实脾固肾化瘀方对阿霉素肾纤维化大鼠肾组织TGF-β1/Smads信号通路的影响

Effects of Shipigushenhuayufang on the TGF-β1/Smads signaling pathway in the kidney tissue of adriamycin induced renal fibrosis rats

目的探讨实脾固肾化瘀方对阿霉素(ADR)肾纤维化大鼠肾组织转化生长因子-β1(TGF-β1)/Smads信号通路的影响。方法研究于2016年8—12月在上海中医药大学附属曙光医院肾病研究所进行,40只雄性SD大鼠随机分成正常对照组、模型组、实脾固肾化瘀方组、福辛普利钠组等4组,每组10只。除正常对照组外,其余3组大鼠均采用尾静脉注射ADR(4mg/kg)法制成肾纤维化大鼠模型。造模成功后实脾固肾化瘀方组予以实脾固肾化瘀方浸膏(43g/kg);福辛普利钠组予以福辛普利钠溶液(2mg/kg);模型组和正常对照组予以生理盐水。实验结束后,留取全部大鼠24 h尿量、血清及肾脏组织,观察各组大鼠肾脏病理变化及肾组织TGF-β1、Smad2及Smad7表达的变化。结果 Masson染色发现,正常对照组大鼠肾组织形态结构正常,未见肾小管、间质纤维化及炎细胞浸润等异常改变。肾间质胶原纤维微弱表达,胶原纤维呈蓝色,肌纤维及红细胞呈红色。模型组大鼠肾组织出现明显的病理损害,实脾固肾化瘀方组、福辛普利钠组大鼠肾组织病理损害明显减轻,且实脾固肾化瘀方组大鼠肾组织病理改善比福辛普利钠组更明显。免疫组化染色并对其灰度值测定后发现,模型组肾组织TGF-β1、Smad2表达较正常对照组明显上调(P<0.01),实脾固肾化瘀方组大鼠肾组织TGF-β1及Smad2表达较模型组明显下调(P<0.01),Smad7表达明显上调(P<0.01),与福辛普利钠组相比,实脾固肾化瘀方组大鼠肾组织TGF-β1、Smad2表达差异无统计学意义(P>0.05),而Smad7表达则明显上调(P<0.05)。结论实脾固肾化瘀方可能通过下调TGF-β1及Smad2表达、上调Smad7表达,干预TGF-β1/Smads信号通路,从而发挥改善ADR肾病大鼠肾纤维化、保护肾功能的作用。

Objective To investigate the effect of Shipigushenhuayufang on transforming growth factor-β1 （TGF-β1） /Smads signaling pathway in renal tissue of adriamycin （ADR） rats with renal fibrosis.Methods The study was conducted at the Institute of nephropathy, affiliated Shuguang Hospital of Shanghai University of Traditional Chinese Medicine from Aug 2016 to December 2016.40 male SD rats were randomly divided into 4 groups （10 rats in each group）： normal control group, model group, Shipigushenhuayufang group, and fosinopril sodium group.Except the normal control group, the rats in the other 3 groups were injected with ADR （4 mg/kg） into the renal fibrosis model.At the beginning of the third week of the model, the Shipigushenhuayufang group was treated with Shipigushenhuayufang （43 g/kg）;the fosinopril sodium group was treated with fosinopril sodium solution （2 mg/kg）;the model group and the normal control group were treated with normal saline.At the end of the experiment, 24 h urine volume, serum and kidney tissue were taken from all rats.The pathological changes of kidney and the changes of the expressions of TGF-β1, Smad2 and Smad7 in renal tissue were observed in each group.Results Masson staining showed that the normal control rats were normal in morphology and structure, and no abnormal changes were found in renal tubules,interstitial fibrosis and inflammatory cell infiltration.The collagen fibers in the kidney were weakly expressed, the collagen fibers were blue, and the muscle fibers and erythrocytes were red.The pathological damage appeared in the renal tissue of the rats in the model group, Shipigushenhuayufang group, fosinopril sodium group had pathological damage significantly reduced, and the Shipigushenhuayufang group rat improve the pathological kidney tissue more obvious than fosinopril sodium group.Immunohistochemical staining showed that, compared with normal control group, TGF-β1 expression was significantly increased in renal tissue of model group （P 〈0.01）, the expression of Smad2 in renal tissue of model group increased significantly （P 〈0.01）, compared with the model group, Shipigushenhuayufang group rat kidney TGF-β1 and the expression of Smad2 was down regulated （P 〈0.01）, Smad7 expression up regulated （P 〈0.01）.Compared with the fosinopril sodium group, the expression of TGF-β1 and Smad2 in the renal tissue of the Shipigushenhuayufang group was not statistically significant different （P 〉0.05）, while the expression of Smad7 was significantly higher （P 〈0.05）.Conclusion The Shipigushenhuayufang may regulate the expression of TGF-β1 and Smad2, and up regulate the expression of Smad7, and interfere with the TGF-β1/Smads signaling pathway, thereby improving the renal fibrosis and protecting the renal function in ADR nephropathy rats.