Thelma Lopes da Silva

Proteomic analysis of kidney in rats chronically exposed to fluoride.

Two-dimensional gel electrophoresis (2-DE) was used to better understand alterations in renal metabolism induced by fluoride (F). Three groups of weanling male Wistar rats were treated with drinking water containing 0 (control), 5, or 50 ppm F for 60 days (n=6/group). Kidneys were collected for proteomic and histological (HE) analysis. After protein isolation, renal proteome profiles were examined using 2-DE and Colloidal Coomassie Blue staining. Protein spots with a 2-fold significant difference as detected by quantitative intensity analysis (Image Master Platinum software) and t-test (p<0.05) were excised and analyzed by MALDI-TOF MS (matrix assisted laser desorption ionization-time-of-flight mass spectrometry). The histological analysis revealed no damage in kidneys induced by F, except for a vascular congestion in the 50 ppm F group. Between control vs 50 ppm F, and control vs 5 ppm F groups, 12 and 6 differentially expressed proteins were detected, respectively. Six proteins, mainly related with metabolism, detoxification and housekeeping, were successfully identified. At the high F group, pyruvate carboxylase, a protein involved in the formation of oxaloacetate was found to be downregulated, while enoyl coenzyme A hydratase, involved in fatty acids oxidation, was found to be upregulated. Thus, proteomic analysis can provide new insights into the alterations in renal metabolism after F exposure, even in low doses.

Demineralization effect of EDTA, EGTA, CDTA and citric acid on root dentin: a comparative study

The purpose of this study was to biochemically compare the decalcifying effects of 1% EDTA (pH 7.4), 1% EGTA (pH 7.4), 1% CDTA (pH 7.4), 1% citric acid solutions (pH 1.0 and 7.4) and saline solution (control) on root dentin. Forty-eight single-rooted teeth were used in this study. The canals were instrumented by the step-back technique and the roots were randomly divided into six equal experimental groups (n = 8) according to the irrigating agent tested. A total of 30 microL of each solution was pipetted into the root canal and allowed to set undisturbed for 5 minutes. After this time, 15 microL of the solutions were removed from each canal using a Hamilton syringe and placed in a container with 5 mL of deionised water. The microg/mL concentration of calcium ion (Ca2+) extracted from the root canal samples was determined using inductively coupled plasma-atomic emission spectrometry (ICP-AES). Data were analysed by means of the Kruskal-Wallis and Moods median tests. Citric acid solution at pH 1.0 removed more calcium than at pH 7.4 and than the other chelating solutions tested (p < 0.05). No differences were observed between EDTA and EGTA. Both EDTA and EGTA removed significantly more calcium than CDTA and citric acid at pH 7.4 (p < 0.05). There were no differences between citric acid at pH 7.4 and saline solution, which had the least efficacy for Ca2+ extraction (p > 0.05). These results indicate that citric acid at pH 1.0 is a good alternative as an irrigating solution to remove the smear layer and facilitate the biomechanical procedures.

Fluorine content of several brands of chocolate bars and chocolate cookies found in Brazil

Chocolate bars and chocolate cookies are foodstuffs highly appreciated by children. The possibility of having fluorine (F) among their components, associated with an excessive consumption, may make them decisive contributors to the total daily F intake. Thus, they could participate in the establishment of dental fluorosis. The aim of this study was to analyze the fluorine concentration [F] of the chocolates bars (CB) Baton, Confeti, Garoto Ball, Kinder Ovo, M&M s, Milkybar, Nescau, Nescau Ball, Surpresa, Surpresa Bichos, Tortuguita; and of the chocolate cookies (CC) Danyt s, Hipop , Nescau, Passatempo, Pokémon, S tio do Pica-Pau Amarelo and Trakinas. Samples were purchased in Bauru, São Paulo, Brazil. Three grams of each product were previously ashed at 525 C (CB and cookies fillings) and at 550 C (cookies dough), during 4 hours. Fluorine was separated from the ash by hexamethyldisiloxane (HMDS)-facilitated diffusion. Fluorine analysis was carried out with the specific electrode. Mean [F]s SD and amplitude (unit mg/g) were: CB = 0.30 0.45 (0.07 - 1.60, n = 12) and CC = 1.08 2.64 (0.04 - 7.10, n = 7). It was concluded that some of the analyzed foods may be important contributors to the total daily F intake. As for the product that had the highest [F] (Danyt s), when only 3 units are consumed just once a day, they may supply up to 40% of the maximum recommended daily F intake (0.07 mg/kg body weight) for a 2-year-old child (12 kg). The [F] in these products should be informed on their labels.

