通塞颗粒对AECOPD大鼠气道上皮屏障及EGFR/ERK通路的影响  

Effect of Tongsai granules on airway epithelial barrier in a rat model of acute exacerbation of chronic obstructive pulmonary disease and role of EGFR/ERK signaling pathway

作　　者：魏焱鑫 魏毓 刘新光[1,2] 田燕歌[1,2] 刘学芳[1,2] 赵迪[1,2] 赵鹏 WEI Yanxin;WEI Yu;LIU Xinguang;TIAN Yange;LIU Xuefang;ZHAO Di;ZHAO Peng(Collaborative Innovation Center for Chinese Medicine and Respiratory Diseases Co-constructed by Henan Province&Minis-try of Education,Henan Key Laboratory of Chinese Medicine for Respiratory Disease,Henan University of Chinese Medicine,Zhengzhou 450046,China;Academy of Chinese Medical Sciences,Henan University of Chinese Medicine,Zhengzhou 450046,China)

机构地区：[1]河南中医药大学呼吸疾病中医药防治省部共建协同创新中心,河南省中医药防治呼吸病重点实验室,河南郑州450046 [2]河南中医药大学中医药科学院,河南郑州450046

出　　处：《中国病理生理杂志》2023年第12期2204-2213,共10页Chinese Journal of Pathophysiology

基　　金：国家自然科学基金资助项目(No.82274274;No.82074406);河南省科技研发计划联合基金资助项目(No.22301420070);河南省重点研发与推广专项科技攻关(No.222102310375);郑州市科技协同创新专项(No.2023XTCX045)。

摘　　要：目的:探讨通塞颗粒(TSG)对慢性阻塞性肺疾病急性加重期(AECOPD)大鼠上皮屏障功能损伤的作用及其机制。方法:24只SD大鼠随机分为对照(control)组、模型(model)组、TSG组和莫西沙星(MXF)+沙丁胺醇(STL)组。第1~8周建立COPD大鼠模型,第9周第3天,经鼻滴入肺炎克雷伯杆菌建立AECOPD大鼠模型。第9周的第1~2天和4~7天,control组和model组给予生理盐水灌胃,TSG组和MXF+STL组分别给予TSG和MXF+STL灌胃,灌胃结束后处死大鼠。检测大鼠呼气峰流速(PEF)的变化;HE染色观察大鼠肺组织病理变化;检测肺组织白细胞介素1β(IL-1β)、IL-6、肿瘤坏死因子α(TNF-α)、基质金属蛋白酶2(MMP2)、MMP9、闭锁小带蛋白1(ZO-1)、上皮钙黏素(E-Cad)和闭合蛋白(OCC)蛋白表达。采用香烟烟雾提取物(CSE)刺激人支气管上皮细胞BEAS-2B,给予TSG不同活性部位干预,检测ZO-1、E-cad、OCC、表皮生长因子受体(EGFR)、磷酸化EGFR(p-EGFR)、细胞外信号调节激酶(ERK)和磷酸化ERK(p-ERK)蛋白水平。结果:TSG可显著降低AECOPD大鼠支气管壁厚度、肺泡平均线性截距,以及肺组织IL-1β、IL-6、TNF-α、MMP2和MMP9表达水平,增加平均肺泡数和PEF,上调ZO-1、E-Cad和OCC蛋白表达(P<0.01)。TSG可以显著抑制CSE诱导的BEAS-2B细胞中ZO-1、E-Cad和OCC的表达(P<0.01)。网络药理分析获得TSG2所含化合物对应328个靶点以及AECOPD相关的3864个基因;整合分析显示TSG2缓解AECOPD可能与ERBB2、ERK、EGFR、IL和WNT等信号通路有关。实验结果显示,TSG2可以显著抑制CSE诱导的BEAS-2B细胞中p-EGFR和p-ERK水平的升高(P<0.05或P<0.01)。结论:TSG可以保护气道上皮屏障功能,缓解AECO-PD,其机制可能与抑制EGFR/ERK信号通路有关。AIM:To investigate the effect of Tongsai granules(TSG)on epithelial barrier dysfunction in acute exacerbation of chronic obstructive pulmonary disease(AECOPD)and the underlying mechanism.METHODS:Twenty-four Sprague-Dawley rats were randomly divided into control group,model group,TSG group,and moxifloxacin(MXF)+salbutamol(STL)group.Rat COPD model was established over 8 weeks.On day 3 of week 9,the rats with COPD were intratracheally administered Klebsiella pneumoniae to establish the AECOPD model.On days 1 to 2 and 4 to 7 in week 9,saline was administered via oral gavage to the rats in control and model groups,and the rats in TSG and MXF+STL groups were treated daily with TSG and MXF+STL by gavage,respectively.Peak expiratory flow(PEF),histopatho-logical changes,and the expression levels of interleukin-1β(IL-1β),IL-6,tumor necrosis factor-α(TNF-α),matrix me-talloproteinase 2(MMP2),MMP9,zonula occludens-1(ZO-1),E-cadherin(E-Cad)and occludin(OCC)were deter-mined.Moreover,human bronchial epithelial BEAS-2B cells were exposed to cigarette smoke extract(CSE)and treated with different TSG fractions,and the protein levels of ZO-1,E-Cad,OCC,epidermal growth factor receptor(EGFR),phosphorylated EGFR(p-EGFR),extracellular signal-regulated kinase(ERK)and phosphorylated ERK(p-ERK)were determined.RESULTS:Treatment with TSG significantly reduced bronchial wall thickness,mean linear intercept,and the levels of IL-1β,IL-6,TNF-α,MMP2 and MMP9(P<0.05 or P<0.01),significantly increased mean alveolar number and PEF(P<0.01),and up-regulated the ZO-1,E-Cad and OCC protein levels(P<0.01)in the lungs of AECOPD rats.Treatment with TSG2,the second TSG fraction,increased the protein levels of ZO-1,E-Cad and OCC in a dose-dependent manner in CSE-exposed BEAS-2B cells(P<0.05 or P<0.01).Network pharmacology analysis of 328 targets of the com-pounds in TSG2 and 3864 genes related to AECOPD suggested that TSG2 relieved AECOPD likely through the regulation of ERBB2,ERK,EGFR,IL and WNT signaling pathways.Treatment with TSG2 also inhibited CS

关 键 词：通塞颗粒 慢性阻塞性肺疾病急性加重期 气道上皮屏障 EGFR/ERK信号通路 

分 类 号：R563[医药卫生—呼吸系统] R285.5[医药卫生—内科学] R363[医药卫生—临床医学]