title: Development of autologous human dermal-epidermal composites based on sterilized human allodermis for clinical use.
abstract: The aim of this study was to identify a sterilization technique for the preparation of human allodermis which could be used as a dermal component in wound healing and as the dermal base for production of dermal-epidermal composites for one-stage grafting in patients. We report that it is possible to produce dermal-epidermal composites which perform well in vitro and in vivo using a standard ethylene oxide sterilization methodology. Prevention of ethylene oxide-induced damage to the dermis was achieved using gentle dehydration of the skin prior to ethylene oxide sterilization. The issue of whether viable fibroblasts are required for composite production was examined in comparative studies using glycerol vs. ethylene oxide sterilized dermis. Where good collagen IV retention was achieved following preparation of acellular de-epidermized dermis there was no advantage to having fibroblasts present in vitro or in vivo; however, where collagen IV retention was poor or where keratinocytes were initially expanded in culture then there was a significant advantage to introducing fibroblasts to the composites during their preparative 10-day period in vitro. The requirement for fibroblasts became less evident when composites were grafted on to nude mice. In conclusion, we report a protocol for the successful sterilization of human allodermis to achieve an acellular dermis with good retention of collagen IV. This acellular dermis would be appropriate for clinical use as a dermal replacement material. It can also be used for the production of dermal-epidermal composites using autologous keratinocytes (with or without fibroblasts).
authors.affiliation: University Division of Clinical Sciences, Section of Medicine, NorthernGeneral Hospital, Sheffield, UK.
authors.forename: K H
authors.initials: KH
authors.lastname: Chakrabarty
authors.forename: R A
authors.initials: RA
authors.lastname: Dawson
authors.forename: P
authors.initials: P
authors.lastname: Harris
authors.forename: C
authors.initials: C
authors.lastname: Layton
authors.forename: M
authors.initials: M
authors.lastname: Babu
authors.forename: L
authors.initials: L
authors.lastname: Gould
authors.forename: J
authors.initials: J
authors.lastname: Phillips
authors.forename: I
authors.initials: I
authors.lastname: Leigh
authors.forename: C
authors.initials: C
authors.lastname: Green
authors.forename: E
authors.initials: E
authors.lastname: Freedlander
authors.forename: S
authors.initials: S
authors.lastname: Mac Neil
mesh_terms: D000328:Adult; D000818:Animals; D018929:Cell Culture Techniques; D003890:Desiccation; D005027:Ethylene Oxide; D005260:Female; D005347:Fibroblasts / Q000166:cytology; D005990:Glycerol; D006801:Humans; D015603:Keratinocytes / Q000166:cytology; D051379:Mice; D008819:Mice, Nude; D012867:Skin / Q000033:anatomy & histology; D016038:Skin Transplantation / Q000379:methods*; D017768:Skin, Artificial; D013242:Sterilization / Q000379:methods*; D014182:Transplantation, Autologous
chemical_list: D005027:Ethylene Oxide; D005990:Glycerol