SiRNA联合介导Livin和Survivin基因沉默诱导人肾癌细胞786-O化疗敏感性的实验研究

目的:观察siRNA联合介导Livin和Survivin基因的沉默诱导人肾癌细胞786-O化疗敏感性的作用。方法:构建Livin和Suvivin联合靶向的小干扰RNA(siRNA)重组表达载体shRNA,然后将目的载体转染于人肾癌细胞株786-O。应用实时荧光定量聚合酶链反应(Real-time PCR)及Western blot方法分别检测转染前细胞经顺铂、5-FU和丝裂霉素处理后的Livin和Survivin基因的mRNA与蛋白水平的变化。应用CCK-8试验检测转染前后细胞对顺铂、5-氟尿嘧啶(5-FU)和丝裂霉素的半数致死量(IC50)、细胞增殖的变化。结果:CCK-8结果显示,联合介导Livin和Survivin基因沉默组细胞较分别单独介导细胞组化疗的敏感性明显增强(P0.05),且转染前后细胞对顺铂、5-FU和丝裂霉素的IC50发生显著变化(P0.05)。结论:SiRNA联合介导Livin和Survivin基因表达下调,发挥协同增强的siRNA作用。

Experimental study of chemotherapy sensitivity in human renal carcinoma cell line 786-O induced by siRNA-mediated co-silencing of Livin and Survivin genes

Objective：To investigate the chemotherapy sensitivity of human renal carcinoma cell line 786-O in- duced by siRNA-mediated co-silencing of Livin and Survivin genes. Method： We constructed Livin and Survivin combined targeting small interfering RNA （siRNA） recombinant expression vector. Then we transfected them in 786-O cell line. Real-time fluorescent quantitative polymerase chain reaction （Real-time PCR） and Western blot methods were used respectively to detect the expression level of Livin and Survivin in mitomycin, 5-FU and cispla- tin treated 786-O cell. Then we investigated the effect of these shRNA in human renal carcinoma cell line 786-O, including the affection of the chemotherapy sensitivity and proliferation. Result：CCK-8 analysis results showed that the sensitivity to chemotherapy of combined mediated Livin and Survivin genes silencing were higher than separate- ly mediated Livin and Survivin （P〈0.05）. Moreover, IC50 showed significant difference between before and after the transfection （P〈0.01）. Conclusion：SiRNA-mediated down-regulation of Livin and Survivin genes enhance the synergistic effect.