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Introduction

  1. Top of page
  2. Introduction
  3. T Cells and Ovariectomy Induced Bone Loss
  4. Estrogen Regulation of T Cell Activation and TNF Production
  5. T Cell Thymic Output and Bone Loss
  6. Effects of Ovariectomy in the Bone Marrow
  7. Interactions Between T Cells and Stromal Cells
  8. A Hypothesis
  9. Conclusions
  10. Disclosures
  11. References

More than a decade has passed since the publication of the first report implicating T cells in the bone loss induced by ovariectomy (ovx).1 Since then there has been extraordinary progress in the understanding of the regulatory network that links the hemopoietic and the mesenchymal compartments of the bone marrow (BM), the interactions between the immune system and bone, and the role of lymphocytes as mediators of the effects of calciotrophic hormones in bone. Collectively this body of knowledge has led to the firm establishment of “osteoimmunology” as a novel discipline and a promising area of investigation. The objective of this perspective is to revisit the exciting hypothesis that T cells play a pivotal role in the mechanism of ovx-induced bone loss.

T Cells and Ovariectomy Induced Bone Loss

  1. Top of page
  2. Introduction
  3. T Cells and Ovariectomy Induced Bone Loss
  4. Estrogen Regulation of T Cell Activation and TNF Production
  5. T Cell Thymic Output and Bone Loss
  6. Effects of Ovariectomy in the Bone Marrow
  7. Interactions Between T Cells and Stromal Cells
  8. A Hypothesis
  9. Conclusions
  10. Disclosures
  11. References

The bone-sparing activity of estrogens is due to a repression of bone remodeling coupled with a balancing effect on bone formation and resorption.2 The dominant acute effect of estrogen is the blockade of new osteoclast formation, cells which arise by cytokine-driven proliferation and differentiation of monocyte precursors that circulate within the hematopoietic cell.3 This process is facilitated by BM stromal cells (SCs), which provide physical support for nascent osteoclasts and produce soluble and membrane-associated factors essential for the proliferation and differentiation of osteoclast precursors. This inhibitory effect on osteoclastogenesis is associated with a repressive effect on osteoblastogenesis.4 Another critical effect of estrogen is that of increasing osteoclast apoptosis5–7 while blocking the apoptosis of osteoblasts and osteocytes.2, 8 It has been proposed that the effects of estrogens on the generation of osteoblasts and the lifespan of osteoblasts and osteoclasts result from extranuclear actions of the endoplasmic reticulum (ER) and activation of cytoplasmic kinases.2, 7

The decline of ovarian function at menopause results in decreased production of estrogen and a parallel increase in pituitary follicle-stimulating hormone (FSH) levels. The combined effects of estrogen deprivation and elevated FSH production cause a marked stimulation of bone resorption9 and a period of rapid bone loss that is central to the pathogenesis of postmenopausal osteoporosis. In mice the acute effects of menopause are modeled by ovx, a procedure that stimulates bone resorption by increasing osteoclast formation3 and lifespan.5–7 This initial phase of bone loss is followed by a slower but more prolonged loss of mainly cortical bone due to incomplete refilling of the resorption cavities due to insufficient osteoblast (OB) activity and lifespan.10 An expansion of the osteoclastic pool is therefore the key mechanism responsible for the bone loss that occurs early after ovx.

The net bone loss caused by ovx is limited, in part, by an increase in bone formation resulting from stimulated osteoblastogenesis.11 This compensation is fueled by an expansion of the pool of BM–SCs, increased commitment of such pluripotent precursors toward the osteoblastic lineage,11 and enhanced proliferation of early OB precursors.4 Subsequent escalations in OB apoptosis12, 13 extensions of osteoclast lifespan5, 6 and increased secretion of cytokines that suppress bone formation, such as interleukin 7 (IL-7) and tumor necrosis factor (TNF), are the likely reasons for why bone formation does not increase as much as resorption after ovx. The stimulatory effect of ovx on SCs is equally relevant for osteoclastogenesis as one of the consequences of estrogen deprivation is the formation of osteoblastic cells with an increased osteoclastogenic activity14; ie, the capacity to support osteoclast formation.

Evidence now suggests that T cells play a pivotal role in the mechanism of ovx-induced bone loss. Core observations include reports showing that ovx fails to induce trabecular and cortical bone loss in: T-cell–deficient nude mice,1, 15–17, wild-type (WT) mice deleted of T cells by injection of anti–T cell antibodies,18 mice treated with Abatacept19 (an agent that blocks T cell costimulation and induces T cell anergy and apoptosis)20, 21 and mice lacking the T cell costimulatory molecule CD40 ligand (CD40L).18 The fact that nude mice are protected against ovx-induced bone loss has been confirmed independently.22 By contrast, Lee and colleagues23 showed that nude mice are protected against the loss of cortical, but not trabecular bone induced by ovx. In another study, T-cell–deficient and B-cell–deficient mice were found to lose bone after ovx.24 The discrepancy between our reports1, 15–19, 25 and those of others23, 24 is likely explained by differences in the experimental design, the lack of B cells in some models, and compensatory mechanisms such as an increase in natural killer (NK) cells producing the osteoclastogenic factor IL-17.26

Both CD4+ and CD8+ cells have been found to play a role in ovx-induced bone loss. CD4+ cells include the TH1, Th2, and Th17 subsets. Th17 are regarded as the most osteoclastogenic subset of T CD4+ cells because they produce high levels of IL-17, receptor activator of NF-κB ligand (RANKL), and TNF, and low levels of interferon gamma (IFNγ).27, 28 The differentiation of Th17 is inhibited by estrogen via a direct effect mediated by estrogen receptor alpha (ERα).29 The role of Th17 in ovx-induced bone loss remains to be determined because one study reported that IL-17R–null mice are more susceptible to ovx-induced bone loss than controls,30 while another group found these mice to be protected from ovx-induced bone loss.26 More abundant information is available about regulatory T cells (Tregs), a population capable of suppressing the effector function of TH1, Th2, and Th17 T cells. Tregs are defined by the expression of the transcription factor FoxP3. Tregs inhibit monocyte differentiation into osteoclasts in vitro and in vivo, and blunt bone resorption31, 32 through the secretion of IL-4, IL-10, and transforming growth factor beta 1 (TGFβ1).33 Attesting to the relevance of Tregs, studies have shown that estrogen increases the relative number of Tregs.27, 34 Moreover, transgenic mice overexpressing Tregs develop progressive high bone mass due to inhibition of bone resorption, and are protected against ovx-induced bone loss.35 Moreover, adoptive transfer of Tregs into T cell–deficient mice increases bone mass, indicating that Tregs directly affect bone homeostasis without the need to engage other T cell lineages.35

Two mechanisms have been described to explain how T cells contribute to ovx-induced bone loss (Fig. 1). The first involves an increase in T cell activation, leading to enhanced production of TNF by BM T cells. The second is a regulatory crosstalk between T cells and SCs, resulting in enhanced production of osteoclastogenic cytokines by SCs.

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Figure 1. Schematic representation of the role of T cells in the mechanism by which ovx promotes osteoclastogenesis, osteoblastogenesis, and hemopoiesis. Estrogen deficiency promotes T cell activation by increasing the interaction of antigen (Ag)-loaded MHC molecules with bone marrow macrophages (BMM) and dendritic cells (DC) with the T cell receptor (TCR). The Ags are likely to be non-self peptides derived from the intestinal macrobiota. T cell activation also requires at least two costimulatory signals provided by the binding of BMM and DC-expressed CD40 and CD80 to the T cell surface molecules CD40L and CD28, respectively. A critical upstream event is the increased production of reactive oxygen species (ROS) that activate DCs by increasing their expression of CD80. The expansion of T cells in the BM is partially driven by an ovx-induced increase in the thymic output of naïve T cells. Activated T cells secrete TNF that stimulates osteoclast formation primarily by potentiating the response to RANKL. In addition, T cell–expressed CD40L and DLK1/FA-1 increase the osteoclastogenic activity of SC by blunting their secretion of OPG and augmenting their production of RANKL, M-CSF, and other proinflammatory factors.

