Introduction:
Cigarette smoking is one of the major risk factors in the pathogenesis of periodontal disease. Despite numerous reports on the general toxic effects of oral smokeless tobacco very few studies have assessed the effect of smokeless tobacco on the viability and inflammatory response of gingival epithelial cells.

Material and Methods:
In this study a reference smokeless tobacco (CORESTA CRP2) was used to stimulate gingival epithelial cells. Once the gingival epithelial cells became confluent, the growth medium were conditioned with aqueous extract of CORESTA CRP2 for 3 and 6 hours. Cell viability was measured by alamar blue assay, and cell culture supernatant was examined for the presence of human interlukin-1beta (IL-1beta) using sandwich enzyme-linked immunosorbant assay (ELISA).

Results:
Cells treated with smokeless tobacco reference product resulted in decreased viability compared to non stimulated group (P<0.05). In addition, the smokeless tobacco extract augmented the production of IL-1beta in a time -dependent manner (P< 0.05). The micrographs showed that smokeless tobacco extract resulted in a significant change in cell appearance, from the cells being in close contact to becoming separated from each other.

Conclusions:
Epithelial cells are the first line of defense against pathogens in the oral cavity. The results suggest that smokeless tobacco not only inhibits the growth of epithelial cells but also induce the generation of inflammatory cytokines which leads to smokeless tobacco-exacerbated disease.