西罗莫司诱导人肝癌细胞株BEL-7402凋亡及其机制

目的探讨西罗莫司（SRL）在体外诱导人肝癌细胞株BEL-7402的凋亡作用及其机制。方法以不同浓度的SRL（5、10、20、30、40和50nmol／L）作用于体外培养的人肝癌细胞株BEL-7402，应用流式细胞仪检测培养24、48和72h时的细胞凋亡情况；Hoechst 33258荧光染色法观察细胞凋亡时的形态学变化；Western Blot法观察BEL-7402细胞中Caspase-3、Bcl-2、Bcl-xl和Bax基因的表达变化；采用Caspase-3试剂盒检测Caspase-3酶的活性；JCl染色法检测细胞线粒体膜电位（△ψbm）。结果SRL可诱导BEL-7402细胞凋亡，并与药物浓度和作用时间呈正相关。SRL作用BEL-7402细胞后48h，在Hoechst33258荧光染色图片上可见核浓缩及核碎裂等典型的细胞凋亡特征，凋亡过程中线粒体膜电位下降及Caspase-3酶原蛋白激活，伴有Bcl-2、Bcl-xl蛋白表达的降低和Bax蛋白上调。结论SRL能使抗凋亡蛋白Bcl-2、gcl-xl表达降低，促凋亡蛋白Bax表达上调，线粒体膜电位下降，激活Caspase-3酶原蛋白，从而诱导细胞凋亡。

Apoptosis of hepatocelluar carcinoma BEL-7402 cells in vitro induced by rapamycin and its action mechanism

Objective To investigate cell apoptosis induced by rapamycin on human hepatocellu- lar carcinoma BEL-7402 cells and its action mechanism.Methods BEL-7402 cells in culture medium in vitro were treated with rapamycin with different concentrations of 5,10,20,30,40 and 50 nmol/L. Cell apoptosis was detected by flow cytometry(FCM)at 24,48 and 72 h,and morphological changes were observed by Hoechst 33258 staining;The protein expression of Caspase-3,Bcl-2,Bcl-x1 and Bax was assayed by Western blotting,the activity of Caspases was determined by Caspase colorimetric as- say kit,and cell mitochondrial membrane potential was investigated by using JC-1 dying standard methods.Results Rapamycin could induce apoptosis of hepatocellular carcinoma BEL-7402 cells sig- nificantly in a time- and dose-dependent manner.Marked morphological changes of cell apoptosis were observed very dearly by Hoechst 33258 staining.Rapamycin could activate Caspase-3 and cause dis- ruption of the mitochondrial membrane potential during apoptosis.Anti-apoptotic proteins Bcl-2 and Bcl-x1 were down-regulated and pro-apoptotic protein Bax was up-regulated remarkably when apoptosis occurred.Conclusion Rapamycin could induce apoptosis by down-regulation of anti-apoptotic proteins Bcl-2,Bcl-x1 and bax up-regulation of pro apoptotic protein,as well as via disruption of mitochondrial membrane potential and activation of Caspase-3.