微小RNA-29b通过转化生长因子-β1/Smad信号通路在间质性肺疾病A549细胞上皮-间质转化中的作用机制研究

Mechanistic studies on the role of microRNA-29b in epithelial-mesenchymal transition of A549 cells in interstitial lung disease through the transforminggrowth factor-β1/Smad signalling pathway

目的 探讨微小RNA(miR)-29b通过调控转化生长因子(TGF)-β1/Smad信号通路对间质性肺疾病(ILD)A549细胞增殖、迁移、侵袭和上皮-间质转化(EMT)的影响机制。方法 将A549细胞分为空白对照组、TGF-β1诱导组(TGF-β1)、miR-29b过表达组(TGF-β1+转染miR-29b mimic)和NC组(TGF-β1+转染miR-NC)。采用实时荧光定量PCR(qRT-PCR)检测miR-29b及TGF-β1 mRNA表达水平;采用CCK-8、划痕实验与Transwell检测细胞增殖活力、迁移和侵袭能力;采用Western blot检测TGF-β/Smad通路相关蛋白表达水平。结果 与空白对照组比较,TGF-β1诱导组和NC组miR-29b表达水平、各时间点细胞增殖抑制率下降,TGF-β1 mRNA表达水平、侵袭细胞数量、划痕愈合率、TGF-β1、p-Smad2、N-cadherin、Vimentin蛋白相对表达水平均上升;与TGF-β1诱导组和NC组比较,miR-29b过表达组miR-29b表达水平、细胞增殖抑制率均升高,TGF-β1 mRNA表达水平、侵袭细胞数量和划痕愈合率、TGF-β1、p-Smad2、N-cadherin、Vimentin蛋白相对表达水平均降低(P<0.05)。结论 miR-29b可通过调节TGF-β1/Smad信号通路阻止EMT进程而抑制A549细胞增殖、迁移、侵袭。

Objective To investigate the mechanism of microRNA(miR)-29b's effect on proliferation,migration,invasion and epithelial-mesenchymal transition(EMT)of A549 cells in interstitial lung disease(ILD)through the regulation of transforming growth factor(TGF)-β1/Smad signalling pathway.Methods A549 cells were divided into blank control group,TGF-β1 induced group(TCF-β1),miR-29b overexpression group(TGF-β1+transfected with miR-29b mimic)and NC group(TGF-β1+transfected with miR-NC).Real-time fluorescence quantitative PCR(qRT-PCR)was used to detect miR-29b and TCF-β1 mRNA expression levels;CCK-8,scratch assay and Transwell were used to detect cell proliferation viability,migration and invasive ability;Western blot was used to detect the expression levels of TGF-β1/Smad pathway related proteins.Results Compared with the blank control group,miR-29b expression level and cell proliferation inhibition rate at all time points were decreased in TGF-β1 group and NC group,and TGF-β1 mRNA expression level,the number of invasive cells,the rate of scratch healing and the relative expression levels of TGF-β1,p-Smad2,N-cadherin,and Vimentin proteins increased(P<0.05).Compared with TGF-β1 group and NC group,miR-29b overexpression group had increased miR-29b expression levels and cell proliferation inhibition rate,TGF-β1 mRNA expression levels,the number of invading cells and scratch healing rate,TGF-β1,p-Smad2,N-cadherin,and Vimentin protein relative expression levels were decreased(P<0.05).Conclusion miR-29b can inhibit A549 cell proliferation,migration,and invasion by blocking the EMT process through regulating the TCF-β1/Smad signalling pathway.