Plasma Signatures Research Articles

Pre- versus post-operative untargeted plasma nuclear magnetic resonance spectroscopy metabolomics of pheochromocytoma and paraganglioma

PurposePheochromocytomas and Paragangliomas (PPGL) result in chronic catecholamine excess and serious health complications. A recent study obtained a metabolic signature in plasma from PPGL patients; however, its targeted nature may have generated an incomplete picture and a broader approach could provide additional insights. We aimed to characterize the plasma metabolome of PPGL patients before and after surgery, using an untargeted approach, and to broaden the scope of the investigated metabolic impact of these tumors.DesignA cohort of 36 PPGL patients was investigated. Blood plasma samples were collected before and after surgical tumor removal, in association with clinical and tumor characteristics.MethodsPlasma samples were analyzed using untargeted nuclear magnetic resonance (NMR) spectroscopy metabolomics. The data were evaluated using a combination of uni- and multi-variate statistical methods.ResultsBefore surgery, patients with a nonadrenergic tumor could be distinguished from those with an adrenergic tumor based on their metabolic profiles. Tyrosine levels were significantly higher in patients with high compared to those with low BMI. Comparing subgroups of pre-operative samples with their post-operative counterparts, we found a metabolic signature that included ketone bodies, glucose, organic acids, methanol, dimethyl sulfone and amino acids. Three signals with unclear identities were found to be affected.ConclusionsOur study suggests that the pathways of glucose and ketone body homeostasis are affected in PPGL patients. BMI-related metabolite levels were also found to be altered, potentially linking muscle atrophy to PPGL. At baseline, patient metabolomes could be discriminated based on their catecholamine phenotype.

On the Growth and Development of Non‐Linear Kelvin–Helmholtz Instability at Mars: MAVEN Observations

AbstractIn this study, we have analyzed Mars Atmosphere and Volatile EvolutioN (MAVEN) observations of fields and plasma signatures associated with an encounter of fully developed Kelvin–Helmholtz (K–H) vortices at the northern polar terminator along Mars' induced magnetosphere boundary. The signatures of the K–H vortices event are: (a) quasi‐periodic, “bipolar‐like” sawtooth magnetic field perturbations, (b) corresponding density decrease, (c) tailward enhancement of plasma velocity for both protons and heavy ions, (d) co‐existence of magnetosheath and planetary plasma in the region prior to the sawtooth magnetic field signature (i.e., mixing region of the vortex structure), and (e) pressure enhancement (minimum) at the edge (center) of the sawtooth magnetic field signature. Our results strongly support the scenario for the non‐linear growth of K–H instability along Mars’ induced magnetosphere boundary, where a plasma flow difference between the magnetosheath and induced‐magnetospheric plasma is expected. Our findings are also in good agreement with 3‐dimensional local magnetohydrodynamics simulation results. MAVEN observations of protons with energies greater than 10 keV and results from the Walén analyses suggests the possibility of particle energization within the mixing region of the K–H vortex structure via magnetic reconnection, secondary instabilities or other turbulent processes. We estimate the lower limit on the K–H instability linear growth rate to be ∼5.84 × 10−3 s−1. For these vortices, we estimate the instantaneous atmospheric ion escape flux due to the detachment of plasma clouds during the late non‐linear stage of K–H instability to be ∼5.90 × 1026 particles/s. Extrapolation of loss rates integrated across time and space will require further work.

Post-translational protein deimination signatures in sea lamprey (Petromyzon marinus) plasma and plasma-extracellular vesicles

