Extracellular vesicle#Apoptotic bodies

Evidence for the existence of EVs and their functions was first gathered by combined applications of ultracentrifugation, electron microscopy, and functional studies during the mid-20th century.{{cite journal |vauthors=Yáñez-Mó M, Siljander PR, Andreu Z, Zavec AB, Borràs FE, Buzas EI, Buzas K, Casal E, Cappello F, Carvalho J, Colás E, Cordeiro-da Silva A, Fais S, Falcon-Perez JM, Ghobrial IM, Giebel B, Gimona M, Graner M, Gursel I, Gursel M, Heegaard NH, Hendrix A, Kierulf P, Kokubun K, Kosanovic M, Kralj-Iglic V, Krämer-Albers EM, Laitinen S, Lässer C, Lener T, Ligeti E, Linē A, Lipps G, Llorente A, Lötvall J, Manček-Keber M, Marcilla A, Mittelbrunn M, Nazarenko I, Nolte-'t Hoen EN, Nyman TA, O'Driscoll L, Olivan M, Oliveira C, Pállinger É, Del Portillo HA, Reventós J, Rigau M, Rohde E, Sammar M, Sánchez-Madrid F, Santarém N, Schallmoser K, Ostenfeld MS, Stoorvogel W, Stukelj R, Van der Grein SG, Vasconcelos MH, Wauben MH, De Wever O |display-authors=6 |title=Biological properties of extracellular vesicles and their physiological functions |journal=Journal of Extracellular Vesicles |volume=4 |issue= |pages=27066 |date=2015 |pmid=25979354 |pmc=4433489 |doi=10.3402/jev.v4.27066 }} Ultracentrifuged pellets from blood plasma were reported to have procoagulant properties by Erwin Chargaff and Randolph West in 1946.{{cite journal |vauthors=Chargaff E, West R |title=The biological significance of the thromboplastic protein of blood |journal=The Journal of Biological Chemistry |volume=166 |issue=1 |pages=189–97 |date=November 1946 |pmid=20273687 |doi=10.1016/S0021-9258(17)34997-9 |doi-access=free }} The platelet derivation and lipid-containing nature of these particles was further articulated by Peter Wolf.{{cite journal |vauthors=Wolf P |title=The nature and significance of platelet products in human plasma |journal=British Journal of Haematology |volume=13 |issue=3 |pages=269–88 |date=May 1967 |pmid=6025241 |doi=10.1111/j.1365-2141.1967.tb08741.x |s2cid=19215210 }} Around the same time, H. Clarke Anderson and Ermanno Bonucci separately described the calcifying properties of EVs in bone matrix.{{cite journal |vauthors=Anderson HC |title=Vesicles associated with calcification in the matrix of epiphyseal cartilage |journal=The Journal of Cell Biology |volume=41 |issue=1 |pages=59–72 |date=April 1969 |pmid=5775794 |pmc=2107736 |doi=10.1083/jcb.41.1.59 }}

Although the extracellular and vesicular properties of EVs had been recognized by numerous groups by the 1970s, the term "extracellular vesicle" was first used in a manuscript title in 1971.{{cite journal |vauthors=Bonucci E |title=Fine structure and histochemistry of "calcifying globules" in epiphyseal cartilage |journal=Zeitschrift für Zellforschung und Mikroskopische Anatomie |volume=103 |issue=2 |pages=192–217 |date=1970 |pmid=5412827 |doi=10.1007/BF00337312 |s2cid=8633696 }} This electron microscopy study of the flagellate freshwater alga Ochromonas danica reported release of EVs from membranes including those of flagella. Soon thereafter, EVs were seen to be released from follicular thyroid cells of the bat Myotis lucifugus during arousal from hibernation, suggesting the possible involvement of EVs in endocrine processes.{{cite journal |vauthors=Nunez EA, Wallis J, Gershon MD |title=Secretory processes in follicular cells of the bat thyroid. 3. The occurrence of extracellular vesicles and colloid droplets during arousal from hibernation |journal=The American Journal of Anatomy |volume=141 |issue=2 |pages=179–201 |date=October 1974 |pmid=4415703 |doi=10.1002/aja.1001410203 }} Reports of EVs in intestinal villi samples and, for the first time, in material from human cancer (adenoma){{cite journal |vauthors=Chandler RL, Bird RG, Bland AP |title=Letter: Particles associated with microvillous border of intestinal mucosa |journal=Lancet |volume=2 |issue=7941 |pages=931–2 |date=November 1975 |pmid=53415 |doi=10.1016/s0140-6736(75)92175-3 |s2cid=40320534 }}{{cite journal |vauthors=De Broe M, Wieme R, Roels F |title=Letter: Membrane fragments with koinozymic properties released from villous adenoma of the rectum |journal=Lancet |volume=2 |issue=7946 |pages=1214–5 |date=December 1975 |pmid=53703 |doi=10.1016/s0140-6736(75)92709-9 |s2cid=32026872 }}{{cite journal |vauthors=Benz EW, Moses HL |title=Small, virus-like particles detected in bovine sera by electron microscopy |journal=Journal of the National Cancer Institute |volume=52 |issue=6 |pages=1931–4 |date=June 1974 |pmid=4834422 |doi=10.1093/jnci/52.6.1931 }}{{cite journal |vauthors=Dalton AJ |title=Microvesicles and vesicles of multivesicular bodies versus "virus-like" particles |journal=Journal of the National Cancer Institute |volume=54 |issue=5 |pages=1137–48 |date=May 1975 |pmid=165305 |doi=10.1093/jnci/54.5.1137 }} referred back to even earlier publications that furnished similar evidence, although conclusions about EV release had not then been drawn. EVs were also described in bovine serum and cell culture conditioned medium with distinctions made between "vesicles of the multivesicular body" and "microvesicles." These studies further noted the similarities of EVs and enveloped viruses.{{cite journal |vauthors=Yim K, Borgoni S, Chahwan R |title=Letter: Serum extracellular vesicles profiling is associated with COVID-19 progression and immune responses |journal=J Extracell Biol |volume=1 |issue=4 |pages=e37 |date=April 2022 |pmid=35574251 |doi=10.1002/jex2.37 |pmc=9088353 }}

In the early- to mid-1980s, the Stahl and Johnstone labs forged a deeper understanding of the release of EVs from reticulocytes,{{cite journal |vauthors=Pan BT, Johnstone RM |title=Fate of the transferrin receptor during maturation of sheep reticulocytes in vitro: selective externalization of the receptor |journal=Cell |volume=33 |issue=3 |pages=967–78 |date=July 1983 |pmid=6307529 |doi=10.1016/0092-8674(83)90040-5 |s2cid=33216388 }}{{cite journal |vauthors=Harding C, Heuser J, Stahl P |title=Endocytosis and intracellular processing of transferrin and colloidal gold-transferrin in rat reticulocytes: demonstration of a pathway for receptor shedding |journal=European Journal of Cell Biology |volume=35 |issue=2 |pages=256–63 |date=November 1984 |pmid=6151502 |doi= }}{{cite journal |vauthors=Johnstone RM, Adam M, Hammond JR, Orr L, Turbide C |title=Vesicle formation during reticulocyte maturation. Association of plasma membrane activities with released vesicles (exosomes) |journal=The Journal of Biological Chemistry |volume=262 |issue=19 |pages=9412–20 |date=July 1987 |pmid=3597417 |doi=10.1016/S0021-9258(18)48095-7 |doi-access=free }} while progress was also made on EVs shed from tumor cells.{{cite journal |vauthors=Dvorak HF, Quay SC, Orenstein NS, Dvorak AM, Hahn P, Bitzer AM, Carvalho AC |title=Tumor shedding and coagulation |journal=Science |volume=212 |issue=4497 |pages=923–4 |date=May 1981 |pmid=7195067 |doi=10.1126/science.7195067 |bibcode=1981Sci...212..923D }} The reticulocyte research, in particular, showed that EVs could be released not only from the plasma membrane or surface of the cell, but also by fusion of the multivesicular body with the plasma membrane. During this time, EVs were described by many names, sometimes in the same manuscript, such as "shedding vesicles," "membrane fragments," "plasma membrane vesicles," "micro-vesicles/microvesicles," "exosomes," (previously used for mobile, transforming DNA elements in model organisms Drosophila and Neurospora{{cite journal |vauthors=Fox AS, Yoon SB |title=DNA-induced transformation in Drosophila: locus-specificity and the establishment of transformed stocks |journal=Proceedings of the National Academy of Sciences of the United States of America |volume=67 |issue=3 |pages=1608–15 |date=November 1970 |pmid=5274483 |pmc=283397 |doi=10.1073/pnas.67.3.1608 |bibcode=1970PNAS...67.1608F |doi-access=free }}{{cite journal |vauthors=Mishra NC, Tatum EL |title=Non-Mendelian inheritance of DNA-induced inositol independence in Neurospora |journal=Proceedings of the National Academy of Sciences of the United States of America |volume=70 |issue=12 |pages=3875–9 |date=December 1973 |pmid=4521213 |pmc=427348 |doi=10.1073/pnas.70.12.3875 |bibcode=1973PNAS...70.3875M |doi-access=free }}), "inclusion vesicles," and more, or referred to by organ of origin, such as "prostasomes" that were found to enhance sperm motility in semen.{{cite journal |vauthors=Stegmayr B, Ronquist G |title=Promotive effect on human sperm progressive motility by prostasomes |journal=Urological Research |volume=10 |issue=5 |pages=253–7 |date=1982 |pmid=6219486 |doi=10.1007/bf00255932 |s2cid=26574697 }}

