miR-133a通过调节Runx2对糖尿病骨质疏松的影响及干预研究

Diabetic osteoporosis (DOP) is a systemic metabolic bone disease with diabetes, which its mechanism has not been fully elucidated. Runx2 promotes osteoblast (OB) differentiation, and miR-133a is closely related to OB differentiation. Our previous studies found that miR-133a downregulated Runx2 protein, whereas, miR-133a inhibition upregulated Runx2 protein. Interestingly, in vivo, miR-133a expression was negative correlated with bone mineral density (BMD). Importantly，ibandronate(IB) decreased miR-133a expression in concentration-dependent manner in vitro. Hereby, we speculated that IB raised Runx2 expression, promoted OB differentiation and osteogenesis by inhibiting miR-133a expression. In this study, osteogenesis disorder was as a center-link of DOP, miR-133a and Runx2 was as a breakthrough point in osteogenesis network, and IB was as the main intervention measures. Real-time RT-PCR, cell transfection, luciferase reporter gene assays, western blot and confocal microscope were used, respectively. This study will further elucidate new mechanism of IB anti-osteoporosis, and whether miR-133a was one of new targets with IB in vivo. Expected results of the research will greatly expand the pathogenesis of DOP and provide new theoretical basis for development of a new anti-osteoporosis drug as miRNAs targets in the future.

糖尿病性骨质疏松症(DOP)是DM的一种全身性代谢性骨病，其机制尚未明。Runx2促进成骨细胞（OB）分化，miＲ-133a与OB分化密切相关。前期发现，miR -133a过表达能下调Runx2蛋白，抑制miＲ-133a则上调Runx2蛋白； miR-133a表达与BMD成负相关；双膦酸盐(IB)能浓度依耐性下调OB中miR-133表达。故推测IB通过抑制miR-133a表达，上调Runx2表达，促进OB分化。本课题针对DOP发病中心环节——骨形成障碍，以骨形成调控网络中的重要成分miR-133a和Runx2为切入点，以IB为干预措施，采用RT-PCR、WB，细胞转染、荧光素酶报告基因检测等技术，进一步阐明miR-133a能否成为IB新的作用靶点和以miR-133a为靶点的抗骨松效应。预期成果将极大拓展DOP发病机制，为将来研发以miRNA为靶点的新型抗骨松药物提供新的理论依据。

糖尿病性骨质疏松症(DOP)是DM的一种全身性代谢性骨病，其机制尚未明确。miR-133a与成骨细胞（OB）的分化密切相关，双膦酸盐类药物能浓度依耐性下调OB中miR-133表达。故推测双膦酸盐类药物通过抑制miR-133a表达，上调Runx2表达，促进OB分化。本课题针对DOP发病中心环节——骨形成障碍，以骨形成调控网络中的重要成分miR-133a和Runx2为切入点，以IB为干预措施，通过动物及细胞实验，采用实时荧光定量PCR、免疫蛋白印记，细胞转染等技术，体外及体内实验均显示：1.在糖尿病性骨质疏松症大鼠的股骨组织及高糖环境下培养的成骨细胞中，调控成骨细胞分化的 miR-133a表达增高，可引起 Runx2、Osterix、OCN 等成骨相关标志物的 mRNA 和蛋白的表达均降低，成骨细胞分化受抑，骨形成降低，这可能是糖尿病性骨质疏松产生的机制之一。2.阿仑膦酸钠在适宜浓度下能够可能通过抑制 miR-133a 的表达来促进 RUNX2 的表达，进而促进 OCN 和 OSX 的表达，积极调节成骨细胞的增殖和分化，进而改善骨的微观结构和骨质疏松状态，综合上述结论，这将为将来研发以 miRNA 为靶点的新型抗骨质疏松药物开发提供新的方向，以及在今后随着微小RNA研究的完善，可在预测及诊断老年性疾病及糖尿病相关并发症基因芯片等技术上应用。