CELLULAR MECHANISMS OF ISCHEMIA PRECONDITIONING

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Ischemic preconditioning (IPC) refers to a phenomenon in which a tissue is rendered resistant to the deleterious effects of prolonged ischemia and reperfusion (I/R) by prior exposure to brief periods of vascular occlusion. Although first described over a decade ago, the mechanisms underlying the beneficial effects of IPC remain uncertain. However, preliminary data from our laboratory indicates that IPC prevents I/R-induced intestinal oxidant production, leukocyte/endothelial cell interactions, and venular protein leakage. Since the production of reactive oxygen metabolites from xanthine oxidase and other sources during reperfusion of ischemic intestine initiates the formation of chemotactic stimuli, induces the expression of endothelial cell adhesion molecules, and reduces intestinal levels of the potent antiadhesive agent nitric oxide, we hypothesize that IPC-induced reductions in leukocyte adhesion are mediated by the effect of preconditioning to reduce the postischemic oxidant production. To address this central postulate, four specific aims are proposed. One aim is to determine whether IPC prevents the increase in mesenteric xanthine oxidase activity or influence of IPC on the elaboration of proinflammatory stimuli (platelet activating factor (PAF), leukotriene B4(LTB4), complement activation) in postischemic small bowel. A third specific aim is to determine whether IPC attenuates the expression of P-selectin by the endothelium in postcapillary venules exposed to I/R. Since the beneficial effects of IPC on postischemic mesenteric microvascular dysfunction are reversed by topical application of adenosine deaminase during IPC or protein kinase C (PKC) inhibitors during the period of prolonged ischemia, fa fourth aim is directed at elucidating the mechanisms that may be involved in coupling adenosine production during IPC to the postischemic reductions in oxidant production, leukocyte adhesion, and venular protein leakage. To accomplish these aims, we will utilize intravital microscopic approaches to quantitate oxidant production, leukocyte/endothelial cell interactions, and venular protein leakage in rat mesentery. Mesenteric xanthine oxidase, nitric oxide synthase, phospholipase C, and PKC activities, nitrite/nitrate production, PAF and LTB4 levels, and the extent of complement activation will also be assessed. In vivo mesenteric P- selectin expression will be quantitated using a dual-radiolabeled monoclonal antibody approach. The proposed studies should not only substantially improve our understanding of the mechanisms whereby IPC reduces postischemic microvascular dysfunction in the intestine but should also provide a rationale for the development of novel therapeutic interventions in I/R that exploit the mediators of the preconditioning phenomenon.