Acquired enamel pellicle and biofilm engineering with a combination of acid-resistant proteins (CaneCPI-5, StN15, and Hemoglobin) for enhanced protection against dental caries - in vivo and in vitro investigations.

OBJECTIVE: This study was designed in two-legs. In the in vivo, we explored the potential of a rinse solution containing a combination (Comb) of 0.1 mg/mL CaneCPI-5 (sugarcane-derive cystatin), 1.88 × 10- 5M StN15 (statherin-derived peptide) and 1.0 mg/mL hemoglobin (Hb) to change the protein profile of the acquired enamel pellicle(AEP) and the microbiome of the enamel biofilm. The in vitro, was designed to reveal the effects of Comb on the viability and bacterial composition of the microcosm biofilm, as well as on enamel demineralization. MATERIALS AND METHODS: In vivo study, 10 participants rinsed (10mL,1 min) with either deionized water (H2O-control) or Comb. AEP and biofilm were collected after 2 and 3 h, respectively, after rinsing. AEP samples underwent proteomics analysis, while biofilm microbiome was assessed via 16 S-rRNA Next Generation Sequencing(NGS). In vitro study, a microcosm biofilm protocol was employed. Ninety-six enamel specimens were treated with: 1)Phosphate-Buffered Solution-PBS(negative-control), 2)0.12%Chlorhexidine, 3)500ppmNaF and 4)Comb. Resazurin, colony-forming-units(CFU) and Transversal Microradiography(TMR) were performed. RESULTS: The proteomic results revealed higher quantity of proteins in the Comb compared to control associated with immune system response and oral microbial adhesion. Microbiome showed a significant increase in bacteria linked to a healthy microbiota, in the Comb group. In the in vitro study, Comb group was only efficient in reducing mineral-loss and lesion-depth compared to the PBS. CONCLUSIONS: The AEP modification altered the subsequent layers, affecting the initial process of bacterial adhesion of pathogenic and commensal bacteria, as well as enamel demineralization. CLINICAL RELEVANCE: Comb group shows promise in shaping oral health by potentially introducing innovative approaches to prevent enamel demineralization and deter tooth decay.

Acquired Pellicle and Biofilm engineering by rinsing with hemoglobin solution.

INTRODUCTION: The identification of acid-resistant proteins, including hemoglobin (Hb), within the acquired enamel pellicle (AEP) led to the proposition of the "acquired pellicle engineering" concept, which involves the modification of the AEP by incorporating specific proteins, presenting a novel strategy to prevent dental demineralization. OBJECTIVE: Combining in vivo and in vitro proof-of-concept protocols we sought to reveal the impact of AEP engineering with Hb protein on the biofilm microbiome and enamel demineralization. METHODS: In the in vivo studies, 10-volunteers, in 2 independent experiments, rinsed (10mL,1min) with: deionized water-negative control or 1.0mg/mL Hb. The AEP and biofilm formed along 2 or 3h, respectively, were collected. AEP was analyzed by quantitative shotgun-label-free proteomics and biofilm by 16S-rRNA Next-Generation-Sequencing (NGS). In vitro study, a microcosm biofilm protocol was employed. Seventy-two bovine enamel specimens were treated with: 1)Phosphate-Buffered Solution-PBS, 2)0.12% Chlorhexidine, 3)500ppm NaF; 4)1.0mg/mL Hb; 5)2.0mg/mL Hb, and 6)4.0mg/mL Hb. The biofilm was cultivated for 5-days. Resazurin, colony-forming-units(CFU) and Transversal Microradiography(TMR) were performed. RESULTS: Proteomics and NGS analysis revealed that Hb increased proteins with antioxidant, antimicrobial, acid-resistance, hydroxyapatite-affinity, calcium-binding properties and showed a reduction in oral pathogenic bacteria. In vitro experiments demonstrated that the lowest Hb concentration was the most effective in reducing bacterial activity, CFU and enamel demineralization compared to PBS. CONCLUSION: These findings suggest that Hb could be incorporated into anticaries dental products to modify the oral microbiome and control caries, highlighting its potential for AEP and biofilm microbiome engineering.

Adaptive responses of the ileum of NOD mice to low-dose fluoride: A proteomic exploratory study.

