TY  - JOUR
AB  - Extracellular vesicles (EVs) hold great promise as circulating biomarkers for diagnostic and therapeutic approaches. Thus, many research groups world-wide investigate important aspects of EVs including their biology and medical significance. For this, a large number of procedures and protocols has been established making it difficult to almost impossible to compare and replicate results. Consequently, diagnostic tests remain problematic to interpret, mainly because the use of reliable reference EVs as a qualified standard has not yet gained widespread acceptance. Beyond doubt, such reference EVs are key to assess EV preparations quantitatively and to establish robust quality control processes to ensure overall quality and validity of data. To further advance the establishment of such controls, we designed and generated a new class of reference EVs expressing horseradish peroxidase (HRP) to facilitate simple and reliable EV tracing during isolation and standardization of EV purification and downstream analysis processes. HRP+ EVs can be quantified easily and with unmatched sensitivity either directly via measuring HRP activity or indirectly via immunodetection of HRP on the EV surface. We demonstrate that HRP+ EVs allow the reliable quantification of absolute EV numbers in biological or medical samples to normalize clinical specimens in liquid biopsies.
AU  - Waas, D.*
AU  - Juraschitz, M.*
AU  - Chen, Y.A.*
AU  - Waltenberger, H.*
AU  - Hammerschmidt, W.
AU  - Zeidler, R.
AU  - Molinaro, S.*
AU  - Gärtner, K.*
C1  - 75270
C2  - 57883
CY  - 111 River St, Hoboken 07030-5774, Nj Usa
TI  - Development of broadly applicable reference EVs expressing horseradish peroxidase.
JO  - J. Extra. Vesicles
VL  - 14
IS  - 8
PB  - Wiley
PY  - 2025
SN  - 2001-3078
ER  - 

TY  - JOUR
AB  - The continuous emergence of multidrug-resistant bacterial pathogens poses a major global healthcare challenge, with Klebsiella pneumoniae being a prominent threat. We conducted a comprehensive study on K. pneumoniae's antibiotic resistance mechanisms, focusing on outer membrane vesicles (OMVs) and polymyxin, a last-resort antibiotic. Our research demonstrates that OMVs protect bacteria from polymyxins. OMVs derived from Polymyxin B (PB)-stressed K. pneumoniae exhibited heightened protective efficacy due to increased vesiculation, compared to OMVs from unstressed Klebsiella. OMVs also shield bacteria from different bacterial families. This was validated ex vivo and in vivo using precision cut lung slices (PCLS) and Galleria mellonella. In all models, OMVs protected K. pneumoniae from PB and reduced the associated stress response on protein level. We observed significant changes in the lipid composition of OMVs upon PB treatment, affecting their binding capacity to PB. The altered binding capacity of single OMVs from PB stressed K. pneumoniae could be linked to a reduction in the lipid A amount of their released vesicles. Although the amount of lipid A per vesicle is reduced, the overall increase in the number of vesicles results in an increased protection because the sum of lipid A and therefore PB binding sites have increased. This unravels the mechanism of the altered PB protective efficacy of OMVs from PB stressed K. pneumoniae compared to control OMVs. The lipid A-dependent protective effect against PB was confirmed in vitro using artificial vesicles. Moreover, artificial vesicles successfully protected Klebsiella from PB ex vivo and in vivo. The findings indicate that OMVs act as protective shields for bacteria by binding to polymyxins, effectively serving as decoys and preventing antibiotic interaction with the cell surface. Our findings provide valuable insights into the mechanisms underlying antibiotic cross-protection and offer potential avenues for the development of novel therapeutic interventions to address the escalating threat of multidrug-resistant bacterial infections.
AU  - Burt, M.*
AU  - Angelidou, G.*
AU  - Mais, C.N.*
AU  - Preußer, C.*
AU  - Glatter, T.*
AU  - Heimerl, T.*
AU  - Groß, R.*
AU  - Serrania, J.*
AU  - Boosarpu, G.*
AU  - Pogge von Strandmann, E.*
AU  - Müller, J.A.*
AU  - Bange, G.*
AU  - Becker, A.*
AU  - Lehmann, M.
AU  - Jonigk, D.*
AU  - Neubert, L.*
AU  - Freitag, H.*
AU  - Paczia, N.*
AU  - Schmeck, B.*
AU  - Jung, A.L.*
C1  - 70703
C2  - 55721
CY  - 111 River St, Hoboken 07030-5774, Nj Usa
TI  - Lipid A in outer membrane vesicles shields bacteria from polymyxins.
JO  - J. Extra. Vesicles
VL  - 13
IS  - 5
PB  - Wiley
PY  - 2024
SN  - 2001-3078
ER  - 

