促凋亡蛋白Bid诱导肝细胞凋亡的机制

为研究促凋亡蛋白Bid对肝细胞凋亡过程中的调节机制 ,在体内和体外分别用TNF α或抗Fas抗体诱导小鼠肝细胞凋亡 .免疫荧光染色观察Bax转位和构象变化 ;采用ELISA检测caspase 3和 8的活性 ;Western印迹测定Bid和Bax的裂解活化及Bax的转位和插入 .结果显示 :TNF α或抗Fas抗体通过激活Bid导致Bax转位和构象变化 ,使Bax得以插入线粒体膜诱导肝细胞凋亡 .阻断Bid的作用 ,则Bax的转位和插入明显被削弱 ,肝细胞的凋亡受到抑制 .提示由死亡受体诱导的肝细胞调亡可能受Bid调节 ,Bax转位和插入依赖于Bid .

Mechanism of Hepatocytes Apoptosis Induced by Proapoptosis Protein Bid

To investigate the mechanism of hepatocytes apoptosis induced by proapoptosis protein Bid, mouse primary hepatocytes were isolated from wild type and Bid deficient mice and treated with TNF-α or Anti-Fas antibody to induce cell apoptosis. Immunofluorescence staining of Bax was processed to recognize Bax translocation and its conformation change. The wild type or wild type mice transfected with the adenovirus carried DN-FADD (dominant negative -Fas associated death domain) and Bid deficient mice were injected with anti-Fas antibody 2 hours before sacrificed. Caspase 3 and 8 activities were measured. Bid cleavage bands and Bax conformation change bands were detected by Western blotting.The results showed that during death receptors including TNF-α or anti-Fas antibody induced hepatocytes apoptosis, Bax translocation and conformation change through activating Bid, which caused Bax to insert to mitochondria membrane of hepatocytes. The translocation and insertion of Bax were blocked when Bid was knocked-out or blocked, and hepatocytes apoptosis was delayed or inhibited. So it is postulated that hepatocytes apoptosis induced by death receptor might be regulated by Bid, and Bax translocation and insertion dependent on Bid.