双硫仑和顺铂通过内质网应激途径协同抗宫颈癌的实验研究

Experimental study on the synergistic anti- cervical cancer effect of disulfiram and cisplatin via endoplasmic reticulum stress

目的:评价双硫仑与顺铂联合用药是否具有协同促进人宫颈癌HeLa细胞凋亡的作用。方法:双硫仑和(或)顺铂处理体外培养的HeLa细胞株后,采用四甲基偶氮唑盐比色法(MTT法)检测细胞的存活率,Annexin V-FITC/PI染色法检测细胞凋亡水平变化,显微镜观察细胞形态变化,Western blotting法检测内质网应激相关蛋白CHOP、凋亡相关蛋白(Caspase-3、Caspase-8、Caspase-9和PARP)的表达变化。结果:与两药单独使用相比,双硫仑与顺铂联合用药能够明显抑制HeLa细胞的存活;可以导致HeLa细胞发生明显的形态学变化,出现细胞凋亡;可以引起内质网应激相关蛋白CHOP表达明显增加,进一步可以活化Caspase-3、Caspase-8、Caspase-9,引起PARP蛋白的切割。结论:双硫仑和顺铂联合用药具有明显的协同作用,有效抑制HeLa细胞的增殖、诱导细胞凋亡,其作用机制可能与诱导内质网应激引起Caspase系统活化相关。

Objective： To evaluate the synergistic anti - apoptosis of human cervical cancer HeLa cells of disulfiram combined with cisplatin. Methods： Human cervical cancer HeLa cells cultured in vitro were treated with disulfiram and （or） cisplatin, the survival rate of HeLa cells were detected by MTT method; Annexin V - FITC/PI staining was used to detect change of cell apoptosis; the morphological change was observed by microscope; the changes of expressions of CHOP, caspase - 3, caspase - 8, caspase - 9 and PARP were detected by Western blotting. Results： Compared with single drug treatment, disulfiram combined with cisplatin could significantly inhibit proliferation of HeLa cells, which induced obviously morphological changes and apoptosis of HeLa ceils, significantly increase of CHOP expression, fur- ther activate caspase - 3, easpase - 8, caspase - 9 and PARP cutting. Conclusion： Disulfiram and cisplatin have synergistic effects on in- hibiting cell proliferation and inducing cell apoptosis of HeLa ceils, and the mechanism may perhaps be related to activation of caspase sys- tem which is induced by endoplasmic reticulum stress.