Modulation of regenerative responses by retinoic and ascorbic acids in human apical papilla cells.

OBJECTIVE: This study investigated the bioactive effects of retinoic acid and ascorbic acid on hSCAPs in vitro. DESIGN: Cells were obtained from human third molars (n=4) and characterized for mesenchymal stem cell markers by flow cytometry. The experimental groups: control (α-MEM); vehicle control group (α-MEM + 0.17 % DMSO); retinoic acid 0.1, 1, and 10 µM; and ascorbic acid 3, 30, and 300 µM (n=8) were tested for cell viability (alamarBlue; 1, 3, and 7 days), total collagen synthesis (Sirius Red; 1 and 7 days), mineralized matrix formation (Alizarin red; 14 days), and the regulation of gene expression related to mineralization (ALPL and DSPP), cell migration (ITGAV and CXCL12) angiogenesis (VEGFA) and collagen synthesis (COL1A1 and COL3A1; RT-qPCR) on 1 and 7 days. ACTB and GAPDH were used as reference genes. Data were analyzed by ANOVA and complementary tests at a 5 % significance level. RESULTS: Ascorbic acid 300 µM increased viability, and retinoic acid reduced it dose-dependently. Retinoic acid 0.1 µM and ascorbic acid 30 and 300 µM increased mineralized matrix formation and total collagen synthesis, and retinoic acid 10 µM decreased. On day 1, 0.1 µM retinoic acid upregulated the gene expression of COL1A1, COL3A1, VEGFA, CXCL12, ALPL, DSPP e ITGAV, and 300 µM ascorbic acid upregulated COL1A1, COL3A1 and DSPP. However, on day 7, retinoic acid downregulated ALPL, COL3A1, CXCL12, and VEGFA and downregulated ITGAV and VEGFA. CONCLUSION: Retinoic acid 0.1 µM and ascorbic acid 300 µM biostimulated hSCAPs to differentiate into pro-regenerative phenotypes with potential application for REPs.

A new approach for professional dental bleaching using a polymeric catalyst primer.

OBJECTIVE: Evaluate the influence of a polymeric catalyst primer (PCP) on esthetic efficacy (EE), degradation kinetics of hydrogen peroxide (H2 O2 ), and trans-amelodentinal cytotoxicity (TC) of bleaching gels. MATERIALS AND METHODS: The following groups were established: G1: No treatment (NC, negative control); G2: PCP; G3: 10% H2 O2 ; G4: PCP + 10% H2 O2 ; G5: 20% H2 O2 ; G6: PCP + 20% H2 O2 ; G7: 35% H2 O2 (positive control); G8: PCP + 35% H2 O2 . To determine EE, enamel/dentin discs (E/DDs) were stained and subjected or not to bleaching protocols for 45 min. To assess TC, the E/DDs were adapted to artificial pulp chambers. The extracts (culture medium + gel components diffused through E/DDs) were applied to odontoblast-like MDPC-23 cells. The viability (VB), oxidative stress (OxS), morphology (SEM), amount of H2 O2 diffused and the production of hydroxyl radical (OH• ) were assessed (two-way ANOVA/Tukey/paired Student t-test; p < 0.05). RESULTS: The highest EE was found in G8 (p < 0.05), and G4, G6, and G7 did not differ statistically (p > 0.05). In G4, the limited H2 O2 diffusion reduced OxS and increased cell VB (p < 0.05). CONCLUSIONS: Coating the enamel with PCP containing 10 mg/ml of manganese oxide before applying the 10% H2 O2 bleaching gel maintains the EE of conventional in-office bleaching and minimizes the toxic effects of this esthetic therapy. CLINICAL SIGNIFICANCE: Coating the enamel with a PCP before applying the bleaching gel may potentiate the EE of the conventional in-office tooth bleaching and reduce the toxicity of this professional therapy to the dental pulp.

The influence of violet LED application time on the esthetic efficacy and cytotoxicity of a 35% H2O2 bleaching gel.

