吴茱萸碱抑制NOD1通路诱导肝癌HepG2和SMMC-7721细胞凋亡  

Evodiamine induces apoptosis in SMMC-7721 and HepG2 cells by inhibiting NOD1 signal pathway

作　　者：郭星娴 李晓朋[1] 吕晓婷[1] 陈益[1] 周鹏 吕艳伟 李静[1] 陈地龙[1,2] GUO Xing-xian;LI Xiao-peng;LYU Xiao-ting;CHEN Yi;ZHOU Peng;LYU Yan-wei;LI Jing;CHEN Di-long(Lab of Stem Cell and Tissue Engineering,Dept of Histology and Embryology,Chongqing Medical University,Chongqing 400016,China;Chongqing Three Gorges Medical College,Chongqing 400016,China)

机构地区：[1]重庆医科大学基础医学院组织细胞工程与干细胞研究室,组织学与胚胎学教研室,重庆400016 [2]三峡医药高等专科学校,重庆400016

出　　处：《中国药理学通报》2018年第11期1588-1593,共6页Chinese Pharmacological Bulletin

基　　金：国家自然科学基金资助项目(No31271368)。

摘　　要：目的探讨吴茱萸碱诱导肝癌HepG2和SMMC-7721细胞凋亡的机制。方法以肝癌HepG2和SMMC-7721细胞为研究对象,CCK-8法检测吴茱萸碱对细胞增殖的影响;Hoechst 33258染色法在显微镜下观察细胞凋亡现象的改变;定量逆转录-聚合酶链反应(qRT-PCR)检测NOD1在正常肝细胞HL-7702与肝癌细胞中的表达; Western blot检测NOD1、NF-κB p65、p-p65,以及凋亡相关蛋白Bcl-2、Bax、p53的表达。结果 CCK-8结果显示,给予吴茱萸碱(0、0. 25、0. 5、1、2、4μmol·L-1) 12、24、48 h后,Hep G2和SMMC-7721细胞抑制率明显增高,并且呈剂量和时间依赖性; Hoechst33258染色结果显示,经吴茱萸碱诱导后,Hep G2和SMMC-7721细胞出现核固缩、核边集等形态学改变; qRT-PCR结果显示,与正常肝细胞HL-7702相比,NOD1在肝癌细胞中的表达明显较高; Western blot结果显示,吴茱萸碱可下调NOD1、p-p65和Bcl-2蛋白水平,上调Bax、p53蛋白表达水平,对p65表达水平无影响。结论吴茱萸碱通过下调NOD1,抑制NF-κB的活性,激活p53信号通路,改变Bcl-2/Bax的比值,诱导肝癌Hep G2和SMMC-7721细胞的凋亡。Aim To investigate the mechanisms of evodiamine in the apoptosis of hepatocellular carcinoma(HCC)cell lines HepG2 and SMMC-7721.Methods Cell counting kit-8(CCK-8)assay was performed to detect proliferation activity of HepG2 and SMMC-7721 cells treated with evodiamine.Hoechst 33258 staining was used for observing morphological changes of apoptosis.qRT-PCR was applied to detect the expression levels of NOD1 in normal liver cells and HCC cells.Western blot was employed to examine the expression of NOD1,NF-κB p65,p-p65,Bcl-2,Bax and p53 proteins.Results The proliferation of HepG2 and SMMC-7721 cells was significantly inhibited by evodiamine treatment for 12 h,24 h and 48 h at concentrations of 0.25,0.5,1,2,4μmol·L^-1,which indicated the inhibitory effect of evodiamine was in dose-dependent and time-dependent manners.Hoechst 33258 staining showed the typical apoptotic morphology in HCC cells treated with evodiamine,such as increased nuclear chromatin concentration and edge accumulation.qRT-PCR showed that NOD1 expression levels in HCC cells were significantly higher than those in normal cell line HL-7702.Western blot showed that the increased expression of Bax and p53,the decreased expression of NOD1,p-p65,Bcl-2 and no change of p65 in HepG2 and SMMC-7721 cells treated with evodiamine.Conclusions Evodiamine can induce the apoptosis of HepG2 and SMMC-7721 cells by down-regulating NOD1,inhibiting the activation of NF-κB,and activating p53 signal pathway,which mediates Bcl-2/Bax balance.

关 键 词：吴茱萸碱 NOD1 肝癌 HEPG2 SMMC-7721 凋亡 

分 类 号：R284.1[医药卫生—中药学] R322.47[医药卫生—中医学]