Safety and In Situ Antierosive Effect of CaneCPI-5 on Dental Enamel.

The sugarcane cystatin (CaneCPI-5) was recently cloned and showed strong binding force to dental enamel and protection against initial erosion. However, evaluations on its safety and efficacy in a situation closer to the clinical condition are necessary. In the present study we analyzed 1) the cytotoxicity of CaneCPI-5 on human gingival fibroblasts (HGFs); 2) the ability of CaneCPI-5 to reduce enamel erosion and erosion+abrasion in situ. In part 1, HGFs were treated with CaneCPI-5 (0.025, 0.05, 0.1, 0.5 or 1.0 mg/mL) or no treatment (control). The cytotoxicity was assessed after 60 s and 24 h by mitochondrial activity (MTT), confocal microscopy, and hematoxylin/eosin staining. In part 2, 15 volunteers participated in a double-blind crossover protocol consisting of 3 phases, according to the following treatments: 1) 0.1 mg/mL CaneCPI-5; 2) SnCl2/NaF/AmF (Elmex; positive control); 3) water (negative control). The volunteers wore an appliance containing 4 bovine enamel specimens for 5 d. Each day, the specimens were individually treated with 50 µL of the tested solutions per 60 s and then subjected to erosive challenges (0.1% citric acid, pH 2.5, for 90 s, 4 times per day). After the first and last erosive challenge each day, 2 samples were abraded (toothbrushing, 15 s). Enamel wear was measured by contact profilometry. One or two-way analysis of variance (ANOVA)/Tukey's or Sidak's tests (P < 0.05) were applied. Regardless of the concentration and the experimental time, CaneCPI-5 did not decrease the cell viability compared to the negative control (P < 0.05). Erosion+abrasion led to significantly greater wear compared to erosion only. For both conditions, the lowest wear was found for SnCl2 and CaneCPI-5, which did not differ significantly from each other, but showed significant protection when compared to the negative control. In conclusion, CaneCPI-5 is safe on HGFs and reduces enamel erosive wear to the same extent as a commercial solution used to control erosive tooth wear (ETW).

Quantitative proteomic analysis in symptomatic and asymptomatic apical periodontitis.

AIM: To quantitatively and qualitatively compare the host proteomic profile in samples of symptomatic and asymptomatic apical periodontitis (AP) using nano-liquid chromatography-electron spray tandem mass spectrometry. METHODOLOGY: Samples were obtained from 18 patients with radiographically evident AP, divided into symptomatic and asymptomatic groups (nine per group) according to clinical characteristics. After sample collection, protein extraction, purification and quantification of the samples were performed, which were analysed by reverse-phase liquid chromatography coupled to mass spectrometry. Label-free quantitative proteomic analysis was performed by Protein Lynx Global Service software. Differences in expression of proteins between the groups were calculated using the Monte Carlo algorithm, considering P < 0.05 for down-regulated proteins and 1 - P > 0.95 for up-regulated proteins. Proteins were identified with the embedded ion accounting algorithm in the software and a search of the Homo sapiens UniProt database. RESULTS: A total of 853 individual human proteins were identified. In the quantitative analysis, common proteins to both groups accounted for 143 proteins. Differences in expression between groups resulted in 51 up-regulated proteins (1 - P > 0.95) in the symptomatic group, including alpha-1-antitrypsin, protein S100-A8, myeloperoxidase, peroxiredoxin and lactotransferrin. This group also had 43 down-regulated proteins (P < 0.05), comprising immunoglobulin, neutrophil defensin, pyruvate kinase and alpha-enolase. The qualitative analysis considered only the exclusive proteins of each group. For the symptomatic group, 318 complete proteins and 29 fragments were identified, such as dedicator of cytokinesis protein, intersectin, prostaglandin, phospholipase DDHD2 and superoxide dismutase. For the asymptomatic group, 326 complete proteins and 37 fragments were identified, including azurocidin, C-reactive protein, collagen alpha, cathepsin, heat shock and laminin. CONCLUSIONS: Quantitative differences in the expression of common proteins in cases of symptomatic and asymptomatic AP were found, which were mostly related to host immune response in both groups. Exclusive proteins in the symptomatic group were mainly related to the host response to the presence of viruses in endodontic infections, oxidative stress and proteolytic enzymes. The results provide a basis for a better understanding of cellular and molecular pathways involved in AP, establishing specific proteomic profiles for symptomatic and asymptomatic conditions.

