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 in association with osteogenic factors as a cell-homing platform for dentin regeneration: Analysis in a pulp-in-a-chip model.

OBJECTIVE: In this paper we propose the association of ß-glycerophosphate (ßGP) and calcium-hydroxide with chitosan (CH) to formulate a porous bioactive scaffold suitable as a cell-homing platform for dentin regeneration. METHODS: Calcium hydroxide and ßGP solutions were incorporated into chitosan to modulate scaffold architecture and composition by a phase separation technique. Architecture, chemical composition, and degradability were evaluated, and biological characterizations were performed by the seeding of dental pulp cells (DPCs) onto scaffolds, or by cultivating them in contact with leachable components (extracts), to determine cytocompatibility and odontoblastic differentiation. Cell-free scaffolds were then positioned in intimate contact with a 3D culture of DPCs in a pulp-in-a-chip platform under simulated pulp pressure. Cell mobilization and odontoblastic marker expression were evaluated. Deposition of mineralized matrix was assessed in direct contact with dentin, in the absence of osteogenic factors. RESULTS: Incorporation of calcium hydroxide and ßGP generated a stable porous chitosan scaffold containing Ca-P nanoglobule topography (CH-Ca-ßGP), which favored cell viability, alkaline phosphatase activity, and mineralized matrix deposition by cells seeded onto the scaffold structure and at a distance. The pulp-in-a-chip assay denoted its chemotactic and bioactive potential, since dentin sialoprotein-positive DPCs from 3D culture adhered to CH-Ca-ßGP more than to plain chitosan. The higher deposition of mineralized matrix onto the scaffold and surrounding dentin was also observed. SIGNIFICANCE: A CH-Ca-ßGP scaffold creates a microenvironment capable of mobilizing DPC migration toward its structure, harnessing the odontogenic potential and culminating in the expression of a highly mineralizing phenotype, key factors for a cell-homing strategy.

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 Tooth Pigmentation on H2O2 Diffusion and Its Cytotoxicity After In-office Tooth Bleaching.

CLINICAL RELEVANCE: Pigments in tooth structures affect the diffusion of H2O2 through enamel and dentin. The bleaching methodology can be impacted. SUMMARY: Objective: The aim of this study was to evaluate the influence of the presence of pigments in tooth structures on the trans-enamel and trans-dentin diffusion of hydrogen peroxide (H2O2) and its cytotoxicity after carrying out an in-office bleaching therapy.Methods and Materials: A bleaching gel with 35% H2O2 was applied for 45 minutes (three times for 15 minutes) on enamel and dentin discs (n=6), either previously submitted to the intrinsic pigmentation protocol with a concentrated solution of black tea, or not, defining the following groups: G1, unbleached untreated discs (control 1); G2, unbleached pigmented discs (control 2); G3, bleached untreated discs; G4, bleached pigmented discs. The discs were adapted to artificial pulp chambers, which were placed in wells of 24-well plates containing 1 mL culture medium (Dulbecco's modified Eagle's medium [DMEM]). After applying the bleaching gel on enamel, the extracts (DMEM + components of bleaching gel that diffused through the discs) were collected and then applied on the cultured MDPC-23 odontoblast-like cells. Cell viability (methyl tetrazolium assay and Live & Dead, Calcein AM, and ethidium homodimer-1 [EthD-1] probes), the amount of H2O2 that diffused through enamel and dentin (leuco-crystal violet product), and the H2O2-mediated oxidative cell stress (SOx) and components of degradation were assessed (analysis of variance/Tukey; α=0.05).Results: There was no significant difference between the groups G1 and G2 for all the parameters tested (p>0.05). Reduction in the trans-enamel and trans-dentin diffusion of H2O2 occurred for G4 in comparison with G3. Significantly lower cell viability associated with greater oxidative stress was observed for G3 (p<0.05). Therefore, in-office tooth bleaching therapy performed in pigmented samples caused lower cytotoxic effects compared with untreated samples submitted to the same esthetic procedure (p<0.05).Conclusion: According to the methodology used in this investigation, the authors concluded that the presence of pigments in hard tooth structures decreases the trans-enamel and trans-dentin diffusion of H2O2 and the toxicity to pulp cells of an in-office bleaching gel with 35% H2O2.

Effects of Enzymatic Activation of Bleaching Gels on Hydrogen Peroxide Degradation Rates, Bleaching Effectiveness, and Cytotoxicity.

