Journal of extracellular biology最新文献

{"title":"Growth phase matters: Boosting immunity via Lacticasebacillus-derived membrane vesicles and their interactions with TLR2 pathways","authors":"Miriam Sandanusova,&nbsp;Kristyna Turkova,&nbsp;Eva Pechackova,&nbsp;Jan Kotoucek,&nbsp;Pavel Roudnicky,&nbsp;Martin Sindelar,&nbsp;Lukas Kubala,&nbsp;Gabriela Ambrozova","doi":"10.1002/jex2.169","DOIUrl":"https://doi.org/10.1002/jex2.169","url":null,"abstract":"<p>Lipid bi-layered particles known as membrane vesicles (MVs), produced by Gram-positive bacteria are a communication tool throughout the entire bacterial growth. However, the MVs characteristics may vary across all stages of maternal culture growth, leading to inconsistencies in MVs research. This, in turn, hinders their employment as nanocarriers, vaccines and other medical applications. In this study, we aimed to comprehensively characterize MVs derived from <i>Lacticaseibacillus rhamnosus</i> CCM7091 isolated at different growth stages: early exponential (6 h, MV6), late exponential (12 h, MV12) and late stationary phase (48 h, MV48). We observed significant differences in protein content between MV6 and MV48 (data are available via ProteomeXchange with identifier PXD041580), likely contributing to their different immunomodulatory capacities. In vitro analysis demonstrated that MV48 uptake rate by epithelial Caco-2 cells is significantly higher and they stimulate an immune response in murine macrophages RAW 264.7 (elevated production of TNFα, IL-6, IL-10, NO). This correlated with increased expression of lipoteichoic acid (LTA) and enhanced TLR2 signalling in MV48, suggesting that LTA contributes to the immunomodulation. In conclusion, we showed that <i>Lacticaseibacillus rhamnosus</i> CCM7091-derived MVs from the late stationary phase boost the immune response the most effectively, which pre-destines them for therapeutical application as nanocarriers.</p>","PeriodicalId":73747,"journal":{"name":"Journal of extracellular biology","volume":"3 8","pages":""},"PeriodicalIF":0.0,"publicationDate":"2024-08-22","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://onlinelibrary.wiley.com/doi/epdf/10.1002/jex2.169","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142041568","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Extracellular vesicles may provide an alternative detoxification pathway during skeletal muscle myoblast ageing","authors":"María Fernández-Rhodes,&nbsp;Emma Buchan,&nbsp;Stephanie D. Gagnon,&nbsp;Jiani Qian,&nbsp;Lee Gethings,&nbsp;Rebecca Lees,&nbsp;Ben Peacock,&nbsp;Andrew J. Capel,&nbsp;Neil R. W. Martin,&nbsp;Pola Goldberg Oppenheimer,&nbsp;Mark P. Lewis,&nbsp;Owen G. Davies","doi":"10.1002/jex2.171","DOIUrl":"10.1002/jex2.171","url":null,"abstract":"<p>Skeletal muscle (SM) acts as a secretory organ, capable of releasing myokines and extracellular vesicles (SM-EVs) that impact myogenesis and homeostasis. While age-related changes have been previously reported in murine SM-EVs, no study has comprehensively profiled SM-EV in human models. To this end, we provide the first comprehensive comparison of SM-EVs from young and old human primary skeletal muscle cells (HPMCs) to map changes associated with SM ageing. HPMCs, isolated from young (24 ± 1.7 years old) and older (69 ± 2.6 years old) participants, were immunomagnetically sorted based on the presence of the myogenic marker CD56 (N-CAM) and cultured as pure (100% CD56⁺) or mixed populations (MP: 90% CD56⁺). SM-EVs were isolated using an optimised protocol combining ultrafiltration and size exclusion chromatography (UF + SEC) and their biological content was extensively characterised using Raman spectroscopy (RS) and liquid chromatography mass spectrometry (LC-MS). Minimal variations in basic EV parameters (particle number, size, protein markers) were observed between young and old populations. However, biochemical fingerprinting by RS highlighted increased protein (amide I), lipid (phospholipids and phosphatidylcholine) and hypoxanthine signatures for older SM-EVs. Through LC-MS, we identified 84 shared proteins with functions principally related to cell homeostasis, muscle maintenance and transcriptional regulation. Significantly, SM-EVs from older participants were comparatively enriched in proteins involved in oxidative stress and DNA/RNA mutagenesis, such as E3 ubiquitin-protein ligase TTC3 (TTC3), little elongation complex subunit 1 (ICE1) and Acetyl-CoA carboxylase 1 (ACACA). These data suggest SM-EVs could provide an alternative pathway for homeostasis and detoxification during SM ageing.