ABSTRACT
In recent reviews, two to six different types of eukaryotic EVs have been distinguished.16-19 Exosomes and microvesicles (also called shedding vesicles, shedding microvesicles, or microparticles) were acknowledged unanimous, and apoptotic vesicles (also called apoptotic blebs or apoptotic bodies) were acknowledged in three reviews.17-19 In one review “ectosomes,” “membrane particles,” and “exosome-like vesicles” were also distinguished.17 1.1.3.2 Revised classification of eukaryotic vesicles
We propose to distinguish four different types of eukaryotic EVs, (1) exosomes, (2) microvesicles, (3) membrane particles, and (4) apoptotic vesicles, thereby omitting “ectosomes” and “exosomelike vesicles,” because in our view there is insufficient evidence to support the existence of these types of vesicles. “Ectosomes” are omitted because their estimated diameter is based on the direct comparison of light scattered from beads, with light scattered from vesicles using flow cytometry, leading to an underestimation of the vesicle diameter.4,5,20 Furthermore, “ectosomes” were observed only in vitro, and the cells initially reported to release “ectosomes” also release microvesicles, which were unknown when the term “ectosomes” was introduced.21 The “exosome-like vesicles” are omitted because they are indistinguishable from exosomes, with electron microscopy pictures showing damaged and disrupted vesicles.22-25 1.2 Properties of Cell-Derived Vesicles
Exosomes are present in many and perhaps all biological fluids, including urine, blood, ascites, and cerebrospinal fluid,26-29 but also, for example, in a conditioned medium of cell cultures. The reported diameter of exosomes is between 30 nm and 100 nm, the density ranges between 1.13 g/mL and 1.19 g/mL, they are usually isolated by ultracentrifugation, and their morphology is often described as “cup shaped” after negative staining and visualization by transmission electron microscopy (TEM). Figure 1.1A shows exosomes from
human plasma isolated by differential centrifugation and imaged by TEM. Exosomes are surrounded by a phospholipid bilayer containing relatively high levels of cholesterol, sphingomyelin, and ceramide and containing detergent-resistant membrane domains (lipid rafts).17,19,30,31 The transmembrane proteins of exosomes have the same outward orientation as the cell, and they have many characteristic proteins, including proteins involved in membrane transport and fusion; components of the endosomal sorting complex required for transport (ESCRT) complex such as Alix; TSG101; heat shock proteins; and tetraspanins, including CD63 and CD81.16-19,30,32-34 Unfortunately, none of the before-mentioned properties is sufficient for specific exosome identification. Because the described properties of exosomes and other types of vesicles overlap considerably, there may be a sort of continuum of vesicles and vesicle types rather than clearly different archetypical types of vesicles, complicating the classification.
