Influence of Manganese Oxide on the Esthetic Efficacy and Toxicity Caused by Conventional In-office Tooth Bleaching Therapy.

OBJECTIVE: This study aimed to evaluate the esthetic efficacy, cytotoxicity, and kinetics of decomposition of hydrogen peroxide (H2O2) present in a commercial bleaching gel with 35% H2O2 (BG35%) chemically activated with manganese oxide (MnO2). METHODS AND MATERIALS: After incorporating 2 mg/mL, 6 mg/mL, and 10 mg/mL of MnO2 into BG35%, the stability of pH and temperature of the products were analyzed. To assess the esthetic efficacy (ΔE and ΔWI), the BG35%s with MnO2 were applied for 45 minutes on enamel/dentin discs (DiE/D). BG35% or no treatment were used as positive (PC) and negative (NC) controls, respectively. To analyze the cell viability (CV) and oxidative stress (OXS), the same bleaching protocols were performed on DiE/D adapted to artificial pulp chambers. The extracts (culture medium + gel components that diffused through the discs) were applied to pulp cells and submitted to H2O2 quantification. BG35% with MnO2 that showed the best results was evaluated relative to kinetic decomposition of H2O2, with consequent generation of free radicals (FR) and hydroxyl radicals (OH•). The data were submitted to the one-way analysis of variance complemented by Tukey post-test (α=0.05). Data on kinetics of H2O2 decomposition were submitted to the Student's-t test (α=0.05). RESULTS: All the BG35%s with MnO2 showed stability of pH and temperature, and the gel with 10 mg/mL of this activator had an esthetic efficacy 31% higher than that of the PC (p<0.05). Reduction in OXS and trans-amelodentinal diffusion of H2O2 occurred when all the BG35%s with MnO2 were used. The addition of 6 and 10 mg/mL of MnO2 to BG35% increased the CV in comparison with PC, confirmed by the cell morphology analysis. An increase in FR and OH• formation was observed when 10 mg/mL of MnO2 was added to BG35%. CONCLUSION: Catalysis of BG35% with MnO2 minimized the trans-amelodentinal diffusion of H2O2 and toxicity of the product to pulp cells. BG35% containing 10 mg/mL of MnO2 potentiated the decomposition of H2O2, enhancing the generation of FR and OH•, as well as the efficacy of the in-office tooth therapy.

Chitosan-Calcium-Simvastatin Scaffold as an Inductive Cell-Free Platform.

The development of biomaterials based on the combination of biopolymers with bioactive compounds to develop delivery systems capable of modulating dentin regeneration mediated by resident cells is the goal of current biology-based strategies for regenerative dentistry. In this article, the bioactive potential of a simvastatin (SV)-releasing chitosan-calcium-hydroxide (CH-Ca) scaffold was assessed. After the incorporation of SV into CH-Ca, characterization of the scaffold was performed. Dental pulp cells (DPCs) were seeded onto scaffolds for the assessment of cytocompatibility, and odontoblastic differentiation was evaluated in a microenvironment surrounded by dentin. Thereafter, the cell-free scaffold was adapted to dentin discs positioned in artificial pulp chambers in direct contact with a 3-dimensional (3D) culture of DPCs, and the system was sealed to simulate internal pressure at 20 cm/H2O. In vivo experiments with cell-free scaffolds were performed in rats' calvaria defects. Fourier-transform infrared spectroscopy spectra proved incorporation of Ca and SV into the scaffold structure. Ca and SV were released upon immersion in a neutral environment. Viable DPCs were able to spread and proliferate on the scaffold over 14 d. Odontoblastic differentiation occurred in the DPC/scaffold constructs in contact with dentin, in which SV supplementation promoted odontoblastic marker overexpression and enhanced mineralized matrix deposition. The chemoattractant potential of the CH-Ca scaffold was improved by SV, with numerous viable and dentin sialoprotein-positive cells from the 3D culture being observed on its surface. Cells at 3D culture featured increased gene expression of odontoblastic markers in contact with the SV-enriched CH-Ca scaffold. CH-Ca-SV led to intense mineralization in vivo, presenting mineralization foci inside its structure. In conclusion, the CH-Ca-SV scaffold induces differentiation of DPCs into a highly mineralizing phenotype in the presence of dentin, creating a microenvironment capable of attracting pulp cells to its surface and inducing the overexpression of odontoblastic markers in a cell-homing strategy.

