金钗石斛脂溶性生物碱提取物诱导人结肠癌 HT-29 细胞凋亡

Fat-soluble alkaloids extracted from Dendrobium Nobile Lindl induced apoptosis of human colorectal cancer HT-29 cells

目的:研究金钗石斛脂溶性生物碱提取物对人结肠癌HT-29细胞生长的抑制作用及其诱导细胞凋亡的分子机制。方法:MTT法检测金钗石斛脂溶性生物碱提取物对HT-29细胞存活率的影响,Hoechst33342染色法观察细胞形态学变化,流式细胞仪检测胞内活性氧水平、线粒体膜电位、细胞周期和细胞凋亡率变化;蛋白免疫印迹检法测蛋白Caspase-3,9和胞内细胞色素C的表达。结果:金钗石斛脂溶性生物碱提取物能降低HT-29结肠癌细胞存活率,呈剂量和时间效应,48 h半数抑制浓度(IC50)为0.72 mg/m L;金钗石斛脂溶性生物碱提取物能诱导HT-29细胞凋亡,并诱导细胞周期阻滞于G2期;金钗石斛脂溶性生物碱提取物诱使线粒体膜电位降低,胞内活性氧浓度升高;激活型Caspase-9,3与胞内细胞色素C表达水平增高。结论:金钗石斛脂溶性生物碱提取物对HT-29结肠癌细胞的生长具有抑制作用并诱导细胞凋亡,其可能通过线粒体凋亡途径诱导细胞凋亡。

Objective：Anti-growth effects of fat-soluble alkaloids extracted from Dendrobium Nobile Lindl on human colorectal cancer HT-29 cells and its possible apoptotic mechanism were studied.Methods：The MTT assay was used to assess the effect of fat-soluble alkaloids extracted from Dendrobium Nobile Lindl on survival rate of HT-29 cells; the morphological changes of HT-29 ceils were observed with Hoechst 33342 staining; the levels of intracellular reactive oxygen species （ROS）, mitochondrial membrane potential（ △ψm）, cell cycle distribution and apoptosis rate were studied using flow cytometry;western blotting analysis was used to detect the expressions of Caspase- 9,3, and intracellular cytochrome C. Results ： Fat- soluble alkaloids extracted from Dendrobium Nobile Lindl decreased survival rate of human colorectal cancer HT-29 cells in a dose- and time-dependent manner; IC50 value for 48 h was 0.72 mg/mL. Fat-soluble alkaloids extracted from Dendrobium Nobile Lindl induced cell apoptosis and cell cycle was arrested in G2 phase ; it caused decline of △ψm and induced ROS accumulation with increased expression levels of apoptotic proteins such as activated Caspase- 9, activated Caspase- 3, and level of intraeellular cytochrome C.Conclusion：Fat-soluble alkaloids extracted from Dendrobium Nobile Lindl can inhibit the growth of HT-29 cells and induce cell apoptosis,which may be associated with the involvement of the mitochondria-mediated apoptotic pathway.