Some of these data were presented at the 19th Annual Meeting of the American Society for Bone and Mineral Research, September 10–14, 1997, Cincinnati, OH, U.S.A. (. Abstract F297). Approved as Journal Paper Number 15795 of the Purdue Agricultural Experiment Station.
Dietary Conjugated Linoleic Acids Alter Serum IGF-I and IGF Binding Protein Concentrations and Reduce Bone Formation in Rats Fed (n-6) or (n-3) Fatty Acids†
Article first published online: 1 JUL 1999
Copyright © 1999 ASBMR
Journal of Bone and Mineral Research
Volume 14, Issue 7, pages 1153–1162, July 1999
How to Cite
Li, Y., Seifert, M. F., Ney, D. M., Grahn, M., Grant, A. L., Allen, K. G. D. and Watkins, B. A. (1999), Dietary Conjugated Linoleic Acids Alter Serum IGF-I and IGF Binding Protein Concentrations and Reduce Bone Formation in Rats Fed (n-6) or (n-3) Fatty Acids. J Bone Miner Res, 14: 1153–1162. doi: 10.1359/jbmr.19126.96.36.1993
- Issue published online: 2 DEC 2009
- Article first published online: 1 JUL 1999
- Manuscript Accepted: 19 JAN 1999
- Manuscript Revised: 28 DEC 1998
- Manuscript Received: 25 SEP 1998
A study was designed to examine the effects of dietary conjugated linoleic acid (CLA) on serum concentrations of insulin-like growth factor-I (IGF-I) and IGF binding proteins (IGFBP) and the relationship of these factors to bone metabolism. Weanling male rats were fed AIN-93G diet containing 70 g/kg of added fat for 42 days. Treatments included 0 g/kg or 10 g/kg of CLA and soybean oil (SBO) or menhaden oil + safflower oil (MSO) following a 2 × 2 factorial design. Serum IGFBP was influenced by dietary polyunsaturated fatty acid (PUFA) type ((n-6) and (n-3)) and CLA (p = 0.01 for 38–43 kDa bands corresponding to IGFBP-3). CLA increased IGFBP level in rats fed SBO (p = 0.05) but reduced it in those fed MSO (p = 0.01). Rats fed MSO had the highest serum IGFBP-3 level. Both (n-3) fatty acids and CLA lowered ex vivo prostaglandin E2 production in bone organ culture. In tibia, rats given CLA had reduced mineral apposition rate (3.69 vs. 2.79 μm/day) and bone formation rate (BFR) (0.96 vs. 0.65 μm3/μm2/day); however, the BFR tended to be higher with MSO. Dietary lipid treatments did not affect serum intact osteocalcin or bone mineral content. These results showed that dietary PUFA type and CLA modulate local factors that regulate bone metabolism.
Conjugated linoleic acid (CLA) is the name given to describe a group of positional and geometric fatty acid isomers derived from linoleic acid (LA). The double bonds in CLA are conjugated and not separated by a methylene group as in LA. Further, CLA will not substitute for LA as an essential fatty acid. CLA isomers occur naturally in ruminant food products during the process of bacterial biohydrogenation(1) and are reported to possess potent beneficial health and biological effects.(2–5) CLA was first recognized as an anticarcinogen after being isolated from extracts of grilled ground beef and shown to reduce skin tumors in mice treated with 7,12-dimethylbenz[a]anthracene, a known carcinogen.(6) In studies to determine its physiological action, CLA was found to reduce prostaglandin E2 (PGE2) concentration in rat serum, spleen,(7,8) cultured keratinocytes,(9) and ex vivo bone organ culture.(10)
We previously reported,(10) as well as other investigators,(7,11) that CLA was found in numerous tissues examined in animals given a dietary CLA supplement. Because CLA is incorporated into membrane phospholipids, it may compete with other polyunsaturated fatty acids (PUFAs) in the formation of arachidonic acid (AA) (the precursor of PGE2) to inhibit PGE2 biosynthesis.(10,12)
Bone cells, primarily osteoblasts, synthesize insulin-like growth factor-I (IGF-I), IGF-II, and six of the seven known IGF binding proteins (IGFBP).(13,14) It is believed that IGF and their binding proteins modulate bone formation.(15) IGFBP can influence the action of IGF and may have direct effects on cell growth and differentiation.(16) It is also known that the presence of IGF and their binding proteins in bone is affected by PGE2. For example, PGE2 stimulated the synthesis of IGF-I and IGFBP-5 in primary osteoblast-enriched cultures from fetal rat bone,(17) increased IGF-I and IGFBP-3 transcript and polypeptide levels in rat calvaria cells,(18,19) and stimulated the expression of IGFBP-3 mRNA,(19,20) and IGFBP-4(21) in human articular chondrocytes and IGFBP-5 in primary rat osteoblasts.(22) Since PGE2 plays an important role in the local regulation of bone formation and resorption,(23) and PGE2 is also a potent regulator of IGF-I and IGFBP in bone,(18,19) we speculate that CLA could exert a regulatory effect on the production and action of IGF-I and IGFBP by modulating PGE2 production to impact local bone metabolism.
