Background: Myocardial infarction (MI) is a severe disease that often associated with dysfunction of angiogenesis. Cell-based therapies for MI using mesenchymal stem cell (MSC)-derived exosomes have been well studied due to their strong proangiogenic effect. Genetic modification is one of the most common methods to enhance exosome therapy. This study investigated the proangiogenic and cardioprotective effect of exosomes derived from hypoxia-inducible factor 1-alpha (HIF-1α)-modified MSCs. Methods: Lentivirus containing HIF-1α overexpressing vector was packaged and used to infect MSCs. Exosomes were isolated from MSC-conditioned medium by ultracentrifugation. Human umbilical vein endothelial cells (HUVECs) were treated under hypoxia condition for 48 h co-cultured with PBS, control exosomes, or HIF-1α-overexpressed exosomes, respectively. Then the preconditioned HUVECs were subjected to tube formation assay, Transwell assay, and EdU assay to evaluate the protective effect of exosomes. Meanwhile, mRNA and secretion levels of proangiogenic factors were measured by RT-qPCR and ELISA assays. In vivo assays were conducted using the rat myocardial infarction model. PBS, control exosomes, or HIF-1α-overexpressed exosomes were injected through tail vein after MI surgery. Heart function was assessed by echocardiography at days 3, 14, and 28. At day 7, mRNA and protein expression levels of proangiogenic factors in the peri-infarction area and circulation were evaluated, respectively. At day 28, hearts were collected and subjected to H&E staining, Masson’s trichrome staining, and immunofluorescent staining. Results: HIF-1α-overexpressed exosomes rescued the impaired angiogenic ability, migratory function, and proliferation of hypoxia-injured HUVECs. Simultaneously, HIF-1α-overexpressed exosomes preserved heart function by promoting neovessel formation and inhibiting fibrosis in the rat MI model. In addition, both in vitro and in vivo proangiogenic factors mRNA and protein expression levels were elevated after HIF-1α-overexpressed exosome application. Conclusion: HIF-1α-overexpressed exosomes could rescue the impaired angiogenic ability, migration, and proliferation of hypoxia-pretreated HUVECs in vitro and mediate cardioprotection by upregulating proangiogenic factors and enhancing neovessel formation.

Mesenchymal stem cells (MSCs) repair infarcted heart through paracrine mechanism. We sought to compare the effectiveness of MSCs and MSC-derived exosomes (MSC-Exo) in repairing infarcted hearts and to identify how MSC-Exo mediated cardiac repair is regulated. In a rat myocardial infarction model, we found that MSC-Exo inhibited cardiac fibrosis, inflammation, and improved cardiac function. The beneficial effects of MSC-Exo were significantly superior compared to that of MSCs. To explore the potential mechanisms underlying MSC-Exo’s effects, we performed several in vitro experiments and miRNA-sequence analysis. MSC-Exo stimulated cardiomyocyte H9C2 cell proliferation, inhibited apoptosis induced by H₂O₂, and inhibited TGF-β induced transformation of fibroblast cell into myofibroblast. Importantly, novel miRNA sequencing results indicated that MSC-Exo and MSCs have similar miRNA expression profile, which could be one of the reasons that MSC-Exo can replace MSCs for cardiac repair. In addition, the expression of several miRNAs from MSC-Exo was significantly different from that of MSCs, which may explain why MSC-Exo has better therapeutic effect than MSCs. In conclusion, this study demonstrates that MSC-Exo could be used alone to promote cardiac repair and are superior to MSCs in repairing injured myocardium.

