Profile Of Saliva Research Articles

Biopolymer-based oral films integrated with probiotic active compounds for improved health applications.

Orally dissolving films (ODFs) have emerged as a versatile platform that combines convenience, efficacy, and patient compliance. In this study, the cell-free supernatant of the oral probiotic Streptococcus salivarius M18 was incorporated into various biopolymer-based ODF formulations, evaluated for demolding, fragility, and flexibility. The combination of carboxymethyl cellulose, sodium alginate, and glycerol successfully formed stable films. The films were characterized by weight, thickness, pH, and disintegration times. Fourier-transform infrared spectroscopy (FTIR) was used to analyze ODF content and release profiles in simulated saliva. Unique absorption peaks in the cell-free product-incorporated ODF samples confirmed the integration of bacterial proteins, lipids, and nucleic acids into the ODF matrix. The biological activity of the ODF carrying M18 bioactive products was assessed by its inhibitory effect on the growth of Streptococcus mutans, a pathogen linked to dental plaque and cavities. Additionally, the anti-proliferative effect on cancer epithelial cells was demonstrated. This study show that probiotic products can be integrated into bio-based thin films without losing activity, making this delivery platform promising for local and potentially systemic effects.

8665 Female Fertility In Patients With Classic And Non-Classic Congenital Adrenal Hyperplasia Under Modified-Release Hydrocortisone (MR-HC)

Abstract Disclosure: L. Tschaidse: None. H.F. Nowotny: None. M. Auer: None. C. Lottspeich: None. M. Bidlingmaier: None. J. Hawley: None. B.G. Keevil: None. N. Reisch: None. Background: Female patients with classic and non-classic (NC) CAH struggle with fertility, due to androgen excess, elevated progesterone and 17-hydroxyprogesterone (17OHP) levels, causing anovulation, impairment of endometrial development and menstrual irregularities. The typical approach to treating infertility often includes an increase in glucocorticoid (GC) dose. However, this poses a potential risk of side effects. Limited observational data on modified-release hydrocortisone (MR-HC), which better mimics the physiological diurnal rhythm of cortisol secretion, indicates improved fertility in female and male patients with classic CAH. Therefore, the aim of this study was to investigate fertility in women with classic and non-classic CAH before and after switching to MR-HC. Methods: 21 adult female patients with CAH (14 classic; 5 NC) with a median (range) age of 33.0 (30.0) years, premenopausal and without hormonal contraception, were enrolled in this prospective, observational, single-centre study. The data collection took place before (conventional GC preparation) and after dose-equivalent switching the medication to MR-HC. In addition to clinical parameters such as menstrual cycle and desire to conceive, the collection of morning serum and daily saliva profiles was conducted, measuring progesterone, 17OHP, androstenedione (A4), testosterone, 11-ketotestosterone (11KT) as well as 11-hydroxyandrostenedione (11OHA4). Results: Before study inclusion, 4/21 patients had an irregular cycle, 5/21 were amenorrhoeic and 10/21 patients had a desire to have children. There was no difference in median (range) hydrocortisone dose equivalent before and after switching to MR-HC (25.0 (50.0) vs. 25.0 (30.0), p=.441). Median (range) morning serum concentrations between 8-10 am were not different before and after switch to MR-HC for progesterone (5.9 (55.5) vs. 5.0 (58.8), p=.823), 17OHP (5.9 (362.1) vs. 5.7 (73.7), p=.280), A4 (2.9 (25.6) vs. 2.5 (6.9), p=.106), testosterone (0.8 (3.5) vs. 0.7 (1.5), p=.063), 11KT (0.4 (7.4) vs. 0.2 (2.6), p=.242) and 11OHA4 (0.8 (12.9) vs. 0.9 (10.4), p=.231). Concerning saliva day profiles, we found a significantly lower median (range) 17OHP level in early morning saliva (43.0 (526.9) vs. 19.4 (264.3), p=.049). 5/9 patients with menstrual irregularities reported menstrual regularisation within and 5/10 patients with a desire to have children became pregnant after switching to MR-HC. Conclusion: Our preliminary data show clinical improvement in fecundity and fertility under MR-HC in both women with classic and NC CAH without a dose increase of glucocorticoids. Under MR-HC replacement lower 17OHP concentrations in early morning saliva reflect better hormonal control over night contributing to ovulatory cycle regulation and improved fertility. Presentation: 6/1/2024

Deciphering disease through glycan codes: leveraging lectin microarrays for clinical insights.

