雷帕霉素对结肠癌细胞增殖和凋亡的影响

Effects of rapamycin on proliferation and apoptosis of colon cancer cells

目的探讨雷帕霉素对结肠癌细胞株HCT-116增殖、凋亡及腺苷酸活化蛋白激酶(AMPK)通路的影响。方法HCT-116细胞分别加入0、2、4、8μg/mL的雷帕霉素处理24、48、72 h,采用MTT法和RT-PCR法分别检测细胞增殖情况和沉默信息调节因子2相关酶类1(SIRT1)、AMPK mRNA的表达。HCT-116细胞分别加入0、2、4、8μg/mL的雷帕霉素处理24 h,采用膜联蛋白V/碘化丙啶双染法和Western blot法分别检测细胞凋亡和SIRT1、AMPK蛋白表达。另取HCT-116细胞分为对照组和转染组(转染SIRT1短发夹RNA),比较两组细胞增殖及AMPK蛋白表达。结果雷帕霉素可呈浓度依赖和时间依赖抑制HCT-116细胞的增殖,降低SIRT1、AMPK mRNA表达(P<0.05)。雷帕霉素可呈浓度依赖促进细胞凋亡,降低SIRT1、AMPK蛋白表达(P<0.05)。转染组细胞AMPK蛋白表达量和细胞增殖能力低于对照组(P<0.05)。结论雷帕霉素可能通过降低SIRT1表达和调节AMPK通路抑制结肠癌细胞的增殖,并促进其凋亡。

Objective To investigate the effects of rapamycin on the proliferation and apoptosis of colon cancer cell line HCT-116.Methods MTT and RT-PCR were used to detect the cell proliferation and SIRT1,adenosine monophosphate-activated protein kinase(AMPK)mRNA expression of HCT-116 cells treated with 0,2,4 and 8μg/mL rapamycin for 24,48 and 72 hours,respectively.Annexin V/propidium iodide double staining method and Western blot were used to detect the cell apoptosis and SIRT1,AMPK protein expressions of HCT-116 cells treated with 0,2,4 and 8μg/mL rapamycin for 24 hours,respectively.The HCT-116 cells were transfected with SIRT1 short hairpin RNA,in which the cell proliferation and AMPK protein expression were compared with those in the cells without transfection.Results Rapamycin inhibited the proliferation of the cells and reduce the mRNA expressions of SIRT1 and AMPK in a concentration-and time-dependent manners(P<0.05).Rapamycin promoted the cell apoptosis and reduced SIRT1 and AMPK protein expressions in a concentration-dependent manner(P<0.05).The AMPK protein expression and cell proliferation ability of the transfected HCT-116 cells were lower than those without transfection(P<0.05).Conclusion Rapamycin can inhibit the proliferation of colon cancer cells and promote the apoptosis of the cells,which may be related to a decrease of SIRT1 expression and AMPK pathway regulation.