Thermodynamic and Kinetic Studies of the Precipitation of Double-Doped Amorphous Calcium Phosphate and Its Behaviour in Artificial Saliva.

Simulated body fluid (SBF) and artificial saliva (AS) are used in biomedical and dental research to mimic the physiological conditions of the human body. In this study, the biomimetic precipitation of double-doped amorphous calcium phosphate in SBF and AS are compared by thermodynamic modelling of chemical equilibrium in the SBF/AS-CaCl2-MgCl2-ZnCl2-K2HPO4-H2O and SBF/AS-CaCl2-MgCl2-ZnCl2-K2HPO4-Glycine/Valine-H2O systems. The saturation indices (SIs) of possible precipitate solid phases at pH 6.5, close to pH of AS, pH 7.5, close to pH of SBF, and pH 8.5, chosen by us based on our previous experimental data, were calculated. The results show possible precipitation of the same salts with almost equal SIs in the two biomimetic environments at the studied pHs. A decrease in the saturation indices of magnesium and zinc phosphates in the presence of glycine is a prerequisite for reducing their concentrations in the precipitates. Experimental studies confirmed the thermodynamic predictions. Only X-ray amorphous calcium phosphate with incorporated Mg (5.86-8.85 mol%) and Zn (0.71-2.84 mol%) was obtained in the experimental studies, irrespective of biomimetic media and synthesis route. Solid-state nuclear magnetic resonance (NMR) analysis showed that the synthesis route affects the degree of structural disorder of the precipitates. The lowest concentration of dopant ions was obtained in the presence of glycine. Further, the behaviour of the selected amorphous phase in artificial saliva was studied. The dynamic of Ca2+, Mg2+, and Zn2+ ions between the solid and liquid phases was monitored. Both direct excitation 31P NMR spectra and 1H-31P CP-MAS spectra proved the increase in the nanocrystalline hydroxyapatite phase upon increasing the incubation time in AS, which is more pronounced in samples with lower additives. The effect of the initial concentration of doped ions on the solid phase transformation was assessed by solid-state NMR.

Salivary Proteins in Human Acquired Enamel Pellicle (AEP) on Eroded and Uneroded Teeth in Patients with Gastro-Oesophageal Reflux Disease (GORD).

The aim of this in vivo study was to compare total protein present in the salivary films (F) and acquired enamel pellicle (AEP) on eroded and non-eroded surfaces in patients suffering from GORD symptoms with and without GORD diagnosis (GORD, No-GORD). Thirty-nine patients suffering from GORD symptoms and erosive tooth wear on lower first molars and an unaffected posterior occlusal surface in the same quadrant were recruited from Guy's hospital, London. Salivary film and AEP were collected from the eroded and uneroded occlusal surfaces, using 0.5% sodium dodecyl sulphate (SDS)-soaked filter papers. Total protein concentration was analysed using bicinchoninic acid assay (BCA). Statistical analysis was conducted using Shapiro-Wilk, ANOVA, and Tukey's tests (p < 0.05), comparing four GDS sample types and GORD vs. No-GORD groups. The level of significance was set as p < 0.05. Data were compared between eroded and uneroded surfaces in the same patient with GORD symptoms, as well as between those with or without a GORD diagnosis (GORD, No-GORD). The AEP total protein concentration from the eroded [2.17 (0.49) mg/mL] and uneroded surfaces [2.24 (0.66) mg/mL] of the GORD group were statistically significantly lower than those on eroded [3.27 (1.01) mg/mL] and uneroded [3.33 (1.57) mg/mL] surfaces in the No-GORD group (p = 0.007) (p = 0.008), respectively. No statistically significant differences were observed for film and AEP between eroded and uneroded surfaces (p > 0.05).

Detection of HPV DNA in Saliva of Patients with HPV-Associated Oropharyngeal Cancer Treated with Radiotherapy.