Lampreys are a jawless vertebrate species belonging to an ancient vertebrate lineage that diverged from a common ancestor with humans ~500 million years ago. The sea lamprey (Petromyzon marinus) has a filter feeding ammocoete larval stage that metamorphoses into a parasitic adult, feeding both on teleost and elasmobranch fish. Lampreys are a valuable comparative model species for vertebrate immunity and physiology due to their unique phylogenetic position, unusual adaptive immune system, and physiological adaptions such as tolerance to salinity changes and urea.Peptidylarginine deiminases (PADs) are a phylogenetically conserved enzyme family which catalyses post-translational deimination/citrullination in target proteins, enabling proteins to gain new functions (moonlighting). The identification of deiminated protein targets in species across phylogeny may provide novel insights into post-translational regulation of physiological and pathophysiological processes. Extracellular vesicles (EVs) are membrane vesicles released from cells that carry cargos of small molecules and proteins for cellular communication, involved in both normal and pathological processes. The current study identified deimination signatures in proteins of both total plasma and plasma-EVs in sea lamprey and furthermore reports the first characterisation of plasma-EVs in lamprey. EVs were poly-dispersed in the size range of 40–500 nm, similar to what is observed in other taxa, positive for CD63 and Flotillin-1. Plasma-EV morphology was confirmed by transmission electron microscopy. Assessment of deimination/citrullination signatures in lamprey plasma and plasma-EVs, revealed 72 deimination target proteins involved in immunity, metabolism and gene regulation in whole plasma, and 37 target proteins in EVs, whereof 24 were shared targets. Furthermore, the presence of deiminated histone H3, indicative of gene-regulatory mechanisms and also a marker of neutrophil extracellular trap formation (NETosis), was confirmed in lamprey plasma. Functional protein network analysis revealed some differences in KEGG and GO pathways of deiminated proteins in whole plasma compared with plasma-EVs. For example, while common STRING network clusters in plasma and plasma-EVs included Peptide chain elongation, Viral mRNA translation, Glycolysis and gluconeogenesis, STRING network clusters specific for EVs only included: Cellular response to heat stress, Muscle protein and striated muscle thin filament, Nucleosome, Protein processing in endoplasmic reticulum, Nucleosome and histone deacetylase complex. STRING network clusters specific for plasma were: Adipokinetic hormone receptor activity, Fibrinogen alpha/beta chain family, peptidase S1A, Glutathione synthesis and recycling-arginine, Fructose 1,6-bisphosphate metabolic process, Carbon metabolism and lactate dehydrogenase activity, Post-translational protein phosphorylation, Regulation of insulin-like growth factor transport and clotting cascade. Overall, for the EV citrullinome, five STRING network clusters, 10 KEGG pathways, 15 molecular GO pathways and 29 Reactome pathways were identified, compared with nine STRING network clusters, six KEGG pathways, two Molecular GO pathways and one Reactome pathway specific for whole plasma; while further pathways were shared. The reported findings indicate that major pathways relevant for immunity and metabolism are targets of deimination in lamprey plasma and plasma-EVs, with some differences, and may help elucidating roles for the conserved PAD enzyme family in regulation of immune and metabolic function throughout phylogeny.

Circulating miRNome of Trachemysscripta after elective gonadectomy under general anesthesia

Post-surgical management is an important issue in veterinary medicine, requiring biomarkers with high sensitivity and specificity for timely and effective treatment. Emerging evidence suggests that miRNAs are promising stress- and pain-related markers. The aims were to profile the circulating miRNA signature in plasma of turtles (Trachemysscripta) and point out potential candidate biomarkers to assess the status of the animal. The plasma of female turtles underwent surgical gonadectomy were collected 24 h pre-surgery, and 2.5 h and 36 h post-surgery. The expression of miRNAs was profiled by Next Generation Sequencing and the dysregulated miRNAs were validated using RT-qPCR. The diagnostic value of miRNAs was calculated by ROC curves. The results showed that 14 miRNAs were differentially expressed over time. RT-qPCR validation highlighted that 2-miR-499-3p and miR-203-5p-out of 8 miRNAs tested were effectively modulated. The Area Under the Curve (AUC) of miR-203-5p was fair (AUC 0.7934) in discriminating pre- and 36 h post-surgery samples and poor for other time points; the AUC of miR-499-3p was excellent (AUC 0.944) in discriminating pre-surgery and 2.5 h post-surgery samples, and fair in discriminating pre-surgery and 36 h post-surgery (AUC 0.7292) and 2.5 h and 36 h post-surgery (AUC 0.7569) samples. In conclusion, we demonstrated for the first time that miRNAs profile changes in plasma of turtles underwent surgical oophorectomy and identified miR-203-5p and miR-499-3p as potential candidate biomarkers to assess animals' status. Further studies are necessary to confirm their diagnostic value and to investigate functional and mechanistic networks to improve our understanding of the biological processes.

Abstract 2606: A nCounter-Based mRNA signature in plasma associates with localized non-small cell lung cancer

