Generation and characterization of leukemia inhibitory factor-dependent equine induced pluripotent stem cells from adult dermal fibroblasts

Stem Cells Dev. 2014 Jul 1;23(13):1515-23. doi: 10.1089/scd.2013.0461. Epub 2014 Apr 1.

Abstract

In this study we have reprogrammed dermal fibroblasts from an adult female horse into equine induced pluripotent stem cells (equiPSCs). These equiPSCs are dependent only on leukemia inhibitory factor (LIF), placing them in striking contrast to previously derived equiPSCs that have been shown to be co-dependent on both LIF and basic fibroblast growth factor (bFGF). These equiPSCs have a normal karyotype and have been maintained beyond 60 passages. They possess alkaline phosphatase activity and express eqNANOG, eqOCT4, and eqTERT mRNA. Immunocytochemistry confirmed that they produce NANOG, REX1, SSEA4, TRA1-60, and TRA1-81. While our equiPSCs are LIF dependent, bFGF co-stimulates their proliferation via the PI3K/AKT pathway. EquiPSCs lack expression of eqXIST and immunostaining for H3K27me3, suggesting that during reprogramming the inactive X chromosome has likely been reactivated to generate cells that have two active X chromosomes. EquiPSCs form embryoid bodies and in vitro teratomas that contain derivatives of all three germ layers. These LIF-dependent equiPSCs likely reflect a more naive state of pluripotency than equiPSCs that are co-dependent on both LIF and bFGF and so provide a novel resource for understanding pluripotency in the horse.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Biomarkers / metabolism
  • Cell Proliferation
  • Coculture Techniques
  • Feeder Cells
  • Female
  • Fibroblast Growth Factors / physiology
  • Fibroblasts / physiology*
  • Gene Expression
  • Histones / metabolism
  • Horses
  • Induced Pluripotent Stem Cells / physiology*
  • Leukemia Inhibitory Factor / physiology*
  • Skin / cytology*
  • Transcription Factors / metabolism
  • X Chromosome / genetics

Substances

  • Biomarkers
  • Histones
  • Leukemia Inhibitory Factor
  • Transcription Factors
  • Fibroblast Growth Factors