5脂氧合酶激活蛋白介导人脐静脉内皮细胞氧化损伤

Five Lipoxygenase Activating Protein-Mediated Human Umbilical Vein Endothelial Cells Oxidation Injury Induced by Oxidized Low Density Lipoprotein

目的探讨5脂氧合酶激活蛋白在人脐静脉内皮细胞氧化损伤中的机制。方法体外分离、培养人脐静脉内皮细胞,分别用10、50、100和200mg/L氧化型低密度脂蛋白处理细胞24h,采用酶联免疫吸附法检测细胞上清液白三烯C4水平,MTT法检测细胞活力,以及实时荧光定量PCR和Western blot检测细胞5脂氧合酶激活蛋白mRNA及蛋白表达的变化。结果100mg/L和200mg/L氧化型低密度脂蛋白作用人脐静脉内皮细胞24h细胞活力显著下降(P<0.01),白三烯C4分泌显著增加(P<0.01),回归分析显示白三烯C4释放量与氧化型低密度脂蛋白浓度显著相关(r=0.953,P<0.01)。PCR和Western blot分析发现,5脂氧合酶激活蛋白mRNA表达和蛋白表达均显著增强(P<0.01和P<0.05)。结论氧化型低密度脂蛋白促进人脐静脉内皮细胞分泌白三烯C4,导致人脐静脉内皮细胞氧化损伤可能与5脂氧合酶激活蛋白异常表达有关。

Aim To investigate whether five lipoxygenase activating protein （FLAP） is involved in the mechanism of human umbilical vein endothelial cells （hUVEC） oxidation injury induced by oxidized low density lipoprotein （ox-LDL）. Methods Isolated and cultured hUVEC were used as experimental model, hUVEC were induced by ox-LDL at 10, 50, 100 and 200 mg/L for 24 hours. Le, dcotriene CA （LTC4） levels in the cell supematant fluid were detected with enzyme-linked immunosorbent assay （ELISA）, cell viability was detected by MTr assay, and FLAP mRNA and FLAP protein were detected with fluo- rescent quantitation PCR and Western blot. Results Ox-LDL at 100 and 200 mg/L significantly induced the release of LTC4 （ P 〈 0.01 ）, the decrease of cell viability in hUVEC （ P 〈 0.01 ） and upregulation of FLAP mRNA expression and FLAP protein expression. The regressive analysis showed that the amounts of released LTC4 had significantly positive correlation with the concentrationofox-LDL（r=0.953, P〈0.01）. Conclusion FLAP could be involved in the LTCA secretion and oxidative injury induced by ox-LDL in h UVEC.