The presence of osteocalcin, osteopontin and reactive oxygen species-positive cells in pulp tissue after dental bleaching.

AIM: To analyse the influence of H2 O2 on pulp repair through osteocalcin and osteopontin immunolabelling and in cellular defence by using the antireactive oxygen species (ROS) antibody. METHODOLOGY: The maxillary molars of 50 rats were treated with 35% H2 O2 (Ble groups) or placebo gel (control groups). At 0 h and 2, 7, 15 and 30 days (n = 10 hemimaxillae), the rats were killed and pulp tissue was evaluated using inflammation and immunolabelling scores (osteocalcin/osteopontin); ROS-positive cells were counted. Paired t-test and Wilcoxon signed-rank test were used (P < 0.05). RESULTS: The Ble group had necrosis in the coronal pulp at 0 h and in the occlusal third of the coronal pulp at 2 days; at 7, 15 and 30 days, no inflammation was noted similar to the controls (P > 0.05). Osteocalcin was absent in the Ble at 0 h, moderate at 2 days and increased thereafter, differing from the controls at all two periods (P < 0.05). Osteopontin was higher principally at 7 and 15 days in Ble groups, but differing with control groups from 2 days after bleaching (P < 0.05). The Ble group had more ROS-positive cells in the pulp at 7 and 15 days (P < 0.05). Tertiary dentine was observed at 7 days, increasing thereafter (P < 0.05). CONCLUSIONS: Post-bleaching pulp repair was associated with increased osteocalcin over time. Osteopontin also participated in this process, and anti-ROS was involved in cellular defence against H2 O2 .

In vivo analysis of the presence of heme oxygenase-1, transcription factor Jun-D and CD90+/CD73+/CD105+/CD45- cells in the pulp of bleached teeth.

AIM: To investigate hydrogen peroxide (H2 O2 )-induced responsiveness in pulp cells using heme oxygenase-1 (HO-1) immunolabelling, Jun-D immunolabelling to study the effects of H2 O2 on odontoblastic differentiation and CD90+/CD73+/CD105+/CD45- cell counting for in vivo identification of mesenchymal stem cells in the pulp. METHODOLOGY: The maxillary molars of 50 rats were treated with a bleaching gel (35% H2 O2 , 1 × 30 min) or placebo gel (control groups). At 2, 3, 7, 15 and 30 days after the treatment (n = 10), inflammation in pulp tissue was analysed by haematoxylin-eosin staining, HO-1- and Jun-D-immunolabelled cells were counted in each third of the pulp chamber, and the number of CD90+/CD73+/CD105+/CD45- cells was quantified by immunofluorescence. The results were assessed using the Paired t-test or Wilcoxon signed-rank test (P < 0.05). RESULTS: Significant H2 O2 -induced inflammation was noted at 2 and 3 days (P < 0.05), with tertiary dentine formation occurring from 7 days. The bleached specimens had greater HO-1 immunolabelling in the middle and cervical thirds of the coronal pulp at 2 and 3 days, in all thirds at 7 days, and in the occlusal third at 15 days (P < 0.05), and significant nuclear Jun-D immunolabelling in the cervical third at 2 and 3 days and in the occlusal and middle thirds at 7 days (P < 0.05). Bleached and control groups had low numbers of CD90+/CD73+/CD105+/CD45- cells in the pulp at all periods (P > 0.05). CONCLUSIONS: Pulp cells responded to oxidative stress by expressing HO-1 during the post-bleaching inflammation phase until the beginning of the repair phase. Jun-D expression occurred during the reduction of inflammation and the beginning of tertiary dentine production. The presence of oxidative stress did not influence the number of CD90+/CD73+/CD105+/CD45- cells identified in vivo in the dental pulp.

The effect of dental bleaching on pulpal tissue response in a diabetic animal model: a study of immunoregulatory cytokines.

