title: Directly visualized glioblastoma-derived extracellular vesicles transfer RNA to microglia/macrophages in the brain.
abstract: BACKGROUND
  To understand the ability of gliomas to manipulate their microenvironment, we visualized the transfer of vesicles and the effects of tumor-released extracellular RNA on the phenotype of microglia in culture and in vivo.
  
  
  METHODS
  Extracellular vesicles (EVs) released from primary human glioblastoma (GBM) cells were isolated and microRNAs (miRNAs) were analyzed. Primary mouse microglia were exposed to GBM-EVs, and their uptake and effect on proliferation and levels of specific miRNAs, mRNAs, and proteins were analyzed. For in vivo analysis, mouse glioma cells were implanted in the brains of mice, and EV release and uptake by microglia and monocytes/macrophages were monitored by intravital 2-photon microscopy, immunohistochemistry, and fluorescence activated cell sorting analysis, as well as RNA and protein levels.
  
  
  RESULTS
  Microglia avidly took up GBM-EVs, leading to increased proliferation and shifting of their cytokine profile toward immune suppression. High levels of miR-451/miR-21 in GBM-EVs were transferred to microglia with a decrease in the miR-451/miR-21 target c-Myc mRNA. In in vivo analysis, we directly visualized release of EVs from glioma cells and their uptake by microglia and monocytes/macrophages in brain. Dissociated microglia and monocytes/macrophages from tumor-bearing brains revealed increased levels of miR-21 and reduced levels of c-Myc mRNA.
  
  
  CONCLUSIONS
  Intravital microscopy confirms the release of EVs from gliomas and their uptake into microglia and monocytes/macrophages within the brain. Our studies also support functional effects of GBM-released EVs following uptake into microglia, associated in part with increased miRNA levels, decreased target mRNAs, and encoded proteins, presumably as a means for the tumor to manipulate its environs.
authors.affiliation: Departments of Neurology and Radiology, Massachusetts General Hospital and NeuroDiscovery Center, Harvard Medical School, Boston, Massachusetts (K.E.v.d.V., E.R.A., X.Z., C.L., S.P., O.M., M.H.W.C., J.S., X.O.B.); Center for Immunology and Inflammatory Diseases, Massachusetts General Hospital, Boston, Massachusetts (E.C., T.R.M., J.E.K., S.E.H.); Neuropathology Service, Massachusetts General Hospital and Department of Pathology, Harvard Medical School, Boston, Massachusetts (D.O., A.S-R.); Department of Neurology, Brigham and Women's Hospital, Harvard Medical School, Boston, Massachusetts (A.M.K.); Division of Molecular Carcinogenesis, The Netherlands Cancer Institute, Amsterdam, the Netherlands (K.E.v.d.V.).
authors.forename: Kristan E
authors.initials: KE
authors.lastname: van der Vos
authors.affiliation: Departments of Neurology and Radiology, Massachusetts General Hospital and NeuroDiscovery Center, Harvard Medical School, Boston, Massachusetts (K.E.v.d.V., E.R.A., X.Z., C.L., S.P., O.M., M.H.W.C., J.S., X.O.B.); Center for Immunology and Inflammatory Diseases, Massachusetts General Hospital, Boston, Massachusetts (E.C., T.R.M., J.E.K., S.E.H.); Neuropathology Service, Massachusetts General Hospital and Department of Pathology, Harvard Medical School, Boston, Massachusetts (D.O., A.S-R.); Department of Neurology, Brigham and Women's Hospital, Harvard Medical School, Boston, Massachusetts (A.M.K.); Division of Molecular Carcinogenesis, The Netherlands Cancer Institute, Amsterdam, the Netherlands (K.E.v.d.V.).
authors.forename: Erik R
authors.initials: ER
authors.lastname: Abels
authors.affiliation: Departments of Neurology and Radiology, Massachusetts General Hospital and NeuroDiscovery Center, Harvard Medical School, Boston, Massachusetts (K.E.v.d.V., E.R.A., X.Z., C.L., S.P., O.M., M.H.W.C., J.S., X.O.B.); Center for Immunology and Inflammatory Diseases, Massachusetts General Hospital, Boston, Massachusetts (E.C., T.R.M., J.E.K., S.E.H.); Neuropathology Service, Massachusetts General Hospital and Department of Pathology, Harvard Medical School, Boston, Massachusetts (D.O., A.S-R.); Department of Neurology, Brigham and Women's Hospital, Harvard Medical School, Boston, Massachusetts (A.M.K.); Division of Molecular Carcinogenesis, The Netherlands Cancer Institute, Amsterdam, the Netherlands (K.E.v.d.V.).
