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Objective: Tissue stem cells in dental pulp are assumed to possess differentiation potentials similar to mesenchymal stem cells (MSCs). The aim of this in vitro study is to examine the differentiation potentials of mouse dental pulp stem cells (DPSCs) and develop the appropriate differentiation assay systems for skeletal myogenic differentiation of these cells., Methods: Dental pulps were extracted from mandible sections of C57/BL6 mice, and adherent dental pulp cells were isolated in culture. These cells were cultured in osteogenic or adipogenic induction medium to induce osteogenic and adipogenic differentiation. On the other hand, the skeletal myogenic differentiation potential of these cells was investigated using different conditions, such as serum-free medium, Myod1 overexpression, or 5-Aza-2'-deoxycytidine (5-Aza) treatment for DNA demethylation. Muscle-specific transcriptional factor expression was evaluated by RT-PCR, and myotube formation and myosin heavy chain expression were evaluated by phase-contrast microscopy and immunofluorescence staining, respectively., Results: The adherent dental pulp cells exhibited a proliferative capacity and they showed osteogenic and adipogenic differentiation as seen in previous studies. Although the expression of Myod1 mRNA and myotube formation was not detected in serum-free conditions, the forced expression of Myod1 up-regulated the expression of Myogenin and Pax7 mRNA. However, myotube formation was not confirmed. Interestingly, myosin heavy chain expression and myotube formation were observed following 5-Aza treatment of these cells., Conclusions: These results demonstrated that mouse DPSCs possess MSC-like differentiation potential. DNA demethylation induced by 5-Aza treatment resulted in the skeletal muscle differentiation in mouse DPSCs, suggesting that DNA demethylation might trigger this differential induction of mouse DPSCs., (Copyright 2010 Elsevier Ltd. All rights reserved.)

Proteases produced by Porphyromonas gingivalis are believed to contribute to the pathogenesis of periodontal diseases. Here the cytotoxic effects of a purified preparation of a P. gingivalis protease with trypsin-like specificity were tested on human gingival fibroblasts in vitro. The active protease induced apoptotic cell death in the fibroblasts, as indicated by DNA fragmentation and the expression of 7A6 antigen. Thus, the production of proteases by periodontopathic bacteria could be an important factor in the induction of apoptosis of host cells in the aetiology of periodontal diseases.

The lipopolysaccharides (LPS) of Porphyromonas gingivalis are implicated in the initiation and development of periodontal diseases. However, the mechanisms underlying P. gingivalis LPS-mediated periodontal destruction are still unknown. Here, it was found that P. gingivalis LPS activates human gingival fibroblasts (HGF) to release interleukin 6 (IL-6) via CD14. Flow-cytometric analysis showed that HGFs bind to fluorescein-isothiocyanate (FITC)-labelled LPS, and express CD14 on their surfaces. The binding of FITC LPS was competitively suppressed by unlabelled synthetic lipid A as well as by LPS. LPS-induced IL-6 production was inhibited by anti-CD14 monoclonal antibody in a dose-dependent manner. The binding of FITC LPS to HGF was abrogated by anti-CD14 monoclonal antibody. Engagement of LPS initiated the protein tyrosine phosphorylation of several intracellular proteins including extracellular signal-regulated kinase (ERK) 1 and 2, and these events were suppressed by the anti-CD14 monoclonal. These results suggest that CD14 is a cell surface binding site for LPS and is involved in the LPS-mediated activation of HGF.

Serum IgG, IgM, and IgA antibody levels to extracts of rat dental plaque and five oral bacteria (Haemophilus actinomycetemcomitans Y-4, Bacteroides gingivalis 381, Bact. intermedius ATCC 25261, Capnocytophaga sp. M-12, Eikenella corrodens ODU) were determined by ELISA. In addition, the presence of rat dental plaque and oral bacterial components in the inflamed gingival tissue was studied using immunofluorescence techniques. Serum and gingival tissue samples were obtained from ODU plaque-susceptible and plaque-resistant rats. In several susceptible rats, IgG, IgM, and IgA antibodies against dental plaque and oral bacteria were detected. There was a correlation between the levels of IgG antibody to dental plaque and pocket probing depth, but not between pocket probing depth and the levels of IgM and IgA. Furthermore, components of rat dental plaque and oral bacteria were detected in the inflamed gingival tissue.

The severity of gingivitis in relation to abnormal macrophage chemotaxis and superoxide (O2-) production was investigated in rats with naturally occurring gingivitis (plaque-susceptible rats). Macrophage chemotaxis was measured by the membrane filter method; zymosan-activated serum was used as a chemo-attractant. O2- production was measured by the reduction of cytochrome c method. Opsonized zymosan, phorbol myristate acetate and calcium ionophore A23187 were used as stimulants. The macrophage chemotaxis and O2- production in the plaque-susceptible rats with deep pockets were significantly lower than those in the susceptible rats with shallow pockets or the normal group. In addition, there was a negative correlation between the severity of gingivitis in plaque-susceptible rats and macrophage chemotaxis or O2- production. These findings suggest that impairement of the host defence mechanisms due to the reduction in macrophage chemotaxis and O2- production is related to the progression of periodontal disease in plaque-susceptible rats.