Reactive oxygen species (ROS) superoxide anion (O2-.) and hydrogen peroxide (H2O2) produced by activated leukocytes and endothelial cells in sites of inflammation or ischemia cause the endothelial barrier dysfunction that may lead to tissue edema. Antioxidant enzymes (AOE) superoxide dismutase (SOD) and catalase conjugated with antibodies to platelet-endothelial cell adhesion molecule-1 (PECAM-1) specifically bind to endothelium, quench the corresponding ROS and alleviate vascular oxidative stress and inflammation. In the present work, we studied effect of anti-PECAM/catalase and anti-PECAM/SOD conjugates on the abnormal permeability manifested by TEER decline, increased FITC-dextran influx and redistribution of VE-cadherin in human umbilical vein endothelial cell (HUVEC) monolayer. Anti-PECAM/catalase protected HUVEC monolayer against H2O2-induced endothelial barrier dysfunction. PEG-conjugated catalase exerted orders of magnitude lower endothelial uptake and no protective effect, similarly to IgG-catalase. Anti-PECAM/catalase, but not anti-PECAM/SOD, alleviated endothelial hyper-permeability caused by exposure to hypoxanthine/xanthine oxidase, implicating primarily H2O2 in disruption of endothelial barrier in this model. Thrombin-induced endothelial permeability was not affected by treatment with anti-PECAM/AOE or NADPH oxidase inhibitor apocynin or over-expression of AOEs, indicating that the endogenous ROS play no key role in thrombin-mediated endothelial barrier dysfunction. In contrast, anti-PECAM/SOD, but not anti-PECAM/catalase, inhibited VEGF-induced increase in endothelial permeability, identifying a key role of endogenous O2-. in the VEGF-mediated regulation of endothelial barrier function. Therefore, AOE targeted to endothelial cells provide versatile molecular tools for testing role of specific ROS in vascular pathology and may be translated into the remedy for these ROS-induced abnormalities.
- Received February 16, 2011.
- Revision received March 31, 2011.
- Accepted April 6, 2011.
- The American Society for Pharmacology and Experimental Therapeutics