Background. To cure ischemic diseases, angiogenesis needs to be improved by various strategies in ischemic area. Considering that microRNA-132 (miR-132) regulates endothelial cell behavior during angiogenesis and the safe and efficacious delivery of microRNAs in vivo is rarely achieved, an ideal vehicle for miR-132 delivery could bring the promise for ischemic diseases. As a natural carrier of biological molecules, exosomes are more and more developed as an ideal vehicle for miRNA transfer. Meanwhile, mesenchymal stem cells could release large amounts of exosomes. Thus, this study aimed to investigate whether MSC-derived exosomes can be used for miR-132 delivery in the treatment of myocardial ischemia. Methods. MSC-derived exosomes were electroporated with miR-132 mimics and inhibitors. After electroporation, miR-132 exosomes were labelled with DiI and added to HUVECs. Internalization of DiI-labelled exosomes was examined by fluorescent microscopy. Expression levels of miR-132 in exosomes and HUVECs were quantified by real-time PCR. The mRNA levels of miR-132 target gene RASA1 in HUVECs were quantified by real-time PCR. Luciferase reporter assay was performed to examine the targeting relationship between miR-132 and RASA1. The effects of miR-132 exosomes on the angiogenic ability of endothelial cells were evaluated by tube formation assay. Matrigel plug assay and myocardial infarction model were used to determine whether miR-132 exosomes can promote angiogenesis in vivo. Results. miR-132 mimics were effectively electroporated and highly detected in MSC-derived exosomes. The expression level of miR-132 was high in HUVECs preincubated with miR-132 mimic-electroporated exosomes and low in HUVECs preincubated with miR-132 inhibitor-electroporated exosomes. The expression level of RASA1, miR-132 target gene, was reversely correlated with miR-132 expression in HUVECs pretreated with exosomes. Luciferase reporter assay further confirmed that RASA1 was a direct target of miR-132. Exosomes loaded with miR-132, as a vehicle for miRNA transfer, significantly increased tube formation of endothelial cells. Moreover, subcutaneous injection of HUVECs pretreated with miR-132 exosomes in nude mice significantly increased their angiogenesis capacity in vivo. In addition, transplantation of miR-132 exosomes in the ischemic hearts of mice markedly enhanced the neovascularization in the peri-infarct zone and preserved heart functions. Conclusions. The findings suggest that the export of miR-132 via MSC-derived exosomes represents a novel strategy to enhance angiogenesis in ischemic diseases.

Inflammation plays crucial roles in the regulation of pathophysiological processes involved in injury, repair and remodeling of the infarcted heart; hence, it has become a promising target to improve the prognosis of myocardial infarction (MI). Mesenchymal stem cells (MSCs) serve as an effective and innovative treatment option for cardiac repair owing to their paracrine effects and immunomodulatory functions. In fact, transplanted MSCs have been shown to accumulate at injury sites of heart, exerting multiple effects including immunomodulation, regulating macrophages polarization, modulating the activation of T cells, NK cells and dendritic cells and alleviating pyroptosis of non-immune cells. Many studies also proved that preconditioning of MSCs can enhance their inflammation-regulatory effects. In this review, we provide an overview on the current understanding of the mechanisms on MSCs and their secretome regulating inflammation and immune cells after myocardial infarction and shed light on the applications of MSCs in the treatment of cardiac infarction.

Generating universal human umbilical mesenchymal stem cells (UMSCs) without immune rejection is desirable for clinical application. Here we developed an innovative strategy using CRISPR/Cas9 to generate B2M^‐UMSCs in which human leucocyte antigen (HLA) light chain β2‐microglobulin (B2M) was deleted. The therapeutic potential of B2M^‐UMSCs was examined in a mouse ischaemic hindlimb model. We show that B2M^‐UMSCs facilitated perfusion recovery and enhanced running capability, without inducing immune rejection. The beneficial effect was mediated by exosomes. Mechanistically, microRNA (miR) sequencing identified miR‐24 as a major component of the exosomes originating from B2M^‐UMSCs. We identified Bim as a potential target of miR‐24 through bioinformatics analysis, which was further confirmed by loss‐of‐function and gain‐of‐function approaches. Taken together, our data revealed that knockout of B2M is a convenient and efficient strategy to prevent UMSCs‐induced immune rejection, and it provides a universal clinical‐scale cell source for tissue repair and regeneration without the need for HLA matching in the future.