Differentiation of human oligodendrocytes from pluripotent stem cells

Nat Protoc. 2009;4(11):1614-22. doi: 10.1038/nprot.2009.186. Epub 2009 Oct 15.


We have developed a four-part protocol to differentiate human embryonic stem cells (hESCs) to oligodendrocyte progenitor cells (OPCs) according to developmental principles. In the first 2 weeks, hESCs are induced to differentiate into neuroepithelial cells, which form neural tube-like rosettes. In the following 10 d, these neuroepithelial cells are specified to OLIG2-expressing progenitors in the presence of retinoic acid (RA) and sonic hedgehog (SHH). Upon treatment with fibroblast growth factor 2 (FGF2) for another 10 d, these progenitors convert to OLIG2 and NKX2.2-expressing pre-OPCs. Finally, the pre-OPCs take 8-9 weeks to differentiate into OPCs, which express additional markers of oligodendrocytes, such as SOX10, platelet-derived growth factor receptor alpha (PDGFRalpha) and NG2. The unique aspects of the protocol are the use of FGF2 to promote the differentiation of gliogenic pre-OPCs in the third part and the removal of FGF2 during the transition of pre-OPCs to OPCs. This 3-month differentiation protocol consistently yields OPCs of high purity capable of producing myelin sheaths in vivo.

Publication types

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

MeSH terms

  • Basic Helix-Loop-Helix Transcription Factors / metabolism
  • Cell Culture Techniques / methods*
  • Cell Differentiation / drug effects
  • Fibroblast Growth Factor 2 / pharmacology
  • Hedgehog Proteins / pharmacology
  • Homeobox Protein Nkx-2.2
  • Homeodomain Proteins / metabolism
  • Humans
  • Nerve Tissue Proteins / metabolism
  • Neuroepithelial Cells / cytology
  • Neuroepithelial Cells / drug effects
  • Neuroepithelial Cells / metabolism
  • Nuclear Proteins
  • Oligodendrocyte Transcription Factor 2
  • Oligodendroglia / cytology*
  • Oligodendroglia / drug effects
  • Oligodendroglia / metabolism
  • Pluripotent Stem Cells / cytology*
  • Pluripotent Stem Cells / drug effects
  • Pluripotent Stem Cells / metabolism
  • Transcription Factors / metabolism
  • Tretinoin / pharmacology
  • Zebrafish Proteins


  • Basic Helix-Loop-Helix Transcription Factors
  • Hedgehog Proteins
  • Homeobox Protein Nkx-2.2
  • Homeodomain Proteins
  • NKX2-2 protein, human
  • Nerve Tissue Proteins
  • Nuclear Proteins
  • OLIG2 protein, human
  • Oligodendrocyte Transcription Factor 2
  • SHH protein, human
  • Transcription Factors
  • Zebrafish Proteins
  • nkx2.2b protein, zebrafish
  • Fibroblast Growth Factor 2
  • Tretinoin