The death of oral keratinocytes is a crucial step in the emergence of recurrent aphthous stomatitis (RAS, also known as aphthae or aphthous ulcers). Since there are no experimental models available to research aphthous ulcers, little is understood about this process. We hypothesize that saliva can be a data bank of information that offers insights on epithelial damage.
In this case-crossover study, we assessed the salivary proteome of patients with RAS (n = 36) in the presence and absence of ulcers using discovery proteomics and bioinformatics. Additionally, we contrasted these patterns with those of healthy individuals (n = 31) who had no prior aphthous ulceration.
Salivary proteome showed that during the ulcerative phase, controlled cell death was downregulated. Due to its ability to distinguish between individuals with and without ulcers, the ATF6B protein raises the possibility that endoplasmic reticulum (ER) stress is responsible for the damage that leads to the death of oral keratinocytes. The high abundance of TRAP1 and ERN1 matches with this biological discovery. The type of death is immunogenic, according to the functional data found in a cell death database.
We identified a cellular process that can lead to the death of oral keratinocytes in the etiopathogenesis process of RAS. Future studies should be conducted to identify what is responsible for the increase in ER stress signaling that would lead to an anti-cell death response.

Objectives: Opiorphin is a physiological inhibitor of peptidases inactivating endogenous opioids displaying strong analgesic properties without undesirable side effects, antidepressant properties or hormonal dependency. It might therefore play an important role in patients with painful diseases related to neuro-hormonal dysregulation of the nervous system, affecting saliva secretion and composition such as anorexia nervosa (AN). The main objective aim of this study was to compare the level of opiorphin in saliva of patients with AN to matched subjects free of eating disorders.Methods: A case-control clinical trial was conducted in 68 AN patients and 43 healthy matched control subjects. Depression symptoms were assessed with the self-scored questionnaire Beck Depression Inventory (BDI) and salivary samples were taken during the acute stage of AN (BMI <15 kg/m2) for measuring opiorphin. Opiorphin levels were measured with a quantitative assay using a commercial immunoenzymatic Elisa kit (cat no. EH1927, Wuchan, Hubei, China).Results: No statistically significant difference was found in salivary opiorphin levels between the AN and control groups, (P = 0.499, Mann-Whitney U-test). Positive correlations to duration of the disease, BDI and bodyweight in AN patients were evidenced.Conclusions: Measurement of salivary opiorphin levels cannot be used as a marker of AN but may allow new perspectives in monitoring AN in its early stages.

Gaining insight into intermolecular interactions between multiple species is possible at an atomic level by looking at different parameters using different NMR techniques. In the specific case of the astringency sensation, in which at least three molecular species are involved, different NMR techniques combined with dynamic light scattering and molecular modeling contribute to decipher the role of each component in the interaction mode and to assess the thermodynamic parameters governing this complex interaction. The binding process between a saliva peptide, a polyphenol, and polysaccharides was monitored by following 1H chemical shift variations, changes in NMR peak areas, and size of the formed complex. These NMR experiments deliver a complete picture of the association pathway, assessed by dynamic light scattering and molecular dynamics simulations: all of the data collected converge toward a comprehensive mode of interaction in which sugars indirectly play a role in astringency by sequestering part of the polyphenols, reducing their effective concentration to bind saliva proteins.

OBJECTIVE: Opiorphin is a pentapeptide isolated from human saliva that suppresses pain from chemically induced inflammation and acute physical pain. Burning mouth syndrome (BMS) is a chronic condition of a burning sensation in the mouth, where no underlying dental or medical cause can be identified. We aimed to measure the level of opiorphin in whole unstimulated (UWS) and stimulated (SWS) saliva of patients with BMS.
METHODS: Originally developed and validated LC-MS/MS method was used for opiorphin quantification. Samples were obtained from 29 BMS patients and 29 age- and sex-matched controls.
RESULTS: The average concentration of opiorphin in UWS and SWS in the BMS group was 8.13 ± 6.45 and 5.82 ± 3.59 ng/ml, respectively. Opiorphin in BMS patients\' UWS was significantly higher, compared to the control group (t = 2.5898; p = 0.0122). SWS opiorphin levels were higher, but not significantly, in BMS patients than in controls.
CONCLUSIONS: Our results indicate that higher quantities of salivary opiorphin in BMS may be a consequence of chronic pain, but we cannot exclude that they occur as a result of emotional and behavioral imbalances possibly associated with BMS. To our knowledge, this is the first original article measuring opiorphin in a pain disorder.
CONCLUSIONS: Opiorphin may be a measurable biomarker for chronic pain, which could help in objectifying otherwise exclusively a subjective experience. Increased opiorphin could serve as a universal objective indicator of painful conditions. Since opiorphin may also reflect emotional and socio-relational imbalances occurring with BMS, it could as well represent a biomarker for BMS. Knowledge on opiorphin\'s involvement in pain pathways could contribute to developing new clinical diagnostic methods for BMS.