Fluoride (F) has been employed worldwide to control dental caries. More recently, it has been suggested that the consumption of low doses of F in the drinking water may reduce blood glucose levels, introducing a new perspective for the use of F for the management of blood glucose. However, the exact mechanism by which F affects blood glucose levels remains largely unexplored. Given that the small gut plays a pivotal role in glucose homeostasis, the aim of this study was to investigate the proteomic changes induced by low doses of F in the ileum of female nonobese-diabetic (NOD) mice. Forty-two female NOD mice were divided into two groups based on the F concentration in their drinking water for 14 weeks: 0 (control) or 10 mgF/L. At the end of the experimental period, the ileum was collected for proteomic and Western blot analyses. Proteomic analysis indicated an increase in isoforms of actin, gastrotropin, several H2B histones, and enzymes involved in antioxidant processes, as well as a decrease in enzymes essential for energy metabolism. In summary, our data indicates an adaptive response of organism to preserve protein synthesis in the ileum, despite significant alterations in energy metabolism typically induced by F, therefore highlighting the safety of controlled fluoridation in water supplies.

Optimized protocol for shotgun label-free proteomic analysis of pancreatic islets.

Pancreatic islets are crucial in diabetes research. Consequently, this protocol aims at optimizing both the protein-extraction process and the proteomic analysis via shotgun methods for pancreatic islets. Six protocols were tested, combining three types of chemical extraction with two mechanical extraction methods. Furthermore, two protocols incorporated a surfactant to enhance enzymatic cleavage. The steps involved extraction and concentration of protein, protein quantification, reduction, alkylation, digestion, purification and desalination, sample concentration to ∼1 µl, and proteomic analysis using the mass spectrometer. The most effective protocol involves either a milder chemical extraction paired with a more intensive mechanical process, or a more robust chemical extraction paired with a gentle mechanical process, tailored to the sample's characteristics. Additionally, it was observed that the use of a surfactant proved ineffective for these types of samples. Protocol 5 was recently used with success to examine metabolic changes in pancreatic islets of non-obese diabetic mice exposed to low doses of fluoride ions (F-) and the primary pathways altered by the treatment.

Increase in plasma resveratrol levels and in acid-resistant proteins in the acquired enamel pellicle after use of resveratrol-containing orodispersible tablets.

OBJECTIVE: This study evaluated the effect of administration of trans-resveratrol-containing orodispersible tablets on the protein composition of the AEP and on blood plasma trans-resveratrol concentrations. METHODS: Ten volunteers participated in two crossover double-blind phases. In each phase, after dental prophylaxis, they received a trans-resveratrol (15 mg) orodispersible tablet, or a placebo tablet (without actives). The AEP formed after 120 min was collected with electrode filter papers soaked in 3 % citric acid. Blood samples were collected 30, 45, 60 and 120 min after the use of the tablet. After protein extraction, AEP samples were analyzed by shotgun labelfree quantitative proteomics and plasma samples were analyzed by high-performance liquid chromatography (HPLC). RESULTS: Eight hundred and two proteins were identified in the AEP. Among them, 336 and 213 were unique to the trans-resveratrol and control groups, respectively, while 253 were common to both groups. Proteins with important functions in the AEP had increased expression in the trans-resveratroltreated group, such as neutrophil defensins, S100 protein isoforms, lysozyme C, cystatin-D, mucin-7, alphaamylase, albumin, haptoglobin and statherin. Trans-resveratrol was detected in the plasma at all the times evaluated, with the peak at 30 min. CONCLUSIONS: The administration of trans-resveratrol in sublingual orodispersible tablets was effective both to increase the bioavailability of the polyphenol and the expression of antibacterial and acid-resistant proteins in the AEP, which might benefit oral and general health.

Hemoglobin Protects Enamel against Intrinsic Enamel Erosive Demineralization.