This laboratory previously reported that a dietary source of anhydrous butterfat stimulated bone formation rates (BFRs) in growing animals presumably by modulating the production of PGE2 in bone.(24) Milk fat might be the richest natural source of CLA with a concentration up to 30 mg/g of fat, though it could be affected by pasture conditions.(25) Together with the fact that dairy products, especially milk, are the most important food source of calcium for bone development in children and adults, the results of the current study would further the understanding of the health benefits of consuming dairy products. Therefore, the purpose of the present investigation was to evaluate the effects of CLA on IGF-I and IGFBP concentrations and bone histomorphometry in rats given a semipurified diet containing a rich source of either (n-6) or (n-3) PUFAs.
MATERIALS AND METHODS
Animals and diets
Forty male weanling Sprague-Dawley (21 days old, mean body weights 46 ± 2.0 g; Harlan Sprague-Dawley, Indianapolis, IN, U.S.A.) rats were housed in individual cages under a 12-h light-dark cycle. Animal care was in compliance with applicable guidelines from the Purdue University policy on animal care and use. The rats were assigned to four dietary groups following a 2 × 2 factorial design. The basal diet (AIN-93G without fat; Dyets, Inc. Bethlehem, PA, U.S.A.) contained one of the following lipid treatments: SBO (soybean oil, diet rich in (n-6) PUFA) or MSO (menhaden oil + safflower oil at a ratio by weight of 56:44, a diet rich in (n-3) PUFA) at 70 g/kg diet with or without added CLA (Table 1). For diets containing CLA (SBOC and MSOC), 10 g/kg of dietary SBO or MSO were replaced with CLA (generously provided by Dr. J.A. Scimeca, Kraft General Foods, Glenview, IL, U.S.A.). The SBO diet (AIN-93G) also served as a control diet since it contained all known nutrients for the growing rat as recommended by the American Institute of Nutrition.(26) The 1% dietary level of CLA was used because 0.5–1.5% of CLA was evaluated in previous studies for anticarcinogenic activity.(8,27–30) All diets were isocaloric and isonitrogenous. Fresh diets were prepared every 14 days and kept at −20°C until fed. Food cups were refilled three times per week and feed consumption measured at these times. Body weights were recorded weekly. All lipids were stored at −80°C before dietary use.