</p>","PeriodicalId":73747,"journal":{"name":"Journal of extracellular biology","volume":"3 8","pages":""},"PeriodicalIF":0.0,"publicationDate":"2024-08-21","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11336379/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142019740","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Increased levels of circulating cell-free double-stranded nucleic acids in the plasma of glioblastoma patients","authors":"Elisabeth Rackles,&nbsp;Elena Zaccheroni,&nbsp;Patricia Hernandez Lopez,&nbsp;Stefania Faletti,&nbsp;Massimiliano Del Bene,&nbsp;Francesco DiMeco,&nbsp;Giuliana Pelicci,&nbsp;Juan M Falcon-Perez","doi":"10.1002/jex2.168","DOIUrl":"10.1002/jex2.168","url":null,"abstract":"<p>Circulating cell-free nucleic acids are considered a promising source of biomarkers for diseases and cancer. Liquid biopsy biomarkers for brain tumours represent a major, still unmet, clinical need. In plasma, nucleic acids can be free or be associated with extracellular vesicles (EVs). Here we report an easy and reproducible method to analyse cell-free nucleic acids in plasma and EVs by conventional flow cytometry easy to translate into the clinics. Nucleic acids associated with the EVs or present in plasma samples are stained by Pyronin Y, which is a fluorescent dye that is preferably binding double-stranded nucleic acids. Fluorescent staining of EVs isolated from cell-conditioned media is suitable for DNA and RNA detection by flow cytometry. The nucleic acids are partially protected from degradation by the EVs’ membrane. Additionally, DNA and RNA can be stained in plasma samples and plasma-derived EVs. Remarkably, analysis of plasma from patients and healthy individuals reveals a difference in their nucleic acid profiles. Taken together, our results indicate that the proposed methodology, which is based on conventional direct flow cytometry, is a promising easy tool for plasma nucleic acid analysis.</p>","PeriodicalId":73747,"journal":{"name":"Journal of extracellular biology","volume":"3 8","pages":""},"PeriodicalIF":0.0,"publicationDate":"2024-08-02","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11294885/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141891165","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Optimization of ultracentrifugation-based method to enhance the purity and proteomic profiling depth of plasma-derived extracellular vesicles and particles","authors":"Zurong Wan,&nbsp;Jinghua Gu,&nbsp;Uthra Balaji,&nbsp;Linda Bojmar,&nbsp;Henrik Molina,&nbsp;Søren Heissel,&nbsp;Alexandra E. Pagano,&nbsp;Christopher Peralta,&nbsp;Lee Shaashua,&nbsp;Dorina Ismailgeci,&nbsp;Hope K. Narozniak,&nbsp;Yi Song,&nbsp;William R. Jarnagin,&nbsp;David P. Kelsen,&nbsp;Jaqueline Bromberg,&nbsp;Virginia Pascual,&nbsp;Haiying Zhang","doi":"10.1002/jex2.167","DOIUrl":"10.1002/jex2.167","url":null,"abstract":"<p>Circulating extracellular vesicles and particles (EVPs) are being investigated as potential biomarkers for early cancer detection, prognosis, and disease monitoring. However, the suboptimal purity of EVPs isolated from peripheral blood plasma has posed a challenge of in-depth analysis of the EVP proteome. Here, we compared the effectiveness of different methods for isolating EVPs from healthy donor plasma, including ultracentrifugation (UC)-based protocols, phosphatidylserine-Tim4 interaction-based affinity capture (referred to as “PS”), and several commercial kits. Modified UC methods with an additional UC washing or size exclusion chromatography step substantially improved EVP purity and enabled the detection of additional proteins via proteomic mass spectrometry, including many plasma membrane and cytoplasmic proteins involved in vesicular regulation pathways. This improved performance was reproduced in cancer patient plasma specimens, resulting in the identification of a greater number of differentially expressed EVP proteins, thus expanding the range of potential biomarker candidates. However, PS and other commercial kits did not outperform UC-based methods in improving plasma EVP purity. PS yielded abundant contaminating proteins and a biased enrichment for specific EVP subsets, thus unsuitable for proteomic profiling of plasma EVPs. Therefore, we have optimized UC-based protocols for circulating EVP isolation, which enable further in-depth proteomic analysis for biomarker discovery.