An important element of treatment for Cleft lip/cleft palate is the attainment of normal occlusion. We recultured human bone tissue-derived mesenchymal stromal cells (hBT-MSCs) in vitro following cryopreservation for ≥10 years, divided the cells into osteogenic differentiation and nondifferentiation MSC groups, and compared the cells in the two groups. In vitro expression of osteoblast markers was then measured. Furthermore, we created a hybrid-type bone substitute and transplanted it into the skulls of 8-week-old male nude rats. Eight weeks later, the skulls were examined by Micro CT. We then performed microscopic analysis of grafts after hematoxylin and eosin (HE) staining and calculated the surface ratio. Immunohistochemical staining with an anti-human osteocalcin antibody was also performed and NEO STEM^™ expression was confirmed by fluorescence microscopy. With regard to osteoblast marker expression, alkaline phosphatase and osterix were significantly elevated in the osteogenic differentiation MSC group (P < 0.05). Micro CT revealed osteogenesis in the hybrid-type bone substitute grafts. HE staining clearly indicated new bone formation within porous hydroxyapatite in the hybrid-type bone substitute grafts. Immunohistochemical staining indicated anti-human osteocalcin antibody expression and NEO STEM^™ expression in the sites exhibiting new bone formation. The volume of bone formation was also significantly higher in the osteogenic differentiation MSC group than in the nondifferentiation MSC group (P < 0.05). We observed excellent osteogenic capability of hBT-MSCs following cryopreservation for ≥10 years, and we believe that our hybrid-type bone substitute can be adapted for future clinical application.

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The aim of this study was to evaluate the efficacy of collagen- or laminin-immobilized titanium implant on surrounding bone formation after implantation in a rat molar model. Collagen or laminin was immobilized onto titanium implants using a tresyl chloride-activated technique. The rat first molars were extracted and then the implants were immediately placed in the tooth sockets. At 4 weeks after implantation, undecalcified sections were prepared and bone response around the implants was evaluated. The value of bone to implant contact was significantly greater around the collagen-immobilized implants than around the titanium- and laminin-immobilized ones. It is assumed that immobilized collagen triggered bone formation around the implant material. These results suggest that collagen immobilization accelerates new bone formation.

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Better understanding of the dentin-pulp complex is now leading to treatment modalities aimed at tooth regeneration, such as pulp capping, in clinical settings. However, the control of infection and biocompatibility of pulp-capping materials are important factors in determining treatment outcomes. The purpose of this study was investigated the effects of CaCO₃ on calcified nodule formation, ALP activity, cell viability and BMP-2 and OCN expression after 30-day culture of human dental pulp cells (hDPCs). The results showed that the number of calcium nodules, ALP activity, cell viability and BMP-2 and OCN expression increased in a CaCO₃-dependent manner. PCR data also confirmed that Smad1 mRNA expression in hDPCs increased in a CaCO₃ dose-dependent manner. This suggests that CaCO₃ is able to induce in vitro cell differentiation of hDPCs into cells capable of mineralization. Thus, treatment of exposed pulp with CaCO₃ is effective for dentinogenesis.

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The purpose of this study was carried out by the embryotoxicity risk assessment of the dental alloys containing indium. We produced Ag-In alloy, Ag-Pd-Au-Cu alloy and gold alloy for porcelain bonding. For the extraction of these alloy components into the ES-D3 cell culture medium, each alloy was reduced to a powder using a diamond point and sterilized after rinsing. In order to facilitate the extraction, each alloy powder was subjected to extraction together with diamond powder using a rotary stirrer. Two-fold diluted test medium was used to estimate the presence of embryotoxicity from the differentiation rate of Embryoid Bodies (EBs) in the extract. Our test results this time suggest that 35Ag-30Pd-20Au-15Cu alloys and gold alloys for porcelain bonding involve virtually no embryotoxicity risk. On the other hand, the alloy of silver and 20% indium may require further examination.

