Journal of extracellular biology最新文献

{"title":"Scalable purification of extracellular vesicles with high yield and purity using multimodal flowthrough chromatography","authors":"Scott E. Bonner,&nbsp;Simonides I. van de Wakker,&nbsp;William Phillips,&nbsp;Eduard Willms,&nbsp;Joost P. G. Sluijter,&nbsp;Andrew F. Hill,&nbsp;Matthew J. A. Wood,&nbsp;Pieter Vader","doi":"10.1002/jex2.138","DOIUrl":"https://doi.org/10.1002/jex2.138","url":null,"abstract":"<p>Extracellular vesicles (EVs) are cell derived membranous nanoparticles. EVs are important mediators of cell–cell communication via the transfer of bioactive content and as such they are being investigated for disease diagnostics as biomarkers and for potential therapeutic cargo delivery to recipient cells. However, existing methods for isolating EVs from biological samples suffer from challenges related to co-isolation of unwanted materials such as proteins, nucleic acids, and lipoproteins. In the pursuit of improved EV isolation techniques, we introduce multimodal flowthrough chromatography (MFC) as a scalable alternative to size exclusion chromatography (SEC). The use of MFC offers significant advantages for purifying EVs, resulting in enhanced yields and increased purity with respect to protein and nucleic acid co-isolates from conditioned 3D cell culture media. Compared to SEC, significantly higher EV yields with similar purity and preserved functionality were also obtained with MFC in 2D cell cultures. Additionally, MFC yielded EVs from serum with comparable purity to SEC and similar apolipoprotein B content. Overall, MFC presents an advancement in EV purification yielding EVs with high recovery, purity, and functionality, and offers an accessible improvement to researchers currently employing SEC.</p>","PeriodicalId":73747,"journal":{"name":"Journal of extracellular biology","volume":"3 2","pages":""},"PeriodicalIF":0.0,"publicationDate":"2024-01-31","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://onlinelibrary.wiley.com/doi/epdf/10.1002/jex2.138","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"139676751","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":"International Society for Extracellular Vesicles workshop. QuantitatEVs: Multiscale analyses, from bulk to single extracellular vesicle","authors":"Manuela Basso,&nbsp;Alessandro Gori,&nbsp;Caterina Nardella,&nbsp;Mari Palviainen,&nbsp;Marija Holcar,&nbsp;Ioannis Sotiropoulos,&nbsp;Sylwia Bobis-Wozowicz,&nbsp;Vito G. D'Agostino,&nbsp;Elena Casarotto,&nbsp;Yari Ciani,&nbsp;Shiro Suetsugu,&nbsp;Alice Gualerzi,&nbsp;Lorena Martin-Jaular,&nbsp;Daniela Boselli,&nbsp;Anna Kashkanova,&nbsp;Pietro Parisse,&nbsp;Lien Lippens,&nbsp;Martina Pagliuca,&nbsp;Martin Blessing,&nbsp;Roberto Frigerio,&nbsp;Thibaut Fourniols,&nbsp;Ana Meliciano,&nbsp;Anna Fietta,&nbsp;Paolo Vincenzo Fioretti,&nbsp;Karolina Soroczyńska,&nbsp;Silvia Picciolini,&nbsp;Amanda Salviano-Silva,&nbsp;Paolo Bergese,&nbsp;Davide Zocco,&nbsp;Marcella Chiari,&nbsp;Guido Jenster,&nbsp;Levi Waldron,&nbsp;Aleksandar Milosavljevic,&nbsp;John Nolan,&nbsp;Marco P. Monopoli,&nbsp;Kenneth W. Witwer,&nbsp;Benedetta Bussolati,&nbsp;Dolores Di Vizio,&nbsp;Juan Falcon Perez,&nbsp;Metka Lenassi,&nbsp;Marina Cretich,&nbsp;Francesca Demichelis","doi":"10.1002/jex2.137","DOIUrl":"10.1002/jex2.137","url":null,"abstract":"<p>The “QuantitatEVs: multiscale analyses, from bulk to single vesicle” workshop aimed to discuss quantitative strategies and harmonized wet and computational approaches toward the comprehensive analysis of extracellular vesicles (EVs) from bulk to single vesicle analyses with a special focus on emerging technologies. The workshop covered the key issues in the quantitative analysis of different EV-associated molecular components and EV biophysical features, which are considered the core of EV-associated biomarker discovery and validation for their clinical translation. The in-person-only workshop was held in Trento, Italy, from January 31^st to February 2^nd, 2023, and continued in Milan on February 3^rd with “Next Generation EVs,” a satellite event dedicated to early career researchers (ECR). This report summarizes the main topics and outcomes of the workshop.