The involvement of EVs in immune responses became increasingly clear in the 1990s with findings of the group of Graça Raposo and others.{{cite journal |vauthors=Raposo G, Nijman HW, Stoorvogel W, Liejendekker R, Harding CV, Melief CJ, Geuze HJ |title=B lymphocytes secrete antigen-presenting vesicles |journal=The Journal of Experimental Medicine |volume=183 |issue=3 |pages=1161–72 |date=March 1996 |pmid=8642258 |pmc=2192324 |doi=10.1084/jem.183.3.1161 }} A clinical trial of dendritic cell-derived EVs was performed in France just before the turn of the century.{{citation needed|date=March 2019}} Cells of the immune system were found capable of transferring transmembrane proteins via EVs. For example, the HIV co-receptors CCR5 and CXCR4 could be transferred from an HIV-susceptible cell to a refractory cell by "microparticles," rendering the recipient cell susceptible to infection.{{cite journal |vauthors=Mack M, Kleinschmidt A, Brühl H, Klier C, Nelson PJ, Cihak J, Plachý J, Stangassinger M, Erfle V, Schlöndorff D |display-authors=6 |title=Transfer of the chemokine receptor CCR5 between cells by membrane-derived microparticles: a mechanism for cellular human immunodeficiency virus 1 infection |journal=Nature Medicine |volume=6 |issue=7 |pages=769–75 |date=July 2000 |pmid=10888925 |doi=10.1038/77498 |s2cid=23027144 }}{{cite journal |vauthors=Rozmyslowicz T, Majka M, Kijowski J, Murphy SL, Conover DO, Poncz M, Ratajczak J, Gaulton GN, Ratajczak MZ |display-authors=6 |title=Platelet- and megakaryocyte-derived microparticles transfer CXCR4 receptor to CXCR4-null cells and make them susceptible to infection by X4-HIV |journal=AIDS |volume=17 |issue=1 |pages=33–42 |date=January 2003 |pmid=12478067 |doi=10.1097/00002030-200301030-00006 |s2cid=6619801 |doi-access=free }}