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Estrogen Regulation of T Cell Activation and TNF Production

  1. Top of page
  2. Introduction
  3. T Cells and Ovariectomy Induced Bone Loss
  4. Estrogen Regulation of T Cell Activation and TNF Production
  5. T Cell Thymic Output and Bone Loss
  6. Effects of Ovariectomy in the Bone Marrow
  7. Interactions Between T Cells and Stromal Cells
  8. A Hypothesis
  9. Conclusions
  10. Disclosures
  11. References

The survival of naïve T cells and some memory T cells requires the low-affinity engagement of the T cell receptor (TCR) by a diverse repertoire of self-antigens (Ags) bound to major histocompatibility complex (MHC) molecules expressed on Ag-presenting cells.36, 37 By contrast, binding of foreign Ags such as bacterial Ags with MHC molecules is followed by high-affinity interactions with TCRs that drive T cell activation. Both low-affinity interactions with self-Ags and high-affinity foreign Ag/MHC/TCR interactions are referred to as “Ag presentation.”

Ovx induces T cell expression of activation markers and promotes T cell proliferation, expansion, and acquisition of effector functions. These are all features of T cells exposed to foreign Ags.36 The gastrointestinal tract is colonized for life with 100 trillion indigenous bacteria, creating a diverse ecosystem known as the microbiota, whose contributions to human health are profound.38 The macrobiota is likely to represent the source of foreign Ag that drives the expansion of T cells induced by ovx. Although direct verification of a role for macrobiota in T cell expansion is still lacking, compelling supportive data are available. For example, adoptive transfer of T cells into T cell–deficient mice is followed by rapid engraftment and expansion of donor T cells into the host. This process is driven by foreign Ags.36 Attesting to a role of the macrobiota, the expansion of transferred T cells into T cell–deficient mice is greatly reduced in host mice raised in a germ-free environment.36 Moreover, mice maintained in germ-free conditions display increased bone mass due to the lack of immune cell activation.39

T cells are key inducers of bone-wasting because ovx increases T cell TNF production to a level sufficient to augment RANKL-induced osteoclastogenesis.1 This effect is due to an increased number of TNF-producing T cells15 and enhanced production of TNF per cell.18, 40 Ovx also increases the population of premature senescent CD4 + CD28–T cells,40 a lineage that produces high levels of TNF.

The presence of increased levels of T cell–produced TNF in the BM of ovx animals is well documented.15, 19, 40, 41 Studies have also shown that menopause increases T cell activation and T cell production of TNF and RANKL in humans.42, 43 The role of TNF in ovx-induced bone loss has been demonstrated in multiple models. For example, ovx fails to induce bone loss in TNF-null mice and in animals lacking the p55 TNF receptor.15 Likewise, transgenic mice insensitive to TNF due to the overexpression of a soluble TNF receptor,44 and mice treated with the TNF inhibitor TNF binding protein45 are protected from ovx-induced bone loss. The specific relevance of T cell TNF production in vivo was demonstrated by the finding that although reconstitution of nude recipient mice with T cells from WT mice restores the capacity of ovx to induce bone loss, reconstitution with T cells from TNF-deficient mice does not.15

The mechanism by which estrogen deficiency expands the pool of TNF-producing T cells is summarized in Figure 1 and involves reactivation of thymic function and induction of T cell activation in the BM. T cell activation is driven by enhanced Ag presentation by macrophages and dendritic cells (DCs).46, 47

T Cell Thymic Output and Bone Loss

  1. Top of page
  2. Introduction
  3. T Cells and Ovariectomy Induced Bone Loss
  4. Estrogen Regulation of T Cell Activation and TNF Production
  5. T Cell Thymic Output and Bone Loss
  6. Effects of Ovariectomy in the Bone Marrow
  7. Interactions Between T Cells and Stromal Cells
  8. A Hypothesis
  9. Conclusions
  10. Disclosures
  11. References

The thymus undergoes progressive structural and functional decline with age, coinciding with increased circulating sex-steroid levels at puberty.48 By middle age most parenchymal tissue is replaced by fat, and in both mice and humans fewer T cells are produced and exported to secondary lymphoid organs. However, the thymus continues to generate new T cells even into old age.49, 50 In fact, active lymphocytic thymic tissue has been documented in adults up to 107 years of age.51 Under severe T cell depletion secondary to human immunodeficiency virus (HIV) infection, chemotherapy, or bone marrow transplant, an increase in thymic output (known as thymic rebound) becomes critical for long-term restoration of T cell homeostasis. For example, middle-aged women treated with autologous bone marrow transplants develop thymic hypertrophy and a resurgence of thymic T cell output, which contributes to the restoration of a wide T cell repertoire,52 although the intensity of thymic rebound declines with age. The mechanism driving thymic rebound is not completely understood, but one factor involved is IL-7.53

Both androgens and estrogen suppress thymic function.54, 55 Accordingly, castration reverses thymic atrophy and increases export of recent thymic emigrants to the periphery,56 whereas sex steroids inhibit thymic regeneration by promoting thymocyte apoptosis and arresting thymocyte/prelymphocyte differentiation.57 Restoration of thymic function after castration occurs in young58 as well as in very old rodents.59, 60

In accordance with the notion that estrogen deficiency induces a rebound in thymic function, ovx increases the thymic export of naïve T cells.61 Indeed, stimulated thymic T cell output accounts for ∼50% of the increase in the number of T cells in the periphery. Moreover, thymectomy decreases the bone loss induced by ovx by ∼50%, thus demonstrating that the thymus plays a previously unrecognized role in the pathogenesis of ovx-induced bone loss in mice.61 The remaining bone loss is a consequence of the peripheral expansion of naïve and memory T cells. This finding suggests the tantalizing hypothesis that estrogen deficiency–induced thymic rebound may be responsible for the exaggerated bone loss in young women undergoing surgical menopause62 or for the rapid bone loss characteristic of women in their first 5 to 7 years after natural menopause.63 Indeed, an age-related decrease in estrogen deficiency–induced thymic rebound could mitigate the stimulatory effects of sex steroid deprivation and explain why the rate of bone loss in postmenopausal women diminishes as aging progresses.63

Effects of Ovariectomy in the Bone Marrow

  1. Top of page
  2. Introduction
  3. T Cells and Ovariectomy Induced Bone Loss
  4. Estrogen Regulation of T Cell Activation and TNF Production
  5. T Cell Thymic Output and Bone Loss
  6. Effects of Ovariectomy in the Bone Marrow
  7. Interactions Between T Cells and Stromal Cells
  8. A Hypothesis
  9. Conclusions
  10. Disclosures
  11. References

The most upstream effects of ovx in the BM are to stimulate the production of reactive oxygen species (ROS) and to impair the generation of antioxidants.10, 19, 64, 65 In response to ovx, ROS are produced by most BM cells, including T cells.41 ROS play an important role in postmenopausal bone loss by generating a more oxidized bone microenvironment.66, 67 Multiple enzymatic pathways regulate the intracellular redox state through modulation of ROS levels.68 Ovx blunts the BM levels of glutathione (GSH), a critical ROS scavenger, and reduces expression of APE1/Ref-1 and Prx-1 proteins, which collectively limit the production of intracellular ROS.69

