Puma基因对HeLa细胞凋亡的作用及机制的初步研究

Effect and mechanism of puma gene on the apoptosis of HeLa cell in vitro

目的研究外源性puma基因表达对HeLa细胞的作用以及线粒体释放的细胞色素C(Cyt c)和凋亡诱导因子(AIF)是否参与了凋亡过程。方法脂质体转染HeLa细胞;RT-PCT鉴定表达效果;AO/EB染色检测puma基因的促凋亡活性;Western blot检测细胞凋亡发生后线粒体中Cyt c和AIF位置是否发生迁移。结果脂质体介导下成功实现了外源性puma基因在HeLa细胞中高表达,AO/EB染色显示转染puma基因后HeLa细胞出现了一系列凋亡形态学改变,这种变化转染48h比24h时更明显;Western blot结果显示,puma基因表达后HeLa细胞的线粒体膜间隙蛋白Cyt c、AIF向胞浆迁移,迁移的蛋白量在转染48h比24h更明显。结论Puma基因表达有促进HeLa细胞凋亡的作用,且Cyt c和AIF参与puma基因促进HeLa细胞凋亡的过程。

Objective To study the effect of puma gene on the apoptosis of HeLa cell in vitro and whether the protein of cytochrome c （Cyt c） and apoptosis inducing factor（AIF） in the chondrosome participate in the process of apoptosis. Methods Liposome HeLa cell was transfected and the result of transfection was assessed by RT-PCR, then the morphologic changes of HeLa cell induced by puma gene were detected by AO/EB dyeing. Finally the position of Cyt c and AIF transferred in the chondrosome was studied by Western blot. Results AO/EB dyeing displayed that HeLa cell had a series of apoptosis morphologic changes after transfecting puma into HeLa cell, and the changes after 48h were more obvious compared to 24h. Western blot displayed that the protein Cyt c and AIF transferred toward to kytoplasm after the expression of puma gene. The mount of transfected protein after 48h was more obvious compared to 24h. Conclusion Puma gene has the effect of promoting apoptosis of HeLa cell in which Cyt c and AIF take part.