Abstract Background: 80% of non-small cell lung cancer (NSCLC) cases are diagnosed at stages IIIB-IV and have a dismal prognosis with a median life expectancy that does not exceed 2 years. In contrast, patients diagnosed at early and locally advanced stages (I-IIIA) can undergo surgery and have the potential to be totally cured. Imaging technologies often detect lung nodules of unknown significance that pose a diagnostic challenge; some patients with benign nodules are submitted to unnecessary surgical interventions while others with small tumors are just kept in observation, risking a significant delay for treatment. A diagnostic test that could differentiate between benign and malignant masses would be of great help in this setting. Methods: Circulating-free RNA (cfRNA) was isolated from the plasma of healthy individuals (N=21), early(I-II) stage (N=22) and stage IIIA (N=12) NSCLC patients, using an automatic extraction method(Qiasymphony, Qiagen). Purified cfRNA was quantified using Qubit, retrotranscribed and pre-amplified (14cycles) using the Low RNA Input Amplification kit (NanoString Technologies). Gene expression analysis was performed on the nCounter platform using the PanCancer IO360TM (NanoString Technologies), which can detect 770 transcripts related to tumor biology, micro-environment and the immune system. Results: Gene expression analysis revealed differential patterns for some cf-mRNAs from localized stage NSCLC patients versus healthy controls. A bioinformatics recursive feature elimination algorithm selected a 16-gene mRNA signature that was able to distinguish between localized NSCLC and control samples with an area under the ROC curve of 0.91 to 0.95. Furthermore, the signature scores derived from the algorithm were significantly different between the two cohorts. Conclusions: We have found an 16-gene signature that can differentiate between cfRNA of localized stages NSCLC patients and control individuals. Our results warrant validation studies in larger cohorts. Citation Format: Ana Giménez Capitán, Jillian Bracht, Nicolas Potie, María González-Cao, Santiago Viteri, Alejandro Martínez-Bueno, Carlos Cabrera-Gálvez, Pablo Rubinstein, Clara Mayo-de-las-Casas, Joselyn Valarezo, Chung-Ying Huang, Carlos Pedraz, Richard Boykind, Sarah Warren, Rafael Rosell, Miguel Ángel Molina-Vilaa, Andrés Aguilar-Hernández. A nCounter-Based mRNA signature in plasma associates with localized non-small cell lung cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 2606.

Identification of a Two-MicroRNA Signature in Plasma as a Novel Biomarker for Very Early Diagnosis of Breast Cancer

Simple SummaryBreast cancer diagnosis at the initial stage of the disease considerably improves prognosis and survival rates. This retrospective study aimed to develop and validate a plasma microRNA signature as a non-invasive biomarker for early-stage breast cancer diagnosis. We confirmed in a testing cohort of 54 BC patients and 89 healthy volunteers the value of a signature based on miR-30b and miR-99a levels in plasma samples for stage I breast cancer detection. Furthermore, our results were blindly validated in a second cohort of 74 breast cancer and 74 healthy samples. The proposed microRNA signature presented high value as a fast, cost-effective, and non-invasive biomarker for early-stage breast cancer detection, which will lead to a better prognosis for breast cancer patients.The early diagnosis of breast cancer is essential to improve patients’ survival rate. In this context, microRNAs have been described as potential diagnostic biomarkers for breast cancer. Particularly, circulating microRNAs have a strong value as non-invasive biomarkers. Herein, we assessed the potential of a microRNA signature based on miR-30b-5p and miR-99a-5p levels in plasma as a diagnostic biomarker for breast cancer. This two-microRNA signature was constructed by Principal Component Analysis and its prognostic value was assessed in a discovery cohort and blindly validated in a second cohort from an independent institution. ROC curve analysis and biomarker performance parameter evaluation demonstrated that our proposed signature presents a high value as a non-invasive biomarker for very early detection of breast cancer. In addition, pathway enrichment analysis identified three of the well-known pathways involved in cancer as targets of the two microRNAs.

Single-cell BCR and transcriptome analysis after influenza infection reveals spatiotemporal dynamics of antigen-specific B cells.

SummaryB cell responses are critical for antiviral immunity. However, a comprehensive picture of antigen-specific B cell differentiation, clonal proliferation, and dynamics in different organs after infection is lacking. Here, by combining single-cell RNA and B cell receptor (BCR) sequencing of antigen-specific cells in lymph nodes, spleen, and lungs after influenza infection in mice, we identify several germinal center (GC) B cell subpopulations and organ-specific differences that persist over the course of the response. We discover transcriptional differences between memory cells in lungs and lymphoid organs and organ-restricted clonal expansion. Remarkably, we find significant clonal overlap between GC-derived memory and plasma cells. By combining BCR-mutational analyses with monoclonal antibody (mAb) expression and affinity measurements, we find that memory B cells are highly diverse and can be selected from both low- and high-affinity precursors. By linking antigen recognition with transcriptional programming, clonal proliferation, and differentiation, these finding provide important advances in our understanding of antiviral immunity.

