A novel multiple marker microarray analyzer and methodology to predict major obstetric syndromes using surface markers of circulating extracellular vesicles from maternal plasma.

Introduction: Placental-derived extracellular vesicles (EVs) are nano-organelles that facilitate intercellular communication between the feto-placental unit and the mother. We evaluated a novel Multiple Microarray analyzer for identifying surface markers on plasma EVs that predict preterm delivery and preeclampsia compared to term delivery controls.

Material And Methods: In this prospective exploratory cohort study pregnant women between 24 and 40 gestational weeks with preterm delivery (n = 16), preeclampsia (n = 19), and matched term delivery controls (n = 15) were recruited from Bnai Zion Medical Center, Haifa, Israel.

New Enhancing MRI Lesions Associate with IL-17, Neutrophil Degranulation and Integrin Microparticles: Multi-Omics Combined with Frequent MRI in Multiple Sclerosis.

Background: Blood-barrier (BBB) breakdown and active inflammation are hallmarks of relapsing multiple sclerosis (RMS), but the molecular events contributing to the development of new lesions are not well explored. Leaky endothelial junctions are associated with increased production of endothelial-derived extracellular microvesicles (EVs) and result in the entry of circulating immune cells into the brain. MRI with intravenous gadolinium (Gd) can visualize acute blood-barrier disruption as the initial event of the evolution of new lesions.

Increased extracellular vesicles (EVs) related to T cell-mediated inflammation and vascular function in familial hypercholesterolemia.

Background And Aims: OxLDL modulates innate and adaptive immunity, and extracellular vesicles (EVs) released from both non-immune and immune cells are proposed key players in atherosclerosis development. In the present study, we aimed to investigate EVs expressing markers related to adaptive immunity-driven inflammation and endothelial activation/dysfunction in hypercholesterolemic patients.

Methods: EVs were phenotyped in thirty patients with familial hypercholesterolemia (FH) and twenty-three healthy controls using the Extracellular Vesicle (EV) Array with antibodies targeting proteins expressed on B and T cells, and endothelial cells.

Hepatocyte-derived biomarkers predict liver-related events at 2 years in Child-Pugh class A alcohol-related cirrhosis.

Background & Aims: In patients with compensated alcohol-related cirrhosis, reliable prognostic biomarkers are lacking. Keratin-18 and hepatocyte-derived large extracellular vesicle (lEV) concentrations reflect disease activity, but their ability to predict liver-related events is unknown.

Methods: We measured plasma keratin-18 and hepatocyte lEV concentrations in 500 patients with Child-Pugh class A alcohol-related cirrhosis.

The Prognostic Value of Plasma Small Extracellular Vesicles' Phenotype in Patients With Gastrointestinal Stromal Tumor.

Background/aim: For patients with local gastrointestinal stromal tumor (GIST), risk stratification is used to assess the prognosis and identify patients to offer adjuvant treatment. For patients with advanced or metastatic GIST, no such risk stratification exists. This study aimed to investigate the prognostic value of 31 different plasma small extracellular vesicles' (SEVs) surface proteins in GIST patients.

Comparative and integrated analysis of plasma extracellular vesicle isolation methods in healthy volunteers and patients following myocardial infarction.

Plasma extracellular vesicle (EV) number and composition are altered following myocardial infarction (MI), but to properly understand the significance of these changes it is essential to appreciate how the different isolation methods affect EV characteristics, proteome and sphingolipidome. Here, we compared plasma EV isolated from platelet-poor plasma from four healthy donors and six MI patients at presentation and 1-month post-MI using ultracentrifugation (UC), polyethylene glycol precipitation, acoustic trapping, size-exclusion chromatography (SEC) and immunoaffinity capture. The isolated EV were evaluated by Nanoparticle Tracking Analysis (NTA), Western blot, transmission electron microscopy (TEM), an EV-protein array, untargeted proteomics (LC-MS/MS) and targeted sphingolipidomics (LC-MS/MS).

Photometric method for dual targeting of surface and surface-associated proteins on extracellular vesicles in the multiparametric test.

Extracellular vesicles (EVs) have become a topic of interest within the field of diagnostic biomarkers; however, recent developments in the study of EVs have increased the need for simpler but still comprehensive methods for characterization. Here, we describe how to simultaneously measure several surface or surface-associated proteins on EVs using a multiparametric microarray-based analysis termed Extracellular Vesicle Array (EV Array), which is developed to catch and phenotypically characterize small EVs. Previously, this analysis has been limited to measuring only one fluorescent signal per analysis.