Glycosylation, a crucial posttranslational modification, plays a significant role in numerous physiological and pathological processes. Lectin microarrays, which leverage the high specificity of lectins for sugar binding, are ideally suited for profiling the glycan spectra of diverse and complex biological samples. In this review, we explore the evolution of lectin detection technologies, as well as the applications and challenges of lectin microarrays in analyzing the glycome profiles of various clinical samples, including serum, saliva, tissues, sperm, and urine. This review not only emphasizes significant advancements in the high-throughput analysis of polysaccharides but also provides insight into the potential of lectin microarrays for diagnosing and managing diseases such as tumors, autoimmune diseases, and chronic inflammation. We aim to provide a clear, concise, and comprehensive overview of the use of lectin microarrays in clinical settings, thereby assisting researchers in conducting clinical studies in glycobiology.

Free Salivary Amino Acid Profile in Breast Cancer: Clinicopathological and Molecular Biological Features.

The study of salivary amino acid profiles has attracted the attention of researchers, since amino acids are actively involved in most metabolic processes, including breast cancer. In this study, we analyzed the amino acid profile of saliva in a sample including all molecular biological subtypes of breast cancer to obtain a more complete picture and evaluate the potential utility of individual amino acids or their combinations for diagnostic purposes. This study included 116 patients with breast cancer, 24 patients with benign breast disease, and 25 healthy controls. From all patients, strictly before the start of treatment, saliva samples were collected, and the quantitative content of 26 amino acids was determined. Statistically significant differences between the three groups are shown in the content of Asp, Gly, Leu + Ile, Orn, Phe, Pro, Thr, and Tyr. To differentiate the three groups from each other, a decision tree was built. To construct it, we selected those amino acids for which the change in concentrations in the subgroups was multidirectional (GABA, Hyl, Arg, His, Pro, and Car). For the first time, it is shown that the amino acid profile of saliva depends on the molecular biological subtype of breast cancer. The most significant differences are shown for the luminal B HER2-positive and TNBC subgroups. In our opinion, it is critically important to consider the molecular biological subtype of breast cancer when searching for potential diagnostic markers.

Comprehensive mapping of saliva by multiomics in children with idiopathic nephrotic syndrome.

Saliva can reflect an individual's physiological status or susceptibility to systemic disease. However, little attention has been given to salivary analysis in children with idiopathic nephrotic syndrome (INS). We aimed to perform a comprehensive analysis of saliva from INS children. A total of 18 children (9 children with INS and 9 normal controls) were recruited. Saliva was collected from each INS patient in the acute and remission phases. 16S rRNA gene sequencing, widely targeted metabolomics, and 4D-DIA proteomics were performed. Actinobacteria and Firmicutes were significantly enriched in the pretreatment group compared with the normal control group, while Bacteroidota and Proteobacteria were significantly decreased. A total of 146 metabolites were identified as significantly different between INS children before treatment and normal controls, which covers 17 of 23 categories. KEGG enrichment analysis revealed three significantly enriched pathways, including ascorbate and aldarate metabolism, pentose and glucuronate interconversions, and terpenoid backbone biosynthesis (P < 0.05). A total of 389 differentially expressed proteins were selected between INS children before treatment and normal controls. According to the KEGG and GO enrichment analyses of the KOGs, abnormal ribosome structure and function and humoral immune disorders were the most prominent differences between INS patients and normal controls in the proteomic analysis. Oral microbiota dysbiosis may modulate the metabolic profile of saliva in children with INS. It is hypothesized that children with INS might have "abnormal ribosome structure and function" and "humoral immune disorders".

Oral and gut microbial profiling in periodontitis and Parkinson’s disease

ABSTRACT Objectives We tested the hypothesis that Parkinson’s disease (PA) alters the periodontitis-associated oral microbiome. Method Patients with periodontitis with Parkinson’s disease (PA+P) and without PA (P) and systemically and periodontally healthy individuals (HC) were enrolled. Clinical, periodontal and neurological parameters were recorded. The severity of PA motor functions was measured. Unstimulated saliva samples and stool samples were collected. Next-generation sequencing of 16S ribosomal RNA (V1-V3 regions) was performed. Results PA patients had mild-to-moderate motor dysfunction and comparable plaque scores as those without, indicating that oral hygiene was efficient in the PA+P group. In saliva, there were statistically significant differences in beta diversity between HC and PA+P (p = 0.001), HC and P (p = 0.001), and P and PA+P (p = 0.028). The microbial profiles of saliva and fecal samples were distinct. Mycoplasma faucium, Tannerella forsythia, Parvimonas micra, and Saccharibacteria (TM7) were increased in P; Prevotella pallens, Prevotella melaninogenica, Neisseria multispecies were more abundant in PA+P group, Ruthenibacterium lactatiformans, Dialister succinatiphilus, Butyrivibrio crossotus and Alloprevotella tannerae were detected in fecal samples in P groups compared to healthy controls. Conclusions No significant differences were detected between Parkinson’s and non-Parkinson’s gut microbiomes, suggesting that Parkinson’s disease modifies the oral microbiome in periodontitis subjects independent of the gut microbiome.

