Journal of Extracellular Vesicles最新文献

{"title":"Stoichiometric constraints for detection of EV-borne biomarkers in blood","authors":"Nataša Zarovni,&nbsp;Danilo Mladenović,&nbsp;Dario Brambilla,&nbsp;Federica Panico,&nbsp;Marcella Chiari","doi":"10.1002/jev2.70034","DOIUrl":"https://doi.org/10.1002/jev2.70034","url":null,"abstract":"<p>Stochiometric issues, encompassing both the quantity and heterogeneity of extracellular vesicles (EVs) derived from tumour or other tissues in blood, pose important challenges across various stages of biomarker discovery and detection, affecting the integrity of data, introducing losses and artifacts during blood processing, EV purification and analysis. These challenges shape the diagnostic utility of EVs especially within the framework of established and emerging methodologies. By addressing these challenges, we aim to delineate crucial parameters and requirements for tumour-specific EV detection, or more precisely, for tumour identification via EV based assays. Our endeavour involves a comprehensive examination of the layers that mask or confound the traceability of EV markers such as nucleic acids and proteins, and focus on ‘low prevalence—low concentration’ scenario. Finally, we evaluate the advantages versus limitations of single-particle analysers over more conventional bulk assays, suggesting that the combined use of both to capture and interpret the EV signals, in particular the EV surface displayed proteins, may ultimately provide quantitative information on their absolute abundance and distribution.</p>","PeriodicalId":15811,"journal":{"name":"Journal of Extracellular Vesicles","volume":"14 2","pages":""},"PeriodicalIF":15.5,"publicationDate":"2025-02-03","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://onlinelibrary.wiley.com/doi/epdf/10.1002/jev2.70034","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143111313","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Small Extracellular Vesicles Engineered Using Click Chemistry to Express Chimeric Antigen Receptors Show Enhanced Efficacy in Acute Liver Failure","authors":"Yen-Ting Lu,&nbsp;Tzu-Yu Chen,&nbsp;Hsin-Hung Lin,&nbsp;Ya-Wen Chen,&nbsp;Yu-Xiu Lin,&nbsp;Duy‑Cuong Le,&nbsp;Yen-Hua Huang,&nbsp;Andrew H.-J. Wang,&nbsp;Cheng-Chung Lee,&nbsp;Thai-Yen Ling","doi":"10.1002/jev2.70044","DOIUrl":"https://doi.org/10.1002/jev2.70044","url":null,"abstract":"<p>Acetaminophen (APAP) overdose can cause severe liver injury and life-threatening conditions that may lead to multiple organ failure without proper treatment. N-acetylcysteine (NAC) is the accepted and prescribed treatment for detoxification in cases of APAP overdose. Nonetheless, in acute liver failure (ALF), particularly when the ingestion is substantial, NAC may not fully restore liver function. NAC administration in ALF has limitations and potential adverse effects, including nausea, vomiting, diarrhoea, flatus, gastroesophageal reflux, and anaphylactoid reactions. Mesenchymal stromal cell (MSC)-based therapies using paracrine activity show promise for treating ALF, with preclinical studies demonstrating improvement. Recently, MSC-derived extracellular vesicles (EVs) have emerged as a new therapeutic option for liver injury. MSC-derived EVs can contain various therapeutic cargos depending on the cell of origin, participate in physiological processes, and respond to abnormalities. However, most therapeutic EVs lack a distinct orientation upon entering the body, resulting in a lack of targeting specificity. Therefore, enhancing the precision of natural EV delivery systems is urgently needed. Thus, we developed an advanced targeting technique to deliver modified EVs within the body. Our strategy aims to employ bioorthogonal click chemistry to attach a targeting molecule to the surface of small extracellular vesicles (sEVs), creating exogenous chimeric antigen receptor-modified sEVs (CAR-sEVs) for the treatment. First, we engineered azido-modified sEVs (N₃-sEVs) through metabolic glycoengineering by treating MSCs with the azide-containing monosaccharide N-azidoacetyl-mannosamine (Ac4ManNAz). Next, we conjugated N₃-sEVs with a dibenzocyclooctyne (DBCO)-tagged single-chain variable fragment (DBCO-scFv) that targets the asialoglycoprotein receptor (ASGR1), thus producing CAR-sEVs for precise liver targeting. The efficacy of CAR-sEV therapy in ALF models by targeting ASGR1 was validated. MSC-derived CAR-sEVs reduced serum liver enzymes, mitigated liver damage, and promoted hepatocyte proliferation in APAP-induced injury. Overall, CAR-sEVs exhibited enhanced hepatocyte specificity and efficacy in ameliorating liver injury, highlighting the significant advancements achievable with cell-free targeted therapy.</p>","PeriodicalId":15811,"journal":{"name":"Journal of Extracellular Vesicles","volume":"14 2","pages":""},"PeriodicalIF":15.5,"publicationDate":"2025-02-03","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://onlinelibrary.wiley.com/doi/epdf/10.1002/jev2.70044","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143111315","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Metabolic Profiling of Brain Tissue and Brain-Derived Extracellular Vesicles in Alzheimer's Disease","authors":"Patricia Hernandez,&nbsp;Elisabeth Rackles,&nbsp;Oihane E. Alboniga,&nbsp;Pablo Martínez-Lage,&nbsp;Emma N. Camacho,&nbsp;Arantza Onaindia,&nbsp;Manuel Fernandez,&nbsp;Ana Talamillo,&nbsp;Juan M. Falcon-Perez","doi":"10.1002/jev2.70043","DOIUrl":"https://doi.org/10.1002/jev2.70043","url":null,"abstract":"<p>Alzheimer´s disease (AD) is the most frequent neurodegenerative disorder in the world and is characterised by the loss of memory and other cognitive functions. Metabolic changes associated with AD are important players in the development of the disease. However, the mechanism underlying these changes is still unknown. Extracellular vesicles (EVs) are nano-sized particles that play an important role in regulating pathophysiological processes and are a non-invasive manner to obtain information of the cell that is secreting them. The analysis of brain-derived EVs (bdEVs) will provide new insights in the metabolic processes associated with AD. To characterize bdEVs in AD, we optimised a method to isolate them from tissue of different brain regions, obtaining the highest enrichment in isolations from the temporal cortex. We performed unbiased untargeted metabolomics analysis on post-mortem human temporal cortex tissue and bdEVs from the same region of AD patients and healthy controls. Both, univariate and multivariate statistical analysis were used to determine the metabolites that influence the separation between AD patients and controls. Interestingly, a clear separation between control and AD groups was obtained with bdEVs, which allowed to select 12 relevant features by a validated PLS-DA model. Furthermore, comparison of tissue and bdEVs identified 68 common features. The pathway enrichment analysis of the common metabolites showed that the alanine, aspartate and glutamate pathway and the arginine, phenylalanine, tyrosine pathway were the most significant ones in the separation between the AD patients and controls. The phenylalanine, tyrosine and tryptophan pathway, still had a very high influence in the separation between groups, albeit not significant. Notably, some metabolites were identified for the first time in bdEVs. For example, the N-acetyl aspartic acid (NAA) metabolite present in bdEVs was suitable to differentiate AD patients from healthy controls. Furthermore, the analysis of the hippocampus, midbrain, temporal and entorhinal cortex and their respective bdEVs indicated that the metabolic profiles of different brain areas were distinct and showed some correlation between the metabolome of the tissue and its respective bdEVs. Thus, our study highlights the potential of bdEVs to understand the metabolic fingerprint associated with AD and their potential use as diagnostic and therapeutic targets.