Bulk Differentiation of Human Induced Pluripotent Stem Cells into Contractile Skeletal Muscles

Effective treatments for various muscular disorders and a clear understanding of their disease pathology remain lacking owing to the limitations of traditional animal models and two-dimensional (2D) cell cultures. Human induced pluripotent stem cells (hiPSCs) offer promising platforms for disease modeling and regenerative medicine. However, existing protocols for skeletal muscle differentiation are often labor-intensive and yield inconsistent results. Here, we utilized a piggyBac transposon-based system to generate bulk MYOD1-hiPSCs, eliminating the need for clonal selection and enabling rapid, efficient, and reproducible skeletal muscle differentiation. Furthermore, we established a protocol for generating functional three-dimensional (3D) skeletal muscle tissues that exert contractile force in response to electrical stimulation, highlighting their physiological relevance. This system provides a simple and valuable tool for disease modeling, drug screening, and the study of sporadic muscle disorders using patient-derived hiPSCs.