BACKGROUND: To investigate the technical feasibility of RT-PCR and direct sequencing to quantify HPV DNA in the saliva of patients with Human-Papilloma-Virus related oropharyngeal cancer (HPV-OPC), the level of which is known to predict prognosis after treatment. METHODS: Nine patients with locally advanced HPV-OPC treated with definitive radiotherapy with chemotherapy or cetuximab, or radiotherapy alone between April 2016 and September 2017, were enrolled, two of whom also received induction chemotherapy. Saliva was collected before (baseline), during (mid-RT) and after (post-RT) radiotherapy. HPV-16 DNAs (E6 and E7) in saliva were quantified by RT-PCR and sequencing, the latter using a custom cancer panel. Correlations between HPV DNA levels and clinical outcomes were assessed. RESULTS: Compared to the baseline, the relative cycle threshold (Ct) value of E6 and E7 reduced at the point of mid-RT in the majority of the patients (100% and 75% for E6 and E7, respectively). Similarly, the relative Ct value from the baseline to post-RT reduced in 86% and 100% of the patients for E6 and E7, respectively. During the follow-up period, three patients (33%) experienced disease progression. The relative baseline Ct values of these three patients were in the top 4 of all the patients. The sequences of HPV DNA were detected in five (83%) of six samples of the baseline saliva that underwent DNA sequencing, along with several gene mutations, such as TP53,CDKN2A and PIK3CA. CONCLUSIONS: This study demonstrates that, in addition to detection and quantification of HPV DNA by RT-PCR, detection by sequencing of HPV-DNA using a customized cancer panel is technically possible.

Changes in cortisol, cortisone and 11ß-hydroxysteroid dehydrogenase type II activity in saliva during pregnancy and lactation in sows.

The activity of the enzyme 11ß-hydroxysteroid dehydrogenase type II, which can be estimated by the combined measurement of cortisol and cortisone, is gaining importance as a marker for the assessment of stress in pigs. The aim of this study was to investigate the activity of this enzyme and the salivary concentrations of cortisol and cortisone in pigs during pregnancy, farrowing and lactation and to compare it with other stress-related biomarkers such as CgA, S100A12 and alpha-amylase. Salivary cortisol concentrations and 11ß-hydroxysteroid dehydrogenase isoenzyme type 2 activity decreased after farrowing, while cortisol concentrations increased. Enzyme activity did not show significant correlations with any of the other stress-related biomarkers measured in this study. Overall, the results of this report indicate a different regulation of 11ß-HSD type II activity and of cortisol and cortisone during pregnancy and lactation, which should be considered when evaluating these analytes in saliva during these periods.

Higher Affinity Enables More Accurate Detection of SARS-CoV-2 in Human Saliva Using Aptamer-based Litmus Test.

Many aptamers have been generated by SELEX to recognize spike proteins of SARS-CoV-2, some of which have been engineered into dimeric and trimeric versions for enhanced affinity for diagnostic applications. However, no studies have been conducted to compare the utilities of monomeric, dimeric and trimeric aptamers in diagnostic assays with real clinical samples to answer the question of what levels of affinity an aptamer must have for accurate clinical diagnostics. Herein, we carried out a comparative study with two monomeric aptamers MSA1 and MSA5, one dimeric aptamer and two homotrimeric aptamers constructed with MSA1 and MSA5, with affinity varying by 1000-fold. Using a colorimetric sandwich assay to analyze 48 human saliva samples, we found that the trimeric aptamer assay (Kd = ~10 pM) can identify the SARS-CoV-2 infection much more accurately than the dimeric aptamer assay (Kd = ~100 pM) and monomeric aptamer assay (Kd = ~10,000 pM). Based on the experimental data, we theoretically predict the levels of affinity an aptamer needs to possess to achieve 80-100% sensitivity and 100% specificity. The findings from this study highlight the need for deriving very high affinity aptamers to enable highly accurate detection of viral infection for future pandemics.

Extraction-Free Testing for SARS-CoV-2 in Nasal Swab and Saliva Samples on a Single High-Throughput Platform.