INTRODUCTION: This study investigated the changes in the acquired enamel pellicle (AEP) proteome when this integument is formed in vivo after treatment with sugarcane-derived cystatin (CaneCPI-5), hemoglobin (HB), and a statherin-derived peptide (StN15), or their combination and then exposed to an intrinsic acid challenge. The effectiveness of these treatments in preventing intrinsic erosion was also evaluated. METHODS: Ten volunteers, after prophylaxis, in 5 crossover phases, rinsed with the following solutions (10 mL, 1 min): control (deionized water-H2O) - group 1, 0.1 mg/mL CaneCPI-5 - group 2, 1.0 mg/mL HB - group 3, 1.88 × 10-5M StN15 - group 4, or a blend of these - group 5. Following this, AEP formation occurred (2 h) and an enamel biopsy (10 µL, 0.01 m HCl, pH 2.0, 10 s) was conducted on one incisor. The biopsy acid was then analyzed for calcium (Arsenazo method). The vestibular surfaces of the other teeth were treated with the same acid. Acid-resistant proteins in the residual AEP were then collected and analyzed quantitatively via proteomics. RESULTS: Compared to control, treatment with the proteins/peptide, mixed or isolated, markedly enhanced acid-resistant proteins in the AEP. Notable increases occurred in pyruvate kinase PKM (11-fold, CaneCPI-5), immunoglobulins and submaxillary gland androgen-regulated protein 3B (4-fold, StN15), Hb, and lysozyme C (2-fold, StN15). Additionally, a range of proteins not commonly identified in the AEP but known to bind calcium or other proteins were identified in groups treated with the tested proteins/peptide either in isolation or as a mixture. The mean (SD, mM) calcium concentrations released from enamel were 3.67 ± 1.48a, 3.11 ± 0.72a, 1.94 ± 0.57b, 2.37 ± 0.90a, and 2.38 ± 0.45a for groups 1-5, respectively (RM-ANOVA/Tukey, p < 0.05). CONCLUSIONS: Our findings demonstrate that all treatments, whether using a combination of proteins/peptides or in isolation, enhanced acid-resistant proteins in the AEP. However, only HB showed effectiveness in protecting against intrinsic erosive demineralization. These results pave the way for innovative preventive methods against intrinsic erosion, using "acquired pellicle engineering" techniques.

Proteomic analysis of stimulated saliva in gastroesophageal reflux disease patients with and without erosive tooth wear: Observational study.

OBJECTIVE: To evaluate the difference in the proteomic profile of stimulated saliva in patients with gastroesophageal reflux disease (GERD) with (GE) and without (GNE) erosive tooth wear (ETW), regarding both human and bacterial proteins. METHODS: Stimulated saliva (SS) was collected from 16 patients (8/group). Samples were centrifuged at 4.500 g for 15 min under refrigeration to remove all debris. The supernatant from each saliva sample was taken and frozen at -80 °C. After extracting the proteins, they were submitted to reverse phase liquid chromatography and mass spectrometry (nLC-ESI-MS/MS). Label-free proteomic quantification was performed using Protein Lynx Global Service (PLGS) software (p < 0.05) for human and bacterial proteins. RESULTS: In total, 67 human proteins were common for GNE and GE groups. GNE group presented, compared to GE group, increase in proteins that confer antimicrobial and acid resistant properties, such as cystatins, histatin and immunoglobulins. However, GNE group had a marked decrease in subunits of hemoglobin (α, ß and delta). Regarding bacterial proteins, for SS, 7 and 10 unique proteins were identified in the GE and GNE groups, respectively. They are related to protein synthesis and energy metabolism and interact with human proteins typically found in saliva and supramolecular complexes of the acquired pellicle. CONCLUSIONS: Our data indicate that the stimulation of the salivary flow increases acid resistant and antimicrobial proteins in saliva, which might protect against ETW. CLINICAL SIGNIFICANCE: This pioneer study showed important differences in the human and bacterial proteome of SS in patients with GERD with or without ETW.

Acquired enamel pellicle protects gastroesophageal reflux disease patients against erosive tooth wear.

The objective of this study was to compare the protein profile of the acquired enamel pellicle (AEP) formed in vivo in patients with or without gastroesophageal reflux disease (GERD), and with or without erosive tooth wear (ETW). Twenty-four volunteers were divided into 3 groups: 1) GERD and ETW; 2) GERD without ETW; and 3) control (without GERD). The AEP formed 120 min after prophylaxis was collected from the lingual/palatal surfaces. The samples were subjected to mass spectrometry (nLC-ESI-MS/MS) and label-free quantification by Protein Lynx Global Service software. A total of 213 proteins were identified, or 119, 92 and 106 from each group, respectively. Group 2 showed a high number of phosphorylated and calcium-binding proteins. Twenty-three proteins were found in all the groups, including 14-3-3 protein zeta/delta and 1-phosphatidylinositol. Several intracellular proteins that join saliva after the exfoliation of oral mucosa cells might have the potential to bind hydroxyapatite, or participate in forming supramolecular aggregates that bind to precursor proteins in the AEP. Proteins might play a central role in protecting the dental surface against acid dissolution.