OBJECTIVE: To assess the potential influence of violet LED (V-LED) application time on the esthetic efficacy and cytotoxicity of a 35% H2O2 bleaching gel. METHODOLOGY: Stained and standardized enamel/dentin discs were subjected to one in-office tooth bleaching session (45 min), and the gel was either irradiated or not with V-LED. Thus, the following groups were established (n = 8): G1: No treatment (negative control, NC); G2: 35% H2O2 (positive control, PC); G3: 35%H2O2 + V-LED/15 min; G4: 35%H2O2 + V-LED/30 min; G5: 35%H2O2 + V-LED/45 min. First, esthetic efficacy was assessed (ΔE00 and ΔWI). Discs assembled in artificial pulp chambers were subjected to the same bleaching treatments. Then, the extracts (culture medium + diffused bleaching gel components) were collected and applied to MDPC-23 pulp cells, which were analyzed for viability (Live/Dead, MTT) and oxidative stress (OxS). The amount of H2O2 in the extracts was also determined (leuco crystal-violet/peroxidase). The data were subjected to ANOVA/Tukey at a 5% significance level. RESULTS: Although esthetic efficacy did not differ among the irradiated groups (G3, G4, and G5) (p > 0.05), their results were higher than in G2 (PC; p < 0.05). In the irradiated groups, the cell viability and OxS as well as the amount of H2O2 in the extracts were statistically similar to G2 (PC), regardless of irradiation time (p > 0.05). CONCLUSION: Although V-LED improves the esthetic outcome of in-office tooth bleaching, increasing irradiation time does not effect the color changes and cytotoxicity of this professional therapy.

Pro-inflammatory mediators expression by pulp cells following tooth whitening on restored enamel surface.

This paper aimed to assess the influence of adhesive restoration interface on the diffusion of hydrogen peroxide (H2O2), indirect toxicity, and pro-inflammatory mediators expression by odontoblast-like cells, after in-office tooth whitening. Dental cavities prepared in bovine enamel/dentin discs were adhesively restored and subjected or not to hydrolytic degradation (HD). A whitening gel with 35% H2O2 (WG) was applied for 45 min onto restored and non-restored specimens adapted to artificial pulp chambers giving rise to the groups: SD- intact discs (control); SD/HP- whitened intact discs; RT/HP- restored and whitened discs; and RT/HD/HP- restored and whitened discs subjected to HD. The extracts (culture medium + WG components diffused through enamel/dentin/restoration interface) were collected and applied to odontoblast-like MDPC-23 cells. The study evaluated the amount of H2O2 in the extracts, as well as the cell viability (CV), cell morphology (CM), and gene expression of inflammatory mediators (TNF-α and COX-2) by the pulp cells exposed to the extracts (ANOVA and Tukey tests; 5% significance). All whitened groups presented lower CV than SD (control; p<0.05). The highest CV reduction and gene expression of TNF-α and COX-2 was observed in the RT/HD/HP group in comparison with SD/HP and RT/HP (control; p<0.05). CM alterations occurred in all whitened groups. The intensity of these cell side effects was directly related with the amount of H2O2 in the extracts. We concluded that adhesive restoration of dental cavity increases the H2O2 diffusion after in-office whitening, enhancing the indirect toxicity of this therapy and trigger pro-inflammatory overexpression by MDPC-23 cells.

Pro-inflammatory mediators expression by pulp cells following tooth whitening on restored enamel surface

Abstract This paper aimed to assess the influence of adhesive restoration interface on the diffusion of hydrogen peroxide (H2O2), indirect toxicity, and pro-inflammatory mediators expression by odontoblast-like cells, after in-office tooth whitening. Dental cavities prepared in bovine enamel/dentin discs were adhesively restored and subjected or not to hydrolytic degradation (HD). A whitening gel with 35% H2O2 (WG) was applied for 45 min onto restored and non-restored specimens adapted to artificial pulp chambers giving rise to the groups: SD- intact discs (control); SD/HP- whitened intact discs; RT/HP- restored and whitened discs; and RT/HD/HP- restored and whitened discs subjected to HD. The extracts (culture medium + WG components diffused through enamel/dentin/restoration interface) were collected and applied to odontoblast-like MDPC-23 cells. The study evaluated the amount of H2O2 in the extracts, as well as the cell viability (CV), cell morphology (CM), and gene expression of inflammatory mediators (TNF-α and COX-2) by the pulp cells exposed to the extracts (ANOVA and Tukey tests; 5% significance). All whitened groups presented lower CV than SD (control; p<0.05). The highest CV reduction and gene expression of TNF-α and COX-2 was observed in the RT/HD/HP group in comparison with SD/HP and RT/HP (control; p<0.05). CM alterations occurred in all whitened groups. The intensity of these cell side effects was directly related with the amount of H2O2 in the extracts. We concluded that adhesive restoration of dental cavity increases the H2O2 diffusion after in-office whitening, enhancing the indirect toxicity of this therapy and trigger pro-inflammatory overexpression by MDPC-23 cells.