The COVID-19 pandemic introduced an urgent need for rapid and high-throughput testing for SARS-CoV-2. RNA extraction is a major bottleneck for RT-qPCR. We describe a semi-automated, extraction-free RT-qPCR assay for detection of SARS-CoV-2 in nasal swab and saliva samples on a single platform. With a limit of detection of 4 copies/mL, this laboratory developed test performed equivalently to established methods requiring nucleic acid extraction. Five technologists staffing two shifts per day (80 person-hours) processed more than 400,000 samples over 10 months. Patients opted to provide nasal swab samples (83.6%) more frequently than saliva (16.4%), creating the added challenge of producing swab collection kits. Real-world testing data indicated a higher frequency of SARS-CoV-2 detection in saliva (10.1%) compared to nasal swab (7.7%). This cost-effective and quickly scalable approach is suitable for pandemic preparedness planning related to surveillance and diagnostic testing.

Detection of Mycobacterium tuberculosis complex in saliva by quantitative PCR: A potential alternative specimen for pulmonary tuberculosis diagnosis.

BACKGROUND: Current tuberculosis (TB) diagnostic tests primarily rely on sputum samples, yet many TB patients cannot produce sputum. This study explored whether saliva could be used instead of sputum to diagnose pulmonary TB (PTB). METHOD: The study included 32 patients with confirmed PTB and 30 patients with other respiratory diseases (ORD). Saliva from all study participants was subjected to quantitative (qPCR) assays targeting the IS1081 gene for detection of M. tuberculosis complex DNA. RESULTS: The sensitivity of saliva IS1081 qPCR was 65.6 % (95 % CI 48.4-80.2 %) with positive results for 21/32 PTB cases, while the specificity was 96.7 % (95 % CI 85.9-99.6 %) with negative results for 29/30 participants with ORD. Sensitivity improved to 72.4 % (95 % CI 54.6-86.0 %) when sputum-Xpert was used as the reference standard, while remaining similar at 65.5 % (95 % CI 47.4-80.7 %) when culture was used as the reference standard. In receiver operating characteristic (ROC) curve analysis, the area under the curve (AUC) for saliva IS1081 qPCR was 82.5 % (95 % CI 71.7-93.3 %). CONCLUSION: Saliva testing offers a promising alternative to sputum for TB diagnosis among confirmed PTB cases. Larger multicenter studies, encompassing diverse clinical TB characteristics, are needed to provide improved estimates of diagnostic sensitivity and specificity.

Salivary Microbiome Relates to Neoadjuvant Immunotherapy Response in OSCC.

Most patients diagnosed with oral squamous cell carcinoma (OSCC) present with locally advanced stages, which are typically associated with poor outcomes. Although immunotherapy offers potential improvements in patient survival, its efficacy is hampered by low response rates. The microbiome is widely involved in tumor immunity and may play a role in immunotherapy. This study aimed to investigate the potential association between the oral (salivary) microbiome and immunotherapy response in patients with OSCC. Salivary metagenome sequencing was performed on 47 patients with OSCC undergoing neoadjuvant immunotherapy (NAIT) in a clinical trial (NCT04649476). Patients were divided into responders and nonresponders based on their pathological responses. The results showed that the species richness of the salivary microbiome was lower in the nonresponders before NAIT than in the responders. Differential analysis revealed that nonresponders exhibited a lower relative abundance of 34 bacterial species and a higher relative abundance of 4 bacterial species. Notably, low levels of Eubacterium infirmum, Actinobaculum, and Selenomas (EAS) in the saliva may be associated with the nonresponse of patients with OSCC to NAIT. A nomogram based on EAS was developed and validated to determine the efficacy of NAIT. The area under the curve for the training cohort was 0.81 (95% confidence interval, 0.66 to 0.81). Quantitative polymerase chain reaction confirmed that low levels of salivary EAS effectively identified nonresponders to NAIT. Furthermore, the low abundance of salivary EAS was closely correlated with a low density of intratumoral CD4+, CD14+, CD68+, and FOXP3+ cells. Metabolic functional annotation revealed numerous biosynthetic processes associated with EAS that were more active in responders. In summary, this study provides valuable data resources for the salivary microbiome and reveals that nonresponders have different salivary microbiome profiles than responders do before NAIT. Low salivary EAS levels can serve as potential biomarkers for distinguishing nonresponders from responders.

