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转化生长因子#beta#1和Snail1参与糖尿病大鼠肾小管上皮细胞向间充质细胞转变

方开云, 娄晶磊, 肖瑛, 石明隽, 桂华珍, 郭兵, 张国忠

贵阳医学院病理生理学教研室.贵州,贵阳 550004

摘要

该文旨在观察转化生长因子#beta#1（transforming growth factor--#beta#1，TGF--#beta#1）和锌指转录因子Snail1在糖尿病（diabetes mellitus，DM）大鼠肾组织中的表达，并初步探讨它们与肾小管上皮细胞向间充质细胞转变的关系。链脲佐菌素（streptozotocin，STZ）诱发大鼠DM，按病程分为2、4、8、12、16、20、24周、16周胰岛素治疗（16wA）、20周胰岛素治疗（20wA）和24周胰岛素治疗（24wA）组（{sl n}=6）。其中胰岛素治疗组动物从第13周起用胰岛素控制血糖至正常水平，每一时点均设鼠龄匹配的正常对照组。测定各组血糖、24h尿蛋白、血肌酐（serum creatinine，Scr）、肾脏指数。PAS染色光镜观察肾脏病理学改变。免疫组织化学检测肾脏Snail1、TGF--#beta#1、#alpha#--平滑肌肌动蛋白（#alpha#--smooth muscle actin，#alpha#--SMA）、E--钙黏素和纤连蛋白（fibronectin,FN）的表达；Western blot检测肾皮质Snail1、TGF--#beta#1和E--钙黏素蛋白表达。RT--PCR检测肾皮质Snail1和E--钙黏素mRNA表达。结果显示：（1）DM各组大鼠的血糖、24h尿蛋白、Scr、肾脏指数均较正常对照组明显升高（{sl P}<0.05，{sl P}<0.01），胰岛素治疗组大鼠上述指标均较DM组显著降低（{sl P}<0.01）。（2）TGF--#beta#1和Snail1免疫组织化学阳性染色见于DM各组大鼠肾小管，正常对照组未见阳性表达，胰岛素治疗组大鼠弱阳性表达，并随治疗时间延长而减少。从16周开始在DM大鼠肾小管上皮细胞可见#alpha#--SMA蛋白阳性表达，胰岛素治疗组大鼠未见#alpha#--SMA蛋白表达；DM组大鼠E--钙黏素蛋白阳性染色明显少于正常对照组。（3）DM组大鼠肾皮质TGF--#beta#1和Snail1蛋白以及Snail1 mRNA表达较正常对照组显著增高（{sl P}<0.01），胰岛素治疗组大鼠则显著低于DM组（{sl P}<0.01）；DM组E--钙黏素mRNA和蛋白表达与TGF--#beta#1和Snail1呈相反变化。结果提示，TGF--#beta#1和Snail1可能参与DM大鼠肾小管上皮细胞向间充质细胞转变，胰岛素治疗可抑制两者表达并阻断肾小管上皮细胞向间充质细胞转变。

关键词： 转化生长因子#beta#1; Snail1; E-钙黏素; #alpha#-平滑肌肌动蛋白; 糖尿病肾病; 大鼠

Transforming growth factor--#beta#1 and Snail1 mediate tubular epithelial--mesenchymal transition in diabetic rats

Fang Kaiyun, Lou Jinglei, Xiao Ying, Shi Mingjuan, Gui Huazhen, Guo Bing, Zhang Guozhong

Department of Pathophysiology, Guiyang Medical College.Guiyang 550004,Guizhou

Abstract

The present study was aimed to explore the expressions of transforming growth factor-#beta#1 （TGF-#beta#1） and Snail1 in renal tissues of diabetic rats, and their role in tubular epithelial-mesenchymal transition （TEMT）. Induced diabetic rats were randomly divided into 2-, 4-, 8-, 12-, 16-, 20-, 24-week and 16wA, 20wA, 24wA groups. The rats in 16wA, 20wA and 24wA groups were treated with insulin to control blood glucose to the normal level from the 13th week. The age-matched rats were set as controls. Blood glucose, 24-hour urine protein, serum creatinine （Scr）, kidney index of rats were measured. PAS staining was used to observe the renal pathological changes. Immunohistochemical staining and （or） Western blot were employed to determine the expressions of TGF-#beta#1, Snail1, E-cadherin, #alpha#-smooth muscle actin （#alpha#-SMA） and fibronectin （FN） proteins. The expressions of Snail1 and E-cadherin mRNAs in renal cortex were examined by RT-PCR. Blood glucose, 24-hour urine protein, Scr and kidney index increased remarkably in diabetic rats as compared with those in the control groups （P〈0.05, P〈0.01） and insulin-treated rats （P〈0.01）. TGF-#beta#1 and Snail1 protein expressions could not be detected by immunohistochemical staining in the normal renal tissues, however, the strongly positive staining was observed in diabetic rat renal tubules. A time-dependent loss of TGF-#beta#1 and Snail1 expressions was detected in the kidney of insulin-treated rats. In diabetic rats tubular #alpha##-SMA positive staining was seen at the 16th week. E-cadherin expression was lost in diabetic rats. The expressions of TGF-#beta#1, Snail1 proteins and Snail1 mRNA were significantly up-regulated in diabetic rats, while down-regulated in insulin-treated rats （P〈0.01）. The expressions of E-cadherin protein and mRNA in the cortex were contrary to the expressions of TGF-#beta#1 and Snail1. Therefore, TGF-#beta#1 and Snail1 are possibly involved in the pathogenesis of TEMT in diabetic nephropathy rats.

Key words: transforming growth factor-#beta#1;Snail1;E-cadherin;#alpha#-smooth muscle actin;diabetic nephropathy;Rat

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引用本文：

方开云, 娄晶磊, 肖瑛, 石明隽, 桂华珍, 郭兵, 张国忠. 转化生长因子#beta#1和Snail1参与糖尿病大鼠肾小管上皮细胞向间充质细胞转变[J]. 生理学报 2008; 60 (1): 125-134.

Fang Kaiyun, Lou Jinglei, Xiao Ying, Shi Mingjuan, Gui Huazhen, Guo Bing, Zhang Guozhong. Transforming growth factor--#beta#1 and Snail1 mediate tubular epithelial--mesenchymal transition in diabetic rats. Acta Physiol Sin 2008; 60 (1): 125-134 (in Chinese with English abstract).