</p>","PeriodicalId":73747,"journal":{"name":"Journal of extracellular biology","volume":"3 7","pages":""},"PeriodicalIF":0.0,"publicationDate":"2024-07-23","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11263976/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141753473","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Evaluation of resazurin phenoxazine dye as a highly sensitive cell viability potency assay for natural killer cell-derived extracellular vesicle-based cancer biotherapeutics","authors":"Frederic St-Denis-Bissonnette,&nbsp;Shirley Qiu,&nbsp;Sarah E. Cummings,&nbsp;Melanie Kirkby,&nbsp;Yohannes Haile,&nbsp;Sarah Wassmer,&nbsp;Gauri Muradia,&nbsp;Jelica Mehic,&nbsp;Andrew Stalker,&nbsp;Amit Shrestha,&nbsp;Michele Ardolino,&nbsp;Seung-Hwan Lee,&nbsp;Dylan Burger,&nbsp;Lisheng Wang,&nbsp;Jessie R. Lavoie","doi":"10.1002/jex2.166","DOIUrl":"10.1002/jex2.166","url":null,"abstract":"<p>Natural killer cell-derived extracellular vesicles (NK-EVs) are candidate biotherapeutics against various cancers. However, standardised potency assays are necessary for a reliable assessment of NK-EVs' cytotoxicity. This study aims to thoroughly evaluate a highly sensitive resazurin phenoxazine-based cell viability potency assay (measurement of the cellular redox metabolism) for quantifying the cytotoxicity of NK-EVs against leukaemia K562 cells (suspension model) and breast cancer MDA-MB-231 cells (adherent model) in vitro. The assay was evaluated based on common analytical parameters setforth by regulatory guidelines, including specificity, selectivity,accuracy, precision, linearity, range and stability. Our results revealed that this resazurin-based cell viability potency assay reliably and reproducibly measured a dose-response of NK-EVs’ cytotoxic activity against both cancer models. The assay showed precision with 5% and 20% variation for intra-run and inter-run variability. The assay signal showed specificity and selectivity of NK-EVs against cancer target cells, as evidenced by the diminished viability of cancer cells following a 5-hour treatment with NK-EVs, without any detectable interference or background. The linearity analysis of target cancer cells revealed strong linearity for densities of 5000 K562 and 1000 MDA-MB-231 cells per test with a consistent range. Importantly, NK-EVs’ dose-response for cytotoxicity showed a strong correlation (|<i>ρ</i>| ∼ 0.8) with the levels of known cytotoxic factors associated with the NK-EVs’ corona (FasL, GNLY, GzmB, PFN and IFN-γ), thereby validating the accuracy of the assay. The assay also distinguished cytotoxicity changes in degraded NK-EVs, indicating the ability of the assay to detect the potential loss of sample integrity. Compared to other commonly reported bioassays (i.e., flow cytometry, cell counting, lactate dehydrogenase release assay, DNA-binding reporter assay and confluence assay), our results support this highly sensitive resazurin-based viability potency assay as a high-throughput and quantitative method for assessing NK-EVs’ cytotoxicity against both suspension and adherent cancer models for evaluating NK-EVs’ biotherapeutics.</p>","PeriodicalId":73747,"journal":{"name":"Journal of extracellular biology","volume":"3 7","pages":""},"PeriodicalIF":0.0,"publicationDate":"2024-07-17","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11253028/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141636023","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Mechanical property estimation of sarcoma-relevant extracellular vesicles using transmission electron microscopy","authors":"Premanshu Kumar Singh,&nbsp;Patricia Sarchet,&nbsp;Catherine Hord,&nbsp;Lucia Casadei,&nbsp;Raphael Pollock,&nbsp;Shaurya Prakash","doi":"10.1002/jex2.158","DOIUrl":"10.1002/jex2.158","url":null,"abstract":"<p>Analysis of single extracellular vesicles (EVs) has the potential to yield valuable label-free information on their morphological structure, biomarkers and therapeutic targets, though such analysis is hindered by the lack of reliable and quantitative measurements of the mechanical properties of these compliant nanoscale particles. The technical challenge in mechanical property measurements arises from the existing tools and methods that offer limited throughput, and the reported elastic moduli range over several orders of magnitude. Here, we report on a flow-based method complemented by transmission electron microscopy (TEM) imaging to provide a high throughput, whole EV deformation analysis for estimating the mechanical properties of liposarcoma-derived EVs as a function of their size. Our study includes extracting morphological data of EVs from a large dataset of 432 TEM images, with images containing single to multiple EVs, and implementing the thin-shell deformation theory. We estimated the elastic modulus, <i>E</i> = 0.16 ± 0.02 MPa (mean±SE) for small EVs (sEVs; 30–150 nm) and <i>E</i> = 0.17 ± 0.03 MPa (mean±SE) for large EVs (<i>l</i>EVs; &gt;150 nm). To our knowledge, this is the first report on the mechanical property estimation of LPS-derived EVs and has the potential to establish a relationship between EV size and EV mechanical properties.