Multiple extracellular vesicle types in peritoneal dialysis effluent are prominent and contain known biomarkers

Peritoneal dialysis inevitability results in activation of inflammatory processes and its efficiency is highly variable between patients. An improved method to isolate biomarkers and study pathophysiological mechanisms in peritoneal dialysis effluent (PDE) is expected to be of much benefit for the development of this treatment approach and help with patient management. Extracellular vesicles (EVs) are released as part of normal cellular processes. Their proteome is expected to reflect both type and health of their cell of origin. Although there is a significant interest in using EVs for "liquid biopsies", little is reported of their presence or composition in plentiful dialysis waste fluids, including peritoneal dialysis effluent (PDE). Here we determined the presence of EVs in PDE and subsequently characterized their proteome. EVs were first isolated from PDE using differential centrifugation, then a further enrichment using size exclusion chromatography (SEC) was performed. The presence of EVs was demonstrated using transmission electron microscopy, and their particle counts were investigated using nanoparticle tracking analysis and dynamic light scattering. Using tandem mass spectrometry, marker proteins from three types of EVs i.e. apoptotic bodies, ectosomes, and exosomes were identified. The proteomic results demonstrated that the isolation of EVs by differential centrifugation helped enrich for over 2,000 proteins normally masked by abundant proteins in PDE such as albumin and SEC markedly further improved the isolation of low abundant proteins. Gene ontology analysis of all identified proteins showed the marked enrichment of exosome and membrane-associated proteins. Over 3,700 proteins were identified in total, including many proteins with known roles in peritoneal pathophysiology. This study demonstrated the prominence of EVs in PDE and their potential value as a source of biomarkers for peritoneal dialysis patients.