Resumo Este trabalho teve como objetivo avaliar a influência da interface de uma restauração adesiva na difusão do peróxido de hidrogênio (H2O2), toxicidade indireta e expressão de mediadores pró-inflamatórios por células odontoblastóides, após clareamento dental em consultório. Cavidades dentárias preparadas em discos de esmalte / dentina foram restauradas com adesivo e submetidas ou não à degradação hidrolítica (HD). Um gel clareador com 35% H2O2 (WG) foi aplicado por 45 min em discos restaurados e não restaurados adaptados às câmaras pulpares artificiais dando origem aos grupos: SD- discos intactos (controle); SD / HP - Discos intactos clareados; RT / HP - discos restaurados e clareados; e RT / HD / HP - discos restaurados, clareados e submetidos a HD. Os extratos (meio de cultura + componentes WG difundidos através da interface esmalte/dentina/restauração) foram coletados e aplicados em células odontoblastóides MDPC-23. Foi avaliada a quantidade de H2O2 nos extratos, bem como a viabilidade (CV), morfologia (CM) e expressão gênica de mediadores inflamatórios (TNF-α e COX-2) pelas células pulpares expostas aos extratos (ANOVA e testes de Tukey; 5% de significância). Todos os grupos clareados apresentaram menor CV do que SD (controle; p <0,05). A maior redução CV e expressão gênica de TNF-α e COX-2 foi observada no grupo RT / HD / HP em comparação com SD / HP e RT / HP (controle; p <0,05). Alterações na CM ocorreram em todos os grupos clareados. A intensidade desses efeitos celulares teve relação direta com a quantidade de H2O2 nos extratos. Concluímos que a presença de uma cavidade contendo restauração adesiva aumenta a difusão de H2O2 após o clareamento em consultório, o que, por sua vez, aumenta a toxicidade indireta dessa terapia e desencadeia a expressão de mediadores pró-inflamatórios pelas células pulpares MDPC-23.

Proliferation rate and expression of stem cells markers during expansion in primary culture of pulp cells.

The aim of the present study was to evaluate the proliferation rate and the expression of stem cells markers during expansion in primary culture of dental pulp stem cells (DPSCs), comparing different techniques (explant and enzymatic digestion), subject ages (up to 40 and over 40) and cell passages (#2, #5 and #8). DPSCs were isolated using either the enzymatic digestion (ED) or explant (EX) technique. The number of days needed for the cells to reach confluence was determined. Immunophenotyping was performed by immunofluorescence and flow cytometry analysis using antibodies specific for nestin, vimentin, CD44, CD146, Oct3/4 and CD34. Data were subjected to three-way analysis of variance (n = 6/group). The ANOVA tests were complemented by Tukey's or t-tests (p < 0.05). The variables "donor age" and "technique" were analyzed to define the optimal desirability value using a response optimization. DPSCs presented a high proliferation rate from passages 2 to 5 while cells from passage 8 proliferated at a slower rate. For all markers, no significant difference was observed among passages, irrespective of the technique used or the donor's age. The mean fraction of specific antibodies was 73.7% (± 11.5), 49.0% (± 18.7), 80.1% (± 8.0), 45.2% (± 13.7), 64.7% (± 5.3) and 2.0% (± 1.5) for CD44, OCT, vimentin, nestin, CD146 and CD34, respectively. The highest optimal desirability value was obtained using the ED technique and cells from younger patients (d = 0.92). However, it was concluded that neither the isolation technique nor the donor age or cell passage significantly interfered with the stem cell phenotype and proliferation rate during cell expansion.

Proliferation rate and expression of stem cells markers during expansion in primary culture of pulp cells

Abstract: The aim of the present study was to evaluate the proliferation rate and the expression of stem cells markers during expansion in primary culture of dental pulp stem cells (DPSCs), comparing different techniques (explant and enzymatic digestion), subject ages (up to 40 and over 40) and cell passages (#2, #5 and #8). DPSCs were isolated using either the enzymatic digestion (ED) or explant (EX) technique. The number of days needed for the cells to reach confluence was determined. Immunophenotyping was performed by immunofluorescence and flow cytometry analysis using antibodies specific for nestin, vimentin, CD44, CD146, Oct3/4 and CD34. Data were subjected to three-way analysis of variance (n = 6/group). The ANOVA tests were complemented by Tukey's or t-tests (p < 0.05). The variables "donor age" and "technique" were analyzed to define the optimal desirability value using a response optimization. DPSCs presented a high proliferation rate from passages 2 to 5 while cells from passage 8 proliferated at a slower rate. For all markers, no significant difference was observed among passages, irrespective of the technique used or the donor's age. The mean fraction of specific antibodies was 73.7% (± 11.5), 49.0% (± 18.7), 80.1% (± 8.0), 45.2% (± 13.7), 64.7% (± 5.3) and 2.0% (± 1.5) for CD44, OCT, vimentin, nestin, CD146 and CD34, respectively. The highest optimal desirability value was obtained using the ED technique and cells from younger patients (d = 0.92). However, it was concluded that neither the isolation technique nor the donor age or cell passage significantly interfered with the stem cell phenotype and proliferation rate during cell expansion.