Transdermal pilocarpine on the skin over salivary glands to increase salivation: an in vivo study.

BACKGROUND: Hyposalivation is treated using oral cholinergic drugs; however, systemic side effects occasionally lead to discontinuation of treatment. We aimed to investigate the effects of transdermal pilocarpine on the salivary gland skin on saliva secretion and safety in rats. METHODS: Pilocarpine was administered to rats orally (0.5 mg/kg) or topically on the salivary gland skin (5 mg/body). Saliva volume, the number of sweat dots, and fecal weight were measured along with pilocarpine concentration in plasma and submandibular gland tissues. RESULTS: Saliva volume significantly increased 0.5 h after oral administration and 0.5, 3, and 12 h after topical administration. Fecal weight and sweat dots increased significantly 1 h after oral administration; however, no changes were observed after topical application. The pilocarpine concentration in the submandibular gland tissues of the topical group was higher than that in the oral group at 0.5, 3, and 12 h of administration. CONCLUSIONS: Pilocarpine application to salivary gland skin persistently increased salivary volume in rats without inducing sweating or diarrhea. Transdermal pilocarpine applied to the skin over the salivary glands may be an effective and safe treatment option for hyposalivation.

Physicochemical properties, biological chemistry and mechanisms of action of caries-arresting diammine-silver(I) fluoride and silver(I)-fluoride solutions for clinical use: a critical review.

This paper serves as a Part II follow-up of our research investigations performed on the molecular structures of silver(I)-fluoride (SF) and diammine-silver(I) fluoride (SDF) complexes in solution-based commercial products for clinical application, their precise chemical compositions, and their nature in aqueous solution, the latter including rapid fluoride-exchange processes at the silver(I) ion centre monitored by 19F NMR analysis (Part I). Part I of this series also explores the mechanisms of action (MoA) of these complexes, and is therefore largely focused on their chemical reactions with constituents of human saliva, which has access to their sites of application. Such reactions were found to slowly promote the generation of potentially physiologically-active Ag/AgCl nanoparticles from primarily-generated discoloured silver(I) chloride (AgCl) precipitates, a process involving salivary electron-donors such as thiocyanate and L-cysteine. Since this research has shed new light on potential MoAs for these products, in this accompanying report (Part II), we have performed a critical review of scientific literature in order to rationalize our results in relation to current views on these mechanisms for SF and SDF products employed for the successful clinical arrest of dental caries. Following an Introduction to the subject matter ( Section 1), this paper comprises a generalized overview of silver coordination chemistry ( Section 2), which is followed by a section focused on the aqueous solution status and equilibria involved in SF chemistry ( Section 3), the latter including results acquired from an original simulation of the electronic absorption spectra of coloured SF complexes in aqueous solution (Section 3.1). Section 4 then investigates detailed rationales for the biologically-relevant ligand-exchange and redox chemistries, disposition and fates of SF, SDF and silver(I)-nitrate when employed for the treatment of dental caries, with emphasis placed on their therapeutic MoAs. This Section is supported by the provision of valuable information centralized on (1) relevant biomolecular chemistry involved in solution- and solid-state matrices ( Section 4.1); (2) SF and perhaps silver(I)-nitrate as more cost-effective alternatives to SDF therapies ( Section 4.2); and (3) the potential therapeutic benefits and effects offered by silver-based nanoparticles and their associated MoAs ( Section 4.3). Recommendations for future investigations in this area are proposed.

B cell senescence promotes age-related changes in oral microbiota.

