知母皂苷对糖尿病肾病大鼠的肾保护作用及机制

The renoprotective effect and mechanism of Zhimu saponin in diabetic nephropathy rats

目的探讨知母皂苷对糖尿病肾病大鼠的肾保护作用及机制。方法采用链脲佐菌素诱导法构建糖尿病肾病大鼠模型,随机分为模型组(等量生理盐水灌胃),知母皂苷低、高剂量组(10、20 mg/kg知母皂苷灌胃),每组16只,同时设置对照组大鼠16只。连续灌胃8 w,称量体质量、肾脏质量,计算肾指数。收集24 h尿液,之后眼眶取血,离心收集血清,全自动分析仪检测血清尿素氮、肌酐及24 h尿蛋白水平。取大鼠肾组织,分成两部分,一部分进行组织匀浆后离心收集上清液,酶联免疫吸附试验检测丙二醛、超氧化物歧化酶(SOD)、晚期糖基化终产物(AGEs)、血管内皮生长因子(VEGF)、Ⅳ型胶原(Ⅳ-Col)水平。Western印迹检测另一部分肾组织中转化生长因子(TGF)-β1、Smad同源物(Smad)3、Smad7蛋白表达。结果知母皂苷低、高剂量组体质量明显高于模型组,肾脏质量、肾指数明显低于模型组(P<0.05);知母皂苷高剂量组上述改变更明显。知母皂苷各组血清尿素氮、肌酐及24 h尿蛋白水平明显低于模型组(P<0.05);知母皂苷高剂量组以上指标明显低于低剂量组(P<0.05)。知母皂苷低、高剂量组丙二醛、AGEs、VEGF、Ⅳ-Col水平明显低于模型组,SOD水平明显高于模型组(P<0.05);且知母皂苷高剂量组丙二醛、AGEs、VEGF、Ⅳ-Col水平明显低于知母皂苷低剂量组,SOD水平明显高于知母皂苷低剂量组(P<0.05)。知母皂苷各组肾组织中TGF-β1、Smad3蛋白表达水平明显低于模型组,Smad7蛋白表达水平明显高于模型组(P<0.05);知母皂苷高剂量组TGF-β1、Smad3蛋白表达水平明显低于知母皂苷低剂量组,Smad7蛋白表达水平明显高于知母皂苷低剂量组(P<0.05)。结论知母皂苷能够抑制糖尿病肾病大鼠肾脏纤维化,降低氧化应激水平,改善肾功能,可能通过调节TGF-β1/Smad信号通路来实现。

Objective To investigate the renoprotective effect and mechanism of Zhimu saponin in diabetic nephropathy rats.Methods Diabetic nephropathy rat models were established using streptozotocin injection and rats were randomly divided into model group(given equivalent saline by gavage),Zhimu saponin low-dose,high-dose groups(given 10,20 mg/kg Zhimu saponin by gavage),with 16 rats in each group and control group(16 rats).The body mass and kidney mass were measured and the kidney index was calculated after 8 w of continuous gavage.Urine samples were collected for 24 h,followed by blood collection via the orbital plexus,and serum was separated by centrifugation.Serum levels of blood urea nitrogen,creatinine,and 24-hour urinary protein were detected using an automatic analyzer.Rat kidney tissues were divided into two parts.One part was homogenized and the supernatant was collected after centrifugation.Levels of malondialdehyde,superoxide dismutase(SOD),advanced glycation end products(AGEs),vascular endothelial growth factor(VEGF),and collagen typeⅣ(Ⅳ-Col)were determined using enzyme-linked immunosorbent assay.Western blot was used to detect the protein expressions of transforming growth factor(TGF)-β1,Smad homologue(Smad)3 and Smad7 in another part of the kidney tissues.Results The body mass of the Zhimu saponin low-dose and high-dose groups were significantly higher than that in the model group,while the kidney weight and renal index were significantly lower than those in the model group(P<0.05);the changes in the Zhimu saponin high-dose group were more pronounced.Serum levels of blood urea nitrogen,creatinine,and 24-hour urinary protein in the Zhimu saponin groups were significantly lower than those in the model group(P<0.05);the above indexes of Zhimu saponin high-dose group were significantly lower than those in the Zhimu saponin low-dose group(P<0.05).Levels of malondialdehyde,AGEs,VEGF,andⅣ-Col in the Zhimu saponin low-dose and high-dose groups were significantly lower than those in the model group,the SOD level was significantly higher than that in the model group(P<0.05);levels of malondialdehyde,AGEs,VEGF,andⅣ-Col in the Zhimu saponin high-dose group were significantly lower than those in the Zhimu saponin low-dose group,the SOD level was significantly higher than that in the Zhimu saponin low-dose group(P<0.05).The expression levels of TGF-β1 and Smad3 protein in the kidney tissues of the Zhimu saponin groups were significantly lower than those in the model group,while the Smad7 protein expression level was significantly higher than that in the model group(P<0.05);the expression levels of TGF-β1 and Smad3 protein in the Zhimu saponin high-dose group were significantly lower than those in the Zhimu saponin low-dose group,while the Smad7 protein expression level was significantly higher than that in the Zhimu saponin low-dose group(P<0.05).Conclu-sions Zhimu saponin could inhibit renal fibrosis in diabetic nephropathy rats,reduce oxidative stress levels,and improve renal function,which might be achieved by regulating the TGF-β1/Smad signaling pathway.