微小RNA-21在小鼠肺成纤维细胞炎症反应中的表达变化及意义

赵红，姚平波，戴利，何军，董素素，张平

(南华大学护理学院，湖南衡阳421001)

摘要：目的观察小鼠肺成纤维细胞炎症反应中微小RNA-21(miR-21)及其下游靶基因含Ⅰ型血小板结合蛋白基序的解聚蛋白样金属蛋白酶(ADAMTS-1)的表达变化,并探讨其临床意义。方法体外培养小鼠NIH3T3成纤维细胞，分为对照组和TGF-β1组。TGF-β1组加入终浓度为5 μg/L的TGF-β1，分别作用24、48、72、96、120 h。对照组不加入TGF-β1。收集两组上清液，采用ELISA法测定TNF-α、IL-1β、IL-6水平，应用qRT-PCR技术检测miR-21及ADAMTS-1的表达。结果TGF-β1组24、48、72、96、120 h时TNF-α、IL-1β、IL-6水平及miR-21表达均高于对照组，ADAMTS-1表达均低于对照组(P均<0.05或0.01)。结论 在TGF-β1诱导的大鼠肺成纤维细胞的炎症反应中，miR-21表达增高、ADAMTS-1表达降低；miR-21可成为预防和治疗肺纤维化新的分子靶点。

关键词：转化生长因子β1；微小RNA；炎症；成纤维细胞；肺

doi：10.3969/j.issn.1002-266X.2015.42.004

中图分类号：R563.9文献标志码：A

基金项目：湖南省自然科学基金资助项目(14JJ7044)；湖南省教育厅高等学校科学研究项目优秀青年项目(14B156)。

作者简介：第一赵红(1978-)，女，讲师，研究方向为呼吸内科疾病的诊断与治疗。E-mail: zhaohong779900@163.com

收稿日期：(2015-08-10)

Expression changes of microRNAs-21 in inflammation reaction of

lung fibroblast cells of mice and the significance

ZHAOHong,YAOPing-bo,DAILi,HEJun,DONGSu-su,ZHANGPing

(SchoolofNursing,UniversityofSouthChina,Hengyang421001,China)

Abstract:ObjectiveTo investigate the expression changes of microRNAs-21 (miR-21) and its downstream target genes (a disintegrin and metalloprotease with thrombospond in type 1 motifs, ADAMTS-1) in inflammation reaction of lung fibroblasts cells of mice and the clinical significance. MethodsThe fibroblast cell line NIH3T3 of mic+9e were cultured in vitro and then were divided into the control group and the TGF-β1 group. The TGF-β1 group was added with 5 μg/L TGF-β1 for stimulating 24, 48, 72, 96 and 120 h. The control group was not treated. The supernatant of the two groups was collected at different time points. Using the enzyme linked immunosorbent assay (ELISA) to detect the levels of tumor necrosis factor-α (TNF-α), inflammatory cytokine interleukin-1β (IL-1β) and interleukin-6 (IL-6). The cells were collected at different time points. We used the real-time fluorescence quantitative polymerase chain reaction (PCR) to detect the expression of miR-21 and ADAMTS-1. Results The expression levels of TNF-a , IL-1β, IL-6 and miR-21 in the TGF-β1 groups at 24, 48, 72, 96 and 120 h were higher than those of the control group; the expression level of ADAMTS-1 was lower than that of the control group (all P<0.05 or P<0.01). Conclusions In the TGF-β1-induced inflammatory responses of lung fibroblast cells of mice, the miR-21 expression is up-regulated, while the ADAMTS-1 expression is down-regulated; miR-21 may be a new molecular target for the prevention and treatment of pulmonary fibrosis

Key words: transforming growth factor -β1; microRNAs-21; inflammation; fibroblast cells; lung

微小RNA(miRNA)是一类能够在转录后水平调控基因表达的内源性非编码RNA[1]。近年研究发现，miRNA参与了机体免疫及炎症反应的调节，其中miRNA-21(以下简称miR-21)与炎症反应密切相关[2]。但miR-21是否参与肺成纤维细胞的炎症反应尚不明确。2014年1月～2015年6月，我们检测了在转化生长因子-β1(TGF-β1)诱导的小鼠肺成纤维细胞炎症反应中，miR-21及其下游靶基因含Ⅰ型血小板结合蛋白基序的解聚蛋白样金属蛋白酶(ADAMTS-1)的变化，探讨miR-21在肺纤维化炎症反应中的作用。

1材料与方法

1.1材料小鼠胚胎成纤维细胞NIH3T3购于中国科学院细胞库；DMEM培养基购于美国Sigma-Aldrich公司；胎牛血清购于美国Gibco公司；……