论文部分内容阅读
目的探讨去甲斑蝥素(Norcantharidin,NCTD)诱导人皮肤鳞状细胞癌A431细胞凋亡作用及其相关分子机制。方法体外培养A431细胞,分为对照组和NCTD低、高剂量组(20,40μmol·L~(-1))。采用MTT法检测对细胞增殖的抑制作用;Annexin V/PI染色法检测细胞凋亡;试剂盒检测细胞活性氧(ROS)水平、丙二醛(MDA)含量、线粒体膜电位(MMP)变化、Caspase-3及Caspase-9酶活性变化;Western Blot法检测对细胞Bcl-2及Bax蛋白表达水平的影响。结果采用NTCD处理A431细胞48 h,与对照组相比,NCTD低、高剂量组均能抑制细胞增殖能力(P<0.05),并明显诱导细胞凋亡(P<0.05),提高细胞内的ROS水平(P<0.05,P<0.01)和MDA含量(P<0.05,P<0.01),降低细胞MMP(P<0.05,P<0.01)。同时,NTCD可以抑制Bcl-2蛋白表达及上调Bax蛋白表达,并能够明显升高细胞的Caspase-3及Caspase-9酶活性。结论 NCTD可能通过提高细胞内ROS水平的途径诱导A431细胞发生凋亡。
Objective To investigate the apoptosis of human skin squamous cell carcinoma A431 cells induced by norcantharidin (NCTD) and its related molecular mechanisms. Methods A431 cells were cultured in vitro and divided into control and NCTD low and high dose groups (20, 40 μmol·L -1). The inhibition of cell proliferation was detected by MTT assay. Apoptosis was detected by Annexin V / PI staining. The levels of reactive oxygen species (ROS), malondialdehyde (MDA), mitochondrial membrane potential (MMP) and Caspase -3 and Caspase-9 enzyme activity changes; Western Blot detection of cell Bcl-2 and Bax protein expression levels. Results Compared with control group, NTCD treatment of A431 cells for 48 h showed that NCTD low and high dose groups could both inhibit cell proliferation (P <0.05) and induce apoptosis (P <0.05), and increase intracellular ROS (P <0.05, P <0.01) and MDA content (P <0.05, P <0.01), and decreased MMP (P <0.05, P <0.01). At the same time, NTCD can inhibit the expression of Bcl-2 protein and up-regulate the expression of Bax protein, and significantly increase the activity of Caspase-3 and Caspase-9. Conclusion NCTD may induce apoptosis of A431 cells by increasing the intracellular ROS level.