UCLA

Skeletal muscle is one of the most regenerative tissues in the human body. Skeletal muscle progenitor cells (SMPCs) contribute to developmental myogenesis, and skeletal muscle stem cells (satellite cells, SCs) contribute to postnatal muscle homeostasis and regeneration. Differentiating human pluripotent stem cells (hPSCs) into SCs is a valuable resource for developing cell replacement therapies for muscle wasting diseases. However, current hPSC directed myogenic differentiation protocols result in immature, embryonic-like SMPCs that do not engraft and promote muscle regeneration as efficiently as postnatal SCs. Because it is unclear how SMPCs and SCs molecularly differ, it is not known how to mature SMPCs into SCs. Chapter 2 provides an overview on the current understandings of muscle development and the state of the field in generating directed differentiation myogenic cultures from hPSCs. In chapter 3, we found that although SMPCs and SCs from across development have many functional similarities in their roles in muscle formation, profiling them from both in vivo and in vitro hPSC directed differentiations using single-cell RNA-sequencing has revealed that they are transcriptionally distinct. However, it was unclear what regulated their differences in cell states. The chromatin landscape has a direct role in dictating the state of a cell, particularly through modulations in accessibility at regulatory regions which mediate transcription factor (TF) binding and control gene expression. In chapter 4, we evaluated differences in chromatin accessibility using single-nucleus ATAC-sequencing in tissue-derived SMPCs and SCs and hPSC-derived SMPCs (hPSC-SMPCs). We identified several TF candidates as potential regulators of development from embryonic SMPCs to postnatal SCs. For example, nuclear factor I family members were found to have roles in establishing stage-specific enhancers in the maturation from embryonic to fetal SMPCs. We also found that reduction of HMGA1 expression during directed differentiation promotes maturation of hPSC-SMPCs, and thus we identified HMGA1 as a TF that maintains SMPC immaturity. Collectively, this work sheds light on the transcriptional differences between muscle progenitor and stem cell states and the gene regulatory mechanisms that mediate these differences. It provides a comprehensive, multiomic view on the progression of human myogenesis in single-cell and single-nucleus resolution. These findings enhance our understanding of how to promote the regenerative potential of hPSC-derived muscle cells and enable the development of improved cell therapies for muscle diseases.