Activity of Matrix Metalloproteinases in Bovine versus Human Dentine

Metalloproteinases (MMPs) have been implicated with metabolism of collagen in physiological and pathological processes in human dentine. As bovine teeth have been used as a substitute for human teeth in laboratory analysis, this study evaluated the activity of MMP-2 and -9 in bovine versus human dentine. Bovine and human dentine fragments, from crowns and roots, were powderized. Protein extraction was performed by two protocols: a neutral extraction with guanidine-HCl/EDTA (pH 7.4) and an acidic extraction with citric acid (pH 2.3). Gelatinolytic activities of extracts were revealed by zymography. MMP-2 and -9 were detected in crown and root dentine from bovine and human teeth. Total activities of MMP-2 were 11.4 +/- 2.2, 14.6 +/- 2.0, 9.7 +/- 1.2 and 12.4 +/- 0.9 ng/ml for bovine root, human root, bovine crown and human crown dentine, respectively. Corresponding activities for MMP-9 were 14.9 +/- 2.0, 15.3 +/- 1.3, 15.4 +/- 1.3 and 15.5 +/- 1.3 ng/ml, respectively. Bovine dentine was found to be a reliable substrate for studies involving the activity of MMP-2 and -9. Copyright (C) 2011 S. Karger AG, Basel

Influence of human dentine on the antibacterial activity of self-etching adhesive systems against cariogenic bacteria

Objectives: The incorporation of antibacterial agents into adhesive systems has been proposed to eliminate residual bacteria from dentine. This study used the agar diffusion method to evaluate the antibacterial activity of Clearfil Protect Bond (CPB), Clearfil SE Bond (CSEB), Clearfil Tri-S Bond (C3SB) and Xeno-III (XIII) self-etching adhesive systems, with or without light-activation, against cariogenic bacteria, and to assess the influence of human dentine on the antibacterial activity of these materials.Methods: An aliquot of 10 mu l per material (and individual components) were pipetted onto paper and dentine discs distributed in Petri dishes containing bacterial culture in BHI agar. Positive control was 0.2% chlorhexidine digluconate (CHX).Results: After incubation, the adhesive components of CPB and CSEB, liquid A of XIII and C3SB did not present antibacterial activity when applied to paper discs. The non-light-activated CPB primer + adhesive promoted the greatest inhibition of Streptococcus mutans (p < 0.05), whereas with light-activation, there was no significant difference between primer + adhesive and primer alone. For Lactobacillus acidophilus, CPB primer presented the greatest antibacterial activity in both light-activation conditions (p < 0.05). Regarding the dentine discs, only CHX promoted an inhibitory effect, though less intense than on paper discs (p < 0.05). CHX presented greater antibacterial activity against S. mutans than against L. acidophilus (p < 0.05).Conclusions: Light-activation significantly reduced the antibacterial activity of the self-etching adhesive systems; MDPB incorporation contributed to the effect of adhesive systems against cariogenic bacteria; the components eluted from the adhesive systems were not capable to diffuse through 400 mu m-thick dentine disc to exert their antibacterial activity against cariogenic bacteria. (C) 2008 Elsevier Ltd. All rights reserved.

Morphological analysis of human and bovine dentine by scanning electron microscope investigation

Objective: The objective of this study was to compare the superficial morphology of bovine and human sclerotic dentine. Design: For the morphological analysis, bovine (n = 3) and human (n = 3) incisors exhibiting exposed dentine were used. Dentine presented characteristics of sclerosis: brownish, smooth and shiny-the vitreous appearance. The teeth were prepared for assessment on a scanning electron microscope (SEM). Three pre-determined areas of each sample were submitted to SEM. The number of open tubules per area was obtained from the electron micrographs (n = 9 per group) for comparison purposes. Results: The number of open tubules in both species compared were similar (p > 0.05). Human dentine presented 31.89 +/- 23.94 open tubules per area, whereas bovine dentine showed 30.33 +/- 18.14 open tubules per area. Conclusion: Based on the results, we concluded that dentine exposed at the incisal surface of human and bovine teeth presented similar clinical and micro-morphological aspects, represented by surfaces with equivalent numbers of open dentinal tubules. (C) 2007 Elsevier Ltd. All rights reserved.

