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 myocardium. The altered venular permeability is induced by endogenous inflammatory mediators, and more importantly, by activated neutrophils. Upon inflammatory stimulation, neutrophils undergo sequential dynamic changes involving adherence to venular endothelium, diapedesis, and release of 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. Therefore, the overall goal of this study is to identify the precise mechanisms of neutrophil-mediated alterations in microvascular permeability. The permeability of venular endothelium in response to neutrophil activation will be quantified using the isolated porcine coronary venule preparation. In addition, in vitro tissue culture and immunoblot analysis will be employed to investigate at a subcellular level the signaling mechanisms of hyperpermeability induced by neutrophil-derived factors. Four major hypotheses will be tested: I) 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 endothelial barrier and subsequent macromolecular leakage; 3) neutrophil-induced venular hyperpermeability is mainly mediated by reactive oxygen species. granule enzymes, and lipid metabolites; and 4) the signaling pathway leading to neutrophil-dependent hyperpermeability involves the production of nitric oxide and phosphorylation of the endothelial cytoskeletal and junctional proteins. The study will provide a unique insight into the physiology of coronary exchange and pathogenesis of inflammatory and ischemic heart diseases.
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