AIM: To evaluate the influence of tooth bleaching on immunoregulatory cytokines production (IL-6, Tumour necrosis factor (TNF)-α and IL-17) in the pulp tissue of normoglycaemic and diabetic rats. METHODOLOGY: Twenty-eight rats were divided into normoglycaemic and diabetic rats (n = 14). Diabetes mellitus (DM) was induced with a single dose of alloxan diluted in citrate buffer via intramuscular injection. After DM confirmation, all rats were sedated and tooth bleaching was performed using 35% hydrogen peroxide on the right maxillary molars for 30 min. Left molars were used as controls. Bleaching resulted in four hemimaxillae groups: normoglycaemic (N), N-bleached (NBle), diabetic (D) and D-bleached (DBle). After 2 and 30 days, rats were euthanized and hemimaxillae processed for analysis by haematoxylin and eosin and immunohistochemistry. Results within and between animals were submitted to Wilcoxon signed-rank and Mann-Whitney tests (P < 0.05). RESULTS: At 2 days, the NBle group had mild, and the DBle had severe inflammatory infiltration in the pulpal tissue (P < 0.05). TNF-α and IL-6 cytokines were associated with increased immunolabelling in the bleached groups compared to nonbleached (P < 0.05). However, IL-17 had increased immunolabelling in the NBle compared to the N and DBle group (P < 0.05). At 30 days, reactionary dentine was observed in the coronal pulp of all bleached teeth and no inflammation was present (P > 0.05). TNF-α cytokines had increased immunolabelling in the DBle group compared to the D group (P < 0.05). However, for IL-6 and IL-17, no difference was observed in this period (P > 0.05). CONCLUSIONS: Tooth bleaching increased IL-6 and TNF-α in the pulp tissue regardless of diabetes mellitus; however, diabetic rats had higher TNF-α levels for longer periods. Tooth bleaching influenced the increase in IL-17 in the early periods in normoglycaemic rats.

Concentration-dependent effect of bleaching agents on the immunolabelling of interleukin-6, interleukin-17 and CD5-positive cells in the dental pulp.

AIM: To evaluate lymphocyte-like cell activation (CD5-positive cells) and the expression of interleukin (IL)-6 and IL-17 in the pulp after tooth bleaching with two concentrations of hydrogen peroxide (H2 O2 ). METHODOLOGY: The right and left maxillary molars from 40 rats were treated randomly with bleaching gel with 20% H2 O2 (BLUE group, 1 application of 50 min), 35% H2 O2 (MAXX group, three applications of 15 min), or placebo gel (control). After 2 and 30 days, the rats were killed (n = 10), and the jaws were processed for histological and immunohistochemistry analysis of the pulp tissue. The scores of inflammation and immunolabelling (IL-6/IL-17) were submitted to Mann-Whitney and Kruskal-Wallis followed Dunn tests, respectively; anova tests were used for comparisons of number of CD5-positive cells and pulp chamber area values (P < 0.05). RESULTS: At 2 days, 60% of specimens of the BLUE group were associated with moderate inflammation in pulp horns, and in the MAXX group with necrosis (P < 0.05). At 30 days, the pulp was organized, and tertiary dentine was formed. The MAXX group had superior immunolabelling of IL-17 at 2 days differing significantly from other groups (P < 0.05). At 2 days, 90% of the specimens of the BLUE group had moderate immunolabelling of IL-6, and 50% of the MAXX group had severe immunolabelling, both significantly different from the control (P < 0.05). There was no significant difference between the groups at 30 days (P > 0.05). CD5-positive cells were present at 2 and 30 days, particularly in the bleached groups (P < 0.05), without significant difference between time periods (P > 0.05). CONCLUSIONS: IL-6 and IL-17 participated in inflammation in the pulp tissue of rats after tooth bleaching, particularly at 2 days. The immunolabelling was greater with increasing H2 O2 concentration. This process was accompanied by the prolonged activation of CD5-positive cells.

The effect of dental bleaching on pulpal tissue response in a diabetic animal model.