Differential effects of flavonoids on bovine kidney low molecular mass protein tyrosine phosphatase

Among the structurally related flavonoids tested on the bovine kidney low molecular weight protein tyrosine phosphatase (LMrPTP) activity, quercetin activated by about 2.6-fold the p-nitrophenyl-phosphate (p-NPP)-directed reaction, in contrast to morin that acted as a competitive inhibitor, with Ki values of 87, 73 and 50 μM for p-NPP, FMN, and tyrosine-phosphate, respectively. Other related flavonoids, such as rutin, kaempferol, catechin, narigin, phloretin and taxifolin did not significantly affect the LMrPTP activity. The positions of the hydroxyl groups in the structures of the flavonoids were important for their distinct effects on LMrPTP activity. The hydroxyl groups at C3′ and C4′ and the presence of a double bond at C2 and C3 were essential for the activating effect of quercetin. The absence of the 3′-OH (kaempferol), absence of the double bond (taxifolin) and the presence of the sugar rutinose at the 3-OH (rutin) suppressed the effect of quercetin. The C2′- and C4′-hydroxyl groups, the presence of the double bond, and a C4-ketone group were important requirements for the inhibitory effects of morin.

Aquaporin 5 Interacts with Fluoride and Possibly Protects against Caries

Aquaporins (AQP) are water channel proteins and the genes coding for AQP2, AQP5, and AQP6 are clustered in 12q13. Since AQP5 is expressed in serous acinar cells of salivary glands, we investigated its involvement in caries. DNA samples from 1,383 individuals from six groups were studied. Genotypes of eight single nucleotide polymorphisms covering the aquaporin locus were tested for association with caries experience. Interaction with genes involved in enamel formation was tested. The association between enamel microhardness at baseline, after creation of artificial caries lesion, and after exposure to fluoride and the genetic markers in AQP5 was tested. Finally, AQP5 expression in human whole saliva, after exposure to fluoride in a mammary gland cell line, which is known to express AQP5, and in Wistar rats was also verified. Nominal associations were found between caries experience and markers in the AQP5 locus. Since these associations suggested that AQP5 may be inhibited by levels of fluoride in the drinking water that cause fluorosis, we showed that fluoride levels above optimal levels change AQP5 expression in humans, cell lines, and rats. We have shown that AQP5 is involved in the pathogenesis of caries and likely interacts with fluoride.

Proteomic analysis of urine in rats chronically exposed to fluoride

Urine is an ideal source of materials to search for potential disease‐related biomarkers as it is produced by the affected tissues and can be easily obtained by noninvasive methods. 2‐DE‐based proteomic approach was used to better understand the molecular mechanisms of injury induced by fluoride (F−) and define potential biomarkers of dental fluorosis. Three groups of weanling male Wistar rats were treated with drinking water containing 0 (control), 5, or 50 ppm F− for 60 days (n = 15/group). During the experimental period, the animals were kept individually in metabolic cages, to analyze the water and food consumption, as well as fecal and urinary F− excretion. Urinary proteome profiles were examined using 2‐DE and Colloidal Coomassie Brilliant Blue staining. A dose‐response regarding F− intake and excretion was detected. Quantitative intensity analysis revealed 8, 11, and 8 significantly altered proteins between control vs. 5 ppm F−, control vs. 50 ppm F− and 5 ppm F− vs. 50 ppm F− groups, respectively. Two proteins regulated by androgens (androgen‐regulated 20‐KDa protein and α‐2μ‐globulin) and one related to detoxification (aflatoxin‐B1‐aldehyde‐reductase) were identified by MALDI‐TOF‐TOF MS/MS. Thus, proteomic analysis can help to better understand the mechanisms underlying F− toxicity, even in low doses.

Xylitol concentrations in artificial saliva after application of different xylitol dental varnishes

Objective The present study analyzed xylitol concentrations in artificial saliva over time after application of varnishes containing 10% and 20% xylitol. Material and Methods Fifteen bovine enamel specimens (8x4 mm) were randomly allocated to 3 groups (n=5/group), according to the type of varnish used: 10% xylitol, 20% xylitol and no xylitol (control). After varnish application (4 mg), specimens were immersed in vials containing 500 µL of artificial saliva. Saliva samples were collected in different times (1, 8, 12, 16, 24, 48 and 72 h) and xylitol concentrations were analyzed. Data were assessed by two-way repeated-measures ANOVA (p<0.05). Results Colorimetric analysis was not able to detect xylitol in saliva samples of the control group. Salivary xylitol concentrations were significantly higher up to 8 h after application of the 20% xylitol varnish. Thereafter, the 10% xylitol varnish released larger amounts of that polyol in artificial saliva. Conclusions Despite the results in short-term, sustained xylitol releases could be obtained when the 10% xylitol varnish was used. These varnishes seem to be viable alternatives to increase salivary xylitol levels, and therefore, should be clinically tested to confirm their effectiveness.

