Colony-stimulating factor-1 (CSF-1) is an essential regulator of the differentiation, proliferation and survival of macrophage lineage cells including bone-resorbing osteoclasts. We have demonstrated that vascular endothelial growth factor (VEGF), a known angiogenic factor, can act as a substitute for CSF-1 function in osteoclastogenesis through the VEGF receptor-1. Osteopetrotic Csf1^op/Csf1^op (op/op) mice exhibit severe osteoclast deficiency owing to the lack of CSF-1 function. However, the deficiency is gradually reversed with aging, suggesting the existence of an alternative factor supporting osteoclastogenesis. We have found that the administration of VEGF to op/op mice induces a sufficient number of osteoclasts to ameliorate the osteopetrosis. Estrogen deficiency induces the acceleration of osteoclastic bone resorption mediated by the upregulation of bone-resorbing factors including CSF-1. Ovariectomized op/op mice exhibited upregulation of VEGF expression and an increase in number of osteoclasts. VEGF antagonists inhibited both spontaneous osteoclast recruitment in the aging op/op mice and estrogen deficiency-dependent increases in osteoclasts in OVX-op/op mice. These results clearly demonstrate an ability of osteoclastogenic activity of VEGF.

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It is speculated more that 400 bacterial species reside in the oral cavity, and the integrity of oral mucosa is maintained in such an environment. Oral mucosal cells such as epithelial cells and fibroblasts express a wide variety of receptors and produce many immune regulators i. e., cytokines, chemokines, proteases, and chemical mediators and actively participate in host defenses by interacting with immune cells either directly or indirectly. Furthermore, it was evident that saliva contains many immune regulatory factors. Thus, a triangle consisting of oral mucosal cells, immune cells and saliva formed by connecting with the immune regulator is considered important to maintaining the integrity of oral mucosa, and dysfunction (disorder) of the triangle leads to onset of oral mucosal diseases and inflammation such as oral lichen planus, pemphigus, Sjögren syndrome, oral cancer, leukoplakia, and periodontitis. We consider oral mucosal diseases to be “oral mucosal signal transduction diseases” and are investigated the molecular mechanism involved in disease onset. To overcome “oral mucosal signal transduction diseases”, we attempted to clarify the underlying molecular mechanisms and develop appropriate therapeutic strategies. This review focuses on our recent study regarding 1) regulation of innate immune responses by host and parasitic proteases in oral mucosa, 2) cross-talk between oral mucosal cells and immune cells, 3) activation of oral mucosal cells by proteases, and 4) regulation of oral mucosal responses by salivary components.

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NF-E2 related factor 2 (Nrf2) is a transcription factor that plays pivotal roles in the protection against the toxicity of electrophiles and reactive oxygen species. Nrf2 possesses a basic region-leucine zipper domain, which is well conserved among members of the CNC (Cap’n’collar) family proteins. Analysis of the regulatory mechanism governing Nrf2 activity led to the identification of a novel cytoplasmic protein named Keap1 (Kelch-like ECH-associated protein1), which inhibits Nrf2 activity by directly binding to the N-terminal Neh2 domain. This Nrf2-Keap1 system is responsible for the protective response against oxidative stress in mammals. Although the regulatory mechanism of Nrf2 has been clarified through many in vitro experiments, the establishment of Nrf2-deficient mice has advanced the analysis of roles that Nrf2 plays in terms of pathophysiology. As Nrf2 regulates a range of cytoprotective genes, Nrf2-deficient mice show susceptibility to toxic electrophiles or oxidative stress. In addition, Nrf2-deficient mice are prone to inflammation and autoimmune diseases. Nrf2-deficient mice show an interesting phenotype in the oral region: decolorization of the incisors. We demonstrated that this decolorization in Nrf2-deficient mice was due to the loss of iron deposition on the enamel surface. In this review, we will discuss the significance of Nrf2 in the protection against diseases, including oral diseases.

