title: Fibroblasts derived from human embryonic stem cells direct development and repair of 3D human skin equivalents.
abstract: BACKGROUND
  Pluripotent, human stem cells hold tremendous promise as a source of progenitor and terminally differentiated cells for application in future regenerative therapies. However, such therapies will be dependent upon the development of novel approaches that can best assess tissue outcomes of pluripotent stem cell-derived cells and will be essential to better predict their safety and stability following in vivo transplantation.
  
  
  METHODS
  In this study we used engineered, human skin equivalents (HSEs) as a platform to characterize fibroblasts that have been derived from human embryonic stem (hES) cell. We characterized the phenotype and the secretion profile of two distinct hES-derived cell lines with properties of mesenchymal cells (EDK and H9-MSC) and compared their biological potential upon induction of differentiation to bone and fat and following their incorporation into the stromal compartment of engineered, HSEs.
  
  
  RESULTS
  While both EDK and H9-MSC cell lines exhibited similar morphology and mesenchymal cell marker expression, they demonstrated distinct functional properties when incorporated into the stromal compartment of HSEs. EDK cells displayed characteristics of dermal fibroblasts that could support epithelial tissue development and enable re-epithelialization of wounds generated using a 3D tissue model of cutaneous wound healing, which was linked to elevated production of hepatocyte growth factor (HGF). Lentiviral shRNA-mediated knockdown of HGF resulted in a dramatic decrease of HGF secretion from EDK cells that led to a marked reduction in their ability to promote keratinocyte proliferation and re-epithelialization of cutaneous wounds. In contrast, H9-MSCs demonstrated features of mesenchymal stem cells (MSC) but not those of dermal fibroblasts, as they underwent multilineage differentiation in monolayer culture, but were unable to support epithelial tissue development and repair and produced significantly lower levels of HGF.
  
  
  CONCLUSIONS
  Our findings demonstrate that hES-derived cells could be directed to specified and alternative mesenchymal cell fates whose function could be distinguished in engineered HSEs. Characterization of hES-derived mesenchymal cells in 3D, engineered HSEs demonstrates the utility of this tissue platform to predict the functional properties of hES-derived fibroblasts before their therapeutic transplantation.
authors.affiliation: Program in Cell, Molecular and Developmental Biology, Sackler School of Graduate Biomedical Sciences, Tufts University School of Medicine, 136 Harrison Avenue, Boston, MA, 02111, USA. Jonathan.Garlick@tufts.edu
authors.forename: Yulia
authors.initials: Y
authors.lastname: Shamis
authors.forename: Kyle J
authors.initials: KJ
authors.lastname: Hewitt
authors.forename: Mark W
authors.initials: MW
authors.lastname: Carlson
authors.forename: Mariam
authors.initials: M
authors.lastname: Margvelashvilli
authors.forename: Shumin
authors.initials: S
authors.lastname: Dong
authors.forename: Catherine K
authors.initials: CK
authors.lastname: Kuo
authors.forename: Laurence
authors.initials: L
authors.lastname: Daheron
authors.forename: Christophe
authors.initials: C
authors.lastname: Egles
authors.forename: Jonathan A
authors.initials: JA
authors.lastname: Garlick
mesh_terms: D018929:Cell Culture Techniques; D019070:Cell Lineage; D049109:Cell Proliferation; D053595:Embryonic Stem Cells / Q000166:cytology*; D005347:Fibroblasts / Q000166:cytology* / Q000637:transplantation; D017228:Hepatocyte Growth Factor / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D006801:Humans; D015603:Keratinocytes / Q000166:cytology; D034622:RNA Interference; D034741:RNA, Small Interfering / Q000378:metabolism; D023822:Tissue Engineering; D014945:Wound Healing
chemical_list: D034741:RNA, Small Interfering; D017228:Hepatocyte Growth Factor