利拉鲁肽对糖尿病肾病大鼠肾脏功能和足细胞损伤的改善作用及其机制

doi:10.13481/j.1671-587X.20220209

摘要/Abstract

摘要： 目的

观察利拉鲁肽对糖尿病肾病（DN）大鼠肾脏功能和足细胞相关蛋白表达及病理形态表现的影响，并探讨其作用机制。

方法

48只雄性SD大鼠随机选取12只作为正常对照组，其余36只大鼠采用腹腔注射链脲佐菌素（STZ）的方法制备糖尿病模型，随机分为模型组（n=12）、利拉鲁肽组（n=12）和利拉鲁肽+LY294002组（n=12）。利拉鲁肽组大鼠腹腔注射利拉鲁肽，利拉鲁肽+LY294002组大鼠提前给予LY294002腹腔注射，30 min后再给予利拉鲁肽腹腔注射。正常对照组和模型组大鼠腹腔注射等量生理盐水，各组每天注射1次。于用药8周后，测定各组大鼠体质量，检测各组大鼠尿白蛋白与肌酐比值（UACR）及空腹血糖（FBG）、血肌酐（Scr）和血尿素氮（BUN）水平；采用HE染色观察各组大鼠肾脏组织病理形态表现，免疫组织化学法检测各组大鼠肾脏足细胞中Synaptopodin蛋白表达水平，电镜观察各组大鼠肾脏组织足细胞形态表现，Western blotting法检测各组大鼠肾脏组织足细胞中Synaptopodin和磷脂酰肌醇3-激酶/蛋白激酶B（PI3K/Akt）信号通路相关蛋白表达水平。

结果

与正常对照组比较，模型组大鼠体质量明显降低（P<0.05），UACR、FBG、血清中Scr和BUN水平明显升高（P<0.05）；与模型组比较，利拉鲁肽组大鼠UACR、FBG、血清中Scr和BUN水平明显降低（P<0.05）。HE染色，模型组大鼠肾小球体积肿大，系膜细胞和基质增多，而利拉鲁肽组大鼠肾脏组织病理形态变化较模型组明显缓解；免疫组织化学染色，与正常对照组比较，模型组大鼠肾脏组织足细胞中Synaptopodin蛋白表达水平明显降低（P<0.05）；与模型组比较，利拉鲁肽组大鼠肾脏组织足细胞中Synaptopodin蛋白表达水平明显升高（P<0.05）。电镜下观察，正常对照组大鼠肾脏组织中足突覆盖于肾小球基底膜外表面，形态正常，分布均匀；模型组大鼠肾脏组织中足突增宽、肿胀、广泛融合，部分可见足突消失，溶酶体坏死，肾间质水肿；利拉鲁肽组大鼠肾脏组织中足突肿胀、融合好转，与正常对照组足突结构相似。Western blotting法检测，与正常对照组比较，模型组大鼠肾脏组织中Synaptopodin、PI3K和p-Akt蛋白表达水平明显降低（P<0.05）；与模型组比较，利拉鲁肽组大鼠肾脏组织中Synaptopodin、PI3K和p-Akt蛋白表达水平明显升高（P<0.05）；与利拉鲁肽组比较，利拉鲁肽+LY294002组大鼠肾脏组织中Synaptopodin、PI3K和p-Akt蛋白表达水平明显降低（P<0.05）。

结论

利拉鲁肽能通过激活PI3K/Akt信号通路改善大鼠肾脏功能，减轻DN大鼠足细胞损伤。

关键词: 糖尿病肾病, 足细胞, 胰高血糖素样肽1, 利拉鲁肽, 磷脂酰肌醇3-激酶, 蛋白激酶B

Abstract: Objective

To observe the effects of liraglutide on the renal function and the expressions of podocyte related proteins and the pathomorphologic changes in the rats with diabetic nephropathy （DN）， and to explore their mechanisms.

