The cardio-protective effects of human induced pluripotent stem cell-derived cardiovascular progenitor cells (CPC) are largely mediated by the paracrine release of extracellular vesicles (EV). We aimed to assess the immunological behavior of EV-CPC, which is a prerequisite for their clinical translation.
Flow cytometry demonstrated that EV-CPC expressed very low levels of immune relevant molecules including HLA class I, CD80, CD274 (PD-L1), CD275 (ICOS-L); and moderate levels of ligands of the natural killer (NK) cell activating receptor, NKG2D. In mixed lymphocyte reactions, EV-CPC neither induced nor modulated adaptive allogeneic T cell immune responses. They also failed to induce NK cell degranulation, even at high concentrations. These in vitro effects were confirmed in vivo as repeated injections of EV-CPC did not stimulate production of immunoglobulins or affect the IFN-γ responses from primed splenocytes. In a mouse model of chronic heart failure, intra-myocardial injections of EV-CPC, 3 weeks after myocardial infarction, decreased both the number of cardiac pro-inflammatory Ly6Chigh monocytes and circulating levels of pro-inflammatory cytokines (IL-1α, TNFα and IFN-γ). In a model of acute infarction, direct cardiac injection of EV-CPC 2 days after infarction reduced pro-inflammatory macrophages, Ly6Chigh monocytes, and neutrophils in heart tissue as compared to controls. EV-CPC also reduced levels of pro-inflammatory cytokines IL-1α, IL-2 and IL-6, and increased levels of the anti-inflammatory cytokine IL-10. These effects on human macrophages and monocytes were reproduced in vitro; EV-CPC reduced the number of pro-inflammatory monocytes and M1 macrophages, while increasing the number of anti-inflammatory M2 macrophages.
EV-CPC do not trigger an immune response either in in vitro human allogeneic models or in immunocompetent animal models. The capacity for orienting the response of monocyte/macrophages towards resolution of inflammation strengthens the clinical attractiveness of EV-CPC as an acellular therapy for cardiac repair.
Mounting evidence suggests that the release of extracellular vesicles (EV) conveys the therapeutic efficacy of various progenitor cells in the failing heart. In the present study, we showed that intra-myocardial delivery of EV isolated from human iPSC-derived cardiovascular progenitor cells does not trigger an allogenic immune response and seems rather to induce a systemic anti-inflammatory effect. Our results therefore suggest that human EV administration might not require immunosuppression. It remains to establish whether the systemic delivery of EV would trigger similar anti-inflammatory effects, positively impacting heart function.

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