FUT8调控TGF-β1介导的肺成纤维细胞纤维化的机制  

FUT8 modulates galectin-3 expression to regulate TGF-β1-mediated fibrosis of lung fibroblasts

作　　者：高崴崴 刘待见[1] 张晓萍[1] 冯青青[1] 刘颖[1] GAO Weiwei;LIU Daijian;ZHANG Xiaoping;FENG Qingqing;LIU Ying(Department of Respiratory and Critical Care Medicine,Second Affiliated Hospital of Zhengzhou University,Zhengzhou 450000,China)

机构地区：[1]郑州大学第二附属医院呼吸与危重症医学科,河南郑州450000

出　　处：《南方医科大学学报》2022年第8期1166-1173,共8页Journal of Southern Medical University

基　　金：河南省医学科技攻关计划联合共建项目(LHGJ20200413)。

摘　　要：目的探讨α-1,6岩藻糖基转移酶(FUT8)对人胚肺成纤维细胞(MRC-5)增殖、迁移和纤维化的作用及可能机制。方法将24只C57/BL6小鼠随机分为对照组、博莱霉素组、sh-NC组和sh-FUT8组,应用Masson染色观察肺纤维化情况。细胞实验分别设置对照组:MRC-5正常培养;TGF-β1组:TGF-β1处理MRC-5;si-NC组:si-NC转染MRC-5后TGF-β1处理;si-FUT8组:siFUT8转染MRC-5后TGF-β1处理;Gal-3组:si-FUT8和pcDNA3.1-Gal转染MRC-5后TGF-β1处理。CCK-8和BrdU方法检测细胞增殖;划痕实验检测细胞迁移;RT-qPCR和Western blot检测α-平滑肌肌动蛋白(α-SM A)、I型胶原蛋白(COLIA1)和细胞外基质纤维连接蛋白(FN)的表达水平。同时,免疫共沉淀检测了FUT8与半乳糖凝集素-3(Gal-3)的作用及沉默FUT8对FAK/Akt信号通路的调节。结果沉默FUT8显著降低博莱霉素诱导的小鼠肺组织细胞外胶原沉积。沉默FUT8抑制TGF-β1介导的细胞增殖(186.81±6.29 vs 118.09±9.48,P<0.05)和迁移。FUT8缺失下调α-SMA、COLIA1和FN的mRNA和蛋白表达水平(P<0.05)。此外,FUT8可直接与Gal-3作用。沉默FUT8下调Gal-3的表达并抑制FAK/Akt信号通路,过表达Gal-3逆转FUT8对细胞增殖、迁移和纤维化的作用(P<0.05)。结论FUT8通过调控Gal-3调控TGF-β1介导的MRC-5细胞增殖、迁移和纤维化,FAK/Akt信号通路可能参与了这个过程。Objective To investigate the regulatory role ofα-1,6-fucosyltransferase(FUT8)in TGF-β1-induced proliferation,migration and fibrosis of human embryonic lung fibroblasts(MRC-5 cells)and explore the underlying molecular mechanism.Methods C57/BL6 mice were randomized into 4 groups for treatment with saline(control group),bleomycin,bleomycin+sh-NC or bleomycin+sh-FUT8,and pulmonary fibrosis was observed using Masson staining.MRC-5 cells were transfected with si-NC,FUT8 siRNA(si-FUT8),or both si-FUT8 and a galectin-3(Gal-3)overexpression plasmid(pcDNA3.1-Gal)prior to TGF-β1 treatment,and the changes in cell proliferation and migration were assessed using CCK-8 assay,BrdU assay,and wound healing assay;the changes in the expression levels ofα-SMA,collagen I(COLIA1)and extracellular matrix fibronectin(FN)were detected with real-time quantitative PCR(RT-qPCR)and Western blotting.The interaction of FUT8 and Gal-3 was tested using coimmunoprecipitation(Co-IP)assay,and the effect of FUT8 silencing on Gal-3 and FAK/Akt signaling pathways was analyzed.Results FUT8 knockdown significantly reduced bleomycin-induced extracellular collagen deposition in the lung tissues of the mice.Silencing FUT8 obviously inhibited cell proliferation(P<0.05)and migration mediated by TGF-β1.FUT8 knockdown down-regulated the mRNA and protein levels ofα-SMA,COLIA1 and FN(P<0.05)in the cells.Coimmunoprecipitation analysis showed that FUT8 interacted with Gal-3.Silencing FUT8 significantly down-regulated Gal-3expression and inhibited the activation of the FAK/Akt signaling pathway(P<0.05).Overexpression of Gal-3 obviously reversed the effects of FUT8 silencing on cell proliferation,migration and fibrosis(P<0.05).Conclusion FUT8 regulates TGF-β1-induced proliferation,migration and fibrosis of MRC-5 cells by modulating Gal-3 expression,in which the FAK/Akt pathway may play a role.

关 键 词：FUT8 GALECTIN-3 增殖 迁移 纤维化 

分 类 号：R563[医药卫生—呼吸系统]