ROS have important direct effects on osteoblasts and osteoclasts; these effects have been addressed elsewhere.7, 70, 71 However, additional pivotal effects of ROS include expanding the pool of mature DCs that express the costimulatory molecule CD80, and increasing DC-mediated Ag presentation.19 Antioxidants potently inhibit DC differentiation and their ability to activate T cells,72, 73 in part by suppressing expression of MHC class II and costimulatory molecules in response to Ag.74 N-acetyl-cysteine (NAC), which acts as an intracellular scavenger of ROS by restoring intracellular concentrations of GSM, can block DC maturation75 and DC-mediated T cell activation.76

In vivo support for a role of ROS is provided by experiments demonstrating that administration of antioxidants prevents ovx-induced bone loss,19, 64, 70 while depletion of glutathione by buthionine sulfoximine (BSO), which inhibits glutathione synthesis, enhances bone loss.64 Bone loss caused by BSO has significant similarities to bone loss induced by estrogen deficiency, as both processes are TNF-dependent.77

A second, direct upstream effect of estrogen deficiency is to blunt BM levels of TGFβ,78 a powerful repressor of T cell activation. TGFβ acts as an immunosuppressant by inhibiting T cell activation and T cell production of inflammatory cytokines, including IFNγ. Demonstrating the relevance of the repressive effects of TGFβ on T cell function, mice with T cell–specific blockade of TGFβ signaling were found to be completely resistant to the bone-sparing effects of estrogen.16 Gain of function experiments confirmed that elevation of the systemic levels of TGFβ prevents ovx-induced bone loss and bone turnover.16

The key downstream mechanism by which ovx increases Ag presentation by macrophages is a stimulatory effect on the expression of the gene encoding Class II Transactivator (CIITA). The product of CIITA is a non-DNA binding factor induced by IFNγ that functions as a transcriptional coactivator at the MHC II promoter.79 Increased CIITA expression in macrophages results from ovx-mediated increases in IFNγ production by CD4+ T cells and the responsiveness of CIITA to IFNγ.46 This cytokine was initially described as an anti-osteoclastogenic cytokine because it is a potent inhibitor of osteoclastogenesis in vitro.80 The notion that IFNγ is an inhibitor of bone resorption was reinforced by the finding that silencing of IFNγR−/− signaling leads to a more rapid onset of collagen-induced arthritis and bone resorption81 as compared to WT controls, and by the report that IFNγ decreases serum calcium and osteoclastic bone resorption in nude mice.82, 83 Several mechanisms have been proposed to explain the anti-osteoclastogenic activity of IFNγ, including inhibition of RANKL signaling through the degradation of TNF receptor–associated factor 6 (TRAF6),80 stimulation of apoptosis mediated by Fas/FasL signals,84 and inhibition of RANK and c-Fms gene expression.85 However, the finding that IFNγ is an effective treatment for osteopetrosis both in humans86 and rodents87 demonstrates that the net effect of IFNγ in vivo is to stimulate osteoclastic bone resorption. In keeping with a net pro-resorptive effect of IFNγ in vivo are reports demonstrating that IFNγ−/− and IFNγR−/− mice are protected against ovx-induced bone loss.17, 46 Mice lacking IFNγ production are also protected against infection-induced alveolar bone loss,88 whereas in erosive tuberculoid leprosy and psoriatic arthritis IFNγ production correlates positively with tissue destruction.89 In addition, randomized controlled trials have shown that IFNγ does not prevent bone loss in patients with rheumatoid arthritis (RA),90 nor the bone-wasting effect of cyclosporine A.91 Finally, disruption of IFNγ signaling in vivo results in a strong and sustained inhibition of markers of osteoclastic activity.92 These opposing in vitro and in vivo effects of IFNγ are explained by the fact that IFNγ influences osteoclast formation via both direct and indirect effects.17 IFNγ directly blocks osteoclast formation through targeting of maturing osteoclast.93 However, IFNγ is also a potent inducer of antigen presentation and thus of T cell activation. Therefore, when IFNγ levels are increased in vivo, activated T cells secrete pro-osteoclastogenic factors and this activity offsets the anti-osteoclastogenic effects of IFNγ.17 It should also be mentioned that it is now recognized that IFNγ affects bone turnover by promoting the commitment of BM–SCs into the osteoblastic lineage and their differentiation into mature osteoblasts.94 Accordingly, treatment with IFNγ reverses ovx-induced bone loss by promoting bone formation.92

Another mechanism by which estrogen regulates T cell TNF production is by repressing the production of IL-7, a potent lymphopoietic cytokine and an inducer of bone destruction in vivo.95 Attesting to the relevance of this factor, IL-7 levels are significantly elevated following ovx,61, 96 and in vivo IL-7 blockade is effective in preventing ovx-induced bone destruction.96

The elevated BM levels of IL-7 contribute to the expansion of the T cell population in peripheral lymphoid organs through several mechanisms. First, IL-7 directly stimulates T cell proliferation.61 Second, IL-7 increases antigen presentation by upregulating the production of IFNγ. Third, IL-7 and TGFβ inversely regulate each other's production.97, 98 The reduction in TGFβ signaling characteristic of estrogen deficiency may serve to further stimulate IL-7 production, thus driving the cycle of osteoclastogenic cytokine production and bone wasting. In estrogen deficiency, IL-7 compounds bone loss by suppressing bone formation and thus uncoupling bone formation from resorption.

Interactions Between T Cells and Stromal Cells

  1. Top of page
  2. Introduction
  3. T Cells and Ovariectomy Induced Bone Loss
  4. Estrogen Regulation of T Cell Activation and TNF Production
  5. T Cell Thymic Output and Bone Loss
  6. Effects of Ovariectomy in the Bone Marrow
  7. Interactions Between T Cells and Stromal Cells
  8. A Hypothesis
  9. Conclusions
  10. Disclosures
  11. References

A clue that the crosstalk between T cells and SCs is relevant for ovx-induced bone loss was provided by the finding that activated T cells induce SC apoptosis via the Fas/Fas ligand pathway, a phenomenon which blunts the compensatory increase in bone formation that limits bone loss in ovx mice.22 More abundant information is available about the T cell/SC crosstalk driven by the CD40L/CD40 system. CD40L, also known as CD154, is a key surface ligand expressed on T cells.99 CD40L binds to CD40100 and several integrins.101, 102 CD40 is expressed on antigen-presenting cells, hemopoietic progenitors, and cells of the osteoblastic lineage.103 CD40L has been linked to postnatal skeletal maturation because T cells, through the CD40L/CD40 system, promote production of the anti-osteoclastogenic factor osteoprotegerin (OPG) by B cells.104 Consequently, CD40L-deficient mice attain a reduced peak bone volume due to exaggerated bone resorption.104 Low bone density has also been found in children affected by X-linked hyper–immunoglobulin M (IgM) syndrome, a condition in which CD40L production is impaired due to a mutation in the CD40L gene.105 However, mice lacking T cell–expressed CD40L are protected against parathyroid hormone (PTH)-induced bone loss,106 raising the possibility that CD40L may exert antiresorptive activities in unstimulated conditions, while promoting bone resorption under conditions of bone stress.

Studies with CD40L-null mice and with WT mice treated with the anti CD40L Ab MR-1 have revealed that silencing of CD40L completely prevents ovx-induced bone loss.18 A dual mechanism was shown to be involved. First, silencing of CD40L blocks the activation of T cells and the resulting production of TNF. Second, CD40L is required for ovx to increase the proliferation and the differentiation of SCs and their capacity to support osteoclast formation through enhanced production of macrophage colony-stimulating factor (M-CSF) and RANKL, and diminished secretion of OPG. Thus, a critical additional mechanism by which T cells dysregulate bone homeostasis in ovx mice is through a CD40L-mediated crosstalk between T cells and SCs that results in enhanced osteoclastogenesis and, to a lesser degree, enhanced osteoblastogenesis.

