The EpiOralTM tissue model consists of normal human keratinocytes differentiated to form a multilayered tissue. The phenotype is analogous to human oral epithelium in vivo providing the opportunity for in vitro irritancy testing on human tissue. This study evaluated the effects of potential snus ingredients as irritants using the MTT tissue viability assay as a surrogate for irritancy. Conditions used were provided by the MatTek Corporation with modifications. Substances were applied topically at 0.3-5000 µg/ml for both 1 and 20h in duplicate. Positive controls, Triton X-100 (1%) and sodium dodecyl sulphate (SDS, 1%), and the negative control, DMSO (0.5%), were used. MTT conversion to formazan, as a measure of tissue viability, was assessed by measuring OD570. Under our experimental conditions, irritancy was defined as a reduction in tissue viability of =25%, relative to negative controls. The tables below show control data and results for the tested compounds:
The MTT viability assay proved to be a reliable and reproducible method to assess EpiOralTM tissue viability with consistent positive and negative control data. Future work will accumulate data for a range of ingredients to collate a database of tissue cytotoxicites (irritancy) or maximum concentrations without an effect.