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This study investigated the in vitro anti-demineralization effects of resin-based temporary filling materials containing surface prereacted glass-ionomer (S-PRG) filler on dentin. Bovine root dentin specimens with a 3×3 mm experimental surface were divided into four treatment groups: DuraSeal (DU) as a control, S-PRG filler-free temporary material (S0), material containing 10% (S10) and 20% (S20) S-PRG filler. Each material was applied to 3×2 mm of the experimental surface, and the specimens were immersed in 8% methylcellulose gel demineralization system for one week at 37˚C. Mineral profiles and integrated mineral loss (IML) of lesions induced on the surface (3×1 mm) adjacent to the materials were computed by transversal microradiography. S10 and S20 yielded thick surface layers and shallow lesion bodies, with significantly lower IML than DU and S0 (p

Salivary macromolecules infiltrate white and brown spot enamel lesions and adsorb onto hydroxyapatite. Calcium-binding salivary proteins such as statherin hinder remineralization of these lesions. We assessed whether bleaching agents can remove salivary components that have infiltrated and bound to experimental subsurface lesions in bovine enamel prepared by immersing specimens in acid and then human saliva. Transversal microradiography showed that such demineralized lesions mimicked incipient carious lesions. Bound proteins to the experimental and untreated control specimens were eluted in a stepwise manner with phosphatebuffered saline, 0.4 M phosphate buffer, and 1 M HCl. SDS-PAGE of dialyzed extracts showed that specific salivary proteins bound to the lesions, while virtually no protein bands were detected if the specimens were bleached. Western blotting showed that even statherin, which was more firmly bound than other proteins, was removed. In-office bleaching agent may be useful in treating enamel lesions for removing proteins bound to these lesions.

The polymerization shrinkage of flowable resin composites was evaluated using air bubbles as traceable markers. Three different surface treatments i.e. an adhesive silane coupling agent, a separating silane coupling agent, and a combination of both, were applied to standard cavities. Before and after polymerization, X-ray micro-computed tomography images were recorded. Their superimposition and comparison allowed position changes of the markers to be visualized as vectors. The movement of the markers in the resin composite was, therefore, quantitatively evaluated from the tomographic images. Adhesion was found to significantly influence shrinkage patterns. The method used here could be employed to visualize shrinkage vectors and shrinkage volume.

The preceramic polymer perhydropolysilazane (PHPS) is an attractive candidate as a coating material to prevent discoloration of veneering resin composites. At the present time, however, a practical method to apply this material is not available. The purpose of this study was to establish a low-temperature method for applying a silica film coating to a veneering resin composite. Two types of PHPS, NP and NL, were coated onto a veneering resin composite. The specimens were exposed to hydrogen peroxide vapor at 97°C, and the state of the conversion process was evaluated using FT-IR. With exposure to the hydrogen peroxide vapor, a 0.5-µm-thick silica film similar to that produced by baking was formed on the surface of the NP samples in 10 min, while a 0.2-µm-thick film was formed on the NL in 15 min. The silica coating method described in this study may mitigate the discoloration of veneering resin composite.

The aims of this study were to evaluate a new restorative method using a carbon dioxide laser (CO(2)-laser) and to evaluate the acid resistance of teeth. Experimental calcium phosphate glass (CPG) powder and two low melting point ceramics (Finesse and zirconium silicate) were fused to enamel surfaces using a CO(2)-laser at an irradiation intensity of 1.0 watt for 30 seconds with a beam size of 0.49 mm at the focal point. The treated teeth were observed with a scanning electron microscope, and the acid resistance of the treated enamel surfaces was evaluated. The CPG fused successfully to the enamel surface, and the treated enamel surface showed high acid resistance compared with the low melting point ceramics and the non-irradiated surfaces. This system may lead to the development of new restorative methods that do not require the use of bonding agents.

The purpose of this study was to evaluate the influence of a bottled green tea beverage on dentin demineralization with a demineralization gel system in vitro . Samples were cut from bovine root dentin. Each sample was immersed in 8% methylcellulose gel onto which a layer of green tea beverage was placed. For comparison, sugar-free coffee beverage, deionized water, and 0.8-ppm F solutions were used. After two weeks, the treatment solutions were replaced with demineralization solution. The mineral profiles and mineral loss values of the lesions were obtained by transversal microradiography (TMR) after one-week demineralization. The green tea, coffee, and fluoride solution treatments induced a significantly thicker surface layer when compared with the deionized water treatment. In particular, the mineral volume % of the demineralized dentin specimens treated with green tea was approximately seven times higher than that of the de-ionized water treatment. The green tea treatment showed significantly lower mineral loss than the other three treatments. TMR measurements clearly showed that the sugar-free bottled green tea beverage inhibited dentin lesion progression, presumably due to the effect of sub-ppm fluoride levels.

This study aimed to investigate the shear bond strength of an autopolymerizing resin to a nylon denture base polymer (Lucitone FRS: LT) subjected to different surface treatments, and the results thereof compared with a heat-polymerizing resin and a polycarbonate polymer. Specimens were divided into five groups according to the surface treatment method: polishing (#600), sandblasting, adhesive primer application (resin primer), sandblasting + adhesive primer application, and tribochemical coating (Rocatec system). Following which, specimens were subjected to a shear bond strength test and Si concentrations were measured using an electron probe microanalyzer (EPMA). On shear bond strength, that of LT with tribochemical coating was significantly higher than the other groups. On EPMA results, the surface of LT with tribochemical coating was found to be covered with a silica film. Therefore, findings in this study indicated that silica-coating by Rocatec system was effective in improving the bond strength of nylon denture base polymer to autopolymerizing repair resin.

Gemini-type hybrid surfactants with two fluorocarbon chains connected through a hydrocarbon spacer, F(CF2)m(CH2)2CH(OSO3Na)(CH2)nCH(OSO3Na)(CH2)2(CF2)mF [Fm(Hn)FmOS, m = 4, 6; n = 5, 6, 7, 8)], were synthesized and their surface chemical properties were examined with the aim to have highly functional and highly water-soluble fluorinated surfactants when compared with the conventional fluorinated surfactants. Comparisons of the surface chemical properties of the synthesized gemini-type hybrid surfactants with those of monounit-type hybrid surfactants, F(CF2)m(CH2)2CH(OSO3Na)(CH2)nH [FmEHnOS, m = 4, 6; n = 3, 5)], revealed that gemination causes a remarkable lowering (about 1/100) in cmc value while it produces little changes in Krafft point (below 0 °C) and surface tension at cmc (γcmc).

