Salivary Proline-rich Proteins Research Articles

#1865 The role of osteoclast-associated receptor in phosphorus metabolism via salivary-bone-kidney interaction

Abstract Background and Aims We have identified osteoclast-associated receptor (OSCAR) in the osteoclasts and salivary proline-rich proteins (S-PRPs), which are specifically secreted from the salivary glands and are carrier proteins for calcium and phosphorus, as new factors involved in phosphorus metabolism using bulk RNA sequencing of all organs in a high-phosphate (HP) loaded mouse model with kidney disease. The roles of OSCAR have been reported to include activation of NFATc1, which is essential for osteoclast differentiation, and inhibition of apoptosis. However, to our knowledge, no study has examined the relationship between OSCAR and chronic kidney disease-mineral bone disorder (CKD-MBD). In this study, we investigated the role of OSCAR in phosphorus metabolism particularly involving the salivary grands (S-PRPs) in mice models of high phosphorus load and kidney disease. Method We used 11- to 13-week C57BL/6N male mice to compare phosphorus metabolism between wild-type (WT) and OSCAR knockout (KO) mice. WT and OSCAR KO mice underwent sham surgery (Ctrl group) or 5/6 nephrectomy (CKD group), followed by HP loading (1.8% phosphate diet) for 1 week. After HP loading, these mice were kept in metabolic gauges for 24 hours, during which urine and fecal samples were collected. Subsequently, blood and organ (parotid grand and kidney) samples were collected. The parotid grands, the main source of S-PRPs, were harvested after 1 week of HP loading, and specific mRNA for S-PRPs (Prb1 and Prpmp5) and for phosphate (Pi) transporter (NaPi2b and Pit-2), responsible for Pi reabsorption in primary saliva, were examined. Results In the non-CKD group, there were no significant difference in Pi homeostasis including plasma Pi, FGF23, PTH, or urinary, fecal, and salivary Pi excretion levels after HP loading. In the CKD group, OSCAR KO mice showed significantly lower urinary Pi excretion than WT mice (WT vs OSCAR KO; 25.43 ± 3.88 mg/day vs 17.65 ± 5.75 mg/day, P = .006) after HP loading, but no difference in fecal Pi excretion was seen between genotypes. HP-induced plasma intact and C-terminal FGF-23 elevations were significantly suppressed in OSCAR KO mice (WT vs OSCAR KO; 6857 ± 4484 pg/mL vs 1573 ± 524 pg/mL, P = .002), indicating that the increased FGF23 “production” was suppressed in HP-loaded CKD/OSCAR KO mice. In contrast, there was no difference in plasma PTH levels between OSCAR KO and WT mice (WT vs OSCAR KO; 4189 ± 1361 pg/mL vs 5082 ± 1120 pg/mL, P = .22). As a result, plasma Pi levels in WT mice were not significantly different from those in OSCAR KO mice after 1-week HP loading (WT vs OSCAR KO; 26.91 ± 7.76 mg/dL vs 28.87 ± 7.49 mg/dL, P = 0.61), suggesting an increased internal Pi burden to organs other than the blood stream such as bones and ectopic calcifications in OSCAR KO mice compared to WT mice after HP loading and CKD. In the parotid gland, S-PRPs encoding mRNA (Prb1 and Prpmp5) expressions were decreased in OSCAR KO mice compared to those in WT mice, with or without CKD. Pi reabsorption transporter mRNA expression in the parotid gland was also decreased in OSCAR KO mice compared to those in WT mice, with or without CKD, after 1-week HP loading, suggesting an increase in salivary Pi concentrations in OSCAR KO mice compared to WT mice. Conclusion These results suggest that OSCAR may regulate urinary Pi excretion via FGF-23 in the bone. OSCAR was also significantly associated with salivary Pi levels via S-PRPs and Pi transporter in the parotid grands. OSCAR regulate Pi homeostasis through interactions with the kidney, salivary grands, bone, and intestine.

