Acute effects of cigarette smoke on three-dimensional cultures of normal human oral mucosa

Context: Human oral mucosa is the combustion chamber of cigarette, but scanty evidence is available about the early smoke effects. Objective: The present work aimed at evaluating from a morphological point of view whole smoke early effects on epithelial intercellular adhesion and keratinocyte terminal differentiation in a three-dimensional model of human oral mucosa. Materials and methods: Biopsies of keratinized oral mucosa of healthy nonsmoking women (n = 5) were collected. After culturing in a Transwell system, one fragment of each biopsy was exposed to the smoke of one single cigarette; the remnant represented the internal control. The distribution of epithelial differentiation markers (keratin-10, K10, and keratin-14, K14, for suprabasal and basal cells respectively), desmosomes (desmoglein-1, desmoglein-3), tight junctions (occludin), adherens junctions (E-cadherin, beta-catenin), and apoptotic cells (p53, caspase 3) were evaluated by immunofluorescence. Results: Quantitative analysis of K14 immunolabeling revealed an overexpression in the suprabasal layers as early as 3 h after smoke exposure, without impairment of the epithelial junctional apparatus and apoptosis induction. Discussion and conclusion: These results suggested that the first significant response to cigarette smoke came from the basal and suprabasal layers of the human oral epithelium. The considered model maintained the three-dimensional arrangement of the human mucosa in the oral cavity and mimicked the inhalation/exhalation cycle during the exposure to cigarette smoke, offering a good possibility to extrapolate the reported observations to humans.