This paper evaluated the wear resistance of resin composite materials with fillers which were modified with a novel hydrophobic silane coupling agent. The novel silane coupling agent containing hydrophobic phenyl group 3-(3-methoxy-4-methacryloyloxyphenyl)propyltrimethoxysilane (p-MBS) was synthesized. The experimental light-cure hybrid composites containing 85wt% of filler modified with this silane were formulated. Twelve specimens were prepared for the three-body-wear test with the ACTA machine and the collected data were analyzed statistically using a one-way ANOVA and Tukey's multiple comparison test as the post hoc test. The wear of the composites containing fillers treated with p-MBS was significantly lower compared with the composite materials containing fillers pretreated with 3-methacryloyloxypropyltrimethoxysilane or the commercially composites (AP-X and ELS extra low shrinkage) after a wear test for 200,000 cycles (p

The effect of fluoride-containing orthodontic resin bonding systems on acid resistance of adjacent enamel was evaluated using transversal microradiography. Four fluoride-containing orthodontic resin bonding systems: BeautyOrtho Bond (BO), BeautyOrtho Bond+Salivatect (BOS), Kurasper F (KP), Transbond XT (TB) and a resin-modified glass-ionomer (Fuji Ortho LC (FO)) were used. Superbond Orthomite (SB) was used as a non-fluoride material. Rectangular bovine enamel specimens (10X6 mm) were prepared. After curing the materials, nail varnish was applied to the enamel surfaces, leaving a gap of 1 mm from the cured material's periphery. The specimens were demineralized with 8% Methocel MC gel and 0.1 M lactic acid. BO, BOS, and FO revealed shallow lesions and distinct surface layers. The mineral losses of BO, BOS, and FO were significantly lower than those of TB, KP, and SB (p

The biocompatibility of periapical tissue with mineral trioxide aggregate (MTA) affects its ability to repair and regenerate itself. Here we report the cytotoxicity of MTA and how it affects the expression of bone extracellular matrix protein in MC3T3-E1 osteoblast cells. We quantified the cytotoxicity of MTA, amalgam, and Dycal (Dentsply/Caulk, Milford, DE) on MC3T3-E1 cells by measuring the ability of cells to cleave a tetrazolium salt to produce formazan dye during a period of 24, 48, or 96 hours. We used reverse-transcriptase polymerase chain reaction with primer sets for type I collagen, osteocalcin, and bone sialoprotein to measure the gene-expression response of MC3T3-E1 cells treated with MTA. MTA, amalgam, and Dycal were less toxic after 48 hours. MC3T3-E1 cell growth with MTA and Dycal was greater than nonstimulated controls. MTA caused an upregulation of type I collagen and osteocalcin messenger RNA expression after 24 hours. These results showed that, in the presence of MTA, cells grow faster and produce more mineralized matrix gene expression in osteoblasts.

The purpose of this study was to evaluate the antibacterial activity against polymicrobial (PM) biofilms of a condensed tannin extracted from astringent persimmon (PS-M), which is contained in refreshing beverages commercially available in Japan. Salivary PM biofilms were formed anaerobically on glass coverslips for 24 and 72 h and were treated for 5 min with sterilized deionized water (DW), 0.05 and 0.2 wt% chlorhexidine digluconate (CHX), and 0.5–4.0 wt% PS-M solution. The colony forming units (CFU/mL) were determined and morphological changes of the biofilms were observed by scanning electron microscopy (SEM). The CFUs were lower in all PS-M and CHX groups compared to the DW group. PS-M exerted a dose-dependent effect. PS-M (1.53 × 107) at a dose of 4.0 wt% had the same effect as 0.2 wt% CHX (2.03 × 107), regardless of the culture period. SEM revealed the biofilm structures were considerably destroyed in the 4.0 wt% PS-M and 0.2 wt% CHX. These findings indicate that the antibacterial effects of PS-M, a naturally derived substance, are comparable to those of CHX. PS-M may keep the oral cavity clean and prevent dental caries and periodontal disease related to dental plaque, as well as systemic disease such as aspiration pneumonitis.

To simulate an oral demineralization environment by multiple species of bacteria by inducing subsurface dentin lesions with a polymicrobial biofilm model.Polymicrobial biofilms consisting of multiple species of bacteria were generated from stimulated saliva using a high-throughput active attachment model. Biofilms were grown on dentin specimens in McBain medium containing 0, 0.2 or 2.5 ppm F and on glass without fluoride for 192 hours. The medium was refreshed twice daily, after 10 and 14 hours, until 72 hours, followed by treatment for 5 minutes with 400 ppm fluoride. Specimens were recovered after 192 hours. The number of colony forming units (CFU) was measured, and integrated mineral loss (IML) was determined by transversal microradiography.Mineral profiles in specimens grown with 0.2F and 2.5F revealed surface layers and initial lesions distinct from those grown with 0F. IML was significantly lower with 0.2F and 2.5F than with 0F (P0.05), although CFUs were similar. CFUs of biofilms grown on dentin in medium containing 0F were 10-fold higher than on glass (P0.05). Subsurface lesions on dentin formed consistently, with their growth progression inhibited by application of fluoride. To our knowledge, this is the first report describing the induction of subsurface dentin lesions by a polymicrobial biofilm model, and this model may be useful for studies of demineralization supporting in situ and in vivo models.

A fluoride-releasing coating material containing surface pre-reacted glass-ionomer (S-PRG) filler has become commercially available. However, there has been no detailed investigation of its remineralization effects at various tooth surface regions. The remineralization effects of S-PRG filler-containing coating material at different sites of demineralized dentin surfaces in vitro were evaluated.Baseline lesions were prepared on bovine root dentin surfaces by immersion in demineralization buffer and divided into four groups: (B)--baseline lesion; (P)--S-PRG filler-containing material; (V)--S-PRG filler-free coating material as negative control; and (X)--resin-modified glass- ionomer as positive control. Material was applied to half the lesion surface, then P, V and X were remineralized in a gel system. Mineral profiles, integrated mineral loss (IML) and lesion depth (LD) at four regions, i.e. 1--exposed dentin surface adjacent to the material; 2--at a distance from the material; 3--beneath the material near to the edge; and 4--at a distance from the edge, were analyzed by transversal microradiography. Data were analyzed using ANOVA and Games-Howell test with α = 0.05.B showed typical artificial demineralized lesion. The IMLs of V, P and X at regions 1 and 2, and P and X at region 3 were significantly lower than that of B, however, those of V at region 3 and the other three groups at region 4 were not significantly different from that of B. At region 1, P and X showed significantly lower IMLs than V. At region 2, the IML of X showed significantly lower IML than V. There was no significant difference between P and X. The LD values of V, P and X at all regions were not significantly different from that of B. Fluoride, strontium and silicate ions released from the S-PRG filler would provide a favorable environment for remineralization of the demineralized dentin in P.

