六味地黄汤调控TGF-β及Smad家族蛋白表达对肾间质纤维化的影响研究

Effects of Liuwei Dihuang Decoction on renal interstitial fibrosis by regulating the expressions of TGF-βand Smad family proteins

目的探究六味地黄汤抑制肾间质纤维化(renal interstitial fibrosis,RIF)的干预作用及机制。方法将30只SPF级SD大鼠随机均分为假手术组、模型组、阳性药物组、中药高剂量组和中药低剂量组,每组6只。除假手术组外,其余各组采用左侧输尿管结扎术制备RIF模型。造模结束后,假手术组和模型组予以蒸馏水灌胃,阳性药物组予以醋酸泼尼松片0.2 mg/kg灌胃,中药低、高剂量组分别予以六味地黄汤0.5、1 mg/kg灌胃,疗程为21 d。治疗结束后,采用HE染色法评估肾间质损伤,Masson染色法检测肾间质胶原蛋白沉积和纤维化,RT-PCR测定肾间质转化生长因子-β1(transforming growth factor-β1,TGF-β1)、针对半身不遂同源物2的重组母体(recombinant mothers against decapentaplegic homolog 2,Smad2)、Smad3、Smad7的mRNA表达,Western blot测定肾间质TGF-β1、Smad2、Smad3、Smad7的蛋白表达。结果与假手术组相比,模型组的病理呈现系膜增生、间质增宽、胶原蛋白大面积沉积和明显纤维化,且TGF-β1、Smad2、Smad3、Smad7的mRNA和蛋白表达均增加(P<0.05)。与模型组相比,阳性药物组和中药低、高剂量组的病理表现均减轻,且TGF-β1、Smad2、Smad3、Smad7的mRNA和蛋白表达均降低(P<0.05)。与中药低剂量组相比,阳性药物组和中药高剂量组的TGF-β1、Smad2、Smad3、Smad7的mRNA和蛋白表达均降低(P<0.05)。结论六味地黄汤能够减轻RIF大鼠的肾间质纤维化程度,且高剂量效果更佳,其机制可能与调控TGF-β及Smad家族蛋白表达有关。

Objective To explore the intervention effects and mechanism of Liuwei Dihuang Decoction(LWDHD)on inhibiting renal interstitial fibrosis(RIF).Methods A total of 30 SPF SD rats were randomly divided into sham-operated group,model group,positive drug group,high and low-dose Chinese medicines groups,with six rats in each group.Except for the sham-operated group,all other groups were prepared with left ureteral ligation to establish RIF models.After modeling,the sham-operated group and model group were given distilled water by gavage,the positive drug group was given acetic acid prednisone tablets 0.2 mg/kg by gavage,and the high and low-dose Chinese medicines groups were given LWDHD 1 mg/kg and 0.5 mg/kg by gavage,respectively,for a treatment course of 21 d.After treatment,renal interstitial injury was evaluated by HE staining,and renal interstitial collagen deposition and fibrosis were measured by Masson staining.The mRNA expressions of transforming growth factor-β1(TGF-β1),recombinant mothers against decapentaplegic homolog 2(Smad2),Smad3,and Smad7 were determined by RT-PCR.The protein expressions of TGF-β1,Smad2,Smad3,and Smad7 in the renal interstitium were determined by Western blot.Results Compared with the sham-operated group,the model group exhibited pathological features of mesangial proliferation,interstitial widening,and extensive collagen deposition and obvious fibrosis,as well as elevated mRNA and protein expressions of TGF-β1,Smad2,Smad3,and Smad7(P<0.05).Compared with the model group,the positive drug group,low and high-dose Chinese medicines groups showed alleviated pathological changes,as well as a decrease in the mRNA and protein expressions of TGF-β1,Smad2,Smad3,and Smad7(P<0.05).Compared with the low-dose Chinese medicines group,the mRNA and protein expressions of TGF-β1,Smad2,Smad3,and Smad7 in the positive drug group and the high-dose Chinese medicines group decreased(P<0.05).Conclusion LWDHD can reduce renal interstitial fibrosis in RIF rats,with higher doses exhibiting more pronounced effects.The mechanism may be related to the regulation of TGF-β1 and Smad family protein expression.