Myocardial-derived small extracellular vesicles spontaneously released from living myocardial slices under biomimetic culture conditions regulate contractility and cardiac remodelling

BACKGROUNDSmall extracellular vesicles (sEVs) released in the cardiac microenvironment are reported to regulate cardiac remodelling, partially via microRNA transfer. Harvesting sEVs produced exclusively from the myocardium remains challenging and a solid research platform for sEV cardiovascular testing needs to be established. Organotypic living myocardial slices (LMS) allow to mimic cardiac disease and to record electrophysiological responses to biological and pharmacological stimuli. This study aims at understanding how cardiac sEVs obtained from donor and failing human LMS and rat LMS under physiological or heart failure-mimicking conditions impact myocardial function and remodelling. METHODS & RESULTSHuman LMS were obtained from the left ventricle (LV) of human donor non-failing and end-stage failing hearts and cultured at 2.2 {micro}m sarcomere length (SL). Rat LV LMS from healthy Sprague-Dawley rats were cultured at a preload of 2.2 or 2.4 {micro}m SL, to recapitulate physiological load and overload, respectively. Following 48-hours biomimetic culture, sEVs were isolated from the culture media by size exclusion chromatography and characterized for their size, concentration, and expression of exosome markers. LMS from human failing hearts presented impaired contractility (P<0.05 vs donor-LMS), which was improved by application of donor heart-derived sEVs at 15 and 20% stretch. Whilst rat overloaded sEVs did not alter the force production of physiological LMS, physiological sEVs significantly increased the active force and decreased their passive force. In rat LMS, 1x108 physiological EVs/slice restored the contractility of overloaded slices, reduced apoptosis, fibrosis-related gene expression and promoted angiogenesis. microRNAs analysis showed significant upregulation of miR-23a-3p and miR-378a-3p in rat physiological sEVs. Finally, to test whether sEVs have a direct effect on cardiomyocytes, we applied sEVs on cultured induced pluripotent stem cell-derived cardiomyocytes (iPSC-CMs). sEVs did not affect the contractility of iPSC-CM monoculture but increased the contractility of iPSC-CM co-cultured with human microvasculature endothelial cells (MVECs). CONCLUSIONSCardiac sEVs isolated from healthy hearts increase the contractility of failing LMS. This effect is associated with, and possibly brought about by, a combination of inhibition of apoptosis, reduction of fibrosis and increased microvascular density, and could involve the transfer of sEV-microRNA into myocardial cells. Our data support the hypothesis that the sEV inotropic action is mediated by endothelial cells.