TGF-β和ERK/GSK3β信号通路激活与辐射诱导放射性肺炎大鼠肺泡上皮细胞间充质转化的关系  

Relationship between activation of TGF-β and ERK/GSK3β signaling pathway and radiation-induced mesenchymal transformation of alveolar epithelial cells in rats with radioactive pneumonia

作　　者：谭化 季辉[2] 李娜 张秀敬 贾营 Tan Hua;Ji Hui;Li Na;Zhang Xiujing;Jia Ying(Cadre Health Department of Shijiazhuang Third Hospital,Shijiazhuang 050061,China;Department of Pulmonary Diseases,Cangzhou Hospital of Integrated Traditional Chinese and Western Medicine,Shijiazhaung 061001,China)

机构地区：[1]河北省石家庄市第三医院干部保健科,石家庄050061 [2]河北省沧州中西医结合医院肺病科,沧州061001

出　　处：《广西医科大学学报》2019年第12期1909-1914,共6页Journal of Guangxi Medical University

基　　金：河北省石家庄市卫健委资助项目(No.181461153)

摘　　要：目的:探究转化生长因子(TGF-β)和细胞外调节蛋白激酶(ERK)/糖原合成酶激酶3β(GSK3β)信号通路激活与辐射诱导放射性肺炎大鼠肺泡上皮细胞间充质转化的关系。方法:6至8周龄的SD大鼠购自上海实验动物中心。90只实验大鼠随机分为3组:对照组(为正常饲养大鼠,每次只进行麻醉处理,n=30),放射诱导组(每次麻醉后进行钴-60伽马射线处理,处理持续两个星期,n=30)和U0126组(先试试辐射诱导再进行U0126腹膜内注射,n=30)。通过透射电子显微镜观察肺泡上皮细胞的超微结构变化。采用PCR测量上皮间充质转化(EMT)相关蛋白mRNA的表达,免疫荧光分析Snail的易位潜力。ELISA测定法测量TGF-β、Smad2和Smad3浓度。采用蛋白质印迹测定ERK和GSK3β的磷酸化和总蛋白表达。结果:对照组中的肺泡上皮细胞含有丰富的线粒体,结构正常,而辐射诱导引起肺泡上皮细胞的超微结构改变,包括许多嵴断裂的线粒体,细胞器减少,以及液泡的增加。对照组中的大鼠肺泡上皮细胞显示出经典的上皮形态。辐射诱导后处理后,形态变为细长纺锤状。与对照组相比,辐射诱导组波形蛋白和α-SMA的mRNA水平增加,但E-钙粘蛋白mRNA的表达降低(P<0.05)。与对照组相比,辐射增加了Snail(绿色荧光)的信号,并且大部分信号位于细胞核中。与对照组相比,辐射诱导组TGF-β、Smad2和Smad3含量显著增加(P<0.05)。辐射诱导增加了p-ERK和p-GSK3β的磷酸化水平(P<0.05),但ERK和GSK3β蛋白质水平没有差异(P>0.05)。U0126抑制ERK的磷酸化并有效地消除了辐射诱导的GSK3β磷酸化。此外,阻断ERK信号传导还抑制了辐射诱导的Snail蛋白水平和α-SMA的增加(P<0.05)。相反,阻断ERK信号传导增加了辐射后E-钙粘蛋白的表达(P<0.05)。结论:辐射导致ERK信号传导途径的激活。活化的ERK1/2引起GSK3β的磷酸化,导致GSK3β和Snail的解离。未结合的Snail迁移到细胞核并作为E-钙粘蛋�Objective:To investigate the relationship between the activation of TGF-βand ERK/GSK3βsignaling pathway and radiation-induced mesenchymal transformation of alveolar epithelial cells in rats with radioactive pneumonia.Methods:Six to eight weeks old Sprague-Dawley(SD)rats were randomly divided into 3 groups:control group(n=30),radiation-induction group(n=30)and U0126 group(n=30,intraperitoneal injection of U0126 after radiation-induction).The ultrastructural changes of alveolar epithelial cells were observed by transmission electron microscopy.The PCR method was used to detect the epithelial mesenchymal transition(EMT)related-protein mRNA expression.Immunofluorescence was used to analyze the translocation potential of Snail.TGF-β,Smad2 and Smad3 concentrations were measured by ELISA assay.Western blotting was performed to determine the phosphoryla tion and total protein expression of ERK and GSK3.Results:Alveolar epithelial cells in the control group were rich in mitochondria and normal in structure,while radiation induced ultrastructural changes in alveolar epithelial cells,including many mitochondria with sputum rupture,decreased organelles,and increased vacuoles.Rat alveolar epithelial cells in the control group showed a classic epithelial morphology.After radiation-induction,the morphology of cells changed to a slender spindle shape.Compared with the control group,the mRNA levels of vimentin andα-SMA were increased in the radiation-induction group,while the E-cadherin mRNA level was decreased(P<0.05).Radiation increased the signal of Snail(green fluorescence)compared to the control group,and most of the signal was located in the nucleus.Compared with the control group,the levels of TGF-β,Smad2 and Smad3 were increased in the radiation-induction group(P<0.05).Radiation-induction increased the phosphorylation levels of ERK and GSK3β(P<0.05),and there were no changes in ERK and GSK3βtotal protein levels(P>0.05).U0126 inhibitd the phosphorylation of ERK and effectively abolished radiation-induced GSK3βphosp

关 键 词：转化生长因子-β ERK/GSK3β信号通路 肺泡上皮细胞 诱导放射性肺炎 上皮间充质转化 

分 类 号：R512.91[医药卫生—内科学]