After 42 days of feeding, rats were killed and tissues collected for IGF-I measurement and bone histomorphometric analysis. For dynamic assessment of bone formation parameters, animals were given double intraperitoneal injections of calcein green (10 mg/kg of body weight; Sigma Chemical Co., St. Louis, MO, U.S.A.) 5 days and 1 day before killing. Blood was collected by bleeding from axillary vessels. Serum was harvested from blood by centrifugation at 1200g for 20 minutes at 4°C and stored at −80°C until analyzed. Bones were removed and carefully freed of soft tissue. Right proximal tibiae, collected for bone histology, were fixed in 10% neutral buffered formalin, dehydrated in ethanol, and embedded undecalcified in methylmethacrylate.(31,32)
Bone histomorphometric analyses were performed on frontal sections of proximal tibial metaphyses cut using a Reichert-Jung 2050 microtome (Heidelberg, Germany). Static parameters were measured on sections (5-μm-thick) stained with von Kossa/tetrachrome, and kinetic measurements were performed on unstained sections (10-μm-thick) viewed using fluorescence microscopy.(31,32) Trabecular tissues were measured in a standard area (2.4 mm2) located just below the growth plate zone of mineralized cartilage, composed of secondary spongiosae. This area excluded trabeculae connected to the osseous cortex. Two sections per animal were analyzed at an objective magnification of 20×, and both static and kinetic parameters were measured in the same area. Sections were analyzed using a semiautomatic image analysis system (Osteomeasure Histomorphometry System; Osteometrics, Inc., Atlanta, GA, U.S.A.).(31) The following static parameters were measured in stained tissue sections: trabecular bone volume (BV/TV,%), trabecular thickness (μm), trabecular separation (μm), and trabecular number (#/mm). Kinetic parameters measured from unstained sections included total fractional labeled surface (LS/BS, %), single-labeled surface (sL.S/BS, %), double-labeled surface (dL.S/BS, %), mineral apposition rate (MAR, μm/day), and bone formation rate (BFR/BS, μm3/μm2/day). These measurements are indicators of bone mass and bone formation, and the formulas used for their calculation are based on the recommendations of the American Society for Bone and Mineral Research Nomenclature Committee.(33)
Bone mineral analyses were performed on rat humeri as described by Watkins et al.(24) Briefly, bones were cleaned of soft tissue, dried, weighed, and measured for length using a caliper. Bones were dry-ashed at 600°C for 48 h, and the ash content was calculated by weight loss on a dry basis. Bone ash was digested with 15.9 mol/l of HNO3 and calcium, magnesium, and phosphorus measured by inductively coupled plasma–atomic emission spectroscopy (Plasma 4000; Perkin-Elmer, Norwalk, CT, U.S.A.). Bone mineral content was expressed as ash weight per bone length (mg/mm).
Lipids in the diet and tissue samples were extracted with chloroform/methanol (2:1, v/v). Lipids from the diet were saponified, and fatty acid methyl esters (FAMEs) were prepared by transesterification using boron trifluoride in methanol (12%, w/w) (Supelco, Inc., Bellefonte, PA, U.S.A.). Lipid extraction was performed on pulverized cortical bone cooled in liquid nitrogen.(24) Lipid extracts from rat tissues were methylated by sodium methoxide in methanol.(10) Briefly, the lipid sample was dissolved in dry toluene (1 ml) in a test tube, 0.5 M sodium methoxide in anhydrous methanol (2 ml) added (50°C for 10 minutes), followed by the additions of glacial acetic acid (0.1 ml) and deionized H2O (5 ml). The FAME were extracted into hexane (2 × 3 ml) using a Pasteur pipette to separate the layers. The hexane layer was dried over anhydrous sodium sulfate and filtered. FAMEs were analyzed using a gas chromatograph (HP 5890 series II, auto sampler 7673, HP 3365 ChemStation; Hewlett-Packard Co., Avondale, PA, U.S.A.) equipped with a DB 225 or DB 23 column (30 m, 0.53 mm internal diameter, film thickness 0.5 μm; J&W Scientific Co., Folsom, CA, U.S.A.) and operated at 140°C for 2 minutes, temperature programmed 1.5°C/minute to 198°C and held for 7 minutes. The injector and flame-ionization detector temperatures were 225°C and 250°C, respectively. FAMEs were identified by comparison of their retention times with authentic standards (GLC-422, CLA [UC-59-A and UC-59-M]; Nu-Chek-Prep, Elysian, MN; CLA [catalog #1245, c9,t11 and catalog #1181, t9,t11] Matreya, Inc., Pleasant Gap, PA, U.S.A.) and FAMEs of menhaden oil (catalog #1177; Matreya Inc.).