Fluoride Mode of Action: Once There Was an Observant Dentist . . .

The discovery and implementation of fluoride in the prevention of dental caries is often praised as one of the most important achievements in health care. In the early 20th century, it took 30 y to identify fluoride as the cause of enamel mottling but also of reduced caries prevalence in a population drinking water containing fluoride. Similarly, from 1960 to 1990, it took major efforts to unravel the working mode of fluoride in such detail that a rational scheme of caries prevention could be formulated. This article describes the scientific struggle leading to a consensus on the topic. For a historic purpose, the field, the actors, and their main research achievements are described. Ultimately it was generally agreed that the effect of fluoride is primarily topical by fluorides in the oral fluids rather than systemic by incorporation of fluoride in the enamel mineral crystals. Fluoride concentrations, even <1 mg/L, enhance the deposition of calcium phosphates during remineralization of enamel (and dentin). Similarly, such low levels of fluoride are effective in reducing the dissolution of the calcified tissues. This understanding has led to the development of fluoride-containing caries-preventive products that had an undisputed beneficial effect on the levels of dental caries.

The use of fluoride for the prevention of dental erosion and erosive tooth wear in children and adolescents.

BACKGROUND: Erosive tooth wear (ETW) has gained increasing clinical relevance. It is estimated that worldwide 30-50% of deciduous and 20-45% of permanent teeth are affected. One of the most important nutritional factors causing ETW is the overconsumption of soft drinks, but also patient-related factors like reflux or eating disorders can lead to erosive lesions. Whether acids lead to erosive demineralisation depends on their degree of saturation with respect to tooth mineral at their actual pH. REVIEW: Fluoride compounds like sodium or amine fluoride seem to be of limited efficacy against erosion, the main reason for this is the missing biofilm in the erosive process as well as the lower pH of the acids compared to bacterial acids. This means that to achieve some kind of preventive effect it would be necessary to use products with higher fluoride concentration, which is not an appropriate option for small children, and/or to increase the frequency of application. In addition, the fluoride compound plays a role as promising effects were found when fluoride is combined with titanium or stannous ions. TiF4 can cause acid-resistant surface coatings and when Sn2+/F- formulations are applied, Sn is not only found on the surface but is also incorporated into enamel and dentine. Both effects make the tooth surface more resistant against acid demineralisation. Different fluoride-containing vehicles have been tested to prevent erosion/ETW, such as toothpastes, rinses, gels and varnishes. Toothpastes offer some degree of protection, especially Sn2+-containing formulations, but effects of the active ingredients are sometimes counteracted by the presence of abrasives. CONCLUSION: Detecting associated factors and influencing them is the main instrument in arresting erosive tooth wear. Additionally, patients at risk for dental erosion should always use an additional fluoride source preferably containing Sn2+.

Microstructure and selected mechanical properties of aged Ti-15Zr-based alloys for biomedical applications.

In this study, Ti-15Zr-xMo (5, 10, 15, and 20 wt%) alloys were submitted to solution and aging treatments and their effects evaluated in terms of phase composition and selected mechanical properties (Vickers microhardness and Young's modulus) for use as biomedical implants. The solution treatment was performed at 1123 K for 2 h, while aging treatments were carried out at 698 K for 4, 8, and 12 h, followed by water quenching. Phase composition and microstructure were dependent of the heat treatments, with Ti-15Zr-5Mo (α +â€¯ß type) and Ti-15Zr-10Mo (metastable ß type) alloys exhibiting intense α phase precipitation. The α-phase precipitates were related to α″ → α and ß → α phase decompositions. The Ti-15Zr-10Mo alloy exhibited an intermediary isothermal ω-phase precipitation after aging for 4 h. Vickers microhardness and Young's modulus values changed gradually with the amount of α phase. Aged Ti-15Zr-15Mo and Ti-15Zr-20Mo alloys presented better combinations of hardness and Young's modulus than CP-Ti and Ti-64 ELI for biomedical applications.

