Background: Although mice have long served as an animal model for periodontitis, information on the composition of their indigenous oral microbiota is limited. The aim of the current study was to characterize mouse oral bacterial flora by applying extensive parallel pyrosequencing using the latest model pyrosequencer, a Roche/454 Genome Sequencer FLX Titanium. In addition, the effect of Toll-like receptor (TLR) 2 deficiency on oral microbiota was evaluated. Results: Eight oral bacterial communities of wild-type (n = 4) and TLR2 knock-out (n = 4) C57BL/6 mice were characterized by analyzing 80,046 reads of 16S rRNA genes obtained by pyrosequencing. Excluding the PCR primers, the average length of each sequencing product was 443 bp. The average species richness of the murine oral bacterial communities was estimated to be about 200, but the communities were dominated by only two main phyla and several species. Therefore, the bacterial communities were relatively simple. The bacterial composition of the murine oral microbiota was significantly different from that of humans, and the lack of TLR2 had a negligible effect on the murine oral microbiota. Conclusion: Pyrosequencing using the Roche/454 FLX Titanium successfully characterized mouse oral bacterial communities. The relatively simple oral bacterial communities of mice were not affected by TLR2 deficiency. These findings will provide a basis for future studies on the role of periodontal pathogens in the murine model of periodontitis.

Host immune response is known to contribute to the progression of periodontitis, and alveolar bone destruction in periodontitis is associated with enhanced osteoclast activity. Therefore, we evaluated the roles of activated lymphocyte subsets in osteoclastogenesis. Osteoclast precursors were co‐cultured with activated lymphocytes (B, CD4⁺ T, CD8⁺ T) in the presence of either macrophage colony‐stimulating factor (M‐CSF) alone or M‐CSF plus soluble receptor activator of NF‐κB ligand (sRANKL), and subsequent differentiation into active osteoclasts was evaluatedby a resorption assay. The activated B and CD4⁺ cells, but not CD8⁺ T cells, induced osteoclast differentiation in the presence of M‐CSF alone. In the presence of M‐CSF and sRANKL, B cells induced the formation of small but highly active osteoclasts and increased resorption, while CD8⁺ T cells profoundly suppressed osteoclastogenesis. Co‐culture using an insert wellor supernatant suggested that both B and CD8⁺ T cells acted on osteoclasts mostly via soluble proteins. Activated B cells expressed many osteoclastogenic factors including RANKL, TNF‐α, IL‐6, MIP‐1α, and MCP‐3. CD8⁺ T cells expressed a substantial amount of osteoprotegerin (OPG) along with RANKL. However, blocking antibody to OPG did not reverse the suppression by CD8⁺ T cells, suggesting that other factor(s) are involved. Taken together, activated B cells promoted osteoclastogenesis, while CD8⁺ T cells inhibited the osteoclast formation via direct interaction. The results imply the importance of lymphocyte subpopulations in the development of periodontitis.

Periodontitis is chronic inflammation of the periodontium caused by the host's inflammatory response to plaque biofilm, which destroys tooth‐supporting soft and hard tissues. Periodontitis is a complex disease that involves interactions among three main features – microbial challenge, the host immune response, and environmental and genetic risk factors – in its pathogenesis. Although periodontitis has been regarded as the result of hyperimmune or hyperinflammatory responses to plaque bacteria, recent studies indicate that periodontal pathogens are rather poor activators and/or suppressors of the host immune response. This raises the question of how periodontal pathogens cause inflammation. To resolve this issue, in the present review we propose that bacterial invasion into gingival tissue is a key event in the initiation of periodontitis and that the persistence of these bacteria within host tissue results in chronic inflammation. In support of this hypothesis, we present the ways in which microbial, environmental and genetic risk factors contribute to bacterial invasion. It is hoped that the current model will instigate active discussion and new research to complete the puzzle of this complex disease process.

Some synthetic lipopeptides, in addition to native lipoproteins derived from both Gram-negative bacteria and mycoplasmas, are known to activate TLR2 (Toll-like receptor 2). However, the native lipoproteins inherent to Gram-positive bacteria, which function as TLR2 ligands, have not been characterized. Here, we have purified a native lipoprotein to homogeneity from Staphylococcus aureus to study as a native TLR2 ligand. The purified 33-kDa lipoprotein was capable of stimulating TLR2 and was identified as a triacylated SitC lipoprotein, which belongs to a family of ATP binding cluster (ABC) transporter substrate-binding proteins. Analyses of the SitC-mediated production of cytokine using mouse peritoneal macrophages revealed that the SitC protein (3 nm) induced the production of tumor necrosis factor-α and interleukin-6. Moreover, analysis of knock-out mice showed that SitC required TLR2 and MyD88, but not TLR1 or TLR6, for the induction of cytokines. In addition to the S. aureus SitC lipoprotein, we purified two other native ABC transporter substrate-binding lipoproteins from Bacillus subtilis and Micrococcus luteus, which were both shown to stimulate TLR2. These results demonstrate that S. aureus SitC lipoprotein is triacylated and that the ABC transporter substrate-binding lipoproteins of Gram-positive bacteria function as native ligands for TLR2.