Abstract
Skeletal muscle xenograft models provide valuable platforms for studying human muscle regeneration and evaluating cell-based therapeutic strategies. However, variability in xenograft outcomes remains a major challenge, limiting the reproducibility and predictive value of human muscle regeneration models. Systematic optimization of parameters is essential to establish robust and reliable platforms. In this study, we optimized key parameters influencing human skeletal muscle xenograft regeneration using human iPSC-derived skeletal myogenic progenitors generated through teratoma differentiation. We first evaluated the effects of extracellular matrix (ECM) substrates on the in vitro culture of these myogenic progenitors, finding that ECM influences their myogenic potential. We next systematically optimized host preconditioning parameters by evaluating irradiation dose and the timing of cardiotoxin-induced muscle injury prior to transplantation. Then, using an optimized xenograft preconditioning platform, we evaluated ECM-assisted cell delivery strategies, finding that ECM co-delivery can improve donor cell retention, engraftment, and donor-derived muscle regeneration compared with cell transplantation alone. Together, these findings demonstrate that optimization of both host preconditioning and cell delivery conditions is critical for establishing reproducible human skeletal muscle xenograft models. This optimized platform enables improved evaluation of human myogenic progenitor regenerative potential and provides a framework for advancing cell-based approaches for skeletal muscle repair.