Astringency sensitivity to tannic acid: Effect of ageing and salivary proline-rich protein levels

The link between salivary composition and sensitivity to astringency as a function of age has still not been established. In this work, we propose the hypothesis that ageing leads to changes in the concentration of salivary proline-rich proteins (PRPs), which alters the astringency perception threshold with age. To test this hypothesis, astringency sensitivity to tannic acid and saliva was assessed in 30 elderly people and 24 young people. Basic PRPs (bPRPs) and glycosylated PRPs (gPRPs) were quantified immunochemically via western blot analysis. The results showed that the amounts of bPRPs and gPRPs were similar between the young and elderly groups. However, a positive correlation between the gPRP amount and astringency threshold was observed only in the young group, while a negative correlation between the bPRP amount and astringency threshold was observed only in the elderly group. This finding suggests differences in the contribution of PRP type to astringency perception as a function of age.

Study on the interaction of tannins and salivary proteins affecting wine aroma volatility: Static HS-SPME and molecular dynamics simulation approaches

The interaction between tannins and salivary proteins might affect intraoral aroma release during wine consumption. In this study, the influence and underlying mechanism of interactions between EGCG and IB5 (salivary proline-rich protein) on wine aroma compounds was analysed by static HS-SPME in vitro and molecular dynamics (MD) simulation. The interaction between IB5 and EGCG could significantly reduce the volatility of most aroma compounds in the model wine by 20 %-70 % (p < 0.05). MD simulations indicated that the energy received by aroma compounds in the mixed system was more pronounced. In addition, the decline rate of rational correlation functions (RCF) of aroma compounds in the mixed system was obviously slower. The analysis of the independent gradient model (IGM) indicated that aroma compounds combined with aggregates of IB5 and EGCG through hydrogen bonds and van der Waals forces. The effect of the interaction between EGCG and IB5 on aroma compounds was confirmed by the volatility and molecular computational simulation. Overall, the results enhance the understanding of the mechanisms affecting retronasal aroma release during wine consumption.

Alcohol-Induced Conformation Changes and Thermodynamic Signatures in the Binding of Polyphenols to Proline-Rich Salivary Proteins.

The first contact of polyphenols (tannins) with the human body occurs in the mouth, where they are known to interact with proline-rich proteins (PRPs). These interactions are important at a sensory level, especially for the development of astringency, but affect also various other biochemical processes. Employing thermodynamic measurements, fluorescence and CD spectroscopy, we investigated the binding process of the prototypical polyphenol ellagic acid (EA) to different IB-PRPs and BSA, also in the presence of ethanol, which is known to influence tannin-protein interactions. Binding of EA to BSA and the small peptide IB7-14 is weak, but very strong to IB9-37. The differences in binding strength and stoichiometry are due to differences in the binding motifs, which also lead to differences in the thermodynamic signatures of the binding process. EA binding to BSA is enthalpy-driven, whereas binding to both IB7-14 and IB9-37 is entropy-driven. The presence of 10 vol.% EtOH, as present in wines, increases the binding constant of EA with BSA and IB7-14 drastically, but not that with IB9-37; however, it changes the binding stoichiometry. These differences can be attributed to the effect of EtOH on the conformation dynamics of the proteins and to changes in hydration properties in alcoholic solution.

Role of Anthocyanins in the Interaction between Salivary Mucins and Wine Astringent Compounds.

Wine astringency is a very complex sensation whose complete mechanism has not been entirely described. Not only salivary proline-rich proteins (PRPs) are involved in its development; salivary mucins can also play an important role. On the other hand, it has been described that anthocyanins can interact with PRPs, but there is no information about their potential role on the interactions with mucins. In this work, the molecular interactions between salivary mucins (M) and different wine phenolic compounds, such as catechin (C), epicatechin (E) and quercetin 3-β-glucopyranoside (QG), as well as the effect of the anthocyanin malvidin 3-O-glucoside (Mv) on the interactions with mucins, were assessed by isothermal titration calorimetry (ITC). Results showed that the interaction between anthocyanin and mucins is stronger than that of both flavanols analyzed, since the affinity constant values were 10 times higher for anthocyanin than for catechin, the only flavanol showing interaction in binary assay. Moreover, at the concentration at which polyphenols are usually found in wine, flavonols seem not to be involved in the interactions with mucins. These results showed, for the first time, the importance of wine anthocyanins in the mechanisms of astringency involving high-molecular-weight salivary proteins like mucins.

