Recapitulation of oral mucosal tissues in long-term organotypic culture.
To test the influence of fibroblasts on epithelial morphology and expression of keratinocyte proteins and barrier lipids, we bioengineered homotypic and heterotypic oral mucosae and skin using cultured adult human cells. Fibroblasts were allowed to modify collagen type I gels for 2 weeks before keratinocytes were added. The organotypic cultures were then grown at the air-liquid interface for 4 weeks. In homotypic combinations, epithelial morphology and protein expression closely mimicked those in vivo. In heterotypic combinations, the morphology resembled that in vivo and keratinocytes expressed their typical markers, except when skin keratinocytes were recombined with alveolar fibroblasts; they expressed K19, K4, and K13, which is similar to oral mucosal epithelia rather than to the epidermis. Morphologically, the stratum corneum layers were typical for the epithelial tissues. Grafting the bioengineered cultures to the backs of Nude mice did not change the results, suggesting that our findings are not merely a culture phenomenon. Lipid profiles of the homotypic combinations mimicked the profiles found in the normal epithelial tissues, except that the engineered alveolar epithelium expressed more ceramide 2 than that in vivo. In the heterotypic combinations, keratinocytes appeared to control the lipid profile, except in the combination of skin keratinocytes with alveolar fibroblasts, wherein the ceramide profile appeared to be partly that of alveolar epithelium and partly that of epidermis. These results suggest that cultured adult fibroblasts and keratinocytes are sufficient to recapitulate graftable oral tissues, and, except for alveolar fibroblasts, the type of fibroblast had little influence on keratinocyte differentiation.
Chinnathambi
Sathivel
S
Department of Anatomy and Cell Biology, University of Iowa, Iowa City, Iowa 52242, USA.
Tomanek-Chalkley
Ann
A
Ludwig
Nicholas
N
King
Elizabeth
E
DeWaard
Rose
R
Johnson
Georgia
G
Wertz
Philip W
PW
Bickenbach
Jackie R
JR
D000328:Adult; D000818:Animals; D002454:Cell Differentiation; D004817:Epidermis / Q000378:metabolism / Q000637:transplantation / Q000648:ultrastructure; D005347:Fibroblasts / Q000166:cytology / Q000378:metabolism / Q000648:ultrastructure; D006801:Humans; D015603:Keratinocytes / Q000166:cytology / Q000378:metabolism / Q000648:ultrastructure; D007633:Keratins / Q000032:analysis; D051379:Mice; D008819:Mice, Nude; D009061:Mouth Mucosa / Q000166:cytology / Q000378:metabolism* / Q000637:transplantation / Q000648:ultrastructure; D009924:Organ Culture Techniques; D013997:Time Factors; D023822:Tissue Engineering; D014183:Transplantation, Heterologous; D016042:Transplantation, Heterotopic
D007633:Keratins