In summary, ovx increases the number of activated CD40L-expressing T cells that promote the expression of M-CSF and RANKL by SCs, and ovx downregulates the SC production of OPG. The net result is a significant increase in the rate of osteoclastogenesis. The interaction of CD40L with CD40 on SCs in the context of estrogen deficiency appears to override the protective effects of CD40/CD40L costimulation on basal B cell OPG production,104 distorting the balance of osteoclast formation in favor of bone loss.

An additional mechanism of T cells/SC crosstalk involving the novel regulator of bone mass delta-like 1/fetal antigen 1(DLK1/FA-1)107, 108 has recently been described.109 DLK1 encodes for a membrane-bound protein known as DLK1. This factor can be cleaved to generate a soluble protein known as FA-1. In physiologic conditions DLK1/FA-1 are produced by SCs, B cells, and T cells. These factors stimulate osteoclastogenesis and block osteoblastogenesis by inducing the production of TNF, IL-7, and other inflammatory cytokines by SCs. Ovx markedly increases the production of DLK1/FA-1 by activated CD4+ and CD8+ T cells, resulting in a further stimulation of the production of osteoclastogenic cytokines by SCs. Attesting to the relevance of DLK1/FA-1, DLK1 null mice are significantly protected against ovx-induced bone loss.109

A Hypothesis

  1. Top of page
  2. Introduction
  3. T Cells and Ovariectomy Induced Bone Loss
  4. Estrogen Regulation of T Cell Activation and TNF Production
  5. T Cell Thymic Output and Bone Loss
  6. Effects of Ovariectomy in the Bone Marrow
  7. Interactions Between T Cells and Stromal Cells
  8. A Hypothesis
  9. Conclusions
  10. Disclosures
  11. References

The hypothesis that the immune system is pivotal for the bone loss brought about by menopause remains to be demonstrated in humans. Meanwhile, it remains interesting to entertain the question of why the immune system in mice, and possibly women, is involved in the mechanism of ovx-induced bone loss. Clues may emerge by regarding postmenopausal bone loss as an unintended recapitulation of an event critical for reproduction, namely the need to stimulate bone resorption in the immediate postpartum period. This process is essential to meeting the markedly increased maternal demand for calcium brought about by lactation. The signal for this event is the acute drop in estrogen levels immediately postpartum. A second adaptation to the postpartum period is changes in breast physiology associated with lactation. Pregnancy is characterized by massive proliferation of the mammary epithelium and formation of lobulo-alveolar structures. These changes are orchestrated by progesterone and prolactin. There is now ample evidence that progesterone regulates breast development and breast adaptation to pregnancy by modulating the local production of RANKL and RANK.110–112 Thus, a system that is absolutely required for bone homeostasis is also a key regulator of lactation.112 The precipitous drop in progesterone levels in the postpartum period results in reduced breast tissue levels of RANK and RANKL, with the effect of finalizing the breasts for lactation. The concomitant increase in the RANKL/OPG ratio and TNF levels in the BM induced by declining estrogen levels accomplishes the goal of transferring calcium from the maternal skeleton to the breast milk, and then to the newborn. During pregnancy, a state of maternal tolerance to the fetus is induced by an estrogen-driven increase in the number of Tregs.113 A third adaptation to the end of pregnancy is the loss of such immune tolerance and the restoration of a normal immune reactivity. This goal is achieved through a decrease in the Treg population, resulting from the acute drop in placental steroid levels. However, a link between immune tolerance to the fetus and bone is provided by the observation that OPG is expressed by human gestational membranes.114 Thus, it is tempting to speculate that cessation of ovarian function induces bone loss through an adaptive immune response because natural selection has integrated these three key adaptations within the immune system to fulfill postpartum requirements (Fig. 2).

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Figure 2. Schematic representation of events taking place in the immediate postpartum period and of the involved cells and cytokines. The figure shows that estrogen and progesterone withdrawal are followed by a stimulation of bone resorption, breast adaptation to lactation, and termination of immune tolerance to the fetus. Activated T cells drive bone resorption by secreting TNF. A decrease in the number of Tregs induces the termination of the immune tolerance developed during pregnancy. The loss of placental OPG may contribute to the tolerance reversal and the increased bone resorption. A decrease in breast levels of RANKL and RANK induce the terminal preparation of the breast for lactation. Together with increased released of calcium from the skeleton, the morphological changes in the breast contribute to successful lactation, an event critical for reproduction. The data suggest the untested hypothesis that cessation of ovarian function induces bone loss through an adaptive immune response because natural selection has centralized key adaptations within the immune system to fulfill postpartum requirements.

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Conclusions

  1. Top of page
  2. Introduction
  3. T Cells and Ovariectomy Induced Bone Loss
  4. Estrogen Regulation of T Cell Activation and TNF Production
  5. T Cell Thymic Output and Bone Loss
  6. Effects of Ovariectomy in the Bone Marrow
  7. Interactions Between T Cells and Stromal Cells
  8. A Hypothesis
  9. Conclusions
  10. Disclosures
  11. References

The data reviewed above strongly support the hypothesis that the bone loss induced by estrogen deficiency is due to a complex interplay of hormones and cytokines that converge to disrupt the process of bone remodeling. Ovx upregulates T cell TNF production by increasing T cell activity in the BM, thymus, and the peripheral lymphoid organs. T cell precursors leave the BM and migrate to the thymus, where T cell differentiation, selection, and expansion take place, in large measure under the control of IL-7. Following release from the thymus, these new T cells home to peripheral lymphoid organs, including the BM itself. Ovx induces T cell activation in the BM in part by directly promoting antigen presentation, and in part via stimulation of IL-7 and IFNγ production and downregulation of TGFβ production. The net result of these actions is an increase in the number of TNF-producing T cells. The elevated levels of TNF increase RANKL-induced osteoclast formation. Estrogen deficiency also amplifies T cell activation and osteoclastogenesis by downregulating antioxidant pathways, leading to an upswing in ROS. The combined effect of IFNγ and ROS markedly enhances Ag presentation, amplifying T cell activation. T cells further stimulate RANKL and M-CSF production by SCs, through CD40L and DLK1/FA-1.

