配对盒基因2对正常肾小管上皮细胞间充质转分化的诱导作用  

Induction of PAX2 on epithelial-mesenchymal transition in normal renal tubular epithelial cells

作　　者：李里[1,2] 南晓娟[1] 吴玉斌[1] 

机构地区：[1]中国医科大学附属盛京医院小儿肾脏风湿免疫科,辽宁沈阳110004 [2]辽宁医学院附属第一医院儿科,辽宁锦州121000

出　　处：《吉林大学学报（医学版）》2013年第3期565-569,F0003,共6页Journal of Jilin University：Medicine Edition

基　　金：辽宁省沈阳市科学技术计划项目资助课题(F10-205-1-29);盛京医院博士优秀课题基金资助课题(ma26)

摘　　要：目的:观察体外转染配对盒基因2(PAX2)对肾小管上皮细胞表型标志物的影响,探讨PAX2诱导转分化的作用,为肾纤维化治疗提供依据。方法:正常肾小管上皮细胞株NRK52E分为对照组、空载组(转染pGC-Fu空质粒)和转染组(采用脂质体转染法将pGC-FU-GFP-PAX2质粒转染至细胞中)。转染72h后应用倒置相差显微镜观察各组细胞形态,应用Western blotting法和Real-time PCR法检测PAX2蛋白和mRNA表达水平;转染72h后应用免疫组织化学法、Western blotting法和Real-time PCR法检测各组细胞E-钙黏蛋白(E-cadherin)和α-肌动蛋白(α-SMA)及其mRNA表达水平。结果:pGC-FU-GFP-PAX2转染72h后,转染组细胞绿色荧光强度较对照组明显增强,且肾小管上皮细胞的形态伸长;Western blotting法检测,转染组PAX2蛋白表达水平明显高于空载组和对照组(P<0.05);Real-time PCR检测,转染组细胞的PAX2mRNA表达水平明显高于空载组和对照组(P<0.05);转染组细胞中的E-cadherin染色较空载组和对照组明显减弱;α-SMA染色较空载组和对照组明显增强。Western blotting法检测,转染组细胞中E-cadherin表达水平明显低于空载组和对照组(P<0.05);α-SMA表达水平明显高于空载组和对照组(P<0.05);Real-time PCR检测,转染组细胞中E-cadherin mRNA表达水平明显低于空载组和对照组(P<0.05);α-SMA mRNA表达水平明显高于空载组和对照组(P<0.05)。结论:PAX2转染正常肾小管上皮细胞后E-cadherin表达减少,α-SMA表达增加,PAX2可能在体外诱导正常肾小管上皮细胞间充质转分化。Objective To observe the effect of paired box gene 2（PAX2） on the surface markers of renal tubular epithelial cells and induction of PAX2 on epithelial-mesenchymal transition（EMT） in vitro,and to provide basis for treatment of renal fibrosis.Methods The rat normal renal tubular epithelial cell line NRK52E were divided into control group,empty vector group（transfected with pGC-FU empty vector）,and transfection group（transfected with pGC-FU-GFP-PAX2 vector）.The morphology of cells 72 h after transfection was evaluated by inverted phase contrast microscope.The expression levels of PAX2 protein and mRNA were evaluated by Western blotting and Real-time PCR methods.72 h after transfection,the expressions of E-cadherin and α-smooth muscle actin（α-SMA） and their mRNA expressions were detected by immunohistochemistry,Western blotting and Real-time PCR methods.Results 72 h after tranfection of pGC-FU-GFP-PAX2,the fluorescence intensity in transfection group was stronger than that in control group,and the shape of renal tubular epithelial cells elongated.The Western blotting results showed that the expression level of PAX2 protein in transfection group was increased compared with control and empty vector groups（P0.05）;the Real-time PCR results showed that the expression level of PAX2 mRNA in transfection group was increased compared with control group and empty vector group（P0.05）.The dyeing of E-cadherin got weaker and the dyeing of α-SMA got stronger in transfection group compared with control and empty vector groups.The Western blotting results showed that the expression level of E-cadherin in transfection group was decreased and α-SMA was increased compared with control and empty vector groups（P0.05）.The Real-time PCR results showed that the expression level of E-cadherin mRNA in transfection group was decreased and the expression level of α-SMA mRNA was increased in transfection group compared with control and empty vector groups（P0.05）.Conclusion PAX2 transfection can reduce E-

关 键 词：配对盒基因2 肾小管上皮细胞 转分化 转染 

分 类 号：Q279[生物学—细胞生物学]