OBJECTIVE: Idiopathic Burning mouth syndrome (iBMS) is a poorly understood affection characterized by persistent pain in the oral cavity without any clinical or biological abnormality. Opiorphin is a natural inhibitor of enkephalin-inactivating ectopeptidases, mainly produced by salivary glands, that has demonstrated analgesic properties. The objective of the present case-control study was to test the hypothesis of a decrease in opiorphin levels in iBMS patients.
METHODS: Twenty-one iBMS patients and 21 matched controls subjects were included between 2011 and 2013. Submandibular and sublingual salivary, blood, and urinary opiorphin levels of iBMS patients were compared to controls.
RESULTS: Results are expressed as mean values ± SD and compared using the Wilcoxon Signed Rank test. Correlations were analyzed with Spearman coefficient. The level of significance was fixed at p < 0.05. Opiorphin levels in iBMS and controls were respectively (in ng/ml) in basal saliva: 37.8 ± 42.5 and 67.6 ± 188.9 (p = NS); stimulated saliva: 28.8 ± 25.3 and 31.1 ± 29.1 (p = NS); blood: 4.6 ± 5.4 and 1.9 ± 1.4 (p < 0.05); and urines: 68.5 ± 259.8 and 8.9 ± 6.2 (p = NS).
CONCLUSIONS: In conclusion, the lack of significative difference in salivary opiorphin levels between iBMS and controls does not favor a direct local role for opiorphin in the etiopathogeny of iBMS. However, higher blood opiorphin levels may reflect a systemic dysregulation in iBMS. Trial registration NCT02686359 https://clinicaltrials.gov/ct2/show/NCT02686359.

This study aimed to compare the biochemical composition of saliva from patients with eating disorders (EDs) with saliva from control subjects with no ED. All patients who initiated outpatient treatment in an ED clinic during a 12-month period were invited to participate. Of the 65 patients who started treatment during the period, 54 (50 female patients/four male patients; mean age: 21.5 yr) agreed to participate. The controls were 54 sex- and age-matched patients from a dental health clinic. All participants completed a questionnaire and underwent dental clinical examinations, including laboratory analyses of saliva. The proportion of subjects with unstimulated salivary hyposalivation was lower in the ED group and not correlated with intake of xerogenic drugs. Significant differences in the biochemical composition of saliva were found almost exclusively in the unstimulated state, with albumin, inorganic phosphate, aspartate aminotransferase (ASAT), chloride, magnesium, and total protein all being significantly higher in the ED group. Conditional logistic regression showed that higher ASAT and total protein concentrations were relatively good predictors of ED, with sensitivity and specificity of 65% and 67%, respectively. In conclusion, elevated salivary concentrations of ASAT and total protein may serve as indicators of ED as well as of disease severity. Future studies are needed to corroborate these initial findings.

BACKGROUND: Current diagnostic criteria for Sjogren\'s syndrome developed by the American College of Rheumatology include the presence of antinuclear antibodies, rheumatoid factor, anti-Ro or anti-La autoantibodies. The purpose of this report is to describe two patients with biopsy-proven Sjogren\'s syndrome lacking these autoantibodies but identified by antibodies to salivary gland protein 1. Diagnosis was delayed until salivary gland tumors developed in these patients because of the lack of the classic autoantibodies. This report emphasizes the existence of patients with primary Sjogren\'s syndrome who lack autoantibodies anti-Ro or anti-La and may therefore be misdiagnosed. Antibodies to salivary gland protein 1 identify some of these patients.
METHODS: Two patients are described and were seen in the autoimmune disease clinics of the State University of New York (SUNY) at the Buffalo School of Medicine. In both patients, chronic dry mouth and dry eye had been dismissed as idiopathic because test results for autoantibodies anti-Ro and anti-La were negative. Both patients had swelling of major salivary glands that prompted biopsies. Biopsies of major salivary glands from both cases demonstrated salivary gland tumors and existence of inflammation consistent with Sjogren\'s syndrome. Serologic testing revealed antibodies to salivary gland protein 1.
CONCLUSIONS: Patients presenting with classic clinical symptoms of dry mouth and eyes do not always show the current serologic markers of Sjogren\'s syndrome, anti-Ro and anti-La. In these cases, investigation for antibodies to salivary gland protein 1 is of importance to make the diagnosis of Sjogren\'s syndrome. Early diagnosis of Sjogren\'s syndrome is necessary for improved management as well as for vigilance regarding potential complications, such as salivary gland tumors as were seen in the described cases.