Changes in saliva protein profile throughout Rhipicephalus microplus blood feeding

BackgroundWhen feeding on a vertebrate host, ticks secrete saliva, which is a complex mixture of proteins, lipids, and other molecules. Tick saliva assists the vector in modulating host hemostasis, immunity, and tissue repair mechanisms. While helping the vector to feed, its saliva modifies the site where pathogens are inoculated and often facilitates the infection process. The objective of this study is to uncover the variation in protein composition of Rhipicephalus microplus saliva during blood feeding.MethodsTicks were fed on calves, and adult females were collected, weighed, and divided in nine weight groups, representing the slow and rapid feeding phases of blood feeding. Tick saliva was collected, and mass spectrometry analyses were used to identify differentially secreted proteins. Bioinformatic tools were employed to predict the structural and functional features of the salivary proteins. Reciprocal best hit analyses were used to identify conserved families of salivary proteins secreted by other tick species.ResultsChanges in the protein secretion profiles of R. microplus adult female saliva during the blood feeding were observed, characterizing the phenomenon known as “sialome switching.” This observation validates the idea that the switch in protein expression may serve as a mechanism for evading host responses against tick feeding. Cattle tick saliva is predominantly rich in heme-binding proteins, secreted conserved proteins, lipocalins, and protease inhibitors, many of which are conserved and present in the saliva of other tick species. Additionally, another remarkable observation was the identification of host-derived proteins as a component of tick saliva.ConclusionsOverall, this study brings new insights to understanding the dynamics of the proteomic profile of tick saliva, which is an important component of tick feeding biology. The results presented here, along with the disclosed sequences, contribute to our understanding of tick feeding biology and might aid in the identification of new targets for the development of novel anti-tick methods.Graphical

Effect of &lt;i&gt;Trivrit Virechana, Yashtyadi Pratisarana&lt;/i&gt; and &lt;i&gt;Panchaksheeri Vriksha Kashaya Gandoosha&lt;/i&gt; in Periodontitis — A Case Series

Periodontitis is a localized inflammation of the teeth most commonly caused by a bacterial infection in the periodontal cavity. The disease is common in adults over the age of 30. The bacteria involved in periodontal disease are mainly Gram-negative anaerobes. According to Ayurveda, the specific symptoms of upakusha have similarities with periodontitis. The objective of this study was to evaluate the combined effect of virechana, pratisarana and gandusha in improving symptoms, gingival bleeding index, plaque index, and restoring normal salivary microbial profiles in patients with periodontitis. A group of 30 patients who met the inclusion criteria were recruited for the study. Each case was observed for prevalence by age, pain, gingival bleeding, halitosis, plaque index, gingival bleeding index. In the results, it was found that Trivrit virechana, Yashtyadi choorna prathisarana and Panchaksheerivriksha kashya gandusha showed significant reduction in pain, bleeding gums, halitosis, plaque index, bleeding gums and normalization of saliva pH and recovered the bacterial profile in saliva with an improvement of 100%, 100%, 98%, 100% and 100% respectively and was statistically significant with p&lt;0.0001. So, Trivrit virechana purifies the koshta in turn keeps the oral cavity healthy and strengthen the periodontium. Yashtyadi choorna havs anti-inflammatory, antibacterial property inhibits the growth of periodontopathogen and Panchaksheerivriksha kashaya helps to normalize the pH of saliva and reduce the microbial load. Hence, Ayurveda management can be considered as a better alternative in the management of periodontitis.

Cytokine profile of oral fluid in patients with food allergy associated with atopic dermatitis and psoriasis