Prevention of erosive tooth wear: targeting nutritional and patient-related risks factors.

This article provides an overview of the nutritional and patient-related risk factors involved in the aetiology of erosive tooth wear (ETW) and the preventive strategies to counteract them. The first step is to diagnose clinical signs of ETW and to recognise causal factors. Low pH and high buffer capacity of foods/drinks are the major risk factors, while the calcium concentration is the main protective factor. Reduction of frequency of consumption and contact time of erosive foods/drinks with the teeth, use of straws appropriately positioned and consumption of dairy products are advisable. Oral hygiene has a role in the development of ETW, however, postponing toothbrushing is not clinically advisable. In cases of drug abuse, chronic alcoholism, GERD or bulimia, the patient must be referred to a doctor. Immediately after vomiting, patients might be advised to rinse the mouth. Saliva has an important protective role and patients with reduced salivary flow can benefit from the use of chewing gum. Recent studies have focused on improving the protective capacity of the acquired pellicle as well as on the role of protease inhibitors on dentine erosion. However, the degree of evidence for these preventive measures is low. Clinical trials are necessary before these measures can be recommended.

Review of Fluoride Intake and Appropriateness of Current Guidelines.

Since the classical epidemiological studies by Dean, it has been known that there should be an optimum level of exposure to fluoride that would be able to provide the maximum protection against caries, with minimum dental fluorosis. The "optimal" daily intake of fluoride for children (0.05-0.07 mg per kilogram bodyweight) that is still accepted worldwide was empirically determined. In the present review, we discuss the appropriateness of the current guidance for fluoride intake, in light of the windows of susceptibility to caries and fluorosis, the modern trends of fluoride intake from multiple sources, individual variations in fluoride metabolism, and recent epidemiological data. The main conclusion is that it is very difficult to think about a strict recommendation for an "optimal" range of fluoride intake at the individual level in light of existing knowledge of 1) the mechanisms of action of fluoride to control caries, 2) the mechanisms involved in dental fluorosis development, 3) the distinct factors that interfere in the metabolism of fluoride, and 4) the windows of susceptibility to both dental caries and fluorosis development. An "optimal" range of fluoride intake is, however, desirable at the population level to guide programs of community fluoridation, but further research is necessary to provide additional support for future decisions on guidance in this area. This list includes the effect of factors affecting fluoride metabolism, clinical trials on the effectiveness of low-fluoride dentifrices to prevent caries in the primary dentition, and validation of biomarkers of exposure to fluoride.

Preparation and characterization of alloys of the Ti-15Mo-Nb system for biomedical applications.

In the development of new metallic biomaterials, the Ti-15Mo alloy has great prominence because of its excellent corrosion resistance and good combination of mechanical properties. In this study, the element niobium was added to the Ti-15Mo alloy, forming the Ti-15Mo-Nb system for the purpose of improving their properties and promoting its application as a biomaterial. These alloys are very promising to use as biomedical implants, because they integrate a new class of titanium alloys without the presence of aluminum and vanadium, which may cause cytotoxic effects. The alloys were produced by arc-melting and characterized by density, X-ray diffraction, scanning electron microscopy, microhardness, elastic modulus, corrosion, and cytotoxicity assays. The developed alloys have ß phase predominance (with bcc crystalline structure). The addition of niobium decreases the microhardness and elastic modulus, with values around 80 GPa, which is well below that of the metallic alloys used commercially for this type of application. Very low passive current densities were found for all alloys studied showing that the passive film on these alloys is highly protective. In vitro cytotoxicity tests revealed that the introduction of niobium did not cause cytotoxic effects in the studied alloys. © 2017 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater, 106B: 639-648, 2018.