Beginning in 2006, several laboratories reported that EVs contain nucleic acids and have the ability to transfer them from cell to cell.{{cite journal |vauthors=Baj-Krzyworzeka M, Szatanek R, Weglarczyk K, Baran J, Urbanowicz B, Brański P, Ratajczak MZ, Zembala M |display-authors=6 |title=Tumour-derived microvesicles carry several surface determinants and mRNA of tumour cells and transfer some of these determinants to monocytes |journal=Cancer Immunology, Immunotherapy |volume=55 |issue=7 |pages=808–18 |date=July 2006 |pmid=16283305 |doi=10.1007/s00262-005-0075-9 |s2cid=25723677 |pmc=11030663 }}{{cite journal |vauthors=Ratajczak J, Wysoczynski M, Hayek F, Janowska-Wieczorek A, Ratajczak MZ |title=Membrane-derived microvesicles: important and underappreciated mediators of cell-to-cell communication |journal=Leukemia |volume=20 |issue=9 |pages=1487–95 |date=September 2006 |pmid=16791265 |doi=10.1038/sj.leu.2404296 |doi-access= |s2cid=6874345 }}{{cite journal |vauthors=Aliotta JM, Sanchez-Guijo FM, Dooner GJ, Johnson KW, Dooner MS, Greer KA, Greer D, Pimentel J, Kolankiewicz LM, Puente N, Faradyan S, Ferland P, Bearer EL, Passero MA, Adedi M, Colvin GA, Quesenberry PJ |display-authors=6 |title=Alteration of marrow cell gene expression, protein production, and engraftment into lung by lung-derived microvesicles: a novel mechanism for phenotype modulation |journal=Stem Cells |volume=25 |issue=9 |pages=2245–56 |date=September 2007 |pmid=17556595 |pmc=3376082 |doi=10.1634/stemcells.2007-0128 }}{{cite journal |vauthors=Valadi H, Ekström K, Bossios A, Sjöstrand M, Lee JJ, Lötvall JO |title=Exosome-mediated transfer of mRNAs and microRNAs is a novel mechanism of genetic exchange between cells |journal=Nature Cell Biology |volume=9 |issue=6 |pages=654–9 |date=June 2007 |pmid=17486113 |doi=10.1038/ncb1596 |s2cid=8599814 }}{{cite journal |vauthors=Skog J, Würdinger T, van Rijn S, Meijer DH, Gainche L, Sena-Esteves M, Curry WT, Carter BS, Krichevsky AM, Breakefield XO |display-authors=6 |title=Glioblastoma microvesicles transport RNA and proteins that promote tumour growth and provide diagnostic biomarkers |journal=Nature Cell Biology |volume=10 |issue=12 |pages=1470–6 |date=December 2008 |pmid=19011622 |pmc=3423894 |doi=10.1038/ncb1800 }}{{cite journal |vauthors=Pegtel DM, Cosmopoulos K, Thorley-Lawson DA, van Eijndhoven MA, Hopmans ES, Lindenberg JL, de Gruijl TD, Würdinger T, Middeldorp JM |display-authors=6 |title=Functional delivery of viral miRNAs via exosomes |journal=Proceedings of the National Academy of Sciences of the United States of America |volume=107 |issue=14 |pages=6328–33 |date=April 2010 |pmid=20304794 |pmc=2851954 |doi=10.1073/pnas.0914843107 |bibcode=2010PNAS..107.6328P |doi-access=free }}{{cite journal |vauthors=Chetty VK, Ghanam J, Anchan S, Reinhardt K, Brenzel A, Gelléri M, Cremer C, Grueso-Navarro E, Schneider M, von Neuhoff N, Reinhardt D, Jablonska J, Nazarenko I, Thakur BK |title=Efficient Small Extracellular Vesicles (EV) Isolation Method and Evaluation of EV-Associated DNA Role in Cell-Cell Communication in Cancer |journal=Cancers (Basel) |volume=14 |issue=9 |date=April 2022 |page=2068 |pmid=35565197 |pmc=9099953 |doi=10.3390/cancers14092068 |url= |doi-access=free }} Nucleic acids including DNAs and RNAs were even found to be functional in the recipient cell. Whether carrying DNA, RNA, surface molecules, or other factors, the involvement of EVs in cancer progression aroused considerable interest,{{cite journal |vauthors=Al-Nedawi K, Meehan B, Rak J |title=Microvesicles: messengers and mediators of tumor progression |journal=Cell Cycle |volume=8 |issue=13 |pages=2014–8 |date=July 2009 |pmid=19535896 |doi=10.4161/cc.8.13.8988 |doi-access=free }} leading to hypotheses that specific EVs could target specific cells due to "codes" displayed on their surface;{{cite journal |vauthors=Hoshino A, Costa-Silva B, Shen TL, Rodrigues G, Hashimoto A, Tesic Mark M, Molina H, Kohsaka S, Di Giannatale A, Ceder S, Singh S, Williams C, Soplop N, Uryu K, Pharmer L, King T, Bojmar L, Davies AE, Ararso Y, Zhang T, Zhang H, Hernandez J, Weiss JM, Dumont-Cole VD, Kramer K, Wexler LH, Narendran A, Schwartz GK, Healey JH, Sandstrom P, Labori KJ, Kure EH, Grandgenett PM, Hollingsworth MA, de Sousa M, Kaur S, Jain M, Mallya K, Batra SK, Jarnagin WR, Brady MS, Fodstad O, Muller V, Pantel K, Minn AJ, Bissell MJ, Garcia BA, Kang Y, Rajasekhar VK, Ghajar CM, Matei I, Peinado H, Bromberg J, Lyden D |display-authors=6 |title=Tumour exosome integrins determine organotropic metastasis |journal=Nature |volume=527 |issue=7578 |pages=329–35 |date=November 2015 |pmid=26524530 |pmc=4788391 |doi=10.1038/nature15756 |bibcode=2015Natur.527..329H }} create or enhance a metastatic niche;{{cite journal |vauthors=Peinado H, Alečković M, Lavotshkin S, Matei I, Costa-Silva B, Moreno-Bueno G, Hergueta-Redondo M, Williams C, García-Santos G, Ghajar C, Nitadori-Hoshino A, Hoffman C, Badal K, Garcia BA, Callahan MK, Yuan J, Martins VR, Skog J, Kaplan RN, Brady MS, Wolchok JD, Chapman PB, Kang Y, Bromberg J, Lyden D |display-authors=6 |title=Melanoma exosomes educate bone marrow progenitor cells toward a pro-metastatic phenotype through MET |journal=Nature Medicine |volume=18 |issue=6 |pages=883–91 |date=June 2012 |pmid=22635005 |pmc=3645291 |doi=10.1038/nm.2753 }} betray the presence of specific cancers;{{cite journal |vauthors=Melo SA, Sugimoto H, O'Connell JT, Kato N, Villanueva A, Vidal A, Qiu L, Vitkin E, Perelman LT, Melo CA, Lucci A, Ivan C, Calin GA, Kalluri R |display-authors=6 |title=Cancer exosomes perform cell-independent microRNA biogenesis and promote tumorigenesis |journal=Cancer Cell |volume=26 |issue=5 |pages=707–21 |date=November 2014 |pmid=25446899 |pmc=4254633 |doi=10.1016/j.ccell.2014.09.005 }} or be used as a therapy to target cancer cells.{{cite journal |vauthors=Kamerkar S, LeBleu VS, Sugimoto H, Yang S, Ruivo CF, Melo SA, Lee JJ, Kalluri R |display-authors=6 |title=Exosomes facilitate therapeutic targeting of oncogenic KRAS in pancreatic cancer |journal=Nature |volume=546 |issue=7659 |pages=498–503 |date=June 2017 |pmid=28607485 |pmc=5538883 |doi=10.1038/nature22341 |bibcode=2017Natur.546..498K }} Meanwhile, strides were made in the understanding of vesicle biogenesis and subtypes.{{cite journal |vauthors=Ostrowski M, Carmo NB, Krumeich S, Fanget I, Raposo G, Savina A, Moita CF, Schauer K, Hume AN, Freitas RP, Goud B, Benaroch P, Hacohen N, Fukuda M, Desnos C, Seabra MC, Darchen F, Amigorena S, Moita LF, Thery C |display-authors=6 |title=Rab27a and Rab27b control different steps of the exosome secretion pathway |journal=Nature Cell Biology |volume=12 |issue=1 |pages=19–30; sup pp 1–13 |date=January 2010 |pmid=19966785 |doi=10.1038/ncb2000 |hdl-access=free |s2cid=13935708 |hdl=10044/1/19574 }}{{cite journal |vauthors=van Niel G, Porto-Carreiro I, Simoes S, Raposo G |title=Exosomes: a common pathway for a specialized function |journal=Journal of Biochemistry |volume=140 |issue=1 |pages=13–21 |date=July 2006 |pmid=16877764 |doi=10.1093/jb/mvj128 |s2cid=43541754 |doi-access=free }}{{cite journal |vauthors=Kowal J, Arras G, Colombo M, Jouve M, Morath JP, Primdal-Bengtson B, Dingli F, Loew D, Tkach M, Théry C |display-authors=6 |title=Proteomic comparison defines novel markers to characterize heterogeneous populations of extracellular vesicle subtypes |journal=Proceedings of the National Academy of Sciences of the United States of America |volume=113 |issue=8 |pages=E968-77 |date=February 2016 |pmid=26858453 |pmc=4776515 |doi=10.1073/pnas.1521230113 |bibcode=2016PNAS..113E.968K |doi-access=free }}{{cite journal |vauthors=Tkach M, Kowal J, Théry C |title=Why the need and how to approach the functional diversity of extracellular vesicles |journal=Philosophical Transactions of the Royal Society of London. Series B, Biological Sciences |volume=373 |issue=1737 |pages=20160479 |date=January 2018 |pmid=29158309 |pmc=5717434 |doi=10.1098/rstb.2016.0479 }}

Rapid growth of the EV research community in the early 2000s led to the creation of the International Society for Extracellular Vesicles (ISEV), which has led efforts for rigor and standardization in the field including establishment of the Journal of Extracellular Vesicles. A plethora of national and regional EV societies have also been formed. In 2012, the Director's Office of the US National Institutes of Health (NIH) announced a program for funding of EV and extracellular RNA studies, the Extracellular RNA Communication Consortium (ERCC),{{cite journal |vauthors=Leslie M |title=Cell Biology. NIH effort gambles on mysterious extracellular RNAs |journal=Science |volume=341 |issue=6149 |pages=947 |date=August 2013 |pmid=23990535 |doi=10.1126/science.341.6149.947 }} which subsequently invested >USD 100 million in EV research. A second round of funding was announced in 2018. Commercial investment in EV diagnostics and therapeutics also grew during this time.{{citation needed|date=February 2023}}

