A Fernández1,2, P Veloso1, J Astorga1, C Rodríguez2, V A Torres3, M Valdés4, M Garrido1, P J Gebicke-Haerter5,6, M Hernández1,7. 1. Laboratory of Periodontal Biology, Faculty of Dentistry, Universidad de Chile, Santiago, Chile. 2. Faculty of Dentistry, Universidad Andres Bello, Santiago, Chile. 3. Faculty of Dentistry, Universidad de Chile, Institute for Research in Dental Sciences, Santiago, Chile. 4. School of Public Health, Faculty of Medicine, Universidad de Chile, Santiago, Chile. 5. Program of Immunology, Faculty of Medicine, Institute of Biomedical Sciences, Universidad de Chile, Santiago, Chile. 6. Institute of Psychopharmacology, Faculty of Medicine, Central Institute of Mental Health, University of Heidelberg, Mannheim, Germany. 7. Department of Pathology and Oral Medicine, Faculty of Dentistry, Universidad de Chile, Santiago, Chile.
Abstract
AIM: To determine the methylation pattern of TLR2 gene promoter and its association with the transcriptional regulation of periapical inflammatory and angiogenic responses in symptomatic and asymptomatic forms of apical periodontitis. METHODOLOGY: In this cross-sectional study, apical lesions were obtained from volunteers with asymptomatic apical periodontitis (AAP) (n = 17) and symptomatic apical periodontitis (SAP) (n = 17) scheduled for tooth extraction, and both total RNA and DNA were extracted. DNA was bisulfite-treated, a region of CpG island within the TLR2 gene was amplified by qPCR and the products were sequenced. Additionally, the mRNA expression of TLR2, TLR4, IL-6, IL-12, TNFalpha, IL-23, IL-10, TGFbeta, VEGFA and CDH5 was analysed by qPCR. The data were analysed with chi-square tests, Mann-Whitney or unpaired t-tests, and Spearman´s correlation; variable adjustments were performed using multiple linear regression (P < 0.05). RESULTS: TLR2 depicted a hypomethylated DNA profile at the CpG island in SAP when compared with AAP, along with upregulated expression of TLR2, with pro-inflammatory cytokines IL-6 and IL-23, and the angiogenesis marker CDH5 (P < 0.05). TLR2 methylation percentage negatively correlated with mRNA levels of IL-23 and CDH5 in apical periodontitis. Lower methylation frequencies of single CpG dinucleotides -8 and -10 localized in close proximity to nuclear factor κB (NFκB) binding within the TLR2 promoter were identified in SAP versus AAP (P < 0.05). Finally, unmethylated -10 and -8 single sites demonstrated up-regulation of IL-23, IL-10 and CDH5 transcripts compared to their methylated counterparts (P < 0.05). CONCLUSIONS: TLR2 gene promoter hypomethylation was linked to transcriptional activity of pro-inflammatory cytokines and angiogenic markers in exacerbated periapical inflammation. Moreover, unmethylated single sites in close proximity to NFκB binding were involved in active transcription of IL-23, IL-10 and CDH5.
AIM: To determine the methylation pattern of TLR2 gene promoter and its association with the transcriptional regulation of periapical inflammatory and angiogenic responses in symptomatic and asymptomatic forms of apical periodontitis. METHODOLOGY: In this cross-sectional study, apical lesions were obtained from volunteers with asymptomatic apical periodontitis (AAP) (n = 17) and symptomatic apical periodontitis (SAP) (n = 17) scheduled for tooth extraction, and both total RNA and DNA were extracted. DNA was bisulfite-treated, a region of CpG island within the TLR2 gene was amplified by qPCR and the products were sequenced. Additionally, the mRNA expression of TLR2, TLR4, IL-6, IL-12, TNFalpha, IL-23, IL-10, TGFbeta, VEGFA and CDH5 was analysed by qPCR. The data were analysed with chi-square tests, Mann-Whitney or unpaired t-tests, and Spearman´s correlation; variable adjustments were performed using multiple linear regression (P < 0.05). RESULTS: TLR2 depicted a hypomethylated DNA profile at the CpG island in SAP when compared with AAP, along with upregulated expression of TLR2, with pro-inflammatory cytokines IL-6 and IL-23, and the angiogenesis marker CDH5 (P < 0.05). TLR2 methylation percentage negatively correlated with mRNA levels of IL-23 and CDH5 in apical periodontitis. Lower methylation frequencies of single CpG dinucleotides -8 and -10 localized in close proximity to nuclear factor κB (NFκB) binding within the TLR2 promoter were identified in SAP versus AAP (P < 0.05). Finally, unmethylated -10 and -8 single sites demonstrated up-regulation of IL-23, IL-10 and CDH5 transcripts compared to their methylated counterparts (P < 0.05). CONCLUSIONS: TLR2 gene promoter hypomethylation was linked to transcriptional activity of pro-inflammatory cytokines and angiogenic markers in exacerbated periapical inflammation. Moreover, unmethylated single sites in close proximity to NFκB binding were involved in active transcription of IL-23, IL-10 and CDH5.
Authors: C Schweitzer; M Garrido; R Paredes; C Stoore; M Reyes; R Bologna-Molina; A Fernández; Marcela Hernández Rios Journal: Clin Oral Investig Date: 2021-01-07 Impact factor: 3.573