Fluoride concentration and amount of dentifrice influence enamel demineralization in situ.

OBJECTIVES: This study evaluated the effect of conventional (CD, 1100ppm F) and low-fluoride (LFD, 550ppm F) dentifrices, applied in different quantities, on enamel demineralization, and on fluoride (F) concentrations in the dental biofilm formed in situ. METHODS: Five combinations of dentifrices and quantities were tested: placebo (P-F-free) applied on all brush bristles; LFD applied by the transversal technique (0.3g-T1) or on all bristles (0.6g-T2); and CD applied in a pea-sized amount (0.15g-T3) or by the transversal technique (0.3g-T4), in order to produce comparable intensities (F concentration in the dentifrice×amount applied to the brush). Volunteers (n=13, 20-36 years old) wore palatal devices containing 4 bovine enamel blocks, and performed cariogenic challenges (30% sucrose solution) 6×/day, and brushing 3×/day, following a double-blind, cross-over and randomized protocol. On the 8th day, biofilm was collected 5 and 60min after brushing. The percentage of surface hardness loss (%SH), integrated loss of subsurface hardness (ΔKHN) and biofilm F concentrations (solid and fluid phases) were determined. Data were analyzed by repeated-measures ANOVA, Student-Newman-Keuls test, and Pearson's correlation coefficient (p<0.05). RESULTS: Significantly lower ΔKHN was observed for treatments with higher intensity (T2 and T4) in comparison with the lower intensity (T1 and T3). A strong correlation was observed between ΔKHN and F concentrations in total biofilm (r=-0.71) and biofilm fluid (r=-0.72) 5min after brushing. CONCLUSIONS: The treatment intensity has a significant influence on the development of caries lesions in situ. CLINICAL SIGNIFICANCE: The intensity of treatment (amount of dentifrice×concentration) during brushing seems to be a more relevant parameter of clinical efficacy than simply observing the F concentration of the product. The use of a small amount of CD significantly reduced the protective effects against enamel demineralization.

Response of carious enamel to TiF4 varnish treatment under diverse cariogenic activities in situ.

OBJECTIVE: To compare the effect of TiF4 and NaF varnishes on demineralized bovine enamel under different cariogenic activities in situ. METHODS: Twenty subjects participated of this in situ study with 3 crossover phases (14days each), in which they wore palatal appliances containing demineralized bovine enamel samples (8 samples/appliance for phase) treated with TiF4, NaF (all with 2.45% F) or placebo varnish. The samples were subjected to different cariogenic conditions (1. absence of biofilm accumulation and sucrose exposure; 2. presence of biofilm and absence of sucrose exposure; 3. presence of biofilm and 20% sucrose exposure 4×/day; 4. presence of biofilm and 20% sucrose exposure 8×/day). All were exposed to fluoride dentifrice (2×/day). The mineral content and lesion depth were evaluated using transverse microradiography (TMR) and the data were subjected to RM two-way ANOVA/Bonferroni tests (p<0.05). RESULTS: TiF4 varnish significantly increased the remineralization of artificial carious lesions compared to placebo, regardless of the cariogenic activity. On the other hand, the remineralizing effect of NaF varnish was dependent on the cariogenic activity. For NaF, remineralization happened only in conditions 1 and 3 compared to placebo varnish (p<0.0001). NaF was unable to prevent further demineralization under biofilm accumulation and sucrose exposure 8×/day (condition 4). In the absence of fluoride treatment, demineralization happened in all conditions, except in the condition 1. CONCLUSION: Therefore, 4% TiF4 varnish was the only treatment able to improve enamel remineralization regardless of the cariogenic activity, while NaF varnish failed in preventing further demineralization under high cariogenic activity in situ. CLINICAL SIGNIFICANCE: 4% TiF4 varnish showed better remineralizing effect compared to NaF varnish, which was seen regardless of the cariogenic activity. This is a promising finding to support the indication of TiF4 in the clinic.