AIM: To evaluate pulpal tissue response after dental bleaching in normal and alloxan-induced diabetic rats. METHODOLOGY: Twenty-eight rats were divided into two groups of normoglycaemic and diabetic rats (n = 14). Diabetes mellitus (DM) was induced with alloxan. After DM confirmation, all rats were anaesthetized and dental bleaching was performed with 35% hydrogen peroxide (H2 O2 ) on the right maxillary molars for 30 min. Left molars were used as controls. Bleaching resulted in four hemimaxillae groups: normoglycaemic (N), N-bleached (NBle), diabetic (D) and D-bleached (DBle). After 2 or 30 days, the animals were euthanized and the hemimaxillae were removed, processed for histopathological analysis and stained with haematoxylin-eosin (HE), Masson's trichrome (MT) and picrosirius red (PSR). Results obtained within animals (normoglycaemic or diabetic rats) were submitted to Wilcoxon or paired t-tests, and between animal (normoglycaemic and diabetic rats), to Mann-Whitney test or t-tests. RESULTS: At 2 days, the NBle group had a mild inflammatory infiltration in the pulpal tissue, whilst the DBle had severe inflammation or necrosis (P < 0.05). At 30 days, no inflammation was present. However, a significant difference in pulp chamber area reduction by reactionary dentine deposition was found between the NBle and DBle groups (P < 0.05). At 2 days, fewer immature collagen fibres and more mature collagen fibres were noted in the NBle, D and DBle groups; this was significantly different when compared to the N group (P < 0.05). At 30 days, significantly fewer immature collagen fibres and more mature collagen fibres were noted in NBle compared with DBle group (P < 0.05). CONCLUSIONS: The inflammatory tissue response in rats' teeth after dental bleaching was greater in diabetic rats. Additionally, the increase in reactionary dentine deposition and mature collagen fibres observed in diabetic rats needs further evaluation to confirm the present results.

In vitro evaluation of surface roughness and microhardness of restorative materials submitted to erosive challenges.

The aim of this study was to evaluate the effect of different acidic solutions on the microhardness and surface roughness of restorative materials. The 120 specimens of restorative materials (Fuji II LC, Vitremer, Supreme XT, and Supreme XT + Biscover LV) were randomly divided into three groups according to the immersion media: hydrochloric acid, soft drink, or distilled water. Over a period of five weeks, the groups were immersed in the solutions, which were changed weekly. Data were tested using analysis of variance and the Fisher protected least significant difference test (p<0.05). The results showed that the glass ionomer materials showed the highest surface roughness values (Fuji II LC: 0.111 ± 0.014 µm before and 0.139 ± 0.016 µm after immersion; Vitremer: 0.177 ± 0.012 µm before and 0.084 ± 0.012 µm after immersion), whereas the lowest values were found for the resin sealed with Biscover LV before (0.047 ± 0.011 µm) and after exposure in distilled water (0.043 ± 0.007 µm), soft drink (0.040 ± 0.005 µm), and hydrochloric acid (0.045 ± 0.005 µm). The Supreme XT showed the highest microhardness values before (44.96 ± 2.51 KHN) and after the aging process (41.26 ± 1.22 KHN in water, 35.96 ± 0.81 KHN in soft drink, and 34.74 ± 0.97 KHN in HCl), with significant differences from the other materials (p<0.0001). The lowest microhardness values were found for glass ionomer materials. The solutions used in this study decreased the microhardness of all studied materials, whereas the sealed surface suffered minor changes in microhardness and surface roughness after exposure to acidic solutions.

Wear, roughness and microhardness analyses of single increment restorative materials submitted to different challenges in vitro.

PURPOSE: To evaluate the microhardness, roughness, profilometry and cross-sectional hardness of single increment materials submitted to different challenges. METHODS: Thirty-six disks of each material, Filtek Supreme XTE (FT), Filtek One Bulk Fill (BK), Ketac Molar Easy Mix (KT) and Equia Forte + Coat (EQ) were immersed in saliva, pH cycling and Coke for 15 days. Half of each surface was used as its own control. Superficial microhardness, roughness, perfilometry analysis were performed. All samples were sectioned, embedded in acrylic resins, polished and cross-sectional hardness were made. Data were analyzed by ANOVA and Tukey's test (p < 0.05). RESULTS: The KT presented superficial microhardness superior than EQ. However, in depth, EQ showed superior values. FT, KT suffered the effects of challenges on microhardness values. The highest roughness and wear values were found for KT. RC do not suffer wear. All materials suffered the effects of Coke and pH challenges in depths 10 µm and 60 µm. CONCLUSION: The single increment restorative material that suffered less action on its surface was the bulk-fill resin. The coat present in the hybrid ionomer was able to resist to the immersion actions. In addition, Coke was the most aggressive challenge.