Ex vivo PGE2 production in bone organ cultures were performed as previously described.(24,32) Briefly, shafts from the right tibia and femur were removed and carefully flushed with 0.9% NaCl to remove marrow cells. A section of bone shaft was immersed in 20 ml of Hank's balanced salt solution (Sigma Chemical Co.) and incubated with shaking for 2 h at 37°C. After incubation, the bone culture medium was collected and frozen at −80°C until analyzed. Values for PGE2 were expressed as nanograms per gram of bone wet or dry weight.
The relative amount of IGFBP in serum was determined by a Western ligand blotting method with modification.(34) Briefly, serum samples were diluted 1:10 with Laemmli sample buffer without β-mercaptoethanol, heated at 60°C for 10 minutes, and 15 μl was loaded onto a discontinuous SDS-PAGE (3.75% stacking and 10% resolving) for electrophoresis. Proteins were transferred onto 0.2 μm nitrocellulose filter paper, and the membranes were blotted with I125–IGF-I (Amersham, Arlington Heights, IL, U.S.A.) using a rotating hybridization tube at 4°C. Dried membranes were exposed to X-ray film (Kodak X-Omat; Eastman Kodak, Rochester, NY) with intensifying screens at −80°C. Band density was quantitated by laser densitometry (Ultrascan XL; Pharmacia LKB Biotechnology Inc., Piscataway, NJ, U.S.A.) and expressed as optical density units. The method for the detection of IGFBP was validated by a competitive binding experiment using125I–IGF-I and excessive cold IGF-I.(34) Serum IGF-I concentration was measured as described.(35) Serum intact osteocalcin was measured by immunoradiometric assay using a commercial kit (Immutopics, Inc., San Clemente, CA, U.S.A.).
Data were statistically analyzed by a two-way analysis of variance (ANOVA), and where significant differences were found, a Tukey's studentized range test or Duncan's multiple-range test at a probability of α = 0.05 was performed using the SAS software package for UNIX (SAS Institute, Inc., Cary, NC, U.S.A.). By using a factorial design, the number of subjects for each main treatment effect ranged from n = 9–16. Student's t-tests were also performed as indicated. Variation between treatment groups was expressed as the pooled standard error of the mean (± SEM) where applicable. Results were expressed as mean ± SD when sample size was not equal among treatment groups.
Although rat body weights between the control and three treatment groups were not different throughout the feeding period, feed efficiency (total body weight [g]/total feed consumed [g]) was significantly improved in rats fed CLA compared with those not given CLA (0.42 vs. 0.39, p < 0.01). The dietary levels of (n-6) or (n-3) PUFA did not influence body weight or feed efficiency.
Serum IGF-I concentration was lower (p < 0.02) in rats given the dietary CLA supplement compared with those not given CLA after 6 weeks of dietary treatment (Fig. 1). The average serum IGF-I concentrations for rats fed CLA were 501 ± 53 ng/ml (SBOC treatment) and 484 ± 53 ng/ml (MSOC treatment). IGF-I concentrations for rats not given the CLA supplement were 607 ± 25 ng/ml and 584 ± 40 ng/ml for the SBO and MSO treatments, respectively. PUFA type did not significantly affect circulating IGF-I levels.
