Elabela改善db/db小鼠肾损伤的作用及机制研究

Protective effects of Elabela on kidney injury in db/db diabetic mice and its possible mechanism

目的研究Elabela(ELA)改善自发性2型糖尿病db/db小鼠肾损伤的作用及其相关机制。方法将16只8周龄雄性db/db小鼠分为db/db组(n=8)和ELA组(n=8),选取db/m小鼠为对照组(n=8)。ELA治疗组按ELA21制剂5 mg·kg^-1·d^-1进行腹腔注射,db/db组和db/m组小鼠以等量生理盐水注射,干预8周。实验结束时采集小鼠血液、尿液,检测HbA1C、尿白蛋白/肌酐比值等指标。免疫组化法观察ELA在小鼠肾脏组织中的表达情况。HE染色、Masson染色观察肾脏组织病理形态变化。蛋白免疫印迹法检测肾脏组织Ⅳ型胶原蛋白(Col-Ⅳ)、转化生长因子β1(TGF-β1)表达及Yes相关蛋白(YAP)磷酸化水平。结果ELA在db/db小鼠肾组织中的表达明显低于db/m小鼠。ELA干预后db/db小鼠血压、尿白蛋白/肌酐水平明显降低(P<0.05),但体重、HbA1C无明显改变。db/db小鼠的肾小管上皮细胞水肿、管腔变小和胶原纤维沉积增加在ELA干预后减轻。db/db组小鼠肾组织TGF-β1、Col-Ⅳ蛋白表达较db/m组明显增加(0.98±0.08对0.68±0.10、1.10±0.14对0.51±0.08,均P<0.05),YAP磷酸化水平升高(3.38±0.72对0.81±0.13,P<0.05),ELA干预后,TGF-β1、Col-Ⅳ表达及YAP磷酸化水平显著下降(0.80±0.06、0.51±0.05、2.21±0.22,均P<0.05)。结论ELA可改善db/db小鼠肾损伤,延缓糖尿病肾病进展,其机制可能与调控YAP信号通路有关。

Objective To investigate the protective effects of Elabela(ELA)on the renal injury of db/db mice and its possible mechanism.Methods Sixteen eight-week-old male db/db mice were intraperitoneally injected with ELA(5 mg·kg^-1·day^-1)or equivalent normal saline(n=8)for 8 weeks.Eight age-matched male db/m mice received equivalent normal saline injection as normal control.At the end of the experiment,blood and urine samples were obtained for HbA1C and urinary albumin/creatinine(ACR)measurements.Immunohistochemistry was used to observe the expression of ELA.Histopathological changes in kidney tissue were observed by HE staining and Masson staining.The levels of collagen typeⅣ(Col-Ⅳ)and transforming growth factor-β1(TGF-β1)as well as Yes-associated protein(YAP)phosphorylation in kidney tissue were examined by western blot.Results Immunohistochemistry results showed that ELA expression was decreased in the renal tissue of db/db mice as compared with that of db/m mice(P<0.05).After ELA treatment,ACR and blood pressure were markedly decreased in db/db mice(P<0.05),but without significant changes in the body weight and HbA1C.Renal tubular epithelial cells edema,basement membrane thickening,and increased collagen fiber in db/db were improved by ELA administration.Compared with db/m mice,the levels of TGF-β1 and Col-Ⅳexpression,as well as YAP phosphorylation were significantly increased in renal tissue of db/db mice(0.98±0.08 vs 0.68±0.10,1.10±0.14 vs 0.51±0.08,3.38±0.72 vs 0.81±0.13,all P<0.05),which were down-regulated after ELA administration(0.80±0.06,0.51±0.05,2.21±0.22,all P<0.05).Conclusion ELA may improve the renal injury of db/db mice by regulating the signaling pathway of YAP,thereby delaying the development of diabetic nephropathy.