Influence of tannic acid concentration on the physicochemical characteristics of saliva of spider monkeys (Ateles geoffroyi).

Tannins are a chemical defense mechanism of plants consumed by herbivores. Variations in salivary physicochemical characteristics such as pH, total protein concentration (TP), and presence of proline-rich proteins (PRPs) in animals have been reported as a mechanism to protect the oral cavity when consuming food with variations in pH and tannins. Variations in salivary physiochemistry as adaptations for consuming tannin-rich foods have been found in omnivorous and folivorous primates, but have not yet been reported in frugivorous species such as spider monkeys. We therefore assessed changes in pH using test strips, TP concentration by measuring absorbance at 595 nm in a spectrophotometer and salivary PRPs using the SDS-PAGE electrophoresis technique in the saliva of nine captive spider monkeys in response to the consumption of solutions with different concentrations of tannic acid. The results showed variations in pH, TP concentration and the presence and variation of possible salivary PRPs associated with tannic acid concentration. These findings suggest that spider monkeys may tailor their salivary physicochemical characteristics in response to the ingestion of potentially toxic compounds.

Association of increased basic salivary proline-rich protein 1 levels in induced sputum with type 2-high asthma.

BackgroundThe aim of this study is to reveal whether basic salivary proline‐rich protein BstNI subfamily 1 (PRB1) may be used as a diagnostic biomarker for type 2‐high asthma.MethodsPRB1 protein levels in the induced sputum of 67 subjects with asthma and 27 controls were determined by an enzyme‐linked immunosorbent assay. Correlation analyses between PRB1 in the induced sputum and airway inflammatory indicators were also performed.ResultsPRB1 protein levels were significantly upregulated in the induced sputum of asthmatic patients (p =0.0098) and correlated with clinical eosinophil‐related indicators and type 2 airway inflammation. These results indicate that PRB1 is a promising biomarker for type 2‐high asthma.ConclusionsThe expression of PRB1 in induced sputum is a potential biomarker for type 2‐high asthma. The results of this study present new insights into the diagnosis and treatment of asthma.

A tribological approach to astringency perception and astringency prevention

Astringency is one of the most complex oral sensations. This dry, puckering mouthfeel occurs when consuming wine, tea, or other foods containing polyphenols. The exact mechanism behind this dry mouthfeel is not completely understood. Here, we describe a systematic tribological approach to measure model and real saliva to understand the specific role of the salivary proteins (mucins and proline-rich proteins (PRPs)) on lubrication-based astringency. Our approach reveals that there are two routes towards lubrication losses, partly involving irreversible molecular mechanisms for which the order of reactivity matters. For human saliva, we find two lubrication mechanisms: (I) Using phenolic compounds, we find aggregation-induced lubrication losses due to hydrogen bond formation, which depend critically on phenol size: large polyphenols allow for aggregation-induced lubrication losses, but small phenols do not. (II) For metal salts combined with saliva, we observe aggregation without lubrication losses as a result of electrostatic interactions. We find that lubrication losses are caused by the specific removal of the salivary PRP layer, whereas mucin aggregation in the presence of PRPs does not lead to lubrication losses. Additionally, we show that the addition of solvents that are able to reduce protein-polyphenol hydrogen bonding (e.g. ethanol) can prevent lubrication losses. Lubrication losses can also be compensated by the addition of highly viscous fluids (glycerol) that can provide viscous lubrication.

Downregulation of Salivary Proteins, Protective against Dental Caries, in Type 1 Diabetes.

