₂D₃ production in the epidermis as a chemopreventive measure.

Mice deficient for the vitamin D receptor (VDR null mice), for which 1,25(OH)₂D₃ is a ligand, have a marked hyperproliferative response in the hair follicle and epidermis and decreased epidermal differentiation. These mice, when treated with 7,12 dimethylbenzanthracene (DMBA), unlike their wildtype littermates, develop skin tumors containing hair follicle elements and/or are of basal cell origin, tumors characteristic of overexpression of the hedgehog (HH) signaling pathway in keratinocytes. We found that the epidermis and hair follicles of the VDR null animals, as well as the DMBA induced tumors in VDR null mice, overexpress elements of the HH signaling pathway [Sonic Hedgehog (Shh), Patched 1 (Ptch 1), Smoothened (Smoh), Gli 1, and Gli 2]. Moreover, keratinocytes in which the VDR has been knocked down with siRNA directed against the VDR show increased expression of Shh.

Interestingly, mice overexpressing Gli1, Gli2, or Shh in their basal keratinocytes or grafted with human keratinocytes overexpressing Shh develop BCC like lesions. Furthermore, BCC show overexpression of Ptch1, Smoh, Gli1 and Gli2, raising the question of a link between the vitamin D system, HH signaling and skin cancer predisposition.

These results suggest then that increased expression of Shh in the keratinocytes of the VDR null animal activates the HH signaling pathway, predisposing the skin to the development of BCC.

Disclosures: A.E. Teichert, None.

This study received funding from NIH.

PEDIATRIC BONE AND MINERAL WORKING GROUP

WG4

Childhood Hyperparathyroidism with Bilateral Slipped Femoral Epiphysis, Severe Bone Disease and Vitamin D Deficiency. R.N. Mehrotra¹, J. Pingle², U K Nayak³. ¹Department of Endocrinology. ²Orthopedics. ³Head & Neck Surgery, Apollo Hospitals, Jubilee Hills, Hyderabad, India.

Primary Hyperparathyroidism (PHPT) in children is an uncommon disease. It constitutes less than 5% of all cases of PHPT. PHPT causing slipped epiphyses has been described, but is an uncommon manifestation. In contrast to asymptomatic disease seen in the west PHPT in Asians presents with more severe bone disease, large glands and are associated with low Vitamin D levels. This report is about a child with PHPT form India who presented with B/L slipped epiphyses and had associated severe bone disease.

12 year old girl had onset of pain in both the hip joints five years. She had associated symptoms of proximal weakness. As X- rays showed B/L slipped epiphyses she under went fixation by screws. Repeat × rays showed no healing. There has been gradually increasing valgus deformity at the knee. Her proximal weakness has increased. So much so that at presentation she was bed ridden because of severe deformity. During admission during transport she suffered a fracture of right fore arm. Initial calcium done previously was 8.2 mg/dl. At our hospital it was 11.9, with Phosphorus of 3 mg/dl. Alkaline phosphates were 4050 u/l. Intact PTH levels was 2867 picogm/ml and 25 Hydroxy Vitamin D was 6.6 nanogm/ml. The × rays showed subperiosteal re-absorption, Brown tumors, and severe deformities. U/S neck revealed parathyroid lesion of 2×3 cm. This was confirmed on Sestamibi scan. She underwent removal of parathyroid tumor. During surgery both neck sides were explored, but there was no evidence of second adenoma. Biopsy from second gland was normal. Immediate post op she had hypocalcaemia because of hungry bone disease. Histopathology showed parathyroid adenoma. She has been on Vitamin D and calcium supplements since then. At last follow up calcium was 9.0 mg/dl, PTH and alkaline phosphatase had declined. Repeat × rays showed healing of Slipped epiphysis. Due to persistent deformities she underwent corrective osteotomies, and now she is able to walk.

Vitamin D deficiency and poor calcium intake modifies the disease presentation of PHPT in Indians. Slipped epiphysis seems to be due to changes in the metaphyseal spongiosa which leads to un locking of interdigitations due to increased PTH levels.

WG5

Tissue-specific Knockout of CaRs in Parathyroid Cells: A Model of Primary Hyperparathyroidism. W. Chang, C. Tu, T-H. Chen, D. Bikle, D. Shoback. Endocrine Research Unit, University of California, Department of Veterans Affairs Medical Center, San Francisco, CA, USA.

