基于Klotho调控的TGF-β1/Egr1信号通路探讨肾元颗粒对db/db糖尿病肾病小鼠肾脏保护作用

Renal protective effects of Shenyuan Granules on db/db diabetic nephropathy mice based on TGF-β1/Egr1 signaling pathway regulated by Klotho

目的:探讨肾元颗粒对db/db糖尿病肾病(DN)小鼠肾脏保护的可能作用机制。方法:将db/db小鼠随机分为模型组、肾元颗粒组、厄贝沙坦组,同窝野生(WT)小鼠作为空白对照组。观察各组小鼠肾脏病理变化;RT-PCR法检测小鼠肾组织Klotho、Egr1、TGF-β1、Fibronectin、CollagenⅠmRNA表达;Western blot检测肾组织Egr1、TGF-β1、Fibronectin、CollagenⅠ蛋白表达;免疫荧光法检测肾脏Klotho蛋白表达。结果:与空白对照组比较,模型组血清BUN、Scr水平显著升高(P<0.01),病理改变较重,Egr1、TGF-β1、Fibronectin、CollagenⅠmRNA和蛋白表达显著增加(P<0.01),Klotho mRNA表达显著减少(P<0.01)。与模型组比较,厄贝沙坦组和肾元颗粒组小鼠血清BUN、Scr水平显著降低(P<0.01,P<0.05),病理改变均较模型组轻,Egr1、TGF-β1、Fibronectin、CollagenⅠmRNA表达显著降低(P<0.01),Klotho mRNA和蛋白表达显著增加(P<0.01)。结论:肾元颗粒能够改善db/db DN小鼠的肾功能,其作用机制可能与Klotho调控的TGF-β1/Egr1信号通路有关。

Objective: To investigate the possible mechanism of Shenyuan Granules on renal protection in db/db diabetic nephropathy(DN) mice. Methods: Mice were randomly divided into model group, Shenyuan Granules group, irbesartan group, and wild type(WT) group. HE and PASM staining were used to observe the pathological changes of kidneys;Klotho, Egr1, TGF-β1, Fibronectin and Collagen Ⅰ mRNA was detected by RT-PCR in mice kidney tissues. Egr1, TGF-β1, Fibronectin and Collagen Ⅰ proteins were detected by Western Blot. Klotho protein in kidney tissues was detected by immunofluorescence assay. Results: Compared with the Western blot group, serum BUN and Scr levels were significantly increased(P<0.01), the pathological changes were more serious, the expressions of Egr1, TGF-β1, Fibronectin, Collagen Ⅰ mRNA and protein were increased significantly(P<0.01) and the expression of Klotho mRNA and protein in model group was significantly decreased(P<0.01). Compared with the model group, the serum BUN and Scr in the irbesartan group and the Shenyuan Granules group were significantly decreased(P<0.01, P<0.05), and the pathological changes were lighter. The expressions of Egr1, TGF-β1, Fibronectin and Collagen Ⅰ mRNA and protein were significantly decreased(P<0.01), and the expression of Klotho mRNA and protein was significantly increased(P<0.01). Conclusion: Shenyuan Granules has improved renal function in db/db mice with diabetic nephropathy, and its mechanism may be related to Klotho-regulated TGF-β1/Egr1 signaling pathway.