Gene expression profiling of skeletal myogenesis in human embryonic stem cells reveals a potential cascade of transcription factors regulating stages of myogenesis, including quiescent/activated satellite cell-like gene expression

PLoS One. 2019 Sep 27;14(9):e0222946. doi: 10.1371/journal.pone.0222946. eCollection 2019.

Abstract

Human embryonic stem cell (hESC)-derived skeletal muscle progenitors (SMP)-defined as PAX7-expressing cells with myogenic potential-can provide an abundant source of donor material for muscle stem cell therapy. As in vitro myogenesis is decoupled from in vivo timing and 3D-embryo structure, it is important to characterize what stage or type of muscle is modeled in culture. Here, gene expression profiling is analyzed in hESCs over a 50 day skeletal myogenesis protocol and compared to datasets of other hESC-derived skeletal muscle and adult murine satellite cells. Furthermore, day 2 cultures differentiated with high or lower concentrations of CHIR99021, a GSK3A/GSK3B inhibitor, were contrasted. Expression profiling of the 50 day time course identified successively expressed gene subsets involved in mesoderm/paraxial mesoderm induction, somitogenesis, and skeletal muscle commitment/formation which could be regulated by a putative cascade of transcription factors. Initiating differentiation with higher CHIR99021 concentrations significantly increased expression of MSGN1 and TGFB-superfamily genes, notably NODAL, resulting in enhanced paraxial mesoderm and reduced ectoderm/neuronal gene expression. Comparison to adult satellite cells revealed that genes expressed in 50-day cultures correlated better with those expressed by quiescent or early activated satellite cells, which have the greatest therapeutic potential. Day 50 cultures were similar to other hESC-derived skeletal muscle and both expressed known and novel SMP surface proteins. Overall, a putative cascade of transcription factors has been identified which regulates four stages of myogenesis. Subsets of these factors were upregulated by high CHIR99021 or their binding sites were significantly over-represented during SMP activation, ranging from quiescent to late-activated stages. This analysis serves as a resource to further study the progression of in vitro skeletal myogenesis and could be mined to identify novel markers of pluripotent-derived SMPs or regulatory transcription/growth factors. Finally, 50-day hESC-derived SMPs appear similar to quiescent/early activated satellite cells, suggesting they possess therapeutic potential.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Cell Differentiation / drug effects
  • Cell Differentiation / genetics
  • Cell Line
  • Gene Expression Profiling
  • Gene Expression Regulation, Developmental*
  • Glycogen Synthase Kinase 3 beta / antagonists & inhibitors
  • Glycogen Synthase Kinase 3 beta / metabolism
  • Human Embryonic Stem Cells / metabolism*
  • Humans
  • Muscle Development / genetics*
  • Muscle, Skeletal / cytology
  • Muscle, Skeletal / growth & development*
  • Pyridines / pharmacology
  • Pyrimidines / pharmacology
  • Satellite Cells, Skeletal Muscle / metabolism
  • Transcription Factors / metabolism*

Substances

  • Chir 99021
  • Pyridines
  • Pyrimidines
  • Transcription Factors
  • GSK3B protein, human
  • Glycogen Synthase Kinase 3 beta