</p>","PeriodicalId":15811,"journal":{"name":"Journal of Extracellular Vesicles","volume":"14 2","pages":""},"PeriodicalIF":15.5,"publicationDate":"2025-02-03","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://onlinelibrary.wiley.com/doi/epdf/10.1002/jev2.70043","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143111312","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Breast Cancer-Derived Extracellular Vesicles Modulate the Cytoplasmic and Cytoskeletal Dynamics of Blood-Brain Barrier Endothelial Cells","authors":"Sara Busatto,&nbsp;Tzu-Hsi Song,&nbsp;Hyung Joon Kim,&nbsp;Caleb Hallinan,&nbsp;Michael N. Lombardo,&nbsp;Anat O. Stemmer-Rachamimov,&nbsp;Kwonmoo Lee,&nbsp;Marsha A. Moses","doi":"10.1002/jev2.70038","DOIUrl":"10.1002/jev2.70038","url":null,"abstract":"<p>Extracellular vesicles (EVs) from brain-seeking breast cancer cells (Br-EVs) breach the blood-brain barrier (BBB) via transcytosis and promote brain metastasis. Here, we defined the mechanisms by which Br-EVs modulate brain endothelial cell (BEC) dynamics to facilitate their BBB transcytosis. BEC treated with Br-EVs show significant downregulation of Rab11fip2, known to promote vesicle recycling to the plasma membrane and significant upregulation of Rab11fip3 and Rab11fip5, which support structural stability of the endosomal compartment and facilitate vesicle recycling and transcytosis, respectively. Using machine learning and quantitative global proteomic, we identified novel Br-EV-induced changes in BECs morphology, motility, and proteome that correlate with decreased BEC cytoplasm and cytoskeletal organization and dynamics. These results define early steps leading to breast-to-brain metastasis and identify molecules that could serve as targets for therapeutic strategies for brain metastasis.</p>","PeriodicalId":15811,"journal":{"name":"Journal of Extracellular Vesicles","volume":"14 1","pages":""},"PeriodicalIF":15.5,"publicationDate":"2025-01-27","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11770372/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143047096","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Intercellular Epigenomic Signalling via Extracellular Vesicles During B Cell Maturation","authors":"Kevin Ho Wai Yim,&nbsp;Ala'a Al Hrout,&nbsp;Richard Chahwan","doi":"10.1002/jev2.70040","DOIUrl":"10.1002/jev2.70040","url":null,"abstract":"<p>B cell maturation is crucial for effective adaptive immunity. It requires a complex signalling network to mediate antibody diversification through mutagenesis. B cells also rely on queues from other cells within the germinal centre. Recently, a novel class of intercellular signals mediated by extracellular vesicles (EVs) has emerged. Studies have shown that B cell EV-mediated signalling is involved in immune response regulation and tumorigenesis. However, the mechanistic role of B cell EVs is not yet established. We herein study the biological properties and physiological function of B cell EVs during B cell maturation. We use emerging technologies to profile B cell EV surface marker signatures at the single particle level, molecular cargo and physiological roles in B cell maturation. EV ncRNA cargo, characterised by RNA-seq, identified an EV-mediated novel non-coding RNA (ncRNA) regulatory network for B cell maturation. We show that a previously uncharacterised micro-RNA (miR-5099) in combination with a set of long ncRNA are carried within B cell EVs and could contribute to antibody diversification. The physiological role of EVs in B cell maturation is investigated using EV blockade assays and complementation studies using diverse EV sources further confirmed the physiological role and mode of action of EVs in B cell maturation.