Rat tissue fatty acid composition was significantly influenced by dietary PUFA treatment and CLA supplementation. In serum (Table 2), rats given the high (n-6) PUFA diets (SBO, SBOC) had higher values for 18:0, 18:1, 18:2(n-6), 18:3(n-6), 18:3(n-3), 20:2(n-6), 20:3(n-6), 20:4(n-6), 22:4(n-6), total monounsaturates, and total (n-6) PUFA but lower 14:0, 16:0, 16:1(n-7), 20:5(n-3), 22:5(n-3), 22:6(n-3), total saturates, and total (n-3) PUFA compared with those fed diets high in (n-3) fatty acids (MSO, MSOC). Dietary CLA supplementation increased the values for 16:0 and total saturates in both PUFA treatment groups while values for 22:5(n-3), 22:6(n-3) and total (n-3) PUFA were increased only in rats fed the high (n-3) diet. Similar treatment effects were observed for bone marrow fatty acids (Table 3). Supplementation with CLA also decreased the values for 18:1, 18:2(n-6), 20:2(n-6), 20:3(n-6), 22:5(n-6), 22:6(n-3), total monounsaturates, total (n-6) PUFA, and total PUFA in rat bone marrow. The addition of CLA to both PUFA treatments resulted in increased values for various CLA isomers (18:2 c-9, t-11; 18:2 t-10, c-12, and/or 18:2 t,t) in serum, bone marrow, and cortical bone (data not shown). Since CLA supplementation decreased both total (n-6) PUFA and total PUFA, but had little effect on total (n-3) PUFA, it seems that CLA to some extent displaced (n-6) PUFA in tissue lipids of the rat.
The values for serum IGFBP in rats were strongly influenced by the interaction between dietary PUFA type and CLA supplementation (Fig. 2). Three distinct groups of125I–IGF-I labeled bands were observed by ligand blotting. The most pronounced bands occurred in the 38–43 kDa range, corresponding to IGFBP-3.(35) Rats given MSO had the highest serum IGFBP-3 and total IGFBP levels. Supplementation with CLA resulted in opposite effects on IGFBP concentration in rats given the different PUFA treatments. CLA tended to increase the serum IGFBP level in rats given SBO but reduced it in rats given MSO.
Histomorphometric analysis of tibia (Table 4) revealed that MAR and BFR were significantly reduced in rats given the CLA supplement (p ≤ 0.05) independent of the dietary PUFA type. Bone static measurements, such as trabecular bone volume, trabecular thickness, trabecular separation, and trabecular number were not affected by either dietary PUFA type or CLA supplementation. Rats given the high (n-6) PUFA treatment tended to have a relatively higher double labeled surface (p = 0.08). Single-labeled surface was not affected by either PUFA type or CLA supplementation.
Bone mineral analyses of right humeri revealed that bone dry weight, ash weight, and mineral density were lower in rats fed SBO (high (n-6) diet) compared with those fed MSO (high (n-3) diet) (Table 5) without the CLA supplement. There was no dietary PUFA type or CLA supplementation effect on bone calcium, magnesium, and phosphorus contents.
Data for ex vivo PGE2 biosynthesis revealed that rats consuming SBO had a higher production of this prostanoid in bone organ culture compared with those given MSO (Table 6). Addition of the CLA treatment to either the SBO or MSO diets dramatically lowered ex vivo PGE2 production (Table 6).
Serum intact osteocalcin was not affected by either dietary PUFA type or CLA supplementation. The mean values for the treatment groups were 47.7 ± 2.1 ng/ml for SBO, 49.0 ± 2.9 ng/ml for SBOC, 45.2 ± 2.1 ng/ml for MSO, and 45.1 ± 1.9 ng/ml for MSOC (n = 10 for each group).
IGF-I, the most abundant growth factor in bone, is believed to function as both a systemic and local growth factor for bone tissue.(36) The anabolic effects of IGF-I in bone include the stimulation of longitudinal growth and increase of bone mass.(36,37) Production of IGF-I is also regarded as a key factor for mediating the effects of a number of independent signaling molecules in bone cells.(38) Thus, understanding the regulation of the production and activity of IGF-I is critical for elucidating the potential impact of dietary factors on bone metabolism. This is the first study to show that CLA, a dietary anticarcinogen present in dairy products and red meat, influenced the serum IGF/IGFBP system and was associated with reduced bone formation.
In the present investigation, CLA was shown to down-regulate the circulating levels of IGF-I in the growing rat. Based on these data, CLA could have exerted its effect by modulating eicosanoid metabolism, which is consistent with previous work showing that CLA reduced ex vivo bone organ culture PGE2 production.(10) Serum IGFBP was also affected by dietary CLA supplementation although it was dependent on the dietary PUFA type. CLA increased the serum IGFBP level in rats given the high (n-6) PUFA diet, but decreased it in rats fed the high (n-3) PUFA diet. Since IGF-I and its binding proteins were affected differently by CLA depending on the (n-6) and (n-3) PUFA type, dietary PUFA may potentially impact bone metabolism via IGF mechanisms depending on the ratio of (n-6) and (n-3) PUFA in the diet.
