Du Li-Jun, Zhou Jing-Guo, Ren Xing-Jun, Yi Ting-Ting, Jiang Xing-Liang
Institute of Rheumatology and Immunology, Affiliated Hospital of North Sichuan Medical College, Nanchong 637000, China.
Sichuan Da Xue Xue Bao Yi Xue Ban. 2013 Sep;44(5):717-21, 726.
To explore the oxidative mechanism of uric acid (UA) induced CRP expression in human umbilical vein endothelial cells.
Different concentrations of UA (0 mg/dL, 4 mg/dL, 8 mg/dL, 12 mg/dL, 16 mg/dl) were incubated 12 h with HUVECs, and HUVECs were stimulated with 12 mg/dl. UA for different times (6 h, 12 h, 24 h, 48 h). CRP mRNA and protein expression were determined by real-time quantitative polymerase chain reaction (RT-qPCR) and Western blot, respectively; the effects of uric acid on the intracellular reactive oxygen species (ROS) production in HUVECs were measured by fluorescence microscope and flow cytometric analysis using a 2', 7'-Dichlorofluorescin diacetate (DCF-DA) fluorescence probe. The effects of N-acetyl cysteine (NAC) on UA-induced levels of ROS, mRNA and protein of CRP in HUVECs were also observed.
The results demonstrated that UA could significantly increase the mRNA and protein expression of CRP in HUVECs in time- and concentration-dependent manners. HUVECs were stimulated with 12 mg/dL UA at 6 h, mRNA and protein levels of CRP significantly higher than that of control level (P<0.05), reached a peak at 12 h (P<0. 01). NAC reduced UA-induced levels of ROS, mRNA and protein of CRP in HUVECs compared with those of 12 mg/dL UA induced group(P<0. 05).
Uric acid significantly increased mRNA and protein expression of CRP in HUVECs in time- and concentration-dependent manners. Its mechanism may be associated with uric acid induced increasing of ROS levels in endothelial cells, which suggested that the uric acid mediated oxidative stress and inflammation may be involved in the injury of endothelial cells.
探讨尿酸（UA）诱导人脐静脉内皮细胞中CRP表达的氧化机制。
将不同浓度的UA（0mg/dL、4mg/dL、8mg/dL、12mg/dL、16mg/dl）与人脐静脉内皮细胞（HUVECs）孵育12小时，并用12mg/dl的UA刺激HUVECs不同时间（6小时、12小时、24小时、48小时）。分别通过实时定量聚合酶链反应（RT-qPCR）和蛋白质印迹法测定CRP mRNA和蛋白表达；使用2',7'-二氯荧光素二乙酸酯（DCF-DA）荧光探针，通过荧光显微镜和流式细胞术分析测量尿酸对HUVECs细胞内活性氧（ROS）产生的影响。还观察了N-乙酰半胱氨酸（NAC）对UA诱导的HUVECs中ROS水平、CRP mRNA和蛋白的影响。
结果表明，UA能以时间和浓度依赖性方式显著增加HUVECs中CRP的mRNA和蛋白表达。在6小时用12mg/dL UA刺激HUVECs，CRP的mRNA和蛋白水平显著高于对照水平（P<0.05），在12小时达到峰值（P<0.01）。与12mg/dL UA诱导组相比，NAC降低了UA诱导的HUVECs中ROS水平、CRP mRNA和蛋白水平（P<0.05）。
尿酸以时间和浓度依赖性方式显著增加HUVECs中CRP的mRNA和蛋白表达。其机制可能与尿酸诱导内皮细胞中ROS水平升高有关，这表明尿酸介导的氧化应激和炎症可能参与内皮细胞损伤。