Simvastatin-Enriched Macro-Porous Chitosan-Calcium-Aluminate Scaffold for Mineralized Tissue Regeneration.

The present study evaluated the odontogenic potential of human dental pulp cells (HDPCs) exposed to chitosan scaffolds containing calcium aluminate (CHAlCa) associated or not with low doses of simvastatin (SV). Chitosan scaffolds received a suspension of calcium aluminate (AlCa) and were then immersed into solutions containing SV. The following groups were established: chitosan-calcium-aluminate scaffolds (CHAlCa - Control), chitosan calcium-aluminate with 0.5 µM SV (CHAlCa-SV0.5), and chitosan calcium-aluminate with 1.0 µM SV (CHAlCa-SV1.0). The morphology and composition of the scaffolds were evaluated by SEM and EDS, respectively. After 14 days of HDPCs culture on scaffolds, cell viability, adhesion and spread, mineralized matrix deposition as well as gene expression of odontogenic markers were assessed. Calcium aluminate particles were incorporated into the chitosan matrix, which exhibited regular pores homogeneously distributed throughout its structure. The selected SV dosages were biocompatible with HDPCs. Chitosan-calcium-aluminate scaffolds with 1 µM SV induced the odontoblastic phenotype in the HDPCs, which showed enhanced mineralized matrix deposition and up-regulated ALP, Col1A1, and DMP-1 expression. Therefore, one can conclude that the incorporation of calcium aluminate and simvastatin in chitosan scaffolds had a synergistic effect on HDPCs, favoring odontogenic cell differentiation and mineralized matrix deposition.

Influence of Bisphosphonates on the Behavior of Osteoblasts Seeded Onto Titanium Discs.

Among other factors, types of bisphosphonates and treatment regimens seem to be strongly associated with the success or failure of installation of osseointegrated implants. This study investigated the influence of two bisphosphonates, sodium alendronate (SA) and zoledronic acid (ZA), on the metabolism of osteoblasts. Human osteoblasts (Saos-2) were seeded onto machined or acid-treated titanium discs previously placed on 24-well plates in complete culture medium. After 24 h, cells were exposed to bisphosphonates at 0.5, 1 or 5 µM for 24 h, 48 h or 7 days. The effects of SA and ZA on osteoblasts were assessed based on the adhesion of these cells to the titanium surfaces by direct fluorescence, cell viability, total protein and collagen synthesis. Alkaline phosphatase activity and mineral nodule deposition by these cells were also evaluated. Data were evaluated by ANOVA and Tukey tests (α=0.05). Decreased adhesion of cells to the titanium discs was observed when exposed to both bisphosphonates; however, this lack of cell adhesion was more evident for ZA-treated cells. In addition, the exposure of osteoblasts to ZA decreased the viability, ALP activity and mineral nodule deposition, which may be related to poor osseointegration after implant installation.

Influence of bisphosphonates on the behavior of osteoblasts seeded onto titanium discs

Abstract Among other factors, types of bisphosphonates and treatment regimens seem to be strongly associated with the success or failure of installation of osseointegrated implants. This study investigated the influence of two bisphosphonates, sodium alendronate (SA) and zoledronic acid (ZA), on the metabolism of osteoblasts. Human osteoblasts (Saos-2) were seeded onto machined or acid-treated titanium discs previously placed on 24-well plates in complete culture medium. After 24 h, cells were exposed to bisphosphonates at 0.5, 1 or 5 µM for 24 h, 48 h or 7 days. The effects of SA and ZA on osteoblasts were assessed based on the adhesion of these cells to the titanium surfaces by direct fluorescence, cell viability, total protein and collagen synthesis. Alkaline phosphatase activity and mineral nodule deposition by these cells were also evaluated. Data were evaluated by ANOVA and Tukey tests (α=0.05). Decreased adhesion of cells to the titanium discs was observed when exposed to both bisphosphonates; however, this lack of cell adhesion was more evident for ZA-treated cells. In addition, the exposure of osteoblasts to ZA decreased the viability, ALP activity and mineral nodule deposition, which may be related to poor osseointegration after implant installation.

