左旋棉酚通过下调X连锁凋亡抑制蛋白增强表柔比星抑制肝细胞癌的体外作用  

(-)-Gossypol enhances epirubicin’s anti-tumor effects on hepatocellular carcinoma in vitro via down-regulating X-linked inhibitor of apoptosis protein

作　　者：蒋文斌[1] 孙梁博 杜烨湘 肖云华 黎伯胜[1] 闫小晶[1] 吴亚冉[2] 连继勤[2] 何凤田[1] JIANG Wenbin;SUN Liangbo;DU Yexiang;XIAO Yunhua;LI Bosheng;YAN Xiaojing;WU Yaran;LIAN Jiqin;HE Fengtian(Department of Biochemistry and Molecular Biology,College of Basic Medical Sciences,Army Medical University(Third Military Medical University),Chongqing,400038,China;Department of Clinical Biochemisty,Faculty of Pharmacy and Laboratory Medicine,Army Medical University(Third Military Medical University),Chongqing,400038,China;Department of Nuclear Medicine,First Affiliated Hospital,Army Medical University(Third Military Medical University),Chongqing,400038,China)

机构地区：[1]陆军军医大学(第三军医大学)基础医学院生物化学与分子生物学教研室,重庆400038 [2]陆军军医大学(第三军医大学)药学与检验医学系临床生物化学教研室,重庆400038 [3]陆军军医大学(第三军医大学)第一附属医院核医学科,重庆400038

出　　处：《陆军军医大学学报》2022年第16期1599-1605,共7页Journal of Army Medical University

基　　金：重庆市博士后自然科学基金项目(cstc2021jcyj-bshX0200);重庆市博士后特别资助项目(2021XM2050);国家自然科学基金面上项目(81872024)。

摘　　要：目的探讨左旋棉酚增强表柔比星杀伤肝细胞癌(hepatocellular carcinoma,HCC)细胞的作用及可能机制。方法将HCC细胞(Hep3B及HuH7)分为溶剂对照组、左旋棉酚处理组、表柔比星处理组、联合用药组、对照siRNA+表柔比星组、沉默X连锁凋亡抑制蛋白(X-linked inhibitor of apoptosis protein,XIAP)+表柔比星处理组、对照质粒+联合用药组、过表达XIAP+联合用药组,采用CCK-8方法检测各组细胞存活情况;流式细胞术检测各组细胞凋亡情况;Western blot检测剪切型聚(腺苷二磷酸-核糖)聚合酶[cleaved poly(ADP-ribose)polymerase,Cle-PARP]水平、XIAP水平及真核起始因子4E(eukaryotic initiation factor 4E,eIF4E)磷酸化水平。结果与溶剂对照相比,低浓度的左旋棉酚(5μmol/L)单独处理对HCC细胞存活无明显抑制作用(P>0.05),而将左旋棉酚与表柔比星联合使用后可显著增强表柔比星对HCC细胞的杀伤(P<0.05)、促进HCC细胞的凋亡及对剪切型PARP的诱导,同时还能抑制表柔比星诱导的XIAP上调及eIF4E磷酸化。此外,表柔比星抑制HCC细胞存活及升高剪切型PARP的能力能被沉默XIAP所增强,而被过表达XIAP所抑制。结论左旋棉酚可增强表柔比星抑制HCC细胞存活促进细胞凋亡的能力,其机制可能与抑制XIAP上调和eIF4E磷酸化有关,提示左旋棉酚可作为表柔比星治疗HCC的潜在增敏剂。Objective To investigate mechanism and effect on(-)-gossypol enhancing epirubicin killing hepatocellular carcinoma(HCC)cells.Methods HCC cells(Hep 3 B and HuH7)were divided into 8 groups:control,(-)-gossypol,epirubicin,combination,si-control+epirubicin,si-X-linked inhibitor of apoptosis protein(XIAP)+epirubicin,empty vector+combination and pEBB-XIAP+combination groups.Cell viability of each group was detected by CCK-8 assay;Cell apoptosis of each group was detected by flow cytometry and the level of cleavaged PARP,XIAP and eIF4 E phosphorylation was detected by Western blot assay.Results Compared with the control group,no major effect was observed on cell viability exposed to(-)-gossypol in 5μmol/L(P>0.05).Combination of(-)-gossypol and epirubicin significantly enhanced the anti-survival effect on HCC cells(P<0.05),promoted cells apoptosis,increased the induction of cleavaged PARP,and inhibited XIAP up-regulation and eIF4 E phosphorylation induced by epirubicin in HCC cells.Silencing XIAP promoted epirubicin to kill HCC cells and induced PARP cleavaged;Furthermore,XIAP over-expression attenuated epirubicin-induced anti-survival and pro-apoptosis of HCC cells enhanced by(-)-gossypol.Conclusion The combination of(-)-gossypol and epirubicin significantly inhibits HCC cells’survival and promotes cell apoptosis via suppressing epirubicin-induced phosphorylation of eIF4 E and up-regulation of XIAP,suggesting that(-)-gossypol holds promise as a sensitizer for epirubicin-based HCC therapy.

关 键 词：表柔比星 左旋棉酚 肝细胞癌 X连锁凋亡抑制蛋白 真核起始因子4E 

分 类 号：R735.7[医药卫生—肿瘤] R966[医药卫生—临床医学] R979.1