雷公藤多苷对阿霉素肾病模型鼠肾组织TGF-β_1/Smad信号通路的干预作用

Effect of Multi-glycoside of Tripterygium wilfordii Hook.f.in Intervening TGF-β1/Smad Signaling Pathway of Adriamycin-induced Nephropathy Model Rat

目的观察雷公藤多苷(multi-glycoside of Tripterygium wilfordii Hook.f.,GTW)对阿霉素肾病(adriamycin-induced nephropathy,ADRN)模型鼠肾组织转化生长因子(transforming growth factor,TGF)-β1/Smad信号通路的干预作用,阐明GTW改善肾小球硬化(glomerulosclerosis,GS)的分子机制。方法将15只雌性SD大鼠随机分为假手术组、模型组和GTW组。对GTW组和模型组大鼠行右侧肾摘除术,并在4周内2次分别经颈静脉注射0.4mL和0.2mL阿霉素(adriamycin,ADR)而建立ADRN模型。造模成功后,GTW组大鼠经口灌胃GTW[50mg/(kg·d)],并同时以蒸馏水干预模型组,共干预6周。术后第2、4、8、10周称量大鼠体重,并检测24h尿蛋白排泄量,术后第10周处死大鼠,抽取血液,摘除左肾,观察血清生化指标、肾小球形态学特征、肾小球α-平滑肌肌动蛋白(α-smooth muscle actin,α-SMA)、I型胶原表达等,并且,借助RT-PCR和Western blotting等方法检测肾组织内TGF-β1、Smad3、Smad7核酸表达和TGF-β1、磷酸化Smad2/3(p-Smad2/3)蛋白表达。结果 GTW减少模型鼠的蛋白尿,改善血清白蛋白,对血尿素氮(BUN)和血清肌酐(SCr)没有影响,也未引起肝细胞损害。GTW抑制模型鼠系膜细胞增殖、细胞外基质(ECM)和胶原沉积,减少α-SMA、I型胶原表达,改善GS特征性病变。GTW下调模型鼠肾组织内TGF-β1、Smad3核酸表达,同时,上调Smad7核酸表达;GTW还可以下调模型鼠肾组织内TGF-β1和p-Smad2/3蛋白表达。结论 GTW在体内具有改善GS的作用;GTW通过调节肾组织TGF-β1/Smad信号通路中关键信号分子——Smad3、p-Smad2/3核酸或蛋白的表达,干预TGF-β1/Smad信号通路的信号转导,从而减轻ECM沉积,改善GS。

Objective To explore the potential molecular mechanisms of multi-glycoside of Tripterygium wilfordii Hook.f.（GTW） for ameliorating glomerulosclerosis（GS） by observing its intervention effect on transforming growth factor（TGF）-β1/Smad signaling pathway in adriamycin-induced nephropathy（ADRN） model rat.Methods Fifteen female Sprague-Dawley（SD） rats were randomly divided into three groups,the sham-operation group（A）,the untreated model group（B）,and the GTW treated model group（C）.Rats in Group B and C were made into ADRN model by right nephrectomy and intravenous injection of adriamycin（ADR,0.4 mL and 0.2 mL respectively in 4 weeks）.After the model was succesfully established,rats in Group C were orally given GTW（50 mg/kg per day）,while rats in Group B were intervened with distilled water.The intervention for two groups was 6 weeks.Rats＇ body weight were weighed and 24 h urinary protein excretion（Upro） detected by the end of the 2nd,4th,8th and 10th week.All rats were sacrificed at the end of 10th week after operation to withdraw blood and kidney tissue to examine serum biochemical parameters,glomerular morphological changes,α-smooth muscle actin（α-SMA）,and collagen type Ⅰ expression.Besides,the mRNA expressions of TGF-β1,Smad3 and Smad7,as well as protein expressions of TGF-β1 and phosphorylated Smad2/3（p-Smad2/3） in glomeruli were detected by RT-PCR or Western blotting.Results As compared with Group B,in Group C,Upro and serum albumin were improved significantly,but no difference between groups was found in levels of bloold urea nitrogen（BUN）,serum creatinine（SCr）,or hepatic cell injury.Mesangial cell proliferation,extracellular matrix（ECM） and collagen deposition were suppressed by GTW.Expressions of α-SMA and collagen type Ⅰ decreased,and the characteristic changes of GS were attenuated.The mRNA expressions of TGF-β1,Smad3 and protein expression of TGF-β1,p-Smad2/3 in renal tissues were down-regulated,while the protein expression of Smad7 mRNA was up-regulated.Conclusion GTW showed effect in ameliorating GS in vivo.It could reduce the ECM deposition and improve GS by way of intervening TGF-β1/Smad signaling pathway in the kidney through regulating the mRNA or protein expressions of key signal molecules,such as Smad3 and p-Smad2/3.