高尿酸对血管内皮细胞eNOS基因表达的调节作用及其对血管新生的影响

Effect of high uric acid on eNOS gene expression and angiogenesis in vascular endothelial cells

目的 探讨高尿酸对血管内皮细胞内皮型一氧化氮合酶(eNOS)基因表达调节作用及其对血管新生的影响.方法 人脐静脉血管内皮细胞(HUVECs)在600μmol/L尿酸溶液中培养,应用RT-PCR和Western印迹分别检测各组细胞eNOS mRNA和蛋白表达,应用硝酸还原法检测各组细胞上清液中一氧化氮(NO)含量,应用人工基底膜检测细胞的管腔形成能力,应用鸡胚尿囊膜试验检测细胞的血管新生能力,应用四甲基偶氮唑蓝(MTT)法检测各组HUVECs的增殖,应用划痕试验检测各组HUVECs的迁移能力.结果 与对照组比较,600μmol/L的高尿酸培养HUVECs 48 h后,细胞eNOS mRNA转录水平、蛋白表达量、NO浓度显著下降(P<0.01);HUVECs有聚集倾向,但未能形成管腔样网络结构;CAM的血管新生面积比率降低49.78%(P<0.01);同时,细胞的增殖能力降低63.68%(P<0.05),细胞的迁移能力下降38.70%(P<0.01).结论 高尿酸抑制血管内皮细胞eNOS基因表达和NO含量;高尿酸抑制细胞的血管新生能力,同时抑制细胞增殖与迁移,其抑制的可能机制与高尿酸下调eNOS的表达有关.

Objective To investigate the effect of hyperuricemia on eNOS gene expression and angiogenesis in vascular endothelial cells.Methods HUVECs were cultured in 600μmol/L uric acid solution,the expression of eNOS gene and protein was detected by RT-PCR and Western blot.The content of NO in cell supernatant was detected by nitric acid reduction method.The tube formation of cell was detected by artificial basement membrane.The angiogenesis of cell was detected by Chicken embryo allantoic membrane test.The proliferation of HUVECs in each group was detected by MTT method and the migration ability of HUVECs was detected by scratch test.Results After culturing HUVECs with 600μmol/L of hyperuricemia for 48 hours,compared with the control group,the transcriptional level of eNOS mRNA was decreased by 48.51%(P<0.01),and the expression of eNOS protein was decreased by 55.00%(P<0.01);the concentration of NO was decreased by 45.09%(P<0.01).HUVECs were tended to aggregate,but failed to form lumen-like network structure.The ratio of neovascularization area of CAM decreased by 49.78%(P<0.01).Meanwhile,the cell proliferation ability was decreased by 63.68%(P<0.05)and the cell migration ability was decreased by 38.70%(P<0.01).Conclusions High uric acid inhibits the expression of eNOS gene and the content of NO in vascular endothelial cells.High uric acid inhibits the angiogenesis of cells,and inhibits the proliferation and migration of cells.The mechanism of the inhibition might be related to the down-regulation of eNOS expression.