Methods

Twelve of the 48 male SD rats were randomly selected as normal control group （n=12）， and the remaining 36 rats were intraperitoneally injected with streptozotocin （STZ） to establish the diabetes models， and then randomly divided into model group （n=12）， liraglutide group（n=12） and liraglutide+LY294002 group（n=12）.The rats in liraglutide group were intraperitoneally injected with liraglutide， while the rats in liraglutide+LY294002 group were intraperitoneally injected with LY294002 in advance and then intravenously injected with liraglutide 30 min later； the rats in normal control group and model group were intraperitoneally injected with equal amount of saline， once a day for each group. After 8 weeks of treatment， the body weights of rats in various groups were determined， and the urinary albumin to creatinine ratios （UACR） and the levels of fasting blood glucose （FBG）， serum creatinine （Scr）and blood urea nitrogen （BUN） of the rats in various groups were detected. HE staining was used to observe the pathomorphology of kidney tissue of the rats in various groups. The expression levels of Synaptopodin protein in the podocytes of the rats in various groups were detected by immunohistochemistry method. The morphology of podocytes in kidney tissue of the rats were observed by electron microscope. The expression levels of Synaptopodin and phosphatidylinositol 3 kinase/ protein kinase B（PI3K/Akt） signaling pathway related proteins in kidney tissue of the rats in various groups were detected by Western blotting method.

Results

Compared with normal control group， the body weight of rats in model group was significantly reduced （P<0.05）， UACR， FBG， Scr and BUN levels were significantly increased （P<0.05）； compared with model group， UACR， FBG， Scr and BUN levels of the rats in liraglutide group were significantly decreased （P<0.05）. The HE staining results showed that the glomerular volumes in model group were enlarged and mesangial cells and matrix were increased， while the pathomorphologic changes in kidney tissue in liraglutide group were significantly ameliorated compared with model group. The immunohistochemical staining results showed that compared with normal control group， the expression level of Synaptopodin protein in the podocytes in kidney tissue of the rats in model group was significantly decreased （P<0.05）.Compared with model group， the expression level of Synaptopodin protein in podocytes in kidney tissue of the rats in liraglutide group was significantly increased （P<0.05）. In normal control group， the podocytic processes covered the outer surface of the glomerular basement membrane with normal shape and uniform distribution； in model group， the podocytic processes were widened， swollen and fused extensively， and some of the podocytic processes disappeared with lysosomal necrosis and renal interstitial edema； the podocytic processes in liraglutide group were swollen and fused better， and the structures of the podocytic processes were similar to those in normal control group. The Western blotting results showed that compared with normal control group， the expression levels of Synaptopodin， PI3K and p-Akt proteins in kidney tissue of the rats in model group were significantly decreased（P<0.05）；compared with model group， the expression levels of Synaptopodin， PI3K and p-Akt proteins in kidney tissue of the rats in liraglutide group were significantly increased （P<0.05）； compared with liraglutide group， the expression levels of Synaptopodin， PI3K and p-Akt proteins in kidney tissue in liraglutide + LY294002 group were significantly decreased （P<0.05）.

Conclusion

Liraglutide can improve the renal function and alleviate the podocyte injury in the rats with DN by activating PI3K/p-Akt signaling pathway.

Key words: Diabetic nephropathy, Podocyte, Glucagon like peptide-1, Liraglutide, Phosphatidylinositol 3-kinase, Protein kinase B

中图分类号:

R587.2

祁冰雪,王杨威,张艺献,赵静波,马彦,孙亚东. 利拉鲁肽对糖尿病肾病大鼠肾脏功能和足细胞损伤的改善作用及其机制[J]. 吉林大学学报(医学版), 2022, 48(2): 331-339.

Bingxue QI,Yangwei WANG,Yixian ZHANG,Jingbo ZHAO,Yan MA,Yadong SUN. Ameliorative effect of liraglutide on renal function and podocyte injury of rats with diabetic nephropathy and its mechanism[J]. Journal of Jilin University(Medicine Edition), 2022, 48(2): 331-339.

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