线粒体凋亡途径在锰致鸡支持-生精细胞凋亡中的作用

The Role of Mitochondria Mediated Apoptosis Pathway in Sertoli-germ Cell Apoptosis Induced by Manganese in Cocks

探讨线粒体凋亡途径在锰致鸡支持-生精细胞凋亡中的作用。取对数生长期鸡支持-生精细胞,用含0、2、3、4mmol.L-1MnCl2的DMEM培养液,培养24h,采用吖啶橙-溴化乙锭(AO/EB)双荧光染色法检测鸡支持-生精细胞凋亡,流式细胞仪检测线粒体膜电位(△ψm),Westernblot法检测胞浆内细胞色素C(Cytc)蛋白表达含量,分光光度法检测半胱氨酸天冬氨酸蛋白酶9,3(Caspase-9,3)活性。用含MnCl2的DMEM培养液培养24h后,处理组细胞与对照组相比,鸡支持-生精细胞凋亡指数逐渐升高(P<0.01);线粒体膜电位显著降低(P<0.01);胞浆中Cytc蛋白表达量逐渐增加;Caspase-9,3活性逐渐升高(P<0.01)。线粒体途径介导的凋亡是锰对鸡生殖毒性作用的主要机制之一。

The aim of this study was to discuss the role of mitochondria mediated apoptosis pathway in Sertoli-germ cell apoptosis induced by manganese in cocks.Cock Sertoli-germ cells were cultivated in the DMEM for 24 h added with MnCl2,whose final concentrations were 0,2,3,and 4 mmol·L-1 respectively.The apoptosis was detected by AO/EB double staining.Mitochondrial transmembrane potential was detected by flow cytometer.Expression of Cytochrome c（Cytc）in cytolymph was detected by western blotting.The activities of Caspase-9,3 were examined by spectrophotography.Compared with control group,the apoptosis index（AI）of cock Sertoli-germ cells was significantly increased（P0.01）,mitochondrial transmembrane potential was significantly decreased（P0.01）,expression of Cytc in cytolymph was increased,and the activity of Caspase-9,3 were increased（P0.01）in 2,3,and 4 mmol·L-1 MnCl2 group.The apoptosis mediated by mitochondria pathway was one of the reproductive toxic mechanisms caused by manganese.