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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 obtained 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 methylation. 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. It was also found that the luciferase activity decreased 2.6 fold in a construct containing an in vitro methylated TNFA promoter when 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 periodontal disease.
Journal of Periodontology 01/2013; · 2.60 Impact Factor
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ABSTRACT: 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.
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.
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.
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.
Journal Of Clinical Periodontology 11/2010; 37(11):953-61. · 3.00 Impact Factor
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ABSTRACT: Although human dental pulp stem cells isolated from healthy teeth have been extensively characterized, it is unknown whether stem cells also exist in clinically compromised teeth with irreversible pulpitis. Here we explored whether cells retrieved from clinically compromised dental pulp have stem cell-like properties.
Pulp cells were isolated from healthy teeth (control group) and from teeth with clinically diagnosed irreversible pulpitis (diseased group). Cell proliferation, stem cell marker STRO-1 expression, and cell odonto-osteogenic differentiation competence were compared.
Cells from the diseased group demonstrated decreased colony formation capacity and a slightly decreased cell proliferation rate, but they had similar STRO-1 expression and exhibited a similar percentage of positive ex vivo osteogenic induction and dentin sialophosphoprotein expression from STRO-1-enriched pulp cells.
Our study provides preliminary evidence that clinically compromised dental pulp might contain putative cells with certain stem cell properties. Further characterization of these cells will provide insight regarding whether they could serve as a source of endogenous multipotent cells in tissue regeneration-based dental pulp therapy.
Journal of endodontics 05/2010; 36(5):820-5. · 2.95 Impact Factor