The dental follicle is a mesenchymal tissue that surrounds the developing tooth germ. During tooth root formation, periodontal components, viz., cementum, periodontal ligament (PDL), and alveolar bone, are created by dental follicle progenitors. Here, we report the presence of PDL progenitors in mouse dental follicle (MDF) cells. MDF cells were obtained from mouse incisor tooth germs and immortalized by the expression of a mutant human papilloma virus type 16 E6 gene lacking the PDZ-domain-binding motif. MDF cells expressing the mutant E6 gene (MDF( E6-EGFP ) cells) had an extended life span, beyond 150 population doublings (PD). In contrast, normal MDF cells failed to proliferate beyond 10 PD. MDF( E6-EGFP ) cells expressed tendon/ligament phenotype-related genes such as Scleraxis (Scx), growth and differentiation factor-5, EphA4, Six-1, and type I collagen. In addition, the expression of periostin was observed. To elucidate the differentiation capacity of MDF( E6-EGFP ) cells in vivo, the cells were transplanted into severe combined immunodeficiency mice. At 4 weeks, MDF( E6-EGFP ) cell transplants had the capacity to generate a PDL-like tissue that expressed periostin, Scx, and type XII collagen and the fibrillar assembly of type I collagen. Our findings suggest that MDF( E6-EGFP ) cells can act as PDL progenitors, and that these cells may be a useful research tool for studying PDL formation and for developing regeneration therapies.

A cementoblast progenitor cell line designated BCPb8 was successfully isolated from dental follicle cells immortalized with Bmi-1 and hTERT. BCPb8 showed the potential to differentiate into cementoblasts on implantation into immunodeficient mice. BCPb8 was confirmed to be the first established cementoblast progenitor cell line and will provide a useful model for investigating cementogenesis. Introduction: The dental follicle is the mesenchymal tissue surrounding the developing tooth germ. During tooth root development, progenitor cells present in the dental follicle are believed to play a central role in the formation of periodontal components (cementum, periodontal ligament, and alveolar bone). However, little more is known about the biology of these progenitors. Previously, we observed that cultured bovine dental follicle cells (BDFCs) contained putative cementoblast progenitors. To further analyze the biology of these cells, we attempted to isolate cementoblast progenitors from immortalized BDFC through expression of the polycomb group protein, Bmi-1, and human telomerase reverse transcriptase (hTERT). Materials and Methods: BDFCs were transduced with replication-deficient retroviruses carrying human Bmi-1(LXSN-Bmi-1), and hTERT (LXSH-hTERT) for immortalization. Single cell clones were established from immortalized BDFC, and differentiation into cementoblasts was assessed by implantation into immunodeficient mice. Results and Conclusion: BDFCs expressing Bmi-1 and hTERT showed an extended life span - 90 population doublings more than normal BDFCs - and still contained cells with the potential to differentiate into cementoblasts on implantation into immunodeficient mice. From these cells, we established a clonal cell line, designated BCPb8, which formed cementum-like tissue that was reactive to the anti-cementum-specific monoclonal antibody 3G9 and expressed mRNA for bone sialoprotein, osteocalcin, osteopontin, and type I collagen on implantation. Thus, by using Bmi-1 and hTERT, we succeeded in immortalizing cementoblast progenitor cells from BDFC without affecting differentiation potential. The BCPb8 cell line is the first immortalized clonal cell line of cementoblast progenitors and could be a useful tool not only to study cementogenesis but also to develop regeneration therapy for patients with periodontitis.

Objectives. The objectives of the current investigation were two-fold: (1) to examine the effect of different powder/liquid (P/L) ratios on the fracture toughness of commercial resin-modified glass ionomer cement and conventional glass ionomer cement, and (2) to evaluate the effect of powder size reduction on the fracture toughness of experimental resin-modified glass ionomers in order to improve their physical properties. Methods. The P/L ratios of the glass ionomer and resin-modified glass ionomers were varied from the manufacturer's recommended ratio to 2.0 and 1.0 by weight. The powder particle sizes for the experimental resin-modified glass ionomers tested were 2, 5, 10 and 25 μm in diameter. Fracture toughness was determined on ring-shaped specimens with a fatigued pre-crack. Results. The fracture toughness of the resin-modified glass ionomers was significantly higher (p

Incadronate (YM175, disodium cycloheptylaminomethylenediphosphonate monohydrate), a bisphosphonate, has been suggested to prevent the bone resorption associated with periodontitis by inhibiting osteoclast activity. The purpose of this study was to investigate the effect of incadronate in preventing periodontal destruction in rats with Porphyromonas gingivalis-induced periodontitis.Periodontitis was induced in 35 Wister rats by inoculating P. gingivalis into the oral cavity and feeding the rats a soft diet for 4 weeks. Incadronate or placebo was administered to the oral cavity of the rats 2 days per week for 2, 4, or 8 weeks.P. gingivalis infection resulted in destruction of the periodontal ligament, reduced bone density, and caused inflammatory cell migration. Radiographic, morphometric, and histological results showed that incadronate had the ability to increase the bone mineral density (quantum level score; cortex 518.9 [placebo 612.8]; sponge 579.8 [placebo 672.0]) and to prevent periodontal ligament destruction (width 0.16 mm [placebo 0.20 mm]; area 0.36 mm2 [placebo 0.54 mm2]) after 8 weeks' administration. Furthermore, the polymorphonuclear leukocyte (PMN) infiltration in gingival tissue was significantly decreased.These results showed that incadronate inhibits bone resorption and PMN migration in P. gingivalis-induced periodontitis.

