Isolation and characterization of urinary extracellular vesicles: implications for biomarker discovery

Key Points Urinary extracellular vesicles comprise a wide range of biologically distinct structures with contents that are a snapshot of the life of a cell Urine is a dynamic biofluid, which changes over hours and days within an individual; therefore, at present, no single approach for the isolation of urinary extracellular vesicles is likely to comprehensively distinguish between healthy and disease states Alterations in the composition of urinary extracellular vesicles are useful experimentally and may provide information about disease pathophysiology as well as provide diagnostic end points for the study of renal disease Perhaps the greatest promise of this 'extracellular organelle' is to open a window for science into a greater understanding of cellular therapeutics Extracellular vesicles in the urine have potential as disease biomarkers. This Review discusses the different types of extracellular vesicles and the optimization of approaches to enable their isolation and purification, and to characterize their composition by high-throughput 'omics' technologies. Urine is a valuable diagnostic medium and, with the discovery of urinary extracellular vesicles, is viewed as a dynamic bioactive fluid. Extracellular vesicles are lipid-enclosed structures that can be classified into three categories: exosomes, microvesicles (or ectosomes) and apoptotic bodies. This classification is based on the mechanisms by which membrane vesicles are formed: fusion of multivesicular bodies with the plasma membranes (exosomes), budding of vesicles directly from the plasma membrane (microvesicles) or those shed from dying cells (apoptotic bodies). During their formation, urinary extracellular vesicles incorporate various cell-specific components (proteins, lipids and nucleic acids) that can be transferred to target cells. The rigour needed for comparative studies has fueled the search for optimal approaches for their isolation, purification, and characterization. RNA, the newest extracellular vesicle component to be discovered, has received substantial attention as an extracellular vesicle therapeutic, and compelling evidence suggests that ex vivo manipulation of microRNA composition may have uses in the treatment of kidney disorders. The results of these studies are building the case that urinary extracellular vesicles act as mediators of renal pathophysiology. As the field of extracellular vesicle studies is burgeoning, this Review focuses on primary data obtained from studi