Senescence modulated cardiac repair with pluripotent stem cell-derived cardiomyocytes

Human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) offer great promise for treating heart failure. However, their clinical application remains limited owing to poor cell engraftment, highlighting the need for scalable production methods that preserve therapeutic efficacy. Although in vitro expansion using glycogen synthase kinase 3β (GSK3β) inhibitors can substantially increase hiPSC-CM yield, the therapeutic potential of these expanded cells is yet to be fully characterized. hiPSCs were differentiated into non-expanded cardiomyocytes (N-CMs) via a conventional monolayer protocol or were further replated and cultured with CHIR99021 to generate expanded cardiomyocytes (E-CMs). Both E-CMs and N-CMs were transplanted into a rat myocardial infarction model to evaluate engraftment efficiency. To prevent senescence in E-CMs, cells were treated with the PAPD5 inhibitor BCH001 (E-CM-B) or DMSO as a control (E-CM-D) and then transplanted in the same manner. In vitro, cellular characteristics, including senescence and maturation, were evaluated by adhesion assays, immunocytochemistry, RNA-sequencing (RNA-seq) analysis, and mitochondrial assays. In vivo, cardiac function was assessed by echocardiography, engraftment was monitored via bioluminescent imaging, and the properties of engrafted cells were examined through histological and immunohistochemical analyses. Although GSK3β inhibitor–mediated expansion increased hiPSC-CM yield, it induced telomere shortening, elevated DNA damage, and upregulation of senescence-associated markers, ultimately compromising engraftment and functional recovery after transplantation. Treatment with BCH001 during expansion attenuated senescence markers and promoted mitochondrial maturation. Transplantation of these pharmacologically rescued hiPSC-CMs significantly improved outcomes in infarcted hearts, including increased graft size, enhanced left ventricular function, reduced fibrosis, and promoted in vivo maturation. This study identifies cellular senescence as a major barrier to the therapeutic efficacy of expanded hiPSC-CMs. Transient pharmacological intervention with a PAPD5 inhibitor during in vitro expansion effectively restores hiPSC-CMs, preserving their regenerative potential. These findings establish a scalable, clinically relevant strategy to overcome current limitations in hiPSC-CM therapy, paving the way for safer and more effective cell-based treatments for heart failure.