References

  1. Top of page
  2. Introduction
  3. T Cells and Ovariectomy Induced Bone Loss
  4. Estrogen Regulation of T Cell Activation and TNF Production
  5. T Cell Thymic Output and Bone Loss
  6. Effects of Ovariectomy in the Bone Marrow
  7. Interactions Between T Cells and Stromal Cells
  8. A Hypothesis
  9. Conclusions
  10. Disclosures
  11. References
  • 1
    Cenci S, Weitzmann MN, Roggia C, Namba N, Novack D, Woodring J, Pacifici R. Estrogen deficiency induces bone loss by enhancing T-cell production of TNF-alpha. J Clin Invest. 2000; 106(10): 122937.
  • 2
    Manolagas SC, Kousteni S, Jilka RL. Sex steroids and bone. Recent Prog Horm Res. 2002; 57: 385409.
  • 3
    Weitzmann MN, Pacifici R. Estrogen deficiency and bone loss: an inflammatory tale. J Clin Invest. 2006; 116(5): 118694.
  • 4
    Di Gregorio GB, Yamamoto M, Ali AA, Abe E, Roberson P, Manolagas SC, Jilka RL. Attenuation of the self-renewal of transit-amplifying osteoblast progenitors in the murine bone marrow by 17 beta-estradiol. J Clin Invest. 2001; 107(7): 80312.
  • 5
    Nakamura T, Imai Y, Matsumoto T, Sato S, Takeuchi K, Igarashi K, Harada Y, Azuma Y, Krust A, Yamamoto Y, Nishina H, Takeda S, Takayanagi H, Metzger D, Kanno J, Takaoka K, Martin TJ, Chambon P, Kato S. Estrogen prevents bone loss via estrogen receptor alpha and induction of Fas ligand in osteoclasts. Cell. 2007; 130(5): 81123.
  • 6
    Krum SA, Miranda-Carboni GA, Hauschka PV, Carroll JS, Lane TF, Freedman LP, Brown M. Estrogen protects bone by inducing Fas ligand in osteoblasts to regulate osteoclast survival. EMBO J. 2008; 27(3): 53545.
  • 7
    Martin-Millan M, Almeida M, Ambrogini E, Han L, Zhao H, Weinstein RS, Jilka RL, O'Brien CA, Manolagas SC. The estrogen receptor-alpha in osteoclasts mediates the protective effects of estrogens on cancellous but not cortical bone. Mol Endocrinol. 2010; 24(2): 32334.
  • 8
    Chen JR, Plotkin LI, Aguirre JI, Han L, Jilka RL, Kousteni S, Bellido T, Manolagas SC. Transient versus sustained phosphorylation and nuclear accumulation of ERKs underlie anti-versus pro-apoptotic effects of estrogens. J Biol Chem. 2005; 280(6): 46328.
  • 9
    Zaidi M. Skeletal remodeling in health and disease. Nat Med. 2007; 13(7): 791801.
  • 10
    Manolagas SC. From estrogen-centric to aging and oxidative stress: a revised perspective of the pathogenesis of osteoporosis. Endocr Rev. 2010; 31(3): 266300.
  • 11
    Jilka RL, Takahashi K, Munshi M, Williams DC, Roberson PK, Manolagas SC. Loss of estrogen upregulates osteoblastogenesis in the murine bone marrow. Evidence for autonomy from factors released during bone resorption. J Clin Invest. 1998; 101(9): 194250.
  • 12
    Kousteni S, Bellido T, Plotkin LI, O'Brien CA, Bodenner DL, Han L, Han K, DiGregorio GB, Katzenellenbogen JA, Katzenellenbogen BS, Roberson PK, Weinstein RS, Jilka RL, Manolagas SC. Nongenotropic, sex-nonspecific signaling through the estrogen or androgen receptors: dissociation from transcriptional activity. Cell. 2001; 104(5): 71930.
  • 13
    Kousteni S, Han L, Chen JR, Almeida M, Plotkin LI, Bellido T, Manolagas SC. Kinase-mediated regulation of common transcription factors accounts for the bone-protective effects of sex steroids. J Clin Invest. 2003; 111(11): 165164.
  • 14
    Kimble RB, Srivastava S, Ross FP, Matayoshi A, Pacifici R. Estrogen deficiency increases the ability of stromal cells to support murine osteoclastogenesis via an interleukin-1and tumor necrosis factor- mediated stimulation of macrophage colony-stimulating factor production. J Biol Chem. 1996; 271(46): 288907.
  • 15
    Roggia C, Gao Y, Cenci S, Weitzmann MN, Toraldo G, Isaia G, Pacifici R. Up-regulation of TNF-producing T cells in the bone marrow: a key mechanism by which estrogen deficiency induces bone loss in vivo. Proc Natl Acad Sci U S A. 2001; 98(24): 139605.
  • 16
    Gao Y, Qian WP, Dark K, Toraldo G, Lin AS, Guldberg RE, Flavell RA, Weitzmann MN, Pacifici R. Estrogen prevents bone loss through transforming growth factor beta signaling in T cells. Proc Natl Acad Sci U S A. 2004; 101(47): 1661823.
  • 17
    Gao Y, Grassi F, Ryan MR, Terauchi M, Page K, Yang X, Weitzmann MN, Pacifici R. IFN-gamma stimulates osteoclast formation and bone loss in vivo via antigen-driven T cell activation. J Clin Invest. 2007; 117(1): 12232.
  • 18
    Li JY, Tawfeek H, Bedi B, Yang X, Adams J, Gao KY, Zayzafoon M, Weitzmann MN, Pacifici R. Ovariectomy disregulates osteoblast and osteoclast formation through the T-cell receptor CD40 ligand. Proc Natl Acad Sci U S A. 2011; 108(2): 76873.
  • 19
    Grassi F, Tell G, Robbie-Ryan M, Gao Y, Terauchi M, Yang X, Romanello M, Jones DP, Weitzmann MN, Pacifici R. Oxidative stress causes bone loss in estrogen-deficient mice through enhanced bone marrow dendritic cell activation. Proc Natl Acad Sci U S A. 2007; 104(38): 1508792.
  • 20
    Moreland L, Bate G, Kirkpatrick P. Abatacept. Nat Rev Drug Discov. 2006; 5(3): 1856.
  • 21
    Ruderman EM, Pope RM. The evolving clinical profile of abatacept (CTLA4-Ig): a novel co-stimulatory modulator for the treatment of rheumatoid arthritis. Arthritis Res Ther. 2005; 7(Suppl 2): S215.
  • 22
    Yamaza T, Miura Y, Bi Y, Liu Y, Akiyama K, Sonoyama W, Patel V, Gutkind S, Young M, Gronthos S, Le A, Wang CY, Chen W, Shi S. Pharmacologic stem cell based intervention as a new approach to osteoporosis treatment in rodents. PLoS One. 2008; 3(7): e2615.
  • 23
    Lee SK, Kadono Y, Okada F, Jacquin C, Koczon-Jaremko B, Gronowicz G, Adams DJ, Aguila HL, Choi Y, Lorenzo JA. T lymphocyte-deficient mice lose trabecular bone mass with ovariectomy. J Bone Miner Res. 2006; 21(11): 17042.
  • 24
    Anginot A, Dacquin R, Mazzorana M, Jurdic P. Lymphocytes and the Dap12 adaptor are key regulators of osteoclast activation associated with gonadal failure. PLoS One. 2007; 2(7): e585.
  • 25
    Robbie-Ryan M, Pacifici R, Weitzmann MN. IL-7 drives T cell-mediated bone loss following ovariectomy. Ann N Y Acad Sci. 2006; 1068: 34851.
  • 26
    DeSelm C, Takahata Y, Chappel J, Khan T, Li K, Zou W, Teitelbaum S. IL-17 mediates estrogen-deficient osteoporosis. J Bone Miner Res. 2011; 27(Abstract FR0299).
  • 27
    Polanczyk MJ, Carson BD, Subramanian S, Afentoulis M, Vandenbark AA, Ziegler SF, Offner H. Cutting edge: estrogen drives expansion of the CD4+CD25+ regulatory T cell compartment. J Immunol. 2004; 173(4): 222730.
  • 28