A New Sugarcane Cystatin Strongly Binds to Dental Enamel and Reduces Erosion.

Cystatin B was recently identified as an acid-resistant protein in acquired enamel pellicle; it could therefore be included in oral products to protect against caries and erosion. However, human recombinant cystatin is very expensive, and alternatives to its use are necessary. Phytocystatins are reversible inhibitors of cysteine peptidases that are found naturally in plants. In plants, they have several biological and physiological functions, such as the regulation of endogenous processes, defense against pathogens, and response to abiotic stress. Previous studies performed by our research group have reported high inhibitory activity and potential agricultural and medical applications of several sugarcane cystatins, including CaneCPI-1, CaneCPI-2, CaneCPI-3, and CaneCPI-4. In the present study, we report the characterization of a novel sugarcane cystatin, named CaneCPI-5. This cystatin was efficiently expressed in Escherichia coli, and inhibitory assays demonstrated that it was a potent inhibitor of human cathepsins B, K, and L ( Ki = 6.87, 0.49, and 0.34 nM, respectively). The ability of CaneCPI-5 to bind to dental enamel was evaluated using atomic force microscopy. Its capacity to protect against initial enamel erosion was also tested in vitro via changes in surface hardness. CaneCPI-5 showed a very large force of interaction with enamel (e.g., compared with mucin and casein) and significantly reduced initial enamel erosion. These results suggest that the inclusion of CaneCPIs in dental products might confer protection against enamel erosion.

In situ Effect of Chewing Gum with and without CPP-ACP on Enamel Surface Hardness Subsequent to ex vivo Acid Challenge.

The erosion-protective effect of casein phosphopeptide-amorphous calcium phosphate (CPP-ACP) is controversial. This study aimed to investigate the ability of CPP-ACP chewing gum to prevent a single event of erosive demineralization in situ. Bovine enamel blocks (n = 120) were randomly assigned to 3 phases according to the baseline surface hardness: phase I (PI) - chewing gum with CPP-ACP, phase II (PII) - chewing gum without CPP-ACP, and control phase (PIII) - salivary effect without stimulation (no gum). Nineteen volunteers participated in this study during 3 crossover phases of 2 h. In PI and PII, the volunteers wore intraoral palatal appliances for 120 min and chewed a unit of the corresponding chewing gum for the final 30 min. In the control phase the volunteers wore the appliance for 2 h, without chewing gum. Immediately after intraoral use, the appliances were extraorally immersed in a cola drink for 5 min to promote erosive demineralization. The percentage of surface hardness loss was calculated. The data were analyzed by ANOVA models and Tukey's test. Lower enamel hardness loss was found after the use of chewing gum with CPP-ACP (PI: 32.7%) and without CPP-ACP (PII: 33.5%) compared to the salivary effect without stimulation (PIII: 39.8%) (p < 0.05). There was no difference between PI and PII (p > 0.05). The results suggest that the use of chewing gum immediately before an erosive demineralization can diminish enamel hardness loss. However, the presence of CPP-ACP in the chewing gum cannot enhance this protective effect.

Effect of xylitol varnishes on remineralization of artificial enamel caries lesions in situ.

