Directed differentiation of ventral spinal progenitors and motor neurons from human embryonic stem cells by small molecules

Stem Cells. 2008 Apr;26(4):886-93. doi: 10.1634/stemcells.2007-0620. Epub 2008 Jan 31.

Abstract

Specification of distinct cell types from human embryonic stem cells (hESCs) is key to the potential application of these naïve pluripotent cells in regenerative medicine. Determination of the nontarget differentiated populations, which is lacking in the field, is also crucial. Here, we show an efficient differentiation of motor neurons ( approximately 50%) by a simple sequential application of retinoid acid and sonic hedgehog (SHH) in a chemically defined suspension culture. We also discovered that purmorphamine, a small molecule that activates the SHH pathway, could replace SHH for the generation of motor neurons. Immunocytochemical characterization indicated that cells differentiated from hESCs were nearly completely restricted to the ventral spinal progenitor fate (NKX2.2+, Irx3+, and Pax7-), with the exception of motor neurons (HB9+) and their progenitors (Olig2+). Thus, the directed neural differentiation system with small molecules, even without further purification, will facilitate basic and translational studies using human motoneurons at a minimal cost.

Publication types

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

MeSH terms

  • Animals
  • Cell Differentiation* / drug effects
  • Cell Differentiation* / physiology
  • Cell Line
  • Cells, Cultured
  • Directed Molecular Evolution / methods*
  • Embryonic Stem Cells / cytology*
  • Embryonic Stem Cells / drug effects
  • Embryonic Stem Cells / physiology
  • Hedgehog Proteins / biosynthesis
  • Hedgehog Proteins / genetics
  • Hedgehog Proteins / physiology
  • Humans
  • Mice
  • Morpholines / pharmacology
  • Motor Neurons / cytology*
  • Motor Neurons / drug effects
  • Motor Neurons / physiology*
  • Purines / pharmacology
  • Spinal Cord / cytology
  • Spinal Cord / drug effects
  • Spinal Cord / physiology
  • Stem Cells / cytology
  • Stem Cells / drug effects
  • Stem Cells / physiology
  • Tretinoin / pharmacology
  • Tretinoin / physiology

Substances

  • Hedgehog Proteins
  • Morpholines
  • Purines
  • Tretinoin
  • purmorphamine