Molecular mechanic sum of the regulation and dies regulation of bone homeostasis and Development of therapeutic approaches

为了阐明维持骨骼系统结构和功能稳态的调控机制，阐明骨骼稳态系统破坏导致疾病发生的分子机制，本研究主要从以下三个方面展开：(1)阐明骨形成机制和骨形成受损引起疾病的发病机制；我们已经确定ΔFosB是骨机械负荷的早期反应基因（JBMR'04）。快速诱导ΔFosB与JunD形成异源二聚体，并刺激其靶基因。IL-11是靶基因之一，ΔFosB/JunD异二聚体与IL-11基因启动子上的AP-1位点结合。我们还发现，在没有BMP-2刺激的情况下，机械刺激通过激活PECd增强了Smadl的磷酸化，磷酸化的Smadl与IL-11基因上的ΔFosB/JunD形成复合物，促进了I-11基因的完全转录激活。（2…更多）阐明多发性骨髓瘤破坏性骨病变的致病机制：我们之前发现多发性骨髓瘤通过产生MIP-1（一种C-C趋化因子）来增强破骨细胞骨吸收。（Blood '02, '04）在本研究中，我们发现骨髓瘤细胞组成性地分泌sFRP-2，一种Wnt信号抑制剂，强烈抑制成骨细胞分化。此外，TGF-β还能抑制成骨细胞的终末分化。由于TGF-β通过增强骨吸收从骨基质中释放，这些结果表明骨髓瘤-骨微环境对骨形成有很强的抑制作用。添加TGF-β拮抗剂后，骨髓瘤细胞的成骨抑制得到改善，骨髓瘤细胞末梢成骨分化的恢复明显抑制骨髓瘤细胞的生长。这些结果与骨髓瘤-骨微环境存在恶性循环的观点一致，骨髓瘤细胞增强骨吸收并抑制骨形成以延长骨破坏，骨髓瘤细胞衍生的sFRP-2以及骨基质通过增强骨吸收释放的TGF-β共同抑制骨形成。此外，抑制成骨细胞成熟为骨髓瘤细胞生长提供了适宜的环境，可称为“骨髓瘤生态位”。Wnt抑制剂和TGF-β拮抗剂可以成为开发新治疗方式的靶点。(3)阐明由骨骼循环紊乱引起的疾病的致病机制：我们之前报道过，糖皮质激素过量通过内皮中超氧化物的增加产生一氧化氮的减少来抑制内皮依赖性血管舒张反应（Circ Res '03）。我们进一步发现糖皮质激素过量显著抑制尿NOx排泄和主动脉eNOS表达。此外，匹伐他汀可拮抗糖皮质激素的作用。这些观察结果支持了我们的假设，即糖皮质激素过量导致股骨颈微血管循环障碍，这在股骨颈特发性骨坏死的发展中起重要作用。他汀类药物具有高的全身血液水平，可以成为预防超氧诱导的循环障碍的新候选药物。少

In order to clarify the regulatory mechanisms for maintaining structural and functional homeostasis of skeletal system, as well as to elucidate the molecular mechanisms for the development of disorders caused by disruption of the skeletal homeostatic system, the present study was conducted in the following three areas :(1) Elucidation of the mechanism of bone formation and the pathogenesis of disorders due to impaired bone formation: We have identified ΔFosB as an early response gene to mechanical load to the bone (JBMR'04). Rapidly induced ΔFosB forms heterodimers with JunD), and stimulated its target genes. IL-11 is one of the target genes, and ΔFosB /JunD heterodimer binds to a putative AP-1 site on the IL-11 gene promoter We also found that echanical stimuli enhances phosphorylation of Smadl via an activation of PECd without stimulation by BMP-2, and that phosphorylated Smadl forms complex with ΔFosB/JunD on the IL-11 gene promote fir full transcriptional activation of I-11 gene.(2 … More ) Elucidation of the pathogenic mechanisms 'for the destructive bone lesions by multiple myeloma: We have previously found that multiple myeloma enhances osteoclastic bone resorption via the production of MIP-1, a C-C chemokine. (Blood '02, '04) In the present study, we found that myeloma cells constitutively secrete sFRP-2, an inhibitor of Wnt signaling, which strongly inhibits osteoblast differentiation. In addition, TGF-β also suppressed terminal differentiation of osteoblasts. Because TGF-β is released from bone matrix by enhanced bone resorption, these results imply that there is a strong suppression ofbone formation in myeloma-bone microenvironment. If a TGF-β antagonist was added, the suppressed bone formation by myeloma cells was ameliorated, and the restoration of terminal osteoblastic differentiation markedly suppressed myeloma cell growth. These results are consistent with the notion that there is a vicious cycle in myeloma-bone microenvironment, in which myeloma cells enhance bone resorption and suppress bone formation to extend bone destruction, and myeloma cell-derived sFRP-2 as well as TGF-β released from the bone matrix by the enhanced bone resorption co-operatively suppress bone formation. Furthermore, the suppression of osteoblast maturation provides a suitable environment for myeloma cell growth which can be called "myeloma niche". Wnt inhibitors and TGF-β antagonists can be targets for the development of new therapeutic modalities.(3) Elucidation of the pathogenic mechanisms for disorders caused by disturbances in skeletal circulation: We have previously reported that glucocorticoid excess suppresses endothelium-dependent vasodilatory responses via a reduction in NO by an enhanced production of superoxides in the endothelium (Circ Res '03). We further found that glucocorticoid excess markedly suppresses urinary NOx excretion and aortic eNOS expression. In addition, pitavastatin antagonizes the effects of glucocorticoids. These observations support our hypothesis that glucocorticoid excess causes circulatory disturbances in microvessels of the femoral neck, which plays an important role in the development of idiopathic osteonecrosis of the femoral neck. Statins with high systemic blood levels can be a new candidate for the prevention of superoxide-induced circulatory disturbances. Less