The lipid and protein content of exosomes has been extensively analyzed by various techniques including Western blotting, fluorescence-activated cell sorting, immuno-EM and mass spectrometry. Exosome composition varies depending on the cell type of origin. Nevertheless, exosomes contain a number of common protein components (26). As shown in Figure 2, the cytosolic proteins present on exosomes include Rabs, which promote exosome docking and the membrane fusion events (27). The annexins, including annexin I, II, V and VI, may regulate membrane cytoskeleton dynamics and membrane fusion events (28). Several adhesion molecules such as intercellular adhesion molecule-1, CD146, CD9, milk-fat-globule EGF-factor VIII (MFG-E8), CD18, CD11a, CD11b, CD11c, CD166 and LFA-3/CD58 have also been identified in exosomal preparations (26,27). In addition, several proteins involved in apoptosis are present on exosomes including thioredoxin peroxidase II, Alix, 14-3-3 and galectin 3. Exosomes also contain heat-shock proteins Hsp70 and Hsp90, which can facilitate peptide loading onto major histocompatibility complex (MHC)I and MHCII (29). One of the characteristic features of exosomes is the tetraspanins, which include CD9, CD63, CD81 and CD82. Exosomes also carry some cell-specific proteins like MHCII and CD86 present only on exosomes isolated from antigen-presenting cells (APCs) (30) and MFG-E8/lactadherin present on exosomes from immature DCs (31). Exosomes are also enriched in proteins that participate in vesicle formation and trafficking like the lyso-bisphosphatidic acid (LBPA)-binding protein Alix (28). Other proteins detected on exosomes are the metabolic enzymes such as peroxidases, pyruvate and lipid kinases and enolase-1 (32). Consistent with their endosomal origin, exosomes typically do not contain endoplasmic reticulum, mitochondria or nuclear proteins.
Figure 2. Protein composition of exosomes indicating their name, their location (i.e. membrane bound or soluble) and in some cases their function. GDI, GTP dissociation inhibitor; ICAM1, intercellular adhesion molecule-1; CAP-1, adenylyl cyclase-associated protein; LAMP, lysosomal associated membrane protein-1; PGRL, PG regulatory-like protein.
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Similar to proteins, lipids present on exosomes are characteristic of the cell origin, with most of the lipid analytical work being performed on exosomes derived from reticulocytes (33), mast cells (34), B lymphocyte cell lines (35) and human DCs (34). The typical lipid composition of mast cell-derived exosomes includes lysophosphatidylcholine, sphingomyelin, phosphatidylcholine, phosphatidylserine, phosphatidylethanolamine, cholesterol and diglyceride (34). Although most of these lipids are also present on exosomes isolated from other cell types, and the ratios of these lipids vary. For instance, the cholesterol/phospholipid ratio is higher in B-cell-derived exosomes compared with exosomes from mast cells and reticulocytes (36). Phospholipids like LBPA accumulate in MVBs and appear to play a key role in ILV formation (37). Recent studies by de Gassart et al. have demonstrated the presence of lipid rafts on the exosomes (18). These low-density, Triton-insoluble fractions are enriched in cholesterol and glycosphingolipids and contain different acylated proteins such as glycosyl-phosphatidylinositol-anchored proteins and tyrosine kinases of the Src family (38).