OBJECTIVES: Analyze the effect of varnishes containing xylitol compared to commercial fluoridated varnishes on the remineralization of artificial enamel caries lesions in situ. METHODS: Twenty subjects took part in this crossover, double-blind study performed in four phases of 5days each. Each subject worn palatal appliances containing four predemineralized bovine enamel specimens. Artificial caries lesions were produced by immersion in 30ml of lactic acid buffer containing 3mM CaCl2·2H2O, 3mM KH2PO4, 6µM tetraetil metil diphosphanate (pH 5.0) for 6days. The specimens in each subject were treated once with the following varnishes: 20% xylitol (experimental); Duofluorid™ (6% NaF, 6% CaF2), Duraphat™ (5% NaF, positive control) and placebo (no-F/xylitol, negative control). The varnishes were applied in a thin layer and removed after 6h. Fifteen subjects were able to finish all phases. The enamel alterations were quantified by surface hardness and transversal microradiography. The percentage of surface hardness recovery (%SHR), the integrated mineral loss and lesion depth were statistically analyzed by Friedmann and Dunn's tests test (p<0.05). RESULTS: Enamel surface remineralization was significantly increased by Duraphat™, Duofluorid™ and 20% xylitol formulations. Significant subsurface mineral remineralization could also be seen for the experimental and commercial varnishes, except for Duraphat™, when the parameter "lesion depth" was considered. CONCLUSIONS: 20% xylitol varnish seem to be a promising alternative to increase surface and subsurface remineralization of artificial caries lesions in situ. CLINICAL SIGNIFICANCE: effective vehicles are desirable for caries control. Xylitol varnishes seem to be promising alternatives to increase enamel remineralization in situ, which should be confirmed by clinical studies.

The role of matrix metalloproteinases and cysteine-cathepsins on the progression of dentine erosion.

OBJECTIVE: To evaluate in vitro the effect of the inhibition of endogenous dentinal enzymes (matrix metalloproteinases-MMPs and cysteine cathepsins-CCs) on dentine erosion. DESIGN: Dentine blocks (4mm×4mm×2mm) from sound human teeth were randomly divided into 7 groups (n=17) according to the treatment: MMP- and CC-inhibitor chlorhexidine digluconate (CHX, 10mM); MMP-inhibitor galardin (G, 0.2mM); specific cathepsin B inhibitor (CCB, 0.2mM); non-specific CC inhibitor (CCE-64, 0.5µM); fluoride (F, 1.23% NaF); placebo (P) and untreated (UT). Inhibitors were applied as gels once for 1min. Specimens were submitted to 5 days of pH cycling including the erosive challenge (Coke, pH 2.64, 90s/day) and remineralisation (artificial saliva). Demineralised organic surface loss was determined profilometrically. Demineralised organic matrix (DOM) was removed with collagenase and the profile was re-evaluated in the absence of collagen fibrils. The differences in profilometric results and DOM thickness among the groups were analysed with ANOVA and Tukey's test (p<0.05). RESULTS: Loss of demineralised tissue (µm, mean±SD) was: CHX 8.4±1.7 b, G 8.6±1.9 b, CCB 9.6±1.4 a, CCE-64 9.9±1.3 a, F 9.9±1.7 a, P 10.9±2.2 a, UT 11.0±1.5 a. Loss of mineralised tissue was: CHX 15.4±2.2 b, G 16.0±1.8 b, CCB 17.6±2.4 a, CCE-64 17.6±2.0 a, F 17.3±2.8 a, P 19.1±2.1 a, UT 18.9±2.4 a. MMP-inhibitors significantly reduced organic matrix and mineral loss in comparison to all the other groups (p<0.05). No statistically significant differences were found in the thickness of the remaining DOM (p=0.845). CONCLUSION: Dentine endogenous MMPs seem to be the main enzymes responsible for DOM loss and erosion.

Low-Level Fluoride Exposure Increases Insulin Sensitivity in Experimental Diabetes.

