siRNA抑制Survivin基因对大肠癌细胞HCT116增殖及凋亡的的影响

INFLUENCE OF siRNA-MEDIATED INHIBITING SURVIVIN GENE ON PROLIFERATION AND APOPTOSIS OF HCT116 CELLS

目的探讨si RNA沉默Survivin基因后对人大肠癌细胞系HCT116的Survivin蛋白表达、细胞增殖及凋亡的影响。方法以脂质体lip-2000为载体,用针对Survivin特异靶点的si RNA转染人大肠癌细胞系HCT116后,应用免疫细胞化学S-P法、Western Blot检测Survivin蛋白表达变化;MTT法检测细胞增殖;流式细胞技术检测细胞凋亡。结果免疫细胞化学结果显示:HCT116的正常对照组、脂质体对照组及阴性错配对照组细胞浆均呈强阳性表达,Survivin-si R-NA组细胞浆Survivin呈弱阳性表达;Western blot结果显示:HCT116细胞系Survivin-si RNA组细胞的蛋白条带亮度均明显低于正常对照组、脂质体对照组和阴性错配对照组;MTT检测结果:与阴性错配对照组相比,Survivin-si RNA组细胞生长出现明显的抑制(P<0.05),不同时段(24h、48h、72h)肿瘤细胞增殖抑制率之间有显著差异(P<0.05)。流式细胞术检测Survivin-si RNA组细胞凋亡比例为9.72%,明显高于空白对照组及阴性错配对照组(P<0.01)。结论si RNA抑制Survivin基因可以抑制大肠癌细胞增殖,促进凋亡;Survivin有望成为大肠癌基因治疗的新靶点。

Objective To explore the expression of survivin protein and the effect of transfection of Survivin-siRNA on the proliferation and apoptosis of human colorectal cancer HCT116 cells.Methods Survivin-targeted siRNA was transfeted into human colorectal cancer HCT116 cells with lipofectamine 2000.Survivin protein expression was detected by immunocytochemistry and Western Blot.Cell proliferation was measured by MTT assay.Cell apoptosis was detected by flow cytometry.Results The HCT116 cells showed strongly positive expression in normal control,Lip-2000 control and mismatch control groups,but weakly positive expression in the Survivin-siRNA interfered group.The protein level of Survivin was significantly lower in the Survivin-siRNA group than that in the control by Western blot.Compared with that of the mismatch control group,the cell growth inhibition was significant in the Survivin-siRNA interfered group by MTT assay（P〈0.05）.The proliferation rate of HCT116 cells was significantly different at different times（24h,48h,72h）（P〈0.05）.The cell apoptosis rate of the Survivin-siRNA interfered group was 9.72%,significantly higher than that of the blank control and mismatch control groups by FCM.Conclusion The siRNA targeting Survivin gene can inhibit the proliferation of colorectal cancer cells and induce apoptosis.Survivin may become a new gene therapy target of colorectal cancer.