</p>","PeriodicalId":15811,"journal":{"name":"Journal of Extracellular Vesicles","volume":"14 1","pages":""},"PeriodicalIF":15.5,"publicationDate":"2025-01-27","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11770373/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143047098","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Extracellular Vesicles From Bone Marrow-Derived Macrophages Enriched in ARG1 Enhance Microglial Phagocytosis and Haematoma Clearance Following Intracerebral Haemorrhage","authors":"Libin Hu,&nbsp;Zihang Chen,&nbsp;Jianglong Lu,&nbsp;Shandong Jiang,&nbsp;Haopu Lin,&nbsp;Jiayin Zhou,&nbsp;Ning Wang,&nbsp;Chao Ding,&nbsp;Weifang Ni,&nbsp;Haitao Peng,&nbsp;Yin Li,&nbsp;Xuchao He,&nbsp;Jianru Li,&nbsp;Chaohui Jing,&nbsp;Yang Cao,&nbsp;Hang Zhou,&nbsp;Feng Yan,&nbsp;Gao Chen","doi":"10.1002/jev2.70041","DOIUrl":"10.1002/jev2.70041","url":null,"abstract":"<p>Microglial phagocytosis of haematomas is crucial for neural functional recovery following intracerebral haemorrhage (ICH), a process regulated by various factors from within and outside the central nervous system (CNS). Extracellular vesicles (EVs), significant mediators of intercellular communication, have been demonstrated to play a pivotal role in the pathogenesis and progression of CNS diseases. However, the regulatory role of endogenous EVs on the phagocytic capacity of microglia post-ICH remains elusive. Utilising multi-omics analysis of brain tissue-derived EVs proteomics and single-cell RNA sequencing, this study identified that bone marrow-derived macrophages (BMDMs) potentially enhance microglial phagocytosis via EVs following ICH. By blocking BMDMs and reducing ARG1 in BMDM-derived EVs, we demonstrated that BMDMs facilitate erythrophagocytosis by delivering ARG1 to microglia via EVs post-ICH. EVs-carried ARG1 was found to augment phagocytosis by promoting RAC1-dependent cytoskeletal remodelling in microglia. Collectively, this research uncovers an intercellular communication pathway from BMDMs to microglia mediated by EVs post-ICH. This provides a novel paradigm for EV-mediated intercellular communication mechanisms and suggests a promising therapeutic potential for BMDM-derived EVs in the treatment of ICH.</p>","PeriodicalId":15811,"journal":{"name":"Journal of Extracellular Vesicles","volume":"14 1","pages":""},"PeriodicalIF":15.5,"publicationDate":"2025-01-27","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11770371/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143047097","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Uncovering a new family of conserved virulence factors that promote the production of host-damaging outer membrane vesicles in gram-negative bacteria","authors":"Audrey Goman,&nbsp;Bérengère Ize,&nbsp;Katy Jeannot,&nbsp;Camille Pin,&nbsp;Delphine Payros,&nbsp;Cécile Goursat,&nbsp;Léa Ravon-Katossky,&nbsp;Kazunori Murase,&nbsp;Camille V. Chagneau,&nbsp;Hélène Revillet,&nbsp;Frédéric Taieb,&nbsp;Sophie Bleves,&nbsp;Laure David,&nbsp;Etienne Meunier,&nbsp;Priscilla Branchu,&nbsp;Eric Oswald","doi":"10.1002/jev2.70032","DOIUrl":"10.1002/jev2.70032","url":null,"abstract":"<p>CprA is a short-chain dehydrogenase/reductase (SDR) that contributes to resistance against colistin and antimicrobial peptides. The <i>cprA</i> gene is conserved across <i>Pseudomonas aeruginosa</i> clades and its expression is directly regulated by the two-component system PmrAB. We have shown that <i>cprA</i> expression leads to the production of outer membrane vesicles (OMVs) that block autophagic flux and have a greater capacity to activate the non-canonical inflammasome pathway. In a murine model of sepsis, a <i>P. aeruginosa</i> strain deleted for <i>cprA</i> was less virulent than the wild-type (WT) strain. These results demonstrate the important role of CprA in the pathogenicity of <i>P. aeruginosa</i>. It is worth noting that CprA is also a functional ortholog of hemolysin F (HlyF), which is encoded by virulence plasmids of <i>Escherichia coli</i>. We have shown that other cryptic SDRs encoded by mammalian and plant pathogens, such as <i>Yersinia pestis</i> and <i>Ralstonia solanacearum</i> are functional orthologs of CprA and HlyF. These SDRs also induce the production of OMVs which block autophagic flux. This study uncovers a new family of virulence determinants in Gram-negative bacteria, offering potential for innovative therapeutic interventions and deeper insights into bacterial pathogenesis.