Extracellular vesicles and particles (EVPs) are released by cells in different shapes and sizes. Diverse EV subtypes have been proposed, with names such as ectosomes, microvesicles, microparticles, exosomes, oncosomes, apoptotic bodies, and more.{{cite journal |vauthors=Gutierrez BC, Ancarola ME, Volpato-Rossi I, Marcilla A, Ramirez MI, Rosenzvit MC, Cucher M, Poncini CV |display-authors=6 |title=Extracellular vesicles from Trypanosoma cruzi-dendritic cell interaction show modulatory properties and confer resistance to lethal infection as a cell-free based therapy strategy |journal=Frontiers in Cellular and Infection Microbiology |volume=12 |pages=980817 |date=2022 |pmid=36467728 |pmc=9710384 |doi=10.3389/fcimb.2022.980817 |doi-access=free }}{{cite journal |vauthors=Bazzan E, Tinè M, Casara A, Biondini D, Semenzato U, Cocconcelli E, Balestro E, Damin M, Radu CM, Turato G, Baraldo S, Simioni P, Spagnolo P, Saetta M, Cosio MG |display-authors=6 |title=Critical Review of the Evolution of Extracellular Vesicles' Knowledge: From 1946 to Today |journal=International Journal of Molecular Sciences |volume=22 |issue=12 |pages=6417 |date=June 2021 |pmid=34203956 |pmc=8232679 |doi=10.3390/ijms22126417 |doi-access=free }}{{cite journal |vauthors=Gurunathan S, Kang MH, Jeyaraj M, Qasim M, Kim JH |title=Review of the Isolation, Characterization, Biological Function, and Multifarious Therapeutic Approaches of Exosomes |journal=Cells |volume=8 |issue=4 |pages=307 |date=April 2019 |pmid=30987213 |pmc=6523673 |doi=10.3390/cells8040307 |doi-access=free }} These EV subtypes have been defined by various, often overlapping, definitions, based mostly on biogenesis (cell pathway, cell or tissue identity, condition of origin).{{cite journal |vauthors=Théry C, Witwer KW, Aikawa E, Alcaraz MJ, Anderson JD, Andriantsitohaina R, Antoniou A, Arab T, Archer F, Atkin-Smith GK, Ayre DC, Bach JM, Bachurski D, Baharvand H, Balaj L, Baldacchino S, Bauer NN, Baxter AA, Bebawy M, Beckham C, Bedina Zavec A, Benmoussa A, Berardi AC, Bergese P, Bielska E, Blenkiron C, Bobis-Wozowicz S, Boilard E, Boireau W, Bongiovanni A, Borràs FE, Bosch S, Boulanger CM, Breakefield X, Breglio AM, Brennan MÁ, Brigstock DR, Brisson A, Broekman ML, Bromberg JF, Bryl-Górecka P, Buch S, Buck AH, Burger D, Busatto S, Buschmann D, Bussolati B, Buzás EI, Byrd JB, Camussi G, Carter DR, Caruso S, Chamley LW, Chang YT, Chen C, Chen S, Cheng L, Chin AR, Clayton A, Clerici SP, Cocks A, Cocucci E, Coffey RJ, Cordeiro-da-Silva A, Couch Y, Coumans FA, Coyle B, Crescitelli R, Criado MF, D'Souza-Schorey C, Das S, Datta Chaudhuri A, de Candia P, De Santana EF, De Wever O, Del Portillo HA, Demaret T, Deville S, Devitt A, Dhondt B, Di Vizio D, Dieterich LC, Dolo V, Dominguez Rubio AP, Dominici M, Dourado MR, Driedonks TA, Duarte FV, Duncan HM, Eichenberger RM, Ekström K, El Andaloussi S, Elie-Caille C, Erdbrügger U, Falcón-Pérez JM, Fatima F, Fish JE, Flores-Bellver M, Försönits A, Frelet-Barrand A, Fricke F, Fuhrmann G, Gabrielsson S, Gámez-Valero A, Gardiner C, Gärtner K, Gaudin R, Gho YS, Giebel B, Gilbert C, Gimona M, Giusti I, Goberdhan DC, Görgens A, Gorski SM, Greening DW, Gross JC, Gualerzi A, Gupta GN, Gustafson D, Handberg A, Haraszti RA, Harrison P, Hegyesi H, Hendrix A, Hill AF, Hochberg FH, Hoffmann KF, Holder B, Holthofer H, Hosseinkhani B, Hu G, Huang Y, Huber V, Hunt S, Ibrahim AG, Ikezu T, Inal JM, Isin M, Ivanova A, Jackson HK, Jacobsen S, Jay SM, Jayachandran M, Jenster G, Jiang L, Johnson SM, Jones JC, Jong A, Jovanovic-Talisman T, Jung S, Kalluri R, Kano SI, Kaur S, Kawamura Y, Keller ET, Khamari D, Khomyakova E, Khvorova A, Kierulf P, Kim KP, Kislinger T, Klingeborn M, Klinke DJ, Kornek M, Kosanović MM, Kovács ÁF, Krämer-Albers EM, Krasemann S, Krause M, Kurochkin IV, Kusuma GD, Kuypers S, Laitinen S, Langevin SM, Languino LR, Lannigan J, Lässer C, Laurent LC, Lavieu G, Lázaro-Ibáñez E, Le Lay S, Lee MS, Lee YX, Lemos DS, Lenassi M, Leszczynska A, Li IT, Liao K, Libregts SF, Ligeti E, Lim R, Lim SK, Linē A, Linnemannstöns K, Llorente A, Lombard CA, Lorenowicz MJ, Lörincz ÁM, Lötvall J, Lovett J, Lowry MC, Loyer X, Lu Q, Lukomska B, Lunavat TR, Maas SL, Malhi H, Marcilla A, Mariani J, Mariscal J, Martens-Uzunova ES, Martin-Jaular L, Martinez MC, Martins VR, Mathieu M, Mathivanan S, Maugeri M, McGinnis LK, McVey MJ, Meckes DG, Meehan KL, Mertens I, Minciacchi VR, Möller A, Møller Jørgensen M, Morales-Kastresana A, Morhayim J, Mullier F, Muraca M, Musante L, Mussack V, Muth DC, Myburgh KH, Najrana T, Nawaz M, Nazarenko I, Nejsum P, Neri C, Neri T, Nieuwland R, Nimrichter L, Nolan JP, Nolte-'t Hoen EN, Noren Hooten N, O'Driscoll L, O'Grady T, O'Loghlen A, Ochiya T, Olivier M, Ortiz A, Ortiz LA, Osteikoetxea X, Østergaard O, Ostrowski M, Park J, Pegtel DM, Peinado H, Perut F, Pfaffl MW, Phinney DG, Pieters BC, Pink RC, Pisetsky DS, Pogge von Strandmann E, Polakovicova I, Poon IK, Powell BH, Prada I, Pulliam L, Quesenberry P, Radeghieri A, Raffai RL, Raimondo S, Rak J, Ramirez MI, Raposo G, Rayyan MS, Regev-Rudzki N, Ricklefs FL, Robbins PD, Roberts DD, Rodrigues SC, Rohde E, Rome S, Rouschop KM, Rughetti A, Russell AE, Saá P, Sahoo S, Salas-Huenuleo E, Sánchez C, Saugstad JA, Saul MJ, Schiffelers RM, Schneider R, Schøyen TH, Scott A, Shahaj E, Sharma S, Shatnyeva O, Shekari F, Shelke GV, Shetty AK, Shiba K, Siljander PR, Silva AM, Skowronek A, Snyder OL, Soares RP, Sódar BW, Soekmadji C, Sotillo J, Stahl PD, Stoorvogel W, Stott SL, Strasser EF, Swift S, Tahara H, Tewari M, Timms K, Tiwari S, Tixeira R, Tkach M, Toh WS, Tomasini R, Torrecilhas AC, Tosar JP, Toxavidis V, Urbanelli L, Vader P, van Balkom BW, van der Grein SG, Van Deun J, van Herwijnen MJ, Van Keuren-Jensen K, van Niel G, van Royen ME, van Wijnen AJ, Vasconcelos MH, Vechetti IJ, Veit TD, Vella LJ, Velot É, Verweij FJ, Vestad B, Viñas JL, Visnovitz T, Vukman KV, Wahlgren J, Watson DC, Wauben MH, Weaver A, Webber JP, Weber V, Wehman AM, Weiss DJ, Welsh JA, Wendt S, Wheelock AM, Wiener Z, Witte L, Wolfram J, Xagorari A, Xander P, Xu J, Yan X, Yáñez-Mó M, Yin H, Yuana Y, Zappulli V, Zarubova J, Žėkas V, Zhang JY, Zhao Z, Zheng L, Zheutlin AR, Zickler AM, Zimmermann P, Zivkovic AM, Zocco D, Zuba-Surma EK |display-authors=6 |title=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 |journal=Journal of Extracellular Vesicles |volume=7 |issue=1 |pages=1535750 |date=2018 |pmid=30637094 |pmc=6322352 |doi=10.1080/20013078.2018.1535750 }} However, EV subtypes may also be defined by size, constituent molecules, function, or method of separation. Because of the bewildering and sometimes contradictory definitions of different EV subtypes, the current scientific consensus is that "extracellular vesicle" and variations thereon are the preferred nomenclature unless specific biogenetic origin can be demonstrated. Subtypes of EVs may be defined by:

{{blockquote|"a) physical characteristics of EVs, such as size ("small EVs" (sEVs) and "medium/large EVs" (m/lEVs), with ranges defined, for instance, respectively, <100nm or <200nm [small], or >200nm [large and/or medium]) or density (low, middle, high, with each range defined); b) biochemical composition (CD63+/CD81+- EVs, Annexin A5-stained EVs, etc.); or c) descriptions of conditions or cell of origin (podocyte EVs, hypoxic EVs, large oncosomes, apoptotic bodies)."}}

The terms "ectosome," "microvesicle" (MV), and "microparticle" (MP) refer to particles released from the surface of cells. Technically, the platelets of certain vertebrates (which bud from megakaryocytes), as well as red blood cells (e.g., of adult humans) also fulfill the consensus definition of EVs. Especially in the field of platelet research, MP has been the standard nomenclature. Formation of ectosomes may in some cases result from directed processes, and in others from shear forces or adherence of the PM to a surface.{{citation needed|date=February 2023}}

{{main|Exosome (vesicle)}}

Exosome biogenesis begins with pinching off of endosomal invaginations into the multivesicular body (MVB), forming intraluminal vesicles (ILVs). If the MVB fuses with the plasma membrane, the ILVs are released as "exosomes." The first publication to use the term "exosome" for EVs presented it as a synonym for "micro-vesicle."{{cite journal |vauthors=Trams EG, Lauter CJ, Salem N, Heine U |title=Exfoliation of membrane ecto-enzymes in the form of micro-vesicles |journal=Biochimica et Biophysica Acta (BBA) - Biomembranes |volume=645 |issue=1 |pages=63–70 |date=July 1981 |pmid=6266476 |doi=10.1016/0005-2736(81)90512-5 }} The term has also been used for EVs within specific size ranges, EVs separated using specific methods, or even all EVs.

