Osteoclasts are the multinucleated giant cells formed by cell fusion of mononuclear osteoclast precursors. Despite the finding of several membrane proteins involving DC-STAMP as regulatory proteins required for fusion among osteoclast precursors, cellular and molecular events concerning this process are still ambiguous. Here we identified Tunneling Nanotubes (TNTs), long intercellular bridges with small diameters, as the essential cellular structure for intercellular communication among osteoclast precursors in prior to cell fusion. Formation of TNTs was highly associated with osteoclastogenesis and it was accompanied with the significant induction of the M-Sec gene, an essential gene for TNT formation. M-Sec gene expression was significantly upregulated by RANKL-treatment in osteoclast precursor cell line. Blockage of TNT formation by Latrunclin B or by M-Sec siRNA significantly suppressed osteoclastogenesis. We have detected the rapid intercellular transport of not only the membrane phospholipids labeled with DiI but also the DC-STAMP-GFP fusion protein through TNTs formed among osteoclast precursors during osteoclastogenesis. Transportation of such regulatory molecules through TNTs would be essential for the process of the specific cell fusion among osteoclast precursors.

The oral cavity provides an entrance to the alimentary tract to serve as a protective barrier against harmful environmental stimuli. The oral mucosa is susceptible to injury because of its location; nonetheless, it has faster wound healing than the skin and less scar formation. However, the molecular pathways regulating this wound healing are unclear. Here, we show that transient receptor potential vanilloid 3 (TRPV3), a thermosensitive Ca 2+ -permeable channel, is more highly expressed in murine oral epithelia than in the skin by quantitative RT-PCR. We found that temperatures above 33°C activated TRPV3 and promoted oral epithelial cell proliferation. The proliferation rate in the oral epithelia of TRPV3 knockout (TRPV3KO) mice was less than that of wild-type (WT) mice. We investigated the contribution of TRPV3 to wound healing using a molar tooth extraction model and found that oral wound closure was delayed in TRPV3KO mice compared with that in WT mice. TRPV3 mRNA was up-regulated in wounded tissues, suggesting that TRPV3 may contribute to oral wound repair. We identified TRPV3 as an essential receptor in heat-induced oral epithelia proliferation and wound healing. Our findings suggest that TRPV3 activation could be a potential therapeutic target for wound healing in skin and oral mucosa.-Aijima, R., Wang, B., Takao, T., Mihara, H., Kashio, M., Ohsaki, Y., Zhang, J.-Q., Mizuno, A., Suzuki, M., Yamashita, Y., Masuko, S., Goto, M., Tominaga, M., Kido, M. A. The thermosensitive TRPV3 channel contributes to rapid wound healing in oral epithelia. FASEB J. 29, 182-192 (2015). www.fasebj.org Key Words: ambient temperature • oral mucosa • wound repair THE ORAL MUCOSA HAS a highly specialized epithelium that performs essential protective functions against diverse changes, such as chemical, thermal, or mechanical stimuli, in the oral environment. The oral cavity is also the site for sentient responses (1, 2). The oral epithelium is a moist lining membrane in the oral cavity and consists of a stratified squamous epithelium and underlying connective tissues similar to the skin. Although it is continuous with the skin, the oral epithelium is more susceptible to injury because it is exposed to more extensive stimuli than the skin. However, wound repair of the oral mucosa is faster than the skin and recovers with less scar formation (3, 4). Although components in the saliva or a rich vascular supply may contribute to this rapid wound healing (4-6), the molecular mechanisms regulating oral mucosa wound repair are still largely unknown.Transient receptor potential (TRP) channels are a family of Ca 2+ -permeable nonselective cation channels that are responsive to a broad range of environmental stimuli such as temperature, tonicity, or pain (7-9). Among the 28 different mammalian TRP channels, transient receptor potential vanilloid 3 (TRPV3) is uniquely expressed predominantly in keratinocytes and is activated by innocuous warm temperatures above 33°C and natural herbs such as oregano or thyme (10-14). Furthermore, it h...