Resumo Entre outros fatores, os tipos de bisfosfonatos bem como os regimes de tratamento parecem estar diretamente associados com o sucesso ou falhas na instalação de implantes osseointegrados. Este estudo avaliou a influência de dois bisfosfonatos, o alendronato de sódio (AS) e o ácido zoledrônico (AZ), no metabolismo de osteoblastos. Osteoblastos humanos (Saos-2) foram cultivados sobre discos de titânio polidos ou submetidos a tratamento ácido superficial, previamente alocados em placas de 24 compartimentos, utilizando meio de cultura completo. Após 24 horas, as células foram expostas aos bisfosfonatos, nas concentrações de 0,5, 1 ou 5 µM, por 24 h, 48 h, ou 7 dias. Os efeitos do AZ e AZ sobre os osteoblastos foram determinados considerando a adesão destas células às superfícies de titânio, por meio de fluorescência direta, a viabilidade celular, produção de proteína total e síntese de colágeno. A atividade de fosfatase alcalina e a deposição de nódulos mineralizados também foram avaliadas. Os dados foram analisados por meio do teste ANOVA complementado por Tukey (α = 0.05). Menor adesão dos osteoblastos foi observada quando estas células foram expostas a ambos os bisfosfonatos, porém, esta falha na adesão foi mais evidente para as células tratadas com AZ. Além disso, a exposição dos osteoblastos ao AZ também resultou em diminuição da viabilidade, atividade de ALP e deposição de nódulos mineralizados, o que pode estar relacionado a uma pobre osseointegração após a instalação do implante.

Proteolytic activity, degradation, and dissolution of primary and permanent teeth.

BACKGROUND: Primary and permanent teeth composition may influence dissolution and degradation rates. AIM: To compare the dissolution and degradation of primary and permanent teeth. DESIGN: Enamel and dentin powders were obtained from primary molars and premolars and incubated within different pH buffers. Calcium and inorganic phosphate release was quantified in the buffers by atomic absorption and light spectrophotometry. A colorimetric assay was used to assess the MMP activity of primary dentin (PrD) and permanent dentin (PeD). Collagen degradation was assessed by dry mass loss, change in elastic modulus (E), and ICTP and CTX release. Data were submitted to ANOVA and Tukey's tests (α = 0.05). RESULTS: Similar dissolution was found between PrD and PeD after 256 hours. At pH 4.5, enamel released more minerals than dentin whereas at pH 5.5 the inverse result was observed. MMP activity was similar for both substrates. PrD showed higher dry mass loss after 1 week. In general, greater reduction in E was recorded for PrD. Higher quantities of ICTP and CTX were released from PrD after 1 week. CONCLUSIONS: Primary and permanent teeth presented similar demineralization rates. Collagen degradation, however, was faster and more substantial for PrD.

Increased whitening efficacy and reduced cytotoxicity are achieved by the chemical activation of a highly concentrated hydrogen peroxide bleaching gel.

OBJECTIVE: This study was designed for the chemical activation of a 35% hydrogen peroxide (H2O2) bleaching gel to increase its whitening effectiveness and reduce its toxicity. METHODOLOGY: First, the bleaching gel - associated or not with ferrous sulfate (FS), manganese chloride (MC), peroxidase (PR), or catalase (CT) - was applied (3x 15 min) to enamel/dentin discs adapted to artificial pulp chambers. Then, odontoblast-like MDPC-23 cells were exposed for 1 h to the extracts (culture medium + components released from the product), for the assessment of viability (MTT assay) and oxidative stress (H2DCFDA). Residual H2O2 and bleaching effectiveness (DE) were also evaluated. Data were analyzed with one-way ANOVA complemented with Tukey's test (n=8. p<0.05). RESULTS: All chemically activated groups minimized MDPC-23 oxidative stress generation; however, significantly higher cell viability was detected for MC, PR, and CT than for plain 35% H2O2 gel. Nevertheless, FS, MC, PR, and CT reduced the amount of residual H2O2 and increased bleaching effectiveness. CONCLUSION: Chemical activation of 35% H2O2 gel with MC, PR, and CT minimized residual H2O2 and pulp cell toxicity; but PR duplicated the whitening potential of the bleaching gel after a single 45-minute session.

Influence of Zirconia-Coated Bioactive Glass on Gingival Fibroblast Behavior.

