Multiomics approaches disclose very-early molecular and cellular switches during insect-venom allergen-specific immunotherapy: an observational study.

Allergen-specific immunotherapy (AIT) induces immune tolerance, showing the highest success rate (>95%) for insect venom while a much lower chance for pollen allergy. However, the molecular switches leading to successful durable tolerance restoration remain elusive. The primary outcome of this observational study is the comprehensive immunological cellular characterization during the AIT initiation phase, whereas the secondary outcomes are the serological and Th2-cell-type-specific transcriptomic analyses.

Differential miRNA and Protein Expression Reveals miR-1285, Its Targets TGM2 and CDH-1, as Well as CD166 and S100A13 as Potential New Biomarkers in Patients with Diabetes Mellitus and Pancreatic Adenocarcinoma.

Early detection of PDAC remains challenging due to the lack of early symptoms and the absence of reliable biomarkers. The aim of the present project was to identify miRNA and proteomics signatures discriminating PDAC patients with DM from nondiabetic PDAC patients. Proteomics analysis and miRNA array were used for protein and miRNA screening.

High-dimensional immune profiles correlate with phenotypes of peanut allergy during food-allergic reactions.

Background: Food challenges carry a burden of safety, effort and resources. Clinical reactivity and presentation, such as thresholds and symptoms, are considered challenging to predict ex vivo.

Aims: To identify changes of peripheral immune signatures during oral food challenges (OFC) that correlate with the clinical outcome in patients with peanut allergy (PA).

Combinatorial analysis reveals highly coordinated early-stage immune reactions that predict later antiviral immunity in mild COVID-19 patients.

While immunopathology has been widely studied in patients with severe COVID-19, immune responses in non-hospitalized patients have remained largely elusive. We systematically analyze 484 peripheral cellular or soluble immune features in a longitudinal cohort of 63 mild and 15 hospitalized patients versus 14 asymptomatic and 26 household controls. We observe a transient increase of IP10/CXCL10 and interferon-β levels, coordinated responses of dominant SARS-CoV-2-specific CD4 and fewer CD8 T cells, and various antigen-presenting and antibody-secreting cells in mild patients within 3 days of PCR diagnosis.

Standard Peripheral Blood Mononuclear Cell Cryopreservation Selectively Decreases Detection of Nine Clinically Relevant T Cell Markers.

Biobanking is an operational component of various epidemiological studies and clinical trials. Although peripheral blood is routinely acquired and stored in biobanks, the effects of specimen processing on cell composition and clinically relevant functional markers of T cells still require a systematic evaluation. In this study, we assessed 25 relevant T cell markers in human PBMCs and showed that the detection of nine membrane markers (e.

Fitness for purpose of stabilized stool samples for bile acid metabolite analyses.

Biobanks and cohort studies are increasingly utilizing chemical stabilizers to collect and store stool samples for downstream DNA-based microbiome analyses. While stabilizers permit ambient-temperature collection and storage of samples for gut microbiome studies, the use of the same sample type for downstream metabolomics assays has not been explored. Microbiome-metabolomics analysis of fecal samples is increasingly getting attention to further elucidate the mechanisms by which the gut microbiota influences the host.

Proficiency Testing to Assess Technical Performance for CTC-Processing and Detection Methods in CANCER-ID.

Background: Multiple technologies are available for detection of circulating tumor cells (CTCs), but standards to evaluate their technical performance are still lacking. This limits the applicability of CTC analysis in clinic routine. Therefore, in the context of the CANCER-ID consortium, we established a platform to assess technical validity of CTC detection methods in a European multi-center setting using non-small cell lung cancer (NSCLC) as a model.

A Year in the Life of the EU-CardioRNA COST Action: CA17129 Catalysing Transcriptomics Research in Cardiovascular Disease.

The EU-CardioRNA Cooperation in Science and Technology (COST) Action is a European-wide consortium established in 2018 with 31 European country members and four associate member countries to build bridges between translational researchers from academia and industry who conduct research on non-coding RNAs, cardiovascular diseases and similar research areas. EU-CardioRNA comprises four core working groups (WG1-4). In the first year since its launch, EU-CardioRNA met biannually to exchange and discuss recent findings in related fields of scientific research, with scientific sessions broadly divided up according to WG.

