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Mechanism Underlying Post-menopausal Osteoporosis: HIF1α is Required for Osteoclast Activation by Estrogen Deficiency
2015 Volume 64 Issue 3 Pages 44-47
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2015 Volume 64 Issue 3 Pages 44-47
The aging of the population worldwide has sharply increased the number of post-menopausal osteoporosis patients. Bone fragility caused by osteoporosis often results in fractures; therefore, controlling osteoporosis is crucial to prevent such injuries. To date, various drugs to treat osteoporosis have been developed and launched; however, the molecular mechanisms underlying post-menopausal osteoporosis have not been fully elucidated, and additional factors that could be targeted to treat patients remain to be characterized. Recently, hypoxia inducible factor 1 alpha (HIF1α) was identified as essential for osteoclast activation, an activity that promotes bone loss following menopausal estrogen deficiency. Although osteoclasts, which are located in hypoxic regions of the bone surface, express HIF1α mRNA, in pre-menopausal conditions the presence of estrogen decreases HIF1α protein levels in these cells. In menopausal conditions, however, estrogen deficiency allows HIF1α protein to accumulate in osteoclasts, leading to osteoclast activation and bone loss. Osteoclast-specific conditional HIF1α inactivation protects mice from estrogen deficiency-induced osteoclast activation and bone loss, as does systemic administration of a HIF1α inhibitor. Therefore, HIF1α represents a potential therapeutic target to prevent osteoclast activation and bone loss in post-menopausal patients.
Bone homeostasis requires a delicate balance of activity between bone-resorbing osteoclasts and bone-forming osteoblasts (Fig. 1). In women from age 20 years to pre-menopause, these activities are mutually regulated, keeping bone volume stable. However, estrogen deficiency occurring at menopause activates both osteoclasts and osteoblasts, with activation of the former being dominant, resulting in decrease of bone mass (Fig. 1). Therefore, osteoclasts are considered targets when devising treatment strategies for post-menopausal osteoporosis patients. Indeed, various osteoclast-inhibiting agents reportedly increase bone mass and prevent fragility fractures to a significantly greater extent than placebos.1,2,3,4,5 However, the mechanisms underlying osteoclast activation induced by estrogen deficiency remain undefined.
Bone homeostasis is regulated by osteoclast and osteoblast activities.
In pre-menopausal conditions, osteoclasts and osteoblasts work in concert to keep bone volume stable (left panel). However, post-menopausally, osteoclast activities outweigh those of osteoblasts, decreasing bone volume (right panel).
Because osteoclasts are activated in estrogen-deficient conditions, administration of estrogen itself could block their activation and subsequent bone loss.6 However, prolonged estrogen administration to post-menopausal patients could stimulate mammary gland or uterine tumor development or promote venous thrombosis.7 Thus, safer osteoclast-specific inhibitors have been sought.
Manipulation of osteoclast fusion has been considered one way to inhibit osteoclast activity, because fusion of mono-nuclear osteoclasts reportedly promotes the multi-nucleation required to remodel the osteoclast cytoskeleton via the creation of ruffled borders or sealing zones, leading to bone resorption. Among the factors that promote osteoclast cell–cell fusion,8 we identified two: the dendritic cell-specific transmembrane protein (DC-STAMP) and the osteoclast stimulatory transmembrane protein (OC-STAMP).9,10 We found that, both in vivo and in vitro, deficiency in either protein completely abrogated osteoclast cell–cell fusion, although formation of tartrate-resistant acid phosphatase-positive mono-nuclear osteoclasts remained unchanged.9,10 These observations indicate that both DC-STAMP and OC-STAMP are essential for osteoclast fusion but not for osteoclast differentiation (Fig. 2). We then observed that osteoclast bone-resorbing activity was indeed significantly down-regulated in DC-STAMP knockout mice and in OC-STAMP knockout mice, both of which exhibited mono-nuclear osteoclasts rather than their multi-nuclear counterparts typically seen in wild-type mice. However, levels of bone volume increase were limited in both DC-STAMP knockout mice and OC-STAMP knockout mice.9,10 Consequently, we sought other targets that might function to increase bone mass more robustly.
