上皮-间充质转化在高氧诱导大鼠支气管肺发育不良模型中的作用及机制研究  

Role and mechanism of epithelial-mesenchymal transition in a rat model of bronchopulmonary dysplasia induced by hyperoxia exposure

作　　者：林雅婷 颜崇兵 洪文超 蔡成 龚小慧[1] LIN Ya-Ting;YAN Chong-Bin;HONG Wen-Chao;CAI Cheng;GONG Xiao-Hui(Department of Neonatology,Shanghai Children's Hospital,Shanghai Jiao Tong University School of Medicine,Shanghai 200062,China)

机构地区：[1]上海交通大学医学院附属儿童医院/上海市儿童医院新生儿科,上海200062

出　　处：《中国当代儿科杂志》2024年第7期765-773,共9页Chinese Journal of Contemporary Pediatrics

基　　金：上海市“科技创新行动计划”医学创新研究专项基金。

摘　　要：目的探讨上皮-间充质转化(epithelial-mesenchymal transition,EMT)在大鼠支气管肺发育不良(bronchopulmonary dysplasia,BPD)模型中的作用及机制。方法实验分为两部分。(1)将48只早产大鼠随机分为常氧组和高氧组,每组24只。高氧组暴露于85%氧气中建立早产大鼠BPD模型,常氧组置于同一室内常压空气中,于实验第1、4、7、14天收集早产大鼠肺组织标本。(2)将大鼠肺泡Ⅱ型上皮细胞系(RLE-6TN)随机分为常氧组(常规空气中培养)和高氧组(在95%氧气中培养),分别于高氧暴露后12 h、24 h、48 h收集细胞标本进行检测。使用苏木精-伊红染色法观察早产大鼠肺泡化情况,免疫荧光检测早产大鼠肺组织和RLE-6TN细胞肺表面活性蛋白C(surfactant protein C,SPC)和α-平滑肌肌动蛋白(α-smooth muscle actin,α-SMA)的共定位。实时荧光定量聚合酶链反应和蛋白免疫印迹法检测早产大鼠肺组织和RLE-6TN细胞EMT相关mRNA和蛋白表达水平。结果(1)与常氧组相比,高氧暴露7 d开始高氧组早产鼠可见肺泡化阻滞和肺泡结构简单化改变;肺组织中SPC和α-SMA存在明显共定位现象,高氧暴露7 d、14 d时,与常氧组比较,高氧组SPC表达减少,α-SMA表达增加;高氧暴露7 d、14 d时,与常氧组比较,高氧组TGF-β1、α-SMA、N-钙黏蛋白mRNA和蛋白表达水平升高(P<0.05),SPC、E-钙黏蛋白mRNA和蛋白表达水平降低(P<0.05)。(2)高氧暴露24 h、48 h时,与常氧组比较,高氧组SPC表达减少,α-SMA表达增加;高氧暴露48 h时,与常氧组比较,高氧组SPC、E-钙黏蛋白mRNA和蛋白表达降低(P<0.05),TGF-β1、α-SMA、N-钙黏蛋白mRNA和蛋白表达升高(P<0.05)。结论EMT破坏了BPD早产大鼠肺泡上皮细胞之间的紧密连接,造成肺泡结构简单化和发育异常,参与了BPD的发生发展。Objective To investigate the role and mechanism of epithelial-mesenchymal transition(EMT)in a rat model of bronchopulmonary dysplasia(BPD).Methods The experiment consisted of two parts.(1)Forty-eight preterm rats were randomly divided into a normoxia group and a hyperoxia group,with 24 rats in each group.The hyperoxia group was exposed to 85%oxygen to establish a BPD model,while the normoxia group was kept in room air at normal pressure.Lung tissue samples were collected on days 1,4,7,and 14 of the experiment.(2)Rat type II alveolar epithelial cells(RLE-6TN)were randomly divided into a normoxia group(cultured in air)and a hyperoxia group(cultured in 95%oxygen),and cell samples were collected 12,24,and 48 hours after hyperoxia exposure.Hematoxylin-eosin staining was used to observe alveolarization in preterm rat lungs,and immunofluorescence was used to detect the co-localization of surfactant protein C(SPC)andα-smooth muscle actin(α-SMA)in preterm rat lung tissue and RLE-6TN cells.Quantitative real-time polymerase chain reaction and protein immunoblotting were used to detect the expression levels of EMT-related mRNA and proteins in preterm rat lung tissue and RLE-6TN cells.Results(1)Compared with the normoxia group,the hyperoxia group showed blocked alveolarization and simplified alveolar structure after 7 days of hyperoxia exposure.Co-localization of SPC andα-SMA was observed in lung tissue,with decreased SPC expression and increasedα-SMA expression in the hyperoxia group at 7 and 14 days of hyperoxia exposure compared to the normoxia group.In the hyperoxia group,the mRNA and protein levels of TGF-β1,α-SMA,and N-cadherin were increased,while the mRNA and protein levels of SPC and E-cadherin were decreased at 7 and 14 days of hyperoxia exposure compared to the normoxia group(P<0.05).(2)SPC andα-SMA was observed in RLE-6TN cells,with decreased SPC expression and increasedα-SMA expression in the hyperoxia group at 24 and 48 hours of hyperoxia exposure compared to the normoxia group.Compared to the normoxia g

关 键 词：支气管肺发育不良 上皮-间充质转化 肺泡Ⅱ型上皮细胞 紧密连接 早产大鼠 

分 类 号：R722.6[医药卫生—儿科]