</p>","PeriodicalId":73747,"journal":{"name":"Journal of extracellular biology","volume":"3 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2024-01-24","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://onlinelibrary.wiley.com/doi/epdf/10.1002/jex2.137","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"139636084","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":"Proteome encoded determinants of protein sorting into extracellular vesicles","authors":"Katharina Waury,&nbsp;Dea Gogishvili,&nbsp;Rienk Nieuwland,&nbsp;Madhurima Chatterjee,&nbsp;Charlotte E. Teunissen,&nbsp;Sanne Abeln","doi":"10.1002/jex2.120","DOIUrl":"https://doi.org/10.1002/jex2.120","url":null,"abstract":"<p>Extracellular vesicles (EVs) are membranous structures released by cells into the extracellular space and are thought to be involved in cell-to-cell communication. While EVs and their cargo are promising biomarker candidates, sorting mechanisms of proteins to EVs remain unclear. In this study, we ask if it is possible to determine EV association based on the protein sequence. Additionally, we ask what the most important determinants are for EV association. We answer these questions with explainable AI models, using human proteome data from EV databases to train and validate the model. It is essential to correct the datasets for contaminants introduced by coarse EV isolation workflows and for experimental bias caused by mass spectrometry. In this study, we show that it is indeed possible to predict EV association from the protein sequence: a simple sequence-based model for predicting EV proteins achieved an area under the curve of 0.77 ± 0.01, which increased further to 0.84 ± 0.00 when incorporating curated post-translational modification (PTM) annotations. Feature analysis shows that EV-associated proteins are stable, polar, and structured with low isoelectric point compared to non-EV proteins. PTM annotations emerged as the most important features for correct classification; specifically, palmitoylation is one of the most prevalent EV sorting mechanisms for unique proteins. Palmitoylation and nitrosylation sites are especially prevalent in EV proteins that are determined by very strict isolation protocols, indicating they could potentially serve as quality control criteria for future studies. This computational study offers an effective sequence-based predictor of EV associated proteins with extensive characterisation of the human EV proteome that can explain for individual proteins which factors contribute to their EV association.</p>","PeriodicalId":73747,"journal":{"name":"Journal of extracellular biology","volume":"3 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2024-01-23","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://onlinelibrary.wiley.com/doi/epdf/10.1002/jex2.120","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"139550502","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":"Activation of the receptor KIT induces the secretion of exosome-like small extracellular vesicles","authors":"Annika Pfeiffer,&nbsp;Geethani Bandara,&nbsp;Jennifer D. Petersen,&nbsp;Guido H. Falduto,&nbsp;Joshua Zimmerberg,&nbsp;Dean D. Metcalfe,&nbsp;Ana Olivera","doi":"10.1002/jex2.139","DOIUrl":"https://doi.org/10.1002/jex2.139","url":null,"abstract":"<p>The receptor tyrosine kinase (RTK) KIT and its ligand stem cell factor (SCF) are essential for human mast cell (huMC) survival and proliferation. HuMCs expressing oncogenic KIT variants secrete large numbers of extracellular vesicles (EVs). The role KIT plays in regulating EV secretion has not been examined. Here, we investigated the effects of stimulation or inhibition of KIT activity on the secretion of small EVs (sEVs). In huMCs expressing constitutively active KIT, the quantity and quality of secreted sEVs positively correlated with the activity status of KIT. SCF-mediated stimulation of KIT in huMCs or murine MCs, or of transiently expressed KIT in HeLa cells, enhanced the release of sEVs expressing exosome markers. In contrast, ligand-mediated stimulation of the RTK EGFR in HeLa cells did not affect sEV secretion. The release of sEVs induced by either constitutively active or ligand-activated KIT was remarkably decreased when cells were treated with KIT inhibitors, concomitant with reduced exosome markers in sEVs. Similarly, inhibition of oncogenic KIT signalling kinases like PI3K, and MAPK significantly reduced the secretion of sEVs. Thus, activation of KIT and its early signalling cascades stimulate the secretion of exosome-like sEVs in a regulated fashion, which may have implications for KIT-driven functions.