内皮细胞微粒对人脐静脉内皮细胞ICAM-1表达和NF-κB活性的影响

Effects of endothelial microparticles on ICAM-1 expression and NF-κB activity in HUVECs

目的探讨内皮细胞微粒(EMPs)对人脐静脉内皮细胞(HUVECs)核转录因子-κB(NF-κB)活性变化及细胞间黏附分子-1(ICAM-1)表达的影响。方法将体外培养的HU-VECs分3大组:①EMPs不同时点观察组:用EMPs(终浓度105/ml)分别刺激细胞0、3、6、12、24 h;②EMPs不同剂量作用组:分别用终浓度为0、102、103、104、105/ml的EMPs刺激细胞24 h;③二硫代氨基甲酸吡咯烷(PDTC)干预组:在EMPs(终浓度105/ml)刺激前,与终浓度为10μmol/L的PDTC共同孵育30 min。用实时荧光定量PCR测定ICAM-1mRNA的表达,Western blot测定核蛋白NF-κB p65和ICAM-1蛋白的表达。结果 EMPs可通过活化NF-κB,使ICAM-1mRNA和蛋白呈剂量和时间依赖性表达上调。NF-κB特异性拮抗剂PDTC可显著抑制EMPs的此作用。结论在EMPs诱导的HUVECs炎性效应中,NF-κB参与调控炎性因子ICAM-1的表达。

Objective To explore the role of nuclear factor-κB （NF-κB） in expressing of intercellular adhesionmolecule 1 （ICAM-1） in cultured human umbilical vein endothelial cells （HUVECs） activated by endothelial mi- croparticles （EMPs）. Methods The cultured HUVECs were divided into three groups： different time points （0, 3, 6, 12, 24 h） of the observation group with EMPs of 105/ml; different doses of EMPs （0,102, 103, 104, 105/ml）with a fixed stimulation time of 24 h ; EMPs with the specific inhibitor of NF-κB p65, pyrrolidine dithiocarbamic acid （PDTC）. The ICAM-1 mRNA level was detected by quantitative real-time reverse transcriptase polymerase chain reaction （qRT-PCR）. Western blot analysis was performed to determine expression levels of NF-κB p65 and ICAM-1 protein. Results NF-κB activity and the expressions of ICAM-1 increased after the HUVECs were treated by EMPs in a dose-dependent and time-dependent manner. The expressions of ICAM-1 weakened after being trea- ted with PDTC. Conclusion NF-κB controlled the expression of proinflammatory cytokine ICAM-1 in the EMPs induced inflammatory effects on 1-IUVECs.