The effect of chronic fluoride (F) exposure from the drinking water on parameters related to glucose homeostasis was investigated. Wistar rats were randomly distributed into 2 groups (diabetic [D] and nondiabetic [ND]; n = 54 each). In D, diabetes was induced with streptozotocin. Each group was further divided into 3 subgroups (0, 10, or 50 mgF/L in drinking water). After 22 days of treatment, plasma and liver samples were collected. No alterations in glycemia, insulinemia, K(ITT), and HOMA2-IR (homeostasis model assessment 2 of insulin resistance) were seen for ND. F-exposure of D rats led to significantly lower insulinemia, without alterations in glycemia (increased %S). Proteomic analysis detected 19, 39, and 16 proteins differentially expressed for the comparisons D0 vs. D10, D0 vs. D50, and D10 vs. D50, respectively. Gene Ontology with the most significant terms in the comparisons D0 vs. D10, D0 vs. D50, and D50 vs. D10 were organic acid metabolic process and carboxylic acid metabolic process, organic acid metabolic process, and cellular ketone metabolic process. Analysis of subnetworks revealed that proteins with fold changes interacted with GLUT4 in comparison D0 vs. D10. Among these proteins, ERj3p was present in D10. Upregulation of this protein in the presence of F might help to explain the higher %S found in these animals. These data suggest that fluoride might enhance glucose homeostasis in diabetes and identify specific biological mechanisms that merit future studies.

Dentifrice pH but not consistency may affect fluoride uptake in plaque.

OBJECTIVE: Test the ability of acidic fluoridated solutions to enhance fluoride (F) bound on bacteria (1) and the effect of dentifrice consistency on plaque fluid F uptake (2). METHODS: (1) Streptococcus mutans isolates were grown in BHI medium (37°C/18h). Bacteria were washed either with EDTA or CaCl2 both at 1mM to remove or add calcium, respectively. Pellets were incubated with 12 mM NaF at pH 4.5 or 7 for 1 min and F was quantified in the lysates and supernatants with the electrode, after HMDS-facilitated diffusion. (2) A randomized, double-blind, crossover clinical trial was performed in three phases with nineteen adults (20-35 years) that used one of the dentifrices: commercial toothpaste (1100 ppm F, pH7.0 and conventional viscosity (Sorriso Fresh(®))); experimental liquid dentifrice (ELD) (1100 ppm F, pH7.0 and low viscosity [1.1% carboxymethylcellulose (CMC)]) and ELD (1100 ppm F and high viscosity pH7.0 (2.2% CMC)). F concentration in plaque fluid was analyzed using an inverted F electrode. RESULTS: (1) Significantly higher F amounts were detected in the lysates of bacteria incubated with NaF solution at pH4.5 compared to the supernatant, which was not seen at pH7.0, being this effect calcium-dependent. (2) Significantly higher F concentrations in plaque fluid were found 1h after toothbrushing compared to 12h, but no significant differences were seen among the toothpastes. CONCLUSIONS: F at low pH binds more efficiently to S. mutans than at neutral pH and dentifrice viscosity does not interfere in plaque fluid fluoride incorporation. CLINICAL SIGNIFICANCE: pH of the dentifrice but not consistency may be important to F uptake in plaque.

Mercury fractionation in dourada (Brachyplatystoma rousseauxii) of the Madeira River in Brazil using metalloproteomic strategies.

This paper presents the results of mercury fractionation in muscle samples of dourada (Brachyplatystoma rousseauxii) from the JIRAU Hydroelectric Power Plant in the Madeira River Basin in the Amazon region of Brazil. The proteome of the dourada muscle was separated by two-dimensional polyacrylamide gel electrophoresis (2D PAGE). The mercury present in the protein spots was determined by graphite furnace atomic absorption spectrometry (GFAAS) after acid mineralisation in an ultrasound bath. The protein spots in which the presence of mercury was detected were characterised by electrospray ionisation tandem mass spectrometry (ESI-MS/MS) after tryptic digestion. The GFAAS determinations indicated that 65% of the mercury was linked to the protein fraction with a molar mass (Mm) of less than 90 kDa. The mercury concentrations in the seven spots in which this protein fraction was present were in the range of 11.40-35.10 µg kg(-1). Based on the mercury concentrations, it was possible to estimate that the protein spots contained approximately 1-3 mercury atoms per protein molecule. The ESI-MS/MS analysis allowed characterisation of the seven protein spots as the following proteins: protein NLRC5 (molar mass=18.10, pI=6.30); 39S ribosomal protein L36 mitochondrial (molar mass=15.40, pI=8.23); N-alpha-acetyltransferase 20 (Mm=15.95, pI=8.80); Mth938 domain-containing protein (Mm=15.01, pI=9.60); ubiquitin-40S ribosomal protein S27a (Mm=9.80, pI=7.60); parvalbumin alpha (Mm=12.40, pI=3.80) and parvalbumin beta (Mm=13.10, pI=3.45).