CLA supplementation reduced MAR and BFR in rats regardless of the dietary source of PUFA in this study. Furthermore, CLA lowered serum IGF-I level. The dietary (n-6) and (n-3) PUFA treatments did not significantly affect IGF-I concentration and bone histomorphometric measurements. Serum intact osteocalcin was not affected by dietary PUFA type or CLA supplementation. PUFA treatments, however, did affect bone mineral density. Rats fed (n-6) PUFA had a lower ash weight per millimeter of bone length in humeri compared with those fed (n-3) PUFA. These findings suggest that excessive consumption of (n-6) PUFA could have a negative effect on bone metabolism by increasing bone resorptive activity through increased endogenous production of PGE2. This is consistent with the results of a previous study wherein chicks fed SBO for 21 days showed decreased total bone and cortical bone areas in cross-sections of tibiae compared with chicks given menhaden oil.(32)
Dietary PUFA treatment and CLA supplementation both had a significant effect on ex vivo PGE2 production in tibia and femur organ cultures. Rats supplemented with CLA had lowered ex vivo PGE2 production in bone. PGE2 is an important factor in regulating local bone metabolism(23) including bone modeling and remodeling.(39) Sugano et al.(7) reported that the concentration of PGE2 in serum and spleen tended to be reduced by CLA. Raisz and Koolemans-Beynen showed that PGE2 inhibited bone matrix formation at a high concentration in bone organ culture(40); however, at lower concentrations, PGE2 stimulated bone formation in vitro and in vivo.(40–43) Therefore, excessive production of PGE2 may adversely affect bone modeling, whereas a lower level of PGE2 is believed to stimulate bone formation in animals fed diets containing moderate levels of (n-6) PUFA. For example, reduced PGE2 production in chicks fed a higher (n-3) diet was associated with increased rate of bone formation.(32,44) Although CLA and (n-3) PUFA may follow different pathways in modulating tissue PGE2 production, the fact that they both lowered ex vivo bone PGE2 level suggests dietary moderation of bone prostanoid production.
In the current study, rats fed CLA had decreased values for MAR and BFR which likely reflect some impact on osteoblastic function. While these CLA effects may be mediated through the regulation of PGE2 synthesis (via the inducible enzyme cyclooxygenase-2 or isomeric analogs of CLA), which in turn influence IGF-I concentrations, they may also be mediated by other potent cytokines, such as interleukins (IL-1 and IL-6), tumor necrosis factor (TNF) or the lipoxygenase product, leukotriene B4 (Fig. 3). Dietary CLA was recently shown to lower basal, and lipopolysaccharide stimulated IL-6 production and basal TNF production by resident peritoneal macrophages in rats.(45) Furthermore, CLA reduced the release of leukotriene B4, a lipoxygenase product of AA, one of the most potent chemotaxins for polymorphonuclear leukocytes and monocytes(46) and a strong bone resorption factor,(47) from peritoneal exudate cells.(8) Assuming CLA has similar effects on these cytokines in bone, together with the fact that CLA reduced the production of PGE2 in bone tissue, one could hypothesize that at a proper dietary level, CLA anti-inflammatory effects could be beneficial for the treatment of inflammatory bone disease. Further study, however, is needed to unequivocally establish how CLA mediates its effect on bone cell metabolism.
This research is the first to show that CLA isomers affect bone metabolism in laboratory animals. The level of CLA used in this study (1% dietary CLA), though higher than that found in conventional diets without supplementation, compares favorably to the range used (0.5–1.5%) in other studies that examined anti-inflammatory and anticarcinogenic properties of CLA.(8,27–30) Further work is needed to evaluate more typical dietary levels of CLA on bone metabolism.
This work was supported by USDA/NRI grant no. 96–35200–3137.
