高浓度尿酸对肾小球内皮细胞氧化应激及炎症反应的影响

Effects of high concentration of uric acid on oxidative stress and inflammatory response of glomerular endothelial cells

目的探讨高浓度尿酸对肾小球内皮细胞氧化应激及炎症反应的影响。方法取传5～15代、对数生长期、生长状态良好的人肾小球内皮细胞(HRGEC),接种于6孔板,随机分为阴性对照组、生理浓度尿酸组、中浓度尿酸组、高浓度尿酸组。各组分别给予0、4、8、16 mg/d L尿酸干预24 h,分别采用RT-PCR法、Western blotting法检测内皮型一氧化氮合酶(e NOS)、细胞间黏附分子1(ICAM-1)、单核细胞趋化蛋白1(MCP-1)、核因子κB(NF-κB)p56 mRNA和蛋白表达,采用荧光探针法检测细胞内活性氧(ROS)含量。结果阴性对照组、生理浓度尿酸组e NOS mRNA和蛋白相对表达量均明显低于中浓度尿酸组、高浓度尿酸组,ICAM-1、MCP-1、NF-κB p65 mRNA和蛋白相对表达量及ROS含量均明显高于中浓度尿酸组、高浓度尿酸组(P均<0. 05),而阴性对照组与生理浓度尿酸组、中浓度尿酸组与高浓度尿酸组上述指标比较P均> 0. 05。结论高浓度尿酸可能通过增加氧化应激途径中ROS产生和抑制e NOS表达,以及激活NF-κB信号通路促进炎症因子分泌等,导致肾小球内皮细胞损伤。

Objective To investigate the effects of high concentration of uric acid on oxidative stress and inflammatory response in glomerular endothelial cells.Methods Human glomerular endothelial cells(HRGECs)subcultured for 5-15 times with well condition were inoculated into 6-well plates in this study.HRGECs were randomly divided into four groups:the negative control group,physiological concentration uric acid group,medium concentration uric acid group,and high concentration uric acid group,which were given 0,4,8,and 16 mg/dL uric acid for 24 h,respectively.Then,we used real-time PCR and Western blotting to detect the mRNA and protein expression levels of endothelial nitric oxide synthase(eNOS),intercellular adhesion molecule 1(ICAM-1),monocyte chemotactic protein-1(MCP-1),and nuclear factor-κB p56(NF-κB p56).The reactive oxygen species(ROS)content was detected by fluorescent probe method.Results Compared with the medium concentration uric acid group and the high concentration uric acid group,the relative mRNA and protein expression levels of eNOS in the negative control group and the physiological concentration uric acid group significantly decreased(both P<0.05);the mRNA and protein expression levels of ICAM-1,MCP-1,NF-κB p65,and ROS content significantly increased in the negative control group and the physiological concentration uric acid group(all P<0.05).However,there was no significant deference in the above parameters between the negative control group,the physiological concentration uric acid group,medium concentration uric acid group and high concentration uric acid group(all P>0.05).Conclusion High concentration of uric acid may increase glomerular endothelial cell injury by increasing ROS production and inhibiting eNOS expression in the oxidative stress pathway,and activating NF-κB signaling pathway to promote inflammatory factor secretion.