Cells release subpopulations of exosomes with distinct molecular and functional properties

Cells release nano-sized membrane vesicles that are involved in intercellular communication by transferring biological information between cells. It is generally accepted that cells release at least three types of these extracellular vesicles (EVs): apoptotic bodies, microvesicles and exosomes. Whilst exosomes are assumed to be a homogenous population of EVs, they have a wide range of putative functions. Therefore, we hypothesized that cells release subpopulations of exosomes with distinct molecular compositions and functional properties. Exosomes were isolated from conditioned medium by differential ultracentrifugation. Sucrose density gradient centrifugation of the resulting exosome pellet revealed the presence of two distinct subpopulations, one smaller, slow migrating population (HD-Exo), and one fast migrating, larger population (LD-Exo). Interestingly, the exosome subpopulations were found to have differential effects on gene expression in recipient endothelial cells. In conclusion, we demonstrate that cells release distinct subpopulations of exosomes with unique biophysical properties and molecular compositions that elicit differential effects on recipient cells. Our data supports the heterogeneous nature of exosomes. It also highlights the importance of better discrimination between subpopulations to allow more detailed studies on exosome biology and function, and assist in the development of exosome-based diagnostics and therapeutics. H5V cells incubated with PBS and two subpopulations of exosomes (LD-Exo and HD-Exo).