title: Fibroblasts show more potential as target cells than keratinocytes in COL7A1 gene therapy of dystrophic epidermolysis bullosa.
abstract: Dystrophic epidermolysis bullosa (DEB) is an inherited blistering skin disorder caused by mutations in the type VII collagen gene (COL7A1). Therapeutic introduction of COL7A1 into skin cells holds significant promise for the treatment of DEB. The purpose of this study was to establish an efficient retroviral transfer method for COL7A1 into DEB epidermal keratinocytes and dermal fibroblasts, and to determine which gene-transferred cells can most efficiently express collagen VII in the skin. We demonstrated that gene transfer using a combination of G protein of vesicular stomatitis virus-pseudotyped retroviral vector and retronectin introduced COL7A1 into keratinocytes and fibroblasts from a DEB patient with the lack of COL7A1 expression. Real-time polymerase chain reaction analysis of the normal human skin demonstrated that the quantity of COL7A1 expression in the epidermis was significantly higher than that in the dermis. Subsequently, we have produced skin grafts with the gene-transferred or untreated DEB keratinocytes and fibroblasts, and have transplanted them into nude rats. Interestingly, the series of skin graft experiments showed that the gene-transferred fibroblasts supplied higher amount of collagen VII to the new dermal-epidermal junction than the gene-transferred keratinocytes. An ultrastructural study revealed that collagen VII from gene-transferred cells formed proper anchoring fibrils. These results suggest that fibroblasts may be a better gene therapy target of DEB treatment than keratinocytes.
authors.affiliation: Department of Dermatology, Hokkaido University Graduate School of Medicine, Sapporo, Japan.
authors.forename: Maki
authors.initials: M
authors.lastname: Goto
authors.forename: Daisuke
authors.initials: D
authors.lastname: Sawamura
authors.forename: Kei
authors.initials: K
authors.lastname: Ito
authors.forename: Masataka
authors.initials: M
authors.lastname: Abe
authors.forename: Wataru
authors.initials: W
authors.lastname: Nishie
authors.forename: Kaori
authors.initials: K
authors.lastname: Sakai
authors.forename: Akihiko
authors.initials: A
authors.lastname: Shibaki
authors.forename: Masashi
authors.initials: M
authors.lastname: Akiyama
authors.forename: Hiroshi
authors.initials: H
authors.lastname: Shimizu
mesh_terms: D000818:Animals; D002478:Cells, Cultured; D024143:Collagen Type VII / Q000235:genetics* / Q000378:metabolism; D004817:Epidermis / Q000378:metabolism; D016108:Epidermolysis Bullosa Dystrophica / Q000628:therapy*; D005347:Fibroblasts / Q000378:metabolism / Q000637:transplantation*; D015316:Genetic Therapy / Q000379:methods*; D005822:Genetic Vectors / Q000235:genetics; D006801:Humans; D015603:Keratinocytes / Q000378:metabolism / Q000637:transplantation; D051379:Mice; D051381:Rats; D012190:Retroviridae / Q000235:genetics; D014162:Transfection
chemical_list: D024143:Collagen Type VII