奥利司他对肝癌SMMC-7721细胞增殖和凋亡的影响及其机制研究

The effect of orlistat on proliferation and apoptosis of SMMC-7721 hepatoma cells and its mechanism

目的:研究奥利司他(orlistat)对肝癌SMMC-7721细胞增殖和凋亡的影响并探讨其作用机制。方法:运用CCK-8法检测不同浓度奥利司他对SMMC-7721细胞增殖的影响;采用流式细胞术检测不同浓度奥利司他(10,20和40μmol·L^-1)对SMMC-7721细胞周期分布与凋亡的影响;运用Western Blot法检测奥利司他对SMMC-7721细胞中p-AKTThr308,p-AKTSer473,特异性周期蛋白-D1(Cyclin D1)及增殖细胞核抗原(PC-NA)等增殖相关蛋白表达的影响;在AKT/c-Met过表达SMMC-7721细胞中进一步探讨奥利司他调控AKT信号通路抑制肝癌细胞恶性转化的作用机制。结果:奥利司他对SMMC-7721细胞增殖具有抑制作用且呈剂量依赖性;与对照组比较,不同浓度奥利司他(10,20和40μmol·L^-1)能够阻滞SMMC-7721细胞周期于G0/G1期,同时能够显著诱导SMMC-7721细胞凋亡,细胞凋亡率分别为(26.5±1.1)%,(31.8±1.6)%和(46.8±3.7)%;奥利司他能够显著降低SMMC-7721细胞中p-AKTThr308,p-AKTSer473,Cyclin D1,PCNA等蛋白的表达水平;在AKT/c-Met过表达SMMC-7721细胞中,奥利司他能够降低AKT磷酸化水平以及Cyclin D1,PCNA等增殖相关蛋白表达水平。结论:奥利司他能够抑制SMMC-7721细胞增殖并诱导其凋亡,其作用机制可能与奥利司他抑制AKT及其下游增殖信号活化有关。

Objective:To investigate the effect of orlistat on proliferation and apoptosis of SMMC-7721 cellsand its mechanism.Methods:CCK-8 assay was used to detect the effect of orlistat at different concentrations onthe proliferation of SMMC-7721 cells.The flow cytometry was used to detect the effect of orlistat (10,20 and 40μmol·L^-1) on the cycle distribution and apoptosis of SMMC-7721 cells.Western blot method was used to detectthe effect of orlistat on the proliferation-related proteins,including p-AKTThr308,p-AKTSer473,Cyclin D1 and proliferatingcell nuclear antigen (PCNA),in SMMC-7721 cells.In SMMC-7721 cells overexpressing AKT/c-Met,the mechanismof orlistat to inhibit malignant transformation of hepatoma cells via regulation of AKT signaling pathway was furtherinvestigated.Results:Orlistat inhibited the proliferation of SMMC-7721 cells in a dose-dependent manner.Compared with control group,treatment with orlistat (10,20 and 40μmol·L^-1) could block the cell cycle of SMMC-7721 cells at G0/G1 phase,and significantly induce the apoptosis of SMMC-7721 cells with the rates of(26.5±1.1)%,(31.8±1.6)%,and (46.8±3.7)%,respectively.In addition,orlistat significantly reducedthe protein expression levels of p-AKTThr308,p-AKTSer473,cyclin D1,and PCNA in SMMC-7721 cells.Moreover,inSMMC-7721 cells overexpressing AKT/c-Met,orlistat significantly reduced the phosphorylation level of AKT andthe expression level of proliferation-related proteins,including cyclin D1 and PCNA.Conclusion:Orlistat caninhibit proliferation and induce apoptosis of hepatocellular carcinoma cells SMMC-7721 by inhibiting the activationof AKT-mediated proliferation signals.