Abstract 461: Prospective Isolation of Heart Field Specific Cardiomyocytes from Differentiating Human Embryonic Stem Cells

Abstract

Cardiovascular disease (CVD) is the leading cause of death worldwide. Human embryonic stem cell (hESC) derived cardiovascular progenitor cells (CPCs) or cardiomyocytes (CMs) represent a promising candidate for cell-based therapies to treat CVD. Following a myocardial infarction there is extensive CM death and damage primarily localized to the left ventricle of the heart, which is predominantly derived from first heart field (FHF) CPCs during development. This is important in future developmental and regenerative therapy studies since heart field specific cardiac cells will be desired. Current hESC differentiation protocols as well as the use of cardiac-specific reporter lines, yield a mixture of cardiac cells without the ability to isolate heart field specific cardiac progenitors and their associated progeny. Here we generated and characterized a FHF specific TBX5- TdTomato/W hESC reporter line to show that TBX5 + cells represent an enriched population of FHF CPCs that can give rise to CMs, endothelial and smooth muscle cells in vitro . Transplantation of hESC-derived TBX5 + and TBX5 - CPCs into the injured left ventricle of mice showed that these cells were able to survive and predominantly gave rise to CMs without functional integration into the host myocardium. Furthermore, RNA-sequencing analysis of hESC-derived cardiac cells at different stages of development suggested that while TBX5 expression leads to an enrichment of FHF- derived CMs, TBX5 - cells also generate cardiac cells that resemble second heart field (SHF) CMs at the transcriptional level. To enable prospective isolation of FHF and SHF CMs we generated a double transgenic hESC reporter line TBX5 TdTomato/W /NKX2-5 eGFP/W . Using immunostaining, qPCR and electrophysiological studies we were able to confirm the isolation of FHF and SHF specific CMs from differentiating hESCs. Finally, we show that using the novel surface marker, CORIN, we can isolate FHF CMs without the use of genetic reporters. Our studies provide valuable information and resources on the production of heart field specific CMs from hESCs for cell-based therapies to treat CVD.