Comprehensive Phenotyping of Extracellular Vesicles in Plasma of Healthy Humans - Insights Into Cellular Origin and Biological Variation.

Despite immense interest in biomarker applications of extracellular vesicles (EVs) from blood, our understanding of circulating EVs under physiological conditions in healthy humans remains limited. Using imaging and multiplex bead-based flow cytometry, we comprehensively quantified circulating EVs with respect to their cellular origin in a large cohort of healthy blood donors. We assessed coefficients of variations to characterize their biological variation and explored demographic, clinical, and lifestyle factors contributing to observed variation. Cell-specific circulating EV subsets show a wide range of concentrations that do not correlate with cell-of-origin concentrations in blood, suggesting steady-state EV subset concentrations are regulated by complex mechanisms, which differ even for EV subsets from the same cell type. Interestingly, tetraspanin+ circulating EVs largely originate from platelets and to a lesser extent from lymphocytes. Principal component analysis (PCA) and association analyses demonstrate high biological inter-individual variation in circulating EVs across healthy humans, which are only partly explained by the influence of sex, menopausal status, age and smoking on specific circulating EV and/or tetraspanin+ circulating EV subsets. No global influence of the explored subject's factors on circulating EVs was detected. Our findings provide the first comprehensive, quantitative data towards the cell-origin atlas of plasma EVs, with important implications in the clinical use of EVs as biomarkers.