Prolonged exposure to high fluoride levels during adolescence to adulthood elicits molecular, morphological, and functional impairments in the hippocampus.

Fluoride is added to water due to its anticariogenic activity. However, due to its natural presence in soils and reservoirs at high levels, it could be a potential environmental toxicant. This study investigated whether prolonged exposure to fluoride from adolescence to adulthood-at concentrations commonly found in artificially fluoridated water and in fluorosis endemic areas-is associated with memory and learning impairments in mice, and assessed the molecular and morphological aspects involved. For this endeavor, 21-days-old mice received 10 or 50 mg/L of fluoride in drinking water for 60 days and the results indicated that the increased plasma fluoride bioavailability was associated with the triggering of short- and long-term memory impairments after high F concentration levels. These changes were associated with modulation of the hippocampal proteomic profile, especially of proteins related to synaptic communication, and a neurodegenerative pattern in the CA3 and DG. From a translational perspective, our data provide evidence of potential molecular targets of fluoride neurotoxicity in the hippocampus at levels much higher than that in artificially fluoridated water and reinforce the safety of exposure to low concentrations of fluoride. In conclusion, prolonged exposure to the optimum fluoride level of artificially fluoridated water was not associated with cognitive impairments, while a higher concentration associated with fluorosis triggered memory and learning deficits, associated with a neuronal density reduction in the hippocampus.

Effect of experimental toothpaste containing hydroxyapatite nanoparticles and propolis, with and without fluoride, on the microcosm biofilm and enamel demineralization.

This study evaluated the antimicrobial and anticaries effects of toothpaste containing hydroxyapatite nanoparticles (nanoHAP - 5 or 10%), xylitol (2 or 3%) and propolis (1 or 2%), associated or not with 1500 ppm fluoride (F). An in vitro model was used with microcosm biofilm produced from a pool of human saliva and McBain saliva (1:50) in the first 8 h of culture on 162 bovine enamel specimens. At the end of the experimental period, analyses of metabolic activity, colony forming units (CFU) and transverse microradiography (TMR) were performed. This study showed a possible decrease in demineralization and increase in remineralization by the commercial toothpaste (1500 ppm F) and for the experimental toothpaste containing the highest concentration of all agents, combined with F. In addition, a reduction in antimicrobial activity possibly caused by propolis and xylitol, mainly in relation to cariogenic bacteria, was observed.

Metabolic effect of low fluoride levels in the islets of NOD mice: integrative morphological, immunohistochemical, and proteomic analyses.

OBJECTIVES: Fluoride (F) has been widely used to control dental caries, and studies suggest beneficial effects against diabetes when a low dose of F is added to the drinking water (10 mgF/L). This study evaluated metabolic changes in pancreatic islets of NOD mice exposed to low doses of F and the main pathways altered by the treatment. METHODOLOGY: In total, 42 female NOD mice were randomly divided into two groups, considering the concentration of F administered in the drinking water for 14 weeks: 0 or 10 mgF/L. After the experimental period, the pancreas was collected for morphological and immunohistochemical analysis, and the islets for proteomic analysis. RESULTS: In the morphological and immunohistochemical analysis, no significant differences were found in the percentage of cells labelled for insulin, glucagon, and acetylated histone H3, although the treated group had higher percentages than the control group. Moreover, no significant differences were found for the mean percentages of pancreatic areas occupied by islets and for the pancreatic inflammatory infiltrate between the control and treated groups. Proteomic analysis showed large increases in histones H3 and, to a lesser extent, in histone acetyltransferases, concomitant with a decrease in enzymes involved in the formation of acetyl-CoA, besides many changes in proteins involved in several metabolic pathways, especially energy metabolism. The conjunction analysis of these data showed an attempt by the organism to maintain protein synthesis in the islets, even with the dramatic changes in energy metabolism. CONCLUSION: Our data suggests epigenetic alterations in the islets of NOD mice exposed to F levels comparable to those found in public supply water consumed by humans.

Unraveling molecular characteristic of fluoride neurotoxicity on U87 glial-like cells: insights from transcriptomic and proteomic approach.