In recent years, there has been increasing attention towards understanding the relationship between age-related alterations in the oral microbiota and age-associated diseases, with reports emphasizing the significance of maintaining a balanced oral microbiota for host health. However, the precise mechanisms underlying age-related changes in the oral microbiota remain elusive. We recently reported that cellular senescence of ileal germinal center (GC) B cells, triggered by the persistent presence of commensal bacteria, results in diminished IgA production with aging and subsequent alterations in the gut microbiota. Consequently, we hypothesize that a similar phenomenon may occur in the oral cavity, potentially contributing to age-related changes in the oral microbiota. Examination of p16-luc mice, wherein the expression of the senescent cell marker p16INK4a can be visualized, raised under specific pathogen-free (SPF) or germ-free (GF) conditions, indicated that, unlike ileal GC B cells, the accumulation of senescent cells in GC B cells of cervical lymph nodes increases with age regardless of the presence of commensal bacteria. Furthermore, longitudinal studies utilizing the same individual mice throughout their lifespan revealed concurrent age-related alterations in the composition of the oral microbiota and a decline in salivary IgA secretion. Further investigation involving Rag1-/- mice transplanted with B cells from wild-type or p16INK4a and p21Waf1/Cip1 -double knockout mice unveiled that B cell senescence leads to reduced IgA secretion and alteration of the oral microbiota. These findings advance our understanding of the mechanism of age-associated changes in the oral microbiota and open up possibilities of their control.

Assessing the influence of sleep and sampling time on metabolites in oral fluid: implications for metabolomics studies.

INTRODUCTION: The human salivary metabolome is a rich source of information for metabolomics studies. Among other influences, individual differences in sleep-wake history and time of day may affect the metabolome. OBJECTIVES: We aimed to characterize the influence of a single night of sleep deprivation compared to sufficient sleep on the metabolites present in oral fluid and to assess the implications of sampling time points for the design of metabolomics studies. METHODS: Oral fluid specimens of 13 healthy young males were obtained in Salivette® devices at regular intervals in both a control condition (repeated 8-hour sleep) and a sleep deprivation condition (total sleep deprivation of 8 h, recovery sleep of 8 h) and their metabolic contents compared in a semi-targeted metabolomics approach. RESULTS: Analysis of variance results showed factor 'time' (i.e., sampling time point) representing the major influencer (median 9.24%, range 3.02-42.91%), surpassing the intervention of sleep deprivation (median 1.81%, range 0.19-12.46%). In addition, we found about 10% of all metabolic features to have significantly changed in at least one time point after a night of sleep deprivation when compared to 8 h of sleep. CONCLUSION: The majority of significant alterations in metabolites' abundances were found when sampled in the morning hours, which can lead to subsequent misinterpretations of experimental effects in metabolomics studies. Beyond applying a within-subject design with identical sample collection times, we highly recommend monitoring participants' sleep-wake schedules prior to and during experiments, even if the study focus is not sleep-related (e.g., via actigraphy).

[Effects of coffee consumption on salivary cortisol and alpha amylase in young adults]. / Efectos del consumo de café sobre el cortisol y la alfa-amilasa salival en adultos jóvenes.

Objective: The purpose of this study was to determine the effects of coffee consumption on salivary cortisol (sCort) and alpha amylase (sAA) in young adults. Materials and methods: Sixty healthy university students, habitual coffee consumers, participated in this descriptive observational study. Participants were divided into three groups: G1 low consumption (≤ 2 cups of coffee per day, n = 20), G2 moderate consumption (2-5 cups of coffee per day, n = 20), and G3 high consumption (>5 cups of coffee per day, n = 20). Saliva self-collection was in the morning (6:30-7:30 AM) and at night (08:00-09:00 PM). sCort was analyzed using chemiluminescence and sAA activity by kinetic method. Statistical analysis of the data was performed using Student's t-test and analysis of variance. Results: The sample consisted of 30 women and 30 men, aged between 20 and 35 years. In all groups, sCort values were higher in the morning (AM 0,29 ± 0,19 vs. PM 0,09 ± 0,05 µg/dl, p < 0.0001). In contrast, sAA levels were higher in the night (PM 160,16 ± 60,42 vs. AM 32,79 ± 12,98 U/ml, p < 0.0001). No significant differences were detected, in the contents of Corts and AAs, between the groups. Conclusion: : Coffee consumption, in non-stressful conditions, did not alter levels and patterns of sCort and sAA in young adults.