    Sato K, Suematsu A, Okamoto K, Yamaguchi A, Morishita Y, Kadono Y, Tanaka S, Kodama T, Akira S, Iwakura Y, Cua DJ, Takayanagi H. Th17 functions as an osteoclastogenic helper T cell subset that links T cell activation and bone destruction. J Exp Med. 2006; 203(12): 267382.
  • 29
    Lelu K, Laffont S, Delpy L, Paulet PE, Perinat T, Tschanz SA, Pelletier L, Engelhardt B, Guery JC. Estrogen receptor alpha signaling in T lymphocytes is required for estradiol-mediated inhibition of Th1 and Th17 cell differentiation and protection against experimental autoimmune encephalomyelitis. J Immunol. 2011; 187(5): 238693.
  • 30
    Goswami J, Hernandez-Santos N, Zuniga LA, Gaffen SL. A bone-protective role for IL-17 receptor signaling in ovariectomy-induced bone loss. Eur J Immunol. 2009; 39(10): 28319.
  • 31
    Kim YG, Lee CK, Nah SS, Mun SH, Yoo B, Moon HB. Human CD4+CD25+ regulatory T cells inhibit the differentiation of osteoclasts from peripheral blood mononuclear cells. Biochem Biophys Res Commun. 357(4): 104652.
  • 32
    Yuan FL, Li X, Lu WG, Xu RS, Zhao YQ, Li CW, Li JP, Chen FH. 2010 Regulatory T cells as a potent target for controlling bone loss. Biochem Biophys Res Commun. 2007; 402(2): 1736.
  • 33
    Luo CY, Wang L, Sun C, Li DJ. Estrogen enhances the functions of CD4(+)CD25(+)Foxp3(+) regulatory T cells that suppress osteoclast differentiation and bone resorption in vitro. Cell Mol Immunol. 2011; 8(1): 508.
  • 34
    Tai P, Wang J, Jin H, Song X, Yan J, Kang Y, Zhao L, An X, Du X, Chen X, Wang S, Xia G, Wang B. Induction of regulatory T cells by physiological level estrogen. J Cell Physiol. 2008; 214(2): 45664.
  • 35
    Zaiss MM, Sarter K, Hess A, Engelke K, Bohm C, Nimmerjahn F, Voll R, Schett G, David JP. Increased bone density and resistance to ovariectomy-induced bone loss in FoxP3-transgenic mice based on impaired osteoclast differentiation. Arthritis Rheum. 2010; 62(8): 232838.
  • 36
    Surh CD, Sprent J. Homeostasis of naive and memory T cells. Immunity. 2008; 29(6): 84862.
  • 37
    Ernst B, Lee DS, Chang JM, Sprent J, Surh CD. The peptide ligands mediating positive selection in the thymus control T cell survival and homeostatic proliferation in the periphery. Immunity. 1999; 11(2): 17381.
  • 38
    Kau AL, Ahern PP, Griffin NW, Goodman AL, Gordon JI. Human nutrition, the gut microbiome and the immune system. Nature. 2011; 474(7351): 32736.
  • 39
    Sjögren K, Engdahl C, Lagerquist M, Bäckhed F, Ohlsson C. Absence of gut microbiota leads to increased bone mass associated with low serum serotonin levels. J Bone Miner Res. 2010; 25: Abstract 1170.
  • 40
    Tyagi AM, Srivastava K, Sharan K, Yadav D, Maurya R, Singh D. Daidzein prevents the increase in CD4CD28null T cells and B lymphopoesis in ovariectomized mice: a key mechanism for anti-osteoclastogenic effect. PLoS One. 2011; 6(6): e21216.
  • 41
    Tyagi AM, Srivastava K, Kureel J, Kumar A, Raghuvanshi A, Yadav D, Maurya R, Goel A, Singh D. Premature T cell senescence in Ovx mice is inhibited by repletion of estrogen and medicarpin: a possible mechanism for alleviating bone loss. Osteoporos Int. Epub 2011 May 12.
  • 42
    D'Amelio P, Grimaldi A, Di Bella S, Brianza SZ, Cristofaro MA, Tamone C, Giribaldi G, Ulliers D, Pescarmona GP, Isaia G. Estrogen deficiency increases osteoclastogenesis up-regulating T cells activity: a key mechanism in osteoporosis. Bone. 2008; 43(1): 92100.
  • 43
    Eghbali-Fatourechi G, Khosla S, Sanyal A, Boyle WJ, Lacey DL, Riggs BL. Role of RANK ligand in mediating increased bone resorption in early postmenopausal women. J Clin Invest. 2003; 111(8): 122130.
  • 44
    Ammann P, Rizzoli R, Bonjour JP, Bourrin S, Meyer JM, Vassalli P, Garcia I. Transgenic mice expressing soluble tumor necrosis factor-receptor are protected against bone loss caused by estrogen deficiency. J Clin Invest. 1997; 99: 1699703.
  • 45
    Kimble R, Bain S, Pacifici R. The functional block of TNF but not of IL-6 prevents bone loss in ovariectomized mice. J Bone Miner Res. 1997; 12: 93541.
  • 46
    Cenci S, Toraldo G, Weitzmann MN, Roggia C, Gao Y, Qian WP, Sierra O, Pacifici R. Estrogen deficiency induces bone loss by increasing T cell proliferation and lifespan through IFN-gamma-induced class II transactivator. Proc Natl Acad Sci U S A. 2003; 100(18): 1040510.
  • 47
    Grassi F, Pacifici R. Ovariectomy increases the formation of T cell niches at the resorption surfaces. J Bone Miner Res. 2005; 20(Suppl 1):Abs.
  • 48
    Haynes BF, Sempowski GD, Wells AF, Hale LP. The human thymus during aging. Immunol Res. 2000; 22(2–3): 25361.
  • 49
    Douek DC, Koup RA. Evidence for thymic function in the elderly. Vaccine. 2000; 18(16): 163841.
  • 50
    Jamieson BD, Douek DC, Killian S, Hultin LE, Scripture-Adams DD, Giorgi JV, Marelli D, Koup RA, Zack JA. Generation of functional thymocytes in the human adult. Immunity. 1999; 10(5): 56975.
  • 51
    Steinmann GG, Klaus B, Muller-Hermelink HK. The involution of the ageing human thymic epithelium is independent of puberty. A morphometric study. Scand J Immunol. 1985; 22(5): 56375.
  • 52
    Hakim FT, Memon SA, Cepeda R, Jones EC, Chow CK, Kasten-Sportes C, Odom J, Vance BA, Christensen BL, Mackall CL, Gress RE. Age-dependent incidence, time course, and consequences of thymic renewal in adults. J Clin Invest. 2005; 115(4): 9309.
  • 53
    Mackall CL, Fry TJ, Bare C, Morgan P, Galbraith A, Gress RE. IL-7 increases both thymic-dependent and thymic-independent T-cell regeneration after bone marrow transplantation. Blood. 2001; 97(5): 14917.
  • 54
    Leposavic G, Perisic M. Age-associated remodeling of thymopoiesis: role for gonadal hormones and catecholamines. Neuroimmunomodulation. 2008; 15(4–6): 290322.
  • 55
    Leposavic G, Perisic M, Kosec D, Arsenovic-Ranin N, Radojevic K, Stojic-Vukanic Z, Pilipovic I. Neonatal testosterone imprinting affects thymus development and leads to phenotypic rejuvenation and masculinization of the peripheral blood T-cell compartment in adult female rats. Brain Behav Immun. 2009; 23(2): 294304.
  • 56
    Utsuyama M, Hirokawa K. Hypertrophy of the thymus and restoration of immune functions in mice and rats by gonadectomy. Mech Ageing Dev. 1989; 47(3): 17585.
  • 57
    Okasha SA, Ryu S, Do Y, McKallip RJ, Nagarkatti M, Nagarkatti PS. Evidence for estradiol-induced apoptosis and dysregulated T cell maturation in the thymus. Toxicology. 2001; 163(1): 4962.
  • 58
