Cytokine expression in response to root canal infection in gnotobiotic mice.

AIM: To examine cytokine expression profiles during periapical lesion development in response to synergetic human pathogens in a gnotobiotic mouse model. METHODOLOGY: Human strains of Fusobacterium nucleatum and Peptostreptococcus prevotii were inoculated into the root canals of germ-free mice in either mono- or bi-association. Animals were killed 7 and 14 days after infection, and periapical tissues were collected. mRNA expression of the cytokines IFN-γ, TNF-α, Receptor activator of nuclear factor kappa-B ligand (RANKL), IL-10, IL-4 and transforming growth factor ß (TGF-ß) was assessed using real-time PCR. Levene's test was used to assess the equality of variance of the data, whereas a t-test for independent samples was used to evaluate the significance of the differences between groups (P < 0.05). RESULTS: The mRNA expression of IFN-γ and TNF-α was up-regulated by F. nucleatum during the acute (day 7) and chronic phase (day 14) of periapical lesion development. However, in bi-infection the expression of IFN-γ and TNF-α were effectively absent at both time-points. RANKL mRNA expression was down-regulated during dual infection at the chronic phase. As IL-4 expression was similar at both time-points, IL-4 does not appear to be involved in the periapical response to these bacterial strains. IL-10 was up-regulated during the chronic phase by mono-infection with either F. nucleatum or P. prevotii. Dual infection increased TGF-ß mRNA expression on day 7, which paralleled the decrease in IFN-γ and TNF-α mRNA levels at the same time-point. F. nucleatum increased TGF-ß mRNA expression during the chronic phase. CONCLUSION: Cytokine profiles depend on the nature of the bacterial challenge. Both TGF-ß and IL-10 appeared to be regulating the proinflammatory cytokine responses at both time-points of the periapical immune response.

Microbiota of deciduous endodontic infections analysed by MDA and Checkerboard DNA-DNA hybridization.

AIMS: To evaluate the microbiota of endodontic infections in deciduous teeth by Checkerboard DNA-DNA hybridization after uniform amplification of DNA in samples by multiple displacement amplification (MDA). METHODOLOGY: Forty samples from the root canal system of deciduous teeth exhibiting pulp necrosis with or without radiographically detectable periradicular/interradicular bone resorption were collected and 32 were analysed, with three individuals contributing two samples; these were MDA-amplified and analysed by Checkerboard DNA-DNA hybridization for levels of 83 bacterial taxa. Two outcome measures were used: the percentage of teeth colonized by each species and the mean proportion of each bacterial taxon present across all samples. RESULTS: The mean amount of DNA in the samples prior to amplification was 5.2 (±4.7) ng and 6.1 (±2.3) µg after MDA. The mean number of species detected per sample was 19 (±4) (range: 3-66) to the nearest whole number. The most prevalent taxa were Prevotella intermedia (96.9%), Neisseria mucosa (65.6%), Prevotella nigrescens (56.2%) and Tannerella forsythia (56.2%). Aggregatibacter (Haemophilus) aphrophilus and Helicobacter pylori were not detected. P. intermedia (10%), Prevotella tannerae (7%) and Prevotella nigrescens (4.3%) presented the highest mean proportions of the target species averaged across the positive samples. CONCLUSION: Root canals of infected deciduous teeth had a diverse bacterial population. Prevotella sp. were commonly found with P. intermedia, Prevotella tannerae and Prevotella nigrescens amongst the most prominent species detected.