Proteotyping of knockout mouse strains reveals sex- and strain-specific signatures in blood plasma

We proteotyped blood plasma from 30 mouse knockout strains and corresponding wild-type mice from the International Mouse Phenotyping Consortium. We used targeted proteomics with internal standards to quantify 375 proteins in 218 samples. Our results provide insights into the manifested effects of each gene knockout at the plasma proteome level. We first investigated possible contamination by erythrocytes during sample preparation and labeled, in one case, up to 11 differential proteins as erythrocyte originated. Second, we showed that differences in baseline protein abundance between female and male mice were evident in all mice, emphasizing the necessity to include both sexes in basic research, target discovery, and preclinical effect and safety studies. Next, we identified the protein signature of each gene knockout and performed functional analyses for all knockout strains. Further, to demonstrate how proteome analysis identifies the effect of gene deficiency beyond traditional phenotyping tests, we provide in-depth analysis of two strains, C8a−/− and Npc2+/−. The proteins encoded by these genes are well-characterized providing good validation of our method in homozygous and heterozygous knockout mice. Ig alpha chain C region, a poorly characterized protein, was among the differentiating proteins in C8a−/−. In Npc2+/− mice, where histopathology and traditional tests failed to differentiate heterozygous from wild-type mice, our data showed significant difference in various lysosomal storage disease-related proteins. Our results demonstrate how to combine absolute quantitative proteomics with mouse gene knockout strategies to systematically study the effect of protein absence. The approach used here for blood plasma is applicable to all tissue protein extracts.

Comprehensive Plasma Metabolomic Profile of Patients with Advanced Neuroendocrine Tumors (NETs). Diagnostic and Biological Relevance.

Simple SummaryMetabolic flexibility is one of the key hallmarks of cancer and metabolites are the final products of this adaptation, reflecting the aberrant changes of tumors. However, the metabolic plasticity of each cancer type is still unknown, and specifically to date, there are no data on metabolic profile in neuroendocrine tumors. The aim of our retrospective study was to assess the metabolomic profile of NET patients to understand metabolic deregulation in these tumors and identify novel biomarkers with clinical potential. We provided, for the first time, a comprehensive metabolic profile of NET patients and identifies a distinctive metabolic signature in plasma of potential clinical use, selecting a reduced set of metabolites of high diagnostic accuracy. We have identified 32 novel enriched metabolic pathways in NETs related with the TCA cycle, and with arginine, pyruvate or glutathione metabolism, which have distinct implications in oncogenesis and may open innovative avenues of clinical research.Purpose: High-throughput “-omic” technologies have enabled the detailed analysis of metabolic networks in several cancers, but NETs have not been explored to date. We aim to assess the metabolomic profile of NET patients to understand metabolic deregulation in these tumors and identify novel biomarkers with clinical potential. Methods: Plasma samples from 77 NETs and 68 controls were profiled by GC−MS, CE−MS and LC−MS untargeted metabolomics. OPLS-DA was performed to evaluate metabolomic differences. Related pathways were explored using Metaboanalyst 4.0. Finally, ROC and OPLS-DA analyses were performed to select metabolites with biomarker potential. Results: We identified 155 differential compounds between NETs and controls. We have detected an increase of bile acids, sugars, oxidized lipids and oxidized products from arachidonic acid and a decrease of carnitine levels in NETs. MPA/MSEA identified 32 enriched metabolic pathways in NETs related with the TCA cycle and amino acid metabolism. Finally, OPLS-DA and ROC analysis revealed 48 metabolites with diagnostic potential. Conclusions: This study provides, for the first time, a comprehensive metabolic profile of NET patients and identifies a distinctive metabolic signature in plasma of potential clinical use. A reduced set of metabolites of high diagnostic accuracy has been identified. Additionally, new enriched metabolic pathways annotated may open innovative avenues of clinical research.

Placenta Accreta Spectrum: Biomarker Discovery Using Plasma Proteomics

(Am J Obstet Gynecol. 2020;223:433.e1–14) Placenta accreta spectrum (PAS), which includes placenta accreta, placenta increta and placenta percreta, often leads to hysterectomy and large-volume maternal hemorrhage and is a major cause of maternal mortality and morbidity. While maternal outcomes can be improved with planning after an accurate diagnosis, more than 50% of pregnancies complicated by PAS are not diagnosed prior to delivery. A method of reliable, early diagnosis of PAS is needed to improve maternal outcomes. This study used a novel aptamer-based platform for biomarker discovery to explore whether women who experience PAS have a unique protein signature in maternal plasma during the third trimester that may be used to diagnose PAS.

Cellular and plasma proteomic determinants of COVID-19 and non-COVID-19 pulmonary diseases relative to healthy aging.