    Roden AC, Moser MT, Tri SD, Mercader M, Kuntz SM, Dong H, Hurwitz AA, McKean DJ, Celis E, Leibovich BC, Allison JP, Kwon ED. Augmentation of T cell levels and responses induced by androgen deprivation. J Immunol. 2004; 173(10): 6098108.
  • 59
    Sutherland JS, Goldberg GL, Hammett MV, Uldrich AP, Berzins SP, Heng TS, Blazar BR, Millar JL, Malin MA, Chidgey AP, Boyd RL. Activation of thymic regeneration in mice and humans following androgen blockade. J Immunol. 2005; 175(4): 274153.
  • 60
    Perisić M, Arsenović-Ranin N, Pilipović I, Kosec D, Pesić V, Radojević K, Leposavić G. Role of ovarian hormones in age-associated thymic involution revisited. Immunobiology. 215(4): 27593.
  • 61
    Ryan MR, Shepherd R, Leavey JK, Gao Y, Grassi F, Schnell FJ, Qian WP, Kersh GJ, Weitzmann MN, Pacifici R. An IL-7-dependent rebound in thymic T cell output contributes to the bone loss induced by estrogen deficiency. Proc Natl Acad Sci U S A. 2005; 102(46): 1673540.
  • 62
    Hreshchyshyn MM, Hopkins A, Zylstra S, Anbar M. Effects of natural menopause, hysterectomy, and oophorectomy on lumbar spine and femoral neck bone densities. Obstet Gynecol. 1988; 72(4): 6318.
  • 63
    Riggs BL, Khosla S, Melton LJ 3rd. Sex steroids and the construction and conservation of the adult skeleton. Endocr Rev. 2002; 23(3): 279302.
  • 64
    Lean JM, Davies JT, Fuller K, Jagger CJ, Kirstein B, Partington GA, Urry ZL, Chambers TJ. A crucial role for thiol antioxidants in estrogen-deficiency bone loss. J Clin Invest. 2003; 112(6): 91523.
  • 65
    Muthusami S, Ramachandran I, Muthusamy B, Vasudevan G, Prabhu V, Subramaniam V, Jagadeesan A, Narasimhan S. Ovariectomy induces oxidative stress and impairs bone antioxidant system in adult rats. Clin Chim Acta. 2005; 360(1–2): 816.
  • 66
    Basu S, Michaelsson K, Olofsson H, Johansson S, Melhus H. Association between oxidative stress and bone mineral density. Biochem Biophys Res Commun. 2001; 288(1): 2759.
  • 67
    Maggio D, Barabani M, Pierandrei M, Polidori MC, Catani M, Mecocci P, Senin U, Pacifici R, Cherubini A. Marked decrease in plasma antioxidants in aged osteoporotic women: results of a cross-sectional study. J Clin Endocrinol Metab. 2003; 88(4): 15237.
  • 68
    Finkel T. Oxidant signals and oxidative stress. Curr Opin Cell Biol. 2003; 15(2): 24754.
  • 69
    Ozaki M, Suzuki S, Irani K. Redox factor-1/APE suppresses oxidative stress by inhibiting the rac1 GTPase. FASEB J. 2002; 16(8): 88990.
  • 70
    Almeida M, Han L, Martin-Millan M, Plotkin LI, Stewart SA, Roberson PK, Kousteni S, O'Brien CA, Bellido T, Parfitt AM, Weinstein RS, Jilka RL, Manolagas SC. Skeletal involution by age-associated oxidative stress and its acceleration by loss of sex steroids. J Biol Chem. 2007; 282(37): 2728597.
  • 71
    Almeida M, Martin-Millan M, Ambrogini E, Bradsher R 3rd, Han L, Chen XD, Roberson PK, Weinstein RS, O'Brien CA, Jilka RL, Manolagas SC. Estrogens attenuate oxidative stress and the differentiation and apoptosis of osteoblasts by DNA-binding-independent actions of the ERalpha. J Bone Miner Res. 2010; 25(4): 76981.
  • 72
    Mizuashi M, Ohtani T, Nakagawa S, Aiba S. Redox imbalance induced by contact sensitizers triggers the maturation of dendritic cells. J Invest Dermatol. 2005; 124(3): 57986.
  • 73
    Rutault K, Alderman C, Chain BM, Katz DR. Reactive oxygen species activate human peripheral blood dendritic cells. Free Radic Biol Med. 1999; 26(1–2): 2328.
  • 74
    Maemura K, Zheng Q, Wada T, Ozaki M, Takao S, Aikou T, Bulkley GB, Klein AS, Sun Z. Reactive oxygen species are essential mediators in antigen presentation by Kupffer cells. Immunol Cell Biol. 2005; 83(4): 33643.
  • 75
    Vosters O, Neve J, De Wit D, Willems F, Goldman M, Verhasselt V. Dendritic cells exposed to nacystelyn are refractory to maturation and promote the emergence of alloreactive regulatory t cells. Transplantation. 2003; 75(3): 3839.
  • 76
    Verhasselt V, Vanden Berghe W, Vanderheyde N, Willems F, Haegeman G, Goldman M. N-acetyl-L-cysteine inhibits primary human T cell responses at the dendritic cell level: association with NF-kappaB inhibition. J Immunol. 1999; 162(5): 256974.
  • 77
    Jagger CJ, Lean JM, Davies JT, Chambers TJ. Tumor necrosis factor-alpha mediates osteopenia caused by depletion of antioxidants. Endocrinology. 2005; 146(1): 1138.
  • 78
    Yang NN, Venugopalan M, Hardikar S, Glasebrook A. Identification of an estrogen response element activated by metabolites of 17b-estradiol and raloxifene. Science. 1996; 273: 12225.
  • 79
    Boss JM, Jensen PE. Transcriptional regulation of the MHC class II antigen presentation pathway. Curr Opin Immunol. 2003; 15(1): 10511.
  • 80
    Takayanagi H, Ogasawara K, Hida S, Chiba T, Murata S, Sato K, Takaoka A, Yokochi T, Oda H, Tanaka K, Nakamura K, Taniguchi T. T-cell-mediated regulation of osteoclastogenesis by signalling cross- talk between RANKL and IFN-gamma. Nature. 2000; 408(6812): 6005.
  • 81
    Vermeire K, Heremans H, Vandeputte M, Huang S, Billiau A, Matthys P. Accelerated collagen-induced arthritis in IFN-gamma receptor-deficient mice. J Immunol. 1997; 158(11): 550713.
  • 82
    Sato K, Satoh T, Shizume K, Yamakawa Y, Ono Y, Demura H, Akatsu T, Takahashi N, Suda T. Prolonged decrease of serum calcium concentration by murine gamma-interferon in hypercalcemic, human tumor (EC-GI)-bearing nude mice. Cancer Res. 1992; 52(2): 4449.
  • 83
    Tohkin M, Kakudo S, Kasai H, Arita H. Comparative study of inhibitory effects by murine interferon gamma and a new bisphosphonate (alendronate) in hypercalcemic, nude mice bearing human tumor (LJC-1-JCK). Cancer Immunol Immunother. 1994; 39(3): 15560.
  • 84
    Kohara H, Kitaura H, Fujimura Y, Yoshimatsu M, Morita Y, Eguchi T, Masuyama R, Yoshida N. IFN-gamma directly inhibits TNF-alpha-induced osteoclastogenesis in vitro and in vivo and induces apoptosis mediated by Fas/Fas ligand interactions. Immunol Lett. 2011; 137(1–2): 5361.
  • 85
    Ji JD, Park-Min KH, Shen Z, Fajardo RJ, Goldring SR, McHugh KP, Ivashkiv LB. Inhibition of RANK expression and osteoclastogenesis by TLRs and IFN-gamma in human osteoclast precursors. J Immunol. 2009; 183(11): 722333.
  • 86
    Key LL Jr, Rodriguiz RM, Willi SM, Wright NM, Hatcher HC, Eyre DR, Cure JK, Griffin PP, Ries WL. Long-term treatment of osteopetrosis with recombinant human interferon gamma. N Engl J Med. 1995; 332(24): 15949.
  • 87