Serum ionized [Ca²⁺] is tightly maintained by the steep inverse relationship between [Ca²⁺] and the level of parathyroid hormone (PTH) secretion. Increases in the extracellular [Ca²⁺] ([Ca²⁺]_c) activate signaling pathways that inhibit PTH secretion by ligating the extracellular Ca²⁺ receptor (CaR) expressed on parathyroid cells. Heterozygous and homozygous loss-of-function mutations in the CaR gene cause familial benign hypocalciuric hyperparathyroidism (FBHH) and neonatal severe hyperparathyroidism (NSHPT), respectively. Patients with these disorders demonstrate both hyperparathyroidism (HPT) and hypocalciuria depending on gene dosage effects. The deletion of both parathyroid and renal CaRs is thought to be necessary for the development of this phenotype. Mutations in the CaRs in FBHH and NSHPT cause reduced responsiveness and or expression of the receptor in both parathyroid (PT) and kidney cells. To determine the contribution of parathyroid vs renal CaRs to the metabolic disturbances in NSHPT and FBHH and to discern the role of CaRs in tissues beyond the parathyroid and kidney, we generated a floxed CaR mouse (CaR^flox/flox) by inserting loxP sites flanking exon 7 of the CaR, which encodes the transmembrane and intracellular loops of the CaR. We bred parathyroid (PT)-specific CaR knockout (^PTCaR^+/−) mice using this CaR^flox/flox mouse crossed to a transgenic PTH promoter Cre mouse line. The resulting heterozygous knockout (^PTCaR^+/−) mice were backcrossed to the CaR^flox/flox mice to generate heterozygous and homozygous (^PTCaR^−/−) knockout mice and their wild-type (WT) littermates for analyses. ^PTCaR^+/− and ^PTCaR^−/− mice were born alive in normal Mendelian ratios. Serum total [Ca²⁺] were significantly elevated in the ^PTCaR^+/− (12.75 ± 0.12 mg/dl) and ^PTCaR^−/− (15.60 ± 0.19 mg/dl) mice, compared to WT controls (11.58 ± 0.34 mg/dl), consistent with the phenotype of HPT and a gene-dosage effect. Urine [Ca²⁺] were also increased by ≈60 and 80% in ^PTCaR^+/− and ^PTCaR^−/− mice, respectively, vs controls, supporting a compensatory hypercalciuria in ^PTCaR^+/− and ^PTCaR^−/− mice. While the ^PTCaR^+/− mice developed normally, the body weight of the ^PTCaR^+/− mice was ≈25% of their WT and heterozygous littermates. The bones of ^PTCaR^−/− mice were significantly smaller and severely undermineralized as indicated by micro-computed tomography (μCT) scans. This work confirms that decrements in parathyroid cell CaR expression are sufficient to induce HPT, growth retardation, and skeletal demineralization after birth. This floxed CaR mouse model will provide the opportunity to understand the role of CaRs in other tissues unhampered by the global metabolic disturbances of the generalized CaR knockout.

WG6

Bone Mass in Adults and Children with Hypoparathyroidism. MT Collins^1.4, JE McKie^1.4, JC Reynolds², KK Winer³. ¹Skeletal Clinical Studies Unit, CSDB, NIDCR. ²Nuclear Medicine Department. ³NICHD, NIH, Bethesda, MD, USA. ⁴Authors Contributed Equally.

Hypoparathyroidism is a rare condition that is associated with high bone mass. The mechanism of the development of high bone mass is unknown. Since adults and children differ in the cellular processes by which they maintain their bone mass, adults by remodeling and children primarily by modeling, differences between adults and children may shed light on the mechanism by which high bone mass is achieved in hypoparathyroidism. Seventy-five subjects were studied. There were 30 children (18/12 M/F, ages 4–19, median 11, length of disease 3mo.-15y, median 6.5y), and 45 adults (13/32 M/F, ages 20–75, median 45, length of disease 3mo.-35y, median 10y). Bone mineral density (BMD) was measured by Hologic DXA and Z-Scores were calculated using the manufacturer's most recent reference database.