authors.forename: Xuan
authors.initials: X
authors.lastname: Zhang
authors.affiliation: Departments of Neurology and Radiology, Massachusetts General Hospital and NeuroDiscovery Center, Harvard Medical School, Boston, Massachusetts (K.E.v.d.V., E.R.A., X.Z., C.L., S.P., O.M., M.H.W.C., J.S., X.O.B.); Center for Immunology and Inflammatory Diseases, Massachusetts General Hospital, Boston, Massachusetts (E.C., T.R.M., J.E.K., S.E.H.); Neuropathology Service, Massachusetts General Hospital and Department of Pathology, Harvard Medical School, Boston, Massachusetts (D.O., A.S-R.); Department of Neurology, Brigham and Women's Hospital, Harvard Medical School, Boston, Massachusetts (A.M.K.); Division of Molecular Carcinogenesis, The Netherlands Cancer Institute, Amsterdam, the Netherlands (K.E.v.d.V.).
authors.forename: Charles
authors.initials: C
authors.lastname: Lai
authors.affiliation: Departments of Neurology and Radiology, Massachusetts General Hospital and NeuroDiscovery Center, Harvard Medical School, Boston, Massachusetts (K.E.v.d.V., E.R.A., X.Z., C.L., S.P., O.M., M.H.W.C., J.S., X.O.B.); Center for Immunology and Inflammatory Diseases, Massachusetts General Hospital, Boston, Massachusetts (E.C., T.R.M., J.E.K., S.E.H.); Neuropathology Service, Massachusetts General Hospital and Department of Pathology, Harvard Medical School, Boston, Massachusetts (D.O., A.S-R.); Department of Neurology, Brigham and Women's Hospital, Harvard Medical School, Boston, Massachusetts (A.M.K.); Division of Molecular Carcinogenesis, The Netherlands Cancer Institute, Amsterdam, the Netherlands (K.E.v.d.V.).
authors.forename: Esteban
authors.initials: E
authors.lastname: Carrizosa
authors.affiliation: Departments of Neurology and Radiology, Massachusetts General Hospital and NeuroDiscovery Center, Harvard Medical School, Boston, Massachusetts (K.E.v.d.V., E.R.A., X.Z., C.L., S.P., O.M., M.H.W.C., J.S., X.O.B.); Center for Immunology and Inflammatory Diseases, Massachusetts General Hospital, Boston, Massachusetts (E.C., T.R.M., J.E.K., S.E.H.); Neuropathology Service, Massachusetts General Hospital and Department of Pathology, Harvard Medical School, Boston, Massachusetts (D.O., A.S-R.); Department of Neurology, Brigham and Women's Hospital, Harvard Medical School, Boston, Massachusetts (A.M.K.); Division of Molecular Carcinogenesis, The Netherlands Cancer Institute, Amsterdam, the Netherlands (K.E.v.d.V.).
authors.forename: Derek
authors.initials: D
authors.lastname: Oakley
authors.affiliation: Departments of Neurology and Radiology, Massachusetts General Hospital and NeuroDiscovery Center, Harvard Medical School, Boston, Massachusetts (K.E.v.d.V., E.R.A., X.Z., C.L., S.P., O.M., M.H.W.C., J.S., X.O.B.); Center for Immunology and Inflammatory Diseases, Massachusetts General Hospital, Boston, Massachusetts (E.C., T.R.M., J.E.K., S.E.H.); Neuropathology Service, Massachusetts General Hospital and Department of Pathology, Harvard Medical School, Boston, Massachusetts (D.O., A.S-R.); Department of Neurology, Brigham and Women's Hospital, Harvard Medical School, Boston, Massachusetts (A.M.K.); Division of Molecular Carcinogenesis, The Netherlands Cancer Institute, Amsterdam, the Netherlands (K.E.v.d.V.).
authors.forename: Shilpa
authors.initials: S
authors.lastname: Prabhakar
authors.affiliation: Departments of Neurology and Radiology, Massachusetts General Hospital and NeuroDiscovery Center, Harvard Medical School, Boston, Massachusetts (K.E.v.d.V., E.R.A., X.Z., C.L., S.P., O.M., M.H.W.C., J.S., X.O.B.); Center for Immunology and Inflammatory Diseases, Massachusetts General Hospital, Boston, Massachusetts (E.C., T.R.M., J.E.K., S.E.H.); Neuropathology Service, Massachusetts General Hospital and Department of Pathology, Harvard Medical School, Boston, Massachusetts (D.O., A.S-R.); Department of Neurology, Brigham and Women's Hospital, Harvard Medical School, Boston, Massachusetts (A.M.K.); Division of Molecular Carcinogenesis, The Netherlands Cancer Institute, Amsterdam, the Netherlands (K.E.v.d.V.).
