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Epigenetic regulation of inflammation in localized aggressive periodontitis.

ABSTRACT: We have previously demonstrated a Toll-like receptor (TLR)-mediated hyper-responsive phenotype in our cohort of localized aggressive periodontitis (LAP) individuals. However, mechanisms related to this phenotype are still not clear in the literature. The objective of this cross-sectional study is to examine the role of epigenetic regulation, specifically DNA methylation status of genes in the TLR pathway in this cohort. Peripheral blood was collected from 20 LAP patients and 20 healthy unrelated controls. Whole blood was stimulated with 1 ?l (100 ng/?l) of purified Escherichia coli lipopolysaccharide (LPS) for 24 h and cyto/chemokines in the supernatants analyzed by Luminex multiplex assays. Genomic DNA extracted from buffy coats prepared from a second tube of whole blood was used for DNA methylation analysis by pyrosequencing of seven TLR signaling genes (FADD, MAP3K7, MYD88, IL6R, PPARA, IRAK1BP1, RIPK2).Significant differences in the methylation status were observed at specific CpG positions in LAP patients compared to healthy controls and interestingly also between severe and moderate LAP. Specifically, subjects with moderate LAP presented hypermethylation of both the upregulating (MAP3K7, MYD88, IL6R, and RIPK2) and downregulating (FADD, IRAK, and PPARA) genes, while severe LAP presented hypomethylation of these genes. Further analysis on CpG sites with significant differences in methylation status correlates with an increased pro-inflammatory cytokine profile for LAP patients.Our findings suggest that epigenetic modifications of genes in the TLR pathway may orchestrate the thresholds for balancing induction and prevention of tissue destruction during the course of disease, and thus differ significantly at different stages of the disease, where moderate LAP shows hypermethylation and severe LAP shows hypomethylation of several genes.https://clinicaltrials.gov, NCT01330719.

SUBMITTER: Shaddox LM

PROVIDER: S-EPMC5581417 | biostudies-literature | 2017

REPOSITORIES: biostudies-literature

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Epigenetic regulation of inflammation in localized aggressive periodontitis.

Shaddox L M LM Mullersman A F AF Huang H H Wallet S M SM Langaee T T Aukhil I I

Clinical epigenetics 20170902

<h4>Background</h4>We have previously demonstrated a Toll-like receptor (TLR)-mediated hyper-responsive phenotype in our cohort of localized aggressive periodontitis (LAP) individuals. However, mechanisms related to this phenotype are still not clear in the literature. The objective of this cross-sectional study is to examine the role of epigenetic regulation, specifically DNA methylation status of genes in the TLR pathway in this cohort. Peripheral blood was collected from 20 LAP patients and 2 ...[more]

PMID: 28883894

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Project description:Localized aggressive periodontitis (LAP) patients possess a systemic hyperinflammatory response after lipopolysaccharide stimulation. However, the levels of inflammatory and bone biomarkers in plasma, as well as possible associations between local and plasma biomarkers, are unknown in LAP. This cross-sectional study aimed to characterize gingival crevicular fluid (GCF) and plasma biomarker profiles in LAP patients, their healthy siblings (HS), and healthy unrelated controls (HC). Fifty-eight LAP subjects, 33 HS, and 49 HC (African Americans, aged 5 to 25 y) were included. Following collection of clinical parameters with GCF and plasma samples, levels of 16 inflammatory and bone resorption biomarkers were determined with Milliplex. Univariate and correlation analyses were performed among all clinical and laboratorial parameters. Discriminant analyses were used to investigate groups of biomarkers discriminating LAP from HS and HC in GCF and plasma. GCF levels of multiple cytokines and chemokines and RANKL:OPG ratio (receptor activator of nuclear factor kappa-B ligand:osteoprotegerin) were higher in LAP disease, most of which positively correlated with probing depth and attachment loss of sampled sites. A group of IL-12p40, IL-6, IL-12p70, IL-2, and MIP-1? discriminated LAP diseased sites from twheir healthy sites, as well as from HS and HC healthy sites. In plasma, only RANKL levels were increased in LAP versus controls, which positively correlated with the percentage of affected sites and deep/bleeding sites. A plasma inflammatory profile including MIP-1?, IL-8, IL-10, and INF-? could significantly discriminate LAP patients from HS and HC. No correlations were found between GCF and plasma levels of biomarkers. In conclusion, an inflammatory profile including groups of specific biomarkers in GCF and plasma may significantly discriminate LAP from healthy individuals. The hyperinflammatory response previously found in the peripheral blood of LAP patients is dependent on lipopolysaccharide stimulation, apparently resulting mostly in local tissue destruction and changes in biomarker profile, with a slight influence in the systemic inflammatory profile (ClinicalTrials.gov NCT01330719). Knowledge Transfer Statement: The results of this study can be possibly used by clinicians in the future as diagnostic tools for localized aggressive periodontitis. Thus, in the future, with proper consideration of cost, patient preference, chair-side feasibility and ultimately further studies validating the role of GCF markers for disease progression and response to treatment, this information could lead to more appropriate therapeutic decisions and the development of preventive approaches for susceptible patients.

Project description:AimThe purpose of this study was to test for the role of the P2X7 receptor in localized aggressive periodontitis (LAP).MethodsPeripheral blood was obtained from 95 subjects with LAP and 76 healthy unrelated controls (HUCs). Three P2RX7 single-nucleotide polymorphisms (rs1718119, rs2230911, and rs3751143) were genotyped from these subjects, and their peripheral blood samples were stimulated with lipopolysaccharide (LPS) from Escherichia coli and tested for inflammatory markers. The 3 P2RX7 single-nucleotide polymorphisms were in found to be in perfect linkage disequilibrium, and a total of 4 haplotypes and 9 diplotypes were identified among all subjects. For both subject populations, the 9 diplotypes were grouped into 4 functional groups and tested for association with subject inflammatory response. To specifically study the effects of extrinsic activation of the P2X7 receptor in LAP, peripheral blood samples from were stimulated under 3 treatments: LPS, LPS + ATP, and LPS +ATP+ P2X7 selective inhibitor. The effects of these treatments on P2X7 receptor activity were measured through Luminex protein assay. Last, to test whether receptor stimulation was related to P2RX7 expression, relative mRNA levels of P2RX7 were quantified with real-time quantitative polymerase chain reaction.ResultsSeveral associations between the P2RX7 diplotypes and LPS-stimulated blood chemokine/cytokine levels were found between the LAP and HUC populations (P < 0.05). P2X7 activation resulted in statistically significant differences in IL-1β and IL-12p40 concentrations for both subject populations. The relative P2RX7 mRNA levels increased significantly after addition of its inhibitor for both LAP and HUC populations.ConclusionsThis study detected an association between P2RX7 functional diplotypes and in vitro immune response of whole blood from subjects with LAP. In addition, we found that inhibition of the activated P2X7 receptor leads to increased P2RX7 mRNA levels, suggesting a feedback loop ( ClinicalTrials.gov NCT01330719).Knowledge transfer statementThe results of this study suggest that P2RX7 functional diplotypes are associated with LAP and their in vitro immune response to bacteria. Ongoing studies to uncover the mechanistic link between P2RX7 and LAP phenotypes could lead to the development of preventive approaches for susceptible subjects.

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Epigenetic regulation of inflammation in localized aggressive periodontitis.

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Epigenetic regulation of inflammation in localized aggressive periodontitis.

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