Galectins are a unique family of lectins bearing one or two carbohydrate recognition domains (CRDs) that have the ability to bind molecules with b-galactoside-containing carbohydrates. It has been shown that galectins regulate not only cell growth and differentiation but also immune responses, as well as inflammation. Galectin-9, a tandem repeat type of galectin, was originally identified as a chemotactic factor for eosinophils, and is also involved in the regulatory process of inflammation. Here, we examined the involvement of galectin-9 and its receptor, T-cell immunoglobulin-and mucin-domain-containing molecule 3 (Tim-3), in the control of osteoclastogenesis and inflammatory bone destruction. Expression of Tim-3 was detected in osteoclasts and its mononuclear precursors in vivo and in vitro. Galectin-9 markedly inhibited osteoclastogenesis as evaluated in osteoclast precursor cell line RAW-D cells and primary bone marrow cells of mice and rats. The inhibitory effects of galectin-9 on osteoclastogenesis was negated by the addition of b-lactose, an antagonist for galectin binding, suggesting that the inhibitory effect of galectin-9 was mediated through CRD. When galectin-9 was injected into rats with adjuvant-induced arthritis, marked suppression of bone destruction was observed. Inflammatory bone destruction could be efficiently ameliorated by controlling the Tim-3/galectin-9 system in rheumatoid arthritis.

Experimentally, temporomandibular joint (TMJ) nerve units respond to capsaicin, which is used clinically to treat TMJ pain. However, the existence of capsaicin receptors in the TMJ has not previously been clearly demonstrated. Immunohistochemical analysis has revealed the presence of transient receptor potential vanilloid subtype 1 (TRPV1) expression in the nerves and synovial lining cells of the TMJ. TRPV1-immunoreactive nerves are distributed in the synovial membrane of the joint capsule and provide branches to the joint compartment. The disc periphery is supplied by TRPV1 nerves that are mostly associated with small arterioles, and occasional nerves penetrate to the synovial lining layer. Double immunofluorescence has shown that many TRPV1-immunoreactive nerves are labeled with neuropeptide calcitonin gene-related peptide, whereas few are labeled with IB4-lectin. The results provide evidence for the presence of TRPV1 in both nerves and synovial lining cells, which might thus be involved in the mechanism of nociception and inflammation in the TMJ.

Background:The innervation of rat incisors, which are continuously erupting teeth, is quite unique. Although many reports have documented the neural control of the pulpal blood flow, only a few studies have examined the structure and distribution of vascular nerves in the rat incisors. This study examined the nerve plexuses and the spatial relationship of nerve terminals to smooth muscle cells of the pulp vessels of rat incisors.Methods: The innervation of the pulp vessels of rat incisors was studied using immunohistochemical evaluations of calcitonin gene-related peptide (CGRP) and neuropeptide-Y (NPY). The three-dimensional ultrastructure of nerve meshworks and terminals on smooth muscle cells of pulp arterioles were examined by scanning electron microscopy (SEM) with a KOH digestion method.Results: The blood vessels were associated with many nerve fibers immunoreactive for CGRP and NPY. Some NPY and frequent CGRPimmunoreactive nerve fibers were observed in the labial odontoblast layer. Three different morphologies of nerve fibers could be distinguished: fine nerve fibers with numerous terminal varicosities, medium-sized fibers associated with occasional or much scarcer varicosities, and thick fibers that had no varicosity and sometimes ran apart from blood vessels. The SEM observations identified five vascular segment types: terminal arterioles, precapillary arterioles, capillaries, postcapillary venules, and muscular venules. Nerve meshworks were observed around the large terminal arterioles; these meshworks were very comparable to those revealed by immunohistochemistry. The vascular smooth muscle cells were closely attached by one or more terminal varicosities.Conclusions: It has become evident that the large terminal arterioles in the rat incisors receive a dense nerve supply and provide well-developed wall architecture, suggesting an important role of neuronal regulation on the vessels. Anat.