OBJECTIVES: The aim of this study was to evaluate the effect of horseradish peroxidase (HRP) on the release of free radicals, bleaching effectiveness, and indirect cytotoxicity of a 35% hydrogen peroxide (HP) bleaching gel. METHODS AND MATERIALS: First, HP degradation rates and free radical release were evaluated for 35% HP in contact or not with HRP (10 mg/mL). The bleaching gel associated or not with HRP was then applied (3 × 15 minutes) to enamel/dentin discs adapted to artificial pulp chambers, and the culture medium in contact with dentin surfaces (extract) was collected and exposed to cultured odontoblast-like cells. Membrane damage and viability of cells as well as oxidative stress were evaluated. Residual HP/free radical diffusion was quantified, and bleaching effectiveness (ΔE) was assessed. Unbleached discs served as negative controls. RESULTS: The addition of HRP to the 35% HP bleaching gel enhanced the release of free radicals in comparison with plain HP gel. The 35% HP-mediated cytotoxicity significantly decreased with HRP in the bleaching gel and was associated with reduced HP/free radical diffusion through the enamel/dentin discs. ΔE values increased every bleaching session for HRP-containing gel relative to positive control, accelerating the whitening outcome. CONCLUSION: The enzymatic activation of a 35% HP bleaching gel with HRP accelerated HP degradation mediated by intensification of free radical release. This effect optimized whitening outcome as well as minimized residual HP and free radical diffusion through enamel and dentin, decreasing the harmful effects on odontoblast-like cells.

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 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.

Influence of Restoration Type on the Cytotoxicity of a 35% Hydrogen Peroxide Bleaching Gel.

OBJECTIVES: The tooth/restoration interface may act as a pathway for hydrogen peroxide (H2O2) diffusion into the pulp chamber. Therefore, the influence of resin-modified glass ionomer cement (RMGIC) and resin composite simulated restorations on the cytotoxicity of an in-office bleaching gel was assessed in vitro. MATERIALS AND METHODS: Cavities in enamel/dentin discs restored with RMGIC Vitremer (3M ESPE) or Single Bond/Filtek Z350 (3M ESPE) resin composite (RC) were subjected or not subjected to hydrolytic degradation (HD). A 35%-H2O2 bleaching gel was applied to simulated restored and nonrestored enamel surfaces, and culture medium in contact with the dentin substrate (extract) was collected and applied to MDPC-23 cells. Nonrestored discs subjected or not subjected to bleaching were used as positive and negative controls, respectively. Cell viability, oxidative stress, interleukin (IL)-1ß expression, alkaline phosphatase (ALP) activity, and mineralized nodule deposition were evaluated. The H2O2 in the extracts was quantified. Data were subjected to statistical analysis. RESULTS: Higher oxidative stress associated with reduced cell viability, ALP activity, and mineralized nodule deposition was observed for all bleached groups compared with the negative control group. The RMGIC/HD group, which presented the highest H2O2 diffusion, had the lowest values of cell viability, ALP activity, and mineralized nodule deposition, as well as significantly increased IL-1ß expression. CONCLUSIONS: Dental cavities restored with the RMGIC subjected to hydrolytic degradation allowed for more intense diffusion of H2O2 into the pulp chamber, intensifying the toxicity of a 35%-H2O2 bleaching gel to pulp cells.

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.

Increased Durability of Resin-Dentin Bonds Following Cross-Linking Treatment.

OBJECTIVES: This study evaluated the long-term effect of carbodiimide treatments of acid-etched dentin on resin-dentin bond strength of a simplified etch-and-rinse adhesive system. METHODS: Forty-eight sound third molars were divided into three groups (n=16) according to the dentin treatment: G1: deionized water; G2: 0.5 mol/L 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide (EDC) applied for 30 seconds; and G3: 0.5 mol/L EDC applied for 60 seconds. Flat dentin surfaces were produced, etched with 37% phosphoric acid for 15 seconds, and then treated with deionized water for 60 seconds or with 0.5 mol/L EDC for 30 or 60 seconds prior to the application of Single Bond 2. Crowns were restored with resin composite, and beam specimens were prepared for microtensile testing. The beams from each group were tested 24 hours or 6 or 12 months after the adhesive procedures. One slab from each tooth was prepared and analyzed for nanoleakage. Bond strength (MPa) data were submitted to analysis of variance and Tukey test (α=0.05). RESULTS: The treatment of dentin with 0.5 mol/L EDC for 30 seconds (24.1±6.2 MPa) and 60 seconds (25.5±5.1 MPa) did not negatively affect the immediate bond strength of Single Bond 2 when compared to the control group (24.6±7.3 MPa). Additionally, EDC prevented resin-dentin bond degradation after 12 months in artificial saliva for both periods of treatment. An increased accumulation of silver ions was seen for the control group over time, while a much lower amount of silver grains was observed for the EDC-treated groups. CONCLUSIONS: 0.5 mol/L EDC was able to prevent resin-dentin bond degradation after 12 months, especially when applied for 60 seconds.

Cytotoxicity of dimethyl sulfoxide (DMSO) in direct contact with odontoblast-like cells.