Salivary biomarkers for the diagnosis of gastric cancer: a systematic review.

Background: Recent advancements reveal saliva as a crucial source of diagnostic biomarkers for various diseases, notably gastric cancer. This systematic review evaluates these biomarkers, emphasizing their clinical applicability and potential in early detection. Methods: An extensive electronic search was conducted across PubMed/MEDLINE, Scopus, and Web of Science to identify relevant studies. Salivary biomarkers were analyzed as independent variables, with gastric cancer as the dependent variable. The study adhered to a protocol registered with PROSPERO (CRD42021259519). Results: Our analysis identified a range of biomarkers, highlighting three proteins - cystatin-B (CSTB), triosephosphate isomerase (TPI1), and deleted in malignant brain tumors 1 protein (DMBT1) - as particularly accurate for gastric cancer diagnosis. Conclusions: Salivary biomarkers hold substantial promise for the early detection of gastric cancer. Future research should aim to refine study design and validation for enhancing the quality and applicability of these biomarkers.

Introducción: Avances recientes revelan la saliva como una fuente crucial de biomarcadores diagnósticos para diversas enfermedades, especialmente el cáncer gástrico. Esta revisión sistemática evalúa estos biomarcadores, con énfasis en su aplicabilidad clínica y potencial para la detección temprana. Métodos: Se realizó una extensa búsqueda electrónica en PubMed/MEDLINE, Scopus y Web of Science para identificar estudios relevantes. Los biomarcadores salivales fueron analizados como variables independientes, con el cáncer gástrico como variable dependiente. El estudio siguió un protocolo registrado en PROSPERO (CRD42021259519). Resultados: Nuestro análisis identificó una gama de biomarcadores entre los que destacan tres proteínas: cistatina-B (CSTB), triosa fosfato isomerasa (TPI1) y proteína 1 eliminada en tumores cerebrales malignos (DMBT1), como particularmente precisas para el diagnóstico del cáncer gástrico. Conclusiones: : Los biomarcadores salivales tienen un gran potencial para la detección temprana del cáncer gástrico. La investigación futura debería apuntar a refinar el diseño del estudio y la validación para mejorar la calidad y aplicabilidad de estos biomarcadores.

Salivary 8-hydroxy-2'-deoxyguanosine levels in patients with oral cancer: a systematic review and meta-analysis.

BACKGROUND: DNA is an important target for oxidative attack and its modification may increase the risk of mutagenesis. The aim of this study was to evaluate and compare salivary levels of the oxidative stress biomarker 8-hydroxy-2'-deoxyguanosine (8-OHdG) in patients with oral cancer (OC) compared to the control group by a comprehensive search of the available literature. METHODS: The present systematic review and meta-analysis followed the Preferred Reporting Items for Systematic Reviews and Meta-Analysis (PRISMA) guidelines and was registered in Open Science Framework (OSF): https://doi.org/10.17605/OSF.IO/X3YMR. Four electronic databases were used to identify studies for this systematic review: PubMed, Scopus, ScienceDirect, and Web of Science from January 15, 2005, to April 15, 2021. The Joanna Briggs Institute (JBI) tool was used to assess article quality. RESULTS: Of the 166 articles identified, 130 articles were excluded on the basis of title and abstract screening (duplicates, reviews, etc.). Thirty-six articles were evaluated at full text and 7 articles met the inclusion criteria. Of these, only 5 studies had compatible data for quantitative analysis. An increase in salivary 8-OHdG levels was found in patients with OC compared to healthy subjects, but without statistical significance. 8-OHdG: SMD = 2,72 (95%CI= -0.25-5.70); *p = 0.07. CONCLUSIONS: This systematic review and meta-analysis suggests a clear trend of increased 8-OHdG levels in saliva of OC patients compared to the control group. However, further studies are required to clarify and understand the altered levels of this oxidative stress marker.

Comparative proteomic analysis of saliva from chewing tobacco users and oral cancer patients reveals shared biomarkers: A case control observational study.