The world is experiencing a rapid increase in the prevalence of allergic and autoimmune diseases. It is known that allergic inflammation is most often systemic, involving various organs and systems in the pathological process, such as the skin, respiratory and gastrointestinal tract with the development of dermatorespiratory, dermato-intestinal and other manifestations. The study of the features of the cytokine profile in oral fluid (saliva) deserves special attention, since these characteristics reflect not only local, but also systemic disorders. Of particular relevance is the study of local cytokine regulation of intercellular interactions in food allergies. Our objective was to study the concentration of IL-4, IL-10, IFNγ, secretory IgA in salivary fluid, the concentrations of total immunoglobulin E and eosinophilic cationic protein in blood serum of the patients with atopic dermatitis and psoriasis with concomitant food allergies.The study included patients with atopic dermatitis (AD, group 1, n = 20), psoriasis with concomitant food allergy (PS, group 2, n = 27), psoriasis without concomitant allergies (PS, comparison group 3, n = 23). Quantitative assessment of the cytokine concentrations (IL-4, IL-10, IFNγ, sIgA) in salivary fluid was carried out by enzyme-linked immunosorbent assay. Concentrations of total immunoglobulin E and eosinophilic cationic protein in blood serum were determined by indirect immunofluorescence. The obtained results were processed using the Statistica 8.0 applied software.In groups of patients with atopic dermatitis (Group 1) and psoriasis with concomitant food allergy (Group 2), we have noted a statistically significant increase of salivary IL-4 and IL-10, as well as of total immunoglobulin E concentrations in blood serum as compared with a group of patients with psoriasis without concomitant allergies (group 3), and with control group. When studying concentrations of IFNγ in saliva, no statistically significant intergroup differences were found. The concentration of sIgA in saliva was significantly higher in the groups of patients with atopic dermatitis and psoriasis accompanied by food allergies in comparison with control group and the group of psoriatic patients without food allergies (group 3).The cytokine profile of saliva is characterized by unidirectional changes in food allergy. Skin seems to be the shock organ in this condition, regardless of nosological form of the disease (atopic dermatitis or psoriasis). Salivary fluid is an easily accessible material when assessing the state of mucosal immunity in food allergies.

SAT334 Hypoxic Reduction Of Adrenal Steroidogenesis- The Cause Of Idiopathic Allergic Reactions

Abstract Disclosure: S. Kalinchenko: None. L. Vorslov: None. D. Gusakova: None. O. Samburskaya: None. Hypoxic reduction of adrenal steroidogenesis - The Cause of Idiopathic Allergic Reactions Background: This thesis discusses the occurrence of idiopathic allergic reactions, with an established absence of allergy to any trigger by allergo-tests, with the sole presence of a history of decompensated chronic stress, which led to a stress-induced decrease in adrenal steroidogenesis called “adrenal fatigue” with decreased production of the main anti-inflammatory, anti-allergic and stress hormone, cortisol. Materials and methods: The study included 10 patients aged 20 to 55 years, 8 women and 2 men, diagnosed with idiopathic allergic reactions (urticaria, toxidermia, erythema multiforme exudative, Quincke's edema). All patients were examined for steroid hormone levels using a steroid profile in saliva by mass spectrometry and performing allergy tests (to identify a possible allergen). Findings: From the anamnesis, all 10 patients had long-term chronic stress, which was already in the stage of decompensation, and according to the Beck depression scale, 8 patients showed mild (11-17 points) and medium (18-23 points) severity of depression in 2 patients. In the steroid profile of saliva in all 10 patients, a decrease below 07\ры the reference level of cortisol (700-2200 pg/ml) and cortisone (27400-39200 pg/ml) was found. After relief of acute symptoms by corticosteroids, all patients were prescribed infusion therapy with ascorbic acid at a dosage of 1,000 mg three times a week for three weeks to stimulate the adrenal glands. As a result, repeated analysis of the steroid profile of saliva showed an increase in the level of cortisol and cortisone within the reference values.Allergotests are negative for all known allergens. Interpretation: As is known, allergens play a primary role in the development of diseases such as urticaria, toxidermia, erythema multiforme, Quincke's edema. However, there are increasingly cases when it is not possible to establish the trigger that caused the development of an allergic reaction, and conducting allergy tests does not reveal the presence of allergen triggers in patients. However, these patients are united by a decrease in adrenal steroidogenesis against the background of decompensated chronic stress called “adrenal fatigue”, which manifests itself in a decrease in the level of cortisol and cortisone in the steroid profile of saliva below the reference values, which are the main anti-inflammatory, antiallergic hormones and hormones that are synthesized in response to stress, and with chronic stress, the reserves of these hormones are depleted, which provokes the development of inflammatory and allergic reactions in response to ordinary stimuli. Conclusion: Stimulation of the adrenal glands with reduced steroidogenesis is an opportunity to reduce the risk of inflammatory and allergic reactions in patients with decompensated chronic stress. Presentation: Saturday, June 17, 2023

Metabolomic profiling of amino acids study reveals a distinct diagnostic model for diabetic kidney disease

