LncRNA FEZF1-AS1通过调控EZH2对肺间质细胞增殖、迁移及侵袭的作用

Effects of lncRNA FEZF1-AS1 on proliferation,migration and invasion through regulating EZH2 of lung interstitial cells

目的研究长链非编码RNA FEZ家族锌指1-反义RNA 1(lncRNA FEZF1-AS1)调控zeste同源物增强子2(EZH2)对肺间质细胞增殖、迁移、侵袭能力及上皮细胞-间质转化(EMT)的影响及其作用机制。方法将人肺腺癌细胞系A549分为对照组(control)和模型组[model,用转化生长因子β1(TGF-β1)20 ng/mL作用48 h,诱导成为肺间质细胞]。用Western blot检测细胞中E-钙黏蛋白(E-cadherin)、N-钙黏蛋白(N-cadherin)及波形蛋白(vimentin)的蛋白表达。RT-qPCR检测细胞中lncRNA FEZF1-AS1和EZH2基因表达。转染组细胞分为转染si NC组、si lncRNA FEZF1-AS1+OE vector组和si lncRNA FEZF1-AS1+OE EZH2组。CCK-8法检测细胞增殖、细胞划痕检测细胞迁移、Transwell小室法检测细胞侵袭;用Western blot检测细胞中E-cadherin、N-cadherin、vimentin及EZH2的蛋白表达,用RNA免疫沉淀(RIP)测定FEZF1-AS1与EZH2的直接结合作用。结果与对照组比较,模型组E-cadherin的蛋白表达水平减少(P<0.05);N-cadherin及vimentin的蛋白表达水平升高(P<0.05);与对照组比较,模型组lncRNA FEZF1-AS1与EZH2基因的表达水平明显升高(P<0.05);与si NC组相比,si lncRNA FEZF1-AS1+OE vector组细胞增殖、迁移、侵袭能力降低,E-cadherin蛋白表达升高,N-cadherin、vimentin、EZH2蛋白表达降低(P<0.05);与si lncRNA FEZF1-AS1+OE vector组比较,si lncRNA FEZF1-AS1+OE EZHZ组细胞增殖、侵袭、迁移能力升高,E-cadherin蛋白表达降低,N-cadherin、vimentin、EZH2蛋白表达升高(P<0.05);RIP实验进一步证实了lncRNA FEZF1-AS1与EZH2具有结合作用。结论LncRNA FEZF1-AS1通过调控EZH2促进肺间质细胞增殖、侵袭、转移和EMT过程。

Objective To investigate the effects of long non-coding RNA FEZ family zinc finger 1 antisense RNA 1(lncRNA FEZF1-AS1)on enhancer of zeste homolog 2(EZH2)in regulation of proliferation,migration,invasion and epithelial-mesenchymal transition(EMT)of pulmonary interstitial cells and its mechanism.Methods The A549 cells human lung adenocarcinoma cell line were divided into control group and model group[model cells were induced into lung interstitial cells after being treated with transforming growth factorβ1(TGF-β1)20 ng/mL for 48 h].The protein expression of E-cadherin,N-cadherin and vimentin in each group was detected by Western blot.The expression of lncRNA FEZF1-AS1 and EZH2 in the two groups was detected by RT-qPCR.Cells in the transfection group were divided into si NC group,lncRNA FEZF1-AS1+OE vector group and si lncRNA FEZF1-AS1+OE EZH2 group.Cell proliferation was examined by CCK-8 method,cell migration was detected by cell scratch,and cell invasion was detected by Transwell assays.The protein expression of E-cadherin,N-cadherin,vimentin and EZH2 in each group was detected by Western blot.The direct binding effect of FEZF1-AS1 and EZH2 was determined by RNA immuno-precipitation(RIP).Results Compared with the control group,the protein expression level of E-cadherin in the model group was significantly decreased(P<0.05),and the protein expression of N-cadherin and vimentin was significantly increased(P<0.05).Compared with the control group,the expression level of lncRNA FEZF1-AS1 and EZH2 genes was significantly increased in the model group(P<0.05).Compared with si NC group,the proliferation,migration and invasion ability of si lncRNA FEZF1-AS1+OE vector group were decreased,the expression of E-cadherin protein was increased while the expression of N-cadherin,vimentin and EZH2 was decreased(P<0.05).Compared with si lncRNA FEZF1-AS1+OE vector group,the proliferation,invasion and migration of si lncRNA FEZF1-AS1+OE EZH2 group were increased(P<0.05).E-cadherin expression was decreased,while N-cadherin,vimentin and EZH2 expressions were increased(P<0.05).RIP experiment further confirmed that lncRNA FEZF1-AS1 had direct binding effect with EZH2.Conclusions LncRNA FEZF1-AS1 can promote the proliferation,invasion,metastasis and EMT process of pulmonary fibrosis cells by regulating EZH2.