糖肾方对db/db小鼠肾脏纤维化的保护作用

Protective Effects of Tangshen Formula in Renal Fibrosis in db/db Mice

目的:观察糖肾方(Tang Shen Fang,TSF)对于自发性2型糖尿病肾病db/db小鼠肾脏纤维化的保护作用。方法:8周龄雄性db/db小鼠随机分为2组,分别为模型组(db/db)和观察组(db/db+TSF),每组9只;8周龄雄性db/m小鼠为正常对照组(db/m),顺应性喂养2周后,给予糖肾方干预12周,处死小鼠,分离小鼠肾脏用于后续检测。利用PAS和Masson染色观察肾脏病理;利用Western Blotting、Real-time PCR和免疫组化等技术检测肾脏组织纤维化TGF-β1/Smad3通路以及miRNA21和miRNA29b的表达。结果:糖肾方可以改善db/db小鼠尿微量白蛋白水平,减轻小鼠肾脏损伤;Western Blotting、Real-time PCR和免疫组化结果显示,糖肾方可下调db/db小鼠肾脏中纤维化标志分子TGF-β1、Smad3、CollagenⅠ和Fibronectin的表达;下调db/db小鼠肾脏miRNA 21的表达,上调miRNA 29b的表达。结论:糖肾方可作用于TGF-β1/Smad3信号通路抑制肾脏纤维化。

Objective： To observe the effects of Tangshen Formula（ TSF） treatment on renal fibrosis and explore its potential mechanism in db/db mice.Methods： Eight weeks old db/db and db/m mice were randomly divided into three subgroups,（ db/m,db/db,db/db ＋ TSF）,and all mice were fed with a normal diet.After two weeks adaptation,mice were maintained on TSF treatment for 12 weeks and were then sacrificed under anesthesia after overnight fast.Animal urine and renal tissues samples were collected for further analyses.The kidneys were washed with cold PBS solution; one part of the renal tissue were stored in liquid nitrogen,and another deposited-80 ℃ directly for subsequent frozen sections; the rest of the kidney were fixed into the neutral formalin for pathological analysis.The renal fibrosis was detected by PAS and Masson staining.The renal expression of TGF-β1/Smad3 and its target genes were assessed by Western Blotting,real-time PCR,and IHC.Results： TSF diminished UACR of db/db mice.Histological analysis using PAS and Masson staining revealed the occurrence of mesangial matrix expansion and extracellular matrix deposition in the kidneys of db/db mice.The treatment with TSF significantly ameliorated these histological renal injuries in db/db mice.Western Blotting and real-time PCR analysis showed that expression levels of TGF-β1,Smad3,Collagen Ⅰ and Fibronectin were significantly downregulated in the db/db mice with TSF treatment.With TSF treatment,the expression levels of miRNA 21 was downregulated,but the expression levels of miRNA 29b was upregulated.Conclusion： TSF attenuated renal fibrosis in db/db mice,through suppression of TGF-β1/Smad3 pathway.