Influence of enamel/dentin thickness on the toxic and esthetic effects of experimental in-office bleaching protocols.

OBJECTIVES: This paper aims to assess the whitening effectiveness and toxicity of tooth-bleaching protocols applied to enamel/dentin disks simulating mandibular incisors (ICs) and premolars (PMs). MATERIALS AND METHODS: A 10% hydrogen peroxide (H2O2) gel was applied for 3 × 15, 1 × 15, or 1 × 5 min to enamel/dentin disks simulating mandibular ICs and PMs, and the trans-enamel and trans-dentinal diffusion products were applied to human dental pulp cells (1 h). Professional therapy (35% H2O2-3 × 15 min) was used as positive control, and non-bleached samples were used as negative control. Cell viability and morphology, oxidative stress generation, and odontoblastic marker expression were assessed. The H2O2 diffusion and enamel color change (ΔE) were also analyzed. RESULTS: The 10% H2O2 gel induced significant cell viability reduction only when applied 3 × 15 min, with the intensity of oxidative stress and down-regulation of odontoblastic markers being higher in the IC group. The other experimental bleaching protocols caused slight alterations regarding the cell parameters evaluated, with intensity being related to enamel/dentin thickness. These effects were also correlated with higher H2O2 diffusion in the IC group. ΔE values similar as positive control were found for the 10% 3 × 15 and 1 × 15 protocols on IC group, after 4 and 6 sessions. CONCLUSION: Application of a 10% H2O2 bleaching gel for 15 or 45 min to thin dental substrate significantly minimizes cell toxicity in comparison with highly concentrated gels associated with similar esthetic outcomes by increasing the number of bleaching sessions. CLINICAL RELEVANCE: Bleaching gels with 10% H2O2 applied in small teeth for short periods may be an interesting alternative to obtain whitening effectiveness without causing toxicity to pulp cells, which may be able to reduce the tooth hypersensitivity claimed by patients.

Nutritional deprivation and LPS exposure as feasible methods for induction of cellular - A methodology to validate for vitro photobiomodulation studies.

Previous studies have demonstrated that high biostimulation takes place when cells under stress are subjected to phototherapy by laser or light-emitting-diode (LED) devices. Several studies selected nutritional deprivation by reducing the concentration of fetal bovine serum (FBS) in the culture medium or the exposure of cultured cells to lipopolysaccharide (LPS) as an in vitro cellular stress condition. However, there are no data certifying that these stimuli cause stressful conditions for cultured cells. This investigation assessed the induction of cellular stress by decreasing the concentration of FBS or adding LPS to culture medium. Odontoblast-like cells (MDPC-23) were cultured in complete culture medium (DMEM) containing 10% FBS. After a 12-hour incubation period, the DMEM was replaced by fresh medium containing 10% FBS (control), low concentrations of FBS (0, 0.2, 0.5, 2, or 5%) or LPS from Escherichia coli (10µg/ml). After an additional 12-hour incubation, cell viability, total cell-counting, total protein production, and gene expression of heat shock protein 70 (HSP70) were assessed. Data were statistically analyzed by ANOVA complemented by the Tukey test, with 5% considered significant. Cell viability was negatively affected only for 0% FBS, while reduced viable cell numbers and total protein production were detected for FBS concentrations lower than 2%. Higher HSP70 gene expression was also observed for FBS concentrations lower than 2% and for cells exposed to LPS. The nutritional deprivation model with culture medium lower than 2% of FBS can be safely used to induce cellular stress for in vitro photobiomodulation studies.

Indirect cytocompatibility of a low-concentration hydrogen peroxide bleaching gel to odontoblast-like cells.