</p>","PeriodicalId":73747,"journal":{"name":"Journal of extracellular biology","volume":"3 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2024-01-23","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://onlinelibrary.wiley.com/doi/epdf/10.1002/jex2.139","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"139550198","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":"Identification of stable reference genes for relative quantification of long RNA expression in urinary extracellular vesicles","authors":"Xiao-Xiao Zhu,&nbsp;An-Ran Shen,&nbsp;Ning Li,&nbsp;Song-Tao Feng,&nbsp;Tao-Tao Tang,&nbsp;Yue Zhang,&nbsp;Jing Jing,&nbsp;Xin Zhong,&nbsp;Li-Jun Xie,&nbsp;Sheng-Lin Huang,&nbsp;Bi-Cheng Liu,&nbsp;Lin-Li Lv","doi":"10.1002/jex2.136","DOIUrl":"https://doi.org/10.1002/jex2.136","url":null,"abstract":"<p>Urinary extracellular vesicles (uEVs) are rich in valuable biomolecule information which are increasingly recognized as potential biomarkers for various diseases. uEV long RNAs are among the critical cargos capable of providing unique transcriptome information of the source cells. However, consensus regarding ideal reference genes for relative long RNAs quantification in uEVs is not available as of date. Here we explored stable reference genes through profiling the long RNA expression by RNA-seq following unsupervised analysis and validation studies. Candidate reference genes were identified using four algorithms: NormFinder, GeNorm, BestKeeper and the Delta Ct method, followed by validation. RNA profile showed uEVs contained abundant long RNAs information and the core transcriptome was related to cellular structures, especially ribosome which functions mainly as translation, protein and RNA binding molecules. Analysis of RNA-seq data identified RPL18A, RPL11, RPL27, RACK1, RPSA, RPL41, H1-2, RPL4, GAPDH, RPS27A as candidate reference genes. RT-qPCR validation revealed that RPL41, RPSA and RPL18A were reliable reference genes for long RNA quantification in uEVs from patients with diabetes mellitus (DM), diabetic nephropathy (DN), IgA nephropathy (IgAN) and prostate cancer (PCA). Interestingly, RPL41 also outperformed traditional reference genes in renal tissues of DN and IgAN, as well as in plasma EVs of several types of cancers. The stable reference genes identified in this study may facilitate development of uEVs as novel biomarkers and increase the accuracy and comparability of biomarker studies.</p>","PeriodicalId":73747,"journal":{"name":"Journal of extracellular biology","volume":"3 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2024-01-17","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://onlinelibrary.wiley.com/doi/epdf/10.1002/jex2.136","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"139488329","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":"Intracellular localisation and extracellular release of Y RNA and Y RNA binding proteins","authors":"Tom A. P. Driedonks,&nbsp;Sarah Ressel,&nbsp;Thi Tran Ngoc Minh,&nbsp;Amy H. Buck,&nbsp;Esther N. M. Nolte-‘t Hoen","doi":"10.1002/jex2.123","DOIUrl":"https://doi.org/10.1002/jex2.123","url":null,"abstract":"<p>Cells can communicate via the release and uptake of extracellular vesicles (EVs), which are nano-sized membrane vesicles that can transfer protein and RNA cargo between cells. EVs contain microRNAs and various other types of non-coding RNA, of which Y RNA is among the most abundant types. Studies on how RNAs and their binding proteins are sorted into EVs have mainly focused on comparing intracellular (cytoplasmic) levels of these RNAs to the extracellular levels in EVs. Besides