DNA methylation profiles of immune response-related genes in apical periodontitis.

AIM: To investigate the DNA methylation profiles of immune response-related genes in apical periodontitis (AP) lesions. METHODOLOGY: The methylation profiles on the cytosine-phosphate-guanine (CpG) regions of 22 gene promoters involved in inflammation and autoimmunity were assessed in 60 human AP lesions and 24 healthy periodontal ligaments (controls) using a pathway-specific real-time polymerase chain reaction array (EpiTect® Methyl Signature PCR Array Human Inflammatory Response). Differentially methylated genes were subsequently assessed for their mRNA expression. Data analyses (One-way anova, Tukey's multiple comparisons tests and Mann-Whitney tests) were performed using GraphPad Prism 6 software. P values ≤ 0.05 were considered statistically significant. RESULTS: Significant DNA hypermethylation was observed for CXCL3 and FADD gene promoters in AP lesions when compared to control tissues (P < 0.001) and among other genes (P < 0.05). In contrast, IL12B and IL4R were associated with significant hypomethylation in comparison to other genes (P < 0.05). IL12B, IL4R, CXCL3 and FADD had differential mRNA expression in AP lesions and controls (P < 0.001). CONCLUSIONS: Differential methylation profiles of immune response-related genes, such as FADD, CXCL3, IL12B and IL4R, may have an influence on individual AP susceptibility and patient treatment outcomes, through their potential contributions to altered expression of disease-relevant genes. Methylation and/or genetic variations in additional genes may also contribute to the dynamics of AP development and should be considered in future studies.

Effect of the combination of several irrigants on dentine surface properties, adsorption of chlorhexidine and adhesion of microorganisms to dentine.

AIM: To investigate the effects of combinations of several irrigants on the roughness and wettability of dentine, adhesion of Enterococcus faecalis and Candida albicans and adsorption of chlorhexidine (CHX) to the dentine. METHODOLOGY: Bovine dentine samples were prepared and their surface roughness standardized. The samples were distributed in groups (n = 10) and subjected to one of the following irrigation protocols: G1 - saline solution; G2 - sodium hypochlorite (NaOCl); G3 - NaOCl + ethylenediaminetetraacetic acid (EDTA); G4 - NaOCl + peracetic acid (PAA); G5 - NaOCl + 1-hydroxyethylidene-1,1-diphosphonic acid (HEDP); G6 - NaOCl + EDTA + CHX; G7 - NaOCl + PAA + CHX; G8 - NaOCl + HEDP + CHX; and G9 - mixture of NaOCl + HEDP. After treatments, roughness and wettability were measured. In order to evaluate the adhesion of microorganisms to dentine, new dentine samples were prepared and after 2 h of contact with the microorganisms, were analysed using a confocal laser scanning microscope and the number of microorganisms adhering to the surfaces were determined. Absorption spectra were collected by attenuated total reflectance of Fourier-transform infrared spectroscopy before and after immersion of other dentine samples in each solution of G6, G7 and G8 and in a solution of 2% CHX at various time intervals. The areas of the band associated with CHX with the peak at 1492 cm-1 were calculated between 1479 and 1500 cm-1 of the spectral range. The data obtained in all experiments were subjected to one-way ANOVA (α < 0.05). The values of the CHX band were also subjected to one-way repeated measures ANOVA (α < 0.05). RESULTS: Saline solution, NaOCl, HEDP and CHX did not alter the roughness of the dentine (P > 0.05), whilst EDTA and PAA did (P < 0.05). Dentine surface wettability increased after the use of all irrigants compared to saline solution (P < 0.05), with HEDP causing the greatest increases (P < 0.05). The adhesion of E. faecalis was favoured on surfaces treated with only saline solution and NaOCl, and on samples that had decalcifying agents as the final irrigant (P < 0.05). The adhesion of C. albicans was highest on surfaces treated with only saline solution and on surfaces that had NaOCl used as the last irrigant (P < 0.05). The use of CHX as a final irrigant reduced the adhesion of both microorganisms. The roughness and wettability did not influence the adhesion of the microorganisms tested. The adsorption of CHX to the dentine was significant after 1 min of immersion of the mineralized samples in the irrigant (P < 0.05), and the use of chelating agents prior to CHX potentiated this adsorption. CONCLUSIONS: The irrigation solutions had a variable effect on the properties of dentine, on the adhesion of E. faecalis and C. albicans and the adsorption of CHX to the dentine surface.

Expression levels of the receptor activator of NF-κB ligand and osteoprotegerin and the number of gram-negative bacteria in symptomatic and asymptomatic periapical lesions.

OBJECTIVE: The study aimed to verify the potential correlation between the detected amount of gram-negative bacteria and the radiographic sizes of the lesions in patients with symptomatic and asymptomatic apical periodontitis. Furthermore, to evaluate whether the expression of receptor activator of NF-κB ligand (RANKL) and osteoprotegerin (OPG) and the RANKL/OPG ratio are differentially regulated in both groups. DESIGN: Twenty patients with periapical lesions were divided into two groups: symptomatic (SYM) n=10 and asymptomatic (ASYM) n=10. After periapical surgery, the lesions were collected and processed for histological examination, and immunohistochemistry. The percentage of RANKL- and OPG-immunopositive areas relative to the total area of the microscopic field was calculated. For gram staining, the number of gram-negative cells per microscopic field was assessed. The radiographs of each patient were processed and measured. The Student's t-test and the Pearson correlation coefficient were performed. RESULTS: The SYM group showed a significantly higher number of gram-negative cells (p=0.007) when compared to the ASYM group. A higher number of gram-negative bacteria occurred more frequently in larger periapical lesions and the SYM group (p=0.03). The expression for RANKL and OPG and the RANKL/OPG ratio were not significantly different between the groups. There was a significant positive correlation between the number of bacteria and OPG levels in the SYM group (p=0.01). CONCLUSION: The number of bacteria seems to influence the symptoms and the radiographic size of a periapical lesion. Gram-negative bacteria may play an important role in OPG activity in the SYM group.