Diabetic kidney disease (DKD), a highly prevalent complication of diabetes mellitus, is a major cause of mortality in patients. However, identifying circulatory markers to diagnose DKD requires a thorough understanding of the metabolic mechanisms of DKD. In this study, we performed ultra-performance liquid chromatography tandem mass spectrometry (UPLC-MS/MS) to reveal altered metabolic profiles of amino acids (AAs) in patients with DKD. We found decreased plasma levels of histidine and valine, increased urine levels of proline, decreased urine levels of histidine and valine, and increased saliva levels of arginine in patients with DKD compared with the levels in patients with type 2 diabetes mellitus (T2DM) and in healthy controls. Our analyses of the key metabolites and metabolic enzymes involved in histidine and valine metabolism indicated that the AAs level alterations may be due to enhanced carnosine hydrolysis, decreased degradation of homocarnosine and anserine, enhanced histidine methylation, and systemic enhancement of valine metabolism in patients with DKD. Notably, we generated a distinct diagnostic model with an AUC of 0.957 and an accuracy up to 92.2% on the basis of the AA profiles in plasma, urine and saliva differing in patients with DKD using logistic regression and receiver operating characteristic analyses. In conclusion, our results suggest that altered AA metabolic profiles are associated with the progression of DKD. Our DKD diagnostic model on the basis of AA levels in plasma, urine, and saliva may provide a theoretical basis for innovative strategies to diagnose DKD that may replace cumbersome kidney biopsies.

Investigating oral biofluid markers as a tool for the chemometric discrimination of different physical exercise intensities utilizing ATR-FTIR spectroscopy

Physical and mental capacity of a person to perform sport activities is affected by exercise intensity, especially noted in athletes who push themselves to the limits of endurance. The present study investigated variations of oral biofluid samples (saliva) during different physical exercise intensities utilizing Attenuated Total Reflection-Fourier Transform Infrared (ATR-FTIR) spectroscopy coupled with multivariate analysis. Samples were collected from 32 high-level athletes at i) rest and after different exercise intensities: ii) walking (4–5 km/h), iii) jogging (9–10 km/h) and iv) running (14–15 km/h). PLS-DA model with ten components succeed to classify spectral profile of saliva in the four (i-iv) different groups with an accuracy of 74.3%. The observed discrimination appears to be attributed to the presence of infrared vibrations related to thiocyanate, phospholipids, lactate, phosphate, and glucose. Second derivative ATR-FTIR spectroscopy was utilized to evaluate biomarkers adaptations during the different exercise conditions, and thiocyanate vibration of ATR-FTIR spectra was investigated as a novel index associated with physical exercise intensity. According to our research, monitoring a biochemical change using ATR-FTIR focusing on thiocyanate could serve as a promising supplementary indicator for non-invasively monitoring athletes' training.

CATION COMPOSITION OF ORAL SECRETION IN PRIMARY SCHOOL-AGED CHILDREN

SUMMARY. Recurrent respiratory infections (RRIs) currently have the greatest medical and social burden among infectious diseases worldwide, with the highest prevalence and incidence rates of acute respiratory infections in children. Recent studies indicate the role of inadequate immune response and the development of recurrent acute respiratory infections (ARI) in children due to deficiencies in essential trace elements in the body, which are crucial for the proper functioning of physical barriers and the immune system. The hypothesis of our study assumed that, considering the recurrence of at least 6 episodes of ARI per year, children with RRIs may have changes in salivary ion levels, which could serve as a basis for local immune suppression of the upper respiratory mucosa. The aim – to investigate the levels of metals in the saliva of children of primary school age with recurrent respiratory infections in comparison with episodically ill peers and establish the relationship of these changes with the state of local immunity. Material and Methods. In the actual study, 40 children participated, with 30 of them (the main group) being children with recurrent respiratory infections and 10 other children who had episodic illnesses were assigned to the control group. The levels of metal cations in the saliva were examined using inductively coupled plasma atomic emission spectrometry, including essential metals such as copper, manganese, zinc, calcium, cobalt, potassium, magnesium, sodium, phosphorus, selenium, iron, chromium, and sulfur. Additionally, levels of conditionally essential metals like lithium, nickel, and boron were measured. The levels of secretory IgA and lysozyme in saliva were determined using the enzyme-linked immunosorbent assay method. Results. The main group was not homogeneous; therefore, we divided it into two clusters. Respondents in one cluster had significantly lower relative levels of essential ion content in saliva. Specifically, calcium was lower in 64 % (p=0.003), copper in 76.5 % (p=0.001), manganese in 41.1 % (p=0.008), magnesium in 64 % (p=0.02), and zinc in 58.8 % (p=0.027). This fact can be explained by their increased losses due to frequent inflammatory processes in the upper respiratory tract, resulting in higher daily replenishment needs compared to their peers. Correlation analysis revealed a connection between illness incidence and the levels of copper (p=0.008), sodium (p=0.00005), and phosphorus (p=0.028), indicating potential associations between saliva metal profiles and the frequency of ARI in children with RRIs. The presence of dental caries correlated with lithium (p=0.02), copper (p=0.032), and nickel levels (p=0.037). Integral indicators of local immunity, such as lysozyme and secretory IgA, exhibited correlations with metal levels in saliva (Li, Cu, Ni, Na, Co, P, Zn), confirming the hypothesis of their role in regulating immune reactions in the oral cavity. Conclusions. Cluster analysis of the metal profiles of the group of children with RRI shows its heterogeneity, 43 % of representatives of this cohort had significantly lower levels of calcium (p=0.003), copper (p=0.001), manganese (p=0.008), magnesium (p=0.02) and zinc (p=0.027) than the control group. Which may indicate the presence of deficient conditions in some children with PRI as a result of the increased need for their recovery. The presence of caries, the level of SIgA, lysozyme (clinical indicators of local immunity of the oral cavity) have certain relationships with the levels of essential and conditionally essential levels of salivary metals (Li, Cu, Ni, Na, Co, P, Zn), which indicates the benefit of the connection between the metal profile of saliva and the state of local immunity.