Saliva, an essential oral secretion involved in protecting the oral cavity’s hard and soft tissues, is readily available and straightforward to collect. Recent studies have analyzed the salivary proteome in children and adolescents with extensive carious lesions to identify diagnostic and prognostic biomarkers. The current study aimed to investigate saliva’s diagnostic ability through proteomics to detect the potential differential expression of proteins specific for the occurrence of carious lesions. For this study, we performed bioinformatics and functional analysis of proteomic datasets, previously examined by our group, from samples of adolescents with regulated and unregulated type 1 diabetes, as they compare with healthy controls. Among the differentially expressed proteins relevant to caries pathology, alpha-amylase 2B, beta-defensin 4A, BPI fold containing family B member 2, protein S100-A7, mucin 5B, statherin, salivary proline-rich protein 2, and interleukin 36 gamma were significantly downregulated in poorly-controlled patients compared to healthy subjects. In addition, significant biological pathways (defense response to the bacterium, beta-defensin activity, proline-rich protein activity, oxygen binding, calcium binding, and glycosylation) were deregulated in this comparison, highlighting specific molecular characteristics in the cariogenic process. This analysis contributes to a better understanding of the mechanisms involved in caries vulnerability in adolescents with unregulated diabetes.

Estimation of salivary proteins in early childhood caries before and after treatment using gel electrophoresis.

Saliva being an important biological fluid of our body contains both specific and nonspecific protective factors which form an integral part of our immune system. Salivary proteins play a substantial role in protecting humans against infection. Their level in oral cavity is subject to constant variations which is dependent on various factors. The purpose of the study was to compare the levels of salivary proline-rich proteins (PRPs), glycoproteins, amylase bands, and salivary pH in children with early childhood caries before and after treatment using gel electrophoresis. The whole salivary pH, mean protein concentrations, and electrophoretic profiles of the salivary proteins were measured using sodium dodecylsulfate-polyacrylamide gel electrophoresis in both pre- and posttreatment groups. Statistical analysis was done using Statistical Package for Social Sciences (SPSS) version 15.0 software. Chi-square test and independent t-test were used to further compare the results. The results were statistically significant in all the groups. There was variation in pre- and posttreatment values seen. Salivary proteins (glycoproteins, PRPs, and amylase) establish an imperative defense mechanism as antimicrobial agents.

Cell Wall Mannoproteins from Yeast Affect Salivary Protein-Flavanol Interactions through Different Molecular Mechanisms.

It is known that interactions between wine flavanols and salivary proline-rich proteins (PRPs) are one of the main factors responsible for wine astringency. The addition of commercial yeast mannoproteins (MPs) to wines has been pointed to as a possible tool to modulate the excessive astringency due to a lack of phenolic maturity at harvest time that might occur as a consequence of global climate change. The aim of this work was to study by isothermal titration calorimetry and molecular dynamics simulation the molecular mechanisms by which mannoproteins could modulate astringency elicited by tannins and if it can be influenced by mannoprotein composition. Results obtained indicate that the MPs assayed had an important impact on astringency through the formation of ternary aggregates with different solubilities or by preventing the flavanol-PRP interaction by a competitive mechanism, although in a different strength, depending on the size and the compositional characteristic of the mannoprotein.

Salivary proteomic profile of patients with renal cell carcinoma.

622 Background: We aimed to explore the salivary proteomic profile (SPP) of patients undergoing surgery for suspected renal cell carcinoma (RCC), then evaluate any significant findings in a validation cohort. Methods: Prospective IRB approved trial of consecutive patients with suspected RCC undergoing curative surgery. Saliva was collected pre- and postoperatively, and from healthy volunteers. Saliva was mixed with sodium orthovanadate and protease inhibitor, aliquoted, and frozen at -80oC. SPP performed using biomarker concentrated saliva via core-shell hydrogel nanoparticles functionalized with molecular baits coupled to high resolution MS/MS electrospray mass spectrometry analysis; output represented as spectral counts. Wilcoxon Rank Sum test was used for comparisons; p-values were adjusted using false discovery rate (FDR) for multiple testing. Results: Exploratory phase: saliva was collected from 27 volunteers and 34 RCC patients (24 clear cell (cc)RCC, 6 chromophobe (ch)RCC, 4 other). Evaluation of cc vs chRCC revealed differential counts for 56 proteins; none were significant after adjustment by FDR. Comparison of matched pre/post-operative SPP in 14 ccRCC patients identified differential counts of 3 proteins: basic salivary proline-rich protein 3 precursor; prolactin-inducible protein precursor; and submaxillary gland androgen-regulated protein 3B precursor. These were not affected by patient age, partial/radical nephrectomy, or tumor size/stage/grade. Validation phase: power analyses indicated a sample size of 38/53 patients = 90% power = effect size of 0.6/0.5, respectively, 2-sided Type 1 error of 0.025. 49 ccRCC patients were enrolled with pre/postop saliva. 611 proteins were identified; none had differential counts after FDR. Conclusions: Using a unique biomarker technology coupled to high resolution MS, interesting SPP’s were seen between RCC subtypes and before/after ccRCC curative resection. We were unable to show significant differences in pre/post-op levels. More research is needed to further study the SPP in RCC patients and the factors impacting their differential expression. In order to aid such efforts, the raw data will be made available for future researchers.

