Driving efficient and pure skeletal muscle cell differentiation from pluripotent stem cells (PSCs) has been challenging. Here, we report an optimized protocol that generates skeletal muscle progenitor cells with high efficiency and purity in a short period of time. Human induced PSCs (hiPSCs) and murine embryonic stem cells (mESCs) were specified into the mesodermal myogenic fate using distinct and species-specific protocols. We used a specific maturation medium to promote the terminal differentiation of both human and mouse myoblast populations, and generated myotubes associated with a large pool of cell-cycle arrested PAX7⁺ cells. We also show that myotube maturation is modulated by dish-coating properties, cell density, and percentage of myogenic progenitor cells. Given the high efficiency in the generation of myogenic progenitors and differentiated myofibers, this protocol provides an attractive strategy for tissue engineering, modeling of muscle dystrophies, and evaluation of new therapeutic approaches in vitro.

驱动多能干细胞 (PSC) 高效、纯的骨骼肌细胞分化一直具有挑战性。在这里，我们报告了一种优化的方案，可以在短时间内以高效率和纯度生成骨骼肌祖细胞。使用不同的物种特异性方案将人类诱导的 PSC (hiPSC) 和鼠胚胎干细胞 (mESC) 指定为中胚层肌源性命运。我们使用特定的成熟培养基来促进人和小鼠成肌细胞群的终末分化，并生成与大量细胞周期停滞的 PAX7 ⁺细胞相关的肌管。我们还表明，肌管成熟受到培养皿涂层特性、细胞密度和肌源性祖细胞百分比的调节。鉴于肌源性祖细胞和分化肌纤维生成的高效率，该协议为组织工程、肌肉营养不良建模和体外新治疗方法评估提供了一种有吸引力的策略。