线粒体-细胞色素C途径在非酒精性脂肪肝大鼠肝细胞凋亡中的作用

目的 研究大鼠非酒精性脂肪肝形成中肝细胞凋亡以及线粒体-细胞色素C途径在其中的作用.方法 Wistar大鼠40只,随机分为正常对照组和高脂饲料2、4、8和12周组(其中每组各8只);HE染色光镜观察肝脏组织病理改变;采用脱氧核糖核苷酸末端转移酶介导的缺口末端标记技术(TUNEL)以及流式细胞仪检测肝细胞凋亡指数;激光共聚焦显微镜检测肝细胞线粒体膜电位;免疫组织化学检查caspase-3在肝脏表达分布,Western blotting检测细胞色素 C以及caspase-3含量变化.结果 高脂饮食组4～12周组肝细胞线粒体膜电位与对照组比较明显下降,差异有统计学意义(P＜0.01);高脂饮食组肝细胞凋亡指数与对照组比较明显增加,随着脂肪肝的加重逐渐增高,差异有统计学意义(P＜0.01);高脂饮食组细胞色素C含量以及caspase-3蛋白表达随着时间延长逐渐增高,以12周增加明显,与对照组比较,差异有统计学意义(P＜0.01).结论 大鼠非酒精性脂肪肝引起的肝细胞损伤与肝细胞凋亡密切相关,线粒体-细胞色素C途径参与了其调控过程.

Effects of mitochondria-cytochrome C in hepatocyte apoptosis of non-alcoholic steatosis rats

Objective To investigate the roles of mitochondria-cytochrome C in hepatocye apoptosis in non-alcoholic steatosis rats and the pathogenesis.Methods Fouty Wistar rats were randomly divided into the control group and the high-fat diet group,and the high-fat diet groups were divided into 4 subgroups(2,4,8,12 week).Each subgroup had 8 rats.Pathological changes in the liver tissue were observed by HE staining and light microscopy.Mitochondrial membrane potential in hepatocyte was determined with the change of intensity of labeling rhodamine 123 by confocal micrographs,hepatocyte apoptosis index(AI) detected by transferase-mediated-UTP-biotin nick-end labeling(TUNEL) technique.Meanwhile,the expression of hepatocyte caspase-3 and cytochrome C were detected with immunohistochemistry or Western blot.Results The mitochondrial membrane potential in nonalcoholic steatosis rat model of 4-12 week were significantly decreased than those in normal group(P<0.01).AI of non-alcoholic steatosis rat was significantly higher than that of normal(P<0.01).The expression of hepatocyte caspase-3 and cytochrome C antigen in rat nonalcoholic steatosis model were markedly increased than in normal group.There was significant positive correlation between those and degree of nonalcoholic steatosis.Conclusion Non-alcoholic steatosis induced liver damage is associated with apoptosis,and mitochondria-cytochrome C mediated apoptotic pathway plays a role in the occurrence of steatosis.