白藜芦醇对糖尿病小鼠肾脏自噬水平及肾间质纤维化的影响

Effect of resveratrol on autophagy and renal interstitial fibrosis in kidney of diabetic mice

目的:观察白藜芦醇(Res)对糖尿病(DM)小鼠肾脏自噬水平及肾间质纤维化的影响,并探讨其可能的作用机制。方法:将清洁级雄性C57BL/6小鼠随机分为正常对照组(NC组)、DM组和Res组,每组各8只。用链脲佐菌素复制DM小鼠模型。从DM模型复制成功8周起,Res组连续给予Res灌胃干预12周后处死各组小鼠测定相应生化指标和肾脏指数(KI)。HE和Masson染色比较各组肾组织的病理学改变;Western blot分别检测各组自噬、上皮-间充质细胞转化(EMT)及纤维化相关指标的蛋白表达水平,real-time PCR检测Ⅳ型胶原(Col Ⅳ)、α-平滑肌肌动蛋白(α-SMA)和上皮钙黏素(E-cadherin)的mRNA表达水平。结果:与NC组相比,DM组的空腹血糖(FBG)、KI、血清肌酐、24 h尿白蛋白排泄率和24 h尿总蛋白均显著增高(P<0.05),HE和Masson染色显示出现肾纤维化改变。DM组小鼠肾组织的E-cadherin、beclin-1和微管相关蛋白1轻链3-Ⅱ(LC3-Ⅱ)蛋白水平较NC组显著降低(P<0.05),α-SMA、Col IV和Snail1蛋白水平较NC组显著升高(P<0.05)。经Res干预后,除了FBG无显著差异(P>0.05)外,Res组中其余生化指标和KI均显著低于DM组(P<0.05);肾纤维化病变明显减轻;E-cadherin、beclin-1和LC3-Ⅱ蛋白水平较DM组显著升高(P<0.05),而α-SMA、Col IV和Snail1蛋白水平较DM组显著降低(P<0.05)。与DM组比较,Res组E-cadherin的mRNA表达增高(P<0.05),Col IV和α-SMA的mRNA表达降低(P<0.05)。结论:白藜芦醇可显著抑制DM小鼠肾组织的EMT,减轻肾间质纤维化病变,其机制可能与促进肾脏自噬水平有关。

AIM:To observe the effect of resveratrol(Res)on renal autophagy level and renal interstitial fibrosis in the mice with diabetes mellitus(DM),and to discuss the possible mechanism.METHODS:The wild-type C57 BL/6 mice were randomly divided into 3 groups,including normal control(NC)group,DM group and Res group(8 in each group).The diabetic mouse model was established by injection of streptozotocin.After 8 weeks of successful replication of the diabetic model,Res was given to the mice in Res group by continuous gavage for 12 weeks,and then the mice in each group were sacrificed to detect the relevant biochemical parameters.The pathological changes of the kidney tissues were observed by HE staining and Masson staining.The levels of the proteins related to autophagy,epithelial-mesenchymal transition(EMT)and fibrosis were determined by Western blot.The mRNA expression of collagen type Ⅳ(Col Ⅳ),α-smooth muscle actin(α-SMA)and E-cadherin were detected by real-time PCR.RESULTS:Compared with NC group,fasting blood glucose(FBG),kidney index(KI),serum creatinine,24-hour urinary albumin excretion rate and 24-hour urine total protein were remarkably increased in DM group(P<0.05).The results of HE and Masson staining indicated that renal tissue presented fibrosis in DM group.The protein levels of E-cadherin,beclin-1,microtubule-associated protein 1 light chain 3-Ⅱ(LC3-Ⅱ)were reduced in DM group,while the levels of α-SMA,Col IV and Snail1 were increased(P<0.05).After intervention with Res for 12 weeks,all the relevant biochemical parameters and KI were reduced(P<0.05)except FBG(P>0.05),and renal fibrosis lesions were obviously alleviated.Compared with DM group,the protein levels of E-cadherin,beclin-1 and LC3-Ⅱ were increased in Res group,but the protein expression levels of α-SMA,Col IV,Snail1 were reduced(P<0.05).Compared with DM group,the mRNA level of E-cadherin was increased in Res group,but the mRNA levels of Col IV and α-SMA were reduced(P<0.05).CONCLUSION:Resveratrol significantly inhibits EMT and reduces renal interstitial fibrosis in diabetic mice,and its mechanism may be related to the promotion of renal autophagy.