The objective of this study was the development of a bioactive glass coating on zirconia (Zr) to modulate the gingival fibroblast phenotype. For this purpose, Biosilicate® (BS) particles in a water/isopropyl alcohol (1:1) vehicle (6 mg/mL) were applied to zirconia discs followed by thermal treatment at 1100 °C for 20 min. The surface topography (SEM), chemical composition (EDX), surface roughness (Ra; confocal microscopy), surface free energy (goniometry), and color alteration (UV-vis spectrophotometry) were assessed (n=6). Thereafter, L929 fibroblasts were seeded onto Zr and Zr+BS discs, and cell proliferation (Alamar Blue; n=6), morphology (SEM; n=2), migration (wound healing; n=4), and collagen synthesis (Sirius Red; n=6) were evaluated up to 7 days. Data were analyzed by ANOVA/Tukey tests (a=5%). A homogeneous coating consisting of Si, Na, O, and Ca was detected on the Zr surface after thermal treatment with BS, which led to a significant increase in surface roughness and free energy (p<0.05). No change in color parameters was observed (p>0.05). Cells seeded on the Zr+BS surface featured increased proliferation, collagen expression, and migration capability in comparison with those cultured on plain Zr (p<0.05). SEM images revealed that cell spreading occurred faster in the presence of BS. Therefore, it was concluded that thermal treatment of the Zr surface with BS led to the deposition of a bioactive coating, which induced gingival fibroblast spread, proliferation, migration, and collagen expression in vitro.

Photodithazine-mediated antimicrobial photodynamic therapy against fluconazole-resistant Candida albicans in vivo.

This study evaluated the efficacy of antimicrobial Photodynamic Therapy (aPDT) against fluconazole-resistant Candida albicans in a murine model of oral candidosis. Mice were inoculated with two clinical isolates (R10, R15) and one reference strain (ATCC) of resistant C. albicans to produce oral candidosis. After inoculation, aPDT mediated by Photodithazine® (PDZ) and LED light was performed. The use of PDZ or light only was also investigated. Additional animals were treated with Nystatin (NYS). Untreated or healthy mice were also evaluated. Microbiological evaluation was performed by recovering C. albicans from the tongue via colony-forming units. Animals were killed 24 hours after treatments, and the tongues were removed for histological analysis. Data were analyzed by one-way ANOVA and Tukey test (P < .05). The results demonstrated that all strains showed the same behavior after aPDT and NYS treatment. A significant reduction in C. albicans viability was achieved after both treatments for R15 and ATCC. No significant reduction was verified for C. albicans R10 submitted to aPDT or NYS. The histological analysis revealed that aPDT did not cause side effects on tissues. aPDT was effective for inactivation of two fluconazole-resistant C. albicans of the three strains evaluated.

Increased whitening efficacy and reduced cytotoxicity are achieved by the chemical activation of a highly concentrated hydrogen peroxide bleaching gel

Abstract Objective This study was designed for the chemical activation of a 35% hydrogen peroxide (H2O2) bleaching gel to increase its whitening effectiveness and reduce its toxicity. Methodology First, the bleaching gel - associated or not with ferrous sulfate (FS), manganese chloride (MC), peroxidase (PR), or catalase (CT) - was applied (3x 15 min) to enamel/dentin discs adapted to artificial pulp chambers. Then, odontoblast-like MDPC-23 cells were exposed for 1 h to the extracts (culture medium + components released from the product), for the assessment of viability (MTT assay) and oxidative stress (H2DCFDA). Residual H2O2 and bleaching effectiveness (DE) were also evaluated. Data were analyzed with one-way ANOVA complemented with Tukey's test (n=8. p<0.05). Results All chemically activated groups minimized MDPC-23 oxidative stress generation; however, significantly higher cell viability was detected for MC, PR, and CT than for plain 35% H2O2 gel. Nevertheless, FS, MC, PR, and CT reduced the amount of residual H2O2 and increased bleaching effectiveness. Conclusion Chemical activation of 35% H2O2 gel with MC, PR, and CT minimized residual H2O2 and pulp cell toxicity; but PR duplicated the whitening potential of the bleaching gel after a single 45-minute session.

Bond Strength and Cytotoxicity of a Universal Adhesive According to the Hybridization Strategies to Dentin

Abstract This study evaluated application protocol (etch-and-rinse/ER and self-etching/SE) and dentin wettability (wet and dry) on microtensile bond strength (μTBS) and transdentinal cytotoxicity of ScotchbondTM Universal (SU) adhesive system. The μTBS values and fracture mode were registered 24 h after adhesive system application and resin composite block build-up (n=5). For analysis of transdentinal cytotoxicity, odontoblast-like MDPC-23 cells were seeded on pulpal surface of dentin discs (0.4 mm thick) adapted to artificial pulp chambers (n=8). The adhesive system was applied to occlusal surface, followed by 24-h incubation time. Cell viability (Alamar Blue) and morphology (SEM) were assessed. Adper Single Bond 2 and Clearfil SE Bond were used as positive controls of the ER and SE application protocols, respectively. No treatment was performed on negative control (NC) group. Data were analyzed by ANOVA and Tukey's tests (α=5%). Higher μTBS values were found for ER mode in comparison with SE protocol (p<0.05). Dentin wettability had no effect on bond strength of SU in both the ER and SE techniques (p>0.05). Most fractures involved hybrid layer and/or adhesive layer. Neither variable prevented the intense toxic effects of adhesive systems on MDPC-23 cultured cells, since intense reduction in cell viability (±88%) and severe alterations in cell morphology were observed for all groups compared to NC, with no differences among them (p>0.05). Therefore, it was concluded that application of SU following the ER protocol had better adhesive performance. However, this adhesive system featured intense transdentinal cytotoxicity to pulp cells, regardless of application protocol and dentin wettability.