COMBINED ANTIBACTERIAL ACTION OF SALIVARY CATIONIC PROLINE-RICH PEPTIDES AND ANTIMICROBIAL PEPTIDES

Saliva is an important biological fluid that reflects human’s health. Its main function is protection of the oral cavity from pathogens. Antimicrobial peptides (AMPs) of the innate immunity may play an important role in anti-infectious defense of the oral cavity, but their relative amount in saliva is low. It’s major component is Proline-rich peptides (PRPs), whose impact in antimicrobial protection remains poorly understood. We suggest that salivary PRPs may reveal their defensive functions upon interaction with other molecules, in particular with AMPs. The aim of this work is an investigation of the combined antibacterial action of salivary PRPs (fragments of Basic salivary proline-rich protein 1: P-H (37-51), IB6 (98-116), p1932) with antimicrobial peptides (histatin 5 and cathelicidin LL-37 and beta-defensin hBD3). Listed PRPs have been obtained by chemical solid-phase synthesis. The method of broth microdilutions was used to compare minimal inhibitory concentrations (MICs) of individual fractions of AMPs and their MICs in the presence of salivary peptides. It was found that in the presence of peptides IB6 (98-116) or P-H (37-51) the activity of defensin hBD3 was increased (reduction of MICs by 2 times) against Staphylococcus aureus SG511. In the presence of IB6 (98-116) or p1932 the activity of this defensin against E. coli ML35p was also improved (MICs of hBD3 was lowered by 2 times). For other combinations of the peptides, this effect was not observed. The obtained data confirm the assumption that the combined action of varied salivary peptides, including cationic Proline-rich peptides, plays an important role in anti-infectious protection of the oral cavity.

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

ObjectiveThis 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. Design324 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. ResultsA 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. ConclusionThe 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.

Interaction between Ellagitannins and Salivary Proline-Rich Proteins.

The first contact of tannins with the human body occurs in the mouth, where some of these tannins are known to interact with salivary proteins, in particular with proline-rich proteins (PRPs). These interactions are important at a sensory level, especially for astringency development, but could also affect the biological activities of the tannins. This study gathers information on the relative affinity of the interaction, complex stoichiometry, and tannin molecular epitopes of binding for the interactions between the families of PRPs (bPRPs, gPRPs, and aPRPs) and three representative ellagitannins (castalagin, vescalagin, and punicalagin). These interactions were studied by saturation-tranfer difference NMR and microcalorimetry. The effect of the PRP-ellagitannin interaction on their antioxidant ability was also assessed by ferric reduction antioxidant power (FRAP) assays. The results support a significant interaction between the studied tannins and PRPs with binding affinities in the micromolar range. Punicalagin was always the ellagitannin with higher affinity. aPRPs were the salivary PRPs with higher affinity. Moreover, it was observed that when ellagitannins are present in low concentrations (5-50 μM), as occurs in food, the antioxidant ability of these tannins when complexed with salivary PRPs could be significantly impaired.