TY  - JOUR
AB  - Cell-cell interactions in the central nervous system are based on the release of molecules mediating signal exchange and providing structural and trophic support through vesicular exocytosis and the formation of extracellular vesicles. The specific mechanisms employed by each cell type in the brain are incompletely understood. Here, we explored the means of communication used by Müller cells, a type of radial glial cells in the retina, which forms part of the central nervous system. Using immunohistochemical, electron microscopic, and molecular analyses, we provide evidence for the release of distinct extracellular vesicles from endfeet and microvilli of retinal Müller cells in adult mice in vivo. We identify VAMP5 as a Müller cell-specific SNARE component that is part of extracellular vesicles and responsive to ischemia, and we reveal differences between the secretomes of immunoaffinity-purified Müller cells and neurons in vitro. Our findings suggest extracellular vesicle-based communication as an important mediator of cellular interactions in the retina.
AU  - Demais, V.*
AU  - Pohl, A.*
AU  - Wunderlich, K.A.*
AU  - Pfaller, A.M.*
AU  - Kaplan, L.*
AU  - Barthélémy, A.*
AU  - Dittrich, R.*
AU  - Puig, B.*
AU  - Giebel, B.*
AU  - Hauck, S.M.
AU  - Pfrieger, F.W.*
AU  - Grosche, A.*
C1  - 66070
C2  - 53083
TI  - Release of VAMP5-positive extracellular vesicles by retinal Müller glia in vivo.
JO  - J. Extra. Vesicles
VL  - 11
IS  - 9
PY  - 2022
SN  - 2001-3078
ER  - 

TY  - JOUR
AB  - Eukaryotic cells, including cancer cells, secrete highly heterogeneous populations of extracellular vesicles (EVs). EVs could have different subcellular origin, composition and functional properties, but tools to distinguish between EV subtypes are scarce. Here, we tagged CD63- or CD9-positive EVs secreted by triple negative breast cancer cells with Nanoluciferase enzyme, to set-up a miniaturized method to quantify secretion of these two EV subtypes directly in the supernatant of cells. We performed a cell-based high-content screening to identify clinically-approved drugs able to affect EV secretion. One of the identified hits is Homosalate, an anti-inflammatory drug found in sunscreens which robustly increased EVs' release. Comparing EVs induced by Homosalate with those induced by Bafilomycin A1, we demonstrate that: (1) the two drugs act on EVs generated in distinct subcellular compartments, and (2) EVs released by Homosalate-, but not by Bafilomycin A1-treated cells enhance resistance to anchorage loss in another recipient epithelial tumour cell line. In conclusion, we identified a new drug modifying EV release and demonstrated that under influence of different drugs, triple negative breast cancer cells release EV subpopulations from different subcellular origins harbouring distinct functional properties.
AU  - Grisard, E.C.*
AU  - Nevo, N.*
AU  - Lescure, A.*
AU  - Doll, S.
AU  - Corbé, M.*
AU  - Jouve, M.*
AU  - Lavieu, G.*
AU  - Joliot, A.*
AU  - Nery, E.D.*
AU  - Martin-Jaular, L.*
AU  - Théry, C.*
C1  - 65626
C2  - 52749
TI  - Homosalate boosts the release of tumour-derived extracellular vesicles with protection against anchorage-loss property.
JO  - J. Extra. Vesicles
VL  - 11
IS  - 7
PY  - 2022
SN  - 2001-3078
ER  - 

