丹蛭降糖胶囊对糖尿病肾病大鼠肾脏TGF-β1/Smad3信号通路和AGEs/RAGE水平的影响

Effects of Danzhi Jiangtang Capsules on renal TGF-β1/Smad3 signaling pathway and AGEs/RAGE level in diabetic nephropathy rats

目的:观察丹蛭降糖胶囊通过调节TGF-β1/Smad3信号通路和AGEs/RAGE水平对糖尿病肾病大鼠肾脏病变的影响。方法:60只雄性SD大鼠随机选取10只作为空白(NC)组,其余大鼠采用高糖高脂饮食联合腹腔注射链脲佐菌素(STZ)55mg/kg建立糖尿病肾病模型,最后42只大鼠造模成功,将其随机分为MC组(等容量蒸馏水)、VC组(缬沙坦胶囊,8mg·kg^(-1)·d^(-1))、DJC组(丹蛭降糖胶囊,540mg·kg^(-1)·d^(-1)),每组14只,NC组给予等量蒸馏水,其余各组给予相应的药物干预8周后,麻醉状态下取材。取血清、尿液,检测大鼠肾功能和24h尿蛋白;采用HE染色法观察肾脏组织病理学变化;用ELISA法检测肾脏组织AGEs、CTGF和RAGE的含量;Western blot法和实时荧光定量PCR法检测肾脏组织转化生长因子-β1(TGF-β1)及其信号蛋白Smad3蛋白和基因表达水平。结果:(1)与NC组比较,MC组大鼠体质量显著下降,右肾质量显著增加,24h尿蛋白、BUN的含量显著升高(P<0.01);与MC组比较,DJC组和VC组大鼠体质量显著增加,右肾质量显著减轻,24h尿蛋白、BUN的含量显著降低(P<0.01,P<0.05);与VC组比较,DJC组大鼠体质量显著增加,右肾质量显著减轻,24h尿蛋白显著降低(P<0.01)。(2)与MC组比较,光镜下VC组和DJC组肾小球和肾小管结构和排列改善明显。(3)VC组和DJC组AGEs、RAGE、CTGF含量和TGF-β1、Smad3和p-Smad3蛋白和基因表达水平均较MC组显著下调(P<0.05,P<0.01),其中DJC组较VC组下调更明显(P<0.05,P<0.01)。结论:丹蛭降糖胶囊可能通过下调肾脏TGF-β1/Smad3信号通路和AGEs/RAGE水平,延缓糖尿病大鼠肾脏纤维化,减轻肾脏损害,发挥肾脏保护作用。

Objective:To observe the effects of Danzhi Jiangtang Capsules on renal disease in diabetic nephnopathy rats by regulating TGF-β1/Smad3 signaling pathway and AGEs/RAGE level.Methods:Ten rats were randomly selected as NC group from 60 mals SD rats,the remaining rats were set up by intraperitoneal injection of high sugar and high fat diet combination Streptozotocin(STZ)55 mg/kg in diabetic nephropathy model,and 42 suceessfully built rats were randomly divided into MC group(equal volumn of distilled water),VC group(valsartan capsules,8 mg·kg^(-1)·d^(-1))and DJC group(Danzhi Jiangtang Capsule,540 mg·kg^(-1)·d^(-1)),14 rats in each group,NC group was given equal volumn of distilled water.After 8 weeks of corresponding drug intervention in the rest groups,materials were drawn under anesthesia.Serum and urine were collected to detect renal function and urinary protein of rats within 24 h.The histopathological changes of kidney were observed by HE staining.The content of AGEs,CTGF and RAGE in renal tissues were detected by ELISA.Western blot and Real-time fluorescence quantitative PCR were used to detect the expression levels of transforming growth factorβ1(TGF-β1)and its signal protein Smad3 in renal tissue.Results:(1)Compared with the NC group,the body mass of the MC group decreased significantly,the weight of the right kidney increased significantly,and the content of 24 h urine protein and BUN increased significantly(P<0.01).Compared with the MC group,the body mass of rats in the DJC group and the VC group was significantly increased,the weight of the right kidney was significantly reduced,and the content of 24 h urine protein was significantly decreased(P<0.01,P<0.05).Compared with the VC group,the body mass of rats in the DJC group was significantly increased,the weight of the right kidney was significantly reduced,and the 24 h urine protein was significantly decreased(P<0.01).(2)Compared with the MC group,the structure and arrangement of glomeruli and tubules in the VC group and DJC group under light microscope were significantly improved.(3)The contents of AGEs,RAGE,CTGF and the protein and gene expression levels of TGF-β1,Smad3 and p-Smad3 in VC and DJC groups were significantly downregulated than MC group(P<0.05,P<0.01),which were more significantly down-regulated in DJC group than those in the VC group(P<0.05,P<0.01).Conclusion:Danzhi Jiangtang Capsules may delay renal fibrosis,reduce renal damage and play a protective role by down-regulating renal TGF-β1/Smad3 signaling pathway and expression of AGEs/RAGE level in diabetic rats.