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ABSTRACT Objective This study aimed to investigate the potential role of CAMK II pathway in the compression-regulated OPG expression in periodontal ligament cells (PDLCs). Material and Methods The PDL tissue model was developed by 3-D culturing human PDLCs in a thin sheet of poly lactic-co-glycolic acid (PLGA) scaffolds, which was subjected to static compression of 25 g/cm2 for 3, 6 and 12 h, with or without treatment of KN-93. After that, the expression of OPG, RANKL and NFATC2 was investigated through real-time PCR and western blot analysis. Results After static compression, the NFATC2 and RANKL expression was significantly up-regulated, while partially suppressed by KN-93 for 6 and 12 h respectively. The OPG expression was significantly down-regulated by compression in 3 h, started to elevate in 6 h, and significantly up-regulated in 12 h. The up-regulation after 12 h was significantly suppressed by KN-93. Conclusions Long-term static compression increases OPG expression in PDLCs, at least partially, via the CAMK II pathway.

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ABSTRACT Low pressure Pain Threshold (PPT) is considered a risk factor for Temporomandibular Disorders (TMD) and is influenced by psychological variables. Objectives To correlate deep pain sensitivity of masticatory muscles with prosthetic factors and Oral-Health-Related Quality of Life (OHRQoL) in completely edentulous subjects. Material and Methods A total of 29 complete denture wearers were recruited. The variables were: a) Pressure Pain Threshold (PPT) of the masseter and temporalis; b) retention, stability, and tooth wear of dentures; c) Vertical Dimension of Occlusion (VDO); d) Oral Health Impact Profile (OHIP) adapted to orofacial pain. The Kolmogorov-Smirnov test, the Pearson Product-Moment correlation coefficient, the Spearman Rank correlation coefficient, the Point-Biserial correlation coefficient, and the Bonferroni correction (α=1%) were applied to the data. Results The mean age (standard deviation) of the participants was of 70.1 years (9.5) and 82% of them were females. There were no significant correlations with prosthetic factors, but significant negative correlations were found between the OHIP and the PPT of the anterior temporalis (r=-0.50, 95% CI-0.73 to 0.17, p=0.005). Discussion The deep pain sensitivity of masticatory muscles in complete dentures wearers is associated with OHRQoL, but not with prosthetic factors.

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ABSTRACT Objective The purpose of this randomized, cross-over, in situ study was to determine the remineralization of demineralized dentin specimens after the application of a 10% fluoride (F-) or a 1% chlorhexidine–1% thymol (CHX–thymol) varnish. Material and Methods Twelve individuals without current caries activity wore removable appliances in the lower jaw for a period of four weeks. Each appliance contained four human demineralized dentin specimens fixed on the buccal aspects. The dentin specimens were obtained from the cervical regions of extracted human third molars. After demineralization, half the surface of each specimen was covered with a nail varnish to serve as the reference surface. The dentin specimens were randomly assigned to one of the three groups: F-, CHX–thymol, and control (no treatment). Before the first treatment period and between the others, there were washout periods of one week. After each treatment phase, the changes in mineral content (vol% µm) and the lesion depths (µm) of the dentin slabs were determined by transverse microradiography (TMR). Data analysis was accomplished by the Kruskal-Wallis test and the Mann-Whitney U test (p<0.05). Results The medians (25th/75th percentile) of integrated mineral loss were 312.70 (203.0-628.7) for chlorhexidine varnish, 309.5 (109.8-665.8) for fluoride varnish, and -346.9 (-128.7 - -596.0) for the control group. The medians (25th/75th percentile) of lesion depth were 13.6 (5.7-34.5) for chlorhexidine varnish, 16.5 (5.6-38.1) for fluoride varnish, and -14.2 (-4.5- -32.9) for the control group. Use of the 10% F- or 1% CHX–1% thymol varnishes resulted in significantly decreased mineral loss and lesion depth in dentin when compared with the control group. There were no statistically significant differences among the test groups. Conclusions Within the limitations of this study, the results suggest that the effect of the treatment of demineralized dentin with 10% F- or 1% CHX–1% thymol is better than without any treatment.

