Effects of long-term fluoride exposure are associated with oxidative biochemistry impairment and global proteomic modulation, but not genotoxicity, in parotid glands of mice.

BACKGROUND: Fluoride has become widely used in dentistry because of its effectiveness in caries control. However, evidence indicates that excessive intake interferes with the metabolic processes of different tissues. Thus, this study aimed to investigate the effects of long-term exposure to F on the parotid salivary gland of mice, from the analysis of oxidative, proteomic and genotoxic parameters. MATERIALS AND METHODS: The animals received deionized water containing 0, 10 or 50 mg/L of F, as sodium fluoride, for 60 days. After, parotid glands were collected for analysis of oxidative biochemistry, global proteomic profile, genotoxicity assessment and histopathological analyses. RESULTS: The results revealed that exposure to fluoride interfered in the biochemical homeostasis of the parotid gland, with increased levels of thiobarbituric acid reactive species and reduced glutathione in the exposed groups; as well as promoted alteration of the glandular proteomic profile in these groups, especially in structural proteins and proteins related to oxidative stress. However, genotoxic assessment demonstrated that exposure to fluoride did not interfere with DNA integrity in these concentrations and durations of exposure. Also, it was not observed histopathological alterations in parotid gland. CONCLUSIONS: Thus, our results suggest that long-term exposure to fluoride promoted modulation of the proteomic and biochemical profile in the parotid glands, without inducing damage to the genetic component. These findings reinforce the importance of rationalizing the use of fluorides to maximize their preventative effects while minimizing the environmental risks.

Effects of Fluoride on Submandibular Glands of Mice: Changes in Oxidative Biochemistry, Proteomic Profile, and Genotoxicity.

Although fluoride (F) is well-known to prevent dental caries, changes in cell processes in different tissues have been associated with its excessive exposure. Thus, this study aimed to evaluate the effects of F exposure on biochemical, proteomic, and genotoxic parameters of submandibular glands. Twenty one old rats (n = 30) were allocated into three groups: 60 days administration of drinking water containing 10 mgF/L, 50 mgF/L, or only deionized water (control). The submandibular glands were collected for oxidative biochemistry, protein expression profile, and genotoxic potential analyses. The results showed that both F concentrations increased the levels of thiobarbituric acid-reactive substances (TBARS) and reduced glutathione (GSH) and changed the proteomic profile, mainly regarding the cytoskeleton and cellular activity. Only the exposure to 50 mgF/L induced significant changes in DNA integrity. These findings reinforce the importance of continuous monitoring of F concentration in drinking water and the need for strategies to minimize F intake from other sources to obtain maximum preventive/therapeutic effects and avoid potential adverse effects.

Effects of acute fluoride exposure on the jejunum and ileum of rats: Insights from proteomic and enteric innervation analysis.

Fluoride (F) is largely employed in dentistry, in therapeutic doses, to control caries. However, excessive intake may lead to adverse effects in the body. Since F is absorbed mostly from the gastrointestinal tract (GIT), gastrointestinal symptoms are the first signs following acute F exposure. Nevertheless, little is known about the mechanistic events that lead to these symptoms. Therefore, the present study evaluated changes in the proteomic profile as well as morphological changes in the jejunum and ileum of rats upon acute exposure to F. Male rats received, by gastric gavage, a single dose of F containing 0 (control) or 25 mg/Kg for 30 days. Upon exposure to F, there was a decrease in the thickness of the tunic muscularis for both segments and a decrease in the thickness of the wall only for the ileum. In addition, a decrease in the density of HuC/D-IR neurons and nNOS-IR neurons was found for the jejunum, but for the ileum only nNOS-IR neurons were decreased upon F exposure. Moreover, SP-IR varicosities were increased in both segments, while VIP-IR varicosities were increased in the jejunum and decreased in the ileum. As for the proteomic analysis, the proteins with altered expression were mostly negatively regulated and associated mainly with protein synthesis and energy metabolism. Proteomics also revealed alterations in proteins involved in oxidative/antioxidant defense, apoptosis and as well as in cytoskeletal proteins. Our results, when analyzed together, suggest that the gastrointestinal symptoms found in cases of acute F exposure might be related to the morphological alterations in the gut (decrease in the thickness of the tunica muscularis) that, at the molecular level, can be explained by alterations in the gut vipergic innervation and in proteins that regulate the cytoskeleton.

Study of proteins with mercury in fish from the Amazon region.

The high concentrations of mercury found in Amazon have been intensively studied by the scientific community in the last decades. These mercurial species bind preferentially to proteins. Therefore, this work proposal sought to obtain the fractionation, identification and study of mercury - bound proteins present in samples of muscular and hepatic tissue from fish collected in the reservoir of the Jirau Hydroelectric Power Plant - on the Madeira River. Two-dimensional electrophoresis (2D-PAGE) for protein fractionation, graphite furnace atomic absorption spectrometry (GFAAS) for the quantification of mercury and Mass Spectrometry (ESI-MS/MS) were used for the identification of proteins. Concluding the work with analysis of graphics from the Blast2go program. Two mercury - bound proteins were identified as triosephosphate isomerase A and Protein FAM45A. The data generated by the bioinformatics programs confirm the tendency of these proteins to be linked to mercury and elucity the possibles existing physiological and cellular interactions.

The influence of fillers and protease inhibitors in experimental resins in the protein profile of the acquired pellicle formed in situ on enamel-resin specimens.