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Aims: This study aimed to elucidate modifications of tooth pulp-driven neuron (TPDN) activities in thalamic ventral posteromedial (VPM) (n=11) and mediodorsal (MD) (n=13) nuclei following chemical conditioning stimulation with the inflammatory irritant mustard oil (allyl-isothiocyanate, MO) applied to pulp in rats. An additional aim was to examine the participation of the endogenous opioid peptide-related central mechanism in this modification. Methods: Single unit activities of TPDNs were recorded extracellularly from the two thalamic nuclei, and changes in their responsiveness were tested during MO and mineral oil (Min) conditioning stimulation to the same pulp (VPM: n=8; MD: n=7). Results: TPDNs in the VPM and MD showed significant increases in pulp-evoked responses following the MO conditioning stimulation (p<0.05, Wilcoxon test), which were also significant compared to baseline following Min conditioning (p<0.05, Mann-Whitney U-test). Systemic administration of the opiate antagonist naloxone with MO conditioning stimulation produced different patterns of significant increases in VPM and MD TPDN responsiveness in respect to the spike numbers evoked. All TPDNs had cutaneous mechanoreceptive fields (CMRFs), and MO conditioning significantly lowered the mechanical threshold of responses to touch and pressure applied to the CMRF (p<0.05, t-test). Conclusion: These results demonstrated that MO conditioning stimulation of pulp affects the responsiveness and cutaneous mechanical threshold of thalamic TPDNs suggesting that responsiveness of neurons in the central nervous system may be sensitized and modified by pulpal inflammation. Different types of modifications in neuronal responsiveness may be closely correlated with different mechanisms related to pain in tooth pulp.

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Enamel matrix derivative, bone morphogenetic protein and platelet-rich protein are used for periodontal regeneration therapy, but they are not always sufficient. Furthermore, new regeneration factors are needed to promote periodontal regeneration therapy. In the present study, monoclonal antibodies (MoAbs) against the periodontium were produced for use in detection of specific factor. A decalcified 3-week-old Sprague-Dawley rat mandible including tooth and periodontium was used as an antigen to immunize BALB/c mice. MoAb against rat periodontium was produced using immunized spleen cells and P3U1 myeloma cells. Antibodies were screened and cloned using an immunohistochemical method and a limiting dilution method, respectively. MoAb against rat periodontium was isolated, and designated as RPT1. The RPT1 reacted with the periodontal ligament, periosteum and fibrous tissue immediately beneath the gingival and hard palatal epithelia of the rat. All other examined tissues were negative for the RPT1. As sections treated with periodic acid were negative for the MoAb, the antigen seemed to be glycoprotein and was stable for heat and other organic solvents. The RPT1-positive sites coincided with hard tissue. Consequently, identification of the antigen reactive with this MoAb could lead to the development of novel agents for use in periodontal regeneration therapy.

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Nerve growth factor (NGF) and neurturin (NTN) are neurotrophic factors that affect the proliferation and survival of neurons. Using a double immunostaining method, we examined whether the taste bud cells in the circumvallate, foliate and fungiform papillae of normal mice expressed NGF, NTN, and their receptors, TrkA and GFRα2, respectively, and if so, what type of cells in the taste buds expressed them. Because NCAM is reported to be expressed in type-III cells in the taste buds, PGP9.5 in type-III and some type-II cells, and α-gustducin, in some type-II cells, we used these markers to identify the types of taste bud cells. Almost all normal taste bud cells expressed NGF, TrkA, NTN, and GFRα2. Confocal laser scanning microscopic observations after double immunostaining showed that almost all anti-NCAM-, anti-PGP9.5-, or anti-α-gustducin immunoreactive cells were positive for NGF. Thus, NGF-immunoreactive cells included type-II and -III cells, and perhaps type-Icells. Almost all anti-PGP9.5-immunoreactive cells were positive for TrkA, NTN, and GFRα2, revealing that type-III cells expressed TrkA, NTN, and GFRα2; although other types of cells were also immunopositive for these receptors and NTN. We suppose that NGF and NTN in the taste bud cells may exert trophic actions on the taste bud cells by binding to their respective receptors, and additionally be involved in the synaptic transmission from the type-III cells to the gustatory nerves.

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An impacted maxillary canine is not a rare anomaly. A disturbance in the normal eruption pattern of one or both of the maxillary canines is found in 1% to 2% of the children aged ten to thirteen years. The material reported in this study was human skeletal remains excavated from the Ashigasaki-nishidaira site, and was probably a female aged 20-40 years old. The site was located on a terraced river embankment by the Shinano river, and the material was recovered from a stratum from the Edo period. The material had symmetrically impacted maxillary canines in the area of the labial maxilla. The right canine was well-preserved, whereas the left one was damaged after death. The maxillary permanent canine usually transposes with the first premolar and occasionally with the lateral incisor. The normal eruption disturbance of the right lateral incisor was obviously associated with the right impacted canine. The malposition of the eruption and/or the dental germ was thought to be the cause of these dental anomalies.

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Streptococcus mutans glucan-binding protein C encoded by the gbpC gene is an important virulent factor in human dental caries. However, a nonsense mutation and several nucleotide substitutions in the gbpC gene have been detected in strain GS-5. Therefore, gbpC nucleotide sequences from 17 additional S. mutans strains were determined and single nucleotide polymorphisms were detected. Both conserved and polymorphic regions of the gbpC gene will be useful for estimation of functional domains of GbpC protein and identification of S. mutans strains.

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