PROJECT SUMMARY/ABSTRACT Lack of oxygenation and nutrients that result from narrowed arteries and reduced blood flow debilitates the normal function of the heart, leading to severe morbidity and early mortality worldwide. This devastating acute medical episode is often seen in patients with obstructive plaques in their arteries. As a result, coronary artery disease is the root cause of ischemic cardiovascular diseases such as myocardial infarction. Despite that a healthy lifestyle and standard care may deter development of these diseases, risk factors such as obesity, dyslipidemia, diabetes, and high blood pressure elevate the odds of recurrent myocardial infarction, leading to heart failure, ischemic cardiomyopathy, or even death as well as a major financial burden to the society. Reperfusion after acute myocardial infarction introduced by coronary angioplasty and direct stenting can cause further damage to the injured heart. Unfortunately, the scarcity of novel molecular targets that stimulate natural new blood vessel growth to restore blood flow and augment heart repair and regeneration creates a daunting fight against this deleterious condition, for which there is currently no cure. The long-term goal of our research is to discern molecular insight into endothelial regulation and heart regeneration in health and disease. In this application, we aim to decipher molecular mechanisms behind the action of cardiac endothelial epsins in regulating in myocardial ischemia. Epsins are a family of prominent endocytic adaptor proteins that were originally discovered by the PI. We recently reported that targeting epsins in macrophages in atherosclerotic plaques by nanoparticle-mediated delivery of epsin siRNAs prevents atheroma progression and promotes atheroma regression, suggesting epsins as an emerging target for treating coronary artery disease in preclinical models. Harnessing newly developed snRNA sequencing technologies, we made a potentially groundbreaking discovery where the loss of cardiac endothelial epsins potentiates endothelial cell expansion and precursor cell transition to fuel vascularization in neonatal hearts. We see heightened neovascularization, bolstered cardiomyocyte proliferation in neonatal hearts of inducible endothelial-specific epsin 1 and 2 double knockout (EC-iDKO) mice. Consistently, elevated revascularization, enhanced cardiac function and improved heart repair and recovery from myocardial infarction in adult EC-iDKO mice. We posit that inhibition of cardiac endothelial epsins fortifies angiocrine signals such as CXCL12-CXCR4 signaling, enable creation of a vast endothelial cell reservoir to build vascular networks in the heart. Our central hypotheses are that epsins in the cardiac endothelium suppress endothelial expansion, in part, by repressing CXCR4 signaling and hindering heart regeneration and repair following myocardial ischemia. If successful, our proposed work will explore uncharted territory, uncover original reg...