The hydrolytic stability of a group of experimental composite materials was evaluated. Seven distinct composites were formed by the mixing of a resin monomer mixture with silica filler that had been pre-treated with one of 7 different ethanol solutions. In one case, the filler was treated with an ethanol solution that contained only 3-methacryloyloxypropyltrimethoxysilane. In 5 cases, it was treated with solution containing a mixture of 3-methacryloyloxypropyltrimethoxysilane and one of the following hydrophobic fluoroalkyltrimethoxysilanes: trifluoropropyl-, nonafluorohexyl-, tridecafluorooctyl-, heptadecafluorodecyl-, and henicosafluorododecyl-trimethoxysilane. The tensile strength, after being immersed in water for 1800 days, of 2 of the experimental composites, whose pre-treatment regimen had included a fluoroalkyltrimethoxysilane, was significantly higher than that of the composite whose pre-treatment regimen had not included a fluoroalkyltrimethoxysilane. Moreover, there was no significant difference between the tensile strength of fresh samples of these 2 composites and the tensile strength of identically produced samples that had remained under water for 1800 days or that had been subjected to 30,000 cycles of thermal stress.

Interactions between leukocyte function-associated antigen-1 (LFA-1) and intercellular adhesion molecule-1 (ICAM-1) influence the development of osteoclasts. However, little is known about how these adhesion molecules are involved in the process of osteoclast development. This study evaluated the role of LFA-1 and its ligands in osteoclast development and bone resorption. Co-cultures of bone marrow cells from LFA-1-deficient mice and MC3T3-G2/PA6 (PA6) cells were cultured in the presence of 1alpha,25(OH)2D3 and dexamethasone for 7 days. The number of TRAP-positive cells that were generated by bone marrow cells from LFA-1-deficient mice was smaller than that generated by bone marrow cells from wild-type mice. In addition, the bone-resorbing activity of osteoclast-like cells that were generated from LFA-1-deficient mice was lower than that generated by osteoclast-like cells from wild-type mice. Immunofluorescence flow cytometry showed that osteoclast stromal PA6 cells expressed the cell adhesion molecules, ICAM-1 and VCAM-1. When monoclonal antibodies to mice VCAM-1, CD11b or CD18 were added separately to the co-culture system, the number of TRAP-positive cells that were generated from LFA-1-deficient mice was 20-30% smaller than that generated from wild-type mice. The formation of TRAP-positive cells from both LFA-1 deficient and wild-type mice was especially inhibited by anti-CD18 antibody, in comparison to the addition of normal IgG serum. These results suggest that LFA-1 adhesion molecules play a role in osteoclast development by affecting adhesion between stromal cells and osteoclast progenitors before the occurrence of ODF-ODF receptor signaling. CD18 appears to be a key adhesion molecule in cell-to-cell contacts during the early stage of osteoclast development.

Cementum-derived attachment protein (CAP) is a 56 kDa collagenous protein that promotes attachment of mesenchymal cells. Previous studies have shown that the presence of CAP is restricted to cementum in adult human tissues. In this study, we report generation of a monoclonal antibody against CAP and its use for the investigation of CAP in developing bovine tooth germs. Mice were immunized with CAP purified from bovine cementum, and a monoclonal antibody, 3G9, was produced. Immunohistochemical staining of bovine tooth germ at root forming stage using 3G9 antibody showed that the tissue distribution of CAP expression was limited to cementum matrix and cementoblasts during cementogenesis. Alveolar bone did not stain with the 3G9 antibody, whereas anti-type I collagen stained positively. CAP was purified from bovine tooth germs with immunoaffinity purification using the 3G9 antibody. Examination of the immunoaffinity-purified fraction showed that CAP existed in tooth germ as a 65 kDa protein. The protein was susceptible to bacterial collagenase. To investigate the possible biological function of CAP during cementogenesis, we isolated dental follicle cells from the bovine tooth germ, and showed that they adhered to surfaces containing CAP. These data demonstrate that CAP is expressed by bovine cementoblasts as a 65 kDa protein and that the CAP may have a function in cementogenesis.

Bone resorption is regulated by the cytokines within marrow cells that mediate osteoclast formation and activation. IL-1 and TNF induce bone resorption by stimulating the production of osteoclast-like multinucleated cells and by increasing the bone-resorbing activity of formed osteoclasts. This study was designed to detect IL-1 and TNF in osteoclasts in vitro and to determine whether these cytokines up-regulate osteoclast differentiation and bone resorption. The production of IL-1 alpha, -beta, and TNF alpha, beta in osteoclasts was examined immunohistochemically and by in situ hybridization. In the co-culture of C57BL/6N mouse bone marrow and MC3T3-G2/PA6 cells, a colony of osteoclasts formed, and IL-1 alpha and TNF alpha were detected. However, IL-1 beta and TNF beta were not detected. To investigate the role of IL-1 alpha and TNF alpha from osteoclasts, we enumerated TRAP-positive cells and measured the resorption pit areas in the presence of antibodies against IL-1 alpha and TNF alpha. The addition of antibodies against IL-1 alpha and TNF alpha to the co-culture system decreased the number of TRAP-positive colonies at seven days after incubation (anti-IL-1 alpha, 25.0 +/- 2.3%; anti-TNF alpha, 41.7 +/- 3.7%; anti-IL-1 alpha + anti-TNF alpha, 40.5 +/- 1.3%; and control, 100%), and the ratio of mononuclear to multinuclear cells had changed (anti-IL-1 alpha, 90:10; anti-TNF alpha, 75:25; anti-IL-1 alpha+ anti-TNF alpha, 88:12; and control, 60:40). The total pit areas per dentin slice also decreased with the addition of antibodies (anti-IL-1 alpha, 28,828; anti-TNF alpha, 49,249; anti-IL-1 alpha + anti-TNF alpha, 30,685; and control, 303,139 mm2). These results suggest that local production of IL-1 alpha and TNF alpha by osteoclasts is an important mechanism for regulating the osteoclast differentiation and bone resorptive process.

