Extracellular RNAs (exRNAs) in human body fluids are emerging as effective biomarkers for detection of diseases. Saliva, as the most accessible and noninvasive body fluid, has been shown to harbor ...exRNA biomarkers for several human diseases. However, the entire spectrum of exRNA from saliva has not been fully characterized.
Using high-throughput RNA sequencing (RNA-Seq), we conducted an in-depth bioinformatic analysis of noncoding RNAs (ncRNAs) in human cell-free saliva (CFS) from healthy individuals, with a focus on microRNAs (miRNAs), piwi-interacting RNAs (piRNAs), and circular RNAs (circRNAs).
Our data demonstrated robust reproducibility of miRNA and piRNA profiles across individuals. Furthermore, individual variability of these salivary RNA species was highly similar to those in other body fluids or cellular samples, despite the direct exposure of saliva to environmental impacts. By comparative analysis of >90 RNA-Seq data sets of different origins, we observed that piRNAs were surprisingly abundant in CFS compared with other body fluid or intracellular samples, with expression levels in CFS comparable to those found in embryonic stem cells and skin cells. Conversely, miRNA expression profiles in CFS were highly similar to those in serum and cerebrospinal fluid. Using a customized bioinformatics method, we identified >400 circRNAs in CFS. These data represent the first global characterization and experimental validation of circRNAs in any type of extracellular body fluid.
Our study provides a comprehensive landscape of ncRNA species in human saliva that will facilitate further biomarker discoveries and lay a foundation for future studies related to ncRNAs in human saliva.
The discovery of disease-specific biomarkers in oral fluids has revealed a new dimension in molecular diagnostics. Recent studies have reported the mechanistic involvement of tumor cells derived ...mediators, such as exosomes, in the development of saliva-based mRNA biomarkers. To further our understanding of the origins of disease-induced salivary biomarkers, we here evaluated the hypothesis that tumor-shed secretory lipidic vesicles called exosome-like microvesicles (ELMs) that serve as protective carriers of tissue-specific information, mRNAs, and proteins, throughout the vasculature and bodily fluids. RNA content was analyzed in cell free-saliva and ELM-enriched fractions of saliva. Our data confirmed that the majority of extracellular RNAs (exRNAs) in saliva were encapsulated within ELMs. Nude mice implanted with human lung cancer H460 cells expressing hCD63-GFP were used to follow the circulation of tumor cell specific protein and mRNA in the form of ELMs in vivo. We were able to identify human GAPDH mRNA in ELMs of blood and saliva of tumor bearing mice using nested RT-qPCR. ELMs positive for hCD63-GFP were detected in the saliva and blood of tumor bearing mice as well as using electric field-induced release and measurement (EFIRM). Altogether, our results demonstrate that ELMs carry tumor cell-specific mRNA and protein from blood to saliva in a xenografted mouse model of human lung cancer. These results therefore strengthen the link between distal tumor progression and the biomarker discovery of saliva through the ELMs.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Saliva is a useful biofluid for the early detection of disease, but how distal tumors communicate with the oral cavity and create disease-specific salivary biomarkers remains unclear. Using an in ...vitro breast cancer model, we demonstrated that breast cancer-derived exosome-like microvesicles are capable of interacting with salivary gland cells, altering the composition of their secreted exosome-like microvesicles. We found that the salivary gland cells secreted exosome-like microvesicles encapsulating both protein and mRNA. We also showed that the interaction with breast cancer-derived exosome-like microvesicles communicated and activated the transcriptional machinery of the salivary gland cells. Thus, the interaction altered the composition of the salivary gland cell-derived exosome-like microvesicles on both the transcriptomically and proteomically.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Morbidly obese patients undergoing general anesthesia are at risk of hypoxemia during anesthesia induction. High-flow nasal oxygenation use during anesthesia induction prolongs safe apnea time in ...nonobese surgical patients. The primary objective of our study was to compare safe apnea time, between patients given high-flow nasal oxygenation or conventional facemask oxygenation during anesthesia induction, in morbidly obese surgical patients.