</p>","PeriodicalId":15811,"journal":{"name":"Journal of Extracellular Vesicles","volume":"14 1","pages":""},"PeriodicalIF":15.5,"publicationDate":"2025-01-22","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11752146/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143006359","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"A Non-Centrifugation Method to Concentrate and Purify Extracellular Vesicles Using Superabsorbent Polymer Followed by Size Exclusion Chromatography","authors":"Markus Bergqvist,&nbsp;Cecilia Lässer,&nbsp;Rossella Crescitelli,&nbsp;Kyong-Su Park,&nbsp;Jan Lötvall","doi":"10.1002/jev2.70037","DOIUrl":"10.1002/jev2.70037","url":null,"abstract":"<p>Extracellular vesicles (EVs) can be isolated and purified from cell cultures and biofluids using different methodologies. Here, we explored a novel EV isolation approach by combining superabsorbent polymers (SAP) in a dialysis membrane with size exclusion chromatography (SEC) to achieve high concentration and purity of EVs without the use of ultracentrifugation (UC). Suspension HEK293 cells transfected with CD63 coupled with Thermo Luciferase were used to quantify the EV yield and purity. The 500 mL conditioned medium volume was initially reduced by pressure ultrafiltration, followed by UC, SAP or a centrifugal filter unit (CFU). Using either of these methods, the EVs were concentrated to a final volume of approximately 1 mL, with retained functionality. The yield, quantified by luciferase activity, was highest with UC (70%–80%), followed by SAP (60%–70%) and CFU (50%–60%). Further purification of the EVs was performed by iodixanol density cushion (IDC) or SEC (Sepharose CL-2B or 6B, in either 10 or 20 mL columns). Although the IDC and Sepharose CL-2B (10 mL) achieved the highest yields, the purity was slightly higher (30%) with IDC. In conclusion, combining SAP concentration with CL-2B SEC is an alternative and efficient way to isolate EVs without using UC.</p>","PeriodicalId":15811,"journal":{"name":"Journal of Extracellular Vesicles","volume":"14 1","pages":""},"PeriodicalIF":15.5,"publicationDate":"2025-01-22","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11752139/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143006655","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Comprehensive Phenotyping of Extracellular Vesicles in Plasma of Healthy Humans – Insights Into Cellular Origin and Biological Variation","authors":"Marija Holcar,&nbsp;Ivica Marić,&nbsp;Tobias Tertel,&nbsp;Katja Goričar,&nbsp;Urška Čegovnik Primožič,&nbsp;Darko Černe,&nbsp;Bernd Giebel,&nbsp;Metka Lenassi","doi":"10.1002/jev2.70039","DOIUrl":"10.1002/jev2.70039","url":null,"abstract":"<p>Despite immense interest in biomarker applications of extracellular vesicles (EVs) from blood, our understanding of circulating EVs under physiological conditions in healthy humans remains limited. Using imaging and multiplex bead-based flow cytometry, we comprehensively quantified circulating EVs with respect to their cellular origin in a large cohort of healthy blood donors. We assessed coefficients of variations to characterize their biological variation and explored demographic, clinical, and lifestyle factors contributing to observed variation. Cell-specific circulating EV subsets show a wide range of concentrations that do not correlate with cell-of-origin concentrations in blood, suggesting steady-state EV subset concentrations are regulated by complex mechanisms, which differ even for EV subsets from the same cell type. Interestingly, tetraspanin+ circulating EVs largely originate from platelets and to a lesser extent from lymphocytes. Principal component analysis (PCA) and association analyses demonstrate high biological inter-individual variation in circulating EVs across healthy humans, which are only partly explained by the influence of sex, menopausal status, age and smoking on specific circulating EV and/or tetraspanin+ circulating EV subsets. No global influence of the explored subject's factors on circulating EVs was detected. Our findings provide the first comprehensive, quantitative data towards the cell-origin atlas of plasma EVs, with important implications in the clinical use of EVs as biomarkers.