© 1997 American Heart Association, Inc.
Articles |
Molecular Mechanisms of Anoxia/Reoxygenation-Induced Neutrophil Adherence to Cultured Endothelial Cells
From the Departments of Physiology and Medicine (H.I., S.F., D.N.G., T.Y.A.) and the Center of Excellence in Arthritis and Rheumatism (R.E.W.), LSU Medical Center, Shreveport, La; the A.C. Burton Vascular Research Laboratory (P.R.K.), Victoria Hospital Research Institute, London, Ontario, Canada; and the First Department of Internal Medicine (T.Y.), Kyoto (Japan) Prefectural University of Medicine.
Correspondence to Dr Tak Yee Aw, Department of Physiology, LSU Medical Center, 1501 Kings Highway, Shreveport, LA 71130-3932. E-mail taw{at}lsumc.edu
Abstract 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-
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.
Key Words: nuclear transcription factor • endothelial cell adhesion molecule • leukocyte–endothelial cell adhesion • selectin • intercellular adhesion molecule-1
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