</p>","PeriodicalId":73747,"journal":{"name":"Journal of extracellular biology","volume":"3 7","pages":""},"PeriodicalIF":0.0,"publicationDate":"2024-07-04","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11222873/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141536086","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Characterisation of extracellular vesicles in baculovirus infection of Spodoptera frugiperda cells","authors":"Lex J. C. Van Es,&nbsp;Robert D. Possee,&nbsp;Linda A. King","doi":"10.1002/jex2.163","DOIUrl":"10.1002/jex2.163","url":null,"abstract":"<p><i>Autographa californica</i> multiple nucleopolyhedrovirus (AcMNPV) is an enveloped DNA virus of the <i>Baculoviridae</i> family. This baculovirus is widely exploited for the biological control of insect pest species and as an expression platform to produce recombinant proteins in insect cells. Extracellular vesicles (EVs) are secreted by all cells and are involved in key roles in many biological processes through their cargo consisting of proteins, RNA or DNA. In viral infections, EVs have been found to transfer both viral and cellular cargo that can elicit either a pro- or antiviral response in recipient cells. Here, small EVs (sEVs) released by <i>Spodoptera frugiperda</i> (Sf) insect cells were characterised for the first time. Using <i>S. frugiperda</i> (SfC1B5) cells stably expressing the baculovirus <i>gp64</i>, the viral envelope protein GP64 was shown to be incorporated into sEVs. Sf9 cells were also transfected with a bacmid AcMNPV genome lacking <i>p6.9</i> (AcΔP6.9) to prevent budded virus production. The protein content of sEVs from both mock- and AcΔP6.9-transfected cells were analysed by mass spectrometry. In addition to GP64, viral proteins Ac-F, ME-53 and viral ubiquitin were identified, as well as many host proteins including TSG101—which may be useful as a protein marker for sEVs.</p>","PeriodicalId":73747,"journal":{"name":"Journal of extracellular biology","volume":"3 7","pages":""},"PeriodicalIF":0.0,"publicationDate":"2024-06-28","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11212295/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141473245","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Towards IVDR-compliance by implementing quality control steps in a quantitative extracellular vesicle-miRNA liquid biopsy assay for response monitoring in patients with classic Hodgkin lymphoma","authors":"Esther E. E. Drees,&nbsp;Nils J. Groenewegen,&nbsp;Sandra A. W. M. Verkuijlen,&nbsp;Monique A. J. van Eijndhoven,&nbsp;Jip Ramaker,&nbsp;Pepijn Veenstra,&nbsp;Mirjam Hussain,&nbsp;Catharina G. M. Groothuis-Oudshoorn,&nbsp;Daphne de Jong,&nbsp;Josée M. Zijlstra,&nbsp;Johan de Rooij,&nbsp;D. Michiel Pegtel","doi":"10.1002/jex2.164","DOIUrl":"10.1002/jex2.164","url":null,"abstract":"<p>Previously, we showed that quantification of lymphoma-associated miRNAs miR-155-5p, -127-3p and let-7a-5p levels in plasma extracellular vesicles (EVs) report treatment response in patients with classic Hodgkin lymphoma (cHL). Prior to clinical implementation, quality control (QC) steps and validation are required to meet international regulatory standards. Most published EV-based diagnostic assays have yet to meet these requirements. In order to advance the assay towards regulatory compliance (e.g., IVDR 2017/746), we incorporated three QC steps in our experimental EV-miRNA quantitative real-time reverse-transcription PCR (q-RT-PCR) assay in an ISO-13485 certified quality-management system (QMS). Liposomes encapsulated with a synthetic (nematode-derived) miRNA spike-in controlled for EV isolation by automated size-exclusion chromatography (SEC). Additional miRNA spike-ins controlled for RNA isolation and cDNA conversion efficiency. After deciding on quality criteria, in total 107 out of 120 samples from 46 patients passed QC. Generalized linear mixed-effect modelling with bootstrapping determined the diagnostic performance of the quality-controlled data at an area under the curve (AUC) of 0.84 (confidence interval [CI]: 0.76–0.92) compared to an AUC of 0.87 (CI: 0.80–0.94) of the experimental assay. After the inclusion of QC steps, the accuracy of the assay was determined to be 78.5% in predicting active disease status in cHL patients during treatment. We demonstrate that a quality-controlled plasma EV-miRNA assay is technically robust, taking EV-miRNA as liquid biopsy assay an important step closer to clinical evaluation.