A silane coupling agent, CF 3 (CF 2 ) 9 CH 2 CH 2 Si(NCO) 3 , prepared by hydrosilylation of trichlorosilane with CF 3 (CF 2 ) 9 CHCH 2 in the presence of hydrogen hexachloroplatinate(IV), followed by reaction with silver cyanate, was used for glass and bovine tooth surface modification. In ESCA of the modified glass surfaces, nitrogen atoms in the silane coupling agent bound to the surfaces were detected only at contamination levels. This ESCA revealed that neither urethane bonds nor NCO groups were present on the modified glass surfaces. Clear AFM images of the modified glass surfaces were also obtained. Though the contact angles of water on the modified glass surfaces were independent of the modification time (20 min or 4 h), the AFM images obtained show that the original glass-surface patterns of unevenness remained on the surfaces in the case of the short modification time (20 min), whereas they disappeared from the surfaces in the case of the long modification time (4 h). In the latter case (4 h), the silane coupling agent reacted sufficiently over the entire glass surface, and no region of unmodified glass surface was detected. The surfaces of bovine tooth specimens were modified with this silane coupling agent by spreading of the agent on the surfaces with a small brush followed by brief drying with a hair drier. The amounts of plaque adhering to the modified and unmodified bovine tooth surfaces after wearing of the specimens in a human oral cavity for five days were compared. The amounts of plaque adhering to these specimens in in vitro tests as determined from SEM images were also compared. It is expected that this silane coupling agent will be useful as a plaque-controlling surface modifier for enhancement of oral health.

To simulate hypersensitive dentin, the smear layer and dentinal plugs of bovine root dentin specimens were removed by immersion in 10% phosphoric acid, polishing with hydroxyapatite particles, and ultrasonic cleansing. The fluoride-tannic acid-lanthanum-apatite (FTLA) group was treated with acidulated phosphate fluoride (APF) containing tannic acid followed by rubbing with a paste of lanthanum chloride (LaCl3) and powdered apatite glass ceramics. The treated specimens were immersed in a remineralizing solution that mimics saliva for 6 weeks. The SEM observations revealed that the treated surfaces of the FTLA group were completely covered with fine spherical compounds and the dentinal tubules were occluded with plugs to a depth of about 3 microns. Fluoride and lanthanum were detected to a depth of over 20 microns by EPMA observation. After the remineralization, the surface of FTLA-treated specimen did not have any opened tubules and showed a remarkable increase in the number of fine spherical deposits in the dentinal tubules. These results suggest that the reaction products produced by sequential treatment with acidic fluoride and LaCl3 and powdered apatite glass ceramics are able to effectively occlude dentinal tubules.

To investigate the laboratory anti-demineralization effect of a novel fluoride-releasing varnish containing surface reaction-type prereacted glass-ionomer (S-PRG) filler.Paired specimens were cut from bovine root dentin. One of each pair was used for the S-PRG group, and the other served as a control (n = 6). A 1 x 3 mm test surface was made on each specimen with the fluoride-releasing varnish. The novel fluoride-releasing varnish is categorized as a two-bottle-type self-etch adhesive. These liquids were mixed, applied on the test surface, and light-cured for 10 seconds. As a control, an S-PRG filler-free varnish was applied in the same manner. Each specimen was immersed in 8% methylcellulose gel demineralization system (1.5 mM Ca, 0.9 mM PO4, 0.1 M acetic acid, pH 5.0) for 7 days at 3 degrees C. The mineral profiles and integrated mineral loss (IML) of the lesions were obtained by transversal microradiography and analytical software.The S-PRG group exhibited significantly thicker surface layer than the control group. Furthermore, the S-PRG group showed significantly lower IML (3,459 vol% xmicropm) than the control group (4,687 vol% xmicropm) ( P0.05, Welch's two-sample t-test). The novel fluoride-releasing varnish increased acid resistance of root dentin in the vicinity of the coated surface.

Three novel aqueous fluoride surfactants (F4, F6, and F8) and a positive control (10F2S-3I) were applied to bovine enamel and the surface free energy was calculated by measuring the surface contact angles of three liquids: distilled water, α-bromonaphthalene and diiodomethane. The specimens were stored in water for 90 days, and then immersed in acetic acid/sodium acetate. The modified specimens recorded higher contact angles and lower surface free energy immediately after treatment than the control (p

われわれはMMP-3 (ストロメライシン) 様活性を持つ59KDのプロテアーゼ (59KDP-ase) を象牙質から抽出したことを報告してきた。この研究の目的は象牙質プロテオグリカン (PG) を抽出し, 59KP-aseによる分解特性を調べることである。PGはウシ象牙質から4MGuHCl (Gl-ext), 0.5MEDTA (E-ext) と4MGuHCL (G2-ext) を用いて連続抽出した。各抽出画分からDEAES ephacel, Superose6とReversed phasedFPLCを用いてPGを精製した。これらPGの分子量はSDS/PAGE電気泳動でGl-extは150～180KDと300KD, E-extは130～150KD, またG2-extは130～150KDと180～210KDであった。59KP-aseはこれらすべてのPGとそのコアプロテインを分解した。さらにE-extのPGがステインズオール染色で青色に染色される数種類のコアプロテインを有することが確認でき, またこれらのコアプロテインはすべて59KP-aseによって分解された。これらの研究から, 数種類の分子量が異なる無機質結合型のPGが象牙質中に存在し, また59KPaseがMMP-3様活性を有していることが証明されたことから, 59KP-aseは象牙質の石灰化と成熟の過程において重要な役割をしている可能性が示唆された。

To address the molecular mechanisms of cementogenesis, we have isolated dental follicle cells and examined them to see if they contain cementoblast progenitors. Dental follicle tissue was dissected from the root surface of bovine tooth germ and cells were released by digestion with bacterial collagenase. The released cells were maintained as a bovine dental follicle cells (BDFC). To elucidate the differentiation capacity of BDFC, they were transplanted into severe combined immunodeficiency (SCID) mice for 4 weeks. Transplanted BDFC formed cementumlike matrix; in contrast, bovine alveolar osteoblast (BAOB) transplants formed bonelike matrix, and bovine periodontal ligament cells (BPDL) formed a small amount of the cementumlike matrix. Immunohistochemical analysis showed that cementumlike matrix was positive for anti-cementum attachment protein monoclonal antibody, whereas bone-like matrix was negative. These results indicated that the BDFC contained cementoblast progenitors that were able to differentiate to cementoblasts in vivo. They also indicated that the BDFC are phenotypically distinct from BAOB and BPDL, and provide a useful model for investigating molecular mechanisms of cementogenesis.