AIM: To assess the initial cytotoxicity and the late phenotype marker expression of odontoblast-like cells (MDPC-23) subjected to less aggressive in-office bleaching therapies. METHODOLOGY: A 17.5% hydrogen peroxide (H2O2) gel was applied for 45, 15 or 5 min to enamel/dentine discs adapted to trans-wells positioned over cultured MDPC-23 cells. No treatment was performed on the negative control. Immediately after bleaching, the cell viability, gene expression of inflammatory mediators and quantification of H2O2 diffusion were evaluated. The ALP activity, DSPP and DMP-1 gene expression and mineralized nodule deposition (MND) were assessed at 7, 14 or 21 days post-bleaching and analysed statistically with Mann-Whitney U-tests (α = 5%). RESULTS: H2O2 diffusion, proportional to treatment time, was observed in all bleached groups. Reductions of approximately 31%, 21% and 13% in cell viability were observed for the 45-, 15- and 5-min groups, respectively. This reduction was significant (P < 0.05) for the 45- and 15-min groups, which also presented significant (P < 0.05) over-expression of inflammatory mediators. The 45-min group was associated with significant (P < 0.05) reductions in DMP-1/DSPP expression at all periods, relative to control. The ALP activity and MND were reduced only in initial periods. The 15-min group had less intense reduction of all markers, with no difference to control at 21 days. CONCLUSIONS: The 17.5% H2O2 applied to tooth specimens for 5 min caused no alteration in the odontoblast-like cells. When this gel was applied for 45 or 15 min, a slight cytotoxicity, associated with alterations in phenotypic markers, was observed. However, cells were able to recover their functions up to 21 days post-bleaching.

Cytocompatibility of HEMA-free resin-based luting cements according to application protocols on dentine surfaces.

AIM: To evaluate the transdentinal cytotoxicity of resin-based luting cements (RBLCs), with no HEMA in their composition, to odontoblast-like cells. METHODOLOGY: Human dentine discs 0.3 mm thick were adapted to artificial pulp chambers (APCs) and placed in wells of 24-well plates containing 1 mL of culture medium (DMEM). Two categories of HEMA-free RBLCs were evaluated: group 1, self-adhesive Rely X Unicem (RU; 3M ESPE), applied directly to the dentine substrate; and group 2, Rely X ARC (RARC; 3M ESPE), applied to dentine previously acid-etched and treated with a bonding agent. In group 3 (control), considered as representing 100% cell metabolic activity, no treatment was performed on dentine. The APC/disc sets were incubated for 24 h or 7 days at 37 °C and 5% CO2 . Then, the extracts (DMEM + dental materials components that diffused through dentine) were applied to cultured odontoblast-like MDPC-23 cells for 24 h. After that, the cell viability (MTT assay), cell morphology (SEM), total protein production (TP) and alkaline phosphatase (ALP) activity were assessed. Data from MTT assay and TP production were analysed by Kruskal-Wallis and Mann-Whitney tests (α = 5%). Data from ALP activity were analysed by one-way anova and Tukey's test (α = 5%). RESULTS: In group 1, a slight reduction in cell viability (11.6% and 16.8% for 24-h and 7-day periods, respectively) and ALP activity (13.5% and 17.9% for 24-h and 7-day periods, respectively) was observed, with no significant difference from group 3 (control) (P > 0.05). In group 2, a significant reduction in cell viability, TP production and ALP activity compared with group 3 (control) occurred (P < 0.05), regardless of incubation time. Alteration in MDPC-23 cell morphology was observed only in group 2. CONCLUSIONS: HEMA-free Rely X ARC cement caused greater toxicity to odontoblast-like MDPC-23 cells than did Rely X Unicem cement when both resin-based luting materials were applied to dentine as recommended by the manufacturer.

Effect of LPS treatment on the viability and chemokine synthesis by epithelial cells and gingival fibroblasts.

