A modified culture system for epidermal cells for grafting purposes: an in vitro and in vivo study.
A fully differentiated epithelium mimicking the features of native epidermis was obtained in vitro by culturing human or porcine epidermal keratinocytes on polyester filter substrate at the air-liquid interface. In addition, after 2 weeks of culture, hemidesmosome-like structures were formed along the basal area of the plasma membrane of the basal cells at the cell-filter interface. When grafted onto full-thickness skin wounds in pigs, the take of cell sheets detached from the filter with dispase was significantly higher (about 70%) in comparison to mechanically detached keratinocytes (about 15%). With dispase-treated keratinocytes alone, basement membrane formation took place within 7 days postgrafting as judged from the presence of a lamina lucida and positive staining for type IV collagen. Also, numerous hemidesmosomes and anchoring fibrils were observed at the basal cell-"neodermis" interface. The fully differentiated epidermis, generated by culturing keratinocytes at the air-liquid interface and detached from the substrate by dispase-treatment, is less fragile and easier to handle than epidermal autografts obtained by conventional culturing methods. Detachment by a short dispase-treatment appeared in our hands the only method for successful and complete epithelial regeneration in full-thickness wounds.
van Dorp
A G
AG
Department of Dermatology, Leiden University Medical Center, Leiden, The Netherlands.
Verhoeven
M C
MC
Nat-Van Der Meij
T H
TH
Koerten
H K
HK
Ponec
M
M
D000818:Animals; D018929:Cell Culture Techniques / Q000379:methods*; D017690:Cell Transplantation; D002478:Cells, Cultured; D000078404:Epidermal Cells; D004817:Epidermis / Q000737:chemistry / Q000637:transplantation* / Q000648:ultrastructure; D005347:Fibroblasts / Q000737:chemistry / Q000166:cytology / Q000637:transplantation / Q000648:ultrastructure; D006801:Humans; D007150:Immunohistochemistry; D015603:Keratinocytes / Q000737:chemistry / Q000166:cytology / Q000637:transplantation / Q000648:ultrastructure; D013552:Swine