Ichikawa H, Flores S, Kvietys P R, Wolf R E, Yoshikawa T, Granger D N, Aw T Y
Department of Physiology, LSU Medical Center, Shreveport, La 71130-3932, USA.
Circ Res. 1997 Dec;81(6):922-31. doi: 10.1161/01.res.81.6.922.
The objectives of this study were to (1) determine the time course of neutrophil adhesion to monolayers of human umbilical vein endothelial cells (HUVECs) that were exposed to 60 minutes of anoxia followed by 30 to 600 minutes of reoxygenation and (2) define the mechanisms responsible for both the early (minutes) and late (hours) hyperadhesivity of postanoxic HUVECs to human neutrophils. The results clearly demonstrate that anoxia/reoxygenation (A/R) leads to a biphasic increase in neutrophil adhesion to HUVECs, with peak responses occurring at 30 minutes (phase 1) and 240 minutes (phase 2) after reoxygenation. Oxypurinol and catalase inhibited phase-1 adhesion, suggesting a role for xanthine oxidase and H2O2. In comparison, platelet activating factor (PAF) contributed to both phases of neutrophil adhesion. Anti-intercellular adhesion molecule-1 (ICAM-1) and anti-P-selectin antibodies (monoclonal antibodies [mAbs]) attenuated phase-1 neutrophil adhesion, consistent with roles for constitutively expressed ICAM-1 and enhanced surface expression of preformed P-selectin. Phase-2 neutrophil adhesion was attenuated by an anti-E-selectin mAb, indicating a dominant role of this adhesion molecule in the late phase response. Pretreatment with actinomycin D and cycloheximide or with competing ds-oligonucleotides containing the nuclear factor-kappa B or activator protein-1 cognate DNA sequences significantly attenuated phase-2 response, suggesting a role for de novo macromolecule synthesis. Surface expression of ICAM-1, P-selectin, and E-selectin on HUVECs correlated with the phase-1 and -2 neutrophil adhesion responses. Collectively, these findings indicate that A/R elicits a two-phase neutrophil-endothelial cell adhesion response that involves transcription-independent and transcription-dependent surface expression of different endothelial cell adhesion molecules.
（1）确定中性粒细胞与人脐静脉内皮细胞（HUVECs）单层的黏附时间进程，其中HUVECs先经历60分钟缺氧，随后再进行30至600分钟复氧；（2）明确缺氧后HUVECs对人中性粒细胞早期（数分钟）和晚期（数小时）高黏附性的机制。结果清楚地表明，缺氧/复氧（A/R）导致中性粒细胞与HUVECs的黏附呈双相增加，复氧后30分钟（第1阶段）和240分钟（第2阶段）出现峰值反应。氧嘌呤醇和过氧化氢酶抑制第1阶段的黏附，提示黄嘌呤氧化酶和H2O2发挥了作用。相比之下，血小板活化因子（PAF）对中性粒细胞黏附的两个阶段均有作用。抗细胞间黏附分子-1（ICAM-1）抗体和抗P-选择素抗体（单克隆抗体[mAbs]）减弱了第1阶段的中性粒细胞黏附，这与组成性表达的ICAM-1以及预先形成的P-选择素表面表达增强的作用一致。第2阶段的中性粒细胞黏附被抗E-选择素单克隆抗体减弱，表明该黏附分子在晚期反应中起主导作用。用放线菌素D和环己酰亚胺或用含有核因子-κB或活化蛋白-1同源DNA序列的竞争性双链寡核苷酸预处理可显著减弱第2阶段反应，提示从头合成大分子发挥了作用。HUVECs上ICAM-1、P-选择素和E-选择素的表面表达与第1阶段和第2阶段的中性粒细胞黏附反应相关。总体而言，这些发现表明A/R引发了两阶段的中性粒细胞-内皮细胞黏附反应，该反应涉及不同内皮细胞黏附分子的转录非依赖性和转录依赖性表面表达。
