No abstract

No abstract

No abstract

Background Tumor necrosis factor-alpha (TNF-α) plays a central role in the molecular pathogenesis of periodontal disease. However, the epigenetic regulation attributable to microbial and inflammatory signals at the biofilm gingival interface are poorly understood. In this study, we investigated the DNA methylation alteration within the TNFA promoter in human gingival biopsies from different stages of periodontal disease, and explored the regulatory mechanism of TNFA transcription by DNA methylation. Methods Gingival biopsies were harvested from 17 chronic periodontitis patients and 18 subjects with periodontal health. Another 11 subjects participated in an experimentally induced gingivitis study, and gingival biopsies were collected at the baseline, induction, and resolution phase. To confirm that TNFA promoter methylation modulated TNFA transcription we treated THP.1 cells with a DNA methyltransferase inhibitor, 5-aza-2-deoxycytidine and used a RAW 294.7 cell line transfected with a TNFA promoter-specific luciferase reporter system with or without methlyaiton, Results In gingival biopsies from subjects with severe chronic periodontitis two individual CpG sites within the TNFA promoter (at -163bp and -161bp) displayed increased methylation in periodontitis samples as compared to gingival health (16.1±5.1% vs. 11.0±4.6%, p=0.02, 19.8±4.1% vs. 15.4±3.6%, p=0.04, respectively). The methylation level at -163bp was inversely associated with the transcription level of TNFA (p=0.018). However, no significant difference in the TNFA promoter methylation pattern was observed in samples biopsied during the induction or resolution phase of experimentally induced gingivitis, which represented a reversible periodontal lesion. THP.1 cells treated with 5-aza-2-deoxycytidine demonstrated a time-dependent increase in TNFA messenger level. We also found that the luciferase activity decreased 2.6 fold in a construct containing an in vitro methylated TNFA promoter as compared to the unmethylated insert (p=0.03). Conclusion Although the biopsy samples represented a mixed cell population, the change in promoter methylation status in chronic periodontal disease suggested that DNA methylation may be an important regulatory mechanism in controlling TNFA transcriptional expression in disease.

No abstract

Levels of prostaglandin E 2 and the prostaglandin-endoperoxide synthase-2 (PTGS2, or COX-2) increase in actively progressing periodontal lesions, but decrease in chronic disease. We hypothesized that chronic inflammation is associated with altered DNA methylation levels within the PTGS2 promoter, with effects on COX-2 mRNA expression. PTGS2 promoter methylation levels from periodontally inflamed gingival biopsies showed a 5.06-fold increase as compared with non-inflamed samples (p = 0.03), and the odds of methylation in a CpG site in the inflamed gingival group is 4.46 times higher than in the same site in the non-inflamed group (p = 0.016). The level of methylation at −458 bp was inversely associated with transcriptional levels of PTGS2 (RT-PCR) (p = 0.01). Analysis of the data suggests that, in chronically inflamed tissues, there is a hypermethylation pattern of the PTGS2 promoter in association with a lower level of PTGS2 transcription, consistent with a dampening of COX-2 expression in chronic periodontitis. These findings suggest that the chronic persistence of the biofilm and inflammation may be associated with epigenetic changes in local tissues at the biofilm-gingival interface.

Aim The goal of this investigation was to determine whether epigenetic modifications in the IFNG promoter are associated with an increase of IFNG transcription in different stages of periodontal diseases. Materials and Methods DNA was extracted from gingival biopsy samples collected from 47 total sites from 47 different subjects: 23 periodontally healthy sites, 12 experimentally induced gingivitis sites and 12 chronic periodontitis sites. Levels of DNA methylation within the IFNG promoter containing six CpG dinucleotides were determined using pyrosequencing technology. Interferon gamma mRNA expression was analysed by quantitative polymerase chain reactions using isolated RNA from part of the biological samples mentioned above. Results The methylation level of all six analysed CpG sites within the IFNG promoter region in the periodontitis biopsies {52% [interquartile range, IQR (43.8%, 63%)]} was significantly lower than periodontally healthy samples {62% [IQR (51.3%, 74%)], p =0.007} and gingivitis biopsies {63% [IQR (55%, 74%)], p =0.02}. The transcriptional level of IFNG in periodontitis biopsies was 1.96-fold and significantly higher than tissues with periodontal health (p =0.04). Although the mRNA level from experimental gingivitis samples exhibited an 8.5-fold increase as compared with periodontally healthy samples, no significant methylation difference was observed in experimental gingivitis sample. Conclusions A hypomethylation profile within IFNG promoter region is related to an increase of IFNG transcription present in the chronic periodontitis biopsies, while such an increase of IFNG in experimentally induced gingivitis seems independent of promoter methylation alteration.

No abstract