Direct Reprogramming of Mouse Embryonic Fibroblasts to Induced Trophoblast Stem Cells

Methods Mol Biol. 2020;2117:285-292. doi: 10.1007/978-1-0716-0301-7_18.

Abstract

Trophoblast cells are the first committed lineage to emerge during mammalian preimplantation embryo development. Trophoblast stem (TS) cells can be derived from the trophectoderm (TE) of blastocyst-stage embryos and differentiate into extraembryonic trophoblast cells of the placenta. While mouse TS cells are an indispensable tool to study placental development, and reproductive diseases such as implantation failure and recurrent miscarriage, human TS cells have not been isolated. To model human trophoblast development and to investigate trophoblast-specific causes of reproductive diseases, it will be important to derive human induced trophoblast stem (iTS) cells. Recent studies have shown that fibroblasts can be reprogrammed to iTS cells by overexpressing four transcription factors (TFs) including TFAP2C, GATA3, EOMES, and ETS2. Here, we describe a protocol to directly convert mouse embryonic fibroblasts (MEFs) to iTS cells following overexpression of 10 TFs. iTS cells are capable of self-renewing using conventional TS cell culture media supplemented with the external signal FGF4 and heparin. iTS cells are also able to differentiate into trophoblast lineages.

Keywords: Blastocyst; Derivation; Differentiation; FGF4; Trophectoderm; Trophoblast stem cells; iTS cells.

MeSH terms

  • Animals
  • Cell Differentiation
  • Cell Self Renewal
  • Cells, Cultured
  • Cellular Reprogramming
  • Cellular Reprogramming Techniques / methods*
  • Embryo, Mammalian / cytology
  • Embryonic Stem Cells / cytology*
  • Embryonic Stem Cells / metabolism
  • Female
  • Fibroblasts / cytology*
  • GATA3 Transcription Factor / genetics
  • GATA3 Transcription Factor / metabolism
  • HEK293 Cells
  • Humans
  • Lentivirus / genetics
  • Mice
  • Pregnancy
  • Proto-Oncogene Protein c-ets-2 / genetics
  • Proto-Oncogene Protein c-ets-2 / metabolism
  • T-Box Domain Proteins / genetics
  • T-Box Domain Proteins / metabolism
  • Transcription Factor AP-2 / genetics
  • Transcription Factor AP-2 / metabolism
  • Transcription Factors / genetics*
  • Transcription Factors / metabolism
  • Transduction, Genetic
  • Trophoblasts / cytology*

Substances

  • GATA3 Transcription Factor
  • Proto-Oncogene Protein c-ets-2
  • T-Box Domain Proteins
  • Transcription Factor AP-2
  • Transcription Factors