Uniform expression of alcohol dehydrogenase 3 in epithelia regenerated with cultured normal, immortalised and malignant human oral keratinocytes

The human oral epithelium is a target for damage from the inhalation of for maldehyde. However, most experimental studies on this chemical have relied on laboratory animals that are obligatory nose breathers, including rats an d mice. Therefore, in vitro model systems that mimic the structure of the h uman oral epithelium and which retain normal tissue-specific metabolic comp etence are desirable. Based on the established role of alcohol dehydrogenas e 3 (ADH3), also known as glutathione-dependent formaldehyde dehydrogenase, as the primary enzyme catalysing the detoxification of formaldehyde, the a im of this study was to investigate the expression of ADH3 in organotypic e pithelia regenerated with normal (NOK), immortalised (SVpgC2a) and malignan t (SqCC/Y1) human oral keratinocytes. Organotypic epithelia, usually consis ting of 5-10 cell layers, were produced at the air-liquid interface of coll agen gels containing human oral fibroblasts, after culture for 10 days in a standardised serum-free medium. Immunochemical staining demonstrated unifo rm expression of ADH3 in these organotypic epithelia, as well as in the epi thelial cells of oral tissue. The specificity of the ADH3 antiserum was asc ertained from the complete neutralisation of the immunochemical reaction wi th purified ADH3 protein. Assessment of the staining intensities indicated that the expression levels were similar among the regenerated epithelia. Fu rthermore, the regenerated epithelia showed similar ADH3 expression to the epithelium in oral tissue. Therefore, a tissue-like expression pattern for ADH3 can he generated from the culture of various oral keratinocyte lines i n an organotypic state. Similar expression levels among the various cell li nes indicate the preservation of ADH3 during malignant transformation, and therefore that NOR, SVpgC2a and SqCC/Y1 represent functional models for in vitro studies of formaldehyde metabolism in human oral mucosa.