authors.forename: Osama
authors.initials: O
authors.lastname: Mardini
authors.affiliation: Departments of Neurology and Radiology, Massachusetts General Hospital and NeuroDiscovery Center, Harvard Medical School, Boston, Massachusetts (K.E.v.d.V., E.R.A., X.Z., C.L., S.P., O.M., M.H.W.C., J.S., X.O.B.); Center for Immunology and Inflammatory Diseases, Massachusetts General Hospital, Boston, Massachusetts (E.C., T.R.M., J.E.K., S.E.H.); Neuropathology Service, Massachusetts General Hospital and Department of Pathology, Harvard Medical School, Boston, Massachusetts (D.O., A.S-R.); Department of Neurology, Brigham and Women's Hospital, Harvard Medical School, Boston, Massachusetts (A.M.K.); Division of Molecular Carcinogenesis, The Netherlands Cancer Institute, Amsterdam, the Netherlands (K.E.v.d.V.).
authors.forename: Matheus H W
authors.initials: MH
authors.lastname: Crommentuijn
authors.affiliation: Departments of Neurology and Radiology, Massachusetts General Hospital and NeuroDiscovery Center, Harvard Medical School, Boston, Massachusetts (K.E.v.d.V., E.R.A., X.Z., C.L., S.P., O.M., M.H.W.C., J.S., X.O.B.); Center for Immunology and Inflammatory Diseases, Massachusetts General Hospital, Boston, Massachusetts (E.C., T.R.M., J.E.K., S.E.H.); Neuropathology Service, Massachusetts General Hospital and Department of Pathology, Harvard Medical School, Boston, Massachusetts (D.O., A.S-R.); Department of Neurology, Brigham and Women's Hospital, Harvard Medical School, Boston, Massachusetts (A.M.K.); Division of Molecular Carcinogenesis, The Netherlands Cancer Institute, Amsterdam, the Netherlands (K.E.v.d.V.).
authors.forename: Johan
authors.initials: J
authors.lastname: Skog
authors.affiliation: Departments of Neurology and Radiology, Massachusetts General Hospital and NeuroDiscovery Center, Harvard Medical School, Boston, Massachusetts (K.E.v.d.V., E.R.A., X.Z., C.L., S.P., O.M., M.H.W.C., J.S., X.O.B.); Center for Immunology and Inflammatory Diseases, Massachusetts General Hospital, Boston, Massachusetts (E.C., T.R.M., J.E.K., S.E.H.); Neuropathology Service, Massachusetts General Hospital and Department of Pathology, Harvard Medical School, Boston, Massachusetts (D.O., A.S-R.); Department of Neurology, Brigham and Women's Hospital, Harvard Medical School, Boston, Massachusetts (A.M.K.); Division of Molecular Carcinogenesis, The Netherlands Cancer Institute, Amsterdam, the Netherlands (K.E.v.d.V.).
authors.forename: Anna M
authors.initials: AM
authors.lastname: Krichevsky
authors.affiliation: Departments of Neurology and Radiology, Massachusetts General Hospital and NeuroDiscovery Center, Harvard Medical School, Boston, Massachusetts (K.E.v.d.V., E.R.A., X.Z., C.L., S.P., O.M., M.H.W.C., J.S., X.O.B.); Center for Immunology and Inflammatory Diseases, Massachusetts General Hospital, Boston, Massachusetts (E.C., T.R.M., J.E.K., S.E.H.); Neuropathology Service, Massachusetts General Hospital and Department of Pathology, Harvard Medical School, Boston, Massachusetts (D.O., A.S-R.); Department of Neurology, Brigham and Women's Hospital, Harvard Medical School, Boston, Massachusetts (A.M.K.); Division of Molecular Carcinogenesis, The Netherlands Cancer Institute, Amsterdam, the Netherlands (K.E.v.d.V.).