TY  - JOUR
AB  - Extracellular vesicles (EVs) are emerging fundamental players in viral infections by shuttling viral components, mediating immune responses and likely the spread of the virus. However, the obstacles involved in purifying EVs and removing contaminating viral particles in a reliable and effective manner bottlenecks the full potential for the development of clinical and diagnostic treatment options targeting EV. Because of the similarities in size, density, membrane composition and mode of biogenesis of EVs and virions there are no standardized approaches for virus-removal from EV preparations yet. Functional EV studies also require EV samples that are devoid of antibody contaminants. Consequently, the study of EVs in virology needs reliable and effective protocols to purify EVs and remove contaminating antibodies and viral particles. Here, we established a protocol for EV purification from hepatitis B virus (HBV)-containing plasma by a combination of size-exclusion chromatography and affinity-based purification. After purification, EV samples were free of virus-sized particles, HBV surface antigen, HBV core antigen, antibodies or infectious material. Viral genomic contamination was also decreased following purification. By using appropriate antibodies and size parameters, this protocol could potentially be applied to purification of EVs from other viral samples. In summary, we established a fast, reproducible and robust approach for the removal of HBV from EV preparations. Looking forward to the point of purifying EVs from clinical samples, this method should enable studies shedding light on the underlying mechanisms of EVs in viral infections and their diagnostic and prognostic potential.
AU  - Jung, S.
AU  - Jakobs, K.F.K.
AU  - Shein, M.*
AU  - Schütz, A.K.*
AU  - Mohr, F.*
AU  - Stadler, H.*
AU  - Stadler, D.
AU  - Lucko, A.
AU  - Altstetter, S.
AU  - Wilsch, F.
AU  - Deng, L.
AU  - Protzer, U.
C1  - 60967
C2  - 49620
CY  - 2-4 Park Square, Milton Park, Abingdon Or14 4rn, Oxon, England
TI  - Efficient and reproducible depletion of hepatitis B virus from plasma derived extracellular vesicles.
JO  - J. Extra. Vesicles
VL  - 10
IS  - 2
PB  - Taylor & Francis Ltd
PY  - 2020
SN  - 2001-3078
ER  - 

TY  - JOUR
AB  - The in vivo detection of dead cells remains a major challenge due to technical hurdles. Here, we present a novel method, where injection of fluorescent milk fat globule-EGF factor 8 protein (MFG-E8) in vivo combined with imaging flow cytometry and deep learning allows the identification of dead cells based on their surface exposure of phosphatidylserine (PS) and other image parameters. A convolutional autoencoder (CAE) was trained on defined pictures and successfully used to identify apoptotic cells in vivo. However, unexpectedly, these analyses also revealed that the great majority of PS+ cells were not apoptotic, but rather live cells associated with PS+ extracellular vesicles (EVs). During acute viral infection apoptotic cells increased slightly, while up to 30% of lymphocytes were decorated with PS+ EVs of antigen-presenting cell (APC) exosomal origin. The combination of recombinant fluorescent MFG-E8 and the CAE-method will greatly facilitate analyses of cell death and EVs in vivo.
AU  - Kranich, J.*
AU  - Chlis, N.-K.
AU  - Rausch, L.*
AU  - Latha, A.*
AU  - Schifferer, M.*
AU  - Kurz, T.*
AU  - Foltyn-Arfa Kia, A.*
AU  - Simons, M.*
AU  - Theis, F.J.
AU  - Brocker, T.*
C1  - 60670
C2  - 49582
TI  - In vivo identification of apoptotic and extracellular vesicle-bound live cells using image-based deep learning.
JO  - J. Extra. Vesicles
VL  - 9
IS  - 1
PY  - 2020
SN  - 2001-3078
ER  - 

TY  - JOUR
AB  - Extracellular vesicles (EVs) are important mediators of cell-cell communication. Intriguingly, EVs can be engineered and thus exploited for the targeted transfer of functional proteins of interest. Thus, engineered EVs may constitute attractive tools for the development of novel therapeutic interventions, like cancer immunotherapies, vaccinations or targeted drug delivery. Here, we describe a novel experimental immunotherapeutic approach for the adjuvant treatment of chronic lymphocytic leukaemia (CLL) based on engineered EVs carrying gp350, the major glycoprotein of Epstein-Barr virus (EBV), CD40L, a central immune accessory molecule and pp65, an immunodominant antigen of the human cytomegalovirus (CMV). We show that these engineered EVs specifically interact with malignant B cells from CLL patients and render these cells immunogenic to allogeneic and autologous EBV- and CMV-specific CD4+ and CD8+ T cells. Collectively, co-opting engineered EVs to re-target the strong herpesviral immunity in CLL patients to malignant cells constitutes an attractive strategy for the adjuvant treatment of a still incurable disease. : CLL: chronic lymphocytic leukaemia; EBV: Epstein-Barr virus; CMV: cytomegalovirus.
AU  - Gärtner, K.
AU  - Luckner, M.*
AU  - Wanner, G.*
AU  - Zeidler, R.
C1  - 55515
C2  - 46366
CY  - 2-4 Park Square, Milton Park, Abingdon Or14 4rn, Oxon, England
TI  - Engineering extracellular vesicles as novel treatment options: exploiting herpesviral immunity in CLL.
JO  - J. Extra. Vesicles
VL  - 8
IS  - 1
PB  - Taylor & Francis Ltd
PY  - 2019
SN  - 2001-3078
ER  - 