This research was originally published in the Journal of Biological Chemistry. Masahiro Saito, Misaki Kurokawa, Masahito Oda, Masamitsu Oshima, Ko Tsutsui, Kazutaka Kosaka, Kazuhisa Nakao, Miho Ogawa, Ri-ichiroh Manabe, Naoto Suda, Ganburged Ganjargal, Yasunobu Hada, Toshihide Noguchi, Toshio Teranaka, Kiyotoshi Sekiguchi, Toshiyuki Yoneda and Takashi Tsuji. ADAMTSL6β Protein Rescues Fibrillin-1 Microfibril Disorder in a Marfan Syndrome Mouse Model through the Promotion of Fibrillin-1 Assembly. J. Biol. Chem. 2011; 286: 38602-38613 © the American Society for Biochemistry and Molecular Biology, Marfan syndrome (MFS) is a systemic disorder of the connective tissues caused by insufficient fibrillin-1 microfibril formation and can cause cardiac complications, emphysema, ocular lens dislocation, and severe periodontal disease. ADAMTSL6β (A disintegrin-like metalloprotease domain with thrombospondin type I motifs-like 6β) is a microfibril-associated extracellular matrix protein expressed in various connective tissues that has been implicated in fibrillin-1 microfibril assembly. We here report that ADAMTSL6β plays an essential role in the development and regeneration of connective tissues. ADAMTSL6β expression rescues microfibril disorder after periodontal ligament injury in an MFS mouse model through the promotion of fibrillin-1 microfibril assembly. In addition, improved fibrillin-1 assembly in MFS mice following the administration of ADAMTSL6β attenuates the overactivation of TGF-β signals associated with the increased release of active TGF-β from disrupted fibrillin-1 microfibrils within periodontal ligaments. Our current data thus demonstrate the essential contribution of ADAMTSL6β to fibrillin-1 microfibril formation. These findings also suggest a new therapeutic strategy for the treatment of MFS through ADAMTSL6β-mediated fibrillin-1 microfibril assembly.

Fourteen novel silane coupling agents with a quaternary ammonium group introduced into the molecule, [CH(2)=CHCH(2)N(+)(CH(3))(C(n)H(2n+1))(CH(2))(3)Si(OCH(3))(3)]X(-) (n-X, n=10-18, X=Cl, Br, or I), were synthesized with the aim of using these agents to make material surfaces antimicrobial for a long period of time. Measurements of the minimum inhibitory concentrations (MIC) against 12 kinds of fungi and bacteria revealed that the coupling agent with a C(10) alkyl chain and Cl- or Br- counter-ion (10-Cl or 10-Br) has the highest antimicrobial activity among the n-X agents synthesized, but 14-Cl and 14-Br showed the highest activity on a modified porcelain plate.

OBJECTIVE: This study tested the hypothesis that the benefit of fluoride-releasing restorative materials continues even after their reserve of fluoride has been depleted.MATERIALS AND METHODS: Pits in perspex blocks simulating cavities were filled with either a fluoride-releasing or a non-fluoride-releasing restorative material and a dentin single-section was placed 1 mm from the edge of the filled pit. These combinations were exposed to an acid gel system. Each demineralized dentin section was separated from the adjacent material and immersed in fresh demineralizing solutions. Transversal microradiographs were taken following the two experimental periods. This study defined ΔΔZ as the increase of integrated mineral loss (ΔZ) during the second acid attack.RESULTS: The first acid attack substantially demineralized the near-surface region (depth < 40 μm) in all samples. The second acid attack, however, did not cause further demineralization in this near-surface region. Instead, it demineralized dentin deeper than 40-60 μm. The ΔΔZ of the material that did not release fluoride was significantly greater than that of fluoride-releasing materials. Negative correlations were found between ΔΔZ and the mineral volume% of the near-surface region and lesion body of the initial lesions. These results indicate that the dentin mineral in the near-surface region is chemically altered to become acid-resistant fluorapatite. In addition, lesion progression during the second period of demineralization, which was fluoride-free, may have been affected for the materials that have high mineral content of the surface layer and lesion body.CONCLUSIONS: It is concluded that dentin surrounding fluoride-releasing materials is protected against demineralization even after the fluoride release has diminished.

Silane coupling agent containing fluorocarbon chain, (1H,1H,2H,2H-henicosafluorododecyl)trimethoxysilane, was applied for the surface modification of some dental materials. The contact angles of water and oleic acid on the modified material surfaces were increased. The modified denture teeth worn in an oral cavity displayed a long-range stain-protecting ability.

enzymogram上におけるみかけの分子量は, ゲル基質の濃度によって異なることが判明した。われわれは, この変化は電気泳動中の酵素と基質の親和性に基づく泳動速度の遅延が原因であり, ミカエリス定数 (Km) 値に依存するのではないかと推察し, それに基づいて電気泳動上での真の分子量を推定する関係式を検討した。みかけの分子量から真の分子量を推定する方法としてゼラチン濃度を変化させ, その関係を調べたところ, 基質濃度およびKm値により導き出される関係式〓が成り立つことが示唆された。したがって本法は, 基質存在下で分子量が本来の値より大きく算出されてしまうenzymogramで, その基質濃度に依存しない, 基質が存在しないところの分子量を導き出す方法である。さらに, 二次元enzymographyを施行したところ, より正確な分子量を導き出せることが判明した。またこうした解析によって分子量の推定のみならず複数の酵素が共存している時でもそれぞれの酵素の基質に対するKm値を求めることが理論的に可能となった。

The flexural strength, flexural modulus, and the amount of fluoride released from four experimental denture base resins containing 5, 10, 20 and 30 wt% surface pre-reacted glass-ionomer (S-PRG) filler added to the powder were evaluated. The mean flexural strength of the experimental resins, except the 30 wt%, and the flexural modulus of all the resins, complied with ISO 1567 requirements. In the 20 wt% resin, the amount of fluoride released in the initial phase was 1.88 microg/cm2/day, after which the level decreased. After recharging in a 9000 ppm fluoride solution for eight hours, the level of released fluoride increased markedly to 40.21 microg/cm2/16 hrs. Our results show that fluoride levels increased as a function of the S-PRG filler content. After the almost completely discharged resins were recharged, similar fluoride release occurred again. These results suggest that denture base resins containing S-PRG filler have great recharge and release capabilities which may assist in preventing root caries of abutment teeth.