Metabolic profiles of saliva in male mouse models of chronic sleep disorders induced by psychophysiological stress

Disordered sleep is a global social problem and an established significant risk factor for psychological and metabolic diseases. We profiled non-targeted metabolites in saliva from mouse models of chronic sleep disorder (CSD). We identified 288 and 55 metabolites using CE-FTMS and LC-TOFMS, respectively, among which concentrations of 58 (CE-FTMS) and three (LC-TOFMS) were significantly changed by CSD. Pathway analysis revealed that CSD significantly suppressed glycine, serine and threonine metabolism. Arginine and proline metabolic pathways were among those that were both upregulated and downregulated. Pathways of alanine, aspartate and glutamate metabolism, genetic information processing, and the TCA cycle tended to be downregulated, whereas histidine metabolism tended to be upregulated in mice with CSD. Pyruvate, lactate, malate, succinate and the glycemic amino acids alanine, glycine, methionine, proline, and threonine were significantly decreased, whereas 3-hydroxybutyric and 2-hydroxybutyric acids associated with ketosis were significantly increased, suggesting abnormal glucose metabolism in mice with CSD. Increases in the metabolites histamine and kynurenic acid that are associated with the central nervous system- and decreased glycine, might be associated with sleep dysregulation and impaired cognitive dysfunction in mice with CSD. Our findings suggested that profiling salivary metabolites could be a useful strategy for diagnosing CSD.

Characteristics of Human and Microbiome RNA Profiles in Saliva

ABSTRACT Saliva is a convenient non-invasive source of liquid biopsy to monitor human health and diagnose diseases. In particular, extracellular vesicles (EVs) in saliva can potentially reveal clinically relevant information for systemic health. Recent studies have shown that RNA in saliva EVs could be exploited as biomarkers for disease diagnosis. However, there is no standardized protocol for profiling RNA in saliva EV nor clear guideline on selecting saliva fractions for biomarker analysis. To address these issues, we established a robust protocol for small RNA profiling from fractionated saliva. With this method, we performed comprehensive small RNA sequencing of four saliva fractions, including cell-free saliva (CFS), EV-depleted saliva (EV-D), exosome (EXO), and microvesicle (MV) from ten healthy volunteers. By comparing the expression profiles of total RNA from these fractions, we found that MV was most enriched in microbiome RNA (76.2% of total reads on average), whereas EV-D was notably enriched in human RNA (70.3% of total reads on average). As for human RNA composition, CFS and EV-D were both enriched in snoRNA and tRNA compared with the two EV fractions (EXO and MV, P < 0.05). Interestingly, EXO and MV had highly correlated expression profiles for various noncoding RNAs such as miRNA, tRNA, and yRNA. Our study revealed unique characteristics of circulating RNAs in various saliva fractions, which provides a guideline on preparing saliva samples to study specific RNA biomarkers of interest.

Salivary Antioxidant Profile in Patients with Systemic Sclerosis and Periodontitis.