Research ethics board approval was obtained. Elective surgical patients ≥18 years with body mass index ≥40 kg·m were included. Patients with severe comorbidity, gastric reflux disease, known difficult airway, or nasal obstruction were excluded. After obtaining informed consent patients were randomized. In the intervention (high-flow nasal oxygenation) group, preoxygenation was provided by 100% nasal oxygen for 3 minutes at 40 L·minute; in the control group, preoxygenation was delivered using a facemask with 100% oxygen, targeting end-tidal O2 >85%. Anesthesia was induced with propofol, remifentanil, and rocuronium. Bag-mask ventilation was not performed. At 2 minutes after rocuronium, videolaryngoscopy was performed. If the laryngoscopy grade was I or II, laryngoscope was left in place and the study was continued; if grade III or IV was observed, the patient was excluded from the study. During the apnea period, high-flow nasal oxygenation patients received nasal oxygen at 60 L·minute; control group patients received no supplemental oxygen. The primary outcome, safe apnea time, was reached when oxygen saturation measured by pulse oximetry (SpO2) fell to 95% or maximum 6 minutes of apnea. The patient was then intubated. T tests and χ analyses were used to compare groups. P < .05 was considered significant.
Forty patients completed the study. Baseline parameters were comparable between groups. Safe apnea time was significantly longer (261.4 ± 77.7 vs 185.5 ± 52.9 seconds; mean difference 95% CI, 75.9 33.3-118.5; P = .001) and the minimum peri-intubation SpO2 was higher (91.0 ± 3.5 vs 88.0 ± 4.8; mean difference 95% CI, 3.1 0.4-5.7; P = .026) in the high-flow nasal oxygenation group compared to the control group.
High-flow nasal oxygenation, compared to conventional oxygenation, provided a longer safe apnea time by 76 seconds (40%) and higher minimum SpO2 in morbidly obese patients during anesthesia induction. High-flow oxygenation use should be considered in morbidly obese surgical patients.
A sensitive assay to identify biomarkers using non-invasively collected clinical specimens is ideal for breast cancer detection. While there are other studies showing disease biomarkers in saliva for ...breast cancer, our study tests the hypothesis that there are breast cancer discriminatory biomarkers in saliva using de novo discovery and validation approaches. This is the first study of this kind and no other study has engaged a de novo biomarker discovery approach in saliva for breast cancer detection. In this study, a case-control discovery and independent preclinical validations were conducted to evaluate the performance and translational utilities of salivary transcriptomic and proteomic biomarkers for breast cancer detection.
Salivary transcriptomes and proteomes of 10 breast cancer patients and 10 matched controls were profiled using Affymetrix HG-U133-Plus-2.0 Array and two-dimensional difference gel electrophoresis (2D-DIGE), respectively. Preclinical validations were performed to evaluate the discovered biomarkers in an independent sample cohort of 30 breast cancer patients and 63 controls using RT-qPCR (transcriptomic biomarkers) and quantitative protein immunoblot (proteomic biomarkers). Transcriptomic and proteomic profiling revealed significant variations in salivary molecular biomarkers between breast cancer patients and matched controls. Eight mRNA biomarkers and one protein biomarker, which were not affected by the confounding factors, were pre-validated, yielding an accuracy of 92% (83% sensitive, 97% specific) on the preclinical validation sample set.
Our findings support that transcriptomic and proteomic signatures in saliva can serve as biomarkers for the non-invasive detection of breast cancer. The salivary biomarkers possess discriminatory power for the detection of breast cancer, with high specificity and sensitivity, which paves the way for prediction model validation study followed by pivotal clinical validation.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
The associations between oral diseases and increased risk of pancreatic cancer have been reported in several prospective cohort studies. In this study, we measured variations of salivary microbiota ...and evaluated their potential associations with pancreatic cancer and chronic pancreatitis.
This study was divided into three phases: (1) microbial profiling using the Human Oral Microbe Identification Microarray to investigate salivary microbiota variation between 10 resectable patients with pancreatic cancer and 10 matched healthy controls, (2) identification and verification of bacterial candidates by real-time quantitative PCR (qPCR) and (3) validation of bacterial candidates by qPCR on an independent cohort of 28 resectable pancreatic cancer, 28 matched healthy control and 27 chronic pancreatitis samples.
