Biological processes occurring in post-ischemic vasculature commonly initiate events that produce failure of endogenous inflammatory systems, precipitating vascular and parenchymal organ injury. Hypoxia Inducible Factor-1 (HIF-1), a heterodimeric protein consisting of alpha and beta subunits activated in vascular endothelium during hypoxia, modulates expression of genes essential for cell survival (i.e., inducible nitric oxide synthase and heme oxygenase-1) in ischemic environments. Little research to date has established a role for HIF-1 in modulating inflammatory events that occur in post ischemic vasculature. New research leading to submission of this revised application reveals that stabilization of the alpha subunit of HIF-1 dramatically reduces infarct size and polymorphonuclear neutrophil infiltration in post-ischemic rabbit and murine hearts, lnterleukin-8 (IL-8), a pro-inflammatory chemokine consistently associated with post-ischemic vascular injury and organ damage was strikingly repressed by HIF-1 activation in our studies. In vitro experiments using human microvascular endothelial cells suggest that HIF-1 modulates secretion of IL-8 acting directly at a transcriptional level or indirectly through generation of nitric oxide (NO) and carbon monoxide (CO) following induction of nitric oxide synthase and heme oxygenase-1 (HO-1) genes respectively. We hypothesize that: Hypoxia inducible factor-1 acting independently or in concert with key effector genes regulates chemokine expression in reoxygenating microvascular endothelium.
Four aims are proposed in this application: (1) To determine the role of HIF-1 activation in attenuation of cardiac chemokine generation following ischemia/reperfusion; (2) To examine molecular interdependency between NO and CO in regulation of IL-8 expression in reoxygenating microvascular endothelium; (3) To evaluate the role of HIF-1 in modulating HO-1-induced IL-8 generation; (4) To determine the functional significance of IL-8 promoter binding by HIF-1 in microvascular endothelium. The research proposed here will produce substantial new knowledge, establishing HIF-1 as a pivotal molecule in modulation of inflammatory events. Further, the results of this work will open a new era of understanding of the regulation of chemokine induced inflammation and permit the rational design of tools targeted specifically to attenuate post-ischemic vascular injury. ? ?
