Dilated cardiomyopathy (DCM) is a leading cause of heart failure around the world. Inherited mutations cause the left ventricle of the heart to enlarge, thinning the heart muscle wall and decreasing the overall function of the heart. In my research project, I will determine if disrupting fibroblast function by knocking out a key developmental signaling factor, p38, can improve, or even reverse, DCM disease characteristics. Specific Aim 1 will be to determine the effects of p38 knockout-induced fibroblast dysfunction on cardiomyocyte function and structural remodeling in late-stage DCM. The rationale is that myocytes in DCM have poor contraction and structurally remodel to longer, thinner morphologies, which occurs in our DCM mouse model around 4 months of age. I expect to see less of these characteristics with the p38 knockout. Specific Aim 2 will assess cardiac fibroblast proliferation and fibrosis in response to disabling cardiac fibroblast function late into the DCM disease process. The rationale is that studying and observing the dynamics of the fibroblast population is critical when understanding the effects of fibroblasts and the p38 knockout model on DCM. In previous studies, the Davis lab identified that cardiac fibroblasts maladaptively respond to inherited DCM mutations in cardiac myocytes, worsening the whole heart. I expect to see less fibroblast proliferation in the p38 model. P38 is essential for fibroblast signaling pathways and functionality, so by knocking it out I will be able to test if fibroblasts are a viable therapeutic target for patients with DCM.

Hypertrophic cardiomyopathy (HCM) is the most common genetic cardiovascular disease. Traditional therapies focus on treating the symptoms of the disease and do not directly treat the underlying functional changes. Myosin modulators are a novel class of pharmaceutical agents designed to treat patients with cardiomyopathies by directly modulating cardiac myosin function in the sarcomere. Compounds including Mavacamten (Mava) and Aficamten (Afi) reduce myosin function, measured through its ATPase activity. In this study, I investigate how these small molecules affect the multiple turnover kinetics of the ATPase cycle. Porcine cardiac heavy meromyosin (pcHMM) is rapidly mixed with a two-times excess of fluorescently labeled ATP (mant.ATP) in the presence of actin and fluorescence is measured over time. Calculating the length of time, tau (τ), until the fluorescence has returned to 50% of the peak value estimates the time taken to hydrolyze all the mant.ATP, and is used to calculate the rate constant of ATP hydrolysis (kcat). Both Afi and Mava increase τ and decrease kcat, while a lower concentration of Mava is required to reach similar inhibition as Afi. I also utilize the In Vitro Motility assay to measure the ability of myosin to move actin and compare the effects of Mava and Afi with stopped-flow data. Preliminary results indicate that both Mava and Afi inhibit actin filament velocity, with Mava requiring a lower concentration, similar to stopped-flow. I will discuss how each modulator affects ATP turnover with a direct effect on catalytic activity and extend those results to the functional consequences of these myosin inhibitors. Insight into the impact of these myosin inhibitors on the myosin actin cross-bridge cycle will help provide tailored treatments to patients who are impacted by HCM.