Unlocking the Molecular Cues for Cardiomyocyte Proliferation
Authors:   , Yuan Qianliang 
Published: March 2024

Human induced pluripotent stem cells (hiPSCs) have demonstrated potential for molecular studies and the generation of human heart models, thereby reducing reliance on animal studies. In contrast, the adult heart, lacking the ability to self-repair after a heart attack, hiPSC-derived cardiomyocytes (hiPSC-CMs) present a promising and potentially limitless source of human embryonic-like CMs. To fully harness this potential, it is essential to delve into the developmental molecular cues governing CM proliferation during embryonic development. Moreover, to achieve the objective of precision medicine, understanding the relevant molecular process on CM maturation during cardiac differentiation becomes vital for improving the practical applications of hiPSC-CMs in drug screening and disease remodeling. In this thesis, we initially utilized hiPSC-CMs to investigate the signaling cues that guide cell fate decisions between proliferation and terminal differentiation. A combinatory screen for CHIR99021/Wnt and Insulin/Akt pathways was conducted on spontaneously beating hiPSC-CMs, revealing the synergistic effects of CHIR99021/Wnt and Insulin/Akt pathways on proliferation and the regulation of sarcomere homeostasis. Moreover, we identified a novel temporal interplay between CHIR99021/Wnt via TCF/LEF and Insulin/Akt via FOXO as regulators of cell-fate decisions in immature hiPSC-CMs. The study contributed to the understanding of the effects of targeting embryonic growth pathways for acute and chronic heart failure treatment. Additionally, we explored sarcomere architecture in high proliferative hiPSC-CMs, we observed sarcomere disassembly during mitosis in proliferating hiPSC-CMs treated with CHIR99021, with an emphasis on Wnt/β-catenin signaling. The presence of these signals is higher in highly proliferative hiPSC-CMs compared to non-proliferative hiPSC-CMs. Furthermore, the study demonstrated that CHIR99021 enhances non-viral vector incorporation in highly proliferative hiPSC-CMs, providing a tool for gene manipulation studies. We further continue to investigate the effect of cell-cell contact on the canonical Wnt/β-catenin pathway in response to hiPSC-CM proliferation. Our results indicate that cell-cell contact negatively influences the canonical Wnt/β-catenin signaling pathway, inhibiting proliferation in immature hiPSC-CMs. The involvement of N-cadherin in cell-contact inhibition was found, resulting in the release of more proliferative hiPSC-CMs due to disrupted N-cadherin. Furthermore, reactivation of Wnt/β-catenin signaling in late-stage hiPSC-CMs led to impaired sarcomere function, providing insights into the role of embryonic signaling pathways in failing/injured heart conditions. We finally explored the impact of specific TNNT2 mutations associated with hypertrophic cardiomyopathy and dilated cardiomyopathy on cardiac function using hiPSC-CMs. We assessed contractile parameters and Ca2+ transients, mitochondrial function, and molecular changes under baseline and isoproterenol stimulation conditions using a well-established High-Throughput Analysis System. These findings offer insights into mutation-specific responses and disease pathways relevant to cardiomyopathies.