The demineralization of dentin was studied when placed adjacent to one of four experimental denture base-resins. These experimental resins contained polymethylmethacrylate (PMMA) and 0, 5, 10, 20 or 30 wt% surface reaction-type prereacted glass-ionomer (S-PRG) filler, respectively. A dentin thin-section was sandwiched between these resins and subjected to demineralization for 1 wk. Microradiographic analyses showed that the mineral vol% of the surface was increased, and lesion formation was inhibited, in a dose-response relationship with the amount of S-PRG filler. Moreover, the mineral loss decreased, by up to 60%, with an increasing amount of filler. These results indicate that denture base-resins containing S-PRG filler possess a substantial inhibition capability against demineralization, and this may thereby assist in preventing root caries of abutment teeth.

ヒト象牙質より内在性プロテァーゼの抽出を試み, その諸性質をSDS/PAGEおよびenzymographyを用いて既知のプロテアーゼと比較検討した。ゼラチンおよびカゼインenzymogramにおいて乳歯象牙質PBS可溶性分画中にMr-59Kのプロテァーゼ (59KP-ase) をメインバンドとする数種類のプロテアーゼが確認された。59KP-aseはヘパリン, ゼラチンアフィニティーカラムを用いることによリゼラチンカラムから1MのNaClで溶出される分画として部分精製された。至適pHは9付近でありEDTAおよびPMSFにより活性が阻害された。分子量はトリプシンや動物性および細菌性コラゲナーゼとは異なっていた。また, このプロテァーゼはオステオポンチンに対する分解能を有していた。以上のことからヒト象牙質中に見出された分子量59Kのプロテアーゼは従来より報告されているようなコラゲナーゼとは異なったメタロプロテアーゼおよびセリンプロテァーゼとしての性格を有する象牙質に特有なリン蛋白分解酵素である可能性が示唆された。

Quaternary ammonium salts are frequently used as antibacterial agent that disrupts cell membrane through the binding of their ammonium cations to anionic sites in the outer layer tissue of bacteria. This article describes the synthesis of quaternary ammonium salt-type antibacterial agents with a phosphate group that strongly binds to hydroxyapatite and bromide ion as counterion. Evaluation of the antibacterial activity of the synthesized compounds in terms of minimum inhibitory concentration (MIC test) showed that the compounds exhibit an excellent antibacterial activity on a variety of bacteria including Gram-positive and Gram-negative bacteria, yeast, and fungi.

This study evaluated the effectiveness of a simplified silica coating method (CoJet System) on the bonding strength of resin cements to dental alloy. Bonding strength of the specimens treated with metal primer after alumina sandblasting was compared with those treated with silica coating and silane coupling agent after alumina sandblasting. Furthermore, the influence of silane coupling agent on bonding strength was compared between one-liquid and two-liquid silane coupling agents. Measurement of shear bond strength before and after thermal cycling revealed that the group treated with silica coating in one step without alumina sandblasting yielded high bonding strength. As for the influence of silane coupling agent, treatment with two-liquid silane coupling agent achieved higher mean shear bond strength than with one-liquid silane coupling agent. Findings in this study indicated that silicatization by means of this simplified silica coating method was effective in improving the bonding strength to dental alloy.

Six silane coupling agents having amide group (biosilanes) were synthesized with the aim to construct the material surface that allows cells to be compatible with it without their destruction. These agents were expected to make a soft landing to cytoplasm through the hydrogen bonding between their amide groups and cells. Evaluations of cell affinity using glass substrates modified with the synthesized biosilanes revealed that many cells remain on the modified glass plate. In addition, the implantation into the body of immunodeficient mouse of a composite material composed of porous hydroxyapatite and osteoblast showed the formation of a bone-like structure.

Specialized connective tissues such as tendon/ligament develop through a series of events that require temporal and spatial expression of numerous genes in mesenchymal progenitors. However, the genes required for tendon/ligament development have not been identified yet. To solve this problem, we made a cDNA library from periodontal ligament and sequenced 11,520 cDNA clones, as a model for investigating tendon/ligament development. The resulting sequence data was assembled to 617 expressed sequence tag (EST) clusters, and an EST database for human periodontal ligament (PDL) was constructed (designated as the KK-Periome database). In the KK-Periome database, the top 13 EST clusters were related to extracellular matrix (ECM) genes. The temporal and spatial expression patterns of these genes during mouse PDL development were examined by in situ hybridization. Among these genes, F-spondin was expressed specifically in dental follicle (DF) cells during tooth germ development, whereas tenascin-N was strongly expressed in the terminally differentiated PDL. This characteristic expression profile was confirmed by in vivo differentiation assay of human PDL (hPDL) cells in the mouse transplant. Thus, the KK-Periome database was proven to be a useful resource for PDL-derived ESTs (transcriptome), and in fact, initial evidence indicated that F-spondin and tenascin-N might serve as markers for DF and PDL, respectively.

This study evaluated the capability of a novel fluoride-releasing, all-in-one adhesive system on forming inhibition layer (radio-opaque layer) as compared with other adhesive systems. Dentin surface was treated with Imperva bond (IB), FL-BOND (FB), Reactmer bond (RE), or FL-BOND S-1 (FS) (which is a novel system). Untreated specimens were categorized as nonbonding group (NB). After storing for 10 days in de-ionized water, the specimens were cut into halves perpendicularly to the pulp chamber and immersed in a buffered demineralizing solution for four days. Longitudinal sections were cut and microradiographed. The width of inhibition layers adjacent to the adhesive surface--at a depth of 50 microm under the demineralization surface--was analyzed. Microradiography revealed distinct inhibition layers adjacent to the experimental surfaces of FB, RE, and FS. No inhibition layers were observed in NB and IB. In particular, the width of the inhibition layer of FS (12.5 microm) was significantly greater than those of FB and RE. These results indicated that a newly developed all-in-one adhesive system, FS, may have a superior ability of forming inhibition layers adjacent to cavity walls, and that it may also protect dentin against further demineralization in case of secondary marginal caries.