Resumo Este estudo avaliou o protocolo de aplicação (convencional/ER e autocondicionante/SE) e o grau de umidade da dentina (úmida e seca) sobre a resistência de união à microtração (μTBS) e a citotoxicidade transdentinária do sistema adesivo ScotchbondTM Universal (SU). Os valores de μTBS e o modo de fratura foram registrados 24 h após aplicação do sistema adesivo e restauração com resina composta pela técnica incremental. Para avaliação da citotoxicidade transdentinária, células odontoblastóides MDPC-23 foram semeadas na face pulpar de discos de dentina (0,4 mm de espessura) adaptados a câmaras pulpares artificiais (n = 8). O sistema adesivo foi aplicado na superfície oclusal, seguido de incubação por 24 h. A viabilidade e morfologia celular foram avaliadas pelo teste de Alamar Blue e MEV, respectivamente. Adper Single Bond 2 e Clearfil SE Bond foram utilizados como controle positivo do protocolo de aplicação ER e SE, respectivamente. Nenhum tratamento foi realizado no grupo controle negativo (NC). Os dados foram analisados pelos testes de ANOVA e Tukey (α = 5%). Maiores valores de μTBS foram encontrados para o modo ER em comparação com o protocolo SE (p < 0,05). O grau de umidade da dentina não apresentou efeito na resistência de união do SU em ambos os protocolos ER e SE (p > 0.05). A maioria das fraturas envolveu a camada híbrida e / ou camada adesiva. Ambas as variáveis não preveniram o intenso efeito citotóxico dos sistemas adesivos sobre as células MDPC-23 em cultura, uma vez que redução intensa na viabilidade celular (± 88%) e alterações severas na morfologia celular foram observadas para todos os grupos quando comparados ao NC, sem diferenças entre eles (p > 0.05). Desta forma, foi concluído que a aplicação do SU seguindo o protocolo ER apresentou melhor performance adesiva. No entanto, esse sistema adesivo promoveu intensa citotoxicidade transdentinária sobre células pulpares, independente do protocolo de aplicação e grau de umidade dentinária.

Análise da citotoxicidade de um gel clareador com 10% de peróxido de hidrogênio sobre células odontoblastóides de acordo com a espessura do substrato dental / Analysis of the cytotoxicity of a 10% hydrogen peroxide bleaching gel to odontoblast-like cells according to the thickness of dental substrate

Objetivo: Avaliar a citotoxicidade de um gel clareador contendo 10% de peróxido de hidrogênio (H2O2), aplicado sobre discos de esmalte/dentina simulando diferentes espessuras dentais. Material e método: Discos com 2,3; 3,5; e 4,0 mm de espessura foram obtidos para simular incisivos centrais inferiores, incisivos centrais superiores e segundos pré-molares superiores, respectivamente. Para cada espessura, o gel com 10% de H2O2 foi aplicado sobre o esmalte por 3x 15 min, 1x 15 min ou 1x 5 min. O protocolo 35% H2O2 3x 15 min foi empregado como controle positivo (CP), e nenhum tratamento foi realizado no controle negativo (CN). Células odontoblastóides MDPC-23 foram expostas por 1 h aos componentes da difusão trans-amelodentinária coletados imediatamente após o clareamento, sendo realizada análise da viabilidade celular, estresse oxidativo, deposição de nódulos de mineralização, bem como a quantificação da difusão de H2O2 pelos discos. Resultados: O gel com 10% de H2O2 não promoveu redução significativa da viabilidade celular em relação ao CN para todos os protocolos e espessuras testadas, resultando em valores de difusão de H2O2 significativamente inferiores ao CP. Apenas o protocolo 10% 3x 15 min aplicado sobre os discos simulando incisivos promoveu aumento no estresse oxidativo e reduziu a deposição de nódulos de mineralização em relação ao CN; porém, estes efeitos foram significativamente inferiores ao CP. Conclusão: De acordo com a metodologia usada neste estudo, foi possível concluir que, independente da espessura dental, a aplicação de um gel clareador com 10% de H2O2 por 5-45 min sobre o esmalte causa limitado efeito citotóxico sobre células pulpares.

