肺岩宁下调转录因子Snail表达抑制人肺腺癌A549细胞EMT发生及侵袭转移  

Feyanning formula down-regulated the expression of transcription factor Snail and inhibited EMT occurrence,invasion and metastasis of human lung adenocarcinoma A549 cells

作　　者：张亚辉 蔡玥娇[1] 邓海滨[1] 王中奇[1] 徐振晔[1] Zhang Yahui;Cai Yuejiao;Deng Haibin;Wang Zhongqi;Xu Zhenye(Longhua Hospital,Shanghai University of Traditional Chinese Medicine,Shanghai 200032,China)

机构地区：[1]上海中医药大学附属龙华医院,上海200032

出　　处：《世界科学技术-中医药现代化》2023年第4期1239-1249,共11页Modernization of Traditional Chinese Medicine and Materia Medica-World Science and Technology

基　　金：上海市教育委员会科研创新计划重大项目(NO.2017-01-07-00-10-E00064):源于精气理论的肺岩宁方抗肺癌生长与转移作用机制的研究,负责人:邓海滨

摘　　要：目的研究肺岩宁抑制人肺腺癌A549细胞侵袭转移的分子机制。方法体外培养A549细胞,细胞计数试剂盒(Cell counting kit 8,CCK-8)检测不同浓度的肺岩宁对A549细胞生长增殖的影响;细胞形态学观察和免疫印迹实验(Western blot)检测转化生长因子β1(Transforming growth factor-β1,TGF-β1)对A549细胞上皮间质转化(Epithelial mesenchymal transformation,EMT)发生的影响;Western blot、实时荧光定量聚合酶链式反应(Real time quantitative polymerase chain reaction,RT-qPCR)实验检测TGF-β1诱导前后,肺岩宁对A549细胞EMT标志蛋白及mRNA表达变化;划痕实验、Transwell实验检测TGF-β1诱导前后,肺岩宁对A549细胞侵袭和迁移能力的影响变化;构建稳定过表达Snail的A549细胞株,Western blot、RT-qPCR实验检测稳定过表达锌指转录因子(Snail)后,肺岩宁对A549细胞EMT标志蛋白及mRNA表达变化;划痕实验、Transwell实验检测稳定过表达Snail后,肺岩宁对A549细胞侵袭和迁移能力的影响变化。结果不同浓度的肺岩宁处理后,A549细胞的生长受到不同程度的抑制(P<0.01);10 ng·mL^(-1)的TGF-β1可以诱导A549细胞EMT发生;与对照组比较,TGF-β1诱导后,A549细胞侵袭迁移能力增加(P<0.01),而与TGF-β1组比较,肺岩宁能够抑制A549细胞的侵袭迁移能力(P<0.01)、上调A549细胞中E钙黏蛋白(E-cadherin)及Ecadherin mRNA的表达(P<0.01),下调N钙黏蛋白(N-cadherin)、波形蛋白(Vimentin)和Snail及其mRNA的表达(P<0.05),抑制A549细胞EMT发生;与TGF-β1诱导组比较,稳定过表达Snail能够减弱肺岩宁对A549细胞的侵袭转移能力的抑制作用(P<0.01)、减弱肺岩宁对A549细胞中E-cadherin及其mRNA表达的上调作用(P<0.01)、N-cadherin、Snail(P<0.01)Vimentin(P>0.05)和及其mRNA表达的下调作用。结论肺岩宁方可能通过下调转录因子Snail的表达抑制A549细胞EMT发生及其侵袭转移。Objective To study the molecular mechanism of inhibiting invasion and metastasis of human lung adenocarcinoma A549 cells by anti-cancer Jingfang Feyanning formula.Methods A549 cells were cultured in vitro,and the effects of different concentrations of Feyanning on the growth and proliferation of A549 cells were detected by CCK-8 experiment.Cell morphological observation and Western blot detection of Transforming growth factor-β1(TGF-β1)on Epithelial mesenchymal transformation(EMT)in A549 cells.Western blot and RT-qPCR were used to detect the expression of EMT marker protein and mRNA in A549 cells induced by Feiyanning formula before and after TGF-β1 induction.Scratch assay and Transwell assay were performed to detect the effects of Feiyanning formula on invasion and migration of A549 cells before and after TGF-β1 induction.Western blot and RT-qPCR were used to detect the expression of EMT marker protein and mRNA of Feyanning formula in A549 cells after stable Snail overexpression.The effect of Feyanning formula on invasion and migration of A549 cells after stable Snail expression was detected by scratch assay and Transwell assay.Results The growth of A549 cells was inhibited by different concentrations of Feiyaning formula(P<0.01);TGF-β1 at 10 ng·mL^(-1)could induce EMT in A549 cells.Compared with the control group,the invasion and migration ability of A549 cells increased after TGF-β1 induction(P<0.01),while compared with TGF-β1 group,Feyanning formula could inhibit the invasion and migration of A549 cells(P<0.01)and up-regulate the expression of E-cadherin and its mRNA in A549 cells(P<0.01)but down-regulated the expression of N-cadherin,Vimentin,Snail and their mRNA(P<0.05),and inhibited the occurrence of EMT in A549 cells.Compared with the TGF-β1-induced group,Stable overexpression of Snail could attenuate the inhibition of Feyanning formula on the invasion and metastasis ability of A549 cells(P<0.01),and attenuate the up-regulation of E-cadherin and its mRNA expression in A549 cells(P<0.01),N-cadheri

关 键 词：肺岩宁方 肺癌 EMT 侵袭转移 SNAIL 

分 类 号：R285.5[医药卫生—中药学]