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目的研究羟基喜树碱诱导肝癌细胞株SMMC-7721凋亡时线粒体凋亡相关蛋白凋亡诱导因子表达及从线粒体发生核转位的变化。方法用80μ／ml羟基喜树碱作用于肝癌细胞株SMMC-7721后, 用吖啶橙／溴化乙啶染色法观察细胞凋亡现象;用电子显微镜观察线粒体超微结构;分别用逆转录聚合酶链反应、Western blot检测凋亡诱导因子在mRNA与蛋白质水平表达的变化;用激光共聚焦显微镜观察凋亡诱导因子在细胞凋亡时从线粒体到核的迁移变化。结果 80μg／ml羟基喜树碱作用SMMC-7721细胞后, 吖啶橙／溴化乙啶荧光双重染色可见细胞体积缩小、细胞皱缩、核碎裂等典型细胞凋亡形态学改变;超微结构观察发现线粒体肿胀;细胞凋亡时凋亡诱导因子在m R N A与蛋白质水平上的表达与对照组细胞相比没有明显变化,但凋亡诱导因子发生了从线粒体到核的迁移。结论羟基喜树碱可以通过线粒体途径诱导人肝癌细胞发生凋亡;在线粒体凋亡途径中,线粒体凋亡相关蛋白凋亡诱导因子从线粒体释放并发生核转位可能与羟基喜树碱诱导细胞凋亡密切相关。
Objective To study the expression of mitochondrial apoptosis-related protein (apoptosis-inducing factor) and its nuclear translocation from mitochondria induced by hydroxycamptothecin in hepatoma cell line SMMC-7721. Methods The apoptosis of hepatoma cell line SMMC-7721 was induced by 80μ / ml hydroxycamptothecin. Apoptosis was observed by acridine orange / ethidium bromide staining. The ultrastructure of mitochondria was observed by electron microscopy. The changes of mRNA and protein expression of apoptosis-inducing factor were detected by Western blotting. The changes of mitochondria-to-nucleus migration were observed by laser scanning confocal microscopy. Results After SMMC-7721 cells were treated with 80μg / ml hydroxycamptothecin, morphological changes such as cell volume reduction, cell shrinkage and nuclear fragmentation were observed by acridine orange / ethidium bromide staining. The ultrastructure The mitochondrial swelling was observed. The expression of apoptosis-inducing factor at the level of m RNA and protein did not change significantly compared with the control group, but the apoptosis-inducing factor migrated from mitochondria to nucleus. CONCLUSION: Hydroxycamptothecin can induce apoptosis in human hepatocarcinoma cells through mitochondria pathway. In mitochondrial apoptosis pathway, mitochondrial apoptosis-inducing factor is released from mitochondria and undergoes nuclear translocation, which may be related to the apoptosis induced by hydroxycamptothecin Closely related.