Evaluation of UCLA Implant-Abutment Sealing

PURPOSE To evaluate the effect of the presence of a prefabricated cobalt-chromium (CoCr) margin in a universal castable long abutment (UCLA) on the sealing capability and fit of the implant-abutment interface. MATERIALS AND METHODS One-hundred twenty external hexagon implants (SIN) were divided into two groups (n=60 each) to receive UCLA abutments from six manufacturers (n=10 each) either with or without a CoCr margin (n=60 each). Abutments were cast and 12 groups were formed: M (Microplant), I (Impladen), S (SIN), Sv (Signo Vinces), T (TitaniumFix), and B (Bionnovation). Sealing was determined by placing 0.7 µL of 0.1% toluidine blue in the implant wells before abutment torquing. Implant-abutment samples were placed into 2.0-mL vials containing 0.7 mL of distilled water to maintain the implant-abutment interface, and aliquots of 100 µL of water were retrieved at 1, 3, 6, 24, 48, 72, 96, and 144 hour incubation times for measurement of absorbance in a spectrophotometer, and returned for repeated measurements. Two-way ANOVA (P<.05) and Tukeys test were used. Scanning electron microscopy (SEM) was used for observation of the implant-abutment fit. RESULTS Groups M, Sv, and T without the CoCr margin resulted in complete release of toluidine blue at 1 hour, whereas I, S, and B did so at 3, 24, and 96 hours, respectively. Complete leakage in abutments with the prefabricated margin occurred at 6 hours for S; 24 hours for Sv, T, and B; and 72 hours for M and I. Implant-abutment gaps were observed in all groups. A poorer fit was depicted for groups M and T without the CoCr margin. CONCLUSION Complete leakage was observed for all UCLA abutments regardless of the presence of the CoCr margin. Implant-abutment gaps were observed in all groups.

Optimizing the procedure for mercury recovery from dental amalgam

Mercury, as any other heavy metal, may cause environmental damages due to its accumulation and biotransformation. Dental offices, whether private or institutional, use dental amalgam as a restorative material on a daily basis. Dental amalgam is composed of mercury (50%), silver (30%) and other metals. Approximately 30% of the amalgam prepared in dental offices (0.6 g per capsule) are wasted and inadequately discarded without any treatment. Methods for mercury recovery have been proposed previously, using high temperatures through exposure to direct flame (650 degrees C), long processing time, and hazardous reagents as potassium cyanide. The purpose of this study was to develop a method to replace the direct flame by an electrical mantle in the process of mercury recovery. Results showed an average mercury recovery of 90% from 2 kg of amalgam after 30 minutes of processing time, thus optimizing the procedure. The proposed modifications allowed a significant reduction in processing time and a mercury recovery with high purity. The modified process also provided minimization of operator exposure to physical, chemical and ergonomic hazards, representing a technological advance compared to the risks inherent to the original method. It also provided environmental health and economy of energy resources by replacing a finite energy source (fossil and organic) by a more environmentally appropriate electric source, resulting in significant improvement of the procedure for mercury recovery from dental amalgam.

Sealing Capability and SEM Observation of the Implant-Abutment Interface

To evaluate the sealing capability of external hexagon implant systems and assess the marginal fit, two groups (n = 10 each) were employed: SIN (Sistema de Implantes Nacional, Brazil) and Osseotite, (Biomet 3i, USA). Sealing capability was determined by placing 0.7 μL of 1% acid-red solution in the implant wells before the torque of their respective abutments. Specimens were then placed into 2.5 mL vials filled with 1.3 mL of distilled water with the implant-abutment interface submerged. Three samples of 100 μL water were collected at previously determinate times. The absorbance was measured with a spectrophotometer, and the data were analyzed by Two-way ANOVA (P < .05) and Tukeys test. Marginal fit was determined using SEM. Leakage was observed for both groups at all times and was significantly higher at 144 hrs. SEM analysis depicted gaps in the implant-abutment interface of both groups. Gaps in the implant-abutment interface were observed along with leakage increased at the 144 hrs evaluation period.