Nicotine-enhanced epithelial differentiation in reconstructed human oral mucosa in vitro

Oral mucosal keratinocytes represent the cells that first encounter tobacco components. Therefore, tobacco-induced abnormal alteration of the mucosal keratinocytes may contribute to the development of oral white lesions. Nicotine is an ingredient of all tobacco products and pharmacologically the most active component of tobacco smoke. To clarify the effects of nicotine on the keratinization of oral mucosal and epidermal keratinocytes, we reconstructed artificial buccal mucosal and skin equivalents using keratinocytes and fibroblasts from noncornifying buccal mucosa and adult foreskin, respectively. The effect of nicotine on keratinization was assessed with morphology, immunohistochemistry and immunoblotting. Long-term treatment with nicotine for 2 weeks enhanced in a dose-dependent manner the expression of differentiation-specific proteins of oral mucosal keratinocytes on living oral mucosal equivalent and epidermal keratinocytes on living skin equivalent, respectively. The effect of nicotine on the cell viability was measured by the 3-(4, 5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide assay. Oral mucosal keratinocytes showed a higher resistance to nicotine toxicity than epidermal keratinocytes. Our results suggest that nicotine stimulates differentiation of both mucosal and epidermal keratinocytes, and this nicotine-induced abnormal differentiation may be associated with the development of oral white lesions.