</p>","PeriodicalId":73747,"journal":{"name":"Journal of extracellular biology","volume":"3 7","pages":""},"PeriodicalIF":0.0,"publicationDate":"2024-06-28","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11213689/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141473247","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Acute neuroinflammation promotes a metabolic shift that alters extracellular vesicle cargo in the mouse brain cortex","authors":"Natasha Vassileff,&nbsp;Jereme G. Spiers,&nbsp;Juliani Juliani,&nbsp;Rohan G. T. Lowe,&nbsp;Keshava K. Datta,&nbsp;Andrew F. Hill","doi":"10.1002/jex2.165","DOIUrl":"10.1002/jex2.165","url":null,"abstract":"<p>Neuroinflammation is initiated through microglial activation and cytokine release which can be induced through lipopolysaccharide treatment (LPS) leading to a transcriptional cascade culminating in the differential expression of target proteins. These differentially expressed proteins can then be packaged into extracellular vesicles (EVs), a form of cellular communication, further propagating the neuroinflammatory response over long distances. Despite this, the EV proteome in the brain, following LPS treatment, has not been investigated. Brain tissue and brain derived EVs (BDEVs) isolated from the cortex of LPS-treated mice underwent thorough characterisation to meet the minimal information for studies of extracellular vesicles guidelines before undergoing mass spectrometry analysis to identify the differentially expressed proteins. Fourteen differentially expressed proteins were identified in the LPS brain tissue samples compared to the controls and 57 were identified in the BDEVs isolated from the LPS treated mice compared to the controls. This included proteins associated with the initiation of the inflammatory response, epigenetic regulation, and metabolism. These results allude to a potential link between small EV cargo and early inflammatory signalling.</p>","PeriodicalId":73747,"journal":{"name":"Journal of extracellular biology","volume":"3 7","pages":""},"PeriodicalIF":0.0,"publicationDate":"2024-06-27","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11212288/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141473244","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"P2RX7 plays a critical role in extracellular vesicle-mediated secretion of pathogenic molecules from microglia and astrocytes","authors":"Mohammad Abdullah,&nbsp;Zhi Ruan,&nbsp;Seiko Ikezu,&nbsp;Tsuneya Ikezu","doi":"10.1002/jex2.155","DOIUrl":"10.1002/jex2.155","url":null,"abstract":"<p>Extracellular vesicle (EV) secretion is mediated by purinergic receptor P2X₇ (P2RX7), an ATP-gated cation channel highly expressed in microglia. We have previously shown that administration of GSK1482160, a P2RX7 selective inhibitor, suppresses EV secretion from murine microglia and prevents tauopathy development, leading to the recovery of the hippocampal function in PS19 mice, expressing P301S tau mutant. It is yet unknown, however, whether the effect of GSK1482160 on EV secretion from glial cells is specifically regulated through P2RX7. Here we tested GSK1482160 on primary microglia and astrocytes isolated from C57BL/6 (WT) and <i>P2rx7^–/–</i> mice and evaluated their EV secretion and phagocytotic activity of aggregated human tau (hTau) under ATP stimulation. GSK1482160 treatment and deletion of <i>P2rx7</i> significantly reduced secretion of small and large EVs in microglia and astrocytes in both ATP stimulated or unstimulated condition as determined by nanoparticle tracking analysis, CD9 ELISA and immunoblotting of Tsg101 and Flotilin 1 using isolated EVs. GSK1482160 treatment had no effect on EV secretion from <i>P2rx7</i>^–/– microglia while we observed significant reduction in the secretion of small EVs from <i>P2rx7</i>^–/– astrocytes, suggesting its specific targeting of P2RX7 in EV secretion except small EV secretion from astrocytes. Finally, deletion of <i>P2rx7</i> suppressed IL-1β secretion and phagocytosed misfolded tau from both microglia and astrocytes. Together, these findings show that GSK1482160 suppresses EV secretion from microglia and astrocytes in P2RX7-dependment manner, and P2RX7 critically regulates secretion of IL-1β and misfolded hTau, demonstrating as the viable target of suppressing EV-mediated neuroinflammation and tau propagation.</p>","PeriodicalId":73747,"journal":{"name":"Journal of extracellular biology","volume":"3 7","pages":""},"PeriodicalIF":0.0,"publicationDate":"2024-06-27","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11212328/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141473246","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}