Caries progression and inhibition in human and bovine root dentine in situ

Since the use of bovine instead of human dentine to evaluate cariogenic and anticariogenic substances is not well established, this in situ study was conducted. Eleven volunteers wore palatal acrylic devices containing 4 dentine slabs (2 human and 2 bovine). Sucrose solution (20%) was dripped over all slabs 4 times a day, simulating a cariogenic challenge. Dentifrice slurries, fluoridated or not, were dripped over specified dentine slabs 3 times a day to evaluate caries reduction. After 14 days, the biofilm formed on the dentine slabs was collected for microbiological analysis. In dentine, mineral loss (DeltaZ) and lesion depth (LD) were determined by cross-sectional microhardness and by polarized light microscopy, respectively. The total streptococci and mutans streptococci counts in the biofilm formed either on human or on bovine slabs, whether treated or not with fluoride dentifrice, were not statistically different. The DeltaZ and the LID values of dentine treated with fluoride dentifrice were significantly lower than the values of dentine treated with non-fluoride dentifrice. The differences in the DeltaZ and LD values between the human and bovine dentine were not statistically significant. The results suggest that bovine dentine can be used instead of human to evaluate caries development and inhibition. Copyright (C) 2003 S. Karger AG, Basel.

Photodynamic antimicrobial therapy of curcumin in biofilms and carious dentine

Photodynamic therapy (PDT) is a technique that involves the activation of photosensitizers by light in the presence of oxygen, resulting in the production of reactive radicals that are capable of inducing cell death. The present study evaluated the susceptibility of Streptococcus mutans and Lactobacillus acidophilus to PDT grown as multi-species in the biofilm phase versus in dentine carious lesions. A brain-heart infusion culture medium supplemented with 1 % glucose, 2 % sucrose, and 1 % young primary culture of L. acidophilus 108 CFU/mL and S. mutans 108 CFU/mL was used to develop multi-species biofilms and to induce caries on human dentine slabs. Five different concentrations of curcumin (0.75, 1.5, 3.0, 4.0, and 5.0 g/L) were used associated with 5.7 J/cm2 light emission diode. Four different groups were analyzed L-D- (control group), L-D+ (drug group), L+D- (light group), and L+D+ (PDT group). ANOVA/Tukey's tests were conducted to compare groups. A significant reduction (p <0.05) in cell viability was observed in the biofilm phase following photosensitization with all curcumin concentrations tested. To achieve significant bacterial reduction (p <0.05) in carious dentine, it was necessary to utilize 5.0 g/L of curcumin in association with blue light. No significant reduction was found for L-D+, supporting the absence of the drug's dark toxicity. S. mutans and L. acidophilus were susceptible to curcumin in the presence of blue light. However, due to light penetration and drug diffusion difficulties, these microorganisms within dentine carious lesions were less affected than they were in the biofilm phase. © 2013 Springer-Verlag London.

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

To evaluate the transdentinal cytotoxicity of resin-based luting cements (RBLCs), with no HEMA in their composition, to odontoblast-like cells. 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%). 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. 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.

Adhesion of Endodontic Sealers to Human Root Dentine Submitted to Different Surface Treatments

Objective: To evaluate the adhesion of the endodontic sealers Epiphany, Apexit Plus, and AH Plus to root canal dentin submitted to different surface treatments, by using the push-out test. Methods: One hundred twenty-eight root cylinders obtained from maxillary canines were embedded in acrylic resin, had the canals prepared, and were randomly assigned to four groups (n = 32), according to root dentin treatment: (I) distilled water (control), (II) 17% EDTAC, (III) 1% NaOCl and (IV) Er:YAG laser with 16-Hz, 400-mJ input (240-mJ output) and 0.32-J/cm(2) energy density. Each group was divided into four subgroups (n = 8) filled with Epiphany (either dispensed from the automix syringe supplied by the manufacturer or prepared by hand mixing), Apexit Plus, or AH Plus. Data (MPa) were analyzed by ANOVA and Tukey's test. Results: A statistically significant difference (p < 0.01) was found among the root-canal sealers, except for the Epiphany subgroups, which had statistically similar results to each other (p > 0.01): AH Plus (4.77 +/- 0.85), Epiphany/hand mixed (3.06 +/- 1.34), Epiphany/automix syringe (2.68 +/- 1.35), and Apexit Plus (1.22 +/- 0.33). A significant difference (p < 0.01) was found among the dentin surface treatments. The highest adhesion values were obtained with AH Plus when root dentin was treated with Er: YAG laser and 17% EDTAC. Epiphany sealer presented the lowest adhesion values to root dentin treated with 17% EDTAC. Conclusions: The resin-based sealers had different adhesive behaviors, depending on the treatment of root canal walls. The mode of preparation of Epiphany (automix syringe or hand mixing) did not influence sealer adhesion to root dentin.

