In vitroengineering of complete autologous oral mucosa equivalents: characterization of a novel scaffold

Abstract

Restoration of oral mucosa defects by means of in vitro-cultured equivalents has become a valid alternative in the field of oral and periodontics surgery. Although different techniques have been described, none has been able to provide an equivalent with an autologous scaffold for the epithelium. The purpose of this study was to obtain complete autologous oral mucosa equivalents (CAOME) using the patient's own fibroblasts and plasma and to characterize these equivalents both morphologically and immunohistochemically. We acquired cell types (keratinocytes and fibroblasts) from the same mucosal samples, which were taken from healthy patients who underwent oral surgery. To construct the CAOME, a small sample of blood was obtained from the patient and subsequently processed to obtain a fibrin glue scaffold. All CAOME thus obtained were stained using the standard hematoxylin and eosin method to study their morphological characteristics. To establish the type of cells in the epithelial layer, CAOME were stained with pancytokeratin AE1/AE3, cytokeratins 5/6 and 13, p-63 and Ki-67. Finally, laminin 5 and collagen IV were used to reveal the presence of a basal membrane. The CAOME featured a monolayer of cube-shaped epithelial cells similar to that found on the basal layer of the oral mucosa. Close to the epithelial layer lay the fibrin and fibroblasts-embedded scaffold. The CAOME was positive to pancytokeratin AE1/AE3, cytokeratin 5/6 and p-63. No reaction was found to cytokeratin 13 and Ki-67. There was staining to laminin 5 but not to collagen IV. It is possible to engineer a CAOME with an epithelium of basal-like and immature keratinocytes, which could potentially reconstruct in vivo loss of tissue.