Wang Chuandong, Shan Shengzhou, Wang Chenglong, Wang Jing, Li Jiao, Hu Guoli, Dai Kerong, Li Qingfeng, Zhang Xiaoling
Department of Orthopedic Surgery, Xin Hua Hospital Affiliated to Shanghai Jiao Tong University School of Medicine (SJTUSM), Shanghai, China.
Department of Plastic & Reconstructive Surgery, Shanghai Ninth People's Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai 200011, China.
Exp Cell Res. 2017 Mar 15;352(2):346-356. doi: 10.1016/j.yexcr.2017.02.021. Epub 2017 Feb 16.
Mechanical unloading leads to bone loss and disuse osteoporosis partly due to impaired osteoblastogenesis of bone marrow stromal cells (BMSCs). However, the underlying molecular mechanisms of this phenomenon are not fully understood. In this study, we demonstrated that cyclic mechanical stretch (CMS) promotes osteoblastogenesis of BMSCs both in vivo and in vitro. Besides, we found that Hedgehog (Hh) signaling pathway was activated in this process. Inhibition of which by either knockdown of Sonic hedgehog (Shh) or treating BMSCs with Hh inhibitors attenuated the osteogenic effect of CMS on BMSCs, suggesting that Hh signaling pathway acts as an endogenous mediator of mechanical stimuli on BMSCs. Furthermore, we demonstrated that Shh expression level was regulated by DNA methylation mechanism. Chromatin Immunoprecipitation (ChIP) assay showed that DNA methyltransferase 3b (Dnmt3b) binds to Shh gene promoter, leading to DNA hypermethylation in mechanical unloading BMSCs. However, mechanical stimulation down-regulates the protein level of Dnmt3b, results in DNA demethylation and Shh expression. More importantly, we found that inhibition of Dnmt3b partly rescued bone loss in HU mice by mechanical unloading. Our results demonstrate, for the first time, that mechanical stimulation regulates osteoblastic genes expression via direct regulation of Dnmt3b, and the therapeutic inhibition of Dnmt3b may be an efficient strategy for enhancing bone formation under mechanical unloading.
机械卸载会导致骨质流失和废用性骨质疏松，部分原因是骨髓间充质干细胞（BMSCs）的成骨细胞生成受损。然而，这一现象的潜在分子机制尚未完全明确。在本研究中，我们证明了周期性机械拉伸（CMS）在体内和体外均能促进BMSCs的成骨细胞生成。此外，我们发现刺猬信号通路（Hh）在此过程中被激活。通过敲低音猬因子（Shh）或用Hh抑制剂处理BMSCs来抑制该通路，会减弱CMS对BMSCs的成骨作用，这表明Hh信号通路是机械刺激作用于BMSCs的内源性介质。此外，我们证明了Shh表达水平受DNA甲基化机制调控。染色质免疫沉淀（ChIP）分析表明，DNA甲基转移酶3b（Dnmt3b）与Shh基因启动子结合，导致机械卸载的BMSCs中DNA高度甲基化。然而，机械刺激会下调Dnmt3b的蛋白水平，导致DNA去甲基化和Shh表达。更重要的是，我们发现抑制Dnmt3b可部分挽救机械卸载的HU小鼠的骨质流失。我们的结果首次表明，机械刺激通过直接调控Dnmt3b来调节成骨基因的表达，对Dnmt3b的治疗性抑制可能是增强机械卸载状态下骨形成的有效策略。
