PROJECT SUMMARY Heart failure (HF) is a leading cause of death in the United States and is often caused by the loss of cardiomyocyte (CM) populations in the heart. Adult mammalian CMs have very limited proliferative potential, and after myocardial infarction (MI) and the death of approximately one billion CMs, damaged cardiac tissue is replaced with fibrotic scar rather than with functioning myocardium. Significant efforts have been made to understand the proliferative capacity of CMs in mouse and zebrafish models of endogenous cardiac regeneration, but these efforts have not led to successful therapies. In 2018, it was shown that the neonatal swine heart has an endogenous capacity to robustly regenerate after cardiac injury, providing a model organism whose heart is anatomically and physiologically highly similar to the human heart; therefore, investigating endogenous cardiac regeneration in this model organism has strong translational potential. As such, the goal of this proposal is to identify novel therapeutic targets for cardiac regeneration by characterizing the molecular landscape of the neonatal swine heart throughout postnatal development, endogenous regeneration, and pathological remodeling using a novel multiomics analysis of the sarcomere, global cardiac proteome, and global cardiac metabolome. Aim 1 uses top-down proteomics, the premier technology to characterize proteoforms – all the protein products that arise from a single gene as a result of genetic variations, alternative splicing, and post- translational modifications – to extensively characterize the molecular composition of sarcomeres that can disassemble, a process that occurs during CM proliferation. These changes in sarcomere proteoform abundances will be correlated to developing, regenerating, and pathologically remodeling hearts to understand the sarcomere composition during various biological states. Aim 2 seeks to characterize how the global proteome, phosphoproteome, and metabolome are altered throughout postnatal swine heart development, regeneration, and pathological remodeling to delineate molecular mechanisms that support the regenerative capacity of neonatal swine hearts. These large -omics data sets will be integrated and bioinformatically analyzed to holistically identify the molecular mechanisms that support myocardial regeneration in swine. The success of this proposed work will elucidate novel targets for developing therapeutic strategies to promote cardiac regeneration in the injured human heart.

PROJECT NARRATIVE While there is a high global prevalence of heart failure (HF), a cardiovascular disease that can arise from a variety of cardiac insults, there are no curative therapies. This proposal aims to characterize endogenous cardiac regeneration in swine, a model system much more closely related to humans than previous mouse and zebrafish models, using a novel multiomics approach integrating both proteomics and metabolomics. The success of the proposed research will offer new insights into the molecular mechanisms supporting endogenous cardiac regeneration, enabling the design of targeted therapeutics to treat and prevent heart failure.