OBJECTIVE: The aim of this study was to compare the salivary proteomic profile of smokeless tobacco users with that of non-users and oral cancer patients using Liquid Chromatography-Mass Spectrometry/ Mass Spectrometry (LC-MS/MS). METHODS: Saliva samples from 65 participants were collected in three groups: control (25 participants), smokeless tobacco users (25 participants), and oral cancer (15 participants). RESULTS: The analysis revealed 343 protein groups with significantly altered abundance in the saliva samples (P < 0.05). Among these, 43 out of 51 dysregulated proteins in the smokeless tobacco group were also dysregulated in the oral cancer group. Notably, Apolipoprotein A1 (ApoA1) and Pon1 were found to be significantly increased in both smokeless tobacco users and oral cancer patients (p < 0.05). Furthermore, six out of the 20 most significantly altered proteins were mitochondrial proteins, and all of these were decreased relative to controls in both smokeless tobacco users and cancer samples. CONCLUSION: The proteomic profile of users of chewing (smokeless) tobacco (SLT) shows substantial overlap in the altered pathways and dysregulated proteins with those altered in oral cancer samples, suggesting that SLT use induces a shift toward an oncogenic state. Specifically indicated pathways included blood microparticles, platelet α-granules and protease inhibitors as well as indicators of oxidative stress and exogenous compound processing. What differentiates oral cancer samples from SLT users is enrichment of alterations related to cytoskeletal organisation and tissue remodelling. CLINICAL SIGNIFICANCE: The findings emphasize the importance of salivary proteomic profiles because changes in certain proteins may be indicators for early oral cancer identification and risk assessment in smokeless tobacco users.

Protein Network Alterations in G-CSF Treated Severe Congenital Neutropenia Patients and Beneficial Effects of Oral Health Intervention.

PURPOSE: Severe congenital neutropenia (SCN) is a raredisorder characterized by diminished neutrophil levels. Despite granulocytecolony-stimulating factor (G-CSF) treatment, SCN patients remain still prone tosevere infections, including periodontal disease-a significant oral healthrisk. This study investigates the host proteome and metaproteome in saliva andgingival crevicular fluid (GCF) of G-CSF-treated patients. EXPERIMENTAL DESIGN: We used label-free quantitative proteomics on saliva and GCF samples from SCN patients before (n = 10, mean age: 10.7 ± 6.6 years) and after a 6-month oral hygiene intervention (n = 9,mean age: 11.6 ± 5.27 years), and from 12 healthy controls. RESULTS: We quantified 894 proteins in saliva (648 human,246 bacterial) and 756 proteins in GCF (493 human, 263 bacterial). Predominant bacterial genera included Streptococcus, Veillonella, Selenomonas, Corynebacterium, Porphyromonas, and Prevotella. SCN patients showed reduced antimicrobial peptides (AMPs) and elevated complement proteins compared tohealthy controls. Oral hygiene intervention improved oral epithelial conditionsand reduced both AMPs and complement proteins. CONCLUSIONS AND CLINICAL RELEVANCE: SCN patients have aunique proteomic profile with reduced AMPs and increased complement proteins, contributing to infection susceptibility. Oral hygiene intervention not onlyimproved oral health in SCN patients but also offers potential overall therapeuticbenefits.

Dysregulation of Porphyromonas gingivalis Agmatine Deiminase Expression in Alzheimer's Disease.

BACKGROUND: Alzheimer's disease (AD) is the most prevalent neurodegenerative disorder, with a significant burden on global health. AD is characterized by a progressive cognitive decline and memory loss. Emerging research suggests a potential link between periodontitis, specifically the presence of oral bacteria such as Porphyromonas gingivalis (P. gingivalis), and AD progression. P. gingivalis produces an enzyme, Agmatine deiminase (AgD), which converts agmatine to N-carbamoyl putrescine (NCP), serving as a precursor to essential polyamines. Recent studies have confirmed the correlation between disruptions in polyamine metabolism and cognitive impairment. OBJECTIVE: This study aims to investigate the dysregulation of P. gingivalis Agmatine deiminase (PgAgD) in the context of AD. METHODS: Saliva samples were collected from a total of 54 individuals, including 27 AD patients and 27 healthy controls. The expression of the PgAgD gene was analyzed using quantitative Real-- Time PCR. RESULTS: The results showed a significant decrease in PgAgD gene expression in the saliva samples of AD patients compared to healthy controls. This downregulation was found in AD patients with advanced stages of periodontitis. Additionally, a correlation was observed between the decrease in PgAgD expression and the 30-item Mini-Mental State Examination (MMSE) score. CONCLUSION: These findings suggest that measuring PgAgD expression in saliva could be a noninvasive tool for monitoring AD progression and aid in the early diagnosis of patients with periodontitis. Further research is needed to validate our results and explore the underlying mechanisms linking periodontitis, PgAgD expression, and AD pathophysiology.