overall transcriptional levels that may regulate sorting of RNAs into EVs, the process may also be driven by local intracellular changes in RNA/RBP concentrations. Changes in extracellular Y RNA have been linked to cancer and cardiovascular diseases. Although the loading of RNA cargo into EVs is generally thought to be influenced by cellular stimuli and regulated by RNA binding proteins (RBP), little is known about Y RNA shuttling into EVs. We previously reported that immune stimulation alters the levels of Y RNA in EVs independently of cytosolic Y RNA levels. This suggests that Y RNA binding proteins, and/or changes in the local Y RNA concentration at EV biogenesis sites, may affect Y RNA incorporation into EVs. Here, we investigated the subcellular distribution of Y RNA and Y RNA binding proteins in activated and non-activated THP1 macrophages. We demonstrate that Y RNA and its main binding protein Ro60 abundantly co-fractionate in organelles involved in EV biogenesis and in EVs. Cellular activation led to an increase in Y RNA concentration at EV biogenesis sites and this correlated with increased EV-associated levels of Y RNA and Ro60. These results suggest that Y RNA incorporation into EVs may be controlled by local intracellular changes in the concentration of Y RNA and their protein binding partners.</p>","PeriodicalId":73747,"journal":{"name":"Journal of extracellular biology","volume":"3 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2024-01-16","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://onlinelibrary.wiley.com/doi/epdf/10.1002/jex2.123","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"139480450","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":"Differential response of placental cells to high D-glucose and its impact on extracellular vesicle biogenesis and trafficking via small GTPase Ras-related protein RAB-7A","authors":"Carlos Palma,&nbsp;Andrew Lai,&nbsp;Katherin Scholz-Romero,&nbsp;Haarika Chittoory,&nbsp;Benjamin Van Haeringen,&nbsp;Flavio Carrion,&nbsp;Aase Handberg,&nbsp;Martha Lappas,&nbsp;Sunil R Lakhani,&nbsp;Amy E McCart Reed,&nbsp; McIntyre,&nbsp;Soumyalekshmi Nair,&nbsp;Carlos Salomon","doi":"10.1002/jex2.135","DOIUrl":"https://doi.org/10.1002/jex2.135","url":null,"abstract":"<p>Placental extracellular vesicles (EVs) can be found in the maternal circulation throughout gestation, and their concentration, content and bioactivity are associated with pregnancy outcomes, including gestational diabetes mellitus (GDM). However, the effect of changes in the maternal microenvironment on the mechanisms associated with the secretion of EVs from placental cells remains to be fully established. Here, we evaluated the effect of high glucose on proteins associated with the trafficking and release of different populations of EVs from placental cells. BeWo and HTR8/SVneo cells were used as placental models and cultured under 5-mM D-glucose (i.e. control) or 25-mM D-glucose (high glucose). Cell-conditioned media (CCM) and cell lysate were collected after 48 h. Different populations of EVs were isolated from CCM by ultracentrifugation (i.e. pellet 2K-g, pellet 10K-g, and pellet 100K-g) and characterised by Nanoparticle Tracking Analysis. Quantitative proteomic analysis (IDA/SWATH) and multiple reaction monitoring protocols at high resolution (MRM^HR) were developed to quantify 37 proteins related to biogenesis, trafficking/release and recognition/uptake of EVs. High glucose increased the secretion of total EVs across the pellets from BeWo cells, an effect driven mainly by changes in the small EVs concentration in the CCM. Interestingly, no effect of high glucose on HTR8/SVneo cells EVs secretion was observed. High glucose induces changes in proteins associated with vesicle trafficking in BeWo cells, including Heat Shock Protein Family A (Hsp70) Member 9 (HSPA9) and Member 8 (HSPA8). For HTR8/SVneo, altered proteins including prostaglandin F2α receptor regulatory protein (FPRP), RAB5A, RAB35, RAB5B, and RB11B, STAM1 and TSG101. These proteins are associated with the secretion and trafficking of EVs, which could explain in part, changes in the levels of