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ABSTRACT Dental composites cured at high temperatures show improved properties and higher degrees of conversion; however, there is no information available about the effect of pre-heating on material degradation. Objectives This study evaluated the effect of pre-heating on the degradation of composites, based on the analysis of radiopacity and silver penetration using scanning electron microscopy/energy-dispersive X-ray spectroscopy (SEM/EDS). Material and Methods Thirty specimens were fabricated using a metallic matrix (2x8 mm) and the composites Durafill VS (Heraeus Kulzer), Z-250 (3M/ESPE), and Z-350 (3M/ESPE), cured at 25°C (no pre-heating) or 60°C (pre-heating). Specimens were stored sequentially in the following solutions: 1) water for 7 days (60°C), plus 0.1 N sodium hydroxide (NaOH) for 14 days (60°C); 2) 50% silver nitrate (AgNO3) for 10 days (60°C). Specimens were radiographed at baseline and after each storage time, and the images were evaluated in gray scale. After the storage protocol, samples were analyzed using SEM/EDS to check the depth of silver penetration. Radiopacity and silver penetration data were analyzed using ANOVA and Tukey’s tests (α=5%). Results Radiopacity levels were as follows: Durafill VS<Z-350<Z-250 (p<0.05). The depth of silver penetration into the composites ranked as follows: Durafill VS>Z-350>Z-250 (p<0.05). After storage in water/NaOH, pre-heated specimens presented higher radiopacity values than non-pre-heated specimens (p<0.05). There was a lower penetration of silver in pre-heated specimens (p<0.05). Conclusions Pre-heating at 60°C mitigated the degradation of composites based on analysis of radiopacity and silver penetration depth.

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ABSTRACT In Aggregatibacter actinomycetemcomitans, different serotypes have been described based on LPS antigenicity. Recently, our research group has reported a differential immunogenicity when T lymphocytes were stimulated with these different serotypes. In particular, it was demonstrated that the serotype b of A. actinomycetemcomitans has a stronger capacity to trigger Th1- and Th17-type cytokine production. Objective This study aimed to quantify the expression of different CC chemokines (CCLs) and receptors (CCRs) in T lymphocytes stimulated with the different A. actinomycetemcomitans serotypes. In addition, the expression of the transcription factors T-bet, GATA-3, RORC2, and Foxp3, master-switch genes implied in the Th1, Th2, Th17, and T-regulatory differentiation, respectively, was analyzed in order to determine T-cell phenotype-specific patterns of CCL and CCR expression upon A. actinomycetemcomitans stimulation. Material and Methods Human naïve CD4+ T lymphocytes were obtained from healthy subjects and stimulated with autologous dendritic cells primed with the different A. actinomycetemcomitans serotypes. The expression levels for the chemokines CCL1, CCL2, CCL3, CCL5, CCL11, CCL17, CCL20, CCL21, CCL25, and CCL28, as well as the chemokine receptors CCR1, CCR2, CCR3, CCR4, CCR5, CCR6, CCR7, CCR8, CCR9, and CCR10 were quantified by qPCR. Similarly, the expression levels for the transcription factors T-bet, GATA-3, RORC2, and Foxp3 were quantified and correlated with the CCL and CCR expression levels. Results Higher expression levels of CCL2, CCL3, CCL5, CCL20, CCL21, CCL28, CCR1, CCR2, CCR5, CCR6, CCR7, and CCR9 were detected in T lymphocytes stimulated with the serotype b of A. actinomycetemcomitans compared with the other serotypes. In addition, these higher expression levels of CCLs and CCRs positively correlated with the increased levels of T-bet and RORC2 when T lymphocytes were stimulated with the serotype b. Conclusion A T-lymphocyte response biased towards a Th1- and Th17-pattern of CCL and CCR expression was detected under stimulation with the serotype b of A. actinomycetemcomitans.

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Objectives To compare three methods of intratubular contamination that simulate endodontic infections using confocal laser scanning microscopy (CLSM).Material and Methods Two pre-existing models of dentinal contamination were used to induce intratubular infection (groups A and B). These methods were modified in an attempt to improve the model (group C). Among the modifications it may be included: specimen contamination for five days, ultrasonic bath with BHI broth after specimen sterilization, use of E. faecalis during the exponential growth phase, greater concentration of inoculum, and two cycles of centrifugation on alternate days with changes of culture media. All specimens were longitudinally sectioned and stained with of LIVE/DEAD® for 20 min. Specimens were assessed using CLSM, which provided images of the depth of viable bacterial proliferation inside the dentinal tubules. Additionally, three examiners used scores to classify the CLSM images according to the following parameters: homogeneity, density, and depth of the bacterial contamination inside the dentinal tubules. Kruskal-Wallis and Dunn’s tests were used to evaluate the live and dead cells rates, and the scores obtained.Results The contamination scores revealed higher contamination levels in group C when compared with groups A and B (p<0.05). No differences were observed between group A and B (p>0.05). The volume of live cells in group C was higher than in groups A and B (p<0.05).Conclusion The new protocol for intratubular infection resulted in high and uniform patterns of bacterial contamination and higher cell viability in all specimens when compared with the current methods.

