Chen Li-Jun, Ye Hong, Zhang Qian, Li Feng-Zhi, Song Lin-Jie, Yang Jie, Mu Qing, Rao Shan-Shan, Cai Peng-Cheng, Xiang Fei, Zhang Jian-Chu, Su Yunchao, Xin Jian-Bao, Ma Wan-Li
Department of Respiratory and Critical Care Medicine, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, Hubei, China.
Department of Pathophysiology, School of Basic Medicine, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, Hubei, China; Key Laboratory of Pulmonary Diseases, Ministry of Health of China, Wuhan, Hubei, China.
Toxicol Appl Pharmacol. 2015 Mar 1;283(2):75-82. doi: 10.1016/j.taap.2015.01.004. Epub 2015 Jan 14.
Idiopathic pulmonary fibrosis (IPF) is a chronic progressive lung disease characterized by the development of subpleural foci of myofibroblasts that contribute to the exuberant fibrosis. Recent studies revealed that pleural mesothelial cells (PMCs) undergo epithelial-mesenchymal transition (EMT) and play a pivotal role in IPF. In animal model, bleomycin induces pulmonary fibrosis exhibiting subpleural fibrosis similar to what is seen in human IPF. It is not known yet whether bleomycin induces EMT in PMCs. In the present study, PMCs were cultured and treated with bleomycin. The protein levels of collagen-I, mesenchymal phenotypic markers (vimentin and α-smooth muscle actin), and epithelial phenotypic markers (cytokeratin-8 and E-cadherin) were measured by Western blot. PMC migration was evaluated using wound-healing assay of culture PMCs in vitro, and in vivo by monitoring the localization of PMC marker, calretinin, in the lung sections of bleomycin-induced lung fibrosis. The results showed that bleomycin induced increases in collagen-I synthesis in PMC. Bleomycin induced significant increases in mesenchymal phenotypic markers and decreases in epithelial phenotypic markers in PMC, and promoted PMC migration in vitro and in vivo. Moreover, TGF-β1-Smad2/3 signaling pathway involved in the EMT of PMC was demonstrated. Taken together, our results indicate that bleomycin induces characteristic changes of EMT in PMC and the latter contributes to subpleural fibrosis.
特发性肺纤维化（IPF）是一种慢性进行性肺部疾病，其特征是肌成纤维细胞在胸膜下灶积聚，导致过度纤维化。最近的研究表明，胸膜间皮细胞（PMC）经历上皮-间质转化（EMT），并在IPF中起关键作用。在动物模型中，博来霉素可诱导肺纤维化，表现出与人类IPF相似的胸膜下纤维化。目前尚不清楚博来霉素是否能诱导PMC发生EMT。在本研究中，培养PMC并用博来霉素进行处理。通过蛋白质印迹法检测I型胶原蛋白、间充质表型标志物（波形蛋白和α-平滑肌肌动蛋白）以及上皮表型标志物（细胞角蛋白-8和E-钙黏蛋白）的蛋白水平。使用体外培养的PMC伤口愈合试验评估PMC迁移，并通过监测博来霉素诱导的肺纤维化小鼠肺组织切片中PMC标志物钙视网膜蛋白的定位，在体内评估PMC迁移。结果显示，博来霉素可诱导PMC中I型胶原蛋白合成增加。博来霉素可显著增加PMC中间充质表型标志物水平，降低上皮表型标志物水平，并促进体外和体内的PMC迁移。此外，还证实了TGF-β1-Smad2/3信号通路参与了PMC的EMT过程。综上所述，我们的结果表明，博来霉素可诱导PMC发生EMT特征性变化，且后者导致胸膜下纤维化。
