肾间质纤维化组织中组织型转谷氨酰胺酶和结缔组织生长因子共表达

Expression and implication of tissue transglutaminase and connective tissue growth factor at fibrotic tubulointerstitium in kidneys from UUO rats

目的:探讨组织型转谷氨酰胺酶(tTG)和结缔组织生长因子 (CTGF)在肾间质纤维化组织中的表达及二者的关系。方法: 采用单侧输尿管结扎(UUO)致肾间质纤维化大鼠模型。将大鼠随机分为假手术组 (5只 )、模型组(6只)、依那普利治疗组(6只)。术后 9天处死各组大鼠,肾组织标本用过碘酸-席夫染色(PAS)确证肾间质纤维化病变。以免疫组织化学染色方法观察肾组织内tTG和CTGF的表达部位,以及细胞外基质成份纤维连接蛋白(FN)的表达变化。用Western蛋白印迹方法分析肾组织内tTG和CTGF蛋白表达水平。结果: 假手术组大鼠tTG,CTGF和FN分别在肾小管、肾间质区仅有微量表达,肾小球内无阳性表达;模型组大鼠tTG、CTGF和FN在肾小管、肾间质区的表达信号比假手术组强;依那普利治疗后三者表达减少。双重染色结果显示,在模型组肾间质tTG阳性表达部位同时表达CTGF,并且强度相似。Westernblot结果显示,模型组tTG和CTGF蛋白水平明显高于假手术组;依那普利治疗组tTG和CTGF蛋白水平明显低于模型组。各组大鼠肾内tTG蛋白水平与CTGF蛋白水平之间(r=0. 683,P<0. 05)以及tTG蛋白水平与肾间质内FN半定量水平之间 (r=0. 737,P<0. 05)均有显著相关关系。结论: tTG和CTGF可能共同参与肾间质纤维化。

Objective: To observe the expression and co-locolization of tissue transglutaminase (tTG) and connective tissue growth factor (CTGF) in kidneys from rats with tubulointerstitial fibrosis. Methods: The animal models of unilateral ureteral obstruction (UUO) were used. The rats were randomly divided into the sham-operation group (n=5), the UUO group (n=6), and the Enalapril-treated UUO group (n=6). All the rats were sacrificed on day 9, the kidneys were collected, and renal interstitial fibrosis was examined by periodic acid-Schiff (PAS) staining. The expression and localization of tTG, CTGF, and Fibronectin(FN)in the obstructed kidneys were detected by immunohistochemistry staining. The protein levels of tTG and CTGF of the whole kidney homogenates were determined by Western blot analysis. Results: Weak signals of tTG, CTGF, and FN-positive immunostaining were observed in renal tubular cells and tubulointerstitium, and very rare positive signals were seen in the glomeruli in the sham-operation group. However, much intensive signals of tTG, CTGF, and FN-positive immunostaining were observed in renal tubular cells and tubulointerstitium in the UUO group, and the intensity of signals were decreased in the Enalapril-treated UUO group. Co-localization of tTG and CTGF immunoreactivity were observed in tubulointerstitium in the UUO group. The levels of tTG and CTGF protein analyzed by Western blotting were increased remarkably in the UUO group compared with the sham-operation group, and a significant reduction of tTG and CTGF protein levels were presented in the Enalapril-treated UUO group compared with the UUO group. The levels of tTG protein had positive correlation with the levels of CTGF protein (r=0.683, P<0.05), and with the semi-quantitative levels of FN expression (r=0.737, P< 0.05). Conclusion: Both tTG and CTGF may play a concordant role in the pathogensis of renal tubu-lointerstitial fibrosis,and Enalapril may suppress the development of fibrosis partially via down-regulation of tTG and CTGF expression.