OBJECTIVES: To evaluate the cytotoxicity of dimethyl sulfoxide (DMSO) on the repair-related activity of cultured odontoblast-like MDPC-23 cells. METHODS: Solutions with different concentrations of DMSO (0.05, 0.1, 0.3, 0.5 and 1.0 mM), diluted in culture medium (DMEM), were placed in contact with MDPC-23 cells (5 × 104 cells/cm(2)) for 24 h. Eight replicates (n = 8) were prepared for each solutions for the following methods of analysis: violet crystal dye for cell adhesion (CA), quantification of total protein (TP), alizarin red for mineralization nodules formation (MN) and cell death by necrosis (flow cytometry); while twelve replicates (n = 12) were prepared for viable cell number (Trypan Blue) and cell viability (MTT assay). Data were analyzed by ANOVA and Tukey or Kruskal-Wallis and Mann-Whitney's tests (p < 0.05). RESULTS: Cell viability, adhesion and percentage of cell death by necrosis were not affected by DMSO at any concentration, with no statistical significant difference among the groups. A significant reduction in total protein production was observed for 0.5 and 1.0 mM of DMSO compared to the control while increased mineralized nodules formation was seen only for 1.0 mM DMSO. SIGNIFICANCE: DMSO caused no or minor cytotoxic effects on the pulp tissue repair-related activity of odontoblast-like cells.

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.

Transdentinal cytotoxicity of carbodiimide (EDC) and glutaraldehyde on odontoblast-like cells.

OBJECTIVE: To evaluate the transdentinal cytotoxicity of three different concentrations of carbodiimide (EDC) or 5% glutaraldehyde (GA) on MDPC-23 cells. METHODS: Seventy 0.4-mm-thick dentin disks obtained from human molars were adapted to artificial pulp chambers. MDPC-23 cells were seeded on the pulpal surface of the disks. After 48 hours, the occlusal dentin was acid-etched and treated for 60 seconds with one of the following solutions (n=10): no treatment (negative control); 0.1 M, 0.3 M, or 0.5 M EDC; 5% GA; Sorensen buffer; or 29% hydrogen peroxide (positive control). Cell viability and morphology were assessed by methyltetrazolium assay and scanning electron microscopy (SEM), respectively. The eluates were collected after the treatments and applied on MDPC-23 seeded in a 24-well plate to analyze cell death, total protein (TP), and collagen production. The last two tests were performed 24 hours and seven days after the challenge. Data were analyzed by Kruskal-Wallis and Mann-Whitney tests (p<0.05). RESULTS: EDC at all test concentrations did not reduce cell viability, while 5% GA did increase cell metabolism. Cell death by necrosis was not elicited by EDC or 5% GA. At the 24-hour period, 0.3 M and 0.5 M EDC reduced TP production by 18% and 36.8%, respectively. At seven days, increased TP production was observed in all groups. Collagen production at the 24-hour period was reduced when 0.5 M EDC was used. After seven days, no difference was observed among the groups. SEM showed no alteration in cell morphology or number, except in the hydrogen peroxide group. CONCLUSIONS: Treatment of acid-etched dentin with EDC or GA did not cause transdentinal cytotoxic effects on odontoblast-like cells.

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.

A phase II trial of high-dose chemotherapy (HDCT) supported by hematopoietic stem-cell transplantation (HSCT) in germ-cell tumors (GCTs) patients failing cisplatin-based chemotherapy: the Multicentric TAXIF II study.

BACKGROUND: High-dose chemotherapy (HDCT) is an effective salvage treatment of germ-cell tumors (GCTs) patients. In the first salvage setting, 30%-70% of patients may achieve durable remissions. Even when HDCT is administered as subsequent salvage treatment, up to 20% of patients may still be definitively cured. However, patients with refractory/relapsed disease still have a very poor long-term prognosis, requiring earlier intervention of HDCT. PATIENTS AND METHODS: This phase II trial was addressed to nonrefractory patients failing Cisplatin-based chemotherapy. Inclusion criteria included seminomatous GCT in relapse after two lines of chemotherapy, nonseminomatous GCT in relapse after first or second lines, partial remission after first line, primary mediastinal GCT in first relapse. Patients received two cycles combining Epirubicin and Paclitaxel (Epi-Tax), followed by three consecutive HDCT, one using a Paclitaxel/Thiotepa (Thio-Tax) association and two using the 5-day Ifosfamide-Carboplatin-Etoposide regimen. The main objective was to determine the complete response rate. RESULTS: Forty-five patients were included between September 2004 and December 2007: 44 received the first HDCT cycle, 39 two HDCT cycles, 29 could receive the whole protocol. Sixteen patients did not receive the entire protocol, including eight (17.7%) for toxic side-effects. Two patients (4.4%) died of toxicities, and 17 (37.7%) of disease progression. With a median follow-up time of 26 months (range, 4-51), the final overall response rate was 48.8% (including a complete response rate of 15.5% and a partial response/negative serum markers rate of 26.6%) in an intent-to-treat analysis. The median progression-free survival (PFS) and overall survival (OS) times were 22 months [95% confidence interval (CI) 2-not reached] and 32 months (95% CI 4-49), respectively. The 2-year PFS was a plateau setup at 50% (95% CI 32-67) and the 2-year OS was 66% (95% CI 44-81). CONCLUSION: The TAXIF II protocol was effective in nonrefractory GCT patients failing Cisplatin-based chemotherapy. The toxic death rate remained acceptable in the field of HDCT regimens. TRIAL REGISTRATION NUMBER: NCT00231582.

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.