We examine the cellular and soluble determinants of coronavirus disease 2019 (COVID-19) relative to aging by performing mass cytometry in parallel with clinical blood testing and plasma proteomic profiling of ~4,700 proteins from 71 individuals with pulmonary disease and 148 healthy donors (25-80 years old). Distinct cell populations were associated with age (GZMK+CD8+ T cells and CD25low CD4+ T cells) and with COVID-19 (TBET-EOMES- CD4+ T cells, HLA-DR+CD38+ CD8+ T cells and CD27+CD38+ B cells). A unique population of TBET+EOMES+ CD4+ T cells was associated with individuals with COVID-19 who experienced moderate, rather than severe or lethal, disease. Disease severity correlated with blood creatinine and urea nitrogen levels. Proteomics revealed a major impact of age on the disease-associated plasma signatures and highlighted the divergent contribution of hepatocyte and muscle secretomes to COVID-19 plasma proteins. Aging plasma was enriched in matrisome proteins and heart/aorta smooth muscle cell-specific proteins. These findings reveal age-specific and disease-specific changes associated with COVID-19, and potential soluble mediators of the physiological impact of COVID-19.

Effects of oxygen plasma treatment on Cd[formula omitted]Zn[formula omitted]Te material and devices

Surface passivation in detectors is designed to improve the performance and stabilize the characteristics over time and ambient. In Cd1−xZnxTe radiation detectors, oxidation by plasma is a well-known passivation method, but its physics is not fully understood. This study focuses on the macroscopic and microscopic effects of plasma treatment. It is shown that plasma oxidation before contact deposition causes a considerable decrease in the device’s dark current and that this reduction is in a strong correlation with the lowering of the surface potential. X-ray photoelectron spectroscopy (XPS) measurements revealed a significant increase in TeO 2 fraction on the plasma-treated surface. The main conclusion of this study is that the plasma-related effects are not confined to the surface layer. Furthermore, the deep penetration of plasma treatment byproduct takes place over long time periods, which may lead to variations in device characteristics. The effects were corroborated by several characterization techniques. DC current–voltage measurements combined with layer-by-layer stripping indicate plasma signature deep into the bulk. Transient current technique (TCT) results demonstrate that the plasma treatment affects the electric field distribution millimeters into the bulk. Photo-induced current transient spectroscopy (PICTS) and thermoelectric emission spectroscopy (TEES) measurements reveal plasma-related traps in-depth of the CdZnTe crystals.

Discovery and Validation of a Three-Cytokine Plasma Signature as a Biomarker for Diagnosis of Pediatric Tuberculosis.

Pediatric TB poses challenge in diagnosis due to the paucibacillary nature of the disease. We conducted a prospective diagnostic study to identify immune biomarkers of pediatric TB and controls (discovery cohort) and obtained a separate “validation” cohort of confirmed cases of pediatric TB and controls. Multiplex ELISA was performed to examine the plasma levels of cytokines. Discovery and validation cohorts revealed that baseline plasma levels of IFNγ, TNFα, IL-2, and IL-17A were significantly higher in active TB (confirmed TB and unconfirmed TB) in comparison to unlikely TB children. Receiver operating characteristics (ROC) curve analysis revealed that IFNγ, IL-2, TNFα, and IL-17A (in the discovery cohort) and TNFα and IL-17A (in the validation cohort) could act as biomarkers distinguishing confirmed or unconfirmed TB from unlikely TB with the sensitivity and specificity of more than 90%. In the discovery cohort, cytokines levels were significantly diminished following anti-tuberculosis treatment. In both the cohorts, combiROC models offered 100% sensitivity and 98% to 100% specificity for a three-cytokine signature of TNFα, IL-2, and IL-17A, which can distinguish confirmed or unconfirmed TB children from unlikely TB. Thus, a baseline cytokine signature of TNFα, IL-2, and IL-17A could serve as an accurate biomarker for the diagnosis of pediatric tuberculosis.

Implementing the MULTI-VP coronal model in EUHFORIA: Test case results and comparisons with the WSA coronal model

