MiR-199a-3p/5p靶向DUSP5/MAP3K11调控翼状胬肉成纤维细胞纤维化和迁移
MiR-199a-3p/5p targets DUSP5/MAP3K11 to regulate fibrosis and migration of pterygium fibroblasts
朱阁 1王陈 2吴细英 3石洋 1郑芳 2袁玥琪 1燕星元 1严明1
作者信息
- 1. 武汉大学中南医院眼科 湖北 武汉 430071
- 2. 武汉大学中南医院检验科/基因诊断中心 湖北 武汉 430071
- 3. 武汉爱尔眼科汉口医院 湖北 武汉 430021
- 折叠
摘要
目的:探讨miR-199a-3p/5p及其靶基因DUSP5/MAP3K11在人翼状胬肉成纤维细胞(HPF)中的作用.方法:采用外植体方法从手术切除的翼状胬肉组织中培养原代HPF,实时定量聚合酶链反应(qRT-PCR)验证其表型.MiR-199a-3p/5p模拟物和抑制剂转染HPF.细胞划痕试验、Transwell法测定细胞迁移能力.免疫蛋白印记分析(Western Blot)和qRT-PCR检测靶基因DUSP5/MAP3K11和纤维化标志物的变化.免疫荧光法验证上皮-间质转化(EMT)相关蛋白的胞内定位.结果:与人结膜成纤维细胞相比,HPF的EMT和纤维化表型显著升高.MiRNA-199a-3p/5p模拟物下调DUSP5/MAP3K11,并促进HPF纤维化和EMT,miR-199a-3p/5p抑制剂上调DUSP5/MAP3K11,抑制HPF纤维化、EMT及迁移.结论:MiRNA-199a-3p/5p可能靶向DUSP5/MAP3K11,以调控翼状胬肉成纤维细胞的纤维化、EMT和迁移能力.
Abstract
Objective:To find out the function of miR-199a-3p/5p and its target gene DUSP5/MAP3K11 in human pterygium fibroblasts(HPFs).Methods:Primary HPFs were cultured from surgically ex-cised pterygium tissue using the explant method and their phenotypes were verified by quantitative real-time polymerase chain reaction(qRT-PCR).MiR-199a-3p/5p mimics and inhibitors were transfected with HPFs,and cell migration ability was determined using cell scratch assay and Transwell.Western Blot and qRT-PCR were used to detect the changes in target genes DUSP5/MAP3K11 and fibrosis markers.Immunofluorescence was used to verify the intracellular localization of epithelial-mesenchymal transition(EMT)-associated proteins.Results:Elevated EMT and fibrosis phenotype were was found in HPFs significantly as compared with that of human conjunctiva fibroblasts.MiR-NA-199a-3p/5p mimic down-regulated DUSP5/MAP3K11,and promoted HPFs'fibrosis and EMT,while miR-199a-3p/5p inhibitor up-regulated DUSP5/MAP3K11,and inhibitd HPFs'fibrosis and EMT and migration.Conclusion:MiRNA-199a-3p/5p may target DUSP5/MAP3K11 to regu-late fibrosis,EMT,and migration ability of pterygium fibroblasts.
关键词
翼状胬肉/纤维化/再上皮化/上皮-间质转化/miR-199a/迁移Key words
Pterygium/Fibrosis/Re-Epithelialization/Epithelial-Mesenchymal Transition/miR-199a/Migration引用本文复制引用
基金项目
国家自然科学基金资助项目(81770898)
武汉大学中南医院2019科技创新培育基金资助项目(znpy2019062)
出版年
2024
NETL
NSTL
万方数据