authors.forename: Anat
authors.initials: A
authors.lastname: Stemmer-Rachamimov
authors.affiliation: Departments of Neurology and Radiology, Massachusetts General Hospital and NeuroDiscovery Center, Harvard Medical School, Boston, Massachusetts (K.E.v.d.V., E.R.A., X.Z., C.L., S.P., O.M., M.H.W.C., J.S., X.O.B.); Center for Immunology and Inflammatory Diseases, Massachusetts General Hospital, Boston, Massachusetts (E.C., T.R.M., J.E.K., S.E.H.); Neuropathology Service, Massachusetts General Hospital and Department of Pathology, Harvard Medical School, Boston, Massachusetts (D.O., A.S-R.); Department of Neurology, Brigham and Women's Hospital, Harvard Medical School, Boston, Massachusetts (A.M.K.); Division of Molecular Carcinogenesis, The Netherlands Cancer Institute, Amsterdam, the Netherlands (K.E.v.d.V.).
authors.forename: Thorsten R
authors.initials: TR
authors.lastname: Mempel
authors.affiliation: Departments of Neurology and Radiology, Massachusetts General Hospital and NeuroDiscovery Center, Harvard Medical School, Boston, Massachusetts (K.E.v.d.V., E.R.A., X.Z., C.L., S.P., O.M., M.H.W.C., J.S., X.O.B.); Center for Immunology and Inflammatory Diseases, Massachusetts General Hospital, Boston, Massachusetts (E.C., T.R.M., J.E.K., S.E.H.); Neuropathology Service, Massachusetts General Hospital and Department of Pathology, Harvard Medical School, Boston, Massachusetts (D.O., A.S-R.); Department of Neurology, Brigham and Women's Hospital, Harvard Medical School, Boston, Massachusetts (A.M.K.); Division of Molecular Carcinogenesis, The Netherlands Cancer Institute, Amsterdam, the Netherlands (K.E.v.d.V.).
authors.forename: Joseph
authors.initials: J
authors.lastname: El Khoury
authors.affiliation: Departments of Neurology and Radiology, Massachusetts General Hospital and NeuroDiscovery Center, Harvard Medical School, Boston, Massachusetts (K.E.v.d.V., E.R.A., X.Z., C.L., S.P., O.M., M.H.W.C., J.S., X.O.B.); Center for Immunology and Inflammatory Diseases, Massachusetts General Hospital, Boston, Massachusetts (E.C., T.R.M., J.E.K., S.E.H.); Neuropathology Service, Massachusetts General Hospital and Department of Pathology, Harvard Medical School, Boston, Massachusetts (D.O., A.S-R.); Department of Neurology, Brigham and Women's Hospital, Harvard Medical School, Boston, Massachusetts (A.M.K.); Division of Molecular Carcinogenesis, The Netherlands Cancer Institute, Amsterdam, the Netherlands (K.E.v.d.V.).
authors.forename: Suzanne E
authors.initials: SE
authors.lastname: Hickman
authors.affiliation: Departments of Neurology and Radiology, Massachusetts General Hospital and NeuroDiscovery Center, Harvard Medical School, Boston, Massachusetts (K.E.v.d.V., E.R.A., X.Z., C.L., S.P., O.M., M.H.W.C., J.S., X.O.B.); Center for Immunology and Inflammatory Diseases, Massachusetts General Hospital, Boston, Massachusetts (E.C., T.R.M., J.E.K., S.E.H.); Neuropathology Service, Massachusetts General Hospital and Department of Pathology, Harvard Medical School, Boston, Massachusetts (D.O., A.S-R.); Department of Neurology, Brigham and Women's Hospital, Harvard Medical School, Boston, Massachusetts (A.M.K.); Division of Molecular Carcinogenesis, The Netherlands Cancer Institute, Amsterdam, the Netherlands (K.E.v.d.V.).
authors.forename: Xandra O
authors.initials: XO
authors.lastname: Breakefield
mesh_terms: D000818:Animals; D001921:Brain / Q000378:metabolism; D001932:Brain Neoplasms / Q000378:metabolism*; D045744:Cell Line, Tumor; D000067128:Extracellular Vesicles / Q000378:metabolism*; D005909:Glioblastoma / Q000378:metabolism*; D006801:Humans; D008264:Macrophages / Q000378:metabolism*; D051379:Mice; D008810:Mice, Inbred C57BL; D035683:MicroRNAs / Q000378:metabolism*; D017628:Microglia / Q000378:metabolism*; D036641:Microscopy, Fluorescence, Multiphoton; D016271:Proto-Oncogene Proteins c-myc / Q000378:metabolism; D012333:RNA, Messenger / Q000378:metabolism; D014407:Tumor Cells, Cultured
chemical_list: C524940:MIRN21 microRNA, human; C531201:MIRN451 microRNA, human; C489427:MYC protein, human; D035683:MicroRNAs; D016271:Proto-Oncogene Proteins c-myc; D012333:RNA, Messenger
keywords: brain tumors; exosomes; extracellular vesicles; microRNA; microglia