Objective: To evaluate the cytotoxicity of a bleaching gel with 10% hydrogen peroxide (H2O2) applied onto enamel/dentin discs simulating different dental thicknesses. Material and methods: Discs with 2.3; 3.5; and 4.0 mm thickness were obtained to simulate low central incisors, upper central incisors and upper second pre-molars, respectively. For each thickness, the 10% H2O2 gel was applied for 3x 15 min, 1x 15 min or 1x 5 min. A gel with 35% H2O2 applied for 3x 15 min was used as positive control (PC) and no treatment was performed in negative control (NC). Odontoblast- like MDPC-23 cells were exposed for 1 h to the transenamel and trans-dentinal components collected immediately after bleaching. Cell viability, oxidative stress and mineralized nodule deposition were assessed as well as the quantification of H2O2 diffused through the discs. Results: The 10% H2O2 gel did not promote significant reduction on cell viability in comparison to NC for all tested protocols and thicknesses, resulting in H2O2 diffusion values significantly lower than PC. Only the protocol 10% H2O2 3x 15 min applied onto discs simulating incisors increased significantly the oxidative stress and reduced mineralized nodule deposition compared to NC; however, these effects were significantly lower than PC. Conclusion: According to the methodology employed in this laboratorial study, the application of a bleaching gel with 10% H2O2 for 5-45 min onto dental structure featured limited cytotoxicity to pulp cells, disregarding the enamel/dentin thicknesses.

Bond Strength and Cytotoxicity of a Universal Adhesive According to the Hybridization Strategies to Dentin.

This study evaluated application protocol (etch-and-rinse/ER and self-etching/SE) and dentin wettability (wet and dry) on microtensile bond strength (µTBS) and transdentinal cytotoxicity of ScotchbondTM Universal (SU) adhesive system. The µTBS values and fracture mode were registered 24 h after adhesive system application and resin composite block build-up (n=5). For analysis of transdentinal cytotoxicity, odontoblast-like MDPC-23 cells were seeded on pulpal surface of dentin discs (0.4 mm thick) adapted to artificial pulp chambers (n=8). The adhesive system was applied to occlusal surface, followed by 24-h incubation time. Cell viability (Alamar Blue) and morphology (SEM) were assessed. Adper Single Bond 2 and Clearfil SE Bond were used as positive controls of the ER and SE application protocols, respectively. No treatment was performed on negative control (NC) group. Data were analyzed by ANOVA and Tukey's tests (α=5%). Higher µTBS values were found for ER mode in comparison with SE protocol (p<0.05). Dentin wettability had no effect on bond strength of SU in both the ER and SE techniques (p>0.05). Most fractures involved hybrid layer and/or adhesive layer. Neither variable prevented the intense toxic effects of adhesive systems on MDPC-23 cultured cells, since intense reduction in cell viability (±88%) and severe alterations in cell morphology were observed for all groups compared to NC, with no differences among them (p>0.05). Therefore, it was concluded that application of SU following the ER protocol had better adhesive performance. However, this adhesive system featured intense transdentinal cytotoxicity to pulp cells, regardless of application protocol and dentin wettability.

Biostimulatory effects of simvastatin on MDPC-23 odontoblast-like cells.

The aim of this study was to evaluate the bioactivity and cytocompatibility of simvastatin (SV) applied to MDPC-23 odontoblast-like cells. For this purpose, MDPC-23 cells were seeded in 96-well plates and submitted to treatments with 0.01 or 0.1 µM of SV for 24 h, 72 h or continuously throughout the experimental protocol. The negative control group (NC) was maintained in DMEM. Cell viability (MTT), ALP activity (thymolphthalein monophosphate), and mineralized matrix deposition (alizarin red) were analyzed at several time points. The data were submitted to ANOVA and Tukey's test (α = 0.05). Although cell viability was observed in the groups treated with SV, these groups did not differ from the NC up to 7 days. There was a reduction in cell viability for the groups treated with 0.1 µM of SV for 72 h, and submitted to continuous mode after 14 days. A significant increase in ALP activity occurred in the group treated with 0.01 µM of SV for 24 h, compared with the NC; however, only the group treated with 0.1 µM of SV in continuous mode reduced the ALP activity, in comparison with the NC. After 14 days, only continuous treatment with 0.1 µM of SV did not differ from NC, whereas the other experimental groups showed increased mineralized matrix deposition. Thus, it was concluded that low concentrations of simvastatin were bioactive and cytocompatible when applied for short periods to cultured MDPC-23 odontoblast-like cells.