沉默PAX2基因对肾间质纤维化大鼠的影响

Influence of PAX2 gene silencing on renal interstitial fibrosis in rats

目的探讨体内沉默PAX2基因对肾间质纤维化大鼠肾小管上皮细胞转分化(EMT)的影响。方法 64只Wistar大鼠麻醉后,单侧输尿管结扎法制作肾间质纤维化大鼠模型,随机分为阴性对照组和PAX2基因沉默组,每组32只。将200μL NC-siRNA-in vivo jetPEI^(TM)混合液转染阴性对照组大鼠,将200μL PAX2-siRNA-in vivo jetPEI^(TM)混合液转染PAX2基因沉默组大鼠,各组分别于转染后3、5、7、14 d分为4个亚组,每组8只。留取各组肾组织标本,应用Real-Time PCR及Western blot法检测肾皮质PAX2 mRNA及其蛋白的沉默情况,以及E-钙粘连素(E-cadherin)、α-平滑肌肌动蛋白(α-SMA)mRNA和蛋白表达情况。结果 PAX2基因沉默组PAX2 mRNA和蛋白表达量较阴性对照组均降低(P<0.05)。随着梗阻时间的延长,两组E-cadhelin mRNA和蛋白表达量逐渐下降,α-SMA mRNA和蛋白表达量逐渐升高;在转染14 d时,PAX2基因沉默组E-cadhelin mRNA和蛋白相对表达量明显高于阴性对照组(P<0.05),而α-SMA mRNA和蛋白相对表达量明显低于阴性对照组(P<0.05)。结论沉默PAX2基因在肾间质纤维化晚期大鼠中可明显抑制肾小管EMT进程,可能对肾间质纤维化有治疗作用。

ObjectiveTo investigate the influence of silencing PAX2 genein vivo on epithelial-mesenchymal transition （EMT） of renal tubular cells in rats with renal interstitial ifbrosis.MethodsA total of 64 Wistar rats were anaesthetized, and unilateral ureteral obstruction （UUO） was performed to establish a rat model of renal interstitial ifbrosis. The 64 rats were randomly divided into negative control and PAX2 gene silencing groups （n=32 each）. The rats in the control group were transfected with 200 μL NC-siRNA-in vivo jetPEITM solution. Those in the PAX2 gene silencing group were transfected with 200 μL PAX2-siRNA-in vivo jetPEITM solution. Each group was further divided into 4 subgroups based on the post-transfection time （3, 5, 7 and 14 days after transfection）, with 8 rats in each subgroup. Renal tissue samples were harvested in each group. Real-time PCR and Western blot were used to measure the mRNA and protein expression of PAX2 in the renal cortex, as well as the mRNA and protein expression of E-cadherin and α-SMA.Results Compared with the control group, the PAX2 gene silencing group showed significantly lower mRNA and protein expression of PAX2 （P〈0.05）. In the two groups, the mRNA and protein expression levels of E-cadherin were gradually reduced over the time of obstruction, while those of α-SMA gradually increased. At 14 days after transfection, the PAX2 gene silencing group had signiifcantly higher mRNA and protein expression of E-cadherin but lower mRNA and protein expression of α-SMA compared with the control group （P〈0.05）.ConclusionsPAX2 gene silencing can signiifcantly inhibit the process of EMT of renal tubular cells in rats with advanced ifbrosis, suggesting that PAX2 gene silencing may have a therapeutic effect on renal interstitial ifbrosis.