- 11992 Dietary sources of conjugated dienoic isomers of linoleic acid, a newly recognized class of anticarcinogens J Food Comp Anal 5:185–197., , , ,
- 21995 The role of phenolics, conjugated linoleic acid, carnosine, and pyrroloquinoline quinone as nonessential dietary antioxidants Nutr Rev 53:49–58.
- 31994 Conjugated linoleic acid suppressed mammary carcinogenesis and proliferative activity of the mammary gland in the rat Cancer Res 54:1212–1215., , ,
- 41989 Antioxidants—Carcinogenic and chemopreventive properties Adv Cancer Res 53:247–302.,
- 51994 Conjugated linoleic acid and atherosclerosis in rabbits Atherosclerosis 108:19–25., ,
- 61987 Anticarcinogens from fried ground beef: Heat altered derivatives of linoleic acid Carcinogenesis 8:1881–1887., ,
- 71997 Lymphatic recovery, tissue distribution, and metabolic effects of conjugated linoleic acid in rats J Nutr Biochem 8:38–43., , , , ,
- 81998 Conjugated linoleic acid modulates tissue levels of chemical mediators and immunoglobulins in rats Lipids 33:521–527., , , ,
- 91998 Conjugated linoleic acid reduces arachidonic acid content and PGE2 synthesis in murine keratinocytes Cancer Lett 127:15–22.,
- 101998 Conjugated linoleic acids alter bone fatty acid composition and reduce ex vivo prostaglandin E2 biosynthesis in rats fed n-6 or n-3 fatty acids Lipids 33:417–425.,
- 111995 Dietary conjugated linoleic acid changes fatty acid composition in different tissues by decreasing monounsaturated fatty acids. 1995 IFT Annual Meeting / Book of Abstracts, 183., ,
- 121989 Effects of d,l-2-difluoromethylornithine and indomethacin on mammary tumor promotion in rats fed high n-3 and/or n-6 fat diets Cancer Res 49:1434–1440., , , , ,
- 131994 Insulin-like growth factors and bone: The osteoporosis connection Proc Soc Exp Biol Med 206:83–102., ,
- 141996 Synthesis and characterization of insulin-like growth factor-binding protein (IGFBP)-7. Recombinant human mac25 protein specifically binds IGF-I and IGF-II J Biol Chem 271:30322–30325., , , , ,
- 151996 Transcriptional regulation of insulin-like growth factor-binding protein-5 by prostaglandin E2 in osteoblast cells Endocrinology 137:2375–2382.,
- 161990 Insulin-like growth factors and their binding proteins Physiol Rev 70:591–614.,
- 171994 Complex pattern of insulin-like growth factor binding protein expression in primary rat osteoblast enriched cultures: Regulation by prostaglandin E2, growth hormone, and the insulin-like growth factors J Cell Physiol 160:163–175., , ,
- 181991 Prostaglandin E2 stimulates insulin-like growth factor I synthesis in osteoblast-enriched cultures from fetal rat bone Endocrinology 128:2895–2900., , ,
- 191992 Prostaglandin E2 stimulates synthesis of insulin-like growth factor binding protein-3 in rat bone cells in vitro J Bone Miner Res 7:1157–1163., , , ,
- 201996 Prostaglandin E2 up-regulates insulin-like growth factor binding protein-3 expression and synthesis in human articular chondrocytes by a c-AMP-independent pathway: Role of calcium and protein kinase A and C J Cell Biochem 63:320–333., , ,
- 211997 Prostaglandin E2 stimulates insulin-like growth factor binding protein-4 expression and synthesis in cultured human articular chondrocytes: possible mediation by Ca (++)-calmodulin regulated processes J Cell Biochem 65:408–419., , , , ,
- 221996 Promoter-dependent and -independent activation of insulin-like growth factor binding protein-5 gene expression by prostaglandin E2 in primary rat osteoblasts J Biol Chem 271:6666–6671., , , , ,
- 231993 Prostaglandins and the skeleton: The legacy and challenges of two decades of research Endocr J 1:337–344.,
- 241997 Dietary lipids modulate bone prostaglandin E2 production, insulin-like growth factor-I concentration and formation rate in chicks J Nutr 127:1084–1091., , , , , ,
- 251996 Milk fat components: Possible chemopreventive agents for cancer and other diseases Aust J Dairy Technol 51:24–32.
