Nestin-EGFP transgenic mice: visualization of the self-renewal and multipotency of CNS stem cells.
We generated transgenic mice carrying enhanced green fluorescent protein (EGFP) under the control of the nestin second-intronic enhancer (E/nestin:EGFP). Flow cytometry followed by in vitro assays revealed that in situ EGFP expression in the embryonic brain correlated with the mitotic index, the cogeneration of both neurons and glia, and the frequency of neurosphere formation in vitro. High-level EGFP expressors derived from embryos included a distinct subpopulation of cells that were self-renewable and multipotent, criteria that define neural stem cells (NSCs). Such cells were largely absent among lower-level or non-EGFP expressors, thereby permitting us to enrich for NSCs using EGFP expression level. In adults, although E/nestin:EGFP-positive cells included the NSC population, the frequency of neurosphere formation did not correlate directly with the level of EGFP expression. However, moderately EGFP-expressing cells in adults gained EGFP intensity when they formed neurospheres, suggesting embryonic and adult NSCs exist in different microenvironments in vivo.
Kawaguchi
A
A
Division of Neuroanatomy, Osaka University Graduate School of Medicine, Suita, Osaka, 565-0871, Japan.
Miyata
T
T
Sawamoto
K
K
Takashita
N
N
Murayama
A
A
Akamatsu
W
W
Ogawa
M
M
Okabe
M
M
Tano
Y
Y
Goldman
S A
SA
Okano
H
H
D000818:Animals; D001253:Astrocytes / Q000166:cytology / Q000502:physiology; D001921:Brain / Q000166:cytology* / Q000196:embryology; D018929:Cell Culture Techniques / Q000379:methods; D002455:Cell Division / Q000502:physiology; D002478:Cells, Cultured; D004742:Enhancer Elements, Genetic / Q000502:physiology; D005434:Flow Cytometry; D018507:Gene Expression Regulation, Developmental; D017930:Genes, Reporter; D049452:Green Fluorescent Proteins; D007202:Indicators and Reagents / Q000378:metabolism*; D007381:Intermediate Filament Proteins / Q000032:analysis / Q000235:genetics*; D007438:Introns; D008164:Luminescent Proteins / Q000235:genetics*; D051379:Mice; D008822:Mice, Transgenic; D008938:Mitosis / Q000502:physiology; D008969:Molecular Sequence Data; D009419:Nerve Tissue Proteins; D064231:Nestin; D009474:Neurons / Q000166:cytology / Q000502:physiology; D051381:Rats; D013234:Stem Cells / Q000166:cytology* / Q000502:physiology; D014404:Tubulin / Q000032:analysis
D007202:Indicators and Reagents; D007381:Intermediate Filament Proteins; D008164:Luminescent Proteins; D009419:Nerve Tissue Proteins; C577485:Nes protein, mouse; C577486:Nes protein, rat; D064231:Nestin; D014404:Tubulin; D049452:Green Fluorescent Proteins