The potential of fluoride (F) as a neurotoxicant in humans is still controversial in the literature. However, recent studies have raised the debate by showing different mechanism of F-induced neurotoxicity, as oxidative stress, energy metabolism and inflammation in the central nervous system (CNS). In the present study, we investigated the mechanistic action of two F concentration (0.095 and 0.22 µg/ml) on gene and protein profile network using a human glial cell in vitro model over 10 days of exposure. A total of 823 genes and 2,084 genes were modulated after exposure to 0.095 and 0.22 µg/ml F, respectively. Among them, 168 were found to be modulated by both concentrations. The number of changes in protein expression induced by F were 20 and 10, respectively. Gene ontology annotations showed that the main terms were related to cellular metabolism, protein modification and cell death regulation pathways, such as the MAP kinase (MAPK) cascade, in a concentration independent manner. Proteomics confirmed the changes in energy metabolism and also provided evidence of F-induced changes in cytoskeleton components of glial cells. Our results not only reveal that F has the potential to modulate gene and protein profiles in human U87 glial-like cells overexposed to F, but also identify a possible role of this ion in cytoskeleton disorganization.

Analysis of Plasma Proteins Involved in Inflammation, Immune Response/Complement System, and Blood Coagulation upon Admission of COVID-19 Patients to Hospital May Help to Predict the Prognosis of the Disease.

The development of new approaches allowing for the early assessment of COVID-19 cases that are likely to become critical and the discovery of new therapeutic targets are urgently required. In this prospective cohort study, we performed proteomic and laboratory profiling of plasma from 163 COVID-19 patients admitted to Bauru State Hospital (Brazil) between 4 May 2020 and 4 July 2020. Plasma samples were collected upon admission for routine laboratory analyses and shotgun quantitative label-free proteomics. Based on the course of the disease, the patients were divided into three groups: (a) mild (n = 76) and (b) severe (n = 56) symptoms, whose patients were discharged without or with admission to an intensive care unit (ICU), respectively, and (c) critical (n = 31), a group consisting of patients who died after admission to an ICU. Based on our data, potential therapies for COVID-19 should target proteins involved in inflammation, the immune response and complement system, and blood coagulation. Other proteins that could potentially be employed in therapies against COVID-19 but that so far have not been associated with the disease are CD5L, VDBP, A1BG, C4BPA, PGLYRP2, SERPINC1, and APOH. Targeting these proteins' pathways might constitute potential new therapies or biomarkers of prognosis of the disease.

Physical Training vs. Perindopril Treatment on Arterial Stiffening of Spontaneously Hypertensive Rats: A Proteomic Analysis and Possible Mechanisms.

(1) Background: Arterial stiffness is an important predictor of cardiovascular events. Perindopril and physical exercise are important in controlling hypertension and arterial stiffness, but the mechanisms are unclear. (2) Methods: Thirty-two spontaneously hypertensive rats (SHR) were evaluated for eight weeks: SHRC (sedentary); SHRP (sedentary treated with perindopril-3 mg/kg) and SHRT (trained). Pulse wave velocity (PWV) analysis was performed, and the aorta was collected for proteomic analysis. (3) Results: Both treatments determined a similar reduction in PWV (-33% for SHRP and -23% for SHRT) vs. SHRC, as well as in BP. Among the altered proteins, the proteomic analysis identified an upregulation of the EH domain-containing 2 (EHD2) protein in the SHRP group, required for nitric oxide-dependent vessel relaxation. The SHRT group showed downregulation of collagen-1 (COL1). Accordingly, SHRP showed an increase (+69%) in the e-NOS protein level and SHRT showed a lower COL1 protein level (-46%) compared with SHRC. (4) Conclusions: Both perindopril and aerobic training reduced arterial stiffness in SHR; however, the results suggest that the mechanisms can be distinct. While treatment with perindopril increased EHD2, a protein involved in vessel relaxation, aerobic training decreased COL1 protein level, an important protein of the extracellular matrix (ECM) that normally enhances vessel rigidity.

Salivary protein candidates for biomarkers of oral disorders in people with a crack cocaine use disorder.