The purpose of this study was to evaluate the durability of tubules occluded with FTLA treatment by toothbrush abrasion test on the applied surface and by measuring fluoride release from the FTLA components. Dentin specimens with simulated hypersensitive surfaces were treated with APF containing tannic acid. After which, the specimens received lanthanum-chloride-with-powdered-fluoroapatite-glass-ceramics treatment. The specimens were subjected to toothbrush abrasion test up to 6,000 strokes. SEM observation revealed that dentinal tubules of the FTLA treated specimens were completely occluded with fine deposits even after toothbrush abrasion of 6,000 strokes. EPMA analysis revealed that fluoride, lanthanum, and aluminum were the main FTLA components on the dentin surface after 6,000-stroke abrasion. To measure fluoride release from the FTLA components, a slurry was enclosed in a cellulose tube and suspended in deionized water at 37 degrees C. After fluoride was dialyzed against deionized water, a high concentration of fluoride was found to be released from FTLA the components, indicating FTLA treatment's prominent durability. These results suggested that FTLA treatment has a superior resistance against toothbrush abrasion and a high fluoride-releasing performance. These characteristics lend much weight to showing that the FTLA method is an effective and durable medicament for dentin hypersensitivity.

This study evaluated clinical and radiographic healing of 236 root-canal treatments in 131 cases obturated with the Obtura II system. One operator performed all canal preparation and obturation with sealer. A standardized apical-coronal preparation technique instrumented all canals. Clinical symptoms, periodontal condition, and radiographic findings were evaluated at 3, 6, and 12 months. Radiographs taken immediately postobturation were compared to recall radiographs. The level of the final root filling was classified as short (more than 2 mm short of the apex), flush (within 2 mm), or over (beyond) in 12.7%, 81.4%, and 5.9% of cases, respectively. More than 96% of cases were treated successfully by the Obtura II system. Where roots were filled flush, over, or short, lesions healed in 97%, 93%, and 93% of cases, respectively, with no significant differences (p < 0.05). Root filling excess had no impact on the healing process.

The purpose of this study was to examine the application of an ultrasonic pulse-reflection technique for the evaluation of adhesive defects. First, the sonic velocities in the enamel and dentin of human molars and bovine incisors were measured with a pulsar receiver attached to an ultrasonic transducer. The identification of the dentino-enamel junction and pulp-dentin interface using the ultrasonic method based on intrinsic sonic velocities showed good agreement with the actual measured thicknesses. Next, a cemented restoration with artificial faults was prepared. Half of the Au-Ag-Pd alloy plate area was cemented to the dentin slab using luting resin cement. The adhesive interface was evaluated with a high-resolution ultrasonic imaging system. Clear internal faults were evident from the ultrasonic tomogram. The findings of this study suggest that the ultrasonic pulse-reflection technique may be useful for inspecting and imaging structural defects of adhesive interfaces.

Dental follicle is the fibrous tissue that surrounds the developing tooth germ, and it is believed to contain progenitors for cementoblasts, periodontal ligament cells, and osteoblasts. In this study, we report the presence of cementoblast progenitors in cultures of bovine dental follicle cells and demonstrate their differentiation capacity. Bovine dental follicle cells (BDFC) obtained from tooth germs by collagenase digestion were compared with bovine alveolar bone osteoblasts (BAOB) and bovine periodontal ligament cells (BPDL) in vitro and in vivo. In culture, BDFC exhibited low levels of alkaline phosphatase activity and expressed mRNA for osteopontin (OP) and type I collagen (COLI), as well as low levels of osteocalcin (OC) mRNA. In contrast, cultured BAOB exhibited high alkaline phosphatase activity levels and expressed mRNA for OC, OP, COLI, and bone sialoprotein (BSP). To elucidate the differentiation capacity of BDFC in vivo, cells were transplanted into severe combined immunodeficiency (SCID) mice and analyzed after 4 weeks. Transplanted BDFC formed fibrous tissue and cementum-like matrix, which stained positive for anti-cementum attachment protein (CAP) monoclonal antibody (3G9), and expressed mRNA for OC, OP, COLI, and BSP. On the other hand, transplanted BAOB formed bone-like matrix, but were negative for anti-CAP monoclonal antibody. The BPDL transplants formed fibrous tissue that contained a few cells expressing CAP. These results indicate that cementoblast progenitors are present in BDFC, which can provide a useful model for investigating the molecular mechanisms of cementogenesis.

In the serum-free culture medium of bovine odontoblasts we detected active gelatinolytic metalloproteinases, matrix metalloproteinase (MMP)-2 and MMP-9 (gelatinases A and B). The activity of MMP-2, in particular, appeared suddenly around day 21 in the culture, coinciding with the development of odontoblastic cell processes and the loss of alkaline phosphatase. Reverse transcriptase-polymerase chain reaction analysis of these odontoblasts demonstrated that messages of MMP-2 but not MMP-9 increased significantly between day 15 and day 21. The in vitro observation indicates that medium conditioned by these odontoblasts and containing significant amounts of MMP-2 degrades not only the collagenous substrates but also purified dentin phosphophoryn as well. We have also observed that dephosphorylated dentin phosphoprotein becomes a better substrate for casein kinase II after limited proteolysis with MMP-2. These results support our working hypothesis that MMP-2-mediated proteolytic processing is an important step in accelerating the process of dentin matrix maturation, which includes phosphorylation and subsequent mineralization. As has been suggested previously, extracellular phosphorylation of matrix proteins is an important step in biomineralization both in bone and in dentin (Mikuni-Takagaki et al., J Bone Miner Res 1995;10:231–42; Zhu et al., Biochem J 1997;323:637–43). Our present histochemical analysis in MMP-2 knockout mice confirms the concept with the delayed formation of mineralized tissues, dentin, and bone.

Silane coupling agents containing a fluorocarbon chain were prepared in high yields. It was found that silanes can be useful modifiers of the surfaces of glass, metals, and resin composites for dental use. The silane coupling agent CF3(CF2)9CH2CH2Si(OCH3)3 was the best modifier of these surfaces in terms of water and oil repellency. Colorants and experimental bacterial plaque detached much more easily from, and adhered less well to, surfaces modified with this silane coupling agent compared with unmodified surfaces. The surfaces of four teeth of a denture were modified with this silane coupling agent by spreading the agent on the surfaces with a small brush followed by brief drying with a hair drier. The modified tooth surfaces of the denture, which was worn for four months in a heavy smoker's oral cavity, were more stain-resistant than the unmodified tooth surfaces. It is expected that silane coupling agents containing a fluorocarbon chain will be useful surface modifiers for enhancement of oral health.