BTB-CNC异体同源体1抑制尿道上皮细胞中转化生长因子-β信号通路的激活从而阻断微小RNA-155-5p诱导的炎性反应  

BTB and CNC homology1 blocks microRNA-155-5p-induced inflammatory response by inhibiting transforming growth factor-βsignaling pathway in urothelial cells

作　　者：孟庆松[1] 张明[1] 贾江华[1] 薛文勇[1] 齐进春[1] Meng Qingsong;Zhang Ming;Jia Jianghua;Xue Wenyong;Qi Jinchun(Department of Urology,the Second Hospital of Hebei Medical University,Shijiazhuang 050000,China)

机构地区：[1]河北医科大学第二医院泌尿外科,石家庄050000

出　　处：《中华实验外科杂志》2022年第6期1058-1061,共4页Chinese Journal of Experimental Surgery

基　　金：河北省省级科技计划(20377724D)。

摘　　要：目的观察尿道上皮细胞中BTB-CNC异体同源体1(BACH1)对转化生长因子-β(TGF-β)信号通路的影响, 从而参与微小RNA(miR)-155-5p诱导的炎症和纤维化的调节。方法采用实时定量反转录聚合酶链反应(RT-qPCR)和蛋白质印迹法(Western blot)检测BACH1敲低或过表达后SV-HUC-1细胞中的TGF-β信号通路TGF-β、信号转导分子7(Smad7)和信号转导分子3(Smad3)的信使RNA(mRNA)和蛋白水平。我们在转染了miR-155-5p的细胞中过表达了BACH1, 酶联免疫吸附试验(ELISA)测定肿瘤坏死因子-α(TNF-α)、白细胞介素(interleukin, IL)-1β和IL-6水平, RT-qPCR检测血管内皮生长因子(VEGF)、纤维连接蛋白(FN)、Ⅳ型胶原(Collagen Ⅳ)、TGF-β、Smad3和Smad7的表达水平。采用单因素方差分析比较两组间差异。结果 TGF-β和Smad3的mRNA表达水平(2.14±0.11比0.99±0.07, F=238.581;1.78±0.06比0.96±0.08, F=201.720, P<0.01), 和蛋白表达水平(1.36±0.08比1.0±0.08, F=32.911;1.34±0.04比1.01±0.08, F=39.361, P<0.05)在BACH1敲低后升高, 在BACH1过表达后下降(mRNA水平分别为0.39±0.06比1.09±0.04, F=246.369;0.56±0.09比1.04±0.11, F=36.668;蛋白水平分别为0.31±0.12比1.00±0.07, F=81.323;0.63±0.05比0.98±0.14, F=16.026, P<0.01), 而Smad7的mRNA水平(0.50±0.07比1.01±0.06, F=105.446)和蛋白水平(0.40±0.08比1.00±0.11, F=58.835)被BACH1小干扰RNA(siRNA)下调, P<0.01, 在BACH1过表达后上调(mRNA水平为1.67±0.11比1.05±0.09, F=55.960;蛋白水平为1.80±0.16比0.99±0.10, F=52.571, P<0.01)。由miR-155-5p mimics引起的IL-1β、IL-6和TNF-α水平的增加因BACH1过表达而减弱(497.60±16.85比287.83±14.89比310.30±13.77, F=171.691;795.17±34.84比396.53±24.59比425.67±21.39, F=195.307;764.43±27.05比401.67±23.06比454.9±24.76, F=184.088, P<0.01)。由miR-155-5p mimics诱导的VEGF、FN和Collagen Ⅳ的上调也被BACH1过表达阻断(1.90±0.10比0.95±0.05比1.21±0.05, F=111.524;2.03±0.11比0.98±0.09比1.11±0.07, F=121.346;1.91±0.11比1.11±0.06Objective To observe the effect of BTB and CNC homology1(BACH1)on transforming growth factor-β(TGF-β)signaling pathway in urothelial cells,thereby participating in the regulation of microRNA(miR)-155-5p-induced inflammation and fibrosis.Methods We used quantitative polymerase chain reaction(qPCR)and Western blotting to detect the mRNA and protein levels of TGF-β,Smad7 and Smad3 of TGF-βsignaling pathways in SV-HUC-1 cells after BACH1 knockdown or overexpression.We overexpressed BACH1 in cells transfected with miR-155-5p,and measured IL-1β,IL-6,and tumor necrosis factor-α(TNF-α)levels by enzyme linked immunosorbent assay(ELISA),and detected the expression level of vascular endothelial growth factor(VEGF),fibronectin(FN),collagenⅣ,TGF-β,Smad3,and Smad7 by qPCR.Results The mRNA expression levels of TGF-βand Smad3(2.14±0.11 vs.0.99±0.07,F=238.581;1.78±0.06 vs.0.96±0.08,F=201.720,P<0.01),and protein expression levels(1.36±0.08 vs.1.0±0.08,F=32.911;1.34±0.04 vs.1.01±0.08,F=39.361,P<0.05)increased after BACH1 knockdown and decreased after BACH1 overexpression(mRNA levels:0.39±0.06 vs.1.00±0.04,F=246.369;0.56±0.09 vs.1.04±0.11,F=36.668 and protein levels:0.31±0.12 vs.1.00±0.07,F=81.323;0.63±0.05 vs.0.98±0.14,F=16.026,P<0.01),and the mRNA level(0.50±0.07 vs.1.01±0.06,F=105.446)and protein level(0.40±0.08 vs.1.00±0.11,F=58.835)of Smad7 were down-regulated by BACH1 small interfering RNA(siRNA)(P<0.01),but up-regulated after BACH1 overexpression(mRNA level:1.67±0.11 vs.1.05±0.09,F=55.960,and protein level:1.80±0.16 vs.0.99±0.10,F=52.571,P<0.01).The IL-1β,IL-6,and TNF-αlevels increased by miR-155-5p mimics,but attenuated by BACH1 overexpression(497.60±16.85 vs.287.83±14.89 vs.310.3±13.77,F=171.691;795.17±34.84 vs.396.53±24.59 vs.425.67±21.39,F=195.307;764.43±27.05 vs.401.67±23.06 vs.454.9±24.76,F=184.088,P<0.01).The upregulation of VEGF,FN and collagenⅣinduced by miR-155-5p mimics was also blocked by BACH1 overexpression(1.90±0.10 vs.0.95±0.05 vs.1.21±0.05,F=111.524;2.03±0.1

关 键 词：微小RNA-155-5p 尿道狭窄 炎性因子 

分 类 号：R69[医药卫生—泌尿科学]