RSV感染HBE细胞后EGFR、FOXA2、MUC5AC表达变化及罗格列酮干预作用  

Changes in EGFR,FOXA2 and MUC5AC expression in respiratory syncytial virus-infected human bronchial epithelium cells and effects of rosiglitazone intervention

作　　者：丁洁 董翌玮 叶文静 李嫦嫦 赵伟 陈小芳 杨康康 李海燕 董琳 Ding Jie;Dong Yiwei;Ye Wenjing;Li Changchang;Zhao Wei;Chen Xiaofang;Yang Kangkang;Li Haiyan;Dong Lin(Children′s Respiratory Department,the Second Affiliated Hospital and Yuying Children′s Hospital of Wenzhou Medical University,Wenzhou 325000,China;Department of Allergy and Immunology,the Affiliated Hospital of Virginia Commonwealth University School of Medicine,Virginia 23284,USA)

机构地区：[1]温州医科大学附属第二医院、育英儿童医院儿童呼吸科,325000 [2]美国弗吉尼亚联邦大学医学院附属医院变态反应和免疫科,23284

出　　处：《中华微生物学和免疫学杂志》2019年第5期358-364,共7页Chinese Journal of Microbiology and Immunology

基　　金：温州市科技局对外合作项目(H20140002).

摘　　要：目的探讨在呼吸道合胞病毒(RSV)感染人支气管上皮(HBE)细胞中表皮生长因子受体(epidermal growth factor receptor,EGFR)-叉头转录因子A2(forkhead transcription factor A2,FOXA2)通路参与黏液分泌的机制,以及过氧化物酶增殖激活受体γ(PPARγ)激动剂罗格列酮、拮抗剂(GW9662)和EGFR抑制剂(AG1478)对该通路的干预作用。方法建立HBE细胞RSV体外感染模型,随机分成6组:A组(AG1478+RSV)、B组(罗格列酮+RSV)、C组(GW9662+RSV)、D组(RSV)、E组(0.1%DMSO+HBE细胞)、F组(HBE细胞对照)。RSV感染HBE细胞造模前2 h,A、B、C组分别予10μmol/L的AG1478、罗格列酮、GW9662预处理。A^D组以RSV感染吸附HBE细胞2 h后加维持液继续培养,各组分别培养至12 h、24 h、48 h收获细胞及上清液。应用RT-PCR检测各组EGFR、PPARγ、FOXA2 mRNA表达情况,Western blot检测p-EGFR及EGFR蛋白水平,ELISA检测上清液中黏蛋白5AC(MUC5AC)水平。结果与F组相比,3个时间点A、B、C、D组的EGFR mRNA、p-EGFR/EGFR蛋白比值、MUC5AC蛋白表达量均明显增加,随时间增加表达量增加,在48 h达到最高;PPARγmRNA均有所增加,其中A和B组表达量随时间逐渐增加,于48 h达到高峰,但D组逐渐减少;FOXA2 mRNA均有降低,其中D组表达量最低。与D组相比,A组和B组EGFR mRNA、p-EGFR/EGFR蛋白比值、MUC5AC蛋白表达量明显降低,FOXA2 mRNA明显升高。结论RSV感染导致MUC5AC蛋白表达增加,PPARγ和EGFR-FOXA2信号通路参与RSV感染后气道黏液高分泌过程。Objective To investigate the mechanism of epidermal growth factor receptor-forkhead transcription factor A2(EGFR-FOXA2)pathway-involved high secretion of mucus in human bronchial epithelium(HBE)cells after respiratory syncytial virus(RSV)infection and to evaluate the effects of intervention using agonist(rosiglitazone)and antagonist(GW9662)of peroxidase proliferation activated receptorγ(PPARγ)and EGFR inhibitor(AG1478).Methods HBE cells were randomly divided into six groups:A group(AG1478+RSV),B group(rosiglitazone+RSV),C group(GW9662+RSV),D group(RSV),E group(0.1%dimethyl sulfoxide DMSO)and F group(HBE cell control group).Two hours before RSV infection,A,B and C groups were respectively treated with 10μmol/L of AG1478,rosiglitazone and GW9662.Expression of EGFR,PPARγand FOXA2 at mRNA level in each group was detected by real-time fluorescence quantitative RT-PCR 12 h,24 h and 48 h after HBE cells were infected with or without RSV.Expression of phosphorylated-EGFR(p-EGFR)and EGFR at protein level was detected by Western blot.ELISA was performed to measure the expression of mucin-5AC(MUC5AC).Results Compared with F group,EGFR expression at mRNA level,p-EGFR/EGFR protein ratio and MUC5AC expression at protein level were increased in a time-dependent manner in A,B,C and D groups at 12 h,24 h and 48 h.Compared with group F,the expression of PPARγat mRNA level in A,B,and D groups increased at each time point.Moreover,PPARγexpression gradually increased over time in A and B groups,reaching the peaks at 48 h,but was in decline in D group.Expression of FOXA2 at mRNA level in RSV-infected HBE cells was declined at each time point compared with that in group F,especially in D group.Compared with group D,A and B groups showed significantly decreased EGFR expression at mRNA level,p-EGFR/EGFR protein ratio and MUC5AC expression at protein level,but markedly increased FOXA2 expression at mRNA level.Conclusions RSV infection increased the expression of MUC5AC at protein level in HBE cells.PPARγand EGFR-FOXA2 signaling pat

关 键 词：呼吸道合胞病毒 黏蛋白5AC 罗格列酮 表皮生长因子受体 叉头框蛋白A2 

分 类 号：R373[医药卫生—病原生物学]