目的:探讨糖尿病与骨代谢的关系以及潜在的表观遗传学机制.方法:运用即时聚合酶链锁反应(RT-PCR)检测糖尿病小鼠与野生型小鼠骨髓中成骨相关因子及组蛋白去乙酰化酶(HDACs)的mRNA表达情况,体外培养小鼠骨髓间充质干细胞(BMSCs),检测高糖培养7,15 d后其成骨相关因子及HDACs的mRNA及蛋白表达变化,染色质免疫共沉淀(ChIP)分析HDAC2与Runx2启动子区域结合情况.结果:糖尿病小鼠骨髓中成骨相关因子mRNA较对照组均出现下调,OCN、Col1降低尤为显著(P<0.05),HDAC2 mRNA表达较对照组升高明显(P<0.05).RT-PCR与Western Blot检测结果可见BMSCs ALP、OCN、Runx2、OSX的mRNA及蛋白表达在7 d及15 d均随着葡萄糖浓度的升高而降低,而HDAC2的表达随着葡萄糖浓度的升高而增加.染色质免疫共沉淀结果示25 mM葡萄糖处理组BMSCs中HDAC2与Runx2基因启动子上、下游结合比例明显高于对照组(P<0.001;P<0.05).结论:糖尿病可通过HDAC2抑制Runx2转录活性,从而抑制骨髓间充质干细胞成骨分化影响骨代谢.%Objective To investigate the relationship between diabetes and bone metabolism and the potential epigenetic mechanisms. Methods BMSCs were cultured for 7 and 15 days in cell culture medium with different concentrations of glucose. The mRNA and protein expression of HDACs and osteogenesis-related genes were detected by RT-PCR and Western blot assay ,respectively. Moreover ,the combination of HDAC to the promoter region of Runx2 was tested by the chromatin immunoprecipitation (ChIP) assay. Results ThemRNA expression of osteogenesis-related genes ,incuding OCN(P < 0.05)and Col1(P < 0.05),in the bone marrow of diabetic mice was significantly reduced compared with the control mice. The mRNA and protein expression of ALP ,OCN ,Runx2 and OSX was gradually reduced with the increasing concentration of glucose ,while HDAC2 mRNA and protein expression was increased. The binding activity of HDAC2 to the upstream and downstream of Runx2 promoter region in 25mM glucose-treated BMSCs was higher than the control group(P < 0.05). Conclusoins Diabetes might repress osteogenesis of BMSCs via inhibiting the activity of Runx2 through upregu-lating the expression of HDAC2.
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