title: Fetal fibroblasts and keratinocytes with immunosuppressive properties for allogeneic cell-based wound therapy.
abstract: Fetal skin heals rapidly without scar formation early in gestation, conferring to fetal skin cells a high and unique potential for tissue regeneration and scar management. In this study, we investigated the possibility of using fetal fibroblasts and keratinocytes to stimulate wound repair and regeneration for further allogeneic cell-based therapy development. From a single fetal skin sample, two clinical batches of keratinocytes and fibroblasts were manufactured and characterized. Tolerogenic properties of the fetal cells were investigated by allogeneic PBMC proliferation tests. In addition, the potential advantage of fibroblasts/keratinocytes co-application for wound healing stimulation has been examined in co-culture experiments with in vitro scratch assays and a multiplex cytokines array system. Based on keratin 14 and prolyl-4-hydroxylase expression analyses, purity of both clinical batches was found to be above 98% and neither melanocytes nor Langerhans cells could be detected. Both cell types demonstrated strong immunosuppressive properties as shown by the dramatic decrease in allogeneic PBMC proliferation when co-cultured with fibroblasts and/or keratinocytes. We further showed that the indoleamine 2,3 dioxygenase (IDO) activity is required for the immunoregulatory activity of fetal skin cells. Co-cultures experiments have also revealed that fibroblasts-keratinocytes interactions strongly enhanced fetal cells secretion of HGF, GM-CSF, IL-8 and to a lesser extent VEGF-A. Accordingly, in the in vitro scratch assays the fetal fibroblasts and keratinocytes co-culture accelerated the scratch closure compared to fibroblast or keratinocyte mono-cultures. In conclusion, our data suggest that the combination of fetal keratinocytes and fibroblasts could be of particular interest for the development of a new allogeneic skin substitute with immunomodulatory activity, acting as a reservoir for wound healing growth factors.
authors.affiliation: Cell and Gene Therapy Unit, CIC Biotherapy INSERM 0503, Hôtel-Dieu University Hospital, Nantes, France.
authors.forename: Thomas
authors.initials: T
authors.lastname: Zuliani
authors.forename: Soraya
authors.initials: S
authors.lastname: Saiagh
authors.forename: Anne-Chantal
authors.initials: AC
authors.lastname: Knol
authors.forename: Julie
authors.initials: J
authors.lastname: Esbelin
authors.forename: Brigitte
authors.initials: B
authors.lastname: Dréno
mesh_terms: D002478:Cells, Cultured; D018920:Coculture Techniques / Q000379:methods; D005260:Female; D005333:Fetus / Q000166:cytology; D005347:Fibroblasts / Q000166:cytology*; D005434:Flow Cytometry; D016178:Granulocyte-Macrophage Colony-Stimulating Factor / Q000378:metabolism; D006801:Humans; D007150:Immunohistochemistry; D050503:Indoleamine-Pyrrole 2,3,-Dioxygenase; D016209:Interleukin-8 / Q000378:metabolism; D015603:Keratinocytes / Q000166:cytology*; D007963:Leukocytes, Mononuclear / Q000378:metabolism; D011247:Pregnancy; D012867:Skin / Q000166:cytology*; D042461:Vascular Endothelial Growth Factor A; D014945:Wound Healing / Q000502:physiology
chemical_list: C576710:IDO1 protein, human; C576711:IDO2 protein, human; D050503:Indoleamine-Pyrrole 2,3,-Dioxygenase; D016209:Interleukin-8; D042461:Vascular Endothelial Growth Factor A; D016178:Granulocyte-Macrophage Colony-Stimulating Factor
references.citation: Nature. 2007 Feb 22;445(7130):874-80
references.pmid: 17314974
references.citation: Transpl Immunol. 2008 Feb;18(4):302-6
references.pmid: 18158115
references.citation: Transpl Int. 2007 Feb;20(2):118-27
references.pmid: 17239019
references.citation: Nature. 2008 May 15;453(7193):314-21
references.pmid: 18480812
references.citation: J Invest Dermatol. 2003 Jun;120(6):1130-7
references.pmid: 12787144