Salivary Cystatin SN Binds to Phytic Acid In Vitro and Is a Predictor of Nonheme Iron Bioavailability with Phytic Acid Supplementation in a Proof of Concept Pilot Study

ABSTRACTBackgroundAcute phytic acid intake has been found to decrease iron bioavailability; however, repeated phytic acid consumption leads to iron absorption adaptation. Salivary proline-rich proteins (PRPs) have been shown to inhibit iron chelation to tannins and may mediate similar iron absorption adaptation with phytic acid intake.ObjectivesThe objectives of this study were to determine whether salivary proteins bind to phytic acid in vitro, and to explore a proof of concept in a pilot study that examined the impact of 4-wk, daily phytic acid supplementation on individuals’ iron status, bioavailability, and salivary PRP concentrations.MethodsHigh-performance liquid chromatography (HPLC) and matrix-assisted laser desorption/ionization–time of flight were used to characterize in vitro salivary protein–phytic acid interactions. Nonanemic women (n = 7) consumed 350 mg phytic acid supplements 3 times daily for 4 wk, and meal challenges were employed to determine iron bioavailability, iron status, and salivary protein concentrations before and after supplementation periods. Enzyme-linked immunosorbent assay (ELISA) analysis of purified protein fractions and participant saliva identified proteins bound to phytic acid.ResultsIn vitro salivary protein–phytic acid interaction identified cystatin SN, a non–proline rich salivary protein, as the specific bound protein to phytic acid. Iron bioavailability (P = 0.32), hemoglobin (P = 0.72), and serum ferritin (P = 0.08) concentrations were not reduced from week 0 to week 4 after phytic acid supplementation. Basic PRPs and cystatin SN concentrations were positively correlated with iron bioavailability at week 4.ConclusionsOverall, results suggest that phytic acid binds to the non-PRP cystatin SN and that salivary protein production may improve iron bioavailability with phytic acid consumption.

Proteomics of acquired pellicle in gastroesophageal reflux disease patients with or without erosive tooth wear

ObjectivesThis in vivo study compared the protein profile of the acquired enamel pellicle (AEP) in volunteers 1) with gastroesophageal reflux disease (GERD) and erosive tooth wear (ETW) (BEWE ≥ 9; GE group); 2) with GERD without ETW (BEWE = 0; GNE group) and 3) control (without GERD and BEWE = 0; C group). Materials and methodsTwenty-four subjects (8/group) participated. AEP was formed during 120 min and collected. After protein extraction, the samples were submitted to reverse phase liquid chromatography coupled to mass spectrometry. Label-free proteomic quantification was performed using Protein Lynx Global Service software. ResultsIn total, 458 proteins were identified. Seventy-six proteins were common to all the groups. The proteomic profile of the AEP was quite different among the distinct groups. The numbers of proteins exclusively found in the C, GE and GNE groups were 113, 110 and 81, respectively. Most of the proteins exclusively identified in the C and GNE groups bind metals, while those in the GE group are mainly membrane proteins. Many proteins were found exclusively in the reflux groups. In the quantitative analyses, when the GNE group was compared with the GE group, the proteins with the highest decreases were Lysozyme C, Antileukoproteinase, Cathepsin G, Neutrophil defensins and Basic salivary proline-rich proteins, while those with the highest increases were subunits of Hemoglobin, Albumin and isoforms of Cystatin. ConclusionProfound alterations in the proteomic profile of the AEP were seen in GNE compared with GE volunteers, which might play a role in the resistance to ETW seen in the first. Clinical significanceThis pioneer study compared the proteomic profile of the AEP of patients with GERD with or without ETW. Increased proteins in those without ETW might be protective and are good candidates to be added to dental products to protect against erosion caused by intrinsic acids.

Effect of malvidin-3-glucoside and epicatechin interaction on their ability to interact with salivary proline-rich proteins.

