Direct Cell Reprogramming of Mouse Fibroblasts into Functional Astrocytes Using Lentiviral Overexpression of the Transcription Factors NFIA, NFIB, and SOX9

Methods Mol Biol. 2021:2352:31-43. doi: 10.1007/978-1-0716-1601-7_3.

Abstract

Astrocytes play an important role in maintaining brain homeostasis and their dysfunction is involved in a number of neurological disorders. An accessible source of astrocytes is essential to model neurological diseases and potential cell therapy approaches. Cell reprogramming techniques offer possibilities to reprogram terminally differentiated cells into other cell types. By overexpressing the three astrocytic transcription factors NFIA, NFIB, and SOX9, we showed that it is possible to directly transdifferentiate fibroblasts into functional astrocytes. These induced astrocytes (iAstrocytes) express glial fibrillary acidic protein (GFAP) and S100 calcium binding protein B (S100B), as well as other astrocytic markers. Moreover, electrophysiological properties indicate that iAstrocytes are functionally comparable to native brain astrocytes. Here we describe an optimized protocol to generate iAstrocytes starting from skin fibroblasts and this approach can be adapted for a wide range of somatic cell types.

Keywords: Astrocytes; Cell reprogramming; Fibroblasts; Lentivirus; Transcription factors.

MeSH terms

  • Animals
  • Astrocytes / cytology*
  • Astrocytes / metabolism*
  • Calcium
  • Cell Line
  • Cell Transdifferentiation / genetics*
  • Cells, Cultured
  • Cellular Reprogramming / genetics*
  • Fibroblasts / cytology*
  • Fibroblasts / metabolism*
  • Gene Expression
  • Genetic Vectors / biosynthesis
  • Genetic Vectors / genetics
  • Humans
  • Lentivirus / genetics
  • Mice
  • Molecular Imaging
  • NFI Transcription Factors / genetics
  • SOX9 Transcription Factor / genetics
  • Transcription Factors / genetics*
  • Transcription Factors / metabolism

Substances

  • NFI Transcription Factors
  • Nfia protein, mouse
  • Nfib protein, mouse
  • SOX9 Transcription Factor
  • Sox9 protein, mouse
  • Transcription Factors
  • Calcium