medical sciences Review The Evolving Role of Caveolin-1: A Critical Regulator of Extracellular Vesicles Kareemah Ni y, Chenghao Wang y, Jonathan M Carnino and Yang Jin * Division of Pulmonary and Critical Care Medicine, Department of Medicine, Boston University Medical Campus, 72 E Concord St. R304. Boston, MA 02118, USA; [email protected] (K.N.); [email protected] (C.W.); [email protected] (J.M.C.) * Correspondence: [email protected]; Tel.: +1-617-358-1356; Fax: +1-617-536-8093 These authors contributed equally to this work and are co-first authors. y Received: 20 August 2020; Accepted: 30 October 2020; Published: 4 November 2020 Abstract: Emerging evidence suggests that extracellular vesicles (EVs) play an essential role in mediating intercellular communication and inter-organ crosstalk both at normal physiological conditions and in the pathogenesis of human diseases. EV cargos are made up of a broad spectrum of molecules including lipids, proteins, and nucleic acids such as DNA, RNA, and microRNAs. The complex EV cargo composition is cell type-specific. A dynamic change in EV cargos occurs along with extracellular stimuli and a change in the pathophysiological status of the host. Currently, the underlying mechanisms by which EVs are formed and EV cargos are selected in the absence and presence of noxious stimuli and pathogens remain incompletely explored. The term EVs refers to a heterogeneous group of vesicles generated via different mechanisms. Some EVs are formed via direct membrane budding, while the others are produced through multivesicular bodies (MVBs) or during apoptosis. Despite the complexity of EV formation and EV cargo selection, recent studies suggest that caveolin-1, a well-known structural protein of caveolae, regulates the formation and cargo selection of some EVs, such as microvesicles (MVs). In this article, we will review the current understanding of this emerging and novel role of cav-1. Keywords: caveolin-1; extracellular vesicle (EVs); EV cargo 1. Introduction 1.1. EV History, Nomenclature, and Categories Although initially described in the 1940s, extracellular vesicles (EVs) were not well characterized until only recently. These nano-sized vesicles were first described as platelet-derived particles in 1946 [1]. In the past several decades, there have been many milestones in EV research. In 1983, two reports published in the Journal of Cell Biology [2] and Cell [3] described ~50 nm-size vesicles released from maturing blood reticulocytes into the extracellular space. These nano-vesicles were then named “exosomes” and later the term “exosome complex” was used. In the 2000s, researchers found that all cells are capable of secreting vesicles, however, extracellular vesicles (EVs) were named differently in previous literature, such as microparticles (MPs), nanoparticles, and exosomes. In 2006–2007, EV cargos were identified. Further discovery has shown that EVs contain nucleotides, including RNA, microRNA (miRNA), and DNAs. Since then, researchers have had an increased interest in studying EVs. So far, EVs have been isolated from most cell types and bodily fluids including saliva, urine, nasal and alveolar bronchial lavage fluid (BALF), amniotic fluid, breast milk, plasma, serum, and seminal fluid [4–8]. Extracellular vesicles (EVs) are a group of heterogeneous vesicles with a broad size range of 20–5000 nm and are released from almost all types of cells [9]. Based on current guidelines issued from Med. Sci. 2020, 8, 46; doi:10.3390/medsci8040046 www.mdpi.com/journal/medsci Med. Sci. 2020, 8, 46 2 of 13 Med. Sci. 2020, 8, x FOR PEER REVIEW 2 of 14 thethe International SocietySociety ofof ExtracellularExtracellular Vesicles Vesicles (ISEV), (ISEV), the the nomenclature nomenclature of of EVs EVs is categorizedis categorized by theby thesize size or origin or origin of the of vesicles, the vesicles, such as such small, as medium,small, medium, and large and EVs, large or epithelial EVs, or EVsepithelial and endothelial EVs and endothelialEVs [10]. However, EVs [10]. many However, researchers many prefer researchers naming pr EVsefer using naming a former EVs category,using a former i.e., exosomes category, (EXOs), i.e., exosomesmicrovesicles (EXOs), (MVs), microvesicles and apoptotic (MVs), bodies and (ABs). apoptoti Thisc three-groupbodies (ABs). category This three-group is defined based category on not is definedonly the based size of on EVs, not only but also the thesize mechanismof EVs, but also of generation the mechanism and their of generation cell surface and markers their cell (Figure surface1). markersGenerally, (Figure ABs are 1). theGenerally, largest EVsABs withare the the largest broadest EV sizes with range, the broadest usually fromsize range, 500 nm usually to 1 µm, from similar 500 