Apoptotic bodies are EVs that are released by dying cells undergoing apoptosis. Since apoptotic cells tend to display phosphatidylserine (PS) in the outer bilayer of the cell membrane, apoptotic bodies tend to externalize PS, although other EVs may also do so. Apoptotic bodies may be quite large (microns in diameter) but may also measure in the submicron range.

In addition to the very large EVs released during apoptosis, micron-sized EVs may be produced by cancer cells, neurons, and other cells. When produced by cancer cells, these particles are termed "large oncosomes"{{cite journal |vauthors=Morello M, Minciacchi VR, de Candia P, Yang J, Posadas E, Kim H, Griffiths D, Bhowmick N, Chung LW, Gandellini P, Freeman MR, Demichelis F, Di Vizio D |display-authors=6 |title=Large oncosomes mediate intercellular transfer of functional microRNA |journal=Cell Cycle |volume=12 |issue=22 |pages=3526–36 |date=November 2013 |pmid=24091630 |pmc=3906338 |doi=10.4161/cc.26539 }}{{cite journal |vauthors=Meehan B, Rak J, Di Vizio D |title=Oncosomes - large and small: what are they, where they came from? |journal=Journal of Extracellular Vesicles |volume=5 |issue= |pages=33109 |date=2016 |pmid=27680302 |pmc=5040817 |doi=10.3402/jev.v5.33109 }} and may reach 20 microns or more in diameter. Large oncosomes can attain sizes comparable to individual cells, but they do not contain full nuclei. They have been shown to contribute to metastasis in a mouse model and a human fibroblast cell culture model of prostate cancer.{{cite journal |vauthors=Minciacchi VR, Spinelli C, Reis-Sobreiro M, Cavallini L, You S, Zandian M, Li X, Mishra R, Chiarugi P, Adam RM, Posadas EM, Viglietto G, Freeman MR, Cocucci E, Bhowmick NA, Di Vizio D |title=MYC Mediates Large Oncosome-Induced Fibroblast Reprogramming in Prostate Cancer |journal=Cancer Research |volume=77 |issue=9 |pages=2306–2317 |date=2017 |pmid=28202510 |doi=10.1158/0008-5472.CAN-16-2942 |doi-access=free }} Cellular internalization of large oncosomes can reprogram non-neoplastic brain cells to divide and migrate in primary tissue culture, and higher numbers of large oncosomes isolated from blood samples from glioblastoma patients were correlated with more advanced disease progression.{{cite journal |vauthors=Bertolini I, Terrasi A, Martelli C, Gaudioso G, Di Cristofori A, Storaci AM, Formica M, Braidotti P, Todoerti K, Ferrero S, Caroli M, Ottobrini L, Vaccari T, Vaira V |title=A GBM-like V-ATPase signature directs cell-cell tumor signaling and reprogramming via large oncosomes |journal=eBioMedicine |volume=41 |pages=225–235 |date=2019 |pmid=30737083 |doi=10.1016/j.ebiom.2019.01.051 |pmc=6441844 }}

{{main|Exopher}}

Exophers are a class of large EV, approximately four microns in diameter, observed in model organisms ranging from Caenorhabditis elegans{{cite journal |vauthors=Melentijevic I, Toth ML, Arnold ML, Guasp RJ, Harinath G, Nguyen KC, Taub D, Parker JA, Neri C, Gabel CV, Hall DH, Driscoll M |display-authors=6 |title=C. elegans neurons jettison protein aggregates and mitochondria under neurotoxic stress |journal=Nature |volume=542 |issue=7641 |pages=367–371 |date=February 2017 |pmid=28178240 |pmc=5336134 |doi=10.1038/nature21362 |bibcode=2017Natur.542..367M }} to mice.{{cite journal |vauthors=Nicolás-Ávila JA, Lechuga-Vieco AV, Esteban-Martínez L, Sánchez-Díaz M, Díaz-García E, Santiago DJ, Rubio-Ponce A, Li JL, Balachander A, Quintana JA, Martínez-de-Mena R, Castejón-Vega B, Pun-García A, Través PG, Bonzón-Kulichenko E, García-Marqués F, Cussó L, A-González N, González-Guerra A, Roche-Molina M, Martin-Salamanca S, Crainiciuc G, Guzmán G, Larrazabal J, Herrero-Galán E, Alegre-Cebollada J, Lemke G, Rothlin CV, Jimenez-Borreguero LJ, Reyes G, Castrillo A, Desco M, Muñoz-Cánoves P, Ibáñez B, Torres M, Ng LG, Priori SG, Bueno H, Vázquez J, Cordero MD, Bernal JA, Enríquez JA, Hidalgo A |display-authors=6 |title=A Network of Macrophages Supports Mitochondrial Homeostasis in the Heart |journal=Cell |volume=183 |pages=94–109 |date=2020 |issue=1 |pmid=32937105 |doi=10.1016/j.cell.2020.08.031 |s2cid=221716195 |doi-access=free |hdl=10261/226682 |hdl-access=free }} When genetically modified to express aggregating proteins, neurons were observed to sequester the aggregates into a portion of the cell and release them within a large EV called an exopher. They are hypothesized to be a mechanism for disposal of unwanted cellular material including protein aggregates and damaged organelles. Exophers can remain connected to the cell body by a thin, membranous filament resembling a tunneling nanotube.

Migrasomes are large membrane-bound EVs, ranging from 0.5 to 3 microns in diameter, that form at the ends of retraction fibers left behind when cells migrate in a process termed "migracytosis." Migrasomes can continue to fill with cytosol and expand even as the originating cell moves away. Migrasomes were first observed in rat kidney cell culture, but they are produced by mouse and human cells as well.{{cite journal |vauthors=Ma L, Li Y, Peng J, Wu D, Zhao X, Cui Y, Chen L, Yan X, Du Y, Yu L |title=Discovery of the migrasome, an organelle mediating release of cytoplasmic contents during cell migration |journal=Cell Research |volume=25 |pages=24–38 |date=2015 |issue=1 |pmid=25342562 |doi=10.1038/cr.2014.135 |pmc=4650581 }} Damaged mitochondria can be expelled from migrating cells inside of migrasomes, suggesting a functional role for this EV in mitochondrial homeostasis.{{cite journal |vauthors=Jiao H, Jiang D, Hu X, Du W, Ji L, Yang Y, Li X, Sho T, Wang X, Li Y, Wu YT, Wei YH, Hu X, Yu L |title=Mitocytosis, a migrasome-mediated mitochondrial quality-control process |journal=Cell |volume=184 |pages=2896–2910 |date=2021 |issue=11 |pmid=34048705 |doi=10.1016/j.cell.2021.04.027 |s2cid=235226529 |doi-access=free }}

