DNA Methylations in Epithelial Rest of Malassez Stimulated With LPS

Objectives: DNA methylation is a major type of epigenetics that are often altered by environmental factors. Epigenetic research has been mainly focused on malignant and premalignant lesions. Recently, it has been shown that epigenetic modifications may cause other common diseases. Epigenetic alterations have also recently been implicated in periodontal diseases. Lipopolysaccharide (LPS) in the dental plaque plays a major pathogenic role. Alteration of DNA methylation can be caused by the stimulation with environmental factor for prolonged periods. In the present study, we examined whether DNA methylations of p14, p15, p16, p53, IL-6 and IL-8 in Epithelial Rest of Malassez (ERM) were altered by the stimulation with LPS derived from P. gingivalis for prolonged periods.
Methods: Epithelial cells derived from porcine epithelial rests of Malassez (ERM). The ERM cells were cultured in DMEM containing 10% FBS. The culture was repeated alternating 3 days with LPS derived from P. gingivalis (WAKO, 1µg/ml) and 3 days without LPS for 1month. No stimulation with LPS was used as the control. Expression levels of p14, p15, p16, p53, IL-6 and IL-8 mRNAs were evaluated by quantitative RT-PCR. The levels of methylation were evaluated by the methylation- specific PCR. Results were compared by Mann-Whitney U test with P-value <0.05 accepted as statistically significant.
Results: The expression levels of IL-6 and IL-8 mRNAs were significantly lower by the stimulation with LPS than the controls. No significant differences were observed in the expression levels of p14, p15, p16 or p53 between the stimulation and the controls. According to the methylation-specific PCR, the levels of methylation of IL-6 were significantly higher than the controls.
Conclusions: The results indicate that the prolonged stimulation with LPS derived from P. gingivalis may induce hypermethylation of IL-6.