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Objective: Human periodontal ligament fibroblasts (HPDLFs) play a crucial role in protecting against oral bacteria in periapical tissue. Toll-like receptors (TLRs) and nucleotide-binding oligomerization domain (NOD) are two major forms of innate immune sensors that recognize microbial pathogens and initiate pro-inflammatory signalling. Tumour necrosis factor receptor-associated factor 6 (TRAF6) is an adapter protein for TLR-mediated nuclear factor-κB (NF-κB) signalling pathway activation that induces the production of pro-inflammatory cytokines. The aim of this study was to investigate the expression of TLR2, TLR4, NOD1, and NOD2 in HPDLFs. We also investigated the expression of TRAF6 and pro-inflammatory cytokines induced by the activation of TLRs and NODs., Methods: The expression of TLR2, TLR4, NOD1, and NOD2 was measured by reverse transcription-polymerase chain reaction (RT-PCR), flow cytometry, and immunostaining. HPDLFs were stimulated with TLR and NOD agonists. Then, the expression of TRAF6 was measured by real-time PCR and western blot. Concentrations of IL-1β, IL-6, and IL-8 in the culture supernatants were determined by enzyme-linked immunosorbent assay (ELISA). Finally, by using small interfering RNA (siRNA) for TRAF6, we analysed the production of IL-1β, IL-6, and IL-8 in HPDLFs upon stimulation with TLRs and NODs agonists., Results: We found clear mRNA and protein expression of TLR2, TLR4, NOD1, and NOD2 in HPDLFs. The expression levels of TRAF6 and pro-inflammatory cytokines (IL-1β, IL-6, and IL-8) were markedly up-regulated upon the activation of TLRs and NODs. Furthermore, the co-activation of TLRs and NODs had synergistic effect on the production of TRAF6 and pro-inflammatory cytokines. We also found TRAF6 suppression resulted in reduced IL-1β, IL-6, and IL-8 expression upon TLR and NOD agonists challenge., Conclusion: These findings indicated that TLR2, TLR4, NOD1, and NOD2 are functional receptors in HPDLFs during innate immune responses to invading bacteria, and a combination of signalling through TLRs and NODs leads to the synergistic enhancement of inflammatory reactions in HPDLFs. In addition, TLR and NOD signalling involving TRAF6 contribute to inflammatory responses in HPDLFs., (Copyright © 2011 Elsevier Ltd. All rights reserved.)

Objective: Periodontitis is a group of inflammatory diseases caused by microorganisms. Porphyromonas gingivalis, a gram-negative bacteria, is strongly associated with the onset of periodontitis. Tumor necrosis factor (TNF) receptor-associated factor 6 (TRAF6) represents an important target in the regulation of many disease processes, including immunity, inflammation, and osteoporosis. The aim of this study was to investigate the role of TRAF6 for inflammatory response in P gingivalis-infected human periodontal ligament cells (HPDLCs)., Method and Materials: HPDLCs were stimulated with 1 x 108 CFU/mL P gingivalis, or 10 ug/mL P gingivalis lipopolysaccharide (LPS), separately in the absence or presence of small interfering RNA (siRNA) for TRAF6. The expression of TRAF6 was examined by real-time polymerase chain reaction and Western blot analysis. Concentrations of IL-1B, IL-6, and IL-8 in the culture supernatants were determined by enzyme-linked immunosorbent assay (ELISA)., Results: In this study, we found that both P gingivalis and its LPS treatment increased the expression of TRAF6 and proinflammatory cytokine production in HPDLCs. In addition, we used siRNA for TRAF6, and the inhibition of TRAF6 expression reduced the production of proinflammatory cytokines in HPDLCs stimulated with P gingivalis and its LPS., Conclusion: The results suggested that TRAF6 may be a key molecule to control proinflammatory cytokine production induced by P gingivalis and its LPS. TRAF6 suppression may inhibit inflammatory responses in HPDLCs infected by P gingivalis and its LPS.

- Vertical root fractures (VRFs) often occur in endodontically treated teeth and in patients older than 40 years of age. However, VRFs in teeth without endodontic treatment are relatively uncommon. VRFs are difficult to diagnose as the symptoms are non-specific or often delayed. The most common radiographic findings are thickening of the periodontal ligament, deep, localized, vertical bone loss, and localized periradicular bone loss. The explicit feature for detecting VRFs is direct visualization of a radiolucent fracture line on radiographs. However, the fracture line can be difficult to directly visualize in conventional diagnostic methods such as periapical radiographs. If unrecognized, VRFs can lead to frustration and inappropriate endodontic treatment. The two cases reported here demonstrate that the use of cone beam computed tomography (CBCT) successfully diagnoses VRFs on teeth without representative clinical and periapical radiographic findings. The clear fracture line can be discerned from the images of CBCT. Thus, CBCT imaging is useful in rapid diagnosis of VRFs and designing of further treatment. [ABSTRACT FROM AUTHOR]