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Original Research ARTICLE Provisionally accepted The full-text will be published soon. Notify me

Front. Oncol. | doi: 10.3389/fonc.2019.00651

Isolation and analysis of plasma-derived exosomes in patients with glioma

  • 1Mayo Clinic Graduate School of Biomedical Sciences, United States
  • 2Department of Immunology, Mayo Clinic, United States
  • 3Department of Neurosurgery, Mayo Clinic, United States
  • 4Department of Urology, Mayo Clinic, United States
  • 5Department of Neurology, Mayo Clinic, United States

Gliomas including glioblastoma (GBM) are the most common primary malignant brain tumors. Glioma extracellular vesicles (EVs) including exosomes have biological effects (e.g. immunosuppression) and contain tumor-specific cargo that could facilitate liquid biopsies. We aimed to develop a simple, reproducible technique to isolate plasma exosomes in glioma patients. Glioma patients’ and normal donors’ plasma exosomes underwent brief centrifugation to remove cells/debris followed by serial density gradient ultracentrifugation (DGU). EV size/concentration was determined by nanoparticle tracking. Protein cargo was screened by array, western blot, and ELISA. Nanoscale flow cytometry analysis quantified exosome and microvesicle populations pre- and post-DGU. One-step DGU efficiently isolates exosomes for nanoparticle tracking. Wild type isocitrate dehydrogenase glioma patients’ (i.e more aggressive tumors) plasma exosomes are smaller but higher concentration than normal donors. A second DGU efficiently concentrates exosomes for subsequent cargo analysis but results in vesicle aggregation that skews nanoparticle tracking. Cytokines and co-stimulatory molecules are readily detected but appeared globally reduced in GBM patients’ exosomes. Surprisingly, immunosuppressive programmed death-ligand 1 (PD-L1) is present in both patients’ and normal donors’ exosomes. Nanoscale flow cytometry confirms efficient exosome (<100nm) isolation post-DGU but also demonstrates increased microvesicles (>100nm) in GBM patients’ plasma pre-DGU. Serial DGU efficiently isolates plasma exosomes with distinct differences between GBM patients and normal donors, suggesting utility for non-invasive biomarker assessment. Initial results suggest global immunosuppression rather than increased circulating tumor-derived immunosuppressive exosomes, though further assessment is needed. Increased glioma patients’ plasma microvesicles suggest these may also be a key source for biomarkers.

Keywords: Glioblastoma, plasma-derived exosomes, Isolation, Immunosuppression, biomarkers

Received: 27 Mar 2019; Accepted: 03 Jul 2019.

Edited by:

Erik P. Sulman, Langone Medical Center, New York University, United States

Reviewed by:

Vinesh Puliyappadamba, UT Southwestern Medical Center, United States
Riccardo Soffietti, University of Turin, Italy  

Copyright: © 2019 Cumba Garcia, Peterson, Cepeda, Johnson and Parney. This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.

* Correspondence: Miss. Luz M. Cumba Garcia, Mayo Clinic Graduate School of Biomedical Sciences, Rochester, United States, cumba-garcia.luz@mayo.edu