Generation of multi-nuclear osteoclasts.
Osteoclasts and macrophages are derived from common precursor cells in the presence of macrophage colony-stimulating factor (M-CSF), a cytokine for macrophage differentiation, and receptor activator of nuclear factor kappa B ligand (RANKL) and M-CSF alone, respectively. Mono-nuclear osteoclasts are generated first, and then multi-nuclear osteoclasts form by fusion of mono-nuclear osteoclasts via DC-STAMP and OC-STAMP. Although mono-nuclear osteoclasts can resorb bone, resorption activity is significantly elevated in multi-nucleated cells.
Osteoclasts localize to bone surfaces that are very hypoxic.11 We found that osteoclasts express mRNA encoding hypoxia inducible factor 1 alpha (HIF1α), a transcription factor regulated by hypoxia.12 Interestingly, HIF1α protein is detected in osteoclasts in ovariectomized (OVX) but not in sham-operated mice,12 suggesting that osteoclast-specific HIF1α protein is suppressed by estrogen.
In vitro, HIF1α protein is found in osteoclasts cultured under hypoxia but not in cells cultured in normoxic conditions.12 Interestingly, HIF1α protein is suppressed in osteoclasts by estrogen derivative estradiol (E2), even under hypoxia, without affecting HIF1α mRNA levels, supporting the idea that HIF1α protein in osteoclasts is normally suppressed by estrogen (Fig. 3). Based on this model, estrogen deficiency would promote HIF1α protein accumulation in osteoclasts, leading to their activation and subsequent bone loss (Fig. 3).
Interaction of hypoxia and estrogen, a mechanism regulating osteoclast activities and bone mass.
Osteoclasts localize at the bone surface in hypoxic regions. Osteoclasts express HIF1α mRNA; however, in estrogen-sufficient pre-menopausal conditions, HIF1α protein is continuously suppressed by the presence of ovary-derived estrogen. Post-menopausally, that suppression is absent, allowing HIF1α protein to accumulate and leading to osteoclast activation and subsequent osteoporosis.
To determine the consequences of HIF1α protein accumulation in osteoclasts in these conditions, we generated osteoclast-specific HIF1α conditional knockout mice12 and found that they were resistant to OVX-induced bone loss. Similarly, in wild-type mice, systemic administration of a HIF1α inhibitor completely abrogated OVX-induced osteoclast activation and bone loss.12 These results suggest that HIF1α could serve as a therapeutic target to prevent osteoclast activation and bone loss in this group of patients. At present, interactions between cell–cell fusion and HIF1α expression are unclear, and the regulation by HIF1α of DC-STAMP or OC-STAMP has not been reported. Because expression of both DC-STAMP and OC-STAMP is regulated by nuclear factor of activated T cells 1,10,13 a transcription factor essential for osteoclastogenesis, cell–cell fusion and HIF1α activity are likely independent.
Not only does aberrant osteoclast activity cause osteoporosis, but osteoclasts in cooperation with osteoblasts are required for normal bone turnover. Thus, inhibition of osteoclasts beyond physiological levels could severely suppress bone turnover. However, our evidence suggests that targeting the osteoclast factors pathologically activated by estrogen deficiency in post-menopausal women is not likely to interfere with physiological bone turnover: we found that systemic administration of a HIF1α inhibitor to sham-operated mice did not inhibit osteoclast activity, likely because HIF1α protein is already sufficiently suppressed by estrogen in normal conditions. HIF1α is reportedly essential for angiogenesis coupled to osteoblastogenesis in bone14,15; however, we found that systemic HIF1α inhibition did not result in decreased bone mass in sham-operated animals in estrogen-sufficient conditions.12 Taken together, we propose that HIF1α is a suitable therapeutic target to inhibit pathological but not physiological osteoclast activities and to maintain bone turnover at levels similar to those in pre-menopausal conditions in post-menopausal osteoporosis patients.
The author has no conflicts of interest to report.
