MicroRNA-21对肺成纤维细胞增殖和分化的影响  

Effect of microRNA-21 on proliferation and differentiation of pulmonary fibroblasts of mice

作　　者：曹嫚[1] 赵红[1] 何军[1] 戴利[1] 殷小成[1] 姚平波[1] 

机构地区：[1]南华大学附属南华医院儿科,湖南衡阳421001

出　　处：《天津医药》2015年第10期1137-1140,共4页Tianjin Medical Journal

基　　金：湖南省自然科学基金资助项目(14JJ7044);湖南省教育厅青年项目(14B156)

摘　　要：目的探讨小鼠肺成纤维化细胞模型中microRNA（miR）-21对肺成纤维细胞增殖和分化的影响。方法24只SPF级C57BIJ6小鼠随机分为假手术组和肺纤维化模型组，各12只。经气管内注入博莱霉素建立小鼠肺纤维化模型，采用荧光定量PCR检测各组肺组织miR-21的表达。提取小鼠成纤维细胞，胰酶消化，接种于6孔板，使细胞浓度达到30％～50％。设随机对照组、空白对照组和miR-21mimic组。各组分别在50μL Opti-MEM加入2．5μLPBS、阴性对照储存液和miR-21mimic储存液（20μmol／L）。采用细胞计数试剂盒-8（CCK-8）法检测细胞增殖活性，蛋白质印迹法检测成纤维细胞含I型血小板结合蛋白基序的解聚蛋白样金属蛋白酶（ADAMTS-1）和转化生长因子（TGF）-β1的表达。结果与假手术组比较，肺纤维化模型组小鼠肺组织中miR-21的表达量明显上调。与随机对照组和空白对照组比较，miR-21mimic组成纤维细胞miR-21基因表达显著上调，成纤维细胞增殖率增加，ADAMTS-1蛋白表达明显下降，而TGF-β1表达显著上调；空白对照组与随机对照组ADAMTS-1及TGF-β1蛋白表达无明显差异。结论在小鼠肺成纤维化细胞模型中上调的miR-21可通过ADAMTS-1／TGF-β1信号通路促进肺纤维化。Objective To investigate the effect of microRNA（miR）-21 on proliferation and differentiation of routine pulmonary fibroblasts. Methods C57BL/6 mice of SPF grade （n=24） were randomly divided into Sham group and Pulmo- nary fibrosis model group with 12 mice in each group. Pulmonary fibrosis model was established by trans-tracheal jet ventila- tion of bleomycin into mice. The transcription levels of miR-21 were examined by quantitative real-time PCR in various pul- monary fibrosis tissues. Primary fibroblast were isolated and digested by Trypsin then inoculated into 6 well plate to reach confluence of 30%-50%. PBS （2.5 μL）, negative control stock solution and miR-21 mimic stock solution （20 μmol/L） were added into Opti-MEM （50 μL） as control group, blank group and miR-21 mimic group respectively.The cell viability was as- sessed by CCK-8. Expressions of ADAMTS-1 and TGF-β1 in the pulmonary fibroblasts were tested using Western blot. Re- sults The expression of miR-21 was significantly increased in lungs of mice in pulmonary fibrosis model group than that in sham group. Expression of miR-21 was higher in miR-21 mimic group than that in control group and blank group. Expres- sion of miR-21 was significantly higher with better cell viability in miR-21 mimic group than that in control group and blank group. The expression of ADAMTS-1 was significantly decreased in miR-2 l mimic group, while the expression of TGF-β1, a target gene of miR-21, was significantly increased in miR-21 mimic group compared with the other two groups. There is no significant different in expressions of ADAMTS- 1 and TGF-β1 between control group and blank group. Conclusion Over- expression of miR-21 in pulmonary fibroblasts disrupts TGF-β1 signaling pathway by reducing expression of ADAMTS-1, which promotes the proliferation and differentiation of pulmonary fibroblast.

关 键 词：肺纤维化 成纤维细胞 转化生长因子Β1 微RNAS 细胞增殖 细胞分化 ADAMTS-1 MICRORNA-21 

分 类 号：R563[医药卫生—呼吸系统]