Interleukin-8 Gene Promoter Polymorphism (rs4073) May Contribute to Chronic Periodontitis

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Background: The proinflammatory chemokine interleukin (IL)-8 is important in the regulation of the inflammatory response. Analyses of the single nucleotide polymorphism (SNP) reference sequence (rs) 4073 showed that the A allele upregulated IL-8 levels after stimulation with lipopolysaccharides. We investigated the association of the SNP rs4073 with chronic periodontitis. Methods: Genotyping was performed by a standard polymerase chain reaction restriction fragment length polymorphism assay in 289 genomic DNA samples of healthy control subjects and patients with chronic periodontitis; analyses were adjusted by multivariate logistic regression modeling. A real-time polymerase chain reaction performance was used to detect levels of the IL-8 mRNA. Results: The analysis pointed to a statistically significant association of chronic periodontitis with the heterozygous TA genotype (P = 0.001); the results showed an increase in the frequency of the A allele in the diseased group (36% in the control group versus 48% in the periodontitis group). The higher levels of the IL-8 mRNA were found in the periodontitis group, mainly in individuals who presented the TA genotype (P = 0.03). Conclusion: The SNP rs4073 was associated with chronic periodontitis in non-smoker Brazilian subjects because the frequency of the A allele was higher in the disease group than in the control group, and the TA genotype was associated with increased levels of IL-8 mRNA transcripts. J Periodontol 2011;82:893-899.826893899Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Coordination and Specialization of Superior Education, Sao Paulo, SP, BrazilNational Council of Research, Sao Paulo, SP, BrazilNational Institutes of Health, Bethesda, Maryland [K99/R00-DE018954]Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)FAPESP [07/02488-0]National Institutes of Health, Bethesda, Maryland [K99/R00-DE018954

DNA Methylation Status of the IL8 Gene Promoter in Aggressive Periodontitis

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Background: Studies evaluating the methylation status of cytokine genes may have relevance for inflammatory diseases in which the expression of some cytokines is altered, such as periodontitis. This study observes the DNA methylation status in the interleukin-8 (IL8) gene promoter in cells of the oral epithelium of subjects affected by generalized aggressive periodontitis (AgP) and compares it to those of control subjects. Methods: Genomic DNA from epithelial oral cells of 37 generalized AgP patients and 37 controls were purified and modified by sodium bisulphite. Modified DNA was submitted by methylation-specific polymerase chain reaction, electrophoresed on 10% polyacrylamide gels, and stained. Results: Subjects who presented generalized AgP have a higher frequency of hypomethylation of the IL8 gene promoter in oral epithelium cells than that of controls (86.5% in the generalized AgP group versus 62% in the control group; P = 0.016; chi(2) test). Conclusions: A marked hypomethylated status is found in the oral epithelial cells of subjects presenting with generalized AgP, compared to controls, in the promoter region of the IL8 gene. This hypomethylated status may reflect a generalized condition of oral epithelial cells, including gingival epithelium, because gingival epithelial cells were also collected during mouthwash use. J Periodontol 2010;81:1336-1341.81913361341Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Coordination and Specialization of Superior Education, Sao Paulo, SP, BrazilNational Council of Research, Sao Paulo, SP, BrazilFundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)FAPESP [07/02488-0

TLR2 and TLR4 gene promoter methylation status during chronic periodontitis

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)Aim: The objective of this study was to analyse the status of DNA methylation in the promoter region of the toll-like receptor (TLR) 2 and TLR4 genes in gingival tissue samples from healthy subjects, smokers and non-smokers affected by chronic periodontitis. Material and Methods: Genomic DNA and total RNA were purified from gingival tissue using the TRIZOL reagent protocol. Genomic DNA was then digested by methylation-sensitive restriction enzymes, amplified by polymerase chain reaction (PCR), electrophoresed on a 10% polyacrylamide gel and stained using SYBR Gold. Real-time PCR was also performed to verify the transcript levels. Results: The CpG dinucleotides analysed were observed to be unmethylated in the majority of DNA samples of the three groups and statistical differences were not found among groups (p > 0.05). However, a trend towards methylation was observed in the TLR2 HhaI site in the samples of the periodontitis non-smoker groups. In fact, the analysis of all CpG sites together shows which complete methylation is observed in the shortest level in the samples of periodontitis non-smoker group. The analysis of transcript levels demonstrated no difference among groups (p > 0.05). Conclusion: The results demonstrated major unmethylation of the TLR4 gene promoter in all groups. However, the results for the TLR2 gene promoter are inconclusive; this gene was found as a mosaic of methylated and unmethylated DNA in the majority of samples of the three groups and we also observed a trend towards the DNA methylation of CpG sites recognized by the HhaI enzyme.3811975983Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)FAPESP [FAPESP 07/02488-0