Enveloped viruses are a type of EV produced under the influence of viral infection. That is, the virion is composed of cellular membranes but contains proteins and nucleic acids produced from the viral genome. Some enveloped viruses can infect other cells even without a functional virion, when genomic material is transferred via EVs. Certain non-enveloped viruses may also reproduce with assistance from EVs.{{cite journal |vauthors=Nolte-'t Hoen E, Cremer T, Gallo RC, Margolis LB |title=Extracellular vesicles and viruses: Are they close relatives? |journal=Proceedings of the National Academy of Sciences of the United States of America |volume=113 |issue=33 |pages=9155–61 |date=August 2016 |pmid=27432966 |pmc=4995926 |doi=10.1073/pnas.1605146113 |bibcode=2016PNAS..113.9155N |doi-access=free }}

Studying EVs and their cargo typically requires separation from a biological matrix (such as a complex fluid or tissue) so that the uniquely EV components can be analyzed. Many approaches have been used, including differential ultracentrifugation, density gradient ultracentrifugation, size exclusion chromatography, ultrafiltration, capillary electrophoresis, asymmetric-flow field-flow fractionation, and affinity/immunoaffinity capture methods.{{cite journal |vauthors=Mateescu B, Kowal EJ, van Balkom BW, Bartel S, Bhattacharyya SN, Buzás EI, Buck AH, de Candia P, Chow FW, Das S, Driedonks TA, Fernández-Messina L, Haderk F, Hill AF, Jones JC, Van Keuren-Jensen KR, Lai CP, Lässer C, Liegro ID, Lunavat TR, Lorenowicz MJ, Maas SL, Mäger I, Mittelbrunn M, Momma S, Mukherjee K, Nawaz M, Pegtel DM, Pfaffl MW, Schiffelers RM, Tahara H, Théry C, Tosar JP, Wauben MH, Witwer KW, Nolte-'t Hoen EN |display-authors=6 |title=Obstacles and opportunities in the functional analysis of extracellular vesicle RNA - an ISEV position paper |journal=Journal of Extracellular Vesicles |volume=6 |issue=1 |pages=1286095 |date=2017 |pmid=28326170 |pmc=5345583 |doi=10.1080/20013078.2017.1286095 }}{{cite journal |vauthors=Multia E, Tear CJ, Palviainen M, Siljander P, Riekkola ML |title=Fast isolation of highly specific population of platelet-derived extracellular vesicles from blood plasma by affinity monolithic column, immobilized with anti-human CD61 antibody |journal=Analytica Chimica Acta |volume=1091 |pages=160–168 |date=December 2019 |pmid=31679569 |doi=10.1016/j.aca.2019.09.022 |bibcode=2019AcAC.1091..160M |hdl-access=free |hdl=10138/321264 |s2cid=203147714 }}{{cite journal |vauthors=Multia E, Liangsupree T, Jussila M, Ruiz-Jimenez J, Kemell M, Riekkola ML |title=Automated On-Line Isolation and Fractionation System for Nanosized Biomacromolecules from Human Plasma |journal=Analytical Chemistry |volume=92 |issue=19 |pages=13058–13065 |date=October 2020 |pmid=32893620 |pmc=7586295 |doi=10.1021/acs.analchem.0c01986 }}{{cite journal |vauthors=Morani M, Mai TD, Krupova Z, Defrenaix P, Multia E, Riekkola ML, Taverna M |title=Electrokinetic characterization of extracellular vesicles with capillary electrophoresis: A new tool for their identification and quantification |journal=Analytica Chimica Acta |volume=1128 |pages=42–51 |date=September 2020 |pmid=32825911 |doi=10.1016/j.aca.2020.06.073 |bibcode=2020AcAC.1128...42M |hdl=10138/332354 |s2cid=221238347 |hdl-access=free }} Each method has its own recovery and purity outcomes: that is, what percentage of input EVs are obtained, and the ratio of "true" EV components to co-isolates. EV separation can also be influenced by pre-analytical variables.{{cite journal |vauthors=Lacroix R, Judicone C, Poncelet P, Robert S, Arnaud L, Sampol J, Dignat-George F |title=Impact of pre-analytical parameters on the measurement of circulating microparticles: towards standardization of protocol |journal=Journal of Thrombosis and Haemostasis |volume=10 |issue=3 |pages=437–46 |date=March 2012 |pmid=22212198 |doi=10.1111/j.1538-7836.2011.04610.x |s2cid=46519893 |doi-access=free }}{{cite journal |vauthors=Witwer KW, Buzás EI, Bemis LT, Bora A, Lässer C, Lötvall J, Nolte-'t Hoen EN, Piper MG, Sivaraman S, Skog J, Théry C, Wauben MH, Hochberg F |display-authors=6 |title=Standardization of sample collection, isolation and analysis methods in extracellular vesicle research |journal=Journal of Extracellular Vesicles |volume=2 |issue= |pages=20360 |date=2013 |pmid=24009894 |pmc=3760646 |doi=10.3402/jev.v2i0.20360 }}{{cite journal |vauthors=Coumans FA, Brisson AR, Buzas EI, Dignat-George F, Drees EE, El-Andaloussi S, Emanueli C, Gasecka A, Hendrix A, Hill AF, Lacroix R, Lee Y, van Leeuwen TG, Mackman N, Mäger I, Nolan JP, van der Pol E, Pegtel DM, Sahoo S, Siljander PR, Sturk G, de Wever O, Nieuwland R |display-authors=6 |title=Methodological Guidelines to Study Extracellular Vesicles |journal=Circulation Research |volume=120 |issue=10 |pages=1632–1648 |date=May 2017 |pmid=28495994 |doi=10.1161/CIRCRESAHA.117.309417 |doi-access=free }}{{cite journal |vauthors=Liangsupree T, Multia E, Riekkola ML |title=Modern isolation and separation techniques for extracellular vesicles |journal=Journal of Chromatography A |volume=1636 |pages=461773 |date=January 2021 |pmid=33316564 |doi=10.1016/j.chroma.2020.461773 |issn=0021-9673 |doi-access=free }}

Separated or concentrated populations of EVs may be characterized by several means. Total concentration of molecules in categories such as protein, lipid or nucleic acid. Total particle counts in a preparation can also be estimated, for example by light-scattering techniques. Each measurement technology may have a specific size range for accurate quantitation, and very small EVs (<100 nm diameter) are not detected by many technologies. Molecular "fingerprints" of populations can be obtained by "omics" technologies like proteomics, lipidomics, and RNomics, or by techniques like Raman spectroscopy. Overall levels of unique molecules can also be measured in the population, such as tetraspanins, phosphatidylserine, or species of RNA. It has been proposed that purity of an EV preparation can be estimated by examining the ratio of one population-level measurement to another, e.g., the ratio of total protein or total lipid to total particles.{{citation needed|date=February 2023}}

Specialized methods are needed to study EVs at the single particle level. The challenge for any putative single-particle method is to identify the individual EV as a single, lipid-bilayer particle, and to provide additional information such as size, surface proteins, or nucleic acid content. Methods that have been used successfully for single-EV analysis include optical microscopy and flow cytometry (for large EVs, usually >200 nm), tunable resistive pulse sensing for evaluating EV size, concentration and zeta potential, as well as electron microscopy (no lower bound) and immuno electron microscopy, single-particle interferometric reflectance imaging (down to about 40 nm), and nano-flow cytometry (also to 40 nm). Some technologies allow the study of individual EVs without extensive prior separation from a biological matrix: to give a few examples, electron microscopy and flow cytometry.

To demonstrate the presence of EVs in a preparation, as well as the relative depletion of non-EV particles or molecules, EV-enriched 'and' -depleted markers are necessary:{{cite journal |vauthors=Lötvall J, Hill AF, Hochberg F, Buzás EI, Di Vizio D, Gardiner C, Gho YS, Kurochkin IV, Mathivanan S, Quesenberry P, Sahoo S, Tahara H, Wauben MH, Witwer KW, Théry C |display-authors=6 |title=Minimal experimental requirements for definition of extracellular vesicles and their functions: a position statement from the International Society for Extracellular Vesicles |journal=Journal of Extracellular Vesicles |volume=3 |issue= |pages=26913 |date=2014 |pmid=25536934 |pmc=4275645 |doi=10.3402/jev.v3.26913 }} For example, the MISEV2018 guidelines recommend:

:At least one membrane-associated marker as evidence of the lipid bilayer (e.g., a tetraspanin protein)

:At least one cytoplasmic but ideally membrane-associated marker to show that the particle is not merely a membrane fragment

:At least one "negative" or "depleted" marker: a "deep cellular" marker, a marker of a non-EV particle, or a soluble molecule not thought to be enriched in EVs.

