Abstract After a heart attack, restoring blood flow to the heart (i.e., reperfusion) is critically necessary to limit the amount of myocardial tissue that is damaged by ischemic injury; however, reperfusion itself leads to its own type of myocardial damage (ischemia-reperfusion [IR] injury) for which there is no treatment and that can lead to chronic heart failure (HF). Thus, knowledge of the molecular mechanisms that are induced by cardiac reperfusion is urgently needed to identify novel strategies for preventing IR injury. Cardiac edema, which is mainly caused by an increase in cardiac vascular permeability, is one of the primary contributors to IR injury, and the results from my preliminary studies indicate that vascular permeability is limited by the expression of Aryl Hydrocarbon Receptor Nuclear Translocator (ARNT), also known as hypoxia-inducible factor 1β, in endothelial cells (ECs). I have recently generated a line of mice carrying an inducible, EC-specific ARNT knockout mutation (ecARNT–/–), and the results from experiments with these mice indicate that the loss of ecARNT expression during reperfusion leads to cardiac hemorrhage, and that when the ecARNT–/– mutation is induced in adult animals, cardiac edema develops and gradually progresses to HF. Thus, ecARNT appears to be a crucial regulator of endothelial barrier function, but the role of ecARNT in heart disease is not currently being investigated. The experiments described in this proposal address this unmet need by studying the mechanisms of ARNT-regulated endothelial barrier function after IR injury and their impact on the progression of HF. Furthermore, our preliminary studies suggest that ARNT also regulates the expression of matrix metalloproteinase 3 (MMP3), which cleaves proteins that form junctions between adjacent ECs; thus, we will conduct in-vitro experiments with cultured ECs to determine whether MMP3 inhibition can reverse the impaired endothelial barrier function associated with ecARNT deletion and if so, elucidate the mechanisms that support this observation. We will also use the MMP3 inhibitors in ecARNT–/– mice and generate a line of ecARNT/MMP3 double-knockout mice to determine whether MMP3 inhibition can restore vascular integrity and limit the progression of IR-induced HF. Upon completion, we expect the results from our studies to have identified new therapeutic targets and strategies for improving patient outcomes by reducing cardiac edema during the early stages of recovery from a heart attack and preventing (or delaying) the progression of heart disease.