Comprehensive comparison of the salivary microbiota between patients with pancreatic cancer and healthy control subjects revealed a significant variation of salivary microflora. Thirty-one bacterial species/clusters were increased in the saliva of patients with pancreatic cancer (n=10) in comparison to those of the healthy controls (n=10), whereas 25 bacterial species/clusters were decreased. Two out of six bacterial candidates (Neisseria elongata and Streptococcus mitis) were validated using the independent samples, showing significant variation (p<0.05, qPCR) between patients with pancreatic cancer and controls (n=56). Additionally, two bacteria (Granulicatella adiacens and S mitis) showed significant variation (p<0.05, qPCR) between chronic pancreatitis samples and controls (n=55). The combination of two bacterial biomarkers (N elongata and S mitis) yielded a receiver operating characteristic plot area under the curve value of 0.90 (95% CI 0.78 to 0.96, p<0.0001) with a 96.4% sensitivity and 82.1% specificity in distinguishing patients with pancreatic cancer from healthy subjects.
The authors observed associations between variations of patients' salivary microbiota with pancreatic cancer and chronic pancreatitis. This report also provides proof of salivary microbiota as an informative source for discovering non-invasive biomarkers of systemic diseases.
ABSTRACT
Tumor exosomes are emerging as antitumor immunity regulators; however, their effects on secondary exosome secretion by distal organs have not been explored. We have previously demonstrated ...that suppression of exosomes at the distal tumor site of pancreatic ductal adenocarcinoma (PDAC) ablated the development of salivary biomarker profile. Here, we explore the function of salivary exosomes from tumor‐bearing mice in immune surveillance. We provide evidence that salivary exosomes from mice with PDAC exhibit a suppressive effect that results in reduced tumor‐killing capacity by NK cells. Salivary exosomes from mice with PDAC where pancreatic tumors were engineered to suppress exosome biogenesis failed to suppress NK cell cytotoxic potential against tumor cells, as opposed to salivary exosomes from mice with PDAC with normal tumor exosome biogenesis. These results reveal an important and previously unknown mechanism of antitumor immune regulation and provide new insights into our understanding of the alterations of this biofluid during tumor development.—Katsiougiannis, S., Chia, D., Kim, Y., Singh, R. P., Wong, D. T. W. Saliva exosomes from pancreatic tumor‐bearing mice modulate NK cell phenotype and antitumor cytotoxicity. FASEB J. 31, 998–1010 (2017). www.fasebj.org
Amperial™ is a novel assay platform that uses immobilized antigen in a conducting polymer gel followed by detection via electrochemical measurement of oxidation-reduction reaction between ...H2O2/Tetrametylbenzidine and peroxidase enzyme in a completed assay complex. A highly specific and sensitive assay was developed to quantify levels of IgG antibodies to SARS-CoV-2 in saliva. After establishing linearity and limit of detection we established a reference range of 5 standard deviations above the mean. There were no false positives in 667 consecutive saliva samples obtained prior to 2019. Saliva was obtained from 34 patients who had recovered from documented COVID-19 or had documented positive serologies. All of the patients with symptoms severe enough to seek medical attention had positive antibody tests and 88% overall had positive results. We obtained blinded paired saliva and plasma samples from 14 individuals. The plasma was analyzed using an EUA-FDA cleared ELISA kit and the saliva was analyzed by our Amperial™ assay. All 5 samples with negative plasma titers were negative in saliva testing. Eight of the 9 positive plasma samples were positive in saliva and 1 had borderline results. A CLIA validation was performed as a laboratory developed test in a high complexity laboratory. A quantitative non-invasive saliva based SARS-CoV-2 antibody test was developed and validated with sufficient specificity to be useful for population-based monitoring and monitoring of individuals following vaccination.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Abstract T helper 17 (Th17), a distinct subset of CD4+ T cells with IL-17 as their major cytokine, orchestrate the pathogenesis of inflammatory and autoimmune diseases. Dysregulated Th17 cells ...contribute to inflammatory and autoimmune diseases. Candidate biologics are in development for targeting IL-17, IL-17 receptors or IL-17 pathways. Several drugs that impact the IL-17 pathway are already in clinical trials for the treatment of autoimmune diseases. In this review we provide evidence for the role of Th17 cells in immune-mediated diseases. An understanding of the role of Th17 in these conditions will provide important insights and unravel novel targets for therapeutic intervention.
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