OBJECTIVE: This study evaluated the influence of the addition of fillers and/or protease inhibitors [(epigallocatechin gallate - EGCG) or (chlorhexidine - CHX)] in experimental resins in the protein profile of the acquired pellicle (AP) formed in situ on enamel-resin specimens. DESIGN: 324 samples of bovine enamel were prepared (6 × 6 × 2 mm). The center of each sample was added with one of the following experimental resins (Bis-GMA+TEGDMA): no filler, no inhibitor (NF-NI); filler no inhibitor (F-NI); no filler plus CHX (NF-CHX); filler plus CHX (F-CHX); no filler plus EGCG (NF-EGCG); filler plus EGCG (F-EGCG). Nine subjects used a removable jaw appliance (BISPM - Bauru in situ pellicle model) with 2 slabs from each group. The AP was formed for 120 min, in 9 days and collected with electrode filter paper soaked in 3% citric acid. The pellicles collected were processed for analysis by LC-ESI-MS/MS. RESULTS: A total of 140 proteins were found in the AP collected from all the substrates. Among them, 16 proteins were found in common in all the groups: 2 isoforms of Basic salivary proline-rich protein, Cystatin-S, Cystatin-AS, Cystatin-SN, Histatin-1, Ig alpha-1 chain C region, Lysozyme C, Mucin-7, Proline-rich protein 4, Protein S100-A9, Salivary acidic proline-rich phosphoprotein ½ and Statherin. Proteins with other functions, such as metabolism and transport, were also identified. CONCLUSION: The composition of the experimental resins influenced the protein profile of the AP. This opens a new avenue for the development of new materials able to guide for AP engineering, thus conferring protection to the adjacent teeth.

Chronic treatment with fluoride affects the jejunum: insights from proteomics and enteric innervation analysis.

Gastrointestinal symptoms are the first signs of fluoride (F) toxicity. In the present study, the jejunum of rats chronically exposed to F was evaluated by proteomics, as well as by morphological analysis. Wistar rats received water containing 0, 10 or 50 mgF/L during 30 days. HuC/D, neuronal Nitric Oxide (nNOS), Vasoactive Intestinal Peptide (VIP), Calcitonin Gene Related Peptide (CGRP), and Substance P (SP) were detected in the myenteric plexus of the jejunum by immunofluorescence. The density of nNOS-IR neurons was significantly decreased (compared to both control and 10 mgF/L groups), while the VIP-IR varicosities were significantly increased (compared to control) in the group treated with the highest F concentration. Significant morphological changes were seen observed in the density of HUC/D-IR neurons and in the area of SP-IR varicosities for F-treated groups compared to control. Changes in the abundance of various proteins correlated with relevant biological processes, such as protein synthesis, glucose homeostasis and energy metabolism were revealed by proteomics.

Exposure to acids changes the proteomic of acquired dentine pellicle.

OBJECTIVES: For the first time, this study characterized the proteome of the acquired pellicle formed on human dentine. The changes in this proteome after exposure to lactic or citric acid were also evaluated. METHODS: Volunteers (n=9) wore a mandibular device containing 6 specimens of human root dentine. After the device remained in the volunteers' oral cavities for 10min or 2h to allow the formation of the acquired pellicle in situ, the specimens were immersed in citric acid (1%, pH 2.5) or lactic acid (0.1M, pH 4.8) or deionized water for 20s. In sequence, the pellicle was collected with an electrode filter paper soaked in 3% citric acid. This procedure was repeated for two additional days following a crossover protocol. After harvest, proteins were subjected to reverse phase liquid chromatography coupled to mass spectrometry (nLC-ESI-MS/MS). MS/MS data were processed and submitted to Proteome Discoverer software. Searches were done using SWISS-PROT and TrEMBL databases for human proteins. RESULTS: In total, 223 distinct proteins were identified in the dentine acquired pellicle in each of the different conditions. Exposure to citric acid dramatically reduced the number of identified proteins. This did not occur for lactic acid. Acid-resistant proteins, such as mucins, were identified after pellicle was exposed to lactic or citric acid. CONCLUSIONS: These proteins could be related to protective effect of tooth homeostasis. Moreover, in the future, they could be candidates to the development of a supplemental therapy for the prevention and treatment of dental caries and dental erosion. CLINICAL SIGNIFICANCE: This study indicates some acid-resistant proteins that could be used in dental products to prevent dental caries and erosion.

Effects of fluoride in bone repair: an evaluation of RANKL, OPG and TRAP expression.

The objective of this study was to evaluate comparatively the effect of fluoride in the expression of the receptor activator of nuclear factor kappa B ligand (RANKL), osteoprotegerin (OPG) and tartrate-resistant acid phosphatase (TRAP) in alveolar bone repair in rats. We used 3 groups of male Wistar rats (n = 5/group), which received drinking water containing different doses of F (NaF): 0, 5 and 50 ppm, for 60 days before the incisors extraction. The upper incisors were extracted and the animals were killed 7, 14, 21 and 30 days after extraction. The hemi-maxillae were collected for microscopic examination (histomorphometric and immunostaining for RANKL, OPG and TRAP). Histomorphometric analysis confirmed an increase in the volume density of neoformed bone between 7 and 30 days for groups control, 5 and 50 ppm of F, with a concomitant decrease in the volume density of connective tissue and blood clot. Higher blood clot for groups 5 and 50 ppm of F at 30 days was observed. The RANKL and OPG expressions were not changed by chronic exposure to fluoride in the drinking water during the studied periods; on the other hand, TRAP expression was changed (at 7 days) by chronic exposure to fluoride (p < 0.05). It was concluded that F in high concentrations can slow the blood clot remission and bone repair, and alter the TRAP expression in the beginning of the bone tissue repair. However, a better understanding about this blood clot remission phenomenon is required.