BMD at the PA spine (PASp) and femoral neck (FN) were increased in adults (mean Z-Score = 1.8±2.0 and 0.98±1.1, respectively), but normal in children (0.36±1.3 and 0 ±1.4, respectively, for PASp adult to child comparison p=0.0008, and FN p=0.0017). To assess whether or not differences in BMD between adults and children were the result of length of hypoparathyroidism, adults and children were grouped and matched by length of disease (≤5 y, and 6–15 y). When there was ≤5 y of hypoparathyroidism, there was no difference between adults and children in BMD Z-Score at either the PASp or the FN (PASp: 0.95±0.89 vs 0.62±1.3, p=0.41, FN: 0.56±0.87 vs 0.37±1.6, p=0.69). However, when adults and children with 6–15 y of hypoparathyroidism were compared, adults had increased mean BMD Z-Score at the PASp and FN (1.9±0.86 and 0.20±1.3, respectively, vs 1.0±.93 and −0.25±1.3, respectively, for PASp adult to child comparison p=0.002, and FN p=0.016). When subjects with hypoparathyroidism due to autoimmune polyglandular syndrome (who often have malabsorption, gonadal failure, and/or are on glucocorticoid treatment for adrenal insufficiency) were excluded from the analysis, there was a significant positive correlation between length of disease and BMD in adults (PASp: R=0.41, p=0.0066, FN: R=0.35, p=0.03), but not in children (PASp R=-0.24, p=0.33, FN R=-0.25, p=0.30). Hypoparathyroidism did not have an effect on growth, as height was normal in children (height Z-Score = −0.21 ± 1.2).

From these data, it appears that normal skeletal modeling is not dependent on PTH, and the elevated BMD seen in hypoparathyroidism occurs only in the adult (remodeling) skeleton. These observations create novel perspectives from which to assess the role of PTH in the formation of the skeleton and the accrual of bone mass.

WG7

Genetic Mosaicism for de novo Mutations of the Calcium Sensing Receptor (CASR) Gene: Implications for Recurrence Risks in Autosomal Dominant Hypoparathyroidism. DEC Cole, BYL Wong, L Canaff, GN Hendy. Depts. of Laboratory Medicine & Pathobiology, Medicine, and Paediatrics (Genetics), University of Toronto, Toronto ON and Depts. of Medicine, Physiology and Human Genetics, McGill University, Montreal QC Canada.

In sporadic cases of autosomal dominant hypoparathyroidism (ADH) and familial hypocalciuric hypercalcemia (FHH), de novo mutations of the calcium sensing receptor gene (CASR) are not uncommon Previously, we described somatic mosaicism in the healthy mother of two siblings with severe ADH due to an activating F788L mutation of CASR (JCEM 2003:88:3674). Here we report on the analysis of nine more parent/proband triads – five with de novo ADH and four with de novo FHH. Analysis of heterozygosity by denaturing high performance liquid chromatography (dHPLC) revealed that CASR exon 4 amplified from leukocyte DNA of the mother of one ADH proband (E228K mutation) had an abnormal melting profile suggestive of mosaicism. This was confirmed by allele-specific amplification for the 228K variant and inspection of the sequencing capillary electro-phoretograms as well as sequence analysis of multiple subclones of the exon 4 amplicon. The degree of heterozygosity was estimated at ∼10%, like the mother of the F788L proband, and is similar to that found in other de novo dominant disorders. In the eight remaining cases, dHPLC profiles were all normal, indicating that if somatic mosaicism was present, the proportion of the mutant allele in peripheral leukocytes was less than 1%. Our second mosaic family emphasizes that de novo mutations in offspring cannot be regarded as eliminating all risk for recurrence. Mosaicism should be factored into the a priori risk estimate and appropriate testing offered as part of the counseling process. Whether there is some significance to the fact that maternal activating mutations account for both instances of mosaicism remains to be seen. Similarly, the absence of any evidence for mosaicism in the other eight cases (ADH or FHH) may well be associated with reduced risk of recurrence, a supposition that can be addressed when a larger case cohort is evaluated.

WG8

Rett Syndrome: Bone Mass Determination in Girls and Mice. J. Shapiro, A. Boskey, G. Bibat, S. Doty, M. Blue, B. Metcalf, A. Khosravi, S. Naidu. Kennedy Krieger Institute, Baltimore, MD, Hospital for Special Surgery, NYC, NY, USA.

