siRNA沉默ERCC1基因表达对结肠癌细胞奥沙利铂耐药的影响和机制

Effect and mechanism of siRNA silencing ERCC1 gene expression on oxaliplatin resistance in colon cancer cells

目的采用小干扰RNA(siRNA)技术作用于人结肠癌细胞HCT116中的ERCC1,探讨其对结肠癌铂类耐药细胞株增殖、凋亡的影响。方法体外培养结肠癌奥沙利铂耐药细胞株HCT116/L-OHP并验证其耐药性;qRT-PCR和蛋白印迹技术检测细胞和组织中ERCC1表达量的变化;采用siRNA-NC和siRNA-ERCC1转染细胞,CCK-8法检测耐药细胞的增殖能力,流式细胞术检测耐药细胞的凋亡变化。结果结肠癌耐药组织和人结肠癌耐药细胞HCT116/L-OHP ERCC1的表达水平明显升高(P<0.05);转染siRNA ERCC1后,HCT116/L-OHP细胞中的ERCC1 mRNA和蛋白水平明显降低(P<0.05);奥沙利铂联合处理后,siRNA ERCC1转染使HCT116/L-OHP细胞的增殖活性下降、凋亡率升高。结论siRNA能有效下调ERCC1基因表达,抑制细胞增殖,增加细胞凋亡,增强奥沙利铂对耐药细胞株HCT116/L-OHP的杀伤作用。

Objective Small interfering RNA(siRNA)technology was used to affect ERCC 1 in human colon cancer cells HCT116 to investigate its effect on the proliferation and apoptosis of platinum-resistant cell lines in colon cancer.Methods To culture the colon cancer oxaliplatin-resistant cell line HCT116/L-OHP and verify its resistance;ERCC1-expression in cells and tissues by qRT-PCR and protein blotting;siRNA-NC and siRNA-ERCC 1-transfection by CCK-8 and apoptotic changes of resistant cells by flow cytometry.Results The expression levels of HCT116/L-OHP ERCC1 in colon cancer resistant tissues and human colon cancer resistant cells were significantly increased(P<0.05);After transfection with siRNA ERCC1,the mRNA and protein levels of ERCC1 in HCT116/L-OHP cells were significantly reduced(P<0.05);After combined treatment with oxaliplatin,siRNA ERCC1 transfection resulted in a decrease in proliferation activity and an increase in apoptosis rate of HCT116/L-OHP cells.Conclusion The siRNA effectively down-regulated ERCC 1 gene expression,inhibited cell proliferation,increased apoptosis,and enhanced the killing effect of oxaliplatin on drug-resistant cell line HCT116/L-OHP.