At red wine pH, malvidin-3-glucoside (mv-3-glc), the major anthocyanin of red wine, is expected to be present mainly in its non-colored hemiketal form. However, due to copigmentation with flavanols (e.g. epicatechin), the stabilization of the colored forms of mv-3-glc occurs. Some flavanols have been linked to astringency, due to their ability to interact/precipitate salivary proteins, namely proline-rich proteins (PRPs). So, a major question is if this copigmentation interaction could affect the ability of flavanols to interact with SP. To answer this, the effect of the interaction between mv-3-glc and epicatechin with basic and acidic PRPs, was investigated by saturation-tranfer difference (STD)-NMR and isothermal titration calorimetry (ITC). The most relevant result was that epicatechin:mv-3-glc mixture presents a synergic effect toward the interaction with both PRPs when compared to individual polyphenols. Furthermore, was observed that epicatechin interaction was driven by hydrophobic and hydrophilic interactions while mv-3-glc interaction was driven by electrostatic interactions.

Oral Lactobacilli and salivary acidic proline-rich proteins (APRP-1/2) in dental caries.

A previous study shows that levels of acidic salivary proline-rich phosphoproteins-1/2 (APRP-1/2) increase with caries severity. The aim of this study was to examine whether this relationship also depends on the presence of H2O2-producing strains of Lactobacillus spp. Adults with severe caries (decayed, missing, and filled teeth (DMFT) > 13.9, n = 28) were compared with similarly aged adults who had minimal caries (DMFT < 5, n = 20). A total of 48 samples of whole unstimulated saliva were collected in the morning and centrifuged. Lactobacillus spp. were isolated from the sediment in Rogosa agar and peroxide (H2O2) production was determined by growing the isolates on TMB-Plus agar. Salivary APRP-1/2 content in the saliva supernatant was estimated using an enzyme-linked immunosorbent sandwich assay (ELISA). Lactobacilli were present in 67% of both caries groups but were H2O2 positive only in the minimal caries group. Irrespective of the presence of Lactobacilli, the total content of APRP-1/2 proteins was 34.5 ± 4.9 ng/ml in severe caries but just under half this in minimal caries. We conclude that Lactobacillus spp. was absent from about a third of the severe and minimal caries groups, and H2O2-producing strains were present only in the minimal caries group. The severe caries group possessed twice the content of salivary APRP 1/2 proteins as the minimal caries group. The implications of these findings for caries development are discussed.

Comparative salivary proteomics analysis of children with and without dental caries using the iTRAQ/MRM approach

BackgroundDental caries is a major worldwide oral disease afflicting a large proportion of children. As an important host factor of caries susceptibility, saliva plays a significant role in the occurrence and development of caries. The aim of the present study was to characterize the healthy and cariogenic salivary proteome and determine the changes in salivary protein expression of children with varying degrees of active caries, also to establish salivary proteome profiles with a potential therapeutic use against dental caries.MethodsIn this study, unstimulated saliva samples were collected from 30 children (age 10–12 years) with no dental caries (NDC, n = 10), low dental caries (LDC, n = 10), and high dental caries (HDC, n = 10). Salivary proteins were extracted, reduced, alkylated, trypsin digested and labeled with isobaric tags for relative and absolute quantitation, and then they were analyzed with GO annotation, biological pathway analysis, hierarchical clustering analysis, and protein–protein interaction analysis. Targeted verifications were then performed using multiple reaction monitoring mass spectrometry.ResultsA total of 244 differentially expressed proteins annotated with GO annotation in biological processes, cellular component and molecular function were identified in comparisons among children with varying degrees of active caries. A number of caries-related proteins as well as pathways were identified in this study. As compared with caries-free children, the most significantly enriched pathways involved by the up-regulated proteins in LDC and HDC were the ubiquitin mediated proteolysis pathway and African trypanosomiasis pathway, respectively. Subsequently, we selected 53 target proteins with differential expression in different comparisons, including mucin 7, mucin 5B, histatin 1, cystatin S and cystatin SN, basic salivary proline rich protein 2, for further verification using MRM assays. Protein–protein interaction analysis of these proteins revealed complex protein interaction networks, indicating synergistic action of salivary proteins in caries resistance or cariogenicity.ConclusionsOverall, our results afford new insight into the salivary proteome of children with dental caries. These findings might have bright prospect in future in developing novel biomimetic peptides with preventive and therapeutic benefits for childhood caries.