**2. EVs—classification and key components**

Classified by their biogenesis, size, morphology and function, there are three main EV categories—exosomes, microvesicles and apoptotic bodies (**Figure 1**) [16–18].

Although exosomes, microvesicles and apoptotic bodies are distinct from one another, there is a partial overlap among their respective size range and composition. Although many different methods have been previously deployed to isolate EVs from their sample sources (a notable example being ultracentrifugation in isolating and purifying exosomes and microvesicles [19–22]), these methods are unable to provide an accurate attribution of unique characteristics to each EV category. This is due to the complex nature of EVs, such that different size ranges can be derived from the same EV source depending on the isolation technique used [23]. As such, this review will mainly elaborate on EVs in general, unless otherwise stated.

Apart from biogenesis, size and morphology, each EV category possesses its own unique set of key proteins, lipids and nucleic acids (**Table 1**). Being able to differentiate EV categories based on their key components is vital in understanding their specific roles in both normal and pathological conditions. In general, all EVs possess cell adhesion proteins [13, 14, 17, 18, 24–28], heat-shock proteins [13, 14, 18, 25, 28–30], biogenesis-associated proteins [13, 14, 17, 18, 24, 25, 28], fusion proteins [13, 14, 18, 25], cell-type specific proteins [13, 14, 18, 27, 28], cytoskeletal proteins [13, 18], signalling molecules [13, 14, 28, 31], enzymes [13, 25, 28], messenger ribonucleic acid (mRNA), micro ribonucleic acid (miRNA), non-coding ribonucleic

#### **Figure 1.**

*Biogenesis, size, morphology and function of exosomes, microvesicles and apoptotic bodies. (A) Exosomes (spheroid shape, 30–200 nm) are mainly involved in regulating intercellular communication. Their formation begins when the plasma membrane undergoes endocytosis to generate an early endosome. Intraluminal vesicles (ILVs) within the endosome are then formed from the inward budding of the endosomal membrane, resulting in a multivesicular body (MVB). This process is facilitated by either endosomal sorting complex required for transport (ESCRT)-dependent or -independent mechanisms [1, 13]. The MVB finally fuses with the plasma membrane to release the ILVs as exosomes. (B) Microvesicles (irregular shape, 100–1000 nm), like exosomes, also regulate intercellular communication. They are formed via budding from the plasma membrane directly without going through endocytic processes. (C) Apoptotic bodies (variable shape, usually >1000 nm but can be as small as 50 nm [14, 15]) are formed only during cell apoptosis, during which the post-apoptotic cell bulges outwards to form vesicles for easier removal by macrophages (created with BioRender.com).*

acid (RNA), phosphatidylethanolamine, sphingolipids and higher levels of phosphatidylserine (PS) than the cell plasma membrane [24, 25, 28, 35].

The distinct protein, lipid and nucleic acid profiles of each category might be correlated with its formation processes and functions. Both exosomes and microvesicles consist of key protein components which are responsible for cell-to-cell communication [18], such as glycoproteins [18, 28], membrane signalling receptors, growth factors and cytokines [18, 25], while apoptotic bodies do not. This is most likely because exosomes and microvesicles are meant to reach target cells, while apoptotic bodies are merely the means for discarding dead cells. Microvesicles and apoptotic bodies consist of other cytoplasmic proteins which seem to be less prominent in exosomes [13]. This might be due to the similar "budding/bulging" nature of the biogenesis of microvesicles and apoptotic bodies from the cytoplasmic membrane directly, a characteristic that differs from the endocytic-driven biogenesis of exosomes. Unlike exosomes and microvesicles, apoptotic bodies are composed of chromosomal deoxyribonucleic acid (DNA) fragments, chromatin remnants, cytosol portions, degraded proteins and cell organelles from dead cells [25, 35], indicative of their role in removing dead cells.

EVs also possess additional key features according to the specific cell line they originate from (**Table 2**). In general, cancer cells consist of higher levels of sphingolipids, glycerophospholipids, sterol lipids, ceramide, phosphatidic acid and matrix metalloproteinases like a disintegrin and metalloproteinase domain-containing protein 10 (ADAM10), while non-cancer cells consist of higher levels of prenol lipids, glycerolipids and fatty acids [24, 83].


#### *Extracellular Vesicles - Role in Diseases, Pathogenesis and Therapy*


**Table 1.** *Classification of key components of EVs by their main categories.*

*Extracellular Vesicles and Their Interplay with Biological Membranes DOI: http://dx.doi.org/10.5772/intechopen.101297*



*Extracellular Vesicles and Their Interplay with Biological Membranes DOI: http://dx.doi.org/10.5772/intechopen.101297*

*Abbreviations: ABC: adenosine triphosphate-binding cassette, ADAM: A disintegrin and metalloproteinase domaincontaining protein, ADP: adenosine diphosphate, AP: adaptor related protein complex, ARFGEF: Rho/Rac guanine nucleotide exchange factor, BROX: BRO1 domain and CAAX motif containing, CA: carbohydrate antigen, CAV: caveolin, CLDN: claudin, CLSTN: calsyntenin, CLT: clathrin light chain, COP: coatomer protein complex, CXCR: CXC chemokine receptor, EDIL: EGF like repeats and discoidin domains, EEA: early endosome antigen, EGFR: epidermal growth factor receptor, EGF: Epidermal growth factor, EpCAM: epithelial cell adhesion molecule, FLOT: flotillin, GN: guanine nucleotide-binding protein, GP: glycoprotein, GRB: growth factor receptor-bound protein, HLA: human leukocyte antigen, HMG: high-mobility group, HNRNP: heterogeneous nuclear ribonucleoprotein, Lamp: lysosomeassociated membrane protein, LDLR: low-density lipoprotein receptor, LDL: low-density lipoprotein, LFA: lymphocyte function-associated antigen, LMP1: Epstein-Barr virus latent membrane protein 1, MAPK: mitogen-activated protein kinase, MCP: membrane cofactor protein, MET: mesenchymal-epithelial transition facror, MFG-E: milk fat globule-EGF factor, MHC: major histocompatibility complex, miRNA: microribonucleic acid, MMP: matrix metalloproteinase, Muc: mucin, PF: platelet factor, RAET: retinoic acid early transcript, Rab: Ras-associated binding, REEP: receptor expression-enhancing protein, RHOG: Ras homolog family member G, RRAS: RAS-related protein R-Ras, SDF: stromal cell-derived factor, SNT: syntrophin, TfR: transferrin receptor, TNF: tumour necrosis factor, TNFR: tumour necrosis factor receptor, TNFRSF: TNF receptor superfamily, TNFAIP: TNF alpha-induced protein, TRAIL: tumour necrosis factor-related apoptosis-inducing ligand, TYRP: tyrosinase-related protein, ULBP: UL16 binding protein.*

#### **Table 2.**

*Classification of additional key components of EVs by their specific cell lines.*
