This 5-year training program proposes the development plan for a career as an independent biomedical researcher in the area of myocardial ischemia and cardiac biology. The principal investigator, is a board certified Anesthesiologist and has completed a Critical Care Fellowship training. With the sponsors and experienced collaborators he will expand on his scientific skills in preparation for career progression as an independent physician-scientist. The program will emphasize skills in molecular biology of intracellular pathways in myocyte function and regulation of ischemia- adaptive processes using in vitro (isolated myocytes exposed to hypoxia) and in vivo (murine in situ myocardial ischemia) models. To advance his knowledge in metabolic biology he will attend courses offered by the Omics Technologies Program at the National Jewish, Denver. Thomas Henthorn, MD, the Departmental Head will provide sponsorship. Holger Eltzschig MD, PhD, an international expert in molecular biology of organ hypoxia and ischemia will be the dedicated mentor and provide sponsorship. The program will benefit from collaborative expertise of Sean Colgan PhD, a world expert in molecular mechanisms leading to adaptation to hypoxia, who will provide consultative support for the studies on intracellular pathways, posttranslational mechanisms and metabolic studies. Additionally, Peter Buttrick, MD and Michael Bristow MD, PhD, both world renowned cardiologists, will collaborate and serve with Drs. Eltzschig and Colgan on an advisory committee every 16 weeks. This committee will review progress and provide close scientific support and career advice. Myocardial ischemia (MI) is a permanent and serious medical problem. Over the last decades, convincing evidence has demonstrated a central role of adenosine generation in signaling in cardiac ischemic preconditioning (IP), a phenomenon known as the strongest in vivo-form of protection against myocardial ischemia. As such, many possible effectors have been identified to be critical for mediating IP of the heart. However, the pathway initiated by IP is completely unknown. In this study, we present data from a microarray where we identified two circadian rhythm proteins, Period 1 and Period 2, as adenosine dependant molecules. In the latter we could confirm an IP and adenosine dependant induction and stabilization of these two rhythm proteins. Following studies in Per2-/- mice revealed a functional role of Period in IP and ischemia of the heart. Per2-/- mice had bigger infarct sizes and abolished cardioprotection mediated by IP. Since circadian rhythm proteins are known to play a dominant role in metabolic processes and are reported to control mitochondrial metabolism, we next pursued studies on the main oxygen consuming enzyme in mitochondria, the ATP Synthase. We found that this enzyme was downregulated due to IP in wildtype, but not in Period deficient mice. Based on these findings we hypothesize that Periods mediate metabolic tissue adaptation which is central to cardiac IP.
Three specific aims were designed to address novel roles for Period in MI. (1) In the first aim, we propose to study regulatory mechanisms and consequences of Period stabilization in vitro by combining pharmacological and genetic approaches. (2) In the second aim, we will combine in vitro and in vivo studies to investigate metabolic pathways initiated by period stabilization. In the third aim, by utilizing tissue specific mice for Period, we will first elucidate the individual contribution of endothelium and cardiomyocytes to MI in vivo. Finally, we will target Period in the heart as therapeutic option to treat MI using intense light exposure. These studies are designed to shed new light on endogenous pathways that regulate cell injury during MI. Targeting such pathways will lay the groundwork for novel and specific therapeutic approaches in the treatment of MI, which are urgently needed to improve morbidity and mortality.
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