白藜芦醇对链脲佐菌素诱导糖尿病小鼠肾脏损伤的保护作用

Protective Effect of Resveratrol on Kidney Injury in Diabetic Mice Induced by Streptozotocin

目的观察白藜芦醇对链脲佐菌素诱导的Ⅰ型糖尿病小鼠肾脏损伤的保护作用,并探讨其可能的作用机制。方法2018年6月—2019年11月期间取6~8周龄C57BL/6小鼠60只,其中正常小鼠20只,腹腔注射链脲霉素(STZ)诱导的糖尿病小鼠40只。实验所用C57BL/6小鼠分为3组:正常对照组(20只)、糖尿病组(20只)和糖尿病白芦藜醇治疗组(20只)。于糖尿病小鼠建模后5个月检测各组小鼠24 h蛋白尿、血液尿素氮(BUN)及血肌酐(Cr)等肾功能相关指标。评估各组小鼠肾脏肥大指数(肾重/体重)水平。利用试剂盒法检测各组小鼠肾脏皮质超氧化物歧化酶(SOD)活性、脂质过氧化代谢产物丙二醛(MDA)含量。取各组小鼠肾脏组织病理切片观察肾脏病理改变。采用TUNEL凋亡试剂盒检测各组小鼠肾脏组织细胞凋亡情况。Western blot及Real time-PCR法检测各组小鼠肾脏皮质血管内皮生长因子(VEGF),和核因子κB(NF-κB)等关键因子表达水平。利用免疫荧光组织化学染色技术观察各组小鼠肾脏组织VEGF表达情况。结果糖尿病白藜芦醇治疗组小鼠24 h尿蛋白、尿素氮(BUN)、血肌酐(Cr)和脂质过氧化代谢产物丙二醛(MDA)含量水平较糖尿病组明显降低,差异有统计学意义(P<0.05);糖尿病白藜芦醇治疗组小鼠肾脏皮质超氧化物歧化酶(SOD)活性较糖尿病组小鼠明显升高;HE病理染色显示白芦藜醇治疗组小鼠肾脏损害较糖尿病组小鼠减轻;TUNEL凋亡染色显示糖尿病白芦藜醇治疗组肾脏细胞凋亡水平较糖尿病组明显减轻;Western blot和RT-PCR结果显示,糖尿病白藜芦醇治疗组VEGF与NF-κB mRNA和蛋白表达水平较糖尿病组明显降低,差异有统计学意义(P<0.05);免疫荧光染色结果显示,糖尿病白藜芦醇治疗组VEGF荧光强度较糖尿病组明显降低。结论白藜芦醇能够明显改善糖尿病小鼠肾脏损伤,其作用机制可能是通过减少肾脏组织细胞凋亡、抗VEGF表达及抗炎作用。

Objective To observe the protective effect of resveratrol on streptozotocin-induced kidney damage in type I diabetic mice,and to explore its possible mechanism of action.Methods 60 c578BL/6 miceaged 6~8 weeks were collected from June 2018 to November 2019,including 20 normal mice and 40 diabetic mice induced by intraperitoneal injection of streptozotocin(STZ).The C57BL/6 mice used in the experiment were divided into three groups:normal control group(20),diabetic group(20)and diabetic resveratrol treatment group(20).5 months after the diabetic mice were modeled,the 24-hour proteinuria,blood urea nitrogen(BUN),blood creatinine(Cr)and other related indicators of renal function were detected in each group.The level of renal hypertrophy index(kidney weight/body weight)of mice in each group was evaluated.The kit method was used to detect the activity of superoxide dismutase(SOD)and the content of malondialdehyde(MDA)of the lipid peroxidation metabolite in the kidney cortex of each group.Pathological sections of the kidney tissues of each group of mice were taken to observe the pathological changes of the kidneys.TUNEL apoptosis kit was used to detect the apoptosis of kidney tissue cells in each group of mice.Western blot and Real time-PCR Methods were used to detect the expression levels of key factors such as vascular endothelial growth factor(VEGF)and nuclear factor kB(NF-κB)in the kidney cortex of each group.Immunofluorescence histochemical staining technique was used to observe the expression of VEGF in the kidney tissue of mice in each group.Results 24 h urine protein,urea nitrogen(BUN),blood creatinine(Cr)and lipid peroxidation metabolite malondialdehyde(MDA)levels of mice in the diabetic resveratrol treatment group were significantly lower than those in the diabetic group,the difference was statistically significant(P<0.05);The activity of superoxide dismutase(SOD)in the renal cortex of diabetic resveratrol-treated mice was significantly higher than that of diabetic mice;HE pathological staining showed that the kidneys of resveratrol-treated mice were more damaged than diabetic mice TUNEL apoptosis staining showed that the level of renal cell apoptosis in the diabetic resveratrol treatment group was significantly less than that in the diabetic group;Western blot and RT-PCR Results showed that the expression of VEGF and NF-κB mRNA and protein in the diabetic resveratrol treatment group was significantly lower than that of the diabetes group,the difference was statistically significant(P<0.05);the immunofluorescence staining results showed that the VEGF fluorescence intensity of the diabetic resveratrol treatment group was significantly lower than that of the diabetes group.Conclusion Resveratrol can significantly improve kidney damage in diabetic mice,and its mechanism of action may be through reducing kidney tissue cell apoptosis,anti-VEGF expression and anti-inflammatory effects.