CAS Key Laboratory of Tissue Microenvironment and Tumor, Laboratory of Molecular Cardiology, Shanghai Institute of Nutrition and Health, University of Chinese Academy of Sciences (CAS), CAS, Shanghai, 200031, P. R. China.
Human Genetics, Genome Institute of Singapore, Singapore, 138672, Singapore.
Cell Death Dis. 2020 May 11;11(5):354. doi: 10.1038/s41419-020-2508-y.
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.
人类多能干细胞（hPSCs）衍生的心血管祖细胞（CVPCs）是心肌修复的有前途的来源，但其机制在很大程度上尚不清楚。细胞外囊泡（EVs）已知介导细胞间通讯，然而，在心肌梗死（MI）的急性期给予 hPSC-CVPC 分泌的 EVs（hCVPC-EVs）时，其在梗死愈合中的功效和机制尚不清楚。在这里，我们报告了在梗死心脏中 hESC-CVPC 下常氧（EV-N）和低氧（EV-H）条件下分泌的 EV 的心脏保护作用，以及长链非编码 RNA（lncRNA）相关机制。通过电子显微镜、纳米颗粒跟踪和免疫印迹分析证实了 hCVPC-EVs 的存在。将 hCVPC-EVs 注射到急性心肌梗死的小鼠心肌中，可显著改善心脏功能并减少梗死 28 天后的纤维化，同时伴有边缘区血管生成和心肌细胞存活的改善。一致地，hCVPC-EVs 增强了人脐静脉内皮细胞（HUVECs）的管形成和迁移，提高了细胞活力，并减轻了缺氧葡萄糖剥夺（OGD）损伤的新生大鼠心肌细胞（NRCMs）的乳酸脱氢酶释放。此外，EV-H 在改善心肌细胞存活和 HUVEC 管形成方面的作用明显优于 EV-N。RNA-seq 分析显示，EV-H 中 MALAT1 的含量很高。在梗死心肌和用 hCVPC-EVs 处理的心肌细胞中，其丰度上调。过表达人 MALAT1 可改善 NRCM 与 OGD 损伤的细胞活力，而 MALAT1 敲低则抑制了 hCVPC-EV 促进的 HUVEC 管形成。此外，荧光素酶活性测定、RNA 下拉和 miR-497 水平的操作表明，MALAT1 通过靶向 miR-497 改善了 NRCM 的存活和 HUVEC 管形成。这些结果表明，hCVPC-EVs 通过改善心肌细胞存活和血管生成来促进梗死愈合。hCVPCs 的低氧条件处理可增强 hCVPC-EVs 的心脏保护作用，而 MALAT1 通过靶向 miRNA 部分贡献。
