Análise microbiológica e imunológica de lesões refratárias ao tratamento endondôntico

Abstract

Introduction: Failure in endodontic treatment is often caused by the persistence of microorganisms in the root canal after therapy. When treatment fails, an immune response develops that is characterized by an extensive network of immunologic mechanisms that lead to the production of cytokines and chemokines.Aims: To combine MDA and Checkerboard DNADNA hybridization to qualitatively and quantitatively evaluate the microbiota of infections refractory to endodontic treatment, and to determine the relative mRNA expression of IFN-, TNF-, IL-1, IL-17A, IL-10, and MCP-1 in these cases.Methodology: Subjects were 40 patients presenting periapical lesions refractory to endodontic treatment (experimental group) and 20 patients presenting healthy teeth with pulp vitality (control group). Microbiologic samples were taken by scraping or filing the root canal walls with a #10 K-type hand file, MDA-amplified and analysed by Checkerboard DNADNA hybridization for levels of 107 bacterial taxa. The taxa were divided in three distinct microbial populations, depending on their mean proportion (%DNA probe counts ± SEM) as follow: dominant (>4%), sub dominant (>2 to 4%) and residual (< 2%) populations. Significance of differences was sought using the Mann Whitney test for microbiologic findings. Immunologic samples were taken experimental and control groups. Three paper points passing through the root apex (2 mm) were used to collect the samples. Total RNA was extracted from each sample, cDNA was synthesized, and quantitative PCR analysis was performed. The Mann-Whitney test was used to determine the statistical significance of immunologic findings (p < 0.05).Results: The dominant taxa were C. diphtheria (7.35 ± 1.22), S. constellatus (6.85 ± 1.15), P. gingivalis (6.19 ± 2.20), A. adjacens (5.94 ± 0.99), and P. denticola (5.79 ± 0.97). Among sub dominant population were S. mutans (4.12 ± 0.61), A. georgiae (4.02 ± 0.60), H. pylori (3.11 ± 0.30), D. peneumosintes (2.25 ± 0.54), and E. corrodens (2.16 ± 0.36). In the residual population, E. coli (0.05 ± 0.01), and L. acidophilus (0.02 ± 0.01) showed the lowest mean proportions. E. faecalis was detected in low mean proportion (0.55 ± 0.27).Significant differences in the levels of IFN-, TNF-, IL-17A, and MCP-1 mRNA expression were observed in cases refractory to endodontic treatment as compared to the control group. The expression of IL-1 mRNA was not significantly different between the groups. Expression of IL-10 mRNA was insignificant in both the experimental and control groups. Conclusions: The microbiota of infections refractory to endodontic treatment is far more complex than previously shown and, despite features of E. faecalis that allow it to survive and persist in treated root canals, this species is only a small part of this microbiota. Significantly increased expression of TNF-, IFN-, IL-17A and MCP-1 mRNA was observed in the periapical immune response in cases of endodontic failure. These results suggest that a pro-inflammatory cytokine profile predominates in these types of dental lesions.