Development and characterization of a tissue-engineered human oral mucosa equivalent produced in a serum-free culture system

A problem maxillofacial surgeons face is a lack of sufficient autogenous or al mucosa for reconstruction of the oral cavity. Split-thickness or oral mu cosa grafts require more than one surgical procedure and can result in dono r site morbidity. Skin has disadvantages of adnexal structures and a differ ent keratinization pattern than oral mucosa. In this study, we successfully assembled, ex vivo, a human oral mucosa equivalent, consisting of epiderma l and dermal components, in a defined, essential-fatty-acid-deficient, seru m-free culture medium without a feeder layer, that could be used for intra- oral grafting in humans. Autogenous oral keratinocytes were seeded onto a c adaveric dermis, AlloDerm(TM). The oral mucosa equivalent was cultured at a n air-liquid interface for 2 wks. The resulting equivalent had a well-strat ified parakeratinized epithelial layer similar to native oral keratinized m ucosa. Expression of differentiation markers, filaggrin and cytokeratin 10/ 13, suggested a premature keratinized state. The presence of proliferation markers, proliferating cell nuclear antigen (PCNA) and Ki-67, suggested a s tate of hyperproliferation. Fatty acid composition of the equivalent was si milar to that of in vitro cultured oral keratinocytes but differed from the that of in vivo native tissue, showing a lower content of 18:2 and 20:4, a nd a higher content of 16:1 and 18:1 fatty acids, respectively. The keratin ocytes of the equivalent appeared to be in a more active and proliferative state than native keratinized mucosa. The dynamic nature of the cell popula tion on the oral mucosa equivalent may be beneficial for intra-oral graftin g procedures and for transfection of the keratinocytes.