Influence of green tea extract in the color of composite resin restorations.

The aim of this study was to evaluate the color change of composite resin restorations in Class I cavity preparation with different depths, submitted to challenge of thermocycling in coffee, after the use of green tea extract (EGCG) as treatment on the dentin surface. Forty-eight human molars were divided into 6 groups according to dentin treatment and depth of restoration (n = 8): Group C3- Control/3 mm; Group C4- Control/4 mm; Group C5- Control/5 mm; Group EGCG3- EGCG/3 mm; Group EGCG4- EGCG/4 mm; and Group EGCG5- EGCG/5 mm. The teeth of the control groups were restored by the bulk fill technique (Filtek Bulk Fill), conditioning the dentin surface only with universal bonding system (Single Bond Universal). The teeth of the EGCG groups were also restored by the bulk filling technique, but conditioning the dentin surface with 0.5% EGCG for 30 s prior to the application of the adhesive system. Initial and final color readings were performed according to the CIE L*a*b* scale in UV-2450 spectroscope, before and after challenge of thermal cycling in coffee. The color change (ΔE) was then calculated based on the formula ΔE = [(ΔL*)2+(Δa*)2+(Δb*)2]½. The ΔE data were submitted to statistical tests of normality, two-way ANOVA and Tukey test to compare the means (p < 0.05). There was no statistically difference for both study factors analyzed (EGCG application and restoration depth), as well as the interaction between both, after aging in coffee (p > 0.05). It was concluded that the previous application of EGCG did not cause a significant color change at the dentin-resin interface.

Effect of Different Light Sources and Enamel Preconditioning on Color Change, H2O2 Penetration, and Cytotoxicity in Bleached Teeth.

This study evaluated the effects of acid etching of the enamel and the combination of different light sources (halogen light, light-emitting diodes [LEDs], and LED/Laser) and the bleaching product on color change, penetration of hydrogen peroxide (H2O2), and cytotoxicity over time. The color change (ΔE) and the amount of H2O2 that permeated the tooth tissue were analyzed using a spectrophotometer. Cell metabolism and morphology were evaluated using the methylthiazol tetrazolium assay and scanning electron microscopy, respectively. The ΔE values and H2O2 permeation were not significantly different under any of the experimental conditions. Tooth whitening significantly reduced cell metabolism, regardless of whether a light source was used. Preconditioning the enamel did not influence the cellular metabolism in any group. In conclusion, combining the bleaching product with different light sources and/or preconditioning the enamel resulted in few significant changes in color, transenamel and transdentinal penetration of H2O2, or cytotoxicity and cell morphology.

Penetration of hydrogen peroxide and degradation rate of different bleaching products.

This study's aim was to evaluate the degradation rate of hydrogen peroxide (H2O2) and to quantify its penetration in tooth structure, considering the residence time of bleaching products on the dental enamel. For this study, bovine teeth were randomly divided according to the bleaching product received: Opalescence Xtra Boost 38%, White Gold Office 35%, Whiteness HP Blue 35%, Whiteness HP Maxx 35%, and Lase Peroxide Sensy 35%. To analyze the degradation of H2O2, the titration of bleaching agents with potassium permanganate was used, while the penetration of H2O2 was measured via spectrophotometric analysis of the acetate buffer solution, collected from the artificial pulp chamber. The analyses were performed immediately as well as 15 minutes, 30 minutes, and 45 minutes after product application. The data of degradation rate of H2O2 were submitted to analysis of variance (ANOVA) and Tukey tests, while ANOVA and Fisher tests were used for the quantification of H2O2, at the 5% level. The results showed that all products significantly reduced the concentration of H2O2 activates at the end of 45 minutes. It was also verified that the penetration of H2O2 was enhanced by increasing the residence time of the product on the tooth surface. It was concluded that the bleaching gels retained substantial concentrations of H2O2 after 45 minutes of application, and penetration of H2O2 in the dental structure is time-dependent.

Effect of sodium ascorbate on dentin bonding after two bleaching techniques.

