Hypoxia/Reoxygenation increases the permeability of endothelial cell monolayers: Role of oxygen radicals

Abstract We assessed the effect of hypoxia/reoxygenation on 14 C-albumin flux across endothelial monolayers. Cultured bovine pulmonary artery endoehtlial cells were grown to confluence on nitrocellulose filters (pore size 12 μm). The endothelialized filters were mounted in Ussing-type chambers which were filled with cell culture medium (M 199). Equimolar amounts (33 nM) of 14 C-labeled and unlabeled albumin were added to the “hot” and “cold” chambers, respectively. The monolayers were then exposed to successive periods (90 min) of normoxia (pO 2 20 mmHg), and reoxygenation (pO 2 145 mmHg). A gas bubbling system was used to control media pO 2 and to ensure adequate mixing. Four aliquots of culture media were taken during each period in order to calculate 14 C-albumin permeability across the endothelialized filter. In some experiments, either the xanthine oxidase inhibitor, oxypurinol (10 μM), or superoxide dismutase (600 U/mL), was added to the media immediately prior to the experiments. As compared to the normoxic control period, albumin permeability was 1.5 times higher during hypoxia ( p p