Novel Mechanisms of Exosome-Mediated Phagocytosis of Dead Cells in Injured Heart

Rationale: After myocardial ischemic injury, improper phagocytic clearance of dying cardiac cells and the ensuing lack of inflammation resolution results in adverse cardiac remodeling and dysfunction that might lead to heart failure. Therefore, therapeutic strategies to ameliorate immune cell phagocytic function are critical for augmenting cardiac repair after injury. Objective: To determine whether mesenchymal stem cell-derived exosomes (MSC-Exos) act as opsonin for apoptotic cells or trigger eat-me phagocytic signaling in resident/recruited phagocytes after myocardial ischemic injury. Methods and Results: We evaluated MSC-Exo-mediated opsonization of apoptotic cardiomyocytes and in vitro and in vivo effects of MFGE8 (milk fat globule epidermal growth factor-factor VIII)-deficient mouse MSC-Exo on macrophage engulfment of apoptotic cardiomyocytes and its implications on cardiac remodeling, repair, and function. Microscopy and FACS analyses show that opsonization of apoptotic cardiomyocytes with MSC-Exo enhances their engulfment by macrophages. Furthermore, preincubation of macrophages with MSC-Exo reprogrammed the signaling pathways involved in phagocytosis and expression of proreparative cytokines. Protein analysis of MSC-Exo reveals expression of MFGE8-a glycoprotein that bridges externalized phosphatidylserine on the apoptotic cell surface to alpha V beta 3 or alpha V beta 5 integrins on the phagocyte. Most intriguingly, siRNA inhibition of MFGE8 significantly reduced the MSC-Exo-mediated augmentation of dead cell engulfment, associated signaling, and proreparative phenotype. After myocardial ischemic injury, intramyocardial administration of MSC-Exo increases macrophage uptake of apoptotic bodies in the border zone of infarct and is associated with reduced proinflammatory response, increase in neovascularization, lower infarct size, and an improvement in cardiac function and MFGE8-deficient MSC-Exo administration failed to protect mice against myocardial infarction. Conclusions: Our data demonstrate that exosome-associated MFGE8 on one hand enhances opsonization of dead cells and on the other activates phagocytic signaling, thus augmenting removal of apoptotic cells, resolution of inflammation, and, therefore, efficient cardiac recovery after injury.