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Updated: Mar 12, 2026
Isolation and Culture of Primary Oral Keratinocytes from the Adult Mouse Palate
Published on: September 24, 2021
Development and Characterization of In Vitro Human Oral Mucosal Equivalents Derived from Immortalized Oral
Luke R Jennings1, Helen E Colley1, Jane Ong2
11 School of Clinical Dentistry, University of Sheffield , Sheffield, United Kingdom .
Abstract:
Tissue-engineered oral mucosal equivalents (OME) are being increasingly used to measure toxicity, drug delivery, and to model oral diseases. Current OME mainly comprise normal oral keratinocytes (NOK) cultured on top of a normal oral fibroblasts-containing matrix. However, the commercial supply of NOK is limited, restricting widespread use of these mucosal models. In addition, NOK suffer from poor longevity and donor-to-donor variability. Therefore, we constructed, characterized, and tested the functionality of OME based on commercial TERT2-immortalized oral keratinocytes (FNB6) to produce a more readily available alternative to NOK-based OME. FNB6 OME cultured at an air-to-liquid interface for 14 days exhibited expression of differentiation markers cytokeratin 13 in the suprabasal layers and cytokeratin 14 in basal layer of the epithelium. Proliferating cells were restricted to the basal epithelium, and there was immuno-positive expression of E-cadherin confirming the presence of established cell-to-cell contacts. The histology and expression of these structural markers paralleled those observed in the normal oral mucosa and NOK-based models. On stimulation with TNFα and IL-1, FNB6 OME displayed a similar global gene expression profile to NOK-based OME, with increased expression of many common pro-inflammatory molecules such as chemokines (CXCL8), cytokines (IL-6), and adhesion molecules (ICAM-1) when analyzed by gene array and quantitative PCR. Similarly, pathway analysis showed that both FNB6 and NOK models initiated similar intracellular signaling on stimulation. Gene expression in FNB6 OME was more consistent than NOK-based OME that suffered from donor variation in response to stimuli. Mucosal equivalents based on immortalized FNB6 cells are accessible, reproducible and will provide an alternative animal experimental system for studying mucosal drug delivery systems, host-pathogen interactions, and drug-induced toxicity.
