Project Summary/Abstract Current therapies for patients with heart failure (HF) have limited efficacy because drugs and surgery can only relieve the symptoms of HF but cannot save necrotic cardiomyocytes. Hence, there is an ongoing need to find novel treatment strategies for HF following Myocardial Infraction (MI). Mesenchymal stem cells (MSCs) have shown to improve cardiac function and reverse remodeling after MI where the underlying mechanism is multifactorial and recently studies suggested that much of this repair can be attributed to extracellular vesicles (EVs) that are released by MSCs. Main challenges impeding the translation of EVs into clinical application are establishing advanced characterization method to document reproducibility and large-scale production for clinical grade EVs. We have developed large-scale manufacture process using Quantum Bioreactors and this project will aid in establishing methods to evaluate batch-to-batch variability. Our centralized hypothesis for this project is that EV restore cardiac function by reducing cardiac remodeling through promotion of tissue homeostasis, inhibition of inflammation, and promotion of angiogenesis. We hypothesize that repeated, systemic delivery of WJMSC EVs will have additive impact for improving cardiac function and reducing cardiac remodeling as compared to single dosing and WJMSC EVs provide underlying molecular mechanisms that is beneficial for cardiac repair. Our study objectives are: 1) to evaluate the feasibility and safety of repeated injections of WJMSC EV administered systemically in small animal model; 2) to observe effect on cardiac structure and function in an experimental MI model; 3) to determine an optimal dose response. The long-term goal is to elucidate the molecular mechanisms of WJMSC EV cargo on cardiac function and remodeling and the optimal dose and regimen showing an improvement in cardiac function and reduced remodeling that will be used to investigate the clinical effectiveness of our established clinical dose of WJMSC EV product in a large animal (porcine) MI model. We are expecting that at least high dose of EV with repeated systemic administration will show greatest improvement in cardiac function and seeing similar outcome with the medium and lower dose of EV. This would coincide with absorption of EV into the heart tissue observed at 24 hours after administration and accompany of finding key molecular mechanism of improving cardiac repair from RNA bioinformatic analysis. In addition, we will be able to establish characterization criteria to reduce donor-to-donor variability. The positive outcome of this study will establish the premise for moving toward large animal MI model and translating the technology and innovation for clinical application.