The aim of the present study was to compare periodontal status and antioxidant profile in unstimulated saliva of systemic sclerosis (SSc) patients with periodontitis and systemically healthy periodontitis patients. Twenty patients with established diagnoses of systemic sclerosis and periodontitis (SSc group) and 20 systemically healthy individuals with periodontitis (P group) were enrolled in the study. Clinical periodontal parameters (clinical attachment level (CAL), gingival recession (GR), periodontal probing depth (PPD), and gingival index (GI)) and concentration of uric acid (UA), superoxide dismutase (SOD), and glutathione peroxidase (GPX) in unstimulated saliva samples were assessed. There were significantly higher mean values of CAL (4.8 ± 0.21 mm versus 3.18 ± 0.17 mm; p ≤ 0.001) and GR (1.66 ± 0.90 mm versus 0.46 ± 0.54 mm; p ≤ 0.001) in the SSc group when compared to the P group. Significantly higher level of GPX (p ≤ 0.001) and SOD (p ≤ 0.001) in unstimulated saliva was detected in the SSc group in comparison with the P group. The specific activity of UA did not significantly differ between the two groups (p = 0.083). The results may indicate higher periodontal destruction and antioxidant perturbations in unstimulated saliva of SSc patients with periodontitis compared to systemically healthy periodontitis patients.

Correlation between caries activity and salivary microbiota in preschool children.

Early childhood caries (ECC) is the most common chronic infectious oral disease in preschool children worldwide. It is closely related to the caries activity (CA) of children. However, the distribution characteristics of oral saliva microbiomes in children with different CA are largely underexplored. The aim of this study was to investigate the microbial community in saliva of preschool children with different CA and caries status, and to analyze the difference of microbial community in saliva of children with different CA and its correlation with ECC. Subjects were divided into 3 groups based on the Cariostat caries activity test: Group H, high CA (n=30); Group M, medium CA (n = 30); Group L, low CA (n=30). Questionnaire survey was used to explore the related influencing factors of CA. According to the caries status (on the basis of decayed mising filled teeth), these subjects were divided into caries-free group (dmft=0, n=19), caries-low group (0 < dmft ≤ 4, n=27) and caries-high group (dmft > 4, n=44). Microbial profiles of oral saliva were analyzed using 16S rRNA gene sequencing. There were significant differences in the microbial structure (P < 0.05). Scardovia and Selenomonas were the biomarkers of both H group and high caries group. The genus Abiotrophia and Lautropia were the biomarkers of both the L group and the low caries group, while the Lactobacillus and Arthrospira spp. were significantly enriched in the M group. The area under the ROC curve of the combined application of dmft score, age, frequency of sugary beverage intake, and the genus Scardovia, Selenomonas, and Campylobacter in screening children with high CA was 0.842. Moreover, function prediction using the MetaCyc database showed that there were significant differences in 11 metabolic pathways of salivary microbiota among different CA groups. Certain bacteria genera in saliva such as Scardovia and Selenomonas may be helpful in screening children with high CA.

Abstract 6528: HPV status and epigenetic profiling of saliva from a cancer-free population

Abstract Identifying environmental factors and their relationship with epigenetic alterations is essential for understanding cancer, a leading cause of death. Currently, cancer screening and early detection development efforts are ongoing and epigenetics-based tests are one of the top choices as strategies. The understanding of how these alterations behave in cancer-free (CF) populations with well annotated data is essential. We randomly selected 601 saliva specimens from two Brazilian countrywide epidemiological studies POP-Brazil and SMESH. Extracted DNAs were sodium bisulfite converted and analyzed by Quantitative Methylation-Specific PCR (QMSP). Five genes previously reported as DNA methylation prone in cancer were selected (ITGA4, EDRNB, PAX5, p16, CCNA1). All participants were interviewed and provided sociodemographic and sexual behavior information as well as genital (gn) and oral biological samples. Variables were categorized and Fisher’s exact test was performed using SAS 9.4. Surprisingly, methylation in 3 cancer associated genes showed considerable frequencies in this CF group. DNA methylation was observed for EDNRB in 75/601 samples (12.5%), PAX5 in 55/601 (9.2%), and ITGA4 in 20/601 (3.3%). 12 samples displayed concomitant methylation on 2 of these 3 genes. p16 and CCNA1 methylation were only present in 1 sample each. Preliminary analyses suggest that specific genes methylation are observed with the presence of low (LR) and high-risk (HR) HPV types. EDNRB methylation groups with the presence of HR HPV type 16, 31, 39, 51, 58 and 59 from gn samples and LR types 6 and 42. ITGA4 methylation and HR HPV types 16 and 59 were detected in oral samples, and types 52, 56 and 68 in gn samples, as well as LR HPV types 40 and 42. In people with early onset of smoking, EDNRB and PAX5 seem to be more frequently methylated. The positivity of PAX5 was higher in participants who smoke, use alcohol and presented a positive diagnosis of syphilis and/or HIV. PAX5 methylation was more frequent in HR HPV 33, 58, 68 and 62 and LR type 6. Further analyses are ongoing. Our study shows methylation in cancer related genes in a CF population. The variables collected include some proven cancer risk factors and others still unknown, thus lead us to uncover directions to identify main cancer related players in a Brazilian population. The presence of cancer related alterations may indicate premalignant stages, exposure and lifestyle consequences, both at the same time or separately. In our pilot analysis, different types of HPV seem to correlate with epigenetic changes, this could bring attention to less studied HPV types and their role in carcinogenesis. Additional analyses of the current data and larger prospective, population-based studies, assessing the use of salivary DNA methylation in cancer-free and cancer groups of subjects are needed to explore the current findings and dissect the potential of saliva as a means of tracking cancer related changes. Citation Format: Ana Paula Muterle Varela, Dieila Giomo De Lima, Laura Palmieri, Juliana Comerlato, Fernando Hayashi Sant’Anna, Isabel C Bandeira, Marina Bessel, Pedro Isaacsson Velho, Eliana M. Wendland, Mariana Brait. HPV status and epigenetic profiling of saliva from a cancer-free population [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 6528.