Method Validation for Extraction of DNA from Human Stool Samples for Downstream Microbiome Analysis.

A formal method validation for biospecimen processing in the context of accreditation in laboratories and biobanks is lacking. A previously optimized stool processing protocol was validated for fitness-for-purpose for downstream microbiome analysis. DNA extraction from human stool was validated with various collection tubes, stabilizing solutions and storage conditions in terms of fitness-for-purpose for downstream microbiome analysis, robustness, and sample stability.

Multicenter Evaluation of Circulating Cell-Free DNA Extraction and Downstream Analyses for the Development of Standardized (Pre)analytical Work Flows.

Background: In cancer patients, circulating cell-free DNA (ccfDNA) can contain tumor-derived DNA (ctDNA), which enables noninvasive diagnosis, real-time monitoring, and treatment susceptibility testing. However, ctDNA fractions are highly variable, which challenges downstream applications. Therefore, established preanalytical work flows in combination with cost-efficient and reproducible reference materials for ccfDNA analyses are crucial for analytical validity and subsequently for clinical decision-making.

Multicenter Evaluation of Circulating Plasma MicroRNA Extraction Technologies for the Development of Clinically Feasible Reverse Transcription Quantitative PCR and Next-Generation Sequencing Analytical Work Flows.

Background: In human body fluids, microRNA (miRNA) can be found as circulating cell-free miRNA (cfmiRNA), as well as secreted into extracellular vesicles (EVmiRNA). miRNAs are being intensively evaluated as minimally invasive liquid biopsy biomarkers in patients with cancer. The growing interest in developing clinical assays for circulating miRNA necessitates careful consideration of confounding effects of preanalytical and analytical parameters.

Small Nucleolar RNA Score: An Assay to Detect Formalin-Overfixed Tissue.

Although there are millions of formalin-fixed paraffin-embedded (FFPE) tissue blocks potentially available for scientific research, many are of questionable quality, partly due to unknown fixation conditions. We analyzed FFPE tissue biospecimens as part of the NCI Biospecimen Preanalytical Variables (BPV) program to identify microRNA (miRNA) markers for fixation time. miRNA was extracted from kidney and ovary tumor FFPE blocks (19 patients, cold ischemia ≤2 hours) with 6, 12, 24, and 72 hours fixation times, then analyzed using the WaferGen SmartChip platform (miRNA chip with 1036 miRNA targets).

Intraindividual Temporal miRNA Variability in Serum, Plasma, and White Blood Cell Subpopulations.

Blood microRNAs (miRNAs) are ideal biomarkers, and blood derivatives are often collected in the scope of miRNA research projects. However, knowledge of temporal variations of miRNAs in healthy individuals is lacking. In this study, miRNA variability was measured over a 1-year period in different blood derivatives, collected every 2-3 months from two healthy donors.

A high rate of telomeric sister chromatid exchange occurs in chronic lymphocytic leukaemia B-cells.

Cancer cells protect their telomere ends from erosion through reactivation of telomerase or by using the Alternative Lengthening of Telomere (ALT) mechanism that depends on homologous recombination. Chronic lymphocytic leukaemia (CLL) B cells are characterized by almost no telomerase activity, shelterin deregulation and telomere fusions. To characterize telomeric maintenance mechanisms in B-CLL patients, we measured their telomere length, telomerase expression and the main hallmarks of the ALT activity i.

Blood DNA Yield but Not Integrity or Methylation Is Impacted After Long-Term Storage.

Collection of human whole blood for genomic DNA extraction is part of numerous clinical studies. Since DNA extraction cannot always be performed at the time of sample collection, whole blood samples may be stored for years before being processed. The use of appropriate storage conditions is then critical to obtain DNA in sufficient quantity and of adequate quality in order to obtain reliable results from the subsequent molecular biological analyses.

Exosomes released by chronic lymphocytic leukemia cells induce the transition of stromal cells into cancer-associated fibroblasts.