    Rodriguiz RM, Key LL Jr, Ries WL. Combination macrophage-colony stimulating factor and interferon-gamma administration ameliorates the osteopetrotic condition in microphthalmic (mi/mi) mice. Pediatr Res. 1993; 33(4 Pt 1): 3849.
  • 88
    Baker PJ, Dixon M, Evans RT, Dufour L, Johnson E, Roopenian DC. CD4(+) T cells and the proinflammatory cytokines gamma interferon and interleukin-6 contribute to alveolar bone loss in mice. Infect Immun. 1999; 67(6): 28049.
  • 89
    Arnoldi J, Gerdes J, Flad HD. Immunohistologic assessment of cytokine production of infiltrating cells in various forms of leprosy. Am J Pathol. 1990; 137(4): 74953.
  • 90
    Cannon GW, Pincus SH, Emkey RD, Denes A, Cohen SA, Wolfe F, Saway PA, Jaffer AM, Weaver AL, Cogen L, et al. Double-blind trial of recombinant gamma-interferon versus placebo in the treatment of rheumatoid arthritis. Arthritis Rheum. 1989; 32(8): 96473.
  • 91
    Mann GN, Jacobs TW, Buchinsky FJ, Armstrong EC, Li M, Ke HZ, Ma YF, Jee WS, Epstein S. Interferon-gamma causes loss of bone volume in vivo and fails to ameliorate cyclosporin A-induced osteopenia. Endocrinology. 1994; 135: 107783.
  • 92
    Duque G, Huang DC, Dion N, Macoritto M, Rivas D, Li W, Yang XF, Li J, Lian J, Marino FT, Barralet J, Lascau V, Deschenes C, Ste-Marie LG, Kremer R. Interferon-gamma plays a role in bone formation in vivo and rescues osteoporosis in ovariectomized mice. J Bone Miner Res. 2011; 26(7): 147283.
  • 93
    Takayanagi H, Kim S, Taniguchi T. Signaling crosstalk between RANKL and interferons in osteoclast differentiation. Arthritis Res. 2002; 4(Suppl 3): S22732.
  • 94
    Duque G, Huang DC, Macoritto M, Rivas D, Yang XF, Ste-Marie LG, Kremer R. Autocrine regulation of interferon gamma in mesenchymal stem cells plays a role in early osteoblastogenesis. Stem Cells. 2009; 27(3): 5508.
  • 95
    Miyaura C, Onoe Y, Inada M, Maki K, Ikuta K, Ito M, Suda T. Increased B-lymphopoiesis by interleukin 7 induces bone loss in mice with intact ovarian function: similarity to estrogen deficiency. Proc Natl Acad Sci U S A. 1997; 19: 93605.
  • 96
    Weitzmann MN, Roggia C, Toraldo G, Weitzmann L, Pacifici R. Increased production of IL-7 uncouples bone formation from bone resorption during estrogen deficiency. J Clin Invest. 2002; 110(11): 164350.
  • 97
    Huang M, Sharma S, Zhu LX, Keane MP, Luo J, Zhang L, Burdick MD, Lin YQ, Dohadwala M, Gardner B, Batra RK, Strieter RM, Dubinett SM. IL-7 inhibits fibroblast TGF-beta production and signaling in pulmonary fibrosis. J Clin Invest. 2002; 109(7): 9317.
  • 98
    Dubinett SM, Huang M, Dhanani S, Economou JS, Wang J, Lee P, Sharma S, Dougherty GJ, McBride WH. Down-regulation of murine fibrosarcoma transforming growth factor-beta 1 expression by interleukin 7. J Natl Cancer Inst. 1995; 87(8): 5937.
  • 99
    Grewal IS, Flavell RA. CD40 and CD154 in cell-mediated immunity. Annu Rev Immunol. 1998; 16: 11135.
  • 100
    Quezada SA, Jarvinen LZ, Lind EF, Noelle RJ. CD40/CD154 interactions at the interface of tolerance and immunity. Annu Rev Immunol. 2004; 22: 30728.
  • 101
    Andre P, Prasad KS, Denis CV, He M, Papalia JM, Hynes RO, Phillips DR, Wagner DD. CD40L stabilizes arterial thrombi by a beta3 integrin–dependent mechanism. Nat Med. 2002; 8(3): 24752.
  • 102
    Leveille C, Bouillon M, Guo W, Bolduc J, Sharif-Askari E, El-Fakhry Y, Reyes-Moreno C, Lapointe R, Merhi Y, Wilkins JA, Mourad W. CD40 ligand binds to alpha5beta1 integrin and triggers cell signaling. J Biol Chem. 2007; 282(8): 514351.
  • 103
    Ahuja SS, Zhao S, Bellido T, Plotkin LI, Jimenez F, Bonewald LF. CD40 ligand blocks apoptosis induced by tumor necrosis factor alpha, glucocorticoids, and etoposide in osteoblasts and the osteocyte-like cell line murine long bone osteocyte-Y4. Endocrinology. 2003; 144(5): 17619.
  • 104
    Li Y, Toraldo G, Li A, Yang X, Zhang H, Qian WP, Weitzmann MN. B cells and T cells are critical for the preservation of bone homeostasis and attainment of peak bone mass in vivo. Blood. 2007; 109(9): 383948.
  • 105
    Lopez-Granados E, Temmerman ST, Wu L, Reynolds JC, Follmann D, Liu S, Nelson DL, Rauch F, Jain A. Osteopenia in X-linked hyper-IgM syndrome reveals a regulatory role for CD40 ligand in osteoclastogenesis. Proc Natl Acad Sci U S A. 2007; 104(12): 505661.
  • 106
    Gao Y, Wu X, Terauchi M, Li JY, Grassi F, Galley S, Yang X, Weitzmann MN, Pacifici R. T cells potentiate PTH-induced cortical bone loss through CD40L signaling. Cell Metab. 2008; 8(2): 13245.
  • 107
    Abdallah BM, Ding M, Jensen CH, Ditzel N, Flyvbjerg A, Jensen TG, Dagnaes-Hansen F, Gasser JA, Kassem M. Dlk1/FA1 is a novel endocrine regulator of bone and fat mass and its serum level is modulated by growth hormone. Endocrinology. 2007; 148(7): 311121.
  • 108
    Abdallah BM, Boissy P, Tan Q, Dahlgaard J, Traustadottir GA, Kupisiewicz K, Laborda J, Delaisse JM, Kassem M. dlk1/FA1 regulates the function of human bone marrow mesenchymal stem cells by modulating gene expression of pro-inflammatory cytokines and immune response-related factors. J Biol Chem. 2007; 282(10): 733951.
  • 109
    Abdallah BM, Ditzel N, Mahmood A, Isa A, Traustadottir GA, Schilling AF, Ruiz-Hidalgo MJ, Laborda J, Amling M, Kassem M. DLK1 is a novel regulator of bone mass that mediates estrogen deficiency-induced bone loss in mice. J Bone Miner Res. 2011; 26(7): 145771.
  • 110
    Fata JE, Kong YY, Li J, Sasaki T, Irie-Sasaki J, Moorehead RA, Elliott R, Scully S, Voura EB, Lacey DL, Boyle WJ, Khokha R, Penninger JM. The osteoclast differentiation factor osteoprotegerin-ligand is essential for mammary gland development. Cell. 2000; 103(1): 4150.
  • 111
    Mulac-Jericevic B, Lydon JP, DeMayo FJ, Conneely OM. Defective mammary gland morphogenesis in mice lacking the progesterone receptor B isoform. Proc Natl Acad Sci U S A. 2003; 100(17): 97449.
  • 112
    Schramek D, Sigl V, Penninger JM. RANKL and RANK in sex hormone-induced breast cancer and breast cancer metastasis. Trends Endocrinol Metab. 2011; 22(5): 18894.
  • 113
    Aluvihare VR, Kallikourdis M, Betz AG. Regulatory T cells mediate maternal tolerance to the fetus. Nat Immunol. 2004; 5(3): 26671.
  • 114
    Lonergan M, Aponso D, Marvin KW, Helliwell RJ, Sato TA, Mitchell MD, Chaiwaropongsa T, Romero R, Keelan JA. Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL), TRAIL receptors, and the soluble receptor osteoprotegerin in human gestational membranes and amniotic fluid during pregnancy and labor at term and preterm. J Clin Endocrinol Metab. 2003; 88(8): 383544.