title: A direct-imaging cryo-EM study of shedding extracellular vesicles from leukemic monocytes.
abstract: 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.
authors.affiliation: Department of Chemical Engineering and the Russell Berrie Nanotechnology Institute, Technion-Israel Institute of Technology, Haifa 3200003, Israel.
authors.forename: Na'ama
authors.initials: N
authors.lastname: Koifman
authors.affiliation: Department of Chemical Engineering and the Russell Berrie Nanotechnology Institute, Technion-Israel Institute of Technology, Haifa 3200003, Israel.
authors.forename: Idan
authors.initials: I
authors.lastname: Biran
authors.affiliation: The Bruce Rappaport Faculty of Medicine, Technion-Israel Institute of Technology, Haifa 3525433, Israel; Department of Hematology and Bone Marrow Transplantation, Rambam Health Care Campus, Haifa 3109601, Israel.
authors.forename: Anat
authors.initials: A
authors.lastname: Aharon
authors.affiliation: The Bruce Rappaport Faculty of Medicine, Technion-Israel Institute of Technology, Haifa 3525433, Israel; Department of Hematology and Bone Marrow Transplantation, Rambam Health Care Campus, Haifa 3109601, Israel.
authors.forename: Benjamin
authors.initials: B
authors.lastname: Brenner
authors.affiliation: Department of Chemical Engineering and the Russell Berrie Nanotechnology Institute, Technion-Israel Institute of Technology, Haifa 3200003, Israel. Electronic address: ishi@technion.ac.il.
authors.forename: Yeshayahu
authors.initials: Y
authors.lastname: Talmon
mesh_terms: D055252:Cell-Derived Microparticles / Q000473:pathology; D020285:Cryoelectron Microscopy / Q000379:methods*; D000067128:Extracellular Vesicles / Q000737:chemistry* / Q000473:pathology; D006801:Humans; D007938:Leukemia / Q000473:pathology*; D008070:Lipopolysaccharides / Q000494:pharmacology; D009000:Monocytes / Q000473:pathology; D013217:Starvation / Q000473:pathology; D000074084:THP-1 Cells
chemical_list: D008070:Lipopolysaccharides
keywords: Blebbing; Cell stimulation; Cryo-EM; Endotoxins; Microparticles; NTA