Effect of xylitol varnishes on remineralization of artificial enamel caries lesions in vitro.

OBJECTIVES: Analyse the effect of varnishes containing xylitol alone or combined with fluoride on the remineralization of artificial enamel caries lesions in vitro. METHODS: Bovine enamel specimens were randomly allocated to 7 groups (n=15/group). Artificial caries lesions were produced by immersion in 30 mL of lactic acid buffer containing 3mM CaCl2·2H2O, 3mM KH2PO4, 6 µM tetraetil metil diphosphanate (pH 5.0) for 6 days. The enamel blocks were treated with the following varnishes: 10% xylitol; 20% xylitol; 10% xylitol plus F (5% NaF); 20% xylitol plus F (5% NaF); Duofluorid™ (6% NaF, 2.71% F+6% CaF2), Duraphat™ (5% NaF, positive control) and placebo (no-F/xylitol, negative control). The varnishes were applied in a thin layer and removed after 6h. The blocks were subjected to pH-cycles (demineralization-2h/remineralization-22 h during 8 days) and enamel alterations were quantified by surface hardness and transversal microradiography. The percentage of surface hardness recovery (%SHR), the integrated mineral loss and lesion depth were statistically analysed by ANOVA/Tukey's test or Kruskal-Wallis/Dunn's test (p<0.05). RESULTS: Enamel surface remineralization was significantly increased by Duraphat™, 10% xylitol plus F and 20% xylitol plus F formulations, while significant subsurface mineral remineralization could be seen only for enamel treated with Duraphat™, Duofluorid™ and 20% xylitol formulations. CONCLUSIONS: 20% xylitol varnishes seem to be promising alternatives to increase remineralization of artificial caries lesions. CLINICAL SIGNIFICANCE: effective vehicles are desirable for caries control. Xylitol varnishes seem to be promising alternatives to increase enamel remineralization in vitro, which should be confirmed by in situ and clinical studies.

Diverse outcomes of Photodynamic Antimicrobial Chemotherapy on five Enterococcus faecalis strains.

OBJECTIVES: In the present study, the effectiveness of Photodynamic Antimicrobial Chemotherapy (PACT) was evaluated on planktonic cells and biofilms of five Enterococcus faecalis clinical isolates. METHODS: Planktonic cells and biofilms of E. faecalis E2, E3, ER3/2s, OS16 and AA-OR34 were grown in SDMY medium plus 0.4% glucose. Approximately 5.0×10(7)CFU planktonic cells and 24h biofilms were subjected to PACT using the combination of Light Emitting Diodes (LEDs, Biotable(®)) and Photogem(®). The metabolic activity of bacterial cells was evaluated by a resazurin assay. Biomass values of the biofilms were determined by a crystal violet assay. RESULTS: Compared to the water-treated control group, gradual increases of light energy led to greater reduction of metabolic activity of planktonic cells and biofilms of E. faecalis when the combination of LEDs and Photogem(®) was applied. Photogem(®) alone significantly reduced the metabolic activity of planktonic cells, whereas LEDs or Photogem(®) alone did not result in biofilm viability changes. PACT yielded similar antimicrobial outcomes on planktonic cells of all tested E. faecalis strains, whereas biofilms of E. faecalis E3, ER3/2s and OS16 were more resistant to PACT than biofilms of E. faecalis E2 and AA-OR34. CONCLUSIONS: The efficacy of PACT on E. faecalis biofilms was strain dependent. PACT demonstrated its potential as an adjuvant antimicrobial treatment by killing of E. faecalis planktonic and biofilm cells.