双mTORC1/2抑制剂AZD2014诱导的自噬对HCCLM3肝癌细胞增殖的影响  

Dual mTORC1/2 inhibitor AZD2014 inhibits proliferation of HCCLM3 cells through induction of autophagy

作　　者：廖晖 徐小平[1] 周陈杰[1] 张健民[1] 钟克波 杨定华[2] Liao Hui;Xu Xiao-Ping;Zhou Chen-Jie;Zhang Jian-Min;Zhong Ke-Bo;Yang Ding-Hua(Department of Hepatobiliary SurgeryⅡ,Zhujiang Hospital of Southern Medical University,Guangzhou 510280,China;Department of Hepatobiliary Surgery,Nanfang Hospital of Southern Medical University,Guangzhou 510515,China)

机构地区：[1]南方医科大学珠江医院肝胆二科,广州510280 [2]南方医科大学南方医院肝胆外科,广州510515

出　　处：《解放军医学杂志》2020年第5期509-512,共4页Medical Journal of Chinese People's Liberation Army

基　　金：广东省自然科学基金(2018A030313214)。

摘　　要：目的探讨双mTORC1/2抑制剂AZD2014诱导的自噬对人肝癌细胞株HCCLM3增殖的影响。方法体外培养人肝癌细胞株HCCLM3,培养过程中分别加入AZD2014、雷帕鸣或3-甲基腺嘌呤(3-MA),使终浓度分别为100 nmol/L、100 nmol/L和10 mmol/L。研究AZD2014对HCCLM3肝癌细胞自噬的影响时,按处理因素分为对照组、雷帕鸣组、AZD2014组、3-MA组、雷帕鸣+3-MA组和AZD2014+3-MA组,采用MDC染色法检测HCCLM3肝癌细胞中自噬囊泡的变化;研究AZD2014对HCCLM3肝癌细胞中自噬相关蛋白表达的影响时,按处理因素分为对照组、雷帕鸣组和AZD2014组,采用Western blotting检测HCCLM3肝癌细胞中自噬相关蛋白LC3B-Ⅱ和Beclin-1的表达;研究AZD2014诱导的自噬对HCCLM3肝癌细胞增殖的影响时,按处理因素分为对照组、3-MA组、AZD2014组和AZD2014+3-MA组,采用CCK-8法检测HCCLM3肝癌细胞的增殖情况。结果MDC染色显示,与对照组比较,AZD2014处理后,HCCLM3肝癌细胞中自噬囊泡的数量较多、体积较大。Western blotting检测结果显示,AZD2014处理后,HCCLM3肝癌细胞中LC3B-Ⅱ和Beclin-1表达水平均明显高于对照组(P<0.05)。CCK-8法检测结果显示,自噬抑制剂(3-MA)可部分逆转AZD2014对HCCLM3肝癌细胞增殖的抑制作用。结论双mTORC1/2抑制剂AZD2014可通过诱导自噬抑制HCCLM3肝癌细胞的增殖,为AZD2014在肝癌分子靶向治疗中的临床应用提供了理论依据。Objective To investigate the effect of mTORC1/2 inhibitor AZD2014 induced autophagy on cell proliferation in HCCLM3 cell lines in vitro.Methods HCCLM3 cells were cultured in the presence or absence of AZD2014 (100 nmol/L) or rapamycin (100 nmol/L) or 3-methyladenine (10 nmol/L).To detect autophagy vacuoles,HCCLM3 cells were divided into the control group,rapamycin group,AZD2014 group,control+3-MA group,rapamycin+3-MA group,and AZD2014+3-MA group,and auto-fluorescent dye monodansylcadaverine (MDC) was used to monitor autophagy vacuoles.To semi-quantify the expression of autophagy-related proteins,HCCLM3 cells were divided into the control group,rapamycin group,and AZD2014 group,and Western blotting analysis was carried out to detect the expression of autophagy-related proteins,LC3B-II and Beclin-1.To evaluate the effect of autophagy induced by AZD2014 on cell proliferation,HCCLM3 cells were divided into the control group,3-MA group,AZD2014 group,and AZD2014+3-MA group,Cell Counting Kit-8 was used.Results AZD2014-treated HCC cells showed an increase in the number of MDC-labeled vacuoles as well as in their size.AZD2014 treatment increased the levels of LC3B-Ⅱ and Beclin-1 (P<0.05).CCK-8 assay revealed that suppression of cell proliferation elicited by AZD2014 in HCCLM3 was partly attenuated by 3-MA (autophagy inhibitor).Conclusion Dual mTORC1/2 inhibitor AZD2014 suppressed cell proliferation in the HCCLM3 cell line with increased autophagy,in vitro.This study provides a potential basis for further investigation of AZD2014 as a clinical molecular targeted agent for HCC treatment.

关 键 词：肝细胞癌 MTOR抑制剂 自噬 细胞增殖 AZD2014 

分 类 号：R735.7[医药卫生—肿瘤]