Extracellular vesicles from human embryonic stem cell-derived cardiovascular progenitor cells promote cardiac infarct healing through reducing cardiomyocyte death and promoting angiogenesis

Qiang Wu,Shenyan Liu,Jiliang Tan,Jinxi Wang,Wilson Lek Wen Tan,Zenia Tiang,Peng Zhang,Zhongyan Chen,Xiujian Yu,Huang-Tian Yang,Yun Jiang,Roger Sik-Yin Foo,Shihui Wang,Qiang Li

doi:10.1038/s41419-020-2508-y

Abstract

Human pluripotent stem cells (hPSCs)-derived cardiovascular progenitor cells (CVPCs) are a promising source for myocardial repair, while the mechanisms remain largely unknown. Extracellular vesicles (EVs) are known to mediate cell–cell communication, however, the efficacy and mechanisms of hPSC-CVPC-secreted EVs (hCVPC-EVs) in the infarct healing when given at the acute phase of myocardial infarction (MI) are unknown. Here, we report the cardioprotective effects of the EVs secreted from hESC-CVPCs under normoxic (EV-N) and hypoxic (EV-H) conditions in the infarcted heart and the long noncoding RNA (lncRNA)-related mechanisms. The hCVPC-EVs were confirmed by electron microscopy, nanoparticle tracking, and immunoblotting analysis. Injection of hCVPC-EVs into acutely infracted murine myocardium significantly improved cardiac function and reduced fibrosis at day 28 post MI, accompanied with the improved vascularization and cardiomyocyte survival at border zones. Consistently, hCVPC-EVs enhanced the tube formation and migration of human umbilical vein endothelial cells (HUVECs), improved the cell viability, and attenuated the lactate dehydrogenase release of neonatal rat cardiomyocytes (NRCMs) with oxygen glucose deprivation (OGD) injury. Moreover, the improvement of the EV-H in cardiomyocyte survival and tube formation of HUVECs was significantly better than these in the EV-N. RNA-seq analysis revealed a high abundance of the lncRNA MALAT1 in the EV-H. Its abundance was upregulated in the infarcted myocardium and cardiomyocytes treated with hCVPC-EVs. Overexpression of human MALAT1 improved the cell viability of NRCM with OGD injury, while knockdown of MALAT1 inhibited the hCVPC-EV-promoted tube formation of HUVECs. Furthermore, luciferase activity assay, RNA pull-down, and manipulation of miR-497 levels showed that MALAT1 improved NRCMs survival and HUVEC tube formation through targeting miR-497. These results reveal that hCVPC-EVs promote the infarct healing through improvement of cardiomyocyte survival and angiogenesis. The cardioprotective effects of hCVPC-EVs can be enhanced by hypoxia-conditioning of hCVPCs and are partially contributed by MALAT1 via targeting the miRNA.

Highlights

Myocardial infarction (MI), characterized by massive cardiomyocyte death followed by cardiac dysfunction and myocardial fibrosis, is a leading cause of death worldwide[1,2,3]
Human pluripotent stem cells, including both human embryonic stem cells and human induced pluripotent stem cells, hold promise for promoting cardiac infarct healing because they can be theoretically produced in unlimited quantities of cells of any lineages, including cardiomyocytes17,19–24. hESC-derived cardiovascular progenitor cells have been shown to improve cardiac function of rodent infarcted hearts when implanted during the subacute stage of ischemia/reperfusion (I/R) hearts[14]
To determine whether the inhibition of cardiomyocyte death is one of mechanisms underlying the cardioprotective effects of hESC-derived cardiovascular progenitor cells (hCVPCs)-Extracellular vesicles (EVs), we examined the number of transferasemediated dUTP nick end labeling (TUNEL)+-cardiomyocytes in the infarcted hearts

Summary

Introduction

Myocardial infarction (MI), characterized by massive cardiomyocyte death followed by cardiac dysfunction and myocardial fibrosis, is a leading cause of death worldwide[1,2,3]. Developing new therapeutic strategies to promote the infarct healing by reducing cardiomyocyte death following acute MI (AMI) and improving the cardiac performance are desirable[11]. Human pluripotent stem cells (hPSCs), including both human embryonic stem cells (hESCs) and human induced pluripotent stem cells (hiPSCs), hold promise for promoting cardiac infarct healing because they can be theoretically produced in unlimited quantities of cells of any lineages, including cardiomyocytes. HESC-derived cardiovascular progenitor cells (hCVPCs) have been shown to improve cardiac function of rodent infarcted hearts when implanted during the subacute stage of ischemia/reperfusion (I/R) hearts[14]. We found that SSEA1+ hCVPCs promote cardiac infarct healing when given during the early phase of MI in mouse[25] and nonhuman primate[26] models, showing the significant improvement in the recovery of left ventricular (LV) function and the amelioration of cardiomyocyte death. We found that the beneficial effects of these cells to the infarcted hearts are related to the hCVPC-

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