A new skin equivalent: keratinocytes proliferated and differentiated on collagen sponge containing fibroblasts.

Three types of artificial skin containing keratinocytic components were prepared and tested for comparison. Keratinocytes were cultured on the artificial skin dermis (collagen sponge) by the air-liquid interface culture method. In order to create continuous keratinocytic layers on the artificial skin dermis, pores of its uppermost layer were filled beforehand with type I collagen gel, Matrigel, or fibroblasts. A band of keratinocytes consisting of two to six cell layers was formed on collagen gel-coated artificial skin dermis. On Matrigel-coated artificial skin dermis, keratinocytes were piled up into about 20 cell layers, but cell differentiation was incomplete; cornified material was not fully developed, and the proportion of cuboidal cells was very high compared with normal epidermis. Keratinocytes formed continuous layers on the fibroblasts-artificial skin dermis complex without gel coating. Keratinocytes proliferated well and differentiated properly on this matrix, and their histologic appearance was similar to that of normal epidermis. Thus keratinocytes cultured on the fibroblast-artificial skin dermis complex seem to be a good skin equivalent.