Cheng Zhe, Dai Ling-Ling, Wang Xi, Jia Liu-Qun, Jing Xiao-Gang, Li Peng-Fei, Liu Meng, Wang Huan, An Lin
Department of Respiratory and Critical Care Medicine, Institute of Clinical Medicine, The First Affiliated Hospital of Zhengzhou University, Zhengzhou 450052, P.R. China.
Oncotarget. 2017 Jul 11;8(28):46312-46325. doi: 10.18632/oncotarget.17933.
This study aims to explore how microRNA-145 (miR-145) affects airway remodeling and cytokine expression by targeting epidermal growth factor receptor (EGFR) to regulate mucin 5AC (MUC5AC).Mice alveolar epithelial cells (AECs) were divided into a control, blank, miR-145 mimics, mimic control, miR-145 inhibitors, inhibitor control, si-EGFR and miR-145 inhibitors + si-EGFR group. Asthma mice models with airway remodeling were induced with an Ovalbumin (OVA) solution and randomly divided into a normal, asthma, asthma + miR-145 mimics, asthma + miR-145 mimic control, asthma + si-EGFR or asthma + si-EGFR NC group. Airway remodeling degree and histomorphology was measured using hematoxylin-eosin (HE), Masson and periodic acid-Schiff (PAS) staining. Flow cytometry was used to detect Th2 and Th17 cells in peripheral blood, ELISA was used to measure inflammatory factors. qRT-PCR and western blotting was adapted to detect the expressions of EGFR and the relevant cytokines that are regulated by miR-145.The control, miR-145 mimics and si-EGFR groups showed a higher expression of miR-145 and a lower expression of EGFR and cytokines than the blank, mimic control, inhibitor control and miR-145 inhibitor + si-EGFR groups. Mice in the asthma + miR-145 mimics and asthma + si-EGFR groups showed lower WAt/Pbm, WAi/Pbm and WAm/Pbm, less inflammatory cells, less airway modeling and alleviated goblet cell hyperplasia and mucus obstruction than the asthma group. Furthermore, the expressions of EGFR and cytokines of transfected cells and lung tissues were negatively related to those of miR-145. MiR-145 can down-regulate MUC5AC by negatively targeting EGFR and thereby relieving airway remodeling.
本研究旨在探讨微小RNA-145(miR-145)如何通过靶向表皮生长因子受体(EGFR)来调节黏蛋白5AC(MUC5AC),从而影响气道重塑和细胞因子表达。将小鼠肺泡上皮细胞(AECs)分为对照组、空白组、miR-145模拟物组、模拟物对照组、miR-145抑制剂组、抑制剂对照组、si-EGFR组和miR-145抑制剂+si-EGFR组。用卵清蛋白(OVA)溶液诱导建立气道重塑的哮喘小鼠模型,并随机分为正常组、哮喘组、哮喘+miR-145模拟物组、哮喘+miR-145模拟物对照组、哮喘+si-EGFR组或哮喘+si-EGFR阴性对照组。采用苏木精-伊红(HE)、Masson和过碘酸-希夫(PAS)染色法检测气道重塑程度和组织形态学。采用流式细胞术检测外周血中Th2和Th17细胞,采用酶联免疫吸附测定(ELISA)法检测炎症因子。采用实时定量聚合酶链反应(qRT-PCR)和蛋白质免疫印迹法检测EGFR及受miR-145调控的相关细胞因子的表达。与空白组、模拟物对照组、抑制剂对照组和miR-145抑制剂+si-EGFR组相比,对照组、miR-145模拟物组和si-EGFR组中miR-145表达较高,EGFR和细胞因子表达较低。与哮喘组相比,哮喘+miR-145模拟物组和哮喘+si-EGFR组小鼠的WAt/Pbm、WAi/Pbm和WAm/Pbm较低,炎症细胞较少,气道重塑较轻,杯状细胞增生和黏液阻塞减轻。此外,转染细胞和肺组织中EGFR和细胞因子的表达与miR-145的表达呈负相关。MiR-145可通过负向靶向EGFR下调MUC5AC,从而减轻气道重塑。