miR-34a调节人卵巢颗粒细胞凋亡

目的: 探讨miR-34a对人卵巢颗粒细胞的作用及其机制。方法: 体外培养人卵泡颗粒细胞，分为模拟物转染组、抑制物转染组、模拟物阴性对照转染组、抑制物阴性对照转染组，分别转染miR-34a模拟物、miR-34a抑制物、模拟物阴性对照和抑制物阴性对照，另设空白对照组不做任何处理；实时定量荧光PCR(qRT-PCR)检测各组细胞miR-34a、Bcl-2、Bax、生存素、半胱氨酸天冬氨酸特异性蛋白酶3(Caspase-3)的mRNA表达水平，蛋白质印迹法检测Bcl-2、Bax、生存素、Caspase-3、剪切体Caspase-3等凋亡相关蛋白的表达；流式细胞术检测细胞凋亡率。结果： miR-34a模拟物转染明显下调Bcl-2 mRNA表达水平，降低Bcl-2、生存素蛋白表达，并增加Bax、Caspase-3蛋白表达，促进人颗粒细胞凋亡；miR-34a抑制物转染作用则与之相反；然而，miR-34a模拟物和抑制物转染对Bax、生存素、Caspase-3 的mRNA水平均无明显影响。结论： miR-34a可通过调控凋亡相关的蛋白与基因的表达，促进人颗粒细胞凋亡。

miR-34a regulates apoptosis of human ovarian granulosa cells

Objective To investigate the effect of miR-34a on human ovarian granulosa cells and potential mechanisms. Methods The cultured human granulosa cells were divided into mimic group, inhibitor group, mimic negative control group, inhibitor negative control group, which were transfected with miR-34a mimic, miR-34a inhibitor, mimic negative control, and inhibitor negative control, respectively. The mRNA expression of Bcl-2, Bax, Survivin, Caspase-3 and miR-34a expression were measured by quantitative real-time PCR(qRT-PCR). Western blotting was used to detect the expression of apoptosis-related proteins, e.g. Bcl-2, Bax, Survivin, Caspase-3 and Cleaved-Caspase-3. Cell apoptosis percentage was evaluated by flow cytometry. Results miR-34a mimic transfection significantly down-regulated Bcl-2 mRNA levels, reduced Bcl-2, and survivin protein levels and increased Bax, Caspase-3 protein levels, and promoted apoptosis of human granulosa cells; however, miR-34a inhibitor transfection was shown to produce the opposite effect; however, Bax, Survivin, Caspase-3 mRNA levels were not significantly affected by miR-34a mimic or inhibitor transfection.  Conclusion miR-34a promoted apoptosis of human granulosa cells by regulating the expression of apoptosis-related proteins or mRNA. ［Key words］miR-34a; ganulosa cells; apoptosis