The purpose of this study was to evaluate osteoporosis in Rett (RTT) syndrome and the RTT murine model. RTT is an X-linked disorder caused by mutations in the Methyl-CpG-Binding Protein (MeCP2) located at Xq28. Some patients with severe seizures and RTT-like features have mutations in cyclin-dependent kinase-like 5 (CDKL5). Features of RTT include deceleration of head growth, early onset cognitive defects, and a movement disorder with stereotypic hand movements. Decreased bone mass on x-rays and an increase in fractures has been observed in RTT children unrelated to their ambulatory status.

Diagnosis was confirmed by mutational analysis in 25 RTT girls ages 1–14. Hologic DXA (n=19) scans were performed. Additionally, 6 MeCP2 deleted (heterozygote-/+) females and (knockout -/-) males were compared to 6 wks control C57BL/6 male and female mice by Faxitron and micro-computed analyses (GE-EVS), histology, and Fourier transform infrared imaging spectroscopy of mineral and matrix. For the girls, 8 of 17 subjects had lumbar spine Z-scores −2.0 or less: Z-scores were within reference range in 9 children. DXA values low for age occurred as early as 6 years. Four children were post pubertal. DXA hip scans could not be obtained because of anatomic variation and motion. Mean BMI ages 1–14 was 16.9 ± 21 . 15/24 used anticonvulsant drugs. Micro CT Analysis of wild type (wt) and mutant mice (females (het) and male ko) are shown in the table as means with n=2/group. 

Table  .  

BVF Fractional bone volume; * Areal moment of inertia: TbN: trabecular number TbS: trabecular separation

Based on the 2 samples/group analyzed to date (-/-) bone has lower BMD and BVF, smaller moments of inertia, lower TbN and greater trabecular separation. The male (-/-) has decreased BMD, BVF, lower TbN and greater TbS. The (+/-) bone did not have altered BMD, but had greater BVF, but lower TbN and increased TbSp. The increased Imin and Imax suggests bone are stronger in the (+/-) females. Histochemical stains indicate a more intense procollagen presence in the (+/+) female compared to the (+/-) female. The intensity of stain was similar in (+/+) vs (-/-) males (n=4).

In conclusion, the low bone mass observed in 50% of Rett children parallels the preliminary data that the MeCP2 murine models provide as models for RTT syndrome.

PHYSICAL ACTIVITY AND FALLS WORKING GROUP

WG9

Bone Adaptation to Impact Loading — Significance of Loading Intensity. A. Vainionpää*¹, R. Korpelainen*², J. Leppäluoto*³, T. Jämsä*¹. ¹Department of Medical Technology, University of Oulu, Oulu, Finland. ²Department of Sports Medicine, Oulu Deaconess Institute, Oulu, Finland. ³Department of Physiology, University of Oulu, Oulu, Finland.

It is important to find efficient strategies for prevention of osteoporosis and fragility fractures as the prevention with medication is impossible at the population level. Exercise is one of the major prevention strategies even though the optimal loading patterns of bone have not yet been clearly defined in terms of bone loading forces in clinical settings. Our aim was to study the effects of impact loading on bone and assess the intensity and amount of efficient impact loading.

We conducted a 12 month population-based exercise intervention in 120 premenopausal women aged 35 to 40 years. The women were randomized equally to exercise (EG) and control groups (CG). The exercise regimen consisted of supervised, progressive high-impact exercises three times per week and an additional home program. Bone mineral density (BMD) in the proximal femur was measured by dual x-ray absorptiometry and bone geometry in the mid-femur was quantified by computed tomography (QCT). Also bone turnover was examined by using biochemical markers (TRACP-5b, PINP and Uncoupling Index). Daily impact loading was continuously recorded with a novel accelerometer-based measurement device and analyzed as the daily number of impacts within five acceleration ranges between 0.3 g to 9.2 g (g=acceleration of gravity, 9.81 m/s2).

Regular impact exercise increased the femoral neck BMD by 1.5% (p=0.003) and led to geometric adaptations by increasing the mid-femur periosteal circumference by 0.2 % (p=0.03) when compared to controls. It also altered bone turnover balance in favour of bone formation. The women with the highest number of impacts at the intensity level above 4 g had the greatest increment in proximal femur BMD. The number of impacts needed exceeding this level was 60 impacts per day. Impacts exceeding 2.5 g were associated with increased periosteal circumference and enhanced bone turnover favouring bone formation.