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ABSTRACT Objective The aim of this study was to investigate the in vitro and in vivo biological responses to nanostructured carbonated hydroxyapatite/calcium alginate (CHA) microspheres used for alveolar bone repair, compared to sintered hydroxyapatite (HA). Material and Methods The maxillary central incisors of 45 Wistar rats were extracted, and the dental sockets were filled with HA, CHA, and blood clot (control group) (n=5/period/group). After 7, 21 and 42 days, the samples of bone with the biomaterials were obtained for histological and histomorphometric analysis, and the plasma levels of RANKL and OPG were determined via immunoassay. Statistical analysis was performed by Two-Way ANOVA with post-hoc Tukey test at 95% level of significance. Results The CHA and HA microspheres were cytocompatible with both human and murine cells on an in vitro assay. Histological analysis showed the time-dependent increase of newly formed bone in control group characterized by an intense osteoblast activity. In HA and CHA groups, the presence of a slight granulation reaction around the spheres was observed after seven days, which was reduced by the 42nd day. A considerable amount of newly formed bone was observed surrounding the CHA spheres and the biomaterials particles at 42-day time point compared with HA. Histomorphometric analysis showed a significant increase of newly formed bone in CHA group compared with HA after 21 and 42 days from surgery, moreover, CHA showed almost 2-fold greater biosorption than HA at 42 days (two-way ANOVA, p<0.05) indicating greater biosorption. An increase in the RANKL/OPG ratio was observed in the CHA group on the 7th day. Conclusion CHA spheres were osteoconductive and presented earlier biosorption, inducing early increases in the levels of proteins involved in resorption.

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ABSTRACT Objective The purpose of this in vitro study was to compare the efficacy of a bleaching toothpaste containing Blue Covarine vs. conventional tooth bleaching techniques using peroxides (both in-office and at-home). Material and Methods Samples were randomly distributed into five experimental groups (n=15): C - Control; BC – Bleaching toothpaste containing Blue Covarine; WBC – Bleaching toothpaste without Blue Covarine; HP35 - In-office bleaching using 35% hydrogen peroxide; and CP10 – At-home bleaching with 10% carbamide peroxide. The dental bleaching efficacy was determined by the color difference (ΔE), luminosity (ΔL), green-red axis (Δa), and blue-yellow axis (Δb). The CIELab coordinates were recorded with reflectance spectroscopy at different times: T0 - baseline, T1 – immediately after bleaching, T2 - 7 days, T3 - 14 days, and T4 - 21 days after the end of treatments. Data were analyzed by a repeated measures mixed ANOVA and post hoc Bonferroni test, with a significance level of 5%. Results No significant differences were found between the treatment groups C, BC, and WBC. The groups HP35 and CP10 showed significantly higher whitening efficacy than groups C, BC, and WBC. Conclusions There were no significant differences in the whitening efficacy between a Blue Covarine containing toothpaste, a standard whitening toothpaste, and a control. Neither of the whitening toothpastes tested were as effective as in-office or at-home bleaching treatments.

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ABSTRACT Plasma technology has the potential to improve the adherence of fibers to polymeric matrices, and there are prospects for its application in dentistry to reinforce the dental particulate composite. Objectives This study aimed to investigate the effect of oxygen or argon plasma treatment on polyethylene fibers. Material and Methods Connect, Construct, InFibra, and InFibra treated with oxygen or argon plasma were topographically evaluated by scanning electron microscopy (SEM), and chemically by X-ray photoelectron spectroscopy (XPS). For bending analysis, one indirect composite (Signum) was reinforced with polyethylene fiber (Connect, Construct, or InFibra). The InFibra fiber was subjected to three different treatments: (1) single application of silane, (2) oxygen or argon plasma for 1 or 3 min, (3) oxygen or argon plasma and subsequent application of silane. The samples (25x2x2 mm), 6 unreinforced and 60 reinforced with fibers, were subjected to three-point loading tests to obtain their flexural strength and deflection. The results were statistically analyzed with ANOVA and the Bonferroni correction for multiple comparison tests. Results SEM analysis showed that oxygen and argon plasma treatments promote roughness on the polyethylene fiber surface. X-ray photoelectron spectroscopy (XPS) analysis shows that both plasmas were effective in incorporating oxygenated functional groups. Argon or oxygen plasma treatment affected the flexural strength and deflection of a fiber reinforced composite. The application of silane does not promote an increase in the flexural strength of the reinforced composites. Conclusions Oxygen and argon plasma treatments were effective in incorporating oxygenated functional groups and surface roughness. The highest strength values were obtained in the group reinforced with polyethylene fibers treated with oxygen plasma for 3 min.

