miR-433在NIH-3T3细胞纤维化和矽尘诱导的小鼠肺纤维化中的作用  

Role of miR-433 in fibrosis of NIH-3T3 cells and silica-induced pulmonary fibrosis in mice

作　　者：郭义威[1] 孙春莉[1] 王树兴[1] GUO Yi-wei;SUN Chun-li;WANG Shu-xing(Department of Anatomy,Xinxiang Medical University,Xinxiang 453003,China)

机构地区：[1]新乡医学院人体解剖学教研室,河南新乡453003

出　　处：《中国病理生理杂志》2020年第8期1458-1464,共7页Chinese Journal of Pathophysiology

基　　金：国家自然科学基金资助项目(No.81571085)。

摘　　要：目的:本研究旨在探究微小RNA-433(miR-433)在纤维化中的作用及机制。方法:采用TargetScan预测miR-433的潜在靶基因。检测转化生长因子β1(TGF-β1)处理小鼠胚胎成纤维细胞(NIH-3T3细胞)24h后miR-433表达的变化。检测miR-433 mimic对TGF-β1处理的NIH-3T3细胞中p-SMAD2、纤维连接蛋白(FN)、α-平滑肌肌动蛋白(α-SMA)和结缔组织生长因子(CTGF)表达水平的影响。采用CCK-8法和流式细胞术检测转染miR-433 mimic对TGF-β1诱导的NIH-3T3细胞生长和S期阻滞的影响。构建矽尘诱导的小鼠肺纤维化模型并使用agomiR-433进行干预,HE染色和Masson染色观察miR-433对小鼠肺纤维化的影响,采用免疫组化检测小鼠肺组织中α-SMA的表达。结果:miR-433能特异性结合SMAD2的3’-UTR,并抑制其蛋白和mRNA的表达。TGF-β1能下调NIH-3T3细胞中miR-433的表达水平,上调p-SMAD2蛋白的水平,同时也增强FN、α-SMA和CTGF蛋白和mRNA的表达,并增强细胞活力,诱导S期细胞数量增加;而miR-433 mimic能逆转TGF-β1对NIH-3T3细胞活力和S期阻滞的影响。在矽尘诱导的小鼠肺纤维化模型中,agomiR-433能抑制肺纤维化进程,减少小鼠肺组织中α-SMA的表达。结论:miR-433可能通过靶向调控SMAD2干预TGF-β1/SMAD2信号通路,从而参与调节纤维化的病理过程。AIM:To study the role and regulatory mechanism of microRNA-433(miR-433)in fibrosis.METHODS:TargetScan was used to predict the potential target genes of miR-433. The changes of miR-433 expression were detected after transforming growth factor β1(TGF-β1)treatment of mouse embryonic fibroblasts(NIH-3 T3 cells)for24 h. The effects of miR-433 mimic on the expression of p-SMAD2,fibronectin(FN),α-smooth muscle actin(α-SMA)and connective tissue growth factor(CTGF)in TGF-β1 treated cells were examined. The effects of miR-433 mimic transfection on the viability and S-phase fraction of NIH-3 T3 cells induced by TGF-β1 were detected by CCK-8 assay and flow cytometry. A model of silica-induced pulmonary fibrosis in mice was established,and agomiR-433 was used for intervention. HE staining and Masson staining were used to observe the effect of miR-433 on pulmonary fibrosis in the mice. The expression of α-SMA in lung tissues was detected by immunohistochemistry.RESULTS:miR-433 specifically bound to the 3’-UTR of SMAD2 and inhibited its expression at protein and mRNA levels. TGF-β1 down-regulated the expression of miR-433 in NIH-3 T3 cells,up-regulated the protein level of p-SMAD2 and the expression of FN,α-SMA and CTGF at protein and mRNA levels,and increased the viability and the number of S-phase cells. miR-433 mimic reversed the effects of TGF-β1 on NIH-3 T3 cell viability and S-phase arrest. In a model of silica-induced pulmonary fibrosis in mice,agomiR-433 inhibited the progress of pulmonary fibrosis and reduced the expression of α-SMA in mouse lung tissues.CONCLUSION:miR-433 may interfere with TGF-β1/SMAD2 signaling pathway through targeting SMAD2,thus participating in the regulation of fibrosis process.

关 键 词：纤维化 微小RNA-433 NIH-3T3细胞 TGF-β1/SMAD2信号通路 

分 类 号：R363.13[医药卫生—病理学] R363.2[医药卫生—基础医学]