Context.In this study, we focus on improving EUHFORIA (European Heliospheric Forecasting Information Asset), a recently developed 3D magnetohydrodynamics space weather prediction tool. The EUHFORIA model consists of two parts covering two spatial domains: the solar corona and the inner heliosphere. For the first part, the semiempirical Wang-Sheeley-Arge (WSA) model is used by default; this model employs the potential field source surface and Schatten current sheet models to provide the necessary solar wind plasma and magnetic conditions above the solar surface, at 0.1 AU, which serve as boundary conditions for the inner heliospheric part. Herein, we present the first results of the implementation of an alternative coronal model in EUHFORIA, the so-called MULTI-VP model.Aims.After we replace the default EUHFORIA coronal setup with the MULTI-VP model, we compare their outputs both at 0.1 AU and 1 AU, for test cases involving high speed wind streams (HSSs). We select two distinct cases in which the standard EUHFORIA setup failed to reproduce the HSS plasma and magnetic signatures at Earth to test the performance of MULTI-VP coupled with EUHFORIA-heliosphere.Methods.To understand the quality of modeling with MULTI-VP in comparison with the default coronal model in EUHFORIA, we considered one HSS case during a period of low solar activity and another one during a period of high solar activity. Moreover, the modeling of the two HSSs was performed by employing magnetograms from different providers: one from the Global Oscillation Network Group (GONG) and the second from theWilcoxSpace Observatory (WSO). This way, we were able to distinguish differences arising not only because of the different models but also because of different magnetograms.Results.The results indicate that when employing a GONG magnetogram, the combination MULTI-VP+EUHFORIA-heliosphere reproduces the majority of HSS plasma and magnetic signatures measured at L1. On the contrary, the standard WSA+EUHFORIA-heliosphere combination does not capture the arrival of the HSS cases at L1. When employing WSO magnetograms, MULTI-VP+EUHFORIA-heliosphere reproduces the HSS that occurred during the period of high solar activity. However, it is unclear if it models the HSS during the period of low solar activity. For the same magnetogram and periods of time, WSA+EUHFORIA-heliosphere is not able to capture the HSSs of interest.Conclusions.The results show that the accuracy of the simulation output at Earth depends on the choice of both the coronal model and input magnetogram. Nevertheless, a more extensive statistical analysis is necessary to determine how precisely these choices affect the quality of the solar wind predictions.

Post-Translational Protein Deimination Signatures in Plasma and Plasma EVs of Reindeer (Rangifer tarandus).

Simple SummaryReindeer are an important wild and domesticated species of the Arctic, Northern Europe, Siberia and North America. As reindeer have developed various strategies to adapt to extreme environments, this makes them an interesting species for studies into diversity of immune and metabolic functions in the animal kingdom. Importantly, while reindeer carry natural infections caused by viruses (including coronaviruses), bacteria and parasites, they can also act as carriers for transmitting such diseases to other animals and humans, so called zoonosis. Reindeer are also affected by chronic wasting disease, a neuronal disease caused by prions, similar to scrapie in sheep, mad cows disease in cattle and Creutzfeldt-Jakob disease in humans. The current study assessed a specific protein modification called deimination/citrullination, which can change how proteins function and allow them to take on different roles in health and disease processes. Profiling of deiminated proteins in reindeer showed that many important pathways for immune defenses, prion diseases and metabolism are enriched in deiminated proteins, both in plasma, as well as in plasma extracellular vesicles. This study provides a platform for the development of novel biomarkers to assess wild life health status and factors relating to zoonotic disease.The reindeer (caribou) Rangifer tarandus is a Cervidae in the order Artiodactyla. Reindeer are sedentary and migratory populations with circumpolar distribution in the Arctic, Northern Europe, Siberia and North America. Reindeer are an important wild and domesticated species, and have developed various adaptive strategies to extreme environments. Importantly, deer have also been identified to be putative zoonotic carriers, including for parasites, prions and coronavirus. Therefore, novel insights into immune-related markers are of considerable interest. Peptidylarginine deiminases (PADs) are a phylogenetically conserved enzyme family which causes post-translational protein deimination by converting arginine into citrulline in target proteins. This affects protein function in health and disease. Extracellular vesicles (EVs) participate in cellular communication, in physiological and pathological processes, via transfer of cargo material, and their release is partly regulated by PADs. This study assessed deiminated protein and EV profile signatures in plasma from sixteen healthy wild female reindeer, collected in Iceland during screening for parasites and chronic wasting disease. Reindeer plasma EV profiles showed a poly-dispersed distribution from 30 to 400 nm and were positive for phylogenetically conserved EV-specific markers. Deiminated proteins were isolated from whole plasma and plasma EVs, identified by proteomic analysis and protein interaction networks assessed by KEGG and GO analysis. This revealed a large number of deimination-enriched pathways for immunity and metabolism, with some differences between whole plasma and EVs. While shared KEGG pathways for whole plasma and plasma EVs included complement and coagulation pathways, KEGG pathways specific for EVs were for protein digestion and absorption, platelet activation, amoebiasis, the AGE–RAGE signaling pathway in diabetic complications, ECM receptor interaction, the relaxin signaling pathway and the estrogen signaling pathway. KEGG pathways specific for whole plasma were pertussis, ferroptosis, SLE, thyroid hormone synthesis, phagosome, Staphylococcus aureus infection, vitamin digestion and absorption, and prion disease. Further differences were also found between molecular function and biological processes GO pathways when comparing functional STRING networks for deiminated proteins in EVs, compared with deiminated proteins in whole plasma. This study highlights deiminated proteins and EVs as candidate biomarkers for reindeer health and may provide information on regulation of immune pathways in physiological and pathological processes, including neurodegenerative (prion) disease and zoonosis.