Effects of copper vapor laser irradiation (λ = 510.6 nm) on the enamel and dentine of human teeth: An ultra-structural morphologic study

Objective: A morphological and ultra-structural study of copper vapor laser (λ = 510.6 nm) effects on enamel and dentine was performed to show the effects of this radiation. Methods: A total of 15 human molars were cut in half; 15 pieces were separated for irradiation on enamel and 15 for dentine. These two groups were further divided into five experimental groups, including a control group, comprised of three half-sections each, irradiated by a CVL laser with a power of 7 W, a repetition rate of 15,000 pulses/sec and exposed at 500, 600, and 800 msec and 1 sec irradiation times with a 5-sec interval between irradiations. Results: In an ultra-structural SEM exam, we observed that on the enamel surfaces irradiated for 1 sec there was morphological alteration that consisted of catering, flaking, and melting on the surfaces. There was no alteration for the other exposure times. On the dentine teeth irradiated for 1 sec, we observed an evident ultra-structural alteration of melted tissue and loss of morphological characteristics. In the dentine group irradiated by 800 msec, we observed ablation and a partial loss of morphological characteristics. In the dentine groups irradiated by 500 and 600 msec, no alteration was observed. Conclusions: The results showed that irradiation with CVL promoted morphologic changes in the enamel as well as in the dentine and demonstrated a need for future studies in order to establish a safe protocol for further use in the odontological practice.

Effect of Nd:YAG Irradiation and Fluoride Application on Dentine Resistance to Erosion in Vitro

Objective: In this paper we evaluated the effect of two fluoridated agents and Nd:YAG irradiation separately and in combination on dentine resistance to erosion. Background Data: The morphological changes in dentin induced by laser treatment may reduce the progression of erosive lesions. Due to the possibility of a synergistic effect of laser with fluoride, this study was conducted. Materials and Methods: Eighty bovine dentine samples (4 x 4 mm) were randomly divided into eight groups, according to the following treatments: G1: untreated (control); G2: acidic phosphate fluoride gel (APF 1.23%) for 4 min; G3: fluoride varnish (NaF 2.26%) for 6 h; G4: 0.5 W Nd: YAG laser (250 mu sec pulse, 10 Hz, 35 J/cm(2), 30 sec); G5: 0.75 W Nd: YAG laser (52.5 J/cm(2)); G6: 1.0 W Nd: YAG laser (70 J/cm(2)); G7: APF + 0.75 W Nd: YAG laser; and G8: NaF + 0.75 W Nd: YAG laser. After the treatments, half of each dentine surface was protected with nail varnish. The samples were stored in artificial saliva (30 mL/sample) for 24 h and submitted to four erosive 1-min cycles. Between the erosive attacks, the blocks were maintained in artificial saliva for 59 min. The erosive wear was evaluated by profilometry. Results: The mean wear (+/- SD, mu m) was: G1: 1.20 +/- 0.20; G2: 0.47 +/- 0.06; G3: 0.81 +/- 0.11; G4: 1.47 +/- 0.32; G5: 1.52 +/- 0.24; G6: 1.49 +/- 0.30; G7: 0.49 +/- 0.11; and G8: 1.06 +/- 0.31 (Tukey's test, p < 0.05). Conclusions: Laser irradiation was not able to reduce dentine erosion. However, fluoride application was able to increase the dentine's resistance to erosion, and APF showed better results than fluoride varnish.

