大黄提取物对脓毒症大鼠急性肾损伤的保护作用和对Keap1/Nrf2信号通路的影响

Protective effect of rhubarb extract on acute kidney injury and its effect on Keap1/Nrf2 signaling pathway in sepsis rats

目的:本文旨在探究大黄提取物对脓毒症大鼠急性肾损伤的保护作用和对Keap1/Nrf2信号通路的影响。方法:建立脓毒症大鼠急性肾损伤模型将大鼠随机分为5组:对照组、急性肾损伤(AKI)组、AKI+RE 50 mg·kg^(-1)组、AKI+RE 100 mg·kg^(-1)组和AKI+RE 200 mg·kg^(-1)组进行后续实验。全自动生化分析仪检测血清中肌酐和尿素氮水平;尿蛋白定量测试盒检测24 h尿蛋白;HE染色检测肾组织病理损伤程度;TUNEL染色检测肾组织凋亡;流式分选检测外周血中F4/80+i NOS+、F4/80+IL-10+含量;ELISA检测外周血中TNF-α、IL-6含量;蛋白免疫印迹检测Caspase-3、cleaved Caspase-3、Bax、Bcl-2、Keap1、Nrf2、p-Nrf2、HO-1蛋白表达水平。结果:结果表明,与对照组相比较,AKI组24 h尿蛋白、血清肌酐、尿素氮水平显著升高(P<0.05),细胞凋亡率显著升高(P<0.05),cleaved Caspase-3/Caspase-3、Bax/Bcl-2比值显著升高(P<0.05),F4/80+i NOS+、F4/80+IL-10+含量显著升高(P<0.05),TNF-α、IL-6含量显著升高(P<0.05),Keap1、HO-1蛋白水平和p-Nrf2/Nrf2比值显著降低(P<0.05)。与AKI组相比较,AKI+RE 100、200mg·kg^(-1)组24h尿蛋白、血清肌酐、尿素氮水平显著降低(P<0.05),病理损伤程度明显改善,细胞凋亡率显著降低(P<0.05),cleaved Caspase-3/Caspase-3、Bax/Bcl-2比值显著降低(P<0.05),F4/80+i NOS+含量显著降低(P<0.05),F4/80+IL-10+含量显著升高(P<0.05),TNF-α、IL-6含量显著降低(P<0.05),Keap1、HO-1蛋白水平和p-Nrf2/Nrf2比值显著升高(P<0.05)。结论:大黄提取物通过介导Keap1/Nrf2信号通路改善肾损伤,抑制肾组织细胞凋亡、炎症因子的释放对脓毒症大鼠急性肾损伤模型起保护作用。

objective:To investigate the protective effect of rhubarb extract on acute kidney injury and the effect of Keap1/Nrf2 signaling pathway in sepsis rats.Methods:The Sepsis rat model of acute kidney injury was established and the rats were randomly divided into 5 groups:Control group,AKI group,AKI+RE 50 mg·kg^(-1)group,AKI+RE 100 mg·kg^(-1)group and AKI+RE 200 mg·kg^(-1)group for follow-up experiments.Serum creatinine and urea nitrogen were measured by automatic biochemical analyzer.24 h urine protein was detected by quantitative test box.HE staining was used to detect the degree of pathological damage of renal tissues.Apoptosis of renal tissue was detected by TUNEL staining.The contents of F4/80+i NOS+and F4/80+IL-10+in peripheral blood were detected by flow cytometry.TNF-αand IL-6 in peripheral blood were detected by ELISA.The expression levels of Caspase-3,cleaved Caspase-3,Bax,Bcl-2,Keap1,Nrf2,p-Nrf2 and HO-1 were detected by western blotting.Results:The results showed as follows:compared with Control group,the levels of 24 h urine protein,serum creatinine and urea nitrogen in AKI group were significantly increased(P<0.05),the cell apoptosis rate was significantly increased(P<0.05),and the ratios of cleaved Caspase-3/Caspase-3 and Bax/Bcl-2 were significantly increased(P<0.05).The contents of F4/80+i NOS+and F4/80+IL-10+were significantly increased(P<0.05),the contents of TNF-αand IL-6 were significantly increased(P<0.05),the protein levels of KEAP1 and HO-1 and the p-Nrf2/Nrf2 ratio were significantly decreased(P<0.05).Compared with AKI group,the levels of 24 h urine protein,serum creatinine and urea nitrogen in AKI+Re 100 and 200 mg·kg^(-1)groups were significantly decreased(P<0.05),the degree of pathological injury was significantly improved,and the rate of apoptosis was significantly decreased(P<0.05).The cleaved Caspase-3/Caspase-3,Bax/Bcl-2 ratio were significantly decreased(P<0.05),the content of F4/80+i NOS+was significantly decreased(P<0.05),and the content of F4/80+IL-10+was significantly increased(P<0.05).The contents of TNF-αand IL-6 were significantly decreased(P<0.05),the protein levels of KEAP1 and HO-1 and the p-Nrf2/Nrf2 ratio were significantly increased(P<0.05).Conclusion:Rhubarb extract can improve renal injury by mediating Keap1/Nrf2 signaling pathway,inhibit renal cell apoptosis and release of inflammatory factors,and have protective effects on sepsis rat model of acute renal injury.