Direct isolation of committed neuronal progenitor cells from transgenic mice coexpressing spectrally distinct fluorescent protemins regulated by stage-specific neural promoters

Kazunobu Sawamoto, Atsuyo Yamamoto, Ayano Kawaguchi, Masahiro Yamaguchi, Kensaku Mori, Steven A. Goldman, Hideyuki Okano

Research output: Contribution to journalArticle

56 Citations (Scopus)

Abstract

Many tissues arise from pluripotent stem cells through cell-type specification and maturation. In the bone marrow, primitive stem cells generate all the different types of blood cells via the sequential differentiation of increasingly committed progenitor cells. Cell-surface markers that clearly distinguish stem cells, restricted progenitors, and differentiated progeny have enabled researchers to isolate these cells and to study the regulatory mechanisms of hematopoiesis. Neuronal differentiation appears to involve similar mechanisms. However, neural progenitor cells that are restricted to a neuronal fate have not been characterized in vivo, because specific cell-surface markers are not available. We have developed an alternative strategy to identify and isolate neuronal progenitor cells based on dual-color fluorescent proteins. To identify and isolate directly progenitor cells from brain tissue without the need for either transfection or intervening cell culture, we established lines of transgenic mice bearing fluorescent transgenes regulated by neural promoters. One set of transgenic lines expressed enhanced yellow fluorescent protein (EYFP) in neuronal progenitor cells and neurons under the control of the Tα1 α-tubulin promoter. Another line expressed enhanced green fluorescent protein (EGFP) in immature neural cells under the control of the enhancer/promoter elements of the nestin gene. By crossing these lines we obtained mice expressing both transgenes. To isolate neuronal progenitor cells directly from the developing brain, we used flow cytometry, selecting cells that expressed EGFP and EYFP simultaneously. We expect this strategy to provide valuable material with which to study the mechanisms of neurogenesis and to develop cell-based therapies for neurological disorders.

Original languageEnglish
Pages (from-to)220-227
Number of pages8
JournalJournal of neuroscience research
Volume65
Issue number3
DOIs
Publication statusPublished - 2001 Aug 1

Keywords

  • Fluorescence-activated cell sorting
  • Green fluorescent protein
  • Nestin
  • Neuronal progenitor cells
  • Transgenic mouse
  • Tα1

ASJC Scopus subject areas

  • Cellular and Molecular Neuroscience

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