circulating EVs in diabetic pregnancies. Further, we identified that proteins RAB11B, PDCD6IP, STAM, HSPA9, HSPA8, SDCBP, RAB5B, RAB5A, RAB7A and ERAP1 regulate EV release in response to high and low glucose when overexpressed in cells. Interestingly, immunohistochemistry analysis of RAB7A revealed distinct changes in placental tissues obtained from women with normal glucose tolerance (NGT, <i>n</i> = 6) and those with GDM (<i>n</i> = 6), influenced by diet or insulin treatment. High glucose regulation of proteins involved in intercellular dynamics and the trafficking of multivesicular bodies to the plasma membrane in placental cells is relevant in the context of GDM pregnancies.</p>","PeriodicalId":73747,"journal":{"name":"Journal of extracellular biology","volume":"3 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2024-01-15","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://onlinelibrary.wiley.com/doi/epdf/10.1002/jex2.135","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"139474006","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":"Analysis of extracellular vesicle microRNA profiles reveals distinct blood and lymphatic endothelial cell origins","authors":"Marianne Pultar,&nbsp;Johannes Oesterreicher,&nbsp;Jaana Hartmann,&nbsp;Moritz Weigl,&nbsp;Andreas Diendorfer,&nbsp;Katharina Schimek,&nbsp;Barbara Schädl,&nbsp;Thomas Heuser,&nbsp;Marlene Brandstetter,&nbsp;Johannes Grillari,&nbsp;Peter Sykacek,&nbsp;Matthias Hackl,&nbsp;Wolfgang Holnthoner","doi":"10.1002/jex2.134","DOIUrl":"https://doi.org/10.1002/jex2.134","url":null,"abstract":"<p>Extracellular vesicles (EVs) are crucial mediators of cell-to-cell communication in physiological and pathological conditions. Specifically, EVs released from the vasculature into blood were found to be quantitatively and qualitatively different in diseases compared to healthy states. However, our understanding of EVs derived from the lymphatic system is still scarce. In this study, we compared the mRNA and microRNA (miRNA) expression in blood vascular (BEC) and lymphatic (LEC) endothelial cells. After characterization of the EVs by fluorescence-triggered flow cytometry, nanoparticle tracking analysis and cryo-transmission electron microscopy (cryo-TEM) we utilized small RNA-sequencing to characterize miRNA signatures in the EVs and identify cell-type specific miRNAs in BEC and LEC. We found miRNAs specifically enriched in BEC and LEC on the cellular as well as the extracellular vesicle level. Our data provide a solid basis for further functional in vitro and in vivo studies addressing the role of EVs in the blood and lymphatic vasculature.</p>","PeriodicalId":73747,"journal":{"name":"Journal of extracellular biology","volume":"3 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2024-01-15","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://onlinelibrary.wiley.com/doi/epdf/10.1002/jex2.134","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"139468436","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 vesicle biogenesis of three-dimensional human pluripotent stem cells in a novel Vertical-Wheel bioreactor","authors":"Laureana Muok,&nbsp;Li Sun,&nbsp;Colin Esmonde,&nbsp;Hannah Worden,&nbsp;Cynthia Vied,&nbsp;Leanne Duke,&nbsp;Shaoyang Ma,&nbsp;Olivia Zeng,&nbsp;Tristan Driscoll,&nbsp;Sunghoon Jung,&nbsp;Yan Li","doi":"10.1002/jex2.133","DOIUrl":"https://doi.org/10.1002/jex2.133","url":null,"abstract":"<p>Extracellular vesicles (EVs) secreted by human-induced pluripotent stem cells (hiPSCs) have great potential as cell-free therapies in various diseases, including prevention of blood–brain barrier senescence and stroke. However, there are still challenges in pre-clinical and clinical use of hiPSC-EVs due to the need for large-scale production of a large quantity. Vertical-Wheel bioreactors (VWBRs) have design features that allow the biomanufacturing of hiPSC-EVs using a scalable aggregate or microcarrier-based culture system under low shear stress. EV secretion by undifferentiated hiPSCs expanded as 3-D aggregates and on Synthemax II microcarriers in VWBRs were investigated. Additionally, two types of EV collection