references.citation: FASEB J. 2010 Nov;24(11):4133-52
references.pmid: 20624931
references.citation: Burns. 2013 Mar;39(2):300-10
references.pmid: 22749444
references.citation: Wound Repair Regen. 2008 Sep-Oct;16(5):585-601
references.pmid: 19128254
references.citation: J Exp Med. 2002 Aug 19;196(4):447-57
references.pmid: 12186837
references.citation: J Invest Dermatol. 2007 May;127(5):1018-29
references.pmid: 17435787
references.citation: J Invest Dermatol. 1996 Dec;107(6):849-55
references.pmid: 8941673
references.citation: Clin Dermatol. 2005 Jul-Aug;23(4):413-8
references.pmid: 16023937
references.citation: Cell. 1990 Oct 19;63(2):425-35
references.pmid: 2145076
references.citation: Cell. 1993 May 21;73(4):713-24
references.pmid: 8500165
references.citation: Plast Reconstr Surg. 2010 Oct;126(4):1172-1180
references.pmid: 20885241
references.citation: J Biol Chem. 2002 Apr 5;277(14):12456-62
references.pmid: 11799124
references.citation: J Invest Dermatol. 2012 May;132(5):1501-5
references.pmid: 22297642
references.citation: Cell. 2000 Nov 22;103(5):745-55
references.pmid: 11114331
references.citation: J Invest Dermatol. 2007 May;127(5):998-1008
references.pmid: 17435785
references.citation: J Cell Biol. 1993 Jul;122(2):417-29
references.pmid: 8320264
references.citation: Burns. 2003 Dec;29(8):837-41
references.pmid: 14636761
references.citation: J Vasc Surg. 2009 Apr;49(4):1013-20
references.pmid: 19341889
references.citation: Tissue Eng Part B Rev. 2008 Mar;14(1):105-18
references.pmid: 18454637
references.citation: Arch Dermatol. 1998 Mar;134(3):293-300
references.pmid: 9521027
references.citation: Wound Repair Regen. 2010 Nov-Dec;18(6):614-23
references.pmid: 20955343
references.citation: Curr Opin Biotechnol. 2009 Oct;20(5):563-7
references.pmid: 19782559
references.citation: J Cell Biol. 1988 Feb;106(2):311-8
references.pmid: 2828381
references.citation: J Exp Med. 2002 Aug 19;196(4):459-68
references.pmid: 12186838
references.citation: Am J Transplant. 2004 May;4(5):698-704
references.pmid: 15084163
references.citation: J Invest Dermatol. 1992 Sep;99(3):294-8
references.pmid: 1512465
references.citation: FEBS Lett. 1992 Jul 6;305(3):241-3
references.pmid: 1299623
references.citation: Wound Repair Regen. 2003 Nov-Dec;11(6):411-8
references.pmid: 14617279
references.citation: J Investig Dermatol Symp Proc. 2006 Sep;11(1):66-72
references.pmid: 17069012
references.citation: Lancet. 2005 Sep 3-9;366(9488):840-2
references.pmid: 16139659
references.citation: Development. 1992 Jan;114(1):253-9
references.pmid: 1576963
references.citation: Ann Surg. 1994 Jan;219(1):65-72
references.pmid: 8297179
references.citation: PLoS One. 2012;7(7):e40951
references.pmid: 22911722
references.citation: Wound Repair Regen. 2012 Jul-Aug;20(4):580-91
references.pmid: 22712462
references.citation: N Engl J Med. 1984 Aug 16;311(7):448-51
references.pmid: 6379456
references.citation: Virchows Arch A Pathol Anat Histol. 1979 Mar 23;381(3):353-61
references.pmid: 155931
references.citation: J Exp Med. 1999 May 3;189(9):1363-72
references.pmid: 10224276
references.citation: Wound Repair Regen. 2007 Sep-Oct;15(5):708-17
references.pmid: 17971017
references.citation: Ann Surg. 1986 Oct;204(4):402-10
references.pmid: 3532970
references.citation: Science. 1998 Aug 21;281(5380):1191-3
references.pmid: 9712583
references.citation: Curr Biol. 2003 Jul 1;13(13):1122-8
references.pmid: 12842011
references.citation: J Cell Biol. 1982 Feb;92(2):584-8
references.pmid: 7061598
references.citation: Exp Gerontol. 2009 Mar;44(3):208-18
references.pmid: 19049860
references.citation: Blood. 2004 Jun 15;103(12):4619-21
references.pmid: 15001472
references.citation: Cell Growth Differ. 2000 Feb;11(2):111-21
references.pmid: 10714767
references.citation: J Invest Dermatol. 1992 May;98(5):805-8
references.pmid: 1569329
references.citation: Physiol Rev. 2003 Jul;83(3):835-70
references.pmid: 12843410