瑞戈非尼联合哌立福辛诱导人肝癌HepG2细胞凋亡的分子机制研究  

Research on the molecular mechanism of regorafenib combined with perifosine in inducing apoptosis of human hepatoma HepG2 cells

作　　者：陈雪健 王伟[1] 周健[1] 李宁 张浩鹏[1] 徐力善[1] CHEN Xuejian;WANG Wei;ZHOU Jian;LI Ning;ZHANG Haopeng;XU Lishan(Department of Oncological and Hepatobiliary Surgery,The Fourth Affiliated Hospital of Harbin Medical University,Harbin 150001,China)

机构地区：[1]哈尔滨医科大学附属第四医院肿瘤外科肝胆外科,哈尔滨150001

出　　处：《实用肿瘤学杂志》2021年第5期408-413,共6页Practical Oncology Journal

基　　金：哈尔滨医科大学附属第四医院火炬计划(编号:HYDSYHJ201903)。

摘　　要：目的探讨瑞戈非尼联合哌立福辛(AKT抑制剂)对人肝癌HepG2细胞凋亡的分子机制。方法不同浓度瑞戈非尼(1、5、10、15、20、25μmol/L)处理肝癌细胞后,通过CCK-8检测其细胞存活率;再设置不同浓度瑞戈非尼(0、5、10、20μmol/L)处理肝癌细胞后,流式细胞仪检测细胞凋亡;用免疫印迹法(Western blot)检测Bax、Bcl-2、p-AKT、AKT、p21、p27、Cleaved-caspase-8、Cleaved-caspase-9的表达情况;设置哌立福辛组(10μmol/L)、瑞戈非尼组(20μmol/L)及双药联合组(哌立福辛10μmol/L联合瑞戈非尼20μmol/L),检测细胞凋亡及相关蛋白表达变化。结果从5μmol/L浓度起,瑞戈非尼可剂量依赖性的抑制肝癌细胞增殖(P<0.05);Bcl-2及p-AKT表达下调,Bax、p21、p27、Cleaved-caspase-8、Cleaved-caspase-9表达上调(P<0.05);瑞戈非尼和哌立福辛双药联合应用后,细胞凋亡率、Bax表达量明显增加(P<0.05),Bcl-2、p-AKT表达量下降程度均较单独应用哌立福辛及瑞戈非尼明显(P<0.05)。结论瑞戈非尼通过增加p21、p27、Bax、caspase-8、caspase-9的表达,减少Bcl-2、p-AKT的表达,来诱导肝癌细胞凋亡,并且瑞戈非尼与哌立福辛双药联合应用后,明显增加肝癌细胞凋亡率,为药物治疗肝癌提供新的理论基础和思路。Objective The objective of this study was to investigate the molecular mechanism of regorafenib combined with perifoxin(AKT inhibitor)in apoptosis induction of human hepatoma HepG2 cells.Methods After treated with different concentrations of regorafenib(1,5,10,15,20 and 25μmol/L),the cell survival rate was detected in hepatoma cells by CCK-8 assay;and then different concentrations of regorafenib(0,5,10 and 20μmol/L)were setup.Flow cytometry was used to detect apoptosis.The expression of Bax,Bcl-2,P-AKT,AKT,p21,p27,Cleaved-caspase-8 and Cleaved-caspase-9 was detected by Western blot.Perifosine group(10μmol/L),regorafenib group(20μmol/L)and the drug combination group(perifosine 10μmol/L combined with regorafenib 20μmol/L)were set to detect cell apoptosis and related protein expression changes.Results Regorafenib inhibited proliferation of hepatoma cells in a dose-dependent manner from the concentration of 5μmol/L(P<0.05).The expression of Bcl-2 and P-AKT was down-regulated and the expression of Bax,p21,p27,Cleaved-caspase-8 and Cleaved-caspase-9 was up-regulated(P<0.05).After the combined application of regorafenib and perifosine,the apoptosis rate and Bax expression were significantly increased(P<0.05),and the decrease of Bcl-2 and P-AKT expression was more obvious than that of perifosine or regorafenib alone(P<0.05).Conclusion Regorafenib can induce apoptosis in hepatma cells by increasing the expression of p21,p27,Bax,Caspase-8 and Caspase-9,and reducing the expression of Bcl-2 and P-AKT.The combined application of regorafenib and perifosine significantly increases the apoptosis rate of hepatma cells,providing a new theoretical basis and ideas for the drug treatment of liver cancer.

关 键 词：瑞戈非尼 AKT抑制剂 肝癌 细胞凋亡 

分 类 号：R735.7[医药卫生—肿瘤]