The use of cocaine and its main derivative, crack, can cause some systemic effects that may lead to the development of some oral disorders. To assess the oral health of people with a crack cocaine use disorder and identify salivary protein candidates for biomarkers of oral disorders. A total of 40 volunteers hospitalized for rehabilitation for crack cocaine addiction were enrolled; nine were randomly selected for proteomic analysis. Intraoral examination, report of DMFT, gingival and plaque index, xerostomia, and non-stimulated saliva collection were performed. A list of proteins identified was generated from the UniProt database and manually revised. The mean age (n=40) was 32 (±8.88; 18-51) years; the mean DMFT index was 16±7.70; the mean plaque and gingival index were 2.07±0.65 and 2.12±0.64, respectively; and 20 (50%) volunteers reported xerostomia. We identified 305 salivary proteins (n=9), of which 23 were classified as candidate for biomarkers associated with 14 oral disorders. The highest number of candidates for biomarkers was associated with carcinoma of head and neck (n=7) and nasopharyngeal carcinoma (n=7), followed by periodontitis (n=6). People with a crack cocaine use disorder had an increased risk of dental caries and gingival inflammation; less than half had oral mucosal alterations, and half experienced xerostomia. As possible biomarkers for 14 oral disorders, 23 salivary proteins were identified. Oral cancer and periodontal disease were the most often associated disorders with biomarkers.

Gingivitis in cattle and supplemental protein diet: Insights from proteomic analysis.

Salivary proteins are essencial in the maintenance of oral homeostasis and can reflect systemic and localized processes, like gingivitis. However, little is known about the relationship between diet and the occurrence of gingivitis in cattle. The present study aimed to characterize the salivary proteomic profile of cattle (n = 12) fed hay (112.19 g/kg of crude protein) cultivated in reformed pastures, and, one group received protein supplement (PS, n = 6); the effect of the protein supplement on the gingival health of the cattle was determined by weekly intraoral examination and periodontal evaluation of the eight (deciduous) incisors. The whole saliva proteome of the two groups was evaluated after 20 and 60 days of confinement. In the periodontal clinical evaluation both groups had episodes of gingivitis; however, the average number of affected sites in the PS group was higher on day 60. The cattle fed exclusively hay, presented a lower average of affected gingival sites on day 60. After 60 days of experimentation, nine biological and 11 immunological processes were altered in bovine saliva. Proteins with multiple functions were detected in the saliva of the cattle; however, differences were observed in their regulation between the two groups. SIGNIFICANCE: In bovine populations, the relationship between diet and increased incidence of gingivitis is theorized. The results of the present pilot study, both diets caused episodes of gingivitis in the primary dentition of cattle and, apparently, diets with protein supplementation stimulate the expression of salivary proteins with a protective role in cattle that can act against infectious-inflammatory processes, such as gingivitis. However, it is plausible that over time, cattle will adapt to these diets and become more vulnerable to gingivitis.

Differential analysis of the dentin soluble proteomic.

OBJECTIVES: To perform a differential analysis of the dentin soluble proteomic and assess the effects of tissue health state and protocol for protein extraction. We hypothesized the dentin soluble proteomic varies according to the tissue physiopathological state (intact vs. caries-affected) and protocol used to extract its proteins. METHODS: Dentin from freshly extracted non-carious and carious teeth were randomly assigned for protein extraction using either guanidine-HCl/ethylenediaminetetraacetic acid (EDTA) or acetic acid. Protein extracts from intact and caries-affected dentin were processed and digested with trypsin for shotgun label-free proteomic analysis (nLC-ESI-MS/MS). Peptides identification was performed on a nanoACQUITY UPLC-Xevo Q-Tof MS system. Peptides identified with scores of confidence greater than 95% were included in the quantitative statistical analysis embedded in the PLGS software. Differences between experimental conditions were calculated using Student test-t with significance pre-set at α=0.05. RESULTS: A total of 158 human proteins were identified. Approximately one-sixth of proteins (24/158) were present in at least two different extracts. Conversely, the greatest number of proteins (134/158) was identified uniquely in only one of the extracts. Overall, a larger number of soluble proteins was retrieved from caries-affected than intact dentin (86/158). Likewise, a greater number of proteins was extracted by the guanidine-HCl/EDTA (106/158) in comparison to acetic acid protocol. Several proteins detected in dentin extracts, mainly those from caries-affected teeth, are biological and/or metabolically involved with tissue turnover/remodeling. CONCLUSION: The identity/abundance of soluble proteins retrieved from and remained in dentin noticeably depend on this tissue physiopathological state and protocol used to remove its minerals. CLINICAL SIGNIFICANCE: The present findings brought new insight into the proteomic phenotype of human dentin and may provide targets for the development of novel caries disease-prevention therapies.