Exosomes derived from solid tumor cells are involved in immune suppression, angiogenesis, and metastasis, but the role of leukemia-derived exosomes has been less investigated. The pathogenesis of chronic lymphocytic leukemia (CLL) is stringently associated with a tumor-supportive microenvironment and a dysfunctional immune system. Here, we explore the role of CLL-derived exosomes in the cellular and molecular mechanisms by which malignant cells create this favorable surrounding.

Method optimization for fecal sample collection and fecal DNA extraction.

Background: This is the third in a series of publications presenting formal method validation for biospecimen processing in the context of accreditation in laboratories and biobanks. We report here optimization of a stool processing protocol validated for fitness-for-purpose in terms of downstream DNA-based analyses.

Methods: Stool collection was initially optimized in terms of sample input quantity and supernatant volume using canine stool.

Method validation for automated isolation of viable peripheral blood mononuclear cells.

Background: This article is part of a series of publications providing formal method validation for biospecimen processing in the context of accreditation in laboratories and biobanks. We report the optimization and validation for fitness-for-purpose of automated and manual protocols for isolating peripheral blood mononuclear cells (PBMCs) from whole blood, and compare the two methods.

Methods: The manual method was optimized for whole blood centrifugation speed, gradient type (Ficoll, Leucosep, CPT), and freezing method (Mr Frosty, Controlled Rate Freezing).

Platelet mitochondrial membrane potential in Parkinson's disease.

Objective: Mitochondrial dysfunction is a hallmark of idiopathic Parkinson's disease (IPD), which has been reported not to be restricted to striatal neurons. However, studies that analyzed mitochondrial function at the level of selected enzymatic activities in peripheral tissues have produced conflicting data. We considered the electron transport chain as a complex system with mitochondrial membrane potential as an integrative indicator for mitochondrial fitness.

Method validation for preparing urine samples for downstream proteomic and metabolomic applications.

Background: Formal validation of methods for biospecimen processing in the context of accreditation in laboratories and biobanks is lacking. A protocol for processing of a biospecimen (urine) was validated for fitness-for-purpose in terms of key downstream endpoints.

Methods: Urine processing was optimized for centrifugation conditions on the basis of microparticle counts at room temperature (RT) and at 4°C.

Method validation for preparing serum and plasma samples from human blood for downstream proteomic, metabolomic, and circulating nucleic acid-based applications.

Background: Formal method validation for biospecimen processing in the context of accreditation in laboratories and biobanks is lacking. Serum and plasma processing protocols were validated for fitness-for-purpose in terms of key downstream endpoints, and this article demonstrates methodology for biospecimen processing method validation.

Methods: Serum and plasma preparation from human blood was optimized for centrifugation conditions with respect to microparticle counts.

Automatic buffy coat extraction: methodology, feasibility, optimization and validation study.

Buffy coat isolation from whole blood has typically been a long, manual process. We tested and evaluated the feasibility, efficiency, and reproducibility of extracting buffy coat by an automated process with the Tecan pipetting robot Freedom Evo200.

Increased Th2 cytokine secretion, eosinophilic airway inflammation, and airway hyperresponsiveness in neurturin-deficient mice.

Neurotrophins such as nerve growth factor and brain-derived neurotrophic factor have been described to be involved in the pathogenesis of asthma. Neurturin (NTN), another neurotrophin from the glial cell line-derived neurotrophic factor family, was shown to be produced by human immune cells: monocytes, B cells, and T cells. Furthermore, it was previously described that the secretion of inflammatory cytokines was dramatically stimulated in NTN knockout (NTN(-/-)) mice.

Mouse natural killer (NK) cells express the nerve growth factor receptor TrkA, which is dynamically regulated.

Background: Nerve growth factor (NGF) is a neurotrophin crucial for the development and survival of neurons. It also acts on cells of the immune system which express the NGF receptors TrkA and p75(NTR) and can be produced by them. However, mouse NK cells have not yet been studied in this context.

Novel method for isolating untouched rat natural killer cells with higher purity compared with positive selection and fluorescence-activated cell sorting.

Natural killer (NK) cells are important effectors of both innate and adaptive immune responses. Although human and mouse NK cells are extensively characterized, much less is known about the rat cells, partly because of the current lack of reliable isolation techniques. We aimed to develop a method for isolating highly pure 'untouched' rat NK cells by negative selection from splenocytes.