Dentine caries inhibition through CO(2) laser (10.6 mu m) irradiation and fluoride application, in vitro

Objective: The purpose of the study was to investigate whether dentine irradiation with a pulsed CO(2) laser (10.6 mu m) emitting pulses of 10 ms is capable of reducing dentine calcium and phosphorus losses in an artificial caries model. Design: The 90 dentine slabs obtained from bovine teeth were randomly divided into six groups (n = 15): negative control group (GC); positive control group, treated with fluoride 1.23% (GF); and laser groups irradiated with 8 J/cm(2) (L8); irradiated as in L8 + fluoride 1.23% (L8F); irradiated with 11j/cm(2) (L11); irradiated as in L11 + fluoride 1.23% (L11F). After laser irradiation the samples were submitted to a pH-cycling model for 9 days. The calcium and phosphorous contents in the de- and remineralization solutions were measured by means of inductively coupled plasma optical emission spectrometer - ICP-OES. Additionally intra-pulpal temperature measurements were performed. The obtained data were analysed by means of ANOVA and Tukey`s test (alpha = 0.05). Results: In the demineralization solutions the groups L11F and GF presented significantly lower means of calcium and phosphorous losses than the control group; and in L11F means were significantly lower than in the fluoride group. Both irradiation parameters tested caused intrapulpal temperature increase below 2 degrees C. Conclusion: It can be concluded that under the conditions of this study, CO(2) laser irradiation (10.6 mu m) with 11J/cm(2) (540 mJ and 10 Hz) of fluoride treated dentine surfaces decreases the loss of calcium and phosphorous in the demineralization process and does not cause excessive temperature increase inside the pulp chamber. (C) 2010 Elsevier Ltd. All rights reserved.

Micro-tensile bond strength to bovine sclerotic dentine: Influence of surface treatment

Objective: The objective of this study was to evaluate the influence of the surface treatment and acid conditioning (AC) time of bovine sclerotic dentine on the micro-tensile bond strength (mu-TBS) to an etch and rinse adhesive system. Materials and method: Thirty-six bovine incisors were divided into six groups (n = 6): G1 sound dentine submitted to AC for 15 s; G2-G6 sclerotic dentine: G2-AC for 15 s; G3-AC for 30 s; G4-EDTA and AC for 15 s; G5-diamond bur and AC for 15 s; G6-diamond paste and AC for 15 s. An adhesive system was applied to the treated dentine surfaces followed by a hybrid composite inserted in increments and light cured. After 24 h storage in water at 37 degrees C, the specimens were perpendicularly cut with a low-speed diamond saw to obtain beams (0.8 mm x 0.8 mm cross-sectional dimensions) for mu-TBS testing. Data was compared by ANOVA followed by Tukey`s test (P <= 0.05). Results: The mean L-TBS was G1: 18.87 +/- 5.36 MPa; G2: 12.94 +/- 2.09 MPa; G3: 11.73 +/- 0.64 MPa; G4: 11.14 +/- 1.50 MPa; G5: 22.75 +/- 4.10 MPa; G6: 22.48 +/- 2.71 MPa. G1, G5 and G6 presented similar bond strengths significantly higher than those of all other groups. Conclusion: The surface treatment of sclerotic dentine significantly influenced the bond strength to an adhesive system. Mechanical treatment, either using a diamond bur or a diamond paste was able to improve bonding to bovine sclerotic dentine, reaching values similar to bonding to sound dentine. (C) 2008 Elsevier Ltd. All rights reserved.

Effect of Iron on Matrix Metalloproteinase Inhibition and on the Prevention of Dentine Erosion

