An improved method of human keratinocyte culture from skin explants: cell expansion is linked to markers of activated progenitor cells.
Exp Dermatol
; 18(8): 720-6, 2009 Aug.
Article
em En
| MEDLINE
| ID: mdl-19558495
ABSTRACT
Human keratinocyte primary cultures are commonly established by tissue dissociation and often rely on feeder cell supports and culture medium that is not defined. Further, contamination by unwanted fibroblasts can be problematic. Here, we developed a skin explant method for growing primary keratinocytes that was rapid, simple, and reliably generated keratinocyte cultures free of fibroblast contamination. The process capitalized on the observation that fibroblasts migrate out of adult skin explants later than epidermal cells, allowing the early harvesting of keratinocytes by trypsinization. When grown subsequently in defined medium in the absence of feeder cells, the explant-derived cells grew rapidly and could be cultured for multiple passages. Immunofluorescence microscopy revealed that a high percentage of cells harvested from the explant outgrowths expressed K15, while very few expressed the differentiation marker K10. Cells that were stained while migrating out from explants strongly expressed markers associated with progenitor cells, including p63, K15 and CD133, and displayed intense K6 expression, indicative of activated keratinocytes in wound-healing epidermis. By replenishing the explants with fresh medium after harvesting, further epidermal outgrowths could be obtained, offering the possibility of greatly increased keratinocyte yields for clinical applications.
Texto completo:
1
Coleções:
01-internacional
Base de dados:
MEDLINE
Assunto principal:
Pele
/
Células-Tronco
/
Queratinócitos
/
Transplante de Pele
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Técnicas de Cultura de Células
/
Dermatologia
Limite:
Adult
/
Female
/
Humans
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Male
/
Middle aged
Idioma:
En
Ano de publicação:
2009
Tipo de documento:
Article