Differentiation of neurons from neural precursors generated in floating spheres from embryonic stem cells.
BACKGROUND
Neural differentiation of embryonic stem (ES) cells is usually achieved by induction of ectoderm in embryoid bodies followed by the enrichment of neuronal progenitors using a variety of factors. Obtaining reproducible percentages of neural cells is difficult and the methods are time consuming.


RESULTS
Neural progenitors were produced from murine ES cells by a combination of nonadherent conditions and serum starvation. Conversion to neural progenitors was accompanied by downregulation of Oct4 and NANOG and increased expression of nestin. ES cells containing a GFP gene under the control of the Sox1 regulatory regions became fluorescent upon differentiation to neural progenitors, and ES cells with a tau-GFP fusion protein became fluorescent upon further differentiation to neurons. Neurons produced from these cells upregulated mature neuronal markers, or differentiated to glial and oligodendrocyte fates. The neurons gave rise to action potentials that could be recorded after application of fixed currents.


CONCLUSIONS
Neural progenitors were produced from murine ES cells by a novel method that induced neuroectoderm cells by a combination of nonadherent conditions and serum starvation, in contrast to the embryoid body method in which neuroectoderm cells must be selected after formation of all three germ layers.
Department of Otolarygology, EENT Hospital of Shanghai Medical College of Fudan University, Institute of Biomedical Sciences, Fudan University Shanghai, 200031, PR China. hwli@shmu.edu.cn
Huawei
H
Li
Hong
H
Liu
C Eduardo
CE
Corrales
Jessica R
JR
Risner
Jeff
J
Forrester
Jeffrey R
JR
Holt
Stefan
S
Heller
Albert S B
AS
Edge
D000818:Animals; D017209:Apoptosis; D001253:Astrocytes / Q000502:physiology; D018929:Cell Culture Techniques / Q000379:methods; D002478:Cells, Cultured; D016895:Culture Media, Serum-Free; D004594:Electrophysiology; D053595:Embryonic Stem Cells / Q000166:cytology*; D007150:Immunohistochemistry; D020287:In Situ Nick-End Labeling; D007381:Intermediate Filament Proteins / Q000378:metabolism; D051379:Mice; D039902:Multipotent Stem Cells / Q000166:cytology* / Q000378:metabolism; D009419:Nerve Tissue Proteins / Q000378:metabolism; D064231:Nestin; D055495:Neurogenesis / Q000502:physiology*; D009474:Neurons / Q000378:metabolism / Q000502:physiology; D018408:Patch-Clamp Techniques; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012815:Signal Processing, Computer-Assisted
D016895:Culture Media, Serum-Free; D007381:Intermediate Filament Proteins; D009419:Nerve Tissue Proteins; C577485:Nes protein, mouse; D064231:Nestin