Michel Record

Extracellular vesicles (EVs) are membraneous vesicles released by a variety of cells into their microenvironment. Recent studies have elucidated the role of EVs in intercellular communication, pathogenesis, drug, vaccine and gene-vector delivery, and as possible reservoirs of biomarkers. These findings have generated immense interest, along with an exponential increase in molecular data pertaining to EVs. Here, we describe Vesiclepedia, a manually curated compendium of molecular data (lipid, RNA, and protein) identified in different classes of EVs from more than 300 independent studies published over the past several years. Even though databases are indispensable resources for the scientific community, recent studies have shown that more than 50% of the databases are not regularly updated. In addition, more than 20% of the database links are inactive. To prevent such database and link decay, we have initiated a continuous community annotation project with the active involvement of EV researchers. The EV research community can set a gold standard in data sharing with Vesiclepedia, which could evolve as a primary resource for the field.

Exosomes are small vesicles secreted from multivesicular bodies, which are able to stimulate the immune system leading to tumour cell eradication. We have analysed lipids of exosomes secreted either upon stimulation from rat mast cells (RBL-2H3 cells), or constitutively from human dendritic cells. As compared with parent cells, exosomes displayed an enrichment in sphingomyelin, but not in cholesterol. Phosphatidylcholine content was decreased, but an enrichment was noted in disaturated molecular species as in phosphatidylethanolamines. Lyso(bis)phosphatidic acid was not enriched in exosomes as compared with cells. Fluorescence anisotropy demonstrated an increase in exosome-membrane rigidity from pH 5 to 7, suggesting their membrane reorganization between the acidic multivesicular body compartment and the neutral outer cell medium. NMR analysis established a bilayer organization of exosome membrane, and ESR studies using 16-doxyl stearic acid demonstrated a higher flip-flop of lipids between the two leaflets as compared with plasma membrane. In addition, the exosome membrane exhibited no asymmetrical distribution of phosphatidylethanolamines. Therefore exosome membrane displays a similar content of the major phospholipids and cholesterol, and is organized as a lipid bilayer with a random distribution of phosphatidylethanolamines. In addition, we observed tight lipid packing at neutral pH and a rapid flip-flop between the two leaflets of exosome membranes. These parameters could be used as a hallmark of exosomes.

Exosomes are bioactive vesicles released from multivesicular bodies (MVB) by intact cells and participate in intercellular signaling. We investigated the presence of lipid-related proteins and bioactive lipids in RBL-2H3 exosomes. Besides a phospholipid scramblase and a fatty acid binding protein, the exosomes contained the whole set of phospholipases (A2, C, and D) together with interacting proteins such as aldolase A and Hsp 70. They also contained the phospholipase D (PLD) / phosphatidate phosphatase 1 (PAP1) pathway leading to the formation of diglycerides. RBL-2H3 exosomes also carried members of the three phospholipase A2 classes: the calcium-dependent cPLA(2)-IVA, the calcium-independent iPLA(2)-VIA, and the secreted sPLA(2)-IIA and V. Remarkably, almost all members of the Ras GTPase superfamily were present, and incubation of exosomes with GTPgammaS triggered activation of phospholipase A(2) (PLA(2))and PLD(2). A large panel of free fatty acids, including arachidonic acid (AA) and derivatives such as prostaglandin E(2) (PGE(2)) and 15-deoxy-Delta(12,14)-prostaglandinJ(2) (15-d PGJ(2)), were detected. We observed that the exosomes were internalized by resting and activated RBL cells and that they accumulated in an endosomal compartment. Endosomal concentrations were in the micromolar range for prostaglandins; i.e., concentrations able to trigger prostaglandin-dependent biological responses. Therefore exosomes are carriers of GTP-activatable phospholipases and lipid mediators from cell to cell.