Usually, but not necessarily, the EV-enriched or -depleted markers are proteins that can be detected by Western blot, flow cytometry, ELISA, mass spectrometry, or other widely-available methods. Assaying for depleted markers is thought to be particularly important, as otherwise the purity of an EV preparation cannot be claimed. However, most studies of EVs prior to 2016 did not support claims of the presence of EVs by showing enriched markers, and <5% measured the presence of possible co-isolates/contaminants.{{cite journal |vauthors=Van Deun J, Mestdagh P, Agostinis P, Akay Ö, Anand S, Anckaert J, Martinez ZA, Baetens T, Beghein E, Bertier L, Berx G, Boere J, Boukouris S, Bremer M, Buschmann D, Byrd JB, Casert C, Cheng L, Cmoch A, Daveloose D, De Smedt E, Demirsoy S, Depoorter V, Dhondt B, Driedonks TA, Dudek A, Elsharawy A, Floris I, Foers AD, Gärtner K, Garg AD, Geeurickx E, Gettemans J, Ghazavi F, Giebel B, Kormelink TG, Hancock G, Helsmoortel H, Hill AF, Hyenne V, Kalra H, Kim D, Kowal J, Kraemer S, Leidinger P, Leonelli C, Liang Y, Lippens L, Liu S, Lo Cicero A, Martin S, Mathivanan S, Mathiyalagan P, Matusek T, Milani G, Monguió-Tortajada M, Mus LM, Muth DC, Németh A, Nolte-'t Hoen EN, O'Driscoll L, Palmulli R, Pfaffl MW, Primdal-Bengtson B, Romano E, Rousseau Q, Sahoo S, Sampaio N, Samuel M, Scicluna B, Soen B, Steels A, Swinnen JV, Takatalo M, Thaminy S, Théry C, Tulkens J, Van Audenhove I, van der Grein S, Van Goethem A, van Herwijnen MJ, Van Niel G, Van Roy N, Van Vliet AR, Vandamme N, Vanhauwaert S, Vergauwen G, Verweij F, Wallaert A, Wauben M, Witwer KW, Zonneveld MI, De Wever O, Vandesompele J, Hendrix A |display-authors=6 |title=EV-TRACK: transparent reporting and centralizing knowledge in extracellular vesicle research |journal=Nature Methods |volume=14 |issue=3 |pages=228–232 |date=February 2017 |pmid=28245209 |doi=10.1038/nmeth.4185 |hdl=1874/359735 |s2cid=205425936 |hdl-access=free }} Despite the high need, a list of EV contaminants is not yet available to the EV research community. A recent study suggested density-gradient-based EV separation from biofluids as an experimental set-up to compile a list of contaminants for EV, based upon differential analysis of EV-enriched fractions versus soluble protein-enriched fractions.{{cite journal |vauthors=Dhondt B, Geeurickx E, Tulkens J, Van Deun J, Vergauwen G, Lippens L, Miinalainen I, Rappu P, Heino J, Ost P, Lumen N, De Wever O, Hendrix A |display-authors=6 |title=Unravelling the proteomic landscape of extracellular vesicles in prostate cancer by density-based fractionation of urine |journal=Journal of Extracellular Vesicles |volume=9 |issue=1 |pages=1736935 |date=11 March 2020 |pmid=32284825 |pmc=7144211 |doi=10.1080/20013078.2020.1736935 |doi-access=free }} Soluble proteins in blood, the Tamm-Horsfall protein (uromodulin) in urine, or proteins of the nucleus, Golgi apparatus, endoplasmic reticulum, or mitochondria in eukaryotic cells. The latter proteins may be found in large EVs or indeed any EVs, but are expected to be less concentrated in the EV than in the cell.

A wide variety of biological functions have been ascribed to EVs.{{citation needed|date=February 2023}}

:"Trash disposal": eliminating unwanted materials

:Transfer of functional proteins

:Transfer of functional DNA and RNA

:Molecular recycling or "nutrition"

:Signaling to the recipient cell via cell-surface or endosomal receptors

:Creation of a metastatic niche for cancer

:Pathfinding through the environment

:Quorum sensing

:Mediating host-commensal or parasite/pathogen interaction

EVs have been implicated in senescence. Extracellular vesicle secretion is generally believed to increase with age due to DNA or mitochondrial damage and lipid peroxidation.{{cite journal |vauthors=Yin Y, Chen H, Wang Y, Zhang L, Wang X |title=Roles of extracellular vesicles in the aging microenvironment and age-related diseases |journal=Journal of Extracellular Vesicles |volume=10 |issue=12 |pages=e12154 |date=October 2021 |pmid=34609061 |pmc=8491204 |doi=10.1002/jev2.12154 }} It has been demonstrated that exosomes released by senescent cells have a miRNA content that contributes to aging.{{cite journal |vauthors=Xu D, Tahara H |title=The role of exosomes and microRNAs in senescence and aging |journal=Advanced Drug Delivery Reviews |volume=65 |issue=3 |pages=368–375 |date=March 2013 |pmid=22820533 |doi=10.1016/j.addr.2012.07.010 }} miRNAs play an essential role in senescence by negatively regulating the suppressors of p53, for example.{{cite journal |vauthors=Suh N |title=MicroRNA controls of cellular senescence |journal=BMB Reports |volume=51 |issue=10 |pages=493–499 |date=October 2018 |pmid=30269742 |pmc=6235093 |doi=10.5483/BMBRep.2018.51.10.209 }} 

Furthermore, EVs play a role in overall chronic inflammation. The interorgan shuttling of EVs can mean that one disease is likely to promote the advancement of another, as is the case with NAFLD and the development of atherosclerosis. EVs released from steatosis-affected hepatocytes induce the release of inflammatory molecules from endothelial cells co-cultured with them. The co-cultured cells also show increased NF-κB activity. It has thus been demonstrated that EVs released by hepatocytes under NAFLD conditions cause vascular endothelial inflammation and promote atherosclerosis.{{cite journal |vauthors=Jiang F, Chen Q, Wang W, Ling Y, Yan Y, Xia P |title=Hepatocyte-derived extracellular vesicles promote endothelial inflammation and atherogenesis via microRNA-1 |journal=Journal of Hepatology |volume=72 |issue=1 |pages=156–166 |date=January 2020 |pmid=31568800 |doi=10.1016/j.jhep.2019.09.014 |s2cid=203622470 }}

EVs also have senolytic potential. EVs harvested from cardio-sphere-derived cells in young rats have been shown to reverse senescent processes in aged rats. The older rats’ endurance and cardiovascular function improved when they received a transfusion of EVs from younger animals. It is therefore believed that EVs hold promise as an anti-aging treatment in humans.{{cite journal |vauthors=Grigorian Shamagian L, Rogers RG, Luther K, Angert D, Echavez A, Liu W, Middleton R, Antes T, Valle J, Fourier M, Sanchez L, Jaghatspanyan E, Mariscal J, Zhang R, Marbán E |display-authors=6 |title=Rejuvenating effects of young extracellular vesicles in aged rats and in cellular models of human senescence |journal=Scientific Reports |volume=13 |issue=1 |pages=12240 |date=July 2023 |pmid=37507448 |pmc=10382547 |doi=10.1038/s41598-023-39370-5 |bibcode=2023NatSR..1312240G }}

Studies indicate that EVs may have a procoagulant effect in various diseases.{{Cite journal |last1=Owens |first1=A. Phillip |last2=Mackman |first2=Nigel |date=13 May 2011 |editor-last=Weber |editor-first=Christian |editor2-last=Mause |editor2-first=Sebastian |title=Microparticles in Hemostasis and Thrombosis |journal=Circulation Research |language=en |volume=108 |issue=10 |pages=1284–1297 |doi=10.1161/CIRCRESAHA.110.233056 |pmid=21566224 |issn=0009-7330 |pmc=3144708 }} EVs can express phosphatidylserine (PS) on their surface. PS is an anionic phospholipid and PS+ EVs therefore provide a negatively charged surface which may facilitate formation of coagulation complexes. Under pathological conditions, EVs can sometimes express tissue factor (TF). TF is the most potent initiator of the coagulation cascade and is under normal conditions mainly contained to subvascular tissue.