</p>","PeriodicalId":15811,"journal":{"name":"Journal of Extracellular Vesicles","volume":"14 1","pages":""},"PeriodicalIF":15.5,"publicationDate":"2025-01-20","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11746918/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143006702","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Reproducibility of extracellular vesicle research","authors":"Rossella Crescitelli,&nbsp;Juan Falcon-Perez,&nbsp;An Hendrix,&nbsp;Metka Lenassi,&nbsp;Le Thi Nguyet Minh,&nbsp;Takahiro Ochiya,&nbsp;Nicole Noren Hooten,&nbsp;Ursula Sandau,&nbsp;Clotilde Théry,&nbsp;Rienk Nieuwland","doi":"10.1002/jev2.70036","DOIUrl":"10.1002/jev2.70036","url":null,"abstract":"&lt;p&gt;Ever since its launch in 2011, the International Society for Extracellular Vesicles (ISEV) has endorsed, initiated, and supported original ideas and solutions to promote reproducibility (Hill et al., &lt;span&gt;2013&lt;/span&gt;; Lötvall et al., &lt;span&gt;2014&lt;/span&gt;; Théry et al., &lt;span&gt;2018&lt;/span&gt;; Welsh et al., &lt;span&gt;2024&lt;/span&gt;) and these efforts have been appreciated by the general scientific community (Abbott, &lt;span&gt;2023&lt;/span&gt;). Improving reproducibility is complex and multifactorial, and involves development of protocols, reference materials and standards, interlaboratory comparison studies, instrument calibration, transparent reporting, and education.&lt;/p&gt;&lt;p&gt;To support reproducibility, ISEV founded the Rigor and Standardization (R&amp;S) Subcommittee in 2019, which now includes fifteen task forces and three inter-societal working groups (https://www.isev.org/rigor-standardization). Within this subcommittee, hundreds of ISEV members have become actively involved in R&amp;S, and together they are working on creative solutions to overcome the challenges of reproducibility in the EV field.&lt;/p&gt;&lt;p&gt;EV research invariably involves collection, handling, and storage of (EV-containing) fluids, such as body fluids and conditioned culture media, and tissues, which are the starting materials for EV research. Collection, handling and storage of (purified) EV-containing materials will affect their composition. For example, preparation of plasma and serum, which are amongst the most commonly used body fluids for EV research (Royo et al., &lt;span&gt;2020&lt;/span&gt;), involves about 40 variables which all may impact the sample composition and downstream analysis of EVs (Clayton et al., &lt;span&gt;2019&lt;/span&gt;). Laboratories and biobanks preparing and storing EV-containing tissues and fluids, commonly use their own ‘in-house’ protocols, which all may differ from each other and have unknown effects on the sample composition and downstream analysis of EVs (López-Guerrero et al., &lt;span&gt;2023&lt;/span&gt;). Moreover, even when the same protocol is used, the sample composition may still vary (Bettin et al., &lt;span&gt;2022&lt;/span&gt;) and these differences can be sufficient to affect the results of downstream EV characterization (Bracht et al., &lt;span&gt;2023&lt;/span&gt;). Taken together, sample preparation and storage invariably leads to variability in sample composition, thereby introducing an ‘uneven playing field’ and bias which can hamper the comparability, interpretation and reproducibility of results on EVs.&lt;/p&gt;&lt;p&gt;At present, the current approach to improve reproducibility is by reporting the protocol of sample preparation in the Materials and Methods section of scientific manuscripts. Unfortunately, this reporting is often incomplete or inconsistent between manuscripts, thereby hampering reproducibility. There can be multiple reasons for incomplete or inconsistent reporting, ranging from researchers not knowing these details themselves, or because the journal has a strict word count, or the details are con","PeriodicalId":15811,"journal":{"name":"Journal of Extracellular Vesicles","volume":"14 1","pages":""},"PeriodicalIF":15.5,"publicationDate":"2025-01-17","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11739893/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143006358","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}