Of the many pathologic changes associated with myocardial ischemia, contractile dysfunction is one of the leading causes of postischemic morbidity and mortality. Contractile dysfunction can generally be classified into two categories, reversible, commonly called stunning, and irreversible, as in infarction. Understanding reversible injury provides the best means of developing interventions to prevent contractile dysfunction. The broad long term goals of this project are to demonstrate how protein oxidation can result in reversible contractile dysfunction. In this application we will examine the concept that contractile functional recovery is dependent on removal of oxidized proteins by the 20S-proteasome setting the stage for de novo synthesis of new proteins; and that apoptosis of stunning is partially related to a disequilibrium between pro- and antiapoptotic proteins as a result of selective inhibition of the 26S-proteasome.
Specific Aim 1 examines the hypothesis that recovery of function of stunned myocardium is partially dependent on 20S- proteasome removal of oxidized actin. Using actin as a model we will determine if oxidation of this protein results in loss of sarcomeric actin and disruption of the myocyte cytoskeleton; and if recovery of function is partially dependent on removal by the 20S-proteasome and de novo synthesis to reestablish proper sarcomeric and cytoskeletal integrity.
Specific Aim 2 examines the hypothesis that the 26S- proteasome is selectively inhibited by oxidative phenomena and exposure to lipofuscin-like materials during ischemia.
Specific Aim 3 examines the hypothesis that inhibition of the 26S-proteosome during stunning leads to disequilibrium between pro- and anti-apoptotic factors. We will examine the role of the 26S-proteasome in dysregulation of the cell cycle protein p53, the cyclin-dependent kinase inhibitor, p27kip1, and the cell death mediator, Bax. These experiments will be performed in a variety of models including an isolated heart preparation, an in vivo model of postischemic recovery, and in cultured neonatal cardiomyocytes. These studies have the potential to explain why some cardiomyocytes are lost and others survive and recover function and ultimately could aid identification of sites for therapeutic intervention. ? ?
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