EVs are believed to play a role in the spreading of different diseases.{{cite journal |vauthors=Yamamoto S, Azuma E, Muramatsu M, Hamashima T, Ishii Y, Sasahara M |title=Significance of Extracellular Vesicles: Pathobiological Roles in Disease |journal=Cell Structure and Function |volume=41 |issue=2 |pages=137–143 |date=November 2016 |pmid=27679938 |doi=10.1247/csf.16014 |doi-access=free }}{{cite journal |vauthors=Yim K, AlHrout A, Borgoni S, Chahwan R |title=Extracellular Vesicles Orchestrate Immune and Tumor Interaction Networks |journal=Cancers |volume=12 |issue=12 |pages=3696 |date=December 2020 |pmid=33317058 |pmc=7763968 |doi=10.3390/cancers12123696 |doi-access=free }} Studies have shown that tumor cells send EVs to send signal to target resident cells, which can lead to tumor invasion and metastasis.{{cite journal |vauthors=Cappariello A, Rucci N |title=Tumour-Derived Extracellular Vesicles (EVs): A Dangerous "Message in A Bottle" for Bone |journal=International Journal of Molecular Sciences |volume=20 |issue=19 |pages=4805 |date=September 2019 |pmid=31569680 |pmc=6802008 |doi=10.3390/ijms20194805 |doi-access=free }}{{Cite journal |last1=Makhijani |first1=Priya |last2=McGaha |first2=Tracy L. |date=2022 |title=Myeloid Responses to Extracellular Vesicles in Health and Disease |journal=Frontiers in Immunology |volume=13 |pages=818538 |doi=10.3389/fimmu.2022.818538 |doi-access=free |issn=1664-3224 |pmc=8934876 |pmid=35320943 }} In vitro studies of Alzheimer's disease have shown that astrocytes that accumulate amyloid beta release EVs that cause neuronal apoptosis.{{cite journal |vauthors=Söllvander S, Nikitidou E, Brolin R, Söderberg L, Sehlin D, Lannfelt L, Erlandsson A |title=Accumulation of amyloid-β by astrocytes result in enlarged endosomes and microvesicle-induced apoptosis of neurons |journal=Molecular Neurodegeneration |volume=11 |issue=1 |pages=38 |date=May 2016 |pmid=27176225 |pmc=4865996 |doi=10.1186/s13024-016-0098-z |doi-access=free }} The content of the EVs was also affected by the exposure to amyloid beta and higher ApoE was found in EVs secreted by astrocyte exposed to amyloid beta.{{cite journal |vauthors=Nikitidou E, Khoonsari PE, Shevchenko G, Ingelsson M, Kultima K, Erlandsson A |title=Increased Release of Apolipoprotein E in Extracellular Vesicles Following Amyloid-β Protofibril Exposure of Neuroglial Co-Cultures |journal=Journal of Alzheimer's Disease |volume=60 |issue=1 |pages=305–321 |date=2017 |pmid=28826183 |pmc=5676865 |doi=10.3233/JAD-170278 }} An oncogenic mechanism illustrates how extracellular vesicles are produced by proliferative acute lymphoblastic leukemia cells and can target and compromise a healthy hematopoiesis system during leukemia development.{{cite journal |vauthors=Georgievski A, Michel A, Thomas C, Mlamla Z, Pais de Barros JP, Lemaire-Ewing S, Garrido C, Quéré R |display-authors=6 |title=Acute lymphoblastic leukemia-derived extracellular vesicles affect quiescence of hematopoietic stem and progenitor cells |journal=Cell Death Dis. |volume=12 |issue=4 |pages=337 |year=2022 |pmid=35414137 |doi=10.1038/s41419-022-04761-5 |pmc=9005650 }}

The fate of T cells can be determined by the transfer of telomeres via EVs from APCs. T cells that acquire telomeres in such a manner regain stem-like characteristics, avoiding senescence. The creation of long-lived memory T cells via an EV injection of telomeres enhances long-term immunological memory.{{cite journal |vauthors=Lanna A, Vaz B, D'Ambra C, Valvo S, Vuotto C, Chiurchiù V, Devine O, Sanchez M, Borsellino G, Akbar AN, De Bardi M, Gilroy DW, Dustin ML, Blumer B, Karin M |display-authors=6 |title=An intercellular transfer of telomeres rescues T cells from senescence and promotes long-term immunological memory |journal=Nature Cell Biology |volume=24 |issue=10 |pages=1461–1474 |date=October 2022 |pmid=36109671 |pmc=7613731 |doi=10.1038/s41556-022-00991-z }}

It has been suggested that EVs carrying nucleic acid cargo could serve as biomarkers for disease, especially in neurological disorders where it is difficult to assess the underlying pathology directly.

EVs facilitate communication between different parts of the CNS,{{cite journal |vauthors=Agnati LF, Guidolin D, Guescini M, Genedani S, Fuxe K |title=Understanding wiring and volume transmission |journal=Brain Research Reviews |volume=64 |issue=1 |pages=137–159 |date=September 2010 |pmid=20347870 |doi=10.1016/j.brainresrev.2010.03.003 |s2cid=36665895 }} and therefore, EVs found in the blood of neurological patients contain molecules implicated in neurodegenerative diseases.{{cite journal |vauthors=Koniusz S, Andrzejewska A, Muraca M, Srivastava AK, Janowski M, Lukomska B |title=Extracellular Vesicles in Physiology, Pathology, and Therapy of the Immune and Central Nervous System, with Focus on Extracellular Vesicles Derived from Mesenchymal Stem Cells as Therapeutic Tools |journal=Frontiers in Cellular Neuroscience |volume=10 |pages=109 |date=2016 |pmid=27199663 |pmc=4852177 |doi=10.3389/fncel.2016.00109 |doi-access=free }} EVs carrying myeloid cargo, for example, have long been recognized as a biomarker of brain inflammation.{{cite journal |vauthors=Verderio C, Muzio L, Turola E, Bergami A, Novellino L, Ruffini F, Riganti L, Corradini I, Francolini M, Garzetti L, Maiorino C, Servida F, Vercelli A, Rocca M, Dalla Libera D, Martinelli V, Comi G, Martino G, Matteoli M, Furlan R |display-authors=6 |title=Myeloid microvesicles are a marker and therapeutic target for neuroinflammation |journal=Annals of Neurology |volume=72 |issue=4 |pages=610–624 |date=October 2012 |pmid=23109155 |doi=10.1002/ana.23627 |s2cid=35702508 |url=https://www.openaccessrepository.it/record/21830 }} Furthermore, nucleic acids corresponding to APP, Aβ42, BACE1, and tau protein biomarkers were found to be associated with different neurodegenerative diseases.{{cite journal |vauthors=Urbanelli L, Buratta S, Sagini K, Ferrara G, Lanni M, Emiliani C |title=Exosome-based strategies for Diagnosis and Therapy |journal=Recent Patents on CNS Drug Discovery |date=2015 |volume=10 |issue=1 |pages=10–27 |pmid=26133463 |doi=10.2174/1574889810666150702124059 }}

Using EVs to profile RNA expression patterns could therefore help diagnose certain diseases before a patient become symptomatic. Exosome Diagnostic (Cambridge, MA, USA), for example, has a patent for detecting neurodegenerative diseases and brain injury based on the measure of RNA-s (mRNA, miRNA, siRNA, or shRNA) associated with CSF-derived EVs.{{cite journal |vauthors=Skog J, Würdinger T, van Rijn S, Meijer DH, Gainche L, Sena-Esteves M, Curry WT, Carter BS, Krichevsky AM, Breakefield XO |display-authors=6 |title=Glioblastoma microvesicles transport RNA and proteins that promote tumour growth and provide diagnostic biomarkers |journal=Nature Cell Biology |volume=10 |issue=12 |pages=1470–1476 |date=December 2008 |pmid=19011622 |pmc=3423894 |doi=10.1038/ncb1800 }}

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