Extracellular vesicles (EVs) are a heterogeneous group of natural particles with relevance for the treatment of cardiovascular diseases. The endogenous properties of these vesicles allow them to survive in the extracellular space, bypass biological barriers and deliver their biologically active molecular cargo to recipient cells. Moreover, EVs can be engineered to enhance their stability, bioactivity, presentation and capacity for on target binding at both cell type and tissue levels. The therapeutic potential of native (i.e., EVs that were not modified via donor cell or direct modulation) and engineered (i.e. EVs that were modified either pre-or post-isolation or whose pharmacokinetics/presentation was altered using engineering methodologies EVs is still limitedly explored in the context of cardiovascular diseases. Efforts to tap into the therapeutic potential of EVs will require innovative approaches and a comprehensive integration of knowledge gathered from decades of molecular compound delivery. In this review, we outline the endogenous properties of EVs that make them natural delivery agents as well as those features that can be improved using bioengineering approaches. We also discuss the therapeutic applications of native and engineered EVs for cardiovascular applications and examine the opportunities and challenges that need to be addressed to advance this research area with an emphasis on clinical translation. Key points• EVs secreted from stem/progenitor cells as well as differentiated somatic cells have regenerative properties in the context of myocardial infarction, ischemic limb, chronic wounds and stroke.

Small extracellular vesicles (sEVs), through their natural ability to interact with biological membranes and exploit endogenous processing pathways to convey biological information, are quintessential for the delivery of therapeutically relevant compounds, such as microRNAs (miRNAs) and proteins. Here, we used a fluorescently‐labelled miRNA to quantify the efficiency of different methods to modulate the cargo of sEVs. Our results showed that, compared with electroporation, heat shock, permeation by a detergent‐based compound (saponin) or cholesterol‐modification of the miRNA, Exo‐Fect was the most efficient method with > 50% transfection efficiency. Furthermore, qRT‐PCR data showed that, compared with native sEVs, Exo‐Fect modulation led to a > 1000‐fold upregulation of the miRNA of interest. Importantly, this upregulation was observed for sEVs isolated from multiple sources. The modulated sEVs were able to delivery miR‐155‐5p into a reporter cell line, confirming the successful delivery of the miRNA to the target cell and, more importantly, its functionality. Finally, we showed that the membrane of Exo‐Fect‐loaded sEVs was altered compared with native sEVs and that enhanced the internalization of Exo‐Fect‐loaded sEVs within the target cells and decreased the interaction of those modulated sEVs with lysosomes.

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