The effect of nicotine on HMGB1 in oral epithelium

Objectives: Cigarette smoking alters overreacting inflammation followed by dental plaque infection, aggravating periodontitis. High mobility group box 1 (HMGB1) is a nuclear protein which not only acts as the transcriptional factor but also as an inflammatory mediator once it is released from cells. Although the level of HMGB1 in periodontal tissues is higher than tissues of healthy subjects, whether smoking affects the level of HMGB1 is unknown. In this study, effects of nicotine (NT) on the activation of HMGB1 and the effects of smoking on the expression of HMGB1 in gingival tissues were determined.

Methods: Normal oral keratinocytes (NOKs) were treated with different concentrations of NT for 24 hours and then stimulated with ATP (0.72mM) and lipopolysaccharides of Porphyromonas gingivalis (Pg-LPS, 1 μg/ml) for another 24 hours. The expression and the intracellular localization of HMGB1 in cells were determined by Western blot analyses and immunofluorescence, respectively. The release of HMGB1 into culture medium was determined by ELISA. Gingival tissues were collected from non-smokers without or with periodontitis or smokers with periodontitis for immunohistochemistry of HMGB1.
Results: NT alone barely affected the expression level and the intracellular localization of HMGB1 in NOKs. The expression levels of HMGB1 increased by stimulation of ATP and Pg-LPS was inhibited by pretreatment of NT. Translocation from nuclei to cytoplasm and release of HMGB1 stimulated by ATP and Pg-LPS were also inhibited by nicotine. In gingival tissues obtained from non-smokers with periodontitis, higher expression levels and less nuclear restriction of HMGB1 was noticed, compared to healthy subjects. In contrast, the expression level and the intracellular localization of HMGB1 in gingival tissues obtained from smokers with periodontitis were similar to healthy subjects.
Conclusions: NT was shown to inhibited the expression and release of HMGB1 in oral keratinocytes. This finding reflected changes of HMGB1 in gingival tissues in smokers.