The acid-base behavior of amino acids is an important subject of study due to their prominent role in enzyme catalysis, substrate binding and protein structure. Due to interactions with the protein environment, their pK(a)s can be shifted from their solution values and, if a protein has two stable conformations, it is possible for a residue to have different \"microscopic\", conformation-dependent pK(a) values. In those cases, interpretation of experimental measurements of the pK(a) is complicated by the coupling between pH, protonation state and protein conformation. We explored these issues using Nitrophorin 4 (NP4), a protein that releases NO in a pH sensitive manner. At pH 5.5 NP4 is in a closed conformation where NO is tightly bound, while at pH 7.5 Asp30 becomes deprotonated, causing the conformation to change to an open state from which NO can easily escape. Using constant pH molecular dynamics we found two distinct microscopic Asp30 pK(a)s: 8.5 in the closed structure and 4.3 in the open structure. Using a four-state model, we then related the obtained microscopic values to the experimentally observed \"apparent\" pK(a), obtaining a value of 6.5, in excellent agreement with experimental data. This value must be interpreted as the pH at which the closed to open population transition takes place. More generally, our results show that it is possible to relate microscopic structure dependent pKa values to experimentally observed ensemble dependent apparent pK(a)s and that the insight gained in the relatively simple case of NP4 can be useful in several more complex cases involving a pH dependent transition, of great biochemical interest.

As noninvasively accessible body fluid, saliva is of growing interest in diagnostics. To exemplify the diagnostic potential of saliva, we used a mass spectrometry-based approach to gain insights into adaptive physiological processes underlying long-lasting endurance work load in a case study. Saliva was collected from male and female athlete at four diurnal time points throughout a 1060 km nonstop cycling event. Total sampling time covered 180 h comprising 62 h of endurance cycling as well as reference samples taken over 3 days before the event, and over 2 days after. Altogether, 1405 proteins and 62 metabolites were identified in these saliva samples, of which 203 could be quantified across the majority of the sampling time points. Many proteins show clear diurnal abundance patterns in saliva. In many cases, these patterns were disturbed and altered by the long-term endurance stress. During the stress phase, metabolites of energy mobilization, such as creatinine and glucose were of high abundance, as well as metabolites with antioxidant functions. Lysozyme, amylase, and proteins with redox-regulatory function showed significant increase in average abundance during work phase compared to rest or recovery phase. The recovery phase was characterized by an increased abundance of immunoglobulins. Our work exemplifies the application of high-throughput technologies to understand adaptive processes in human physiology.

BACKGROUND: In the last years human proteomic has represented a promising tool to promote the communication between basic and clinical science.
METHODS: To explore the correspondence between salivary proteomic profile and clinical response, herein, we used a proteomic approach to analyse the whole saliva of a patient with primary Sjögren\'s Syndrome (pSS) and non-Hodgkin\'s-MALT type parotid lymphoma before, during and after a standard treatment with cyclophosphamide (CTX) and rituximab (RTX). To identify any discriminatory therapeutic salivary biomarker patient\'s whole saliva was collected at the baseline, after the fourth infusion of rituximab, and on remission and analysed combining two-dimensional electrophoresis (2DE) and MALDI-TOF/TOF mass spectrometry.
RESULTS: Proteomic results obtained from the comparison of salivary samples indicated several qualitative and quantitative modifications in the salivary expression of putative albumin, immunoglobulin J chain, Ig kappa chain C region, alpha-1-antitrypsin, haptoglobin and Ig alpha-1 chain C region.
CONCLUSIONS: This study suggests that clinical and functional changes of the salivary glands driven by autoimmune and lymphoproliferative processes might be reflected in patients\' whole saliva proteins, shedding new light on the potential usefulness of salivary proteomic analysis in the identification of prognostic and therapeutic biomarkers for patients with pSS and non Hodgkin\'s lymphomas.