Adhesion of flowing neutrophils to cultured endothelial cells after
hypoxia and re-oxygenation in vitro.
Rainger, G. E ., A. Fisher, C. Shearman, and G. B. Nash.
Department of Physiology, The Medical School, University of
Birmingham, Birmingham, Bioengineering Unit, Wolfson Centre,
University of Strathclyde, Glasgow and Department of Vascular
Surgery, Queen Elizabeth Hospital, Edgbaston, Birmingham, U.K.
APStracts 2:0212H, 1995.
Using a novel in line de-oxygenating system linked to an in vitro flow
based adhesion assay and video microscopy, we have studied neutrophil
recruitment and migration after hypoxia and re-oxygenation of
cultured human umbilical vein endothelial cells (HUVEC).
Unstimulated, purified neutrophils were perfused over re-oxygenating
HUVEC immediately following various periods of endothelial hypoxia.
Adhesion to HUVEC was dependent upon the duration of hypoxia, with
30, 60 and 100 minutes of exposure causing graded increments in
neutrophil recruitment. The degree of hypoxia also markedly
influenced the endothelial response. Severe hypoxia (O2 {SYMBOL 60 \f
"Symbol"} 2.5 %) induced stationary attachment followed by
migration of neutrophils, in contrast to rolling adhesion alone under
a less intense regime (O2 = 2.5 - 4.0 %). Judged from studies with
monoclonal antibodies, P-selectin was essential for adhesion after
severe hypoxia, and neutrophil immobilisation was attributable to the
activation of neutrophil {SYMBOL 98 \f "Symbol"}2 integrin.
Perfusion of neutrophils with an antibody against interleukin-8 (IL
-8) or a platelet activating factor (PAF) antagonist both reduced
levels of adhesion. However, IL-8 appeared to be the dominant agent
involved in the immobilisation from flow, while PAF was the more
potent in initiating sub-endothelial migration. Thus endothelial
cells alone can initiate all stages of adhesion and migration of
flowing neutrophils after hypoxia and reperfusion.
Received 29 December 1994; accepted in final form 18 May 1995.
APS Manuscript Number H1146-4.
Article publication pending Am. J. Physiol. (Heart Circ. Physiology).
ISSN 1080-4757 Copyright 1995 The American Physiological Society.
Published in APStracts on 30 May 1995.