Profiling Blood Serum Extracellular Vesicles in Plaque Psoriasis and Psoriatic Arthritis Patients Reveals Potential Disease Biomarkers.

Psoriasis vulgaris (PsV) and psoriatic arthritis (PsA) are inflammatory diseases with unresolved pathophysiological aspects. Extracellular vesicles (EVs) play an important role in intercellular communication. We compared the miRNA contents and surface proteome of the EVs in the blood serum of PsV and PsA patients to healthy controls.

The Role of Plasma Extracellular Vesicles in Remote Ischemic Conditioning and Exercise-Induced Ischemic Tolerance.

Ischemic conditioning and exercise have been suggested for protecting against brain ischemia-reperfusion injury. However, the endogenous protective mechanisms stimulated by these interventions remain unclear. Here, in a comprehensive translational study, we investigated the protective role of extracellular vesicles (EVs) released after remote ischemic conditioning (RIC), blood flow restricted resistance exercise (BFRRE), or high-load resistance exercise (HLRE).

Rapid neutrophil mobilization by VCAM-1+ endothelial cell-derived extracellular vesicles.

Aims: Acute myocardial infarction rapidly increases blood neutrophils (<2 h). Release from bone marrow, in response to chemokine elevation, has been considered their source, but chemokine levels peak up to 24 h after injury, and after neutrophil elevation. This suggests that additional non-chemokine-dependent processes may be involved.

Optimization of High-Throughput Multiplexed Phenotyping of Extracellular Vesicles Performed in 96-Well Microtiter Plates.

Extracellular vesicles (EVs) are promising biomarkers for several diseases, however, no simple and robust methods exist to characterize EVs in a clinical setting. The EV Array analysis is based on a protein microarray platform, where antibodies are printed onto a solid surface that enables the capture of small EVs (sEVs) by their surface or surface-associated proteins. The EV Array analysis was transferred to an easily handled microtiter plate (MTP) format and a range of optimization experiments were performed within this study.

Extracellular Vesicles: An Important Biomarker in Recurrent Pregnancy Loss?

Recurrent pregnancy loss (RPL) has an estimated incidence of 1-3% of all couples. The etiology is considered to be multifactorial. Extracellular vesicles (EVs) take part in numerous different physiological processes and their contents show the originating cell and pathophysiological states in different diseases.

Cardioprotection by remote ischemic conditioning is transferable by plasma and mediated by extracellular vesicles.

Background: Remote ischemic conditioning (RIC) by brief periods of limb ischemia and reperfusion protects against ischemia-reperfusion injury. We studied the cardioprotective role of extracellular vesicles (EV)s released into the circulation after RIC and EV accumulation in injured myocardium.

Methods: We used plasma from healthy human volunteers before and after RIC (pre-PLA and post-PLA) to evaluate the transferability of RIC.

Molecular characteristics of porcine alpha-synuclein splicing variants.

Alpha-synuclein (α-syn) is a 140 amino acid, intrinsically disordered protein with a potential role in neurotransmitter vesicle release. The protein is natively unfolded under physiological conditions, and is expressed predominantly in neural tissue. α-syn is associated with neuropathological conditions in Parkinson's disease, where the protein misfolds into oligomers and fibrils resulting in aggregates in Lewy bodies.

Treatment with intravenous immunoglobulin increases the level of small EVs in plasma of pregnant women with recurrent pregnancy loss.

Extracellular vesicles (EVs), which are small cell-derived compartments, take part in numerous different physiological processes. The contents of EVs reveal the cell of origin and indicates pathophysiological states in different diseases. In pregnancy disorders, changes have been reported in the composition, bioactivity and concentration of placental and non-placental EVs.

Individually cultured bovine embryos produce extracellular vesicles that have the potential to be used as non-invasive embryo quality markers.

Extracellular vesicles (EVs) are membrane-bound biological nanoparticles (NPs) and have gained wide attention as potential biomarkers. We aimed to isolate and characterize EVs from media conditioned by individually cultured preimplantation bovine embryos and to assess their relationship with embryo quality. Presumptive zygotes were cultured individually in 60 μl droplets of culture media, and 50 μl of media were collected from the droplets either on day 2, 5 or 8 post-fertilization.

