Large-Scale Expansion of Human iPSC-Derived Skeletal Muscle Cells for Disease Modeling and Cell-Based Therapeutic Strategies

Summary Although skeletal muscle cells can be generated from human induced pluripotent stem cells (iPSCs), transgene-free protocols include only limited options for their purification and expansion. In this study, we found that fluorescence-activated cell sorting-purified myogenic progenitors generated from healthy controls and Pompe disease iPSCs can be robustly expanded as much as 5 × 1011-fold. At all steps during expansion, cells could be cryopreserved or differentiated into myotubes with a high fusion index. In vitro, cells were amenable to maturation into striated and contractile myofibers. Insertion of acid α-glucosidase cDNA into the AAVS1 locus in iPSCs using CRISPR/Cas9 prevented glycogen accumulation in myotubes generated from a patient with classic infantile Pompe disease. In vivo, the expression of human-specific nuclear and sarcolemmar antigens indicated that myogenic progenitors engraft into murine muscle to form human myofibers. This protocol is useful for modeling of skeletal muscle disorders and for using patient-derived, gene-corrected cells to develop cell-based strategies.
Highlights • Transgene-free protocol for generation and expansion of myogenic progenitors • Differentiation into contractile skeletal muscle cells in vitro • Correction of glycogen accumulation in Pompe disease using CRISPR/cas9 • Contribution to muscle regeneration in vivo
Van der Wal et al. present a robust protocol for the transgene-free generation and purification of myogenic progenitors from human iPSCs and for their expansion up to 5 × 1011-fold. After gene editing in vitro, these myogenic progenitors matured into contractile skeletal muscle cells, reversing Pompe disease pathology. In vivo, myogenic progenitors contributed to muscle regeneration.