桦木酸对人胃癌MKN-45细胞上皮-间充质转化的影响  

Effects of betulinic acid on epithelial-mesenchymal transition of human gastric cancer MKN-45 cells

作　　者：李怀玉 陈云[3] 胡子毅[4] 万朝星 邓慧灵 叶祯 叶菁 LI Huai-yu;CHEN Yun;HU Zi-yi;WAN Zhao-xing;DENG Hui-ling;YE Zhen;YE Jing(Jiangxi University of Chinese Medicine,Nanchang 330004,China;The First Affiliated Hospital of Guangzhou University of Chinese Medicine,Guangzhou 510405,China;Dept of Gastroenterology,First Affiliated Hospital of Gannan Medical University,Ganzhou,Jiangxi 341002,China;Dept of TCM,the Affiliated Hospital of Jiangxi University of Chinese Medicine,Nanchang 330006,China;School of Clinical Medicine,Jiangxi University of Chinese Medicine,Nanchang 330004,China)

机构地区：[1]江西中医药大学,江西南昌330004 [2]广州中医药大学第一附属医院,广东广州510405 [3]赣南医学院第一附属医院消化科,江西赣州341002 [4]江西中医药大学附属医院中医经典科,江西南昌330006 [5]江西中医药大学临床医学院,江西南昌330004

出　　处：《中国药理学通报》2023年第8期1438-1443,共6页Chinese Pharmacological Bulletin

基　　金：江西省教育厅科学技术研究项目(No GJJ201238);江西省中医药中青年骨干人才培养计划(赣中医药综合字[2020]9号,赣财社指[2020]59号)。

摘　　要：目的观察桦木酸(betulinic acid,BA)对转化生长因子-β1(transforming growth factor beta1,TGF-β1)诱导的人胃癌MKN-45细胞迁移与侵袭能力的影响,并从上皮-间充质转化(epithelial-mesenchymal transition,EMT)方面探讨相关作用机制。方法体外培养MKN-45细胞,采用CCK-8法分别检测24、48和72 h不同浓度BA对MKN-45细胞增殖的影响;通过细胞划痕实验和Transwell侵袭实验观察BA(5、10、20μmol·L^(-1))和TGF-β1抑制剂LY2109761(10μmol·L^(-1))对TGF-β1(10μg·L^(-1))诱导后MKN-45细胞侵袭与转移水平的影响;采用Western blot法检测各组MKN-45细胞中TGF-β1、E-钙黏蛋白(E-Cadherin)、N-钙黏蛋白(N-Cadherin)和基质金属蛋白酶-2(MMP-2)蛋白的表达情况。结果BA能以时间及浓度依赖性地抑制MKN-45细胞的增殖,干预24、48、72 h后的IC 50值分别为212.8、22.72、13.17μmol·L^(-1)。与空白组相比,TGF-β1诱导后MKN-45细胞的划痕迁移(P<0.05)与Transwell侵袭(P<0.01)水平明显增加;MKN-45细胞中TGF-β1(P<0.01)、N-Cadherin(P<0.05)、MMP-2(P<0.01)明显上调,E-Cadherin(P<0.01)明显下调。与TGF-β1组相比,LY2109761能明显逆转TGF-β1诱导的MKN-45细胞迁移与侵袭能力(P<0.01),下调TGF-β1、N-Cadherin、MMP-2(P<0.01),并上调E-Cadherin(P<0.01);BA能呈浓度依赖性地抑制TGF-β1诱导下MKN-45细胞的迁移与侵袭能力(P<0.01),下调TGF-β1(P<0.01)、N-Cadherin(P<0.01)、MMP-2(P<0.05或P<0.01),并上调E-Cadherin(P<0.01)。结论BA能抑制胃癌MKN-45细胞的迁移与侵袭能力,其机制可能与阻滞TGF-β1诱导的EMT进程有关。Aim To observe the effect of betulinic acid(BA)on the migration and invasion of human gastric cancer MKN-45 cells induced by transforming growth factor-β1(TGF-β1),and to explore the effect of BA on epithelial-mesenchymal transition(EMT)and the potential mechanism.Methods The MKN-45 cells were cultivated in vitro,and the effects of different concentrations of BA on the proliferation of MKN-45 cells at 24,48 and 72 h were detected using CCK-8 method.The effects of BA(5,10,20μmol·L^(-1))and TGF-β1 inhibitor LY2109761(10μmol·L^(-1))on MKN induced by TGF-β1(10μg·L^(-1))on the level of invasion and metastasis by were observed by cell scratch assay and Transwell invasion assay.The expressions of E-Cadherin,N-Cadherin and matrix metalloproteinase-2(MMP-2)proteins in MKN-45 cells in each group were detected by Western blotting.Results BA inhibited the proliferation of MKN-45 cells in a time-and concentration-dependent manner,with IC 50 values of 212.8,22.72,and 13.17μmol·L^(-1) after intervention for 24,48,and 72 h,respectively.Compared with the control group,the scratch migration(P<0.05)and Transwell invasion(P<0.01)levels of MKN-45 cells were significantly raised after TGF-β1 induction.TGF-β1 in MKN-45 cells(P<0.01),N-Cadherin(P<0.05)and MMP-2(P<0.01)were significantly up-regulated,and E-Cadherin(P<0.01)was significantly down-regulated.Compared with the TGF-β1 group,LY2109761 could significantly reverse the migration and invasion ability of MKN-45 cells induced by TGF-β1(P<0.01),down-regulate TGF-β1,N-Cadherin,MMP-2(P<0.01),and up-regulate E-Cadherin(P<0.01).5,10,and 20μmol·L^(-1) concentrations of BA could reduce the migration and invasion of MKN-45 cells induced by TGF-β1 in a concentration-dependent manner(P<0.01),down-regulate TGF-β1(P<0.01),N-Cadherin(P<0.01),MMP-2(P<0.05,P<0.01),and up-regulate E-Cadherin(P<0.01).Conclusions BA could inhibit the migration and invasion ability of MKN-45 cells,and its mechanism may be related to the EMT process induced by TGF-β1.

关 键 词：桦木酸 胃癌 上皮-间充质转化 TGF-Β1 迁移 侵袭 

分 类 号：R329.28[医药卫生—人体解剖和组织胚胎学] R73-37[医药卫生—基础医学] R735.2R916.4