It is known that some metal salts can inhibit matrix metalloproteinase (MMP) activity, but the effect of iron has not been tested yet. On the other hand, it has recently been suggested that MMP inhibition might influence dentine erosion. Based on this, the aims of this study were: (1) to test in vitro the effect of FeSO(4) on MMP-2 and -9 activity, and (2) to evaluate in situ the effect of FeSO(4) gel on dentine erosion. MMP-2 and -9 activities were analysed zymographically in buffers containing FeSO(4) in concentrations ranging between 0.05 and 1.5 mmol/l or not. Volunteers (n = 10) wore devices containing bovine dentine blocks (n = 60) previously treated with the following gel treatments: FeSO(4) (1 mmol/l FeSO(4)), F (NaF 1.23%; positive control) and placebo (negative control). The gels were applied once and removed after 1 min. Erosion was performed extraorally with Coca-Cola 4 times per day for 5 min over 5 days. Dentine wear was evaluated by profilometry. The data were analysed by Kruskal-Wallis and Dunn`s tests (p < 0.05). FeSO(4) inhibited both MMP-2 (IC(50) = 0.75 mmol/l) and MMP-9 (IC(50) = 0.50 mmol/l) activities. In the in situ experiment, the mean wear (+/- SD) found for the F gel (0.79 8 +/- 0.08 mu m) was significantly reduced in more than 50% when compared to the placebo gel (1.77 +/- 0.33 mu m), but the FeSO(4) gel completely inhibited the wear (0.05 +/- 0.02 mu m). Since FeSO(4) was able to inhibit MMP in vitro, it is possible that the prevention of dentine wear by the FeSO(4) gel in situ might be due to MMP inhibition, which should be investigated in further studies. Copyright (C) 2010 S. Karger AG, Basel

Resistance to degradation of resin-modified glass-ionomer cements dentine bonds

Objective: To evaluate the effect of EDTA pre-treatment of dentine on resistance to degradation of the bond between dentine and resin-modified glass-ionomer cements. Methods: Sixty non-carious human molars underwent cavity preparations. Teeth were restored with Fuji II LC or Vitremer. Half of the cavities were restored following manufacturers` instructions whereas the other half was pre-treated with EDTA (0.1 M, pH 7.4) for 60 s. Teeth were stored in water at 37 degrees C for 24 h, 3 months or submitted to 10% NaOCl immersion for 5 h. Teeth were sectioned into beams (1 +/- 0.1 mm) and tested to failure in tension at 0.5 mm/min. Bond strength data (MPa) were analyzed by ANOVA and SNK multiple-comparisons tests (p < 0.05). Results: When EDTA was used for pre-treatment of dentine, higher bond strengths were observed for both cements. Degradation challenges produced a decrease in bond strength values only in the Vitremer group. This decrease was avoided when EDTA was used for dentine treatment before restoring with Vitremer. Conclusions: EDTA pre-treatment of dentine increases bond strength of resin modified glass-ionomers cements to dentine and improves resistance to degradation of the bond between Vitremer and dentine. (C) 2009 Elsevier Ltd. All rights reserved.

Influence of glutaraldehyde priming on bond strength of an experimental adhesive system applied to wet and dry dentine

Objectives: This study tested the following null hypotheses: (1) there is no difference in resin-dentine bond strength when an experimental glutaraldehyde primer solution is added prior to bonding procedures and (2) there is no difference in resin-dentine bond strength when experimental glutaraldehyde/adhesive system is applied under dry or wet demineralized dentine conditions. Methods: Extracted human maxillary third molars were selected. Flat, mid-coronal dentine was exposed for bonding and four groups were formed. Two groups were designated for the dry and two for the wet dentine technique: DRY: (1) Group GD: acid etching + glutaraldehyde primer (primer A) + HEMA/ethanol primer (primer B)-under dried dentine + unfilled resin; (2) Group D: the same as GD, except for primer A application; WET: (3) Group GW: the same as GD, but primer B was applied under wet dentine condition; (4) Group W: the same as GW, except for primer A application. The bonding resin was light-cured and a resin core was built up on the adhesive layer. Teeth were then prepared for microtensile bond testing to evaluate bond strength. The data obtained were submitted to ANOVA and Tukey`s test (alpha = 0.05). Results: Glutaraldehyde primer application significantly improved resin-dentine bond strength. No significant difference was observed when the same experimental adhesive system was applied under either dry or wet dentine conditions. These results allow the first null hypothesis to be rejected and the second to be accepted. Conclusion: Glutaraldehyde may affect demineralized dentine properties leading to improved resin bonding to wet and dry substrates. (C) 2008 Elsevier Ltd. All rights reserved.