According to our results, impact loading has significant dose- and intensity dependent effect on weight-bearing bones. The threshold for improving BMD in the hip is 60 impacts per day exceeding 4 g, while geometric adaptation and bone turnover activation can be achieved with lower intensity levels. Impacts required for osteogenic effects can be obtained during habitual exercise training. These thresholds for osteogenic exercise give tools to promote efficient exercise patterns to be included in everyday routines and personal activities, thus increasing the potential to affect in community level.

Disclosures: A. Vainionpää, None.

WG10

Timing of Peak Bone Mass and Bone Mineral Density Are Influenced by High-Level Physical Activity in Young Adults of Both Sexes. S. Breban*, C. Chappard, C. Jaffre*, C. Benhamou. Chr orleans, INSERM Unit U658, Orleans, France.

Optimization of Peak Bone Mass may help to prevent osteoporosis and sport is known to be beneficial for bone tissue by increasing Bone Mineral Density (BMD). No study has reported effects of physical activity on Peak bone Mass. Thus, the aim of this cross-sectional study was to analyse the influence of physical activity on timing and level of Peak Bone Mass.

70 girls and 90 boys aged 17–28 years participated in this study. The sample included 40 athlete-girls and 60 athlete-boys participating in weightlifting, ball collective sports or judo for more than 6 hours per week (9.6±3.5 hours per week for women and 10.4±3.7 for men). They were age and sex matched with control girls (n=30) and boys (n=30). Bone Mineral Content (BMC, g) and BMD (g/cm²) were measured by DXA (Delphi, Hologic ®, Waltham, MA), at lumbar spine (LS) and non dominant femur (total and femoral neck (FN)). We also evaluated whole body (WB) BMC and BMD, with derived analysis of trunk, lower and upper limbs. Height, weight and body composition were derived from these DXA measurements. The timing of Peak Bone Mass was analysed by the inflexions of the age/BMD curves at all bone sites. Our variables were best fitted by second order polynomial regression equations. Thus, the model was expressed as y=ax²⁺bx+c. The age of maximal bone mass was determined as the maximum of the curve, i.e. when x = -b/2a. Athletes had higher BMD measurements than controls in both sexes at all bone sites (p<0.05). Consequently all the Peak Bone Mass levels were significantly higher in athletes than controls. BMD peaks were achieved around 21.5 (FN), 22 years (LS), and 24-25 years (WB, upper and lower limbs) in athlete girls; BMD peaks were reached before 17 years for all bone sites except for LS (21.8 years) in control girls. In athlete boys, BMD peaks were reached before 17 years for upper and lower limbs, and ranged from 21.0 years (FN) to 25.5 and 26.0 years respectively for WB and LS. Finally, in control boys, BMD peaks were reached before 17 years at all bone sites. Timing of Peak Bone Mass was different according to bone sites, sex and athletic or non athletic status. Thus, those results confirm that weight-bearing physical activity lead to a better bone status compared to non athletes, in both sexes and even after puberty. Moreover, our data suggest that the Peak Bone Mass was not reached at all bone sites before 17 years: thus, it is yet possible to optimize BMD and Peak Bone Mass in young adults between 17 and 28 years. High level weight-bearing physical activity seemed to induce a delayed and higher Peak Bone Mass compared to controls.

Disclosures: S. Breban, None.

WG11

Exercise Prevents Falls and Maintains Bone Mineral Density in Elderly Postmenopausal Women. Preliminary data of the Erlangen Senior Fitness and Prevention (SEFIP) Study. W. Kemmler*, S. von Stengel*, W. A. Kalender*, K. Engelke. Institute of Medical Physics, University of Erlangen, Erlangen, Germany.

The aim of the study was to demonstrate that an intensive aerobic and strength training can maintain BMD at the spine and the hip and can prevent falls in elderly community-living women.