nmto the to 1 size µm, range similar of to platelets. the size range However, of platelets. ABs are However, often generated ABs are often by actively generated dying by cells.actively As dying more cells.interest As focusesmore interest on the focuses smaller on vesicles the smaller generated vesicles from generated live cells, from in the live narrower cells, in sense, the narrower EVs often sense, only EVsrefer often to MVs only and refer exosomes. to MVs Inand this exosomes. article, we In use this EVs article, as the we generic use EVs term as tothe avoid generic confusion. term to Inavoid this confusion.review, EVs In refer this review, to the previous EVs refer MVs to the and previous exosomes, MVs a narrowerand exosomes, sense a of narrower the cell-secreted sense of the vesicles. cell- secretedPreviously vesicles. named Previously MVs are the named middle-size MVs are EVs the ranging middle-size from 200 EVs to ranging 1000 nm from and resulting200 to 1000 from nm direct and resultingplasma budding. from direct Previously plasma namedbudding. EXOs Previously range from named 20 to EXOs 200 nmrange in sizefrom and 20 areto 200 generated nm in size through and aremultivesicular generated through bodies (MVBs) multivesicular in the endosomal bodies (MVBs) pathway in [ 9the]. These endosomal two subpopulations pathway [9]. ofThese EVs havetwo subpopulationsdifferent particle of sizes EVs withhave somedifferent overlap, particle a distinct sizes wi biogenesisth some overlap, pathway, a distinct and specific biogenesis surface pathway, markers and(Figure specific1 and surface Table1 ).markers (Figure 1 and Table 1). FigureFigure 1. SchematicSchematic diagram diagram of of extracellular extracellular vesicle vesicle size size and and biogenesis. biogenesis. Extracellular Extracellular vesicles vesicles (EVs) (EVs) referrefer to to a a group of heterogeneous vesicles vesicles ranging ranging from from 20 20 nm to 5 µm.µm. Small vesicles ranging ranging from from 2020 to to 200 nm used to be named exosomes, while sm smallall to medium-size vesicles vesicles ranging ranging from from 200 200 to to 500500 nm nm used to be called microvesiclesmicrovesicles (MVs). The The largest largest EVs, EVs, whic whichh used used to to be be called apoptotic bodiesbodies (ABs), (ABs), are are often often generated generated fr fromom dying dying cells. cells. The The biogenesis biogenesis of of these these EVs EVs has has distinct distinct pathways. pathways. As illustrated illustrated here, here, small small EVs EVs are are often often generated generated via via a a long long journey journey including including endosomes, endosomes, ER/Golgi, ER/Golgi, andand multivesicular multivesicular bodies (MVBs). On On the the other other hand hand,, the the other other larger larger EVs EVs can can be be produced produced via via direct direct buddingbudding from from the the plasma membrane. It It is is commonly commonly facilitated facilitated by by lipid lipid raft raft proteins proteins as as discussed discussed in in thisthis review. TheThe largest-sizelargest-size EVs, EVs, or or previously previously referred referred to asto ABs,as ABs, are are broken broken down down from from apoptotic apoptotic cells. cells. Med. Sci. 2020, 8, 46 3 of 13 Table 1. List of well-documented markers for exosomes and microvesicles. Gene Name Protein TSG101 Tumor susceptibility gene 101 CD63 CD63 antigen Small EV TSPAN3 Tetraspanin-3 markers TSPNA6 Tetraspanin-6 ADAM10 Disintegrin and metalloproteinase domain-containing protein 10 HNRNPH1 Heterogeneous nuclear ribonucleoprotein H HNRNPL heterogeneous nuclear ribonucleoprotein L VDAC1 voltage-dependent anion channel 1 VDAC2 Voltage-dependent anion-selective channel protein 2 Small-medium PHB2 Prohibitin-2 EV markers PDIA4 Protein disulfide-isomerase A4 ATP5O ATP synthase subunit O, mitochondrial SLC25A3 Phosphate carrier protein, mitochondrial RACGAP1 Rac GTPase activating protein 1 KIF23 Kinesin-like protein KIF23 1.2. EV Biogenesis ++ With the complex and various naming systems for EVs, we hereby use the term EVs to discuss extracellular vesicles as a whole. Furthermore, EVs in this article only refer to live cell-generated EVs, and not the apoptotic bodies resulting from dying cells. Despite their well-known heterogeneity, EVs share some common characteristics. First, all EVs have a lipid bilayer membrane similar to the plasma membrane of cells. Second, they carry a diverse pattern of cargos which indicate their unique pathways during biogenesis. The biogenesis of the smallest EVs (formerly called EXOs) is the most complex one. Initially, inward protrusion of the late endosomal membrane results in continuous accumulation of intraluminal vesicles (ILVs) in multivesicular bodies (MVBs), a type of late endosome