Exposure to tolerable concentrations of aluminum triggers systemic and local oxidative stress and global proteomic modulation in the spinal cord of rats.

The tolerable aluminum (Al) intake levels for humans are constantly under review by regulatory agencies due to novel pre-clinical evidence on the neurotoxicity of prolonged Al exposure; however, little is known about the effects of Al on the spinal cord. This study aimed to investigate potential adverse effects on both spinal cord and systemic biochemical balance after prolonged exposure to a low dose of Al. Twenty adult rats were distributed in the control (distilled water) and exposed group (8.3 mg of AlCl3/kg/day). After 60 days, both blood and spinal cord samples were collected for oxidative stress and proteomic analyses. In plasma and erythrocytes, glutathione level was not different between groups; however, exposure to AlCl3 significantly decreased glutathione level in the spinal cord. Thiobarbituric acid reactive substances levels in the plasma and spinal cord of animals from the control group were significantly lower than those animals exposed to AlCl3. Exposure to AlCl3 significantly modulated the expression of proteins associated with the cell cycle, stimulus-response, cytoskeleton, nervous system regulation, protein activity, and synaptic signaling. Therefore, prolonged exposure to a low dose of Al triggered oxidative stress and proteomic changes that may affect spinal cord homeostasis.

Acquired enamel pellicle protects gastroesophageal reflux disease patients against erosive tooth wear

Abstract The objective of this study was to compare the protein profile of the acquired enamel pellicle (AEP) formed in vivo in patients with or without gastroesophageal reflux disease (GERD), and with or without erosive tooth wear (ETW). Twenty-four volunteers were divided into 3 groups: 1) GERD and ETW; 2) GERD without ETW; and 3) control (without GERD). The AEP formed 120 min after prophylaxis was collected from the lingual/palatal surfaces. The samples were subjected to mass spectrometry (nLC-ESI-MS/MS) and label-free quantification by Protein Lynx Global Service software. A total of 213 proteins were identified, or 119, 92 and 106 from each group, respectively. Group 2 showed a high number of phosphorylated and calcium-binding proteins. Twenty-three proteins were found in all the groups, including 14-3-3 protein zeta/delta and 1-phosphatidylinositol. Several intracellular proteins that join saliva after the exfoliation of oral mucosa cells might have the potential to bind hydroxyapatite, or participate in forming supramolecular aggregates that bind to precursor proteins in the AEP. Proteins might play a central role in protecting the dental surface against acid dissolution.

Metabolic effect of low fluoride levels in the islets of NOD mice: integrative morphological, immunohistochemical, and proteomic analyses

Abstract Fluoride (F) has been widely used to control dental caries, and studies suggest beneficial effects against diabetes when a low dose of F is added to the drinking water (10 mgF/L). Objectives This study evaluated metabolic changes in pancreatic islets of NOD mice exposed to low doses of F and the main pathways altered by the treatment. Methodology In total, 42 female NOD mice were randomly divided into two groups, considering the concentration of F administered in the drinking water for 14 weeks: 0 or 10 mgF/L. After the experimental period, the pancreas was collected for morphological and immunohistochemical analysis, and the islets for proteomic analysis. Results In the morphological and immunohistochemical analysis, no significant differences were found in the percentage of cells labelled for insulin, glucagon, and acetylated histone H3, although the treated group had higher percentages than the control group. Moreover, no significant differences were found for the mean percentages of pancreatic areas occupied by islets and for the pancreatic inflammatory infiltrate between the control and treated groups. Proteomic analysis showed large increases in histones H3 and, to a lesser extent, in histone acetyltransferases, concomitant with a decrease in enzymes involved in the formation of acetyl-CoA, besides many changes in proteins involved in several metabolic pathways, especially energy metabolism. The conjunction analysis of these data showed an attempt by the organism to maintain protein synthesis in the islets, even with the dramatic changes in energy metabolism. Conclusion Our data suggests epigenetic alterations in the islets of NOD mice exposed to F levels comparable to those found in public supply water consumed by humans.