Effect of inflammatory cytokines on DNA methylation and demethylation

Objective: To determine the effects of inflammatory cytokines and mediators on expression of DNMT and TETenzymes in human gingival fibroblasts isolated from patients with periodontitis as well as normal and transformed cell lines.Methods: Expression levels of DNA methylatransferase (DNMT1 and 3a) and demethylase (TET1) were examined by RT-PCR in cells treated for up to 24 hours with IL-1, IL-6 or prostaglandin E2. The role of PGE2 in IL-1 suppression of DNMT3a expression was assessed by treating cells with IL-1 in the presence and absence of NS-398. Global levels of DNA methylation following treatment with IL-1 were measured in an ELISA-like assay.Results: Treatment of HGF with IL-1 caused a three-fold increase in levels of DNMT1 mRNA over 24 hours, but a two-fold decrease in DNMT3a mRNA in the same samples. Global levels of 5mC were increased in HGF treated with IL-1 for 72 hours relative to untreated controls. PGE2 caused a dose-dependent decrease in levels of DNMT3asimilar to IL-1, but also slightly decreased levels of DNMT1. Conclusion: These data are consistent with the hypothesis that exposure to inflammatory cytokines or PGE2 alters levels and/or activity of enzymes involved in DNA methylation, resulting in global and possibly gene-specific changes which can lead to changes in cellular characteristics over time. Further experiments will be aimed at testing the effects of these cytokines on expression of DNMT and TET enzymes in normal and transformedelucidating mechanisms involved in cytokine regulation of these enzymes

Interleukin 1β and Prostaglandin E2 Affect Expression of DNA Methylating and Demethylating Enzymes in Human Gingival Fibroblasts

Periodontitis is a common chronic inflammatory condition that results in increased levels of inflammatory cytokines and inflammatory mediators. In addition to oral disease and tooth loss, it also causes low-grade systemic inflammation that contributes to development of systemic conditions including cardiovascular disease, pre-term birth, diabetes and cancer. Chronic inflammation is associated with epigenetic change, and it has been suggested that such changes can alter cell phenotypes in ways that contribute to both ongoing inflammation and development of associated pathologies. Here we show that exposure of human gingival fibroblasts to IL-1β increases expression of maintenance methyltransferase DNMT1 but decreases expression of de novo methyltransferase DNMT3a and the demethylating enzyme TET1, while exposure to PGE2 decreases expression of all three enzymes. IL-1β and PGE2 both affect global levels of DNA methylation and hydroxymethylation, as well as methylation of some specific CpG in inflammation-associated genes. The effects of IL-1β are independent of its ability to induce production of PGE2, and the effects of PGE2 on DNMT3a expression are mediated by the EP4 receptor. The finding that exposure of fibroblasts to IL-1β and PGE2 can result in altered expression of DNA methylating/demethylating enzymes and in changing patterns of DNA methylation suggests a mechanism through which inflammatory mediators might contribute to the increased risk of carcinogenesis associated with inflammation

Interleukin 1 and Prostaglandin E2 Affect Expression of DNA Methyltransferases and Global DNA Methylation Levels in Human Gingival Fibroblasts

Objective: To determine the effects of inflammatory cytokines and mediators on expression of DNMT and TET enzymes in human gingival fibroblasts isolated from patients with periodontitis. Methods: Expression of DNA methyltransferase (DNMT1 and 3a) and demethylase (TET1) was examined by RT-PCR. Protein levels of DNMT3a were determined by ELISA. Global levels of DNA methylation and hydroxymethylation were measured in ELISA-like assays. Results: Treatment of HGF with IL-1 increased levels of DNMT1 , but decreased in DNMT3a mRNA in the same samples. Global levels of 5mC were increased in HGF treated with IL-1 for 72 hours relative to untreated controls. PGE2 caused a dose-dependent decrease in levels of DNMT3a similar to IL-1, but also decreased levels of DNMT1 and TET1. DNMT3a protein levels were decreased by IL-1 and PGE2 in HGF. Inhibition of IL-1 induced PGE2 production with NS398 partially reversed the inhibitory effects of PGE2 on DNMT3a and TET expression. Use of EP receptor agonists showed that EP4 stimulation resulted in decreased expression of DNMT3a. Global levels of 5hmC were not significantly changed in response to treatment with IL-1 or PGE2. Conclusion: IL-1 and PGE2 alter expression of DNA methylating and demethylating enzymes, and result in changes in global DNA methylation. Such changes have the potential to alter gene expression patterns over time and could be involved in the predisposition to cancer associated with chronic inflammation. IL-1 inhibition of DNMT3a and TET1 expression in HGF is likely mediated at least in part through increased production of PGE2, which exerts its inhibitory activity through the EP4 receptor