Coronary microvascular exchange is important for the maintenance of normal cardiac function. In acute inflammation and myocardial ischemia- reperfusion, coronary venular permeability to macromolecules is increased leading to dysfunction of the myocardium. Altered venular permeability is induced by two groups of inflammatory mediators that are either neutrophil-dependent or -independent. This project will extend the investigation to the complex effects of neutrophil-induced permeability changes in the heart. Neutrophils undergo kinetic and metabolic changes during inflammation and ischemia-reperfusion. The sequence of events involves adherence to venular endothelium, diapedesis, and release of various cytotoxic mediators. Venular hyperpermeability could be induced mechanically by the disruption of the endothelial barrier in the process of neutrophil adhesion and migration or chemically by the effects of neutrophil-derived factors. However, neither the relative importance of each event nor the specific contribution of each mediator released by neutrophils has been established. The primary reason for this lack of understanding is that current experimental approaches do not allow unequivocal identification of the mechanisms because of the difficulties to distinguish each event from the sequential changes and the limitations to eliminate the influencing factors (e.g. microvascular pressure and inflammatory cells). Therefore, the overall goal of this study is to identify the precise mechanisms of neutrophil-mediated alterations in microvascular permeability. Porcine coronary venules ranging from 20 to 50 micromoles in diameter will be isolated, cannulated, and perfused with fluorescently-labeled porcine neutrophils. Venular permeability to albumin and neutrophil dynamics will be simultaneously measured using fluorescent microscopic techniques. Three specific hypotheses will be tested: 1). Adherence of neutrophils to the venular wall may modify the endothelial barrier function, but is not a prerequisite for neutrophil-mediated hyperpermeability; 2). Neutrophil transvenular migration does not cause disruption of the endothelial barrier and subsequent macromolecular leakage; and 3). Neutrophil-induced venular hyperpermeability is mainly mediated by four groups of neutrophil- derived factors: toxic oxygen species, granule enzymes, membrane lipid metabolites, and nitrogen-oxygen radicals. Because the isolated vessel preparation allows precise measurements of neutrophil dynamics and venular permeability under controlled physical forces and chemical factors, the study will provide a unique insight into the mechanisms of physiological and pathophysiologicaI regulation of microvascular exchange.
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