Blood flow-restricted resistance exercise alters the surface profile, miRNA cargo and functional impact of circulating extracellular vesicles.

Ischemic exercise conducted as low-load blood flow restricted resistance exercise (BFRE) can lead to muscle remodelling and promote muscle growth, possibly through activation of muscle precursor cells. Cell activation can be triggered by blood borne extracellular vesicles (EVs) as these nano-sized particles are involved in long distance signalling. In this study, EVs isolated from plasma of healthy human subjects performing a single bout of BFRE were investigated for their change in EV surface profiles and miRNA cargos as well as their impact on skeletal muscle precursor cell proliferation.

Surface Proteome of Plasma Extracellular Vesicles as Biomarkers for Pneumonia and Acute Exacerbation of Chronic Obstructive Pulmonary Disease.

Background: Community-acquired pneumonia (CAP) and acute exacerbation of chronic obstructive pulmonary disease (AECOPD) represent a major burden of disease and death and their differential diagnosis is critical. A potential source of relevant accessible biomarkers are blood-borne small extracellular vesicles (sEVs).

Methods: We performed an extracellular vesicle array to find proteins on plasma sEVs that are differentially expressed and possibly allow the differential diagnosis between CAP and AECOPD.

Altered Levels of Toll-Like Receptors in Circulating Extracellular Vesicles in Multiple Sclerosis.

Extracellular vesicles (EVs) are involved in inter-cellular communication and their cargo may provide prognostic/diagnostic biomarkers. To discover EV-associated biomarkers for Multiple Sclerosis (MS), we used an immune marker array to identify surface proteins on circulating EVs that differ between MS patients and controls (n = 3 each). We identified toll-like receptor-3 (TLR3) as a potential target for further validation.

Elevated blood plasma levels of tissue factor-bearing extracellular vesicles in patients with atrial fibrillation.

Background: The risk of thrombus formation in the left atrial appendage (LAA) in patients with atrial fibrillation (AF) may result from blood stasis, local endocardial changes, and/or changed blood composition. Extracellular vesicles (EVs), especially subtypes exposing tissue factor (TF), have procoagulant capacity. We hypothesized that blood concentrations of TF-bearing EVs and other procoagulant biomarkers are elevated in AF patients, particularly in the LAA lumen.

Postprandial Increase in Blood Plasma Levels of Tissue Factor-Bearing (and Other) Microvesicles Measured by Flow Cytometry: Fact or Artifact?

Tissue factor (TF)-bearing microvesicles (MVs) and exosomes may play a role in hemostasis and thrombosis. MVs may be quantified by flow cytometry (FC)-based detection of phosphatidylserine (PS)-positive submicron particles carrying specific antigens, although interference from lipoproteins complicates this approach. In this study, we evaluated the effect of food intake on blood levels of TF-bearing particles measured by FC and small extracellular vesicles (EVs) measured by a protein microarray-based test termed EV Array.

Induction of a Regulatory Phenotype in CD3+ CD4+ HLA-DR+ T Cells after Allogeneic Mixed Lymphocyte Culture; Indications of Both Contact-Dependent and -Independent Activation.

Although the observation of major histocompatibility complex II (MHCII) receptors on T cells is longstanding, the explanation for this occurrence remains enigmatic. Reports of an inducible, endogenous expression exist, as do studies demonstrating a protein acquisition from other cells by mechanisms including vesicle transfer. Irrespective of origin, the presence of the human MHCII isotype, human leukocyte antigen DR (HLA-DR), potentially identifies a regulatory T cell population.

Prospects and limitations of antibody-mediated clearing of lipoproteins from blood plasma prior to nanoparticle tracking analysis of extracellular vesicles.

: Nanoparticle tracking analysis (NTA) enables measurement of extracellular vesicles (EVs) but lacks the ability to distinct between EVs and lipoproteins which are abundantly present in blood plasma. Limitations in ultracentrifugation and size exclusion chromatography applied for EV isolation may result in inadequate EV purification and preservation. In this proof of concept study, we aimed to evaluate the potential of antibody-mediated removal of lipoproteins from plasma prior to extracellular vesicle (EV) analysis by nanoparticle tracking analysis (NTA).