Notch1信号通路对肺腺癌细胞侵袭和顺铂耐药的影响及其机制  

Effects of Notch1 signaling pathway on invasion and cisplatin resistance of lung adenocarcinoma cells and their mechanisms

作　　者：陈立松[1] 房惠 王鹤霏 何程远 盖晓东[1] 历春[1] CHEN Lisong;FANG Hui;WANG Hefei;HE Chengyuan;GAI Xiaodong;LI Chun(Laboratory of Molecular Medicine,Basic Medical College,Beihua University,Jilin 132013,China;Department of Gynecology,Third Hospital of Xiang Ya,Central South University,Changsha 410013,China)

机构地区：[1]北华大学基础医学院分子医学实验室,吉林吉林132013 [2]中南大学湘雅第三医院妇科,湖南长沙410013

出　　处：《吉林大学学报（医学版）》2021年第5期1171-1177,共7页Journal of Jilin University：Medicine Edition

基　　金：吉林省科技厅自然科学基金项目(YDZJ202101ZYTS089);吉林省卫健委重点实验室项目(2020J017)。

摘　　要：目的:观察阻断Notch1信号通路对肺腺癌细胞侵袭、血管生成和上皮-间质转化(EMT)及其对顺铂敏感性的影响,阐明Notch1信号通路在肺腺癌侵袭和转移及顺铂耐药中的作用及相关机制。方法:以γ-分泌酶抑制剂(DAPT)作为Notch1信号通路阻断剂作用A549细胞,分为0μmol·L^(-1)DAPT组(对照组)、10μmol·L^(-1)DAPT组和20μmol·L^(-1)DAPT组。采用Transwell小室实验检测各组A549细胞中侵袭细胞数,酶联免疫吸附测定(ELISA)法检测各组细胞培养上清液中基质金属蛋白酶9(MMP-9)和血管内皮生长因子(VEGF)水平,Western blotting法检测各组A549细胞中Notch1、Hes1、波形蛋白(Vimentin)、N钙黏附蛋白(N-cadherin)和E钙黏附蛋白(E-cadherin)蛋白表达水平,CCK-8法检测顺铂(0.625~5.000 mg·L^(-1))作用各组A549细胞的生长抑制率,Western blotting法检测0和1.250 mg·L^(-1)顺铂单独作用A549细胞后Notch1和Hes1蛋白表达水平以及其单独和联合10μmol·L^(-1)DAPT作用A549细胞后P-糖蛋白(P-gp)和多药耐药相关蛋白1(MRP1)蛋白表达水平。结果:与对照组比较,10和20μmol·L^(-1)DAPT组A549细胞中侵袭细胞数明显减少(P<0.05或P<0.01),细胞培养上清液中MMP-9和VEGF水平明显降低(P<0.05或P<0.01),A549细胞中Notch1、Hes1、Vimentin和N-cadherin蛋白表达水平明显降低(P<0.05或P<0.01),E-cadherin蛋白表达水平明显升高(P<0.05)。与对照组比较,10和20μmol·L^(-1)DAPT组A549细胞的生长抑制率明显升高(P<0.05或P<0.01)。与0 mg·L^(-1)顺铂组比较,1.250 mg·L^(-1)顺铂组A549细胞中Notch1、Hes1、P-gp和MRP1蛋白表达水平明显升高(P<0.05或P<0.01);与1.250 mg·L^(-1)顺铂组比较,1.250 mg·L^(-1)顺铂联合10μmol·L^(-1)DAPT组A549细胞中P-gp和MRP1蛋白表达水平明显降低(P<0.01)。结论:阻断Notch1信号通路可抑制肺腺癌细胞的侵袭并降低其对顺铂的耐药作用。Objective:To observe the effects of blocking Notch1 signaling pathway on the cell invasion,angiogenesis,epithelial mesenchymal transition(EMT),and chemosensitivity to cisplatin of the lung adenocarcinoma cells,and to elucidate the role and related mechanism of Notch1 signaling pathway in the cell invasion,metastasis and cisplatin resistance of lung adenocarcinoma.Methods:The A549 cells were treated with DAPT(N-[N-(3,5-Difluorophenacetyl)-l-alanyl]-S-phenylglycine t-butyl ester)as an inhibitor of Notch1 signaling pathway and divided into 0μmol·L^(-1)DAPT group(control group),10μmol·L^(-1)DAPT group and 20μmol·L^(-1)DAPT group.Transwell chamber assay was used to detect the number of invasion cells of A549 cells in various groups;ELISA assay was used to detect the levels of matrix metalloproteinase-9(MMP-9)and vascular endothelial growth factor(VEGF)in the supernatant of cells in various groups;Western blotting method was performed to detect the expression levels of Notch1,Hes1,Vimentin,N-cadherin and E-cadherin proteins in the cells in various groups;CCK-8 method was used to detect the inhibitory rates of growth of the A549 cells after treated with cisplatin(0.625-5.000 mg·L^(-1))in various groups;Western blotting method was used to detect the expression levels of Notch1 and Hes1 proteins in the A549 cells after treated with 0 and 1.250 mg·L^(-1)cisplatin alone,and the expression levels of P-glycoprotein(P-gp)and multidrug resistance-associated protein 1(MRP1)proteins in the A549 cells after treated with 1.250 mg·L^(-1)cisplatin alone or combination with 10μmol·L^(-1)DAPT.Results:Compared with control group,the number of invasion cells in 10 and 20μmol·L^(-1)DAPT groups was significantly decreased(P<0.05 or P<0.01),the levels of MMP-9 and VEGF in in the cell supernatant were significantly decreased(P<0.05 or P<0.01),the expression levels of Notch1,Hes1,Vimentin and N-cadherin proteins in the A549 cells were significantly decreased(P<0.05 or P<0.01),and the expression levels of E-cadherin protein were s

关 键 词：肺肿瘤 NOTCH1 细胞侵袭 细胞转移 顺铂 多药耐药 

分 类 号：R734.2[医药卫生—肿瘤]