羟基喜树碱通过线粒体途径诱导肝癌SMMC-7721细胞凋亡的观察

Hydroxycamptothecine induces apoptosis in human hepatoma SMMC-7721 cells through mitochondrial pathway

目的:研究羟基喜树碱是否可通过线粒体途径诱导人肝癌细胞SMMC-7721凋亡.方法:将不同浓度的羟基喜树碱作用于肝癌细胞SMMC-7721,用MTT法检测细胞增殖,光镜观察细胞形态;用荧光染料MitocaptureTM检测线粒体跨膜电位、用Western blot法检测细胞色素C从线粒体的释放;用AnnexinⅤ-FITC,hoechst法及电镜检测细胞的凋亡状况.结果:羟基喜树碱对肝癌细胞SMMC-7721的增殖有明显的抑制作用,IC50约是80 mg/L.当用80 mg/L羟基喜树碱作用不同时间后:线粒体膜跨电位下降并伴随有细胞色素C从线粒体至细胞质的释放;细胞膜的磷脂酰丝氨酸外翻;细胞核染色质固缩,呈致密浓染的凋亡状态.结论:羟基喜树碱通过线粒体途径引起线粒体跨膜电位下降与细胞色素C的释放可能是其诱导肿瘤细胞凋亡的途径之一.

AIM： To study whether hydroxycamptothecine （HCPT） could induce apoptosis through mitochondrial pathway by reducing mitochondrial transmembrane potential and cytochrome C release in human hepatoma SMMC-7721 cells. METHODS： SMMC-7721 cells were incubated in vitro with HCPT at different concentrations. The effect of HCPT on cell proliferation was measured by MTT assay; cell morphology was observed under a light microscope; mitochondrial transmembrane potential was detected by Mitocapture^TM Mitochondrial Apoptosis Detection Kit; cytochrome C release of cellular fraction was examined by Western blot; cell apoptosis were ascertained by Annexin V-FITC, hoechst and electron microscopy. RESULTS： HCPT noticeably inhibited the proliferation of SMMC-7721 cells and the IC90 dose was about 80μg/mL; when cells treated by 80μg/mL HCPT, mitochondrial transmembrane potential were reduced and cytochrome C was released from mitochondria to cytosol; at the same time, phosphatidylserine （PS） was exposed from the inner to the outer leaflet of the plasma membrane, and nucleus showed nuclear pyknosis, chromatin condensation and cell apoptosis. CONCLUSION： HCPT could inhibit proliferation and induce apoptosis of human hepatoma SMMC-7721 cells through the change of mitochondrial transmembrane potential and the release of cytochrome C.