- 261993 AIN-93 purified diets for laboratory rodents: Final report for the American Institute of Nutrition Ad Hoc Writing Committee on the reformulation of the AIN-76A rodent diet J Nutr 126:1939–1951., ,
- 271994 Conjugated linoleic acid is a growth factor for rats as shown by enhanced weight gain and improved feed efficiency J Nutr 124:2344–2349., , , ,
- 281991 Mammary cancer prevention by conjugated dienoic derivative of linoleic acid Cancer Res 51:6118–6124., , ,
- 291996 The efficacy of conjugated linoleic acid in mammary cancer prevention is independent of the level or type of fat in the diet Carcinogenesis 17:1045–1050., , , , ,
- 301997 Dietary conjugated linoleic acid induces peroxisome-specific enzyme accumulation and ornithine decarboxylase activity in mouse liver J Nutr Biochem 8:579–584., , ,
- 311994 Lack of evidence for rickets in the osteopetrotic rat mutation, toothless J Bone Miner Res 9:1813–1821.
- 321996 Dietary (n-3) and (n-6) polyunsaturates and acetylsalicylic acid alter ex vivo PGE2 biosynthesis, tissue IGF-I levels, and bone morphometry in chicks J Bone Miner Res 11:1321–1332., , ,
- 331987 Bone histomorphometry: Standardization of nomenclature, symbols and units. Report of the ASBMR Histomorphometry Nomenclature Committee J Bone Miner Res 2:595–610., , , , ,
- 341986 Analysis of serum insulin-like growth factor binding proteins using western blotting: Use of the method for titration of the binding proteins and competitive binding studies Anal Biochem 154:138–143., , , ,
- 351995 High-calorie total parenteral nutrition reduces hepatic insulin-like growth factor-I mRNA and alters serum levels of insulin-like growth factor-binding protein-1, −3, −5, and −6 in the rats Metabolism 44:152–160., , ,
- 361994 Cellular and clinical perspectives on skeletal insulin-like growth factor I J Cell Biochem 55:328–333., ,
- 371994 In vivo and in vitro effects of insulin-like growth factor-I (IGF-I) on femoral mRNA expression in old rats Bone 15:647–653., , , ,
- 381996 Invited review of a workshop: Anabolic hormones in bone: Basic research and therapeutic potential J Clin Endocrinol Metab 81:872–877., ,
- 391990 Biphasic effects of prostaglandin E2 on bone formation in cultured fetal rat calvariae: Interaction with cortisol Endocrinology 126:1654–1659.,
- 401974 Inhibition of bone collagen synthesis by prostaglandin E2 in organ culture Prostaglandins 8:377–385.,
- 411984 Stimulation of bone formation by prostaglandin E2 Prostaglandins 27:97–103.,
- 421990 Prostaglandin E2 enhances cortical bone mass and activates intracortical bone remodeling in intact and ovariectomized female rats Bone 11:253–266., , ,
- 431990 Effects of prostaglandin E2 on production of new cancellous bone in the axial skeleton of ovariectomized Rats Bone 11:103–113., , , ,
- 441994 Dietary lipids modify the fatty acid composition of cartilage, isolated chondrocytes and matrix vesicles Lipids 29:619–625., ,
- 451998 Modulation of macrophage cytokine production by conjugated linoleic acids is influenced by the dietary n-6:n-3 fatty acid ratio J Nutr Biochem 9:258–266., , , ,
- 461980 Chemokinetic activity of arachidonic and lipoxygenase products on leukocytes of different species Prostaglandins 20:411–418., , ,
- 471996 Leukotriene B4 stimulates osteoclastic bone resorption both in vitro and in vivo J Bone Miner Res 11:1619–1627., , , , , ,