OBJECTIVE: Several local factors can affect the wound-healing process, delaying its progression and postponing tissue homeostasis. It is known that local inflammation is related to wound healing; however, the maintenance of the inflammatory reaction can impair the proliferation and migration of oral mucosal cells. The aim of this study was to evaluate the viability and chemokine expression of epithelial cells and gingival fibroblasts exposed to long-term lipopolysaccharide (LPS) treatment. DESIGN: Epithelial cells (HaCaT, Cell Lines Service, 300493) and human gingival fibroblasts (HGFs) were seeded (1×10(5) cells/well) in 24-well plates and incubated for 24h. To simulate the responses of cells to a local chronic oral mucosal inflammation, we added LPS of Escherichia coli (10 µg/ml) to Dulbecco's modified Eagle's medium (DMEM), kept in contact with fibroblasts and epithelial cells for 24, 48, and 72h. Then the cells were assessed for viability (alamarBlue assay), number (trypan blue assay), and expression of CCL2 and CCL5 inflammatory chemokines (enzyme-linked immunosorbent assay (ELISA)). Data were statistically analyzed by nonparametric Kruskal-Wallis and Mann-Whitney tests at a significance level of 5%. RESULTS: Cell treatment with LPS caused significant decrease in viability for both cell lines. No time-dependent effect was observed for epithelial cells. However, reduction in fibroblast viability was greater at 48 and 72 h. CCL2 and CCL5 synthesis was significantly increased for both LPS-treated cells, and this expression decreased with time. CONCLUSION: The maintenance of an inflammatory cell stimulus by LPS decreases the number and viability of cultured oral mucosal cells, which may be related to delayed wound healing.

Intensive chemotherapy as salvage treatment for solid tumors: focus on germ cell cancer

Germ cell tumors present contrasting biological and molecular features compared to many solid tumors, which may partially explain their unusual sensitivity to chemotherapy. Reduced DNA repair capacity and enhanced induction of apoptosis appear to be key factors in the sensitivity of germ cell tumors to cisplatin. Despite substantial cure rates, some patients relapse and subsequently die of their disease. Intensive doses of chemotherapy are used to counter mechanisms of drug resistance. So far, high-dose chemotherapy with hematopoietic stem cell support for solid tumors is used only in the setting of testicular germ cell tumors. In that indication, high-dose chemotherapy is given as the first or late salvage treatment for patients with either relapsed or progressive tumors after initial conventional salvage chemotherapy. High-dose chemotherapy is usually given as two or three sequential cycles using carboplatin and etoposide with or without ifosfamide. The administration of intensive therapy carries significant side effects and can only be efficiently and safely conducted in specialized referral centers to assure optimum patient care outcomes. In breast and ovarian cancer, most studies have demonstrated improvement in progression-free survival (PFS), but overall survival remained unchanged. Therefore, most of these approaches have been dropped. In germ cell tumors, clinical trials are currently investigating novel therapeutic combinations and active treatments. In particular, the integration of targeted therapies constitutes an important area of research for patients with a poor prognosis.

Intensive chemotherapy as salvage treatment for solid tumors: focus on germ cell cancer.

Germ cell tumors present contrasting biological and molecular features compared to many solid tumors, which may partially explain their unusual sensitivity to chemotherapy. Reduced DNA repair capacity and enhanced induction of apoptosis appear to be key factors in the sensitivity of germ cell tumors to cisplatin. Despite substantial cure rates, some patients relapse and subsequently die of their disease. Intensive doses of chemotherapy are used to counter mechanisms of drug resistance. So far, high-dose chemotherapy with hematopoietic stem cell support for solid tumors is used only in the setting of testicular germ cell tumors. In that indication, high-dose chemotherapy is given as the first or late salvage treatment for patients with either relapsed or progressive tumors after initial conventional salvage chemotherapy. High-dose chemotherapy is usually given as two or three sequential cycles using carboplatin and etoposide with or without ifosfamide. The administration of intensive therapy carries significant side effects and can only be efficiently and safely conducted in specialized referral centers to assure optimum patient care outcomes. In breast and ovarian cancer, most studies have demonstrated improvement in progression-free survival (PFS), but overall survival remained unchanged. Therefore, most of these approaches have been dropped. In germ cell tumors, clinical trials are currently investigating novel therapeutic combinations and active treatments. In particular, the integration of targeted therapies constitutes an important area of research for patients with a poor prognosis.

ATP-dependent chromatin remodeling and histone acetyltransferases in 5-FU cytotoxicity in Saccharomyces cerevisiae.

