Senior Research Fellow Lee Kong Chian School of Medicine, Singapore
Abstract: Ischemic heart disease remains the leading cause of mortality worldwide, highlighting the urgent need for regenerative therapies. Human pluripotent stem cell-derived cardiomyocytes (hPSC-CMs) offer a promising approach. Here, we used a chemically defined, xeno-free laminin-221-based differentiation protocol to generate committed cardiac progenitors (CCPs) and investigated their maturation at later stages. Single-cell RNA sequencing of hPSC-CMs at days 21 and 42 revealed a progressive shift towards a cardiomyocyte-dominant population compared to fibroblast, myofibroblasts, epithelial, and endothelial cells. Gene expression analysis highlighted the upregulation of key cardiac maturation markers, including those involved in sarcomere organization, metabolism, and ion handling, consistent with known hallmarks of cardiac maturation. While day 21 cells exhibited features of immature cardiomyocytes. Day 42 cells showed enhanced RYR2, CACNB2, and MYL2 expression, indicating partial maturation. Functional analysis further highlighted a transition from RNA processing pathways at day 21 to cardiac-specific pathways at day 42. Furthermore, transcriptional regulatory analysis identified ESRR and TEAD isoforms as potential key regulators driving cardiomyocyte maturation. This study provides a single-cell transcriptomic profile of the critical transition phase during cardiomyocyte differentiation, offering insights into molecular mechanisms that can be leveraged to enhance cardiomyocyte maturation for regenerative applications.
Funding Source: This work has been supported by grants from the MOE, and NRF of Singapore (MOE-RS06/23, CRP24-2020-0083, RS06/23 and MOE start-up grant, 022976-00001)
