线粒体损伤对NAFL大鼠肝细胞凋亡的影响  

Effects of mitochondrial damage on hepatocyte apoptosis in nonalcoholic fatty liver rats

作　　者：康敏[1] 钟德君[1] 李鹏[2] 刘翼[1] 李昌平[1] 

机构地区：[1]泸州医学院附属医院,四川泸州646000 [2]绵阳市中心医院

出　　处：《山东医药》2010年第17期22-24,共3页Shandong Medical Journal

基　　金：泸州医学院基金项目(07087)

摘　　要：目的研究肝细胞凋亡及线粒体损伤在大鼠非酒精性脂肪肝(NAFL)形成中的作用,探讨NAFL的发病机理。方法 将30只雄性SD大鼠随机分为正常对照组和高脂组;HE染色观察肝组织光镜下的病理改变;电镜观察肝细胞超微结构变化;流式细胞仪检测肝细胞凋亡及线粒体膜电位;实时荧光定量PCR法检查caspase-3的mR-NA表达;免疫组化法检查细胞色素C(CytC)含量及caspase-3的蛋白表达。结果 HE染色结果显示,正常对照组大鼠肝脏无异常发现,高脂组肝脏在4周时出现脂肪变,8周出现轻度脂肪肝,12周出现中、重度脂肪肝;电镜下观察高脂组肝组织中肝细胞线粒体肿胀,嵴变短、减少甚至消失,粗面内质网减少,核型欠规则;与正常对照组比较,高脂组4～12周肝细胞凋亡指数增加,线粒体膜电位明显下降(P<0.05);高脂组caspase-3的mRNA表达随着时间延长逐渐增高;与正常对照组比较,高脂组CytC、caspase-3的蛋白表达随着时间延长逐渐增高,以12周增加明显(P<0.05)。结论大鼠NAFL引起的肝细胞损伤与肝细胞凋亡密切相关,线粒体膜电位下降启动了CytC由线粒体向胞质释放,激活caspase家族的级联反应,最终导致肝细胞凋亡,线粒体功能损伤在NAFL发病机制中起关键作用。Objective To investigate the roles of mitochondrial damage in hepatocye apoptosis in nonalcoholic fatty liver（NAFL） rats and the pathogenesis.Methods A total of 30 SD rats was randomly divided into basic diet-control group and high-fat diet group.Each of the two groups was subdivided into three subgroups（4-,8-and 12-week group）（n=5 in each group）.Pathological changes in the liver tissue were observed by HE staining and light microscopy.Ultrastructural changes in the hepatocyte were observed by electron microscopy.Hepatocyte apoptosis index（AI） and mitochondrial membrane potential were detected by flow cytometry（FCM）.Meanwhile,the expressions of caspase-3 were detected by real-time flurescent quantitive polymerase chain reaction.The expressions of hepatocyte cytochrome C（CytC） and caspase-3 were detected by immunohistochemistry method.Results HE staining revealed that fatty degeneration at 4 w,mild fatty liver at 8 w,moderate to severe fatty liver at 12 w in high-fat diet group,and electron microscopy showed ultrastructural changes in hepatocytes high-fat diet group（12 w）,including swelling of hepatic mitochondria,shortened or even disappearance of crests,decreased of rough surfaced endoplasmic reticulum.Apoptotic indexes of nonalcoholic steatosis rat model of 4-12 week were significantly higher than those of normal.Mitochondrial membrane potential in nonalcoholic steatosis rat model of 4-12 week were significantly decreased than those in normal group（P0.05）.The expressions of hepatocyte CytC and caspase-3 in rat nonalcoholic steatosis model were markedly increased compared with normal group,especially in 12-w group.Conclusions Nonalcoholic steatosis inducing liver damage associates with apoptosis.Decline of mitochondrial membrane potential,release of CytC and activation of caspase result in hepatocyte apoptosis.The damage to the mitochondrial function may play an important role in the genesis and progression of NAFL.

关 键 词：脂肪肝 凋亡 线粒体膜电位 细胞色素C CASPASE-3 

分 类 号：R575.5[医药卫生—消化系统]