Abstract 15544: Manipulating Oxygen-Releasing Levels and Oxidative Stress to Enhance Viability and Functionality of Human Pluripotent Stem Cell-Derived Cardiomyocytes for Myocardial Infarction Therapy

Abstract

Introduction: Human pluripotent stem cell-derived cardiomyocytes (hPSC-CMs) hold great promise for myocardial infarction (MI) treatment. Although O 2 -releasing engineering approaches have been developed to enhance stem cell engraftment, a significant gap persists in knowledge regarding the oxidative stress controlling viability and functionality of hPSC-CMs. Hypothesis: Preconditioning hPSC-CMs with appropriate oxidative levels will enhance their adaptation and engraftment in the infarcted heart. Aims: To develop a novel system that produces a controlled release of O 2 to supply engineered heart tissue for regenerative therapies. Methods: An O 2 -releasing hydrogel system (ORHS) was engineered using peroxides (sodium percarbonate) and antioxidants (β-carotene) encapsulated in PLGA-based particles. The effect of ORHS on hPSC-CMs was evaluated through oxidative stress assays, cell viability assessments, and contractility measurements. RNA-seq profiling was performed to analyze the gene expression of hPSC-CMs in response to ORHS. After implantation of ORHS-hPSC-CMs in infarcted hearts of immunodeficient mice, heart functions were monitored with echocardiography. Implanted cell engraftment was assessed using in vivo imaging systems and immunostaining. Results: The dissolved O 2 levels were increased in ORHS for 7 days, while the oxidative stress was minimized by β-carotene. The optimal ORHS significantly enhanced the viability and mitigated cell death of hPSC-CMs under hypoxic stress conditions. ORHS also enhanced cell contractility and improved calcium signaling in hPSC-CMs. Genes related to maturation and functionality (such as TNNT2, TNNI3, GJA1, and RYR2) were upregulated by ORHS in hPSC-CMs. Implantation of ORHS-hPSC-CMs for 4 weeks resulted in improved ejection fraction, reduced ventricular remodeling, alleviated infarct size, and scar formation. Furthermore, the engraftment and survival of hPSC-CMs were significantly increased by ORHS compared to control hydrogels, as demonstrated by luciferase reporter assays and troponin immunostaining with TUNEL. Conclusion: The development of ORHS offers a promising approach for inducing oxidative preconditioning and achieving enhanced regenerative outcomes of hPSC-CMs.