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Recently, it has been suggested that topical application of basic fibroblast growth factor (bFGF) is an effective treatment for periodontal tissue regeneration. Gingival fibroblasts (GFs) play an important role in periodontal tissue regeneration. Many patients who receive periodontal therapy take amlodipine (AML). Since AML inhibits cell proliferation via cell cycle arrest, it may hinder the periodontal tissue regeneration induced by bFGF. In this study, we investigated whether AML affects the cell cycle and the expression of its control genes in the presence of bFGF in human GFs (hGFs). Semi-confluent cells were synchronized at the G₀/G₁ phase in DMEM containing 0.5% serum (DMEM-0.5) for 24 hrs. After pretreatment with or without 3 or 10 μM AML in DMEM-0.5 for 24 hrs, cells were stimulated with 10 ng/ml bFGF in serum-free DMEM. Then, analyses of the cell cycle and the cell number, and RT-PCR analysis were performed. Our results showed that AML inhibited the bFGF-induced G₁/S cell cycle transition through suppressing the mRNA expression of Cdk 1, 2, 4, and 6 and cyclins A, D1, and E in hGFs. In conclusion, AML may exert a negative influence on periodontal tissue regeneration induced by bFGF.

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Dental adhesive monomers are extensively used in clinical dentistry and are indispensable for repairing hard tissue. Products commercially available in Japan have passed a number of biological safety evaluation tests, including a cytotoxicity test, and are included as dentin adhesives in managed-care equipment. However, these embryotoxicity risks of each dental adhesive are unknown. Thus, an in vitro embryotoxicity test was conducted using the Embryo Stem Test (EST). As a result, all products were within the range of "non-embryotoxicity", while some products were near "weak embryotoxicity". The products containing Bis-GMA tended to show lower IC50 and ID50 values, with some exceptions. In contrast, those containing MMA compositions tended to have few effects on cell differentiation. These dental adhesive should be investigated further regarding their biological properties.

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In Orthodontic therapy, osteoclast precursors such as monocytes and macrophages migrate from vessels into the periodontium and then differentiate into osteoclasts, causing bone resorption. Focal adhesion kinase (FAK) is a 125-kDa non-receptor type tyrosine kinase that localizes to focal adhesions. FAK is involved in osteoclastic bone resorption. Vascular cell adhesion molecule-1 (VCAM -1) is bound to VLA-4(very late antigen-4) which is kind of the α₄ β1-integrin, and recruits monocytic osteoclast progenitors and elevates local osteoclast activity.In our present study we focused on VCAM-1 /α₄ integrin-mediated the differentiation into osteoclasts and found that this type of the differentiation was mediated through FAK. RAW267.4 expressed both α₄ integrins, and it was reported that expression of α₄ integrin and its counterreceptor, VCAM-1, was not enhanced in response to receptor activator of NF-B ligand (RANKL). Neutralizing antibodies against integrin α4 effectively inhibit the differentiation into osteoclasts and phosphorylation of FAK. These findings establish VCAM-1 regulate the differentiation into osteoclasts in bone.

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Natural killer (NK) cells play a key role in inflammation and tumor regression through their ability to migrate into tissues. CXCL8 is one of the chemokines that promote leukocytes invasion and migration into tissues, while the exact molecular mechanisms are not clear at present. In this study, we showed that CXCL8 significantly enhanced CD16⁺CD56⁺ human peripheral NK cells invasion into type I collagen by the catalytic activity of MMP-1. MMP-1 inhibitor, GM6001 and Gi-protein inhibitor, PTX blocked the invasion of CD16⁺CD56⁺ human peripheral NK cells into type I collagen. GM6001 and PTX did not inhibit the production of MMP-1 in CD16⁺CD56⁺ human peripheral NK cells. PTX inhibited the association of MMP-1 but not GM6001, with cell surface in CXCL-8 in CD16⁺CD56⁺ human peripheral NK cells. The association of MMP-1 with cell surface on CXCL-8-stimulated NK cells suggests that this integrin plays a role not in promoting cell migration to type I collagen but cell invasion into type I collagen. These results suggest that the selective regulations of production and/or localization of MMP-1 in NK cells may lead to effective strategies to control inflammation and tumor elimination.

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As a sequel to a dental care survey of foreign patients conducted in 2011, the authors did a follow-up study on what determines the choice of foreigner's dental treatment in Japan. To make the research more comprehensive, people who deliberately choose not to go to dentists in this country were included as respondents of the questionnaire. Of those surveyed, 63% stated they visit dental clinics on a regular basis. These clinics were mostly found through referrals from other foreign acquaintances. What they had in common was the availability of English as well as a sense of acceptance to the foreign patient. The remaining 37% of those surveyed postpone their dental treatment until their next chance to return to their native countries. Unlike the first group, these people were not able to get the right referrals and other information that would prompt them to visit a dentist in Japan. It is interesting to note that looking at their personal oral care habits, these people do care very much about their oral health. They form not a small potential market for new dental patients.

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