Extracellular vesicles, such as exosomes and microvesicles, are host cell-derived packages of information that allow cell-cell communication and enable cells to rid themselves of unwanted substances. The release and uptake of extracellular vesicles has important physiological functions and may also contribute to the development and propagation of inflammatory, vascular, malignant, infectious and neurodegenerative diseases. This Review describes the different types of extracellular vesicles, how they are detected and the mechanisms by which they communicate with cells and transfer information. We also describe their physiological functions in cellular interactions, such as in thrombosis, immune modulation, cell proliferation, tissue regeneration and matrix modulation, with an emphasis on renal processes. We discuss how the detection of extracellular vesicles could be utilized as biomarkers of renal disease and how they might contribute to disease processes in the kidney, such as in acute kidney injury, chronic kidney disease, renal transplantation, thrombotic microangiopathies, vasculitides, IgA nephropathy, nephrotic syndrome, urinary tract infection, cystic kidney disease and tubulopathies. Finally, we consider how the release or uptake of extracellular vesicles can be blocked, as well as the associated benefits and risks, and how extracellular vesicles might be used to treat renal diseases by delivering therapeutics to specific cells.

Extracellular vesicles are cell-derived membrane particles ranging from 30 to 5,000 nm in size, including exosomes, microvesicles, and apoptotic bodies. They are released under physiological conditions, but also upon cellular activation, senescence, and apoptosis. They play an important role in intercellular communication. Their release may also maintain cellular integrity by ridding the cell of damaging substances. This review describes the biogenesis, uptake, and detection of extracellular vesicles in addition to the impact that they have on recipient cells, focusing on mechanisms important in the pathophysiology of kidney diseases, such as thrombosis, angiogenesis, tissue regeneration, immune modulation, and inflammation. In kidney diseases, extracellular vesicles may be utilized as biomarkers, as they are detected in both blood and urine. Furthermore, they may contribute to the pathophysiology of renal disease while also having beneficial effects associated with tissue repair. Because of their role in the promotion of thrombosis, inflammation, and immune-mediated disease, they could be the target of drug therapy, whereas their favorable effects could be utilized therapeutically in acute and chronic kidney injury.

BackgroundThe complement and kallikrein-kinin systems (KKS) are activated during vascular inflammation. The aim of this study was to investigate if blockade of the KKS can affect complement activation on the endothelium during inflammation.MethodsComplement deposition on endothelial microvesicles was assayed in vasculitis patient plasma samples and controls. Plasma was perfused over glomerular endothelial cells and complement deposition assayed by flow cytometry. The effect of the kinin system was assessed using kinin receptor antagonists and C1-inhibitor. The in vivo effect was assessed in kidney sections from mice with nephrotoxic serum-induced glomerulonephritis treated with a kinin receptor antagonist.FindingsVasculitis patient plasma had significantly more C3- and C9-positive endothelial microvesicles than controls. Perfusion of patient acute-phase plasma samples over glomerular endothelial cells induced the release of significantly more complement-positive microvesicles, in comparison to remission or control plasma. Complement activation on endothelial microvesicles was reduced by kinin B1- and B2-receptor antagonists or by C1-inhibitor (the main inhibitor of the classical pathway and the KKS). Likewise, perfusion of glomerular endothelial cells with C1-inhibitor-depleted plasma induced the release of complement-positive microvesicles, which was significantly reduced by kinin-receptor antagonists or C1-inhibitor. Mice with nephrotoxic serum-induced glomerulonephritis exhibited significantly reduced glomerular C3 deposition when treated with a B1-receptor antagonist.InterpretationExcessive complement deposition on the endothelium will promote endothelial injury and the release of endothelial microvesicles. This study demonstrates that blockade of the KKS can reduce complement activation and thereby the inflammatory response on the endothelium.FundingFull details are provided in the Acknowledgements/Funding section.

Shiga toxin is the main virulence factor of non-invasive enterohemorrhagic Escherichia coli strains capable of causing hemolytic uremic syndrome. Our group has previously shown that the toxin can reach the kidney within microvesicles where it is taken up by renal cells and the vesicles release their cargo intracellularly, leading to toxin-mediated inhibition of protein synthesis and cell death. The aim of this study was to examine if recipient cells must express the globotriaosylceramide (Gb3) toxin receptor for this to occur, or if Gb3-negative cells are also susceptible after uptake of Gb3-positive and toxin-positive microvesicles. To this end we generated Gb3-positive A4GALT-transfected CHO cells, and a vector control lacking Gb3 (CHO-control cells), and decreased Gb3 synthesis in native HeLa cells by exposing them to the glycosylceramide synthase inhibitor PPMP. We used these cells, and human intestinal DLD-1 cells lacking Gb3, and exposed them to Shiga toxin 2-bearing Gb3-positive microvesicles derived from human blood cells. Results showed that only recipient cells that possessed endogenous Gb3 (CHO-Gb3 transfected and native HeLa cells) exhibited cellular injury, reduced cell metabolism and protein synthesis, after uptake of toxin-positive microvesicles. In Gb3positive cells the toxin introduced via vesicles followed the retrograde pathway and was inhibited by the retrograde transport blocker Retro-2.1. CHO-control cells, HeLa cells treated with PPMP and DLD-1 cells remained unaffected by toxin-positive microvesicles. We conclude that Shiga toxin-containing microvesicles can be taken up by Gb3-negative cells but the recipient cell must express endogenous Gb3 for the cell to be susceptible to the toxin.

Shiga toxin (Stx)-stimulated blood cells shed extracellular vesicles (EVs) which can transfer the toxin to the kidneys and lead to hemolytic uremic syndrome. The toxin can be taken up by renal cells within EVs wherein the toxin is released, ultimately leading to cell death. The mechanism by which Stx is taken up, translocated, and sequestered in EVs was addressed in this study utilizing the B-subunit that binds to the globotriaosylceramide (Gb3) receptor. We found that Stx1B was released in EVs within minutes after stimulation of HeLa cells or red blood cells, detected by live cell imaging and flow cytometry. In the presence of Retro-2.1, an inhibitor of intracellular retrograde trafficking, a continuous release of Stx-positive EVs occurred. EVs from HeLa cells possess the Gb3 receptor on their membrane, and EVs from cells that were treated with a glycosylceramide synthase inhibitor, to reduce Gb3, bound significantly less Stx1B. Stx1B was detected both on the membrane and within the shed EVs. Stx1B was incubated with EVs derived from blood cells, in the absence of cells, and was shown to bind to, and be taken up by, these EVs, as demonstrated by electron microscopy. Using a membrane translocation assay we demonstrated that Stx1B was taken up by blood cell- and HeLa-derived EVs, an effect enhanced by chloropromazine or methyl-ß-cyclodextrin, suggesting toxin transfer within the membrane. This is a novel mechanism by which EVs derived from blood cells can sequester their toxic content, possibly to evade the host response.