title: Proteomic comparison defines novel markers to characterize heterogeneous populations of extracellular vesicle subtypes.
abstract: Extracellular vesicles (EVs) have become the focus of rising interest because of their numerous functions in physiology and pathology. Cells release heterogeneous vesicles of different sizes and intracellular origins, including small EVs formed inside endosomal compartments (i.e., exosomes) and EVs of various sizes budding from the plasma membrane. Specific markers for the analysis and isolation of different EV populations are missing, imposing important limitations to understanding EV functions. Here, EVs from human dendritic cells were first separated by their sedimentation speed, and then either by their behavior upon upward floatation into iodixanol gradients or by immuno-isolation. Extensive quantitative proteomic analysis allowing comparison of the isolated populations showed that several classically used exosome markers, like major histocompatibility complex, flotillin, and heat-shock 70-kDa proteins, are similarly present in all EVs. We identified proteins specifically enriched in small EVs, and define a set of five protein categories displaying different relative abundance in distinct EV populations. We demonstrate the presence of exosomal and nonexosomal subpopulations within small EVs, and propose their differential separation by immuno-isolation using either CD63, CD81, or CD9. Our work thus provides guidelines to define subtypes of EVs for future functional studies.
authors: Institut Curie, PSL Research University, INSERM U932, Department "Immunité et Cancer", 75248 Paris, France;
authors: Joanna
authors: J
authors: Kowal
authors: Institut Curie, PSL Research University, Centre de Recherche, Laboratoire de Spectrométrie de masse Protéomique, 75248 Paris, France.
authors: Guillaume
authors: G
authors: Arras
authors: Institut Curie, PSL Research University, INSERM U932, Department "Immunité et Cancer", 75248 Paris, France;
authors: Marina
authors: M
authors: Colombo
authors: Institut Curie, PSL Research University, INSERM U932, Department "Immunité et Cancer", 75248 Paris, France;
authors: Mabel
authors: M
authors: Jouve
authors: Institut Curie, PSL Research University, INSERM U932, Department "Immunité et Cancer", 75248 Paris, France;
authors: Jakob Paul
authors: JP
authors: Morath
authors: Institut Curie, PSL Research University, INSERM U932, Department "Immunité et Cancer", 75248 Paris, France;
authors: Bjarke
authors: B
authors: Primdal-Bengtson
authors: Institut Curie, PSL Research University, Centre de Recherche, Laboratoire de Spectrométrie de masse Protéomique, 75248 Paris, France.
authors: Florent
authors: F
authors: Dingli
authors: Institut Curie, PSL Research University, Centre de Recherche, Laboratoire de Spectrométrie de masse Protéomique, 75248 Paris, France.
authors: Damarys
authors: http://orcid.org/0000-0002-9111-8842
authors: D
authors: Loew
authors: Institut Curie, PSL Research University, INSERM U932, Department "Immunité et Cancer", 75248 Paris, France;
authors: Mercedes
authors: M
authors: Tkach
authors: Institut Curie, PSL Research University, INSERM U932, Department "Immunité et Cancer", 75248 Paris, France; clotilde.thery@curie.fr.
authors: Clotilde
authors: C
authors: Théry
mesh_terms: D015703:Antigens, CD / Q000378:metabolism*; D015415:Biomarkers / Q000378:metabolism; D055252:Cell-Derived Microparticles / Q000378:metabolism*; D018840:HSP70 Heat-Shock Proteins / Q000378:metabolism*; D006801:Humans; D008565:Membrane Proteins / Q000378:metabolism*; D040901:Proteomics
chemical_list: D015703:Antigens, CD; D015415:Biomarkers; D018840:HSP70 Heat-Shock Proteins; D008565:Membrane Proteins; C106354:flotillins
keywords: dendritic cells; ectosomes; exosomes; extracellular vesicles; microvesicles