A direct-imaging cryo-EM study of shedding extracellular vesicles from leukemic monocytes.

The human leukemia monocytic cell line (THP-1) is known to shed extracellular vesicles (EVs) under various stimulations. We studied the effects of two types of common stimulation types, lipopolysaccharide (LPS) and starvation conditions by high resolution cryogenic electron microscopy, namely, cryo-SEM and cryo-TEM. Cryo-SEM data of cells undergoing EV blebbing and shedding is presented here for the first time. The high-resolution images show good agreement with models describing the membrane processes of shedding. Cells that underwent a 48-h starvation treatment exhibited differing morphological features, including shrunken nucleus and elongated membrane protrusions. LPS treated cells, however, showed extensive blebbing originating from the cell membrane, in good agreement with the sizes of EVs imaged by cryo-TEM. EVs isolated from both types of stimulations were measured by nanoparticle tracking analysis (NanoSight), by which LPS-EVs samples exhibited higher concentration and smaller mean diameter, as compared to starvation-EVs. Our results suggest a difference in the effects of the two stimulation types on the shedding process and possibly on the type of EVs shed. Our unique methodologies provide an important and innovative outlook of the shedding process and on its products, paving the way to further discoveries in this developing field of research, in which much is still unknown. [ABSTRACT FROM AUTHOR]