Different miRNA Profiles in Plasma Derived Small and Large Extracellular Vesicles from Patients with Neurodegenerative Diseases

Identifying biomarkers is essential for early diagnosis of neurodegenerative diseases (NDs). Large (LEVs) and small extracellular vesicles (SEVs) are extracellular vesicles (EVs) of different sizes and biological functions transported in blood and they may be valid biomarkers for NDs. The aim of our study was to investigate common and different miRNA signatures in plasma derived LEVs and SEVs of Alzheimer’s disease (AD), Parkinson’s disease (PD), Amyotrophic Lateral Sclerosis (ALS) and Fronto-Temporal Dementia (FTD) patients. LEVs and SEVs were isolated from plasma of patients and healthy volunteers (CTR) by filtration and differential centrifugation and RNA was extracted. Small RNAs libraries were carried out by Next Generation Sequencing (NGS). MiRNAs discriminate all NDs diseases from CTRs and they can provide a signature for each NDs. Common enriched pathways for SEVs were instead linked to ubiquitin mediated proteolysis and Toll-like receptor signaling pathways and for LEVs to neurotrophin signaling and Glycosphingolipid biosynthesis pathway. LEVs and SEVs are involved in different pathways and this might give a specificity to their role in the spreading of the disease. The study of common and different miRNAs transported by LEVs and SEVs can be of great interest for biomarker discovery and for pathogenesis studies in neurodegeneration.

Abstract PS2-17: Identification of cell-free microRNA signatures in pre-operative plasma from breast cancer patients

Abstract Background: Breast cancer (BC) is a heterogeneous disease and difficult to treat once metastasis occurs. Thus, determining blood molecular biomarkers that allow for early diagnosis would help to identify BC patients at higher risk. MicroRNAs (miRs) are short sequences (18-22 base pairs) and promising candidates as biomarkers due to their high stability in blood. Using a direct blood cell-free miR (cfmiR) next generation sequencing (NGS) -based assay that requires no extraction, we assessed 2083 cfmiRs in plasma samples from BC patients. We hypothesize that specific cfmiR signatures detected in plasma from BC patients are associated with early BC diagnosis. Those cfmiRs represent unique patterns that may allow for monitoring BC tumor burden and early detection.Methods: Two cohorts of pre-operative plasma samples from BC patients (n= 158), one cohort of pre-operative plasma samples from patients with BC brain metastasis (BCBM, n= 5), and a cohort of female normal donors’ plasma samples (n=20) were assessed by HTG EdgeSeq miR whole transcriptome assay. All primary BC tumors were histopathology staged (TNM, AJCC 8th) after surgery and divided into two different cohorts: the BC-1 cohort (n= 80, including stage I= 39, II= 37, and III= 4 patients), and the BC-2 cohort (n= 78, including stage I= 42, II= 33, and III= 3 patients). All of the samples were analyzed using DESeq2 normalization and interrogated for differentially expressed (DE) miRs using a cut-off of 30 counts, FC>1.2 or <-1.2, and FDR (p<0.05). We utilized the BC-1 group as the training cohort and the BC-2 and the BCBM cohorts were used to validate the observations in BC-1. Results: In a retrospective study the cfmiR profiles obtained from BC-1 patients were compared to the cfmiR profiles obtained from normal donors’ plasma samples. Briefly, 328 cfmiRs were DE in plasma samples from the BC-1 cohort compared to normal donors’ plasma samples, of which 184 were upregulated and 144 were downregulated. To validate our findings, we screened the plasma samples from the BC-2 cohort (n=78). The results showed that 181 of 361 DE cfmiRs were downregulated and 180 were upregulated. By comparing both BC cohorts we found 269 DE cfmiRs consistently changing, which included 82% and 74.5% of the cfmiRs identified in BC-1 and BC-2 cohorts, respectively. In addition, we compared the cfmiR expression of BCBM (n=5) and normal donors’ plasma samples. Of the 300 DE cfmiRs, 30 were downregulated and 270 were upregulated. By integrative analysis, a 59 cfmiR signature observed in BCBM plasma samples was consistently identified in both BC-1 and BC-2 cohorts. Also, 172 cfmiRs were found exclusively in BCBM and not in primary BC tumors. Receiving operative characteristic curve analysis showed that the 59 cfmiR signature was able to distinguish primary BC from female normal donors’ (BC-1 AUC= 0.910, p<0.05 and BC-2 AUC= 0.922, p<0.05, respectively). Using the 59 cfmiR signature, the first five components of the principal component analysis showed 85.26% and 85.27% mean cumulative variance for BC-1 and BC-2 respectively.Conclusions: Specific cfmiR patterns were associated with primary BC tumors and may have potential utility for early BC diagnosis, including disease recurrence. A 59 cfmiR signature detected in primary BC patients was able to detect BCBM and may have potential clinical utility in monitoring patients at higher risk of metastasis. Further analysis are in progress to validate our observations. Citation Format: Matias Alberto Bustos, Negin Rahimzadeh, Xiaoqing Zhang, Kevin D. Tran, Grace Chang, Carol Nishikubo, Dave S.B. Hoon. Identification of cell-free microRNA signatures in pre-operative plasma from breast cancer patients [abstract]. In: Proceedings of the 2020 San Antonio Breast Cancer Virtual Symposium; 2020 Dec 8-11; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2021;81(4 Suppl):Abstract nr PS2-17.

