title: Characterization of vasculogenic potential of human adipose-derived endothelial cells in a three-dimensional vascularized skin substitute.
abstract: OBJECTIVE
  The need for clinically applicable skin substitutes continues to be a matter of fact. Hypothetically, a laboratory grown autologous skin analog with near normal architecture might be a suitable approach to yield both satisfactory functional and cosmetic long-term results. In this study, we explored the use of human endothelial cells derived from freshly isolated adipose stromal vascular fraction (SVF) in a three-dimensional (3D) co-culture model of vascularized bio-engineered skin substitute.
  
  
  METHODS
  The SVF was isolated from human white adipose tissue samples and keratinocytes from human skin biopsies. The SVF, in particular endothelial cells, were characterized using flow cytometry and immuofluorescence analysis. Endothelial and mesenchymal progenitors from the SVF formed blood capillaries after seeding into a 3D collagen type I hydrogel in vitro. Subsequently, human keratinocytes were seeded on the top of those hydrogels to develop a vascularized dermo-epidermal skin substitute.
  
  
  RESULTS
  Flow cytometric analysis of surface markers of the freshly isolated SVF showed the expression of endothelial markers (CD31, CD34, CD146), mesenchymal/stromal cell-associated markers (CD44, CD73, CD90, CD105), stem cell markers (CD49f, CD117, CD133), and additionally hematopoietic markers (CD14, CD15, CD45). Further analysis of white adipose-derived endothelial cells (watECs) revealed the co-expression of CD31, CD34, CD90, CD105, and partially CD146 on these cells. WatECs were separated from adipose-stromal cells (watASCs) using FACS sorting. WatASCs and watECs cultured separately in a 3D hydrogel for 3 weeks did not form any vascular structures. Only if co-cultured, both cell types aligned to develop a ramified vascular network in vitro with continuous endothelial lumen formation. Transplantation of those 3D-hydrogels onto immuno-incompetent rats resulted in a rapid connection of human capillaries with the host vessels and formation of functional, blood-perfused mosaic human-rat vessels within only 3-4 days.
  
  
  CONCLUSIONS
  Adipose tissue represents an attractive cell source due to the ease of isolation and abundance of endothelial as well as mesenchymal cell lineages. Adipose-derived SVF cells exhibit the ability to form microvascular structures in vitro and support the accelerated blood perfusion in skin substitutes in vivo when transplanted.
authors.affiliation: Tissue Biology Research Unit, Department of Surgery, University Children's Hospital Zurich, Zurich, Switzerland.|Children's Research Center, University Children's Hospital Zurich, Zurich, Switzerland.
authors.forename: Agnes S
authors.initials: AS
authors.lastname: Klar
authors.affiliation: Department of Biomedicine, University Hospital of Basel, University of Basel, Basel, Switzerland.
authors.forename: Sinan
authors.initials: S
authors.lastname: Güven
authors.affiliation: Tissue Biology Research Unit, Department of Surgery, University Children's Hospital Zurich, Zurich, Switzerland.|Children's Research Center, University Children's Hospital Zurich, Zurich, Switzerland.
authors.forename: Jakub
authors.initials: J
authors.lastname: Zimoch
authors.affiliation: Tissue Biology Research Unit, Department of Surgery, University Children's Hospital Zurich, Zurich, Switzerland.|Children's Research Center, University Children's Hospital Zurich, Zurich, Switzerland.
authors.forename: Natalia A
authors.initials: NA
authors.lastname: Zapiórkowska
authors.affiliation: Tissue Biology Research Unit, Department of Surgery, University Children's Hospital Zurich, Zurich, Switzerland.|Children's Research Center, University Children's Hospital Zurich, Zurich, Switzerland.
authors.forename: Thomas
authors.initials: T
authors.lastname: Biedermann
authors.affiliation: Tissue Biology Research Unit, Department of Surgery, University Children's Hospital Zurich, Zurich, Switzerland.|Children's Research Center, University Children's Hospital Zurich, Zurich, Switzerland.|Department of Surgery, University Children's Hospital Zurich, Steinwiesstrasse 75, 8032, Zurich, Switzerland.
authors.forename: Sophie
authors.initials: S
authors.lastname: Böttcher-Haberzeth
authors.affiliation: Department of Plastic, Reconstructive, Esthetical and Hand Surgery, Kantonsspital Aarau, Aarau, Switzerland.
authors.forename: Claudia
authors.initials: C
authors.lastname: Meuli-Simmen
authors.affiliation: Department of Biomedicine, University Hospital of Basel, University of Basel, Basel, Switzerland.
authors.forename: Ivan
authors.initials: I
authors.lastname: Martin
authors.affiliation: Department of Biomedicine, University Hospital of Basel, University of Basel, Basel, Switzerland.
authors.forename: Arnaud
authors.initials: A
authors.lastname: Scherberich
authors.affiliation: Tissue Biology Research Unit, Department of Surgery, University Children's Hospital Zurich, Zurich, Switzerland.|Children's Research Center, University Children's Hospital Zurich, Zurich, Switzerland.
authors.forename: Ernst
authors.initials: E
authors.lastname: Reichmann
authors.affiliation: Department of Surgery, University Children's Hospital Zurich, Steinwiesstrasse 75, 8032, Zurich, Switzerland. Martin.Meuli@kispi.uzh.ch.
authors.forename: Martin
authors.initials: M
authors.lastname: Meuli
mesh_terms: D000273:Adipose Tissue / Q000098:blood supply* / Q000166:cytology / Q000378:metabolism*; D000293:Adolescent; D000328:Adult; D000368:Aged; D000818:Animals; D002478:Cells, Cultured; D002648:Child; D002675:Child, Preschool; D018920:Coculture Techniques; D042783:Endothelial Cells / Q000166:cytology / Q000378:metabolism*; D005260:Female; D005434:Flow Cytometry; D005455:Fluorescent Antibody Technique; D006801:Humans; D020136:Hydrogel, Polyethylene Glycol Dimethacrylate / Q000378:metabolism; D015603:Keratinocytes / Q000378:metabolism; D008297:Male; D008875:Middle Aged; D051381:Rats; D017768:Skin, Artificial; D017154:Stromal Cells / Q000166:cytology / Q000378:metabolism; D055815:Young Adult
chemical_list: D020136:Hydrogel, Polyethylene Glycol Dimethacrylate
keywords: Adipose stem cells; Angiogenesis; Blood capillaries; Endothelial cells; Mesenchymal cells; Skin tissue engineering; Stromal vascular fraction; Vascular network formation