The purpose of this study was to analyze the influence of 10% sodium ascorbate (SA) on the hybrid layer, resin tag length, and bond strength to dentin after bleaching. Six groups were tested: G C, control; G SA, sodium ascorbate (SA) + restoration; G CP, bleaching with carbamide peroxide (CP) + restoration; G CP+SA, bleaching with CP + SA+ restoration; G HP, bleaching with 35% hydrogen peroxide (HP) + restoration; and G HP+SA, HP + SA + restoration. After dental bleaching, the dentin was exposed and the antioxidant solution was applied to groups G SA, G CP+SA, and G HP+SA, before bonding procedures. The teeth were sectioned in the mesiodistal direction. One section was decalcified, and the specimens were embedded in paraffin and sectioned in the longitudinal direction with a thickness of 6 µm. Fifteen slices of each specimen were selected according to a systematic sample of slices with an interval proportional to the total number of slices obtained for each tooth. The specimens were stained using the Brown & Brenn method, and an optic microscope was used to analyze the hybrid layer thickness and resin tag length. The remaining tooth segment was sectioned into stick-shaped specimens and used for microtensile bond strength testing (0.5 mm/min). Statistical analysis was performed using two-way analysis of variance and Fisher test. The results for hybrid layer + tag formation (in micrometers) were G C, 13.27 Aa; G SA, 11.85 Ba; G CP, 6.84 Bb; G CP+SA, 9.02 Ab; G HP, 7.28 Bb; and G HP+SA, 9.22 Ab; bond strength results (in MPa) were G C, 49.5 Aa; G SA, 51.7 Aa; G CP, 37.16 Bb; G CP+SA, 47.69 Aa; G HP, 32.39 Ab; and G HP+SA, 39.67 Ab. Tooth bleaching with CP or HP impairs the formation of the hybrid layer and resin tags and reduces the microtensile bond strength. Statistically, the use of SA significantly increases the hybrid layer thickness and resin tag length. The microtensile bond strength values for carbamide peroxide increased, but the microtensile bond strength for hydrogen peroxide was not affected.

Transenamel and transdentinal penetration of hydrogen peroxide applied to cracked or microabrasioned enamel.

The present study evaluated transenamel and transdentinal penetration of hydrogen peroxide during tooth whitening recognized in altered enamel by the presence of cracks or microabrasion. We used 72 experimental units (n=20) obtained from bovine incisors: GI-sound enamel; GII-teeth showing visible enamel cracks (4 mm to 5.7 mm in length); and GIII-microabrasioned enamel. The 12 remaining specimens were used to analyze the enamel surface morphology using scanning electron microscopy. The specimens were cylindrical and 5.7 mm in diameter and 3.5 mm thick. A product based on 35% hydrogen peroxide was used for bleaching, following the manufacturer's recommendations for use. To quantify the H2O2 penetration, the specimens were placed in artificial pulp chambers containing an acetate buffer solution. After bleaching, the solution was collected and adequately proportioned with leucocrystal violet, peroxidase enzyme, and deionized water. The resulting solution was evaluated using ultraviolet visible reflectance spectrophotometer equipment. The data were analyzed by analysis of variance (ANOVA) and Fisher's PLSD at a significance level of 0.05, and significant differences in the penetration of peroxide in different substrate conditions were observed (p<0.0001). The penetration of hydrogen peroxide was more intense in cracked teeth. The group in which the enamel was microabraded showed intermediate values when compared to the control group. Microabrasion and the presence of cracks in the enamel make this substrate more susceptible to penetration of hydrogen peroxide during in-office whitening.

Effect of hydrogen peroxide at 35% on the morphology of enamel and interference in the de-remineralization process: an in situ study.

This study evaluated the microhardness and histomorphology of bovine enamel when 35% hydrogen peroxide is used. A total of 44 specimens were adapted to removable devices used by 11 individuals subjected to dental caries challenge. A decrease in microhardness was observed for all groups after the cariogenic challenge. Microscopic analysis revealed that fragments subjected to cariogenic challenge associated with bleaching had more intense superficial histologic changes, but the depth of the lesions remained unchanged. It was concluded that 35% hydrogen peroxide enhanced the reduction in hardness and histomorphologic changes in the enamel surface exposed to cariogenic challenge.