Via population registers we recruited 246 community-living women of 65 years and older. Exclusion criteria were medication or diseases related to bone metabolism, low aerobic capacity (<75 W on ergometry), and medication potentially increasing the likelihood of falls. There were no inclusion or exclusion conditions based on BMD or prevalent fractures. Subjects were randomly assigned to an exercise intervention group (EG: 2-3 sessions/week) or to a wellness control group (WCG: 1 session/week for three 10 week blocks). The exercise training consisted of 20 min of intense aerobic dancing (70–85% HF_max), 5 min of specific balance exercises and 35 min of isometric and dynamic (60–70% IRM) strength training. No machines were used for the strength training part. All subjects were individually supplemented with calcium and cholecalciferol up to a maximum of 1500 mg/d and 500 IU/d. Bone mineral density at the lumbar spine and the hip was measured with DXA and QCT. Falls as defined by the PROFANE workgroup were counted using daily protocols conducted by all participants. Here we present initial 12 month DXA and fall results for 182 participants (EG: n=90; WCG: n=92).

After 12 months there were significant differences between exercise and wellness control group of BMD of the spine (EG: +1.4% vs. WCG: −0.3%) and of the hip (EG: −0.2% vs. WCG: −1.3%). The number of falls was significantly higher in the WCG (n=124) than in the EG (n=72). The same was true for injurious falls (EG: n=28 vs. WCG: n=42). In the exercise group 5 and in the wellness control group 8 fractures occurred but the power of the subset of patients included in this preliminary analysis was too low for the difference to be significant.

In conclusion preliminary results of our randomized exercise trial show that exercise training positively impact BMD at the hip and spine as well as fall frequency and may prevent fall related fractures in elderly subjects. The training scheme used in the study can easily be transferred into rehabilitation programs as no special equipment was used.

Disclosures: K. Engelke, Synarc 3.

WG12

Relationship Between Serum 25-hydroxyvitamin D₃ Concentration and Walking Ability, Leg Strength, or Balance in Community-Dwelling Japanese Frail Elderlies. J. Okuno*, S. Tomura*, H. Yanagi*, N. Yabushita*, T. Okura*, K. Tanaka*. Graduate school of Comprehensive Human Sciences, University of Tsukuba, Tsukuba City, Japan.

The aim of this study was to evaluate the relationship between serum 25-hydroxyvitamin D₃ (25OHD) concentration and walking ability, muscle strength, or balance in community-dwelling Japanese frail elderlies. The study was a longitudinal study conducted in a town near Tsukuba city (latitude 36°north) from June to September, in 2005 and 2006. Forty-five participants were community dwelling elderlies aged 65 years and over who required minimum support or nursing care to maintain their ADL. Their mean age was 76.5±5.9 years (mean±SD, range: 65–90). Of the participants, 32 attended a 3-month exercise program for nursing care prevention once per week (exercise group). The remaining 13 received only advice for nursing care prevention (control group). The Ethics Committee of University of Tsukuba approved the study. An interview was conducted based on a questionnaire including score of functional capacity of ADL (Tokyo Metropolitan Institute of Gerontology (TMIG) score), experiences of fall, stumbling and body sway during the past one year, walking ability, and the frequency of going outside of the home. The serum levels of 25OHD, intact parathyroid hormone (iPTH) and calcium were measured. The following physical tests were performed at baseline and 3 months: Timed Up & Go (TUG) and a 5-meter walk for walking ability, functional reach and foot balance with open eyes for balance, trunk flexion for flexibility, and ankle strength and grip strength for muscle strength. Among 45 patients who underwent the tests, 66.7% and 40% had difficulties in standing up and in walking, respectively. 54.3% experienced falls, 73.8% experienced stumbling and body sway more than once during the past one year. The mean level of 25OHD was 62.1±13.7 nmol/L (mean±SD, range: 27.5–87.5). At baseline, the rate of 25OHD levels less than 50 nmol/L that is the cut-off point of 25OHD resulting from the significant difference of intact PTH level was 18.0%. At baseline and 3 months there were no significant differences in physical tests between the control group and the exercise group. The subjects with 25OHD levels more than 50 nmol/L at baseline improved their walking ability and ankle strength for a period of three months in both groups. On the other hand, the subjects with 25OHD levels less than 50 nmol/L did not improve their physical performance even in the exercise group. It is suggested that serum 25OHD level is related to walking ability and muscle strength in Japanese frail elderlies, and that serum 25OHD level more than 50 nmol/L may be needed for maintaining their mobility or balance.

Disclosures: J. Okuno, None.