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ABSTRACT The ability of hemostatic agents to promote bone repair has been investigated using in vitro and in vivo models but, up to now, the results are inconclusive. Objective In this context, the aim of this study was to compare the potential of bone repair of collagen sponge with fibrin glue in a rat calvarial defect model. Material and Methods Defects of 5 mm in diameter were created in rat calvariae and treated with either collagen sponge or fibrin glue; untreated defects were used as control. At 4 and 8 weeks, histological analysis and micro-CT-based histomorphometry were carried out and data were compared by two-way ANOVA followed by Student-Newman-Keuls test when appropriated (p≤0.05). Results Three-dimensional reconstructions showed increased bone formation in defects treated with either collagen sponge or fibrin glue compared with untreated defects, which was confirmed by the histological analysis. Morphometric parameters indicated the progression of bone formation from 4 to 8 weeks. Additionally, fibrin glue displayed slightly higher bone formation rate when compared with collagen sponge. Conclusion Our results have shown the benefits of using collagen sponge and fibrin glue to promote new bone formation in rat calvarial bone defects, the latter being discreetly more advantageous.

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ABSTRACT Objective The study aimed to evaluate effects of air abrasion with alumina or glass beads on bond strengths of resin cements to CAD/CAM composite materials. Material and Methods CAD/CAM composite block materials [Cerasmart (CS) and Block HC (BHC)] were pretreated as follows: (a) no treatment (None), (b) application of a ceramic primer (CP), (c) alumina-blasting at 0.2 MPa (AB), (d) AB followed by CP (AB+CP), and (e) glass-beads blasting at 0.4 MPa (GBB) followed by CP (GBB+CP). The composite specimens were bonded to resin composite disks using resin cements [G-CEM Cerasmart (GCCS) and ResiCem (RC)]. The bond strengths after 24 h (TC 0) and after thermal cycling (TC 10,000 at 4–60°C) were measured by shear tests. Three-way ANOVA and the Tukey compromise post hoc tests were used to analyze statistically significant differences between groups (α=0.05). Results For both CAD/CAM composite materials, the None group exhibited a significant decrease in bond strength after TC 10,000 (p<0.05). AB showed significantly higher bond strength after TC 10,000 than the None group, while CP did not (p<0.05). GBB exhibited smaller surface defects than did AB; however, their surface roughnesses were not significantly different (p>0.05). The AB+CP group showed a significantly higher bond strength after TC 10,000 than did the AB group for RC (p<0.05), but not for GCCS. The GBB+CP group showed the highest bond strength for both thermal cyclings (p<0.05). Conclusions Air abrasion with glass beads was more effective in increasing bond durability between the resin cements and CAD/CAM composite materials than was using an alumina powder and a CP.

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ABSTRACT To preserve oral health and to maintain the prosthetic devices, it is important not only to improve the properties of commonly known hygiene products, but also to investigate new materials with antimicrobial action. Objectives This study evaluated the antimicrobial activity of sodium hypochlorite (0.25% and 0.50%) and 10% Ricinus communis’ solutions against specific microorganisms. Material and Methods Sixty four maxillary complete denture wearers were instructed to brush their dentures three times a day and to soak them (20 min/day) in the solutions: SH1: 0.25% sodium hypochlorite; SH2: 0.5% sodium hypochlorite; RC: 10% R. communis oil; and C: 0.85% saline (control). The solutions were used for 7 days in a randomized sequence. Following each period of use, there was a 1-week washout period. Antimicrobial activity was determined by Colony Forming Units (CFU) counts of Streptococcus mutans, Candida spp., and gram-negative microorganisms. For collecting biofilm, the internal surface of maxillary dentures was brushed with saline solution, and biofilm suspension obtained. After dilutions (100 - 10-3), aliquots were seeded in Mitis salivarius, CHROMagar Candida®, and MacConkey agar for detecting S. mutans, Candida spp., or gram-negative microorganisms, respectively. After incubation, colonies were counted, and CFU/mL values were calculated. Then, transformation - log10 (CFU+1) - data were analyzed using the Friedman test (α=0.05). Results showed significant differences between the solutions (p<0.001). Results All three solutions showed antimicrobial activity against S. mutans. Against Candida spp., RC and SH1 solutions showed similar effect while SH2 showed superior activity. SH1 and SH2 solutions showed antimicrobial action against gram-negative microorganisms. The Candida species most frequently isolated was C. albicans, followed by C. tropicalis and C. glabrata. Conclusions The 0.5% sodium hypochlorite solution was the most effective and might be used to control denture biofilm. C. albicans was the most frequently isolated Candida sp.