Ischemic heart disease (IHD) is a leading cause of death worldwide. Current treatments help with managing symptoms of the disease. However, the damaged myocardial tissue remains unfunctional, leading to progression of the disease to heart failure. Therefore, minimizing infarct size and salvaging the infarcted myocardium is crucial for preventing heart failure in IHD patients. Extracellular vesicles (EVs) derived from various stem cell sources induce cardioprotective effects during ischemia-reperfusion injury (IRI). These have been attributed mainly to the anti-apoptotic, pro-angiogenic, miRNA cargo within the stem cell derived EVs. However, endothelial cells, which are naturally abundant in the heart and are sensitive to oxygen deprivation, may hold therapeutic potential toward ischemic myocardial injury via EV-mediated signaling. To test this, human endothelial EVs (EEVs) were isolated and characterized. To assess their human-specific cardioprotection in vitro, we developed a human heart-on-a-chip (hHOC) IRI assay using human stem cell-derived, engineered cardiac tissues. EEVs alleviated cardiac cell death as well as the loss in contractile capacity during and after simulated IRI, in an uptake- and dose-dependent manner. Moreover, EEVs increased the respiratory capacity of cardiomyocytes. To further understand the EEV-mediated cardio-protection their effects on cardiomyocytes proteome was investigated. EEVs partially restored protein profiles of the injured myocytes towards healthy profiles, while presenting enrichment of various metabolic processes related to cellular respiration and modulation of the cellular response to stress. EEV protein cargo was characterized and lead cardioactive proteins were identified. Specifically, EEVs contained proteins that were previously associated with cellular metabolism, redox state, and calcium handling, among other processes, corresponding with the modifications EEVs induced to the myocytes’ proteome. These results suggest that vascular EEVs rescue human cardiac tissues exposed to IRI possibly by supplementing injured myocytes with cargo that supports multiple metabolic and salvage pathways, and therefore may serve as a multi-targeted therapy for IRI. Since the cellular manifestations of IRI are not unique to cardiomyocyte, and are generally common to various cell types, EEVs may provide similar protection against IRI in other organ tissues. In the future, deeper understanding of their mechanisms of action will enable engineering of EEV mimetics that are optimized for treating IRI in different clinical scenarios.