Detection of autoantibodies to heat shock protein 70 in the saliva and urine of normal individuals.

Cells exposed to stressors of various origin activate protective mechanisms that include the expression of heat shock proteins (Hsps)/molecular chaperones belonging to several families. Well-characterized inducible Hsp70 is present in all human cell-types and biological fluids, including blood, urine, and saliva. The presence of anti-Hsp70 autoantibodies in the serum of healthy individuals has already been confirmed, and their elevated titers positively correlated with the severity of several pathological conditions, including coeliac disease and dermatitis herpetiformis - a cutaneous manifestation of coeliac disease. Here, using an indirect enzyme-linked immunosorbent assay, we demonstrate, for the first time, that anti-Hsp70 autoantibodies are present in the saliva and urine of healthy individuals. Although the occurrence of anti-Hsp70 autoantibodies in the biological fluids of healthy individuals is intriguing, their physiological role is currently unknown. It is believed that antibodies reacting with self-molecules present in the serum of healthy individuals are part of natural autoantibody pool with multiple regulatory functions. On the other hand, some autoantibodies (e.g., typical of autoimmune bullous skin diseases or systemic lupus erythematosus) may be present before the onset of the disease and serve as specific predictive biomarkers. Therefore, we would like to initiate a discussion or future research direction on the use of anti-Hsp70 autoantibodies as a potential "biomarker" in the diagnosis or prediction of autoimmune diseases. Our findings can be considered in biomedical research to develop noninvasive, inexpensive and easy-to-use tests. Nevertheless, large-scale comparative studies should be initiated, involving the collection and analysis of biological samples such as saliva or urine from patients suffering from autoimmune diseases or other inflammatory or neoplastic diseases, to determine whether the levels of anti-Hsp70 autoantibodies are indeed elevated and whether they correlate with the clinical picture of any disease or established biomarkers.

Salivary cortisol measurements in brachycephalic dog breeds as part of a standardized stress test.

Introduction: Brachycephalic obstructive airway syndrome (BOAS) is a common condition in brachycephalic dogs, with Pugs (PG) and French Bulldogs (FB) appearing to be particularly typically affected. Objective and easy-to-perform tests are necessary to detect the disease at an early stage and to exclude dogs affected by BOAS from breeding. Methods: The present study investigated the extent to which vital signs and salivary cortisol concentrations can be used to distinguish between healthy and BOAS-affected dogs in a standardized fitness test. A total of 57 PG, 56 FB and 27 meso- and dolichocephalic dogs were studied as control group (CG). In addition to vital signs, salivary cortisol concentrations were measured before and after the exercise test. Results: It emerged that non-brachycephalic dogs showed a higher fitness level than brachycephalic dogs. The PG recovered significantly slower than the FB after the exercise test. In unaffected PG, cortisol levels rose significantly after the test and then fell again, in unaffected FB they fell significantly during the test. Unexpectedly, cortisol levels remained constant in BOAS affected dogs of both breeds. Discussion: A possible explanation could be a disturbance of the pituitary-hypothalamic-adrenal axis, which could be due to the chronic stress of affected animals. This would have to be clarified in further studies. In conclusion, a submaximal fitness test may be a useful method to identify dogs suffering from BOAS as it is imperative to prevent the breeding and reproduction of affected dogs.