Mechanisms involved in the contraction of endothelial cells by hydrogen peroxide

Abstract

The importance of endothelial contraction in the genesis of inflammatory edema has been reported. ROS are metabolites synthesized in pathological conditions in that a significant intravascular fluid leak occurs, such as ischemia-reperfusion. Present experiments were designed to test the hypothesis that ROS, particularly H 2O 2, may elicit the contraction of endothelial cells, and to explore the mechanisms involved. Bovine aortic endothelial cells incubated with H 2O 2 showed a significant reduction in planar cell surface area (PCSA), and a significant increase in myosin light chain phosphorylation (MLCP), with a time- and dose-dependent pattern, without any significant toxicity. This effect of H 2O 2 was not blocked by sulotroban (TxA 2 antagonist) or BN 52021 (PAF antagonist). Lanthanum chloride (calcium channel blocker) and EGTA partially inhibited the increase in MLCP induced by H 2O 2. H7 and staurosporine, PKC inhibitors, and PKC down-regulation (phorbol myristate acetate treatment, 24 h) also blocked H 2O 2-dependent endothelial contraction, measured as PCSA or MLCP. H 2O 2 increased the intracellular calcium concentration, an effect blunted by EGTA and lanthanum chloride. H 2O 2 also increased the phosphorylation of an 80 kD polypeptide, probably MARCKS, a PKC substrate. In summary, the present results demonstrate the ROS-dependent contraction of endothelial cells, an effect that could explain the intravascular fluid leak observed in some pathophysiological situations. Calcium and PKC may be involved in the development of this contraction.