Oxygen deprivation initiates a series of metabolic, biochemical and molecular adaptations to the hypoxic environment. Upregulation of key genes linked to the inflammatory response and thrombosis follow induction of local hypoxemia. In diabetes, the heart is exquisitely sensitive to hypoxic stresses. Increased in the diabetic heart is accumulation both of Advanced Glycation Endproducts (AGEs), the products of nonenzymatic glycation/oxidation of proteins, and S100/calgranulins, a family of proinflammatory cytokines, ligands of Receptor for AGE (RAGE). RAGE is expressed to enhanced degrees in the diabetic heart, particularly in endothelial cells (EC), and infiltrating inflammatory cells, such as mononuclear phagocytes (MP). To assess the impact of oxygen deprivation on the diabetic heart, diabetic and euglycemic ape E (0) mice were subjected to hypoxia (O2 equal approximately to 6 mm Hg) for 6 hrs, or normoxia. Hearts were retrieved; microarray gene analysis revealed marked upregulation of mRNA for early growth response-1 (egr-1), especially in the hearts of diabetic mice exposed to hypoxia versus hypoxic hearts retrieved animals or normoxic controls (confirmed by Northern blotting). Egr-1 regulates expression of divergent gene families linked to the inflammatory/prothrombic response in hypoxia. Consistent with a key role for RAGE in modulating upregulation of egr-1in the hypoxic diabetic heart, daily administration of truncated soluble (s)RAGE (the extracellular ligand-binding domain of RAGE) suppressed hypoxia-induced upregulation of egr-1 mRNA/antigen, as well as its nuclear translocation in hearts retrieved from diabetic apo E (0) mice. Further, expression of mRNA and antigen for a key target gene of egr-1 in hypoxia, JE/MCP-1, was similarly blunted. We hypothesize that RAGE-dependent mechanisms contribute to rapid upregulation of egr-1 in the hypoxic heart, and, thereby, critically modulate expression of proinflammatory and prothrombotic mediators in the hypoxic milieu. Our goaI will be to test the pathways by which activation of the ligand/RAGE axis regulates expression of and the biologic impact of egr-1 in hypoxia. Using homozygous RAGE (0) mice and mice bearing signalling deficient mutants of RAGE in EC and MP, we will test these concepts in global hypoxia and regional ischemia/reperfusion. If successful, our findings will elucidate novel roles forRAGE in the response to hypoxia, and, possibly, extend the potential impact of RAGE blockade to disorders characterized by hypoxic stress. This Project will share mouse models and ligand-RAGE reagents with the other projects. This Project will employ all three Cores of this program.
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