Chromatin is thought to modulate access of repair proteins to DNA lesions, and may be altered by chromatin remodelers to facilitate repair. We investigated the participation of chromatin remodelers and DNA repair in 5-fluorouracil (5-FU) cytotoxicity in Saccharomyces cerevisiae. 5-FU is an antineoplastic drug commonly used in clinical settings. Among the several strains tested, only those with deficiencies in ATP-dependent chromatin remodeling (CR) and some histone acetyltransferases (HAT) exhibited sensitivity to 5-FU. CR and HAT double-mutants exhibited increased resistance to 5-FU in comparison to the wild-type mutant, but were still arrested in G2/M, as were the sensitive strains. The participation of Htz1p in 5-FU toxicity was also evaluated in single- and double-mutants of CR and HAT; the most significant effect was on cell cycle distribution. 5-FU lesions are repaired by different DNA repair machineries, including homologous recombination (HR) and post-replication repair (PRR). We investigated the role of CR and HAT in these DNA repair pathways. Deficiencies in Nhp10 and CR combined with deficiencies in HR or PRR increased 5-FU sensitivity; however, combined deficiencies of HAT, HR, and PRR did not. CRs are directly recruited to DNA damage and lead to chromatin relaxation, which facilitates access of HR and PRR proteins to 5-FU lesions. Combined deficiencies in HAT with defects in HR and PRR did not potentiate 5-FU cytotoxicity, possibly because they function in a common pathway.

Contaminant concentrations, biochemical and hematological biomarkers in blood of West Indian manatees Trichechus manatus from Brazil.

The West Indian manatee Trichechus manatus is threatened with extinction in Brazil, and this study focused on nondestructive blood samples analyzed for metals, polychlorinated biphenyls (PCBs) and organochlorine pesticides (OCPs), as well as biochemical and hematological biomarkers. Studied manatees were kept at Projeto Peixe-Boi headquarters in Pernambuco State, and at two natural areas in estuaries where they are released to the wild. Manatees kept at the natural estuary in Paraiba State have blood concentrations of Al, Pb, Cd, Sn that are 11, 7, 8 and 23 times greater, respectively, than the concentrations found in blood of animals from the same species in Florida, USA. An inhibition of butyrylcholinesterase in manatees kept at the two reintroduction sites in Alagoas and Paraiba States indicated possible exposure of the animals to cholinesterase inhibitor insecticides. PCBs and OCPs were not detected. Results from this study will help delineate conservation efforts in the region.

Transenamel and transdentinal cytotoxicity of carbamide peroxide bleaching gels on odontoblast-like MDPC-23 cells.

AIM: To evaluate the transenamel and transdentinal cytotoxicity of bleaching gels based on carbamide peroxide (CP) on odontoblast-like cells after different contact times of the products with enamel. METHODOLOGY: Enamel/dentine discs were obtained from bovine incisors and placed in artificial pulp chambers. Bleaching gels containing 10% or 16% CP were applied for 8 h day(-1) on the enamel side of the discs during periods of 1, 7 or 14 days. Deionized water and artificial saliva served as controls. The extracts (culture medium plus bleaching gel products that diffused through the discs) were collected and applied on previously cultured MDPC-23 cells for 1 h. Cell metabolism was evaluated by the MTT assay, and the data were analysed statistically by one-way anova and Tukey's test (α=0.05). Cell morphology was analysed by SEM. RESULTS: There was no significant difference (P>0.05) between the controls and the groups bleached with 10% CP gel. In the groups bleached with 16% CP gel, however, cell metabolism decreased significantly (P<0.05) by 40.32%, 30.16% and 26.61% at 1, 7 and 14 days, respectively. There was no significant difference (P>0.05) between 1, 7 or 14 applications of the gels for either of the CP concentrations. CONCLUSION: Regardless of the number of applications on an enamel surface, the 10% CP bleaching gel did not cause transenamel and transdentinal cytotoxicity to the MDPC-23 cell cultures. However, diffusion of products from the 16% CP gel through enamel and dentine and cytopathic effects to the pulp cells occurred even after a single application of this product on enamel.