title: Differentiation of equine induced pluripotent stem cells into a keratinocyte lineage.
abstract: BACKGROUND
  Skin trauma in horses often leads to the development of chronic nonhealing wounds that lack a keratinocyte cover, vital for healing. Reports in mouse and man confirm the possibility of generating functional keratinocytes from induced pluripotent stem cells (iPSC), thus presenting myriad potential applications for wound management or treatment of skin disease. Similarly, differentiation of equine iPSC (eiPSC) into a keratinocyte lineage should provide opportunities for the advancement of veterinary regenerative medicine.
  
  
  OBJECTIVE
  The purpose of this study was to develop an efficient method for the differentiation of eiPSC into a keratinocyte lineage. It was hypothesised that eiPSC can form differentiated keratinocytes (eiPSC-KC) comparable with primary equine keratinocytes (PEK) in their morphological and functional characteristics.
  
  
  METHODS
  Experimental in vitro study.
  
  
  METHODS
  Equine iPSC established using a nonviral system were treated for 30 days with retinoic acid and bone morphogenetic protein-4 to induce directed differentiation into iPSC-KC. Temporospatial gene and protein expression by eiPSC-KC was measured at weekly intervals of differentiation and in response to calcium switch. Proliferative and migratory capacities of eiPSC-KC were compared with those of PEK.
  
  
  RESULTS
  Equine iPSC, upon directed differentiation, showed loss of pluripotency genes and progressive increase in pancytokeratin expression indicating ectodermal specification into keratinocytes. High differentiation efficiency was achieved, with 82.5% of eiPSC expressing keratin 14, a marker of epidermal-specific basal stem cells, after 30 days of directed differentiation. Moreover, the proliferative capacity of eiPSC-KC was superior, while the migratory capacity (measured as the ability to epithelise in vitro wounds) was comparable with that of PEK.
  
  
  CONCLUSIONS
  This proof of concept study suggests that eiPSC can successfully be differentiated into equine keratinocytes (eiPSC-KC) with features that are promising to the development of a stem cell-based skin construct, with the potential to regenerate lost or damaged skin.
authors.affiliation: Département de Biomédecine Vétérinaire, Faculté de Médecine Vétérinaire, Université de Montréal, Saint-Hyacinthe, Québec, Canada.
authors.forename: C
authors.initials: C
authors.lastname: Aguiar
authors.affiliation: Département de Biomédecine Vétérinaire, Faculté de Médecine Vétérinaire, Université de Montréal, Saint-Hyacinthe, Québec, Canada.
authors.forename: J
authors.initials: J
authors.lastname: Therrien
authors.affiliation: Département de Pathologie et Microbiologie, Faculté de Médecine Vétérinaire, Université de Montréal, Saint-Hyacinthe, Québec, Canada.
authors.forename: P
authors.initials: P
authors.lastname: Lemire
authors.affiliation: Département de Pathologie et Microbiologie, Faculté de Médecine Vétérinaire, Université de Montréal, Saint-Hyacinthe, Québec, Canada.
authors.forename: M
authors.initials: M
authors.lastname: Segura
authors.affiliation: Département de Biomédecine Vétérinaire, Faculté de Médecine Vétérinaire, Université de Montréal, Saint-Hyacinthe, Québec, Canada.
authors.forename: L C
authors.initials: LC
authors.lastname: Smith
authors.affiliation: Département de Biomédecine Vétérinaire, Faculté de Médecine Vétérinaire, Université de Montréal, Saint-Hyacinthe, Québec, Canada.
authors.forename: C L
authors.initials: CL
authors.lastname: Theoret
mesh_terms: D000818:Animals; D015415:Biomarkers; D002454:Cell Differentiation / Q000502:physiology*; D002478:Cells, Cultured; D003470:Culture Media; D005786:Gene Expression Regulation / Q000502:physiology; D006736:Horses; D057026:Induced Pluripotent Stem Cells / Q000502:physiology*; D015603:Keratinocytes / Q000502:physiology*
chemical_list: D015415:Biomarkers; D003470:Culture Media
keywords: horse; induced pluripotent stem cells; keratinocytes; regenerative medicine; wound healing