Identification and validation of a multivariable prediction model based on blood plasma and serum metabolomics for the distinction of chronic pancreatitis subjects from non-pancreas disease control subjects

ObjectiveChronic pancreatitis (CP) is a fibroinflammatory syndrome leading to organ dysfunction, chronic pain, an increased risk for pancreatic cancer and considerable morbidity. Due to a lack of specific biomarkers, diagnosis...

Radial Evolution of a CIR: Observations From a Nearly Radially Aligned Event Between Parker Solar Probe and STEREO‐A

AbstractThe addition of Parker Solar Probe (PSP) to the Heliophysics System Observatory has allowed for the unprecedented ability to study Corotating Interaction Regions (CIRs) at multiple radial distances without significant temporal/longitudinal variations. On September 19, 2019, PSP observed a CIR at ∼0.5 au when it was nearly radially aligned with the Solar Terrestrial Relations Observatory‐Ahead (STEREO‐A) spacecraft at ∼1 au, allowing for an unambiguous assessment of the radial evolution of a single CIR. Bulk plasma and magnetic field signatures of the CIR evolve in a fashion characteristic to previous observations; however, the suprathermal ions are enhanced over a larger longitudinal range at PSP than at STEREO‐A, although at much lower intensities. The longitudinal spread appears to be largely a consequence of magnetic field line topology at CIRs between the compressed slow solar wind upstream and high‐speed stream following the CIR, underscoring the importance of the large‐scale topology of these structures.

MicroRNA Signatures in Plasma of Patients With Venous Thrombosis: A Preliminary Report

BackgroundVenous thromboembolism (VTE) is a frequent and potentially fatal disease, but its pathophysiology is incompletely understood. microRNAs (miR) dysregulate hemostatic proteins and influence thrombotic pathology by posttranscriptional regulation of gene expression. Consensus in defining VTE-related miR clusters and functionally relevant miR has not been reached. We aimed to generate a miR database in patients at high thrombotic risk of VTE and explored their biological functions by seeking information on their messenger RNA targets. MethodsBy high-throughput screening (Affymetrix miRNA Microarray), we identified 159 miR in venous blood of male patients who had two unprovoked VTE and in age-matched male controls. ResultsOf the 159 miR, 41 were significantly higher expressed in patients compared to controls. Six miR (hsa-miR-6798-3p, hsa-miR-6789-5p hsa-miR-4651, hsa-miR-6765-5p, hsa-miR-6816-5p, hsa-miR-4734) were modulated ≥ 5.0-fold higher. Higher expression levels of 4 of these miR (hsa-miR-6789-5p, hsa-miR-4651, hsa-miR-6765-5p, and hsa-miR-6816-5p; primers were unavailable for hsa-miR-6798-3p and hsa-miR-4734) were confirmed by quantitative real-time polymerase chain reaction in 10 independent patients and 10 control samples. Ingenuity Pathway Analysis identified 23 altered miR including hsa-miR-6789-5p, hsa-miR-4651, hsa-miR-6765-5p and hsa-miR-4734 as the main regulators of messenger RNAs involved in the pathology of VTE. Seven messenger RNA targets including thrombomodulin and four targets related to platelet function had a direct relationship to 4 identified miR. ConclusionsWe provide evidence of distinct, independently validated miR signatures in patients with VTE and identified a subset of miR as main regulators of messenger RNA involved in disorders related to pathophysiologic processes in venous thrombosis development.