NMR-Based Metabolomics Demonstrates a Metabolic Change during Early Developmental Stages from Healthy Infants to Young Children.

The present study aims to identify the salivary metabolic profile of healthy infants and young children, and to correlate this with age, salivary gland maturation, and dentition. Forty-eight children were selected after clinical evaluation in which all intraoral structures were examined. Total unstimulated saliva was collected, and salivary metabolites were analyzed by 1H Nuclear Magnetic Resonance (NMR) at 25 °C. Partial least squares discriminant analysis (PLS-DA), orthogonal PLS-DA (O-PLS-DA), and univariate analysis were used, adopting a 95% confidence interval. The study showed a distinct salivary metabolomic profile related to age and developmental phase. The saliva of children in the pre-eruption teeth period showed a different metabolite profile than that of children after the eruption. However, more evident changes were observed in the saliva profile of children older than 30 months. Alanine, choline, ethanol, lactate, and sugar region were found in higher levels in the saliva of patients before 30 months old. Acetate, N-acetyl sugar, butyrate, caproate, creatinine, leucine, phenylalanine, propionate, valine, succinate, and valerate were found to be more abundant in the saliva of children after 30 months old. The saliva profile is a result of changes in age and dental eruption, and these findings can be useful for monitoring the physiological changes that occur in infancy.

ЦИТОКИНОВЫЙ ПРОФИЛЬ СЛЮНЫ ПРИ НАРУШЕНИИ МУКОЗАЛЬНОГО ИММУНИТЕТА У ПАЦИЕНТОВ С ХРОНИЧЕСКИМ ГЕНЕРАЛИЗОВАННЫМ ПАРОДОНТИТОМ И САХАРНЫМ ДИАБЕТОМ II ТИПА

Subject. The subject of the study is the cytokine profile of saliva in violation of mucosal immunity in patients with chronic generalized periodontitis and concomitant type II diabetes mellitus.&#x0D; Objectives. The goal was to assess local levels of cytokines: tumor necrosis factor-alpha (TNF-α), interferon-gamma (IFN-γ), transforming growth factor-beta 1 (TGF-β1), interleukin-4 (IL-4), interleukin-17 (IL-17) in patients with chronic generalized periodontitis without concomitant pathology (group I), with chronic generalized periodontitis and type II diabetes mellitus (group II), as well as in patients without signs of periodontitis, but with established type II diabetes mellitus ( III group).&#x0D; Methodology. 126 patients were examined, including: 47 people – group I, 49 people – group II and 30 people – group III. The control group consisted of healthy volunteers (30 people). Patient saliva was used as the research material. The levels of the studied cytokines TNF-α, IFN-γ, TGF-β1, IL-4, IL-17 were determined by the sandwich variant of enzyme-linked immunosorbent assay using specific reagents from R&amp;D Diagnostics Inc (USA).&#x0D; Results. In patients of all studied groups, a significant increase in TNF-α, IFN-γ, IL-4 and IL-17 was found. In the group of patients with chronic periodontitis, a more pronounced increase in TNF-α, IFN-γ, TGF-β1 and IL-17, as well as an increase in the ratio of pro- and anti-inflammatory cytokines, was registered.&#x0D; Conclusion. The data obtained can be considered as promising markers for the personification of the prognosis for the development of chronic generalized periodontitis in type II diabetes mellitus.