TY  - JOUR
AB  - The last decade has seen a sharp increase in the number of scientific publications describing physiological and pathological functions of extracellular vesicles (EVs), a collective term covering various subtypes of cell-released, membranous structures, called exosomes, microvesicles, microparticles, ectosomes, oncosomes, apoptotic bodies, and many other names. However, specific issues arise when working with these entities, whose size and amount often make them difficult to obtain as relatively pure preparations, and to characterize properly. The International Society for Extracellular Vesicles (ISEV) proposed Minimal Information for Studies of Extracellular Vesicles ("MISEV") guidelines for the field in 2014. We now update these "MISEV2014" guidelines based on evolution of the collective knowledge in the last four years. An important point to consider is that ascribing a specific function to EVs in general, or to subtypes of EVs, requires reporting of specific information beyond mere description of function in a crude, potentially contaminated, and heterogeneous preparation. For example, claims that exosomes are endowed with exquisite and specific activities remain difficult to support experimentally, given our still limited knowledge of their specific molecular machineries of biogenesis and release, as compared with other biophysically similar EVs. The MISEV2018 guidelines include tables and outlines of suggested protocols and steps to follow to document specific EV-associated functional activities. Finally, a checklist is provided with summaries of key points.
AU  - Théry, C.*
AU  - Witwer, K.W.*
AU  - Aikawa, E.*
AU  - Alcaraz, M.J.*
AU  - Anderson, J.D.*
AU  - Andriantsitohaina, R.*
AU  - Antoniou, A.C.*
AU  - Arab, T.*
AU  - Archer, F.*
AU  - Atkin-Smith, G.K.*
AU  - Ayre, D.C.*
AU  - Bach, J.M.*
AU  - Bachurski, D.*
AU  - Baharvand, H.*
AU  - Balaj, L.*
AU  - Baldacchino, S.*
AU  - Bauer, N.N.*
AU  - Baxter, A.A.*
AU  - Bebawy, M.*
AU  - Beckham, C.*
AU  - Bedina Zavec, A.*
AU  - Benmoussa, A.*
AU  - Berardi, A.C.*
AU  - Bergese, P.*
AU  - Bielska, E.*
AU  - Blenkiron, C.*
AU  - Bobis-Wozowicz, S.*
AU  - Boilard, E.*
AU  - Boireau, W.*
AU  - Bongiovanni, A.*
AU  - Borràs, F.E.*
AU  - Bosch, S.*
AU  - Boulanger, C.M.*
AU  - Breakefield, X.*
AU  - Breglio, A.M.*
AU  - Brennan, M.*
AU  - Brigstock, D.R.*
AU  - Brisson, A.*
AU  - Broekman, M.L.D.*
AU  - Bromberg, J.F.*
AU  - Bryl-Górecka, P.*
AU  - Buch, S.*
AU  - Buck, A.H.*
AU  - Burger, D.*
AU  - Busatto, S.*
AU  - Buschmann, D.*
AU  - Bussolati, B.*
AU  - Buzás, E.I.*
AU  - Byrd, J.B.*
AU  - Camussi, G.*
AU  - Carter, D.R.F.*
AU  - Gärtner, K.
AU  - Kawamura, Y.*
AU  - Keller, E.T.*
AU  - Khamari, D.*
AU  - Chen, C.*
AU  - Chen, S.*
AU  - Cheng, L.*
AU  - Chin, A.R.*
AU  - Clayton, A.*
AU  - Clerici, S.P.*
AU  - Cocks, A.*
AU  - Cocucci, E.*
AU  - Coffey, R.J.*
AU  - Cordeiro-da-Silva, A.*
AU  - Couch, Y.*
AU  - Coumans, F.A.W.*
AU  - Coyle, B.*
AU  - Crescitelli, R.*
AU  - Criado, M.F.*
AU  - D’Souza-Schorey, C.*
AU  - Das, S.*
AU  - de Candia, P.*
AU  - De Santana, E.F.*
C1  - 54858
C2  - 45903
CY  - 2-4 Park Square, Milton Park, Abingdon Or14 4rn, Oxon, England
SP  - 1535750
TI  - Minimal information for studies of extracellular vesicles 2018 (MISEV2018): A position statement of the International Society for Extracellular Vesicles and update of the MISEV2014 guidelines.
JO  - J. Extra. Vesicles
VL  - 7
IS  - 1
PB  - Taylor & Francis Ltd
PY  - 2018
SN  - 2001-3078
ER  -