media, mTeSR and HBM, were compared. The hiPSCs were characterized by metabolite and transcriptome analysis as well as EV biogenesis markers. Protein and microRNA cargo were analysed by proteomics and microRNA-seq, respectively. The <i>in vitro</i> functional assays of microglia stimulation and proliferation were conducted. HiPSCs expanded as 3-D aggregates and on microcarriers had comparable cell number, while microcarrier culture had higher glucose consumption, higher glycolysis and lower autophagy gene expression based on mRNA-seq. The microcarrier cultures had at least 17–23 fold higher EV secretion, and EV collection in mTeSR had 2.7–3.7 fold higher yield than HBM medium. Microcarrier culture with mTeSR EV collection had a smaller EV size than other groups, and the cargo was enriched with proteins (proteomics) and miRNAs (microRNA-seq) reducing apoptosis and promoting cell proliferation (e.g. Wnt-related pathways). hiPSC-EVs demonstrated the ability of stimulating proliferation and M2 polarization of microglia <i>in vitro</i>. HiPSC expansion on microcarriers produces much higher yields of EVs than hiPSC aggregates in VWBRs. EV collection in mTeSR increases yield compared to HBM. The biomanufactured EVs from microcarrier culture in mTeSR have exosomal characteristics and are functional in microglia stimulation, which paves the ways for future in vivo anti-aging study.</p>","PeriodicalId":73747,"journal":{"name":"Journal of extracellular biology","volume":"3 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2024-01-09","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://onlinelibrary.wiley.com/doi/epdf/10.1002/jex2.133","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"139406842","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":"Distinct targeting and uptake of platelet and red blood cell-derived extracellular vesicles into immune cells","authors":"Petra Ilvonen,&nbsp;Reetta Pusa,&nbsp;Kai Härkönen,&nbsp;Saara Laitinen,&nbsp;Ulla Impola","doi":"10.1002/jex2.130","DOIUrl":"https://doi.org/10.1002/jex2.130","url":null,"abstract":"<p>Blood-derived extracellular vesicles (EVs) hold great therapeutic potential. As blood contains mixed EV populations, it is challenging to study EVs originating from different cells separately. Blood cell concentrates manufactured in blood banks offer an excellent non-invasive source of blood cell-specific EV populations. To study blood cell-specific EVs, we isolated EVs from platelet (TREVs) and red blood cell (EryEVs) concentrates and characterized them using nanoparticle tracking analysis, imaging flow cytometry, electron microscopy and western blot analysis and co-cultured them with peripheral blood mononuclear cells (PBMCs). Our aim was to use imaging flow cytometry to investigate EV interaction with PBMCs as well as study their effects on T-lymphocyte populations to better understand their possible biological functions. As a conclusion, TREVs interacted with PBMCs more than EryEVs. Distinctively, TREVs were uptaken into CD11c+ monocytes rapidly and into CD19+ B-lymphocytes in 24 h. EryEVs were not uptaken into CD11c+ monocytes before the 24-h time point, and they were only seen on the surface of lymphocytes. Neither TREVs nor EryEV were uptaken into CD3+ T-lymphocytes and no effect on T-cell populations was detected. We have previously seen similar differences in targeting PC-3 cancer cells. Further studies are needed to address the functional properties of blood cell concentrate-derived EVs. This study demonstrates that imaging flow cytometry can be used to study the distinctive differences in the interaction and uptake of EVs. Considering our current and previous results, EVs present a new valuable component for the future development of blood-derived therapeutics.</p>","PeriodicalId":73747,"journal":{"name":"Journal of extracellular biology","volume":"3 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2024-01-02","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://onlinelibrary.wiley.com/doi/epdf/10.1002/jex2.130","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"139099503","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}