Ischemia and reperfusion trigger the rapid release of endothelial granules containing mediators of vascular inflammation and thrombosis, including P-selectin, interleukin-8, and von Willebrand factor. P-selectin is translocated from the interior of the endothelial cell to the exterior surface, where it mediates leukocyte adherence to the endothelial wall. Interleukin-8 is released into the blood, where it can activate leukocytes, increasing the interactions between leukocytes and endothelial cells. Endothelial exocytosis is thus an early step in leukocyte trafficking into the ischemic myocardium. The over-all goal of this project is to characterize the mechanisms by which exocytosis of endothelial granules leads to injury in the post-ischemic myocardium. We have previously identified components of the exocytic machinery of endothelial cells. We subsequently discovered that nitric oxide inhibits vascular inflammation by inhibiting specific proteins that mediate exocytosis. We then developed a novel polypeptide that interferes with the exocytic machinery, inhibits exocytosis, and decreases vascular inflammation. We now propose to extend these studies to explore the mechanisms of exocytosis in post-ischemic myocardium. We first plan to study how hypoxia activates exocytosis. We will focus on the role of signaling intermediates such as calcineurin which regulate exocytosis by post-translational modifications of the exocytic machinery. We will then characterize the role of one particular component of the exocytic machinery in postischemic inflammation. We will finally define a novel mechanism which regulates a major stress response pathway in hypoxic endothelial cells. Each of these aims includes a wide range of studies, from recombinant proteins to transduced cells to animal experiments. These studies will increase our understanding of the molecular mechanisms regulating endothelial exocytosis, and will lead to novel therapies that inhibit inflammation and injury to the post-ischemic myocardium.

National Institute of Health (NIH)
National Heart, Lung, and Blood Institute (NHLBI)
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Heart, Lung, and Blood Initial Review Group (HLBP)
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Johns Hopkins University
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