title: Modulation of Immune Responses by Extracellular Vesicles From Retinal Pigment Epithelium.
abstract: Extracellular vesicles (EV), such as exosomes, are important mediators of intercellular communication and have been implicated in modulation of the immune system. We investigated if EV released from retinal pigment epithelium (RPE) modulate immune responses in vitro.
  
  
  
  Extracellular vesicles were isolated from ARPE-19 cultures stimulated or not with the inflammatory cytokines IL-1β, IFN-γ, and TNF-α. Isolated EV were characterized by nanoparticle flow and Western blot analyses. Retinal pigment epithelium-derived EV were cultured with human peripheral blood mononuclear cells, which were assayed for T-cell proliferation by 3H-thymidine incorporation. Retinal pigment epithelium-derived EV were also independently cultured with enriched lymphocytes or monocytes. Cell phenotype and cell death were evaluated by flow cytometric analysis. Cytokine levels were assayed in culture supernatants by multiplex bead analysis.
  
  
  
  The concentration of ARPE-derived EV from cytokine-stimulated cultures was slightly higher than from nonstimulated cultures. The size of EV was approximately 100 nm in both groups. Extracellular vesicles from both nonstimulated and cytokine-stimulated ARPE-19 significantly inhibited T-cell proliferation without affecting T-cell viability. Culture of EV from nonstimulated ARPE-19 with undifferentiated human monocytes induced an immunoregulatory phenotype with a significantly higher percentage of CD14++CD16+ monocytes and upregulation of TGF-β1. Culture of EV from cytokine-stimulated ARPE-19 cells with human monocytes induced upregulation of several proinflammatory cytokines and monocyte death.
  
  
  
  Retinal pigment epithelium cells constitutively secrete EV in the size range of exosomes, with increased release from RPE cells stimulated with inflammatory cytokines. Extracellular vesicles from both nonstimulated and cytokine-stimulated RPE inhibited T-cell stimulation. Extracellular vesicles from nonstimulated ARPE-19 cells promoted an immunoregulatory CD14++CD16+ phenotype in human monocytes, while EV from cytokine-stimulated ARPE-19 cells caused human monocyte death. These findings suggest that RPE cells use EV to induce a downregulatory immune environment under homeostatic conditions. In an inflammatory milieu, RPE-derived EV may mitigate a potentially harmful inflammatory response through killing of monocytes.
authors.affiliation: Laboratory of Immunology, National Eye Institute, National Institutes of Health, Bethesda, Maryland, United States.
authors.forename: Jared E
authors.initials: JE
authors.lastname: Knickelbein
authors.affiliation: Laboratory of Immunology, National Eye Institute, National Institutes of Health, Bethesda, Maryland, United States.
authors.forename: Baoying
authors.initials: B
authors.lastname: Liu
authors.affiliation: Section of Intercellular Interactions, Eunice Kennedy-Shriver National Institute of Child Health and Human Development, Bethesda, Maryland, United States.
authors.forename: Anush
authors.initials: A
authors.lastname: Arakelyan
authors.affiliation: Section of Intercellular Interactions, Eunice Kennedy-Shriver National Institute of Child Health and Human Development, Bethesda, Maryland, United States.
authors.forename: Sonia
authors.initials: S
authors.lastname: Zicari
authors.affiliation: Laboratory of Immunology, National Eye Institute, National Institutes of Health, Bethesda, Maryland, United States.
authors.forename: Susan
authors.initials: S
authors.lastname: Hannes
authors.affiliation: Laboratory of Immunology, National Eye Institute, National Institutes of Health, Bethesda, Maryland, United States.
authors.forename: Ping
authors.initials: P
authors.lastname: Chen
authors.affiliation: Laboratory of Immunology, National Eye Institute, National Institutes of Health, Bethesda, Maryland, United States.
authors.forename: Zhiyu
authors.initials: Z
authors.lastname: Li
authors.affiliation: Laboratory of Immunology, National Eye Institute, National Institutes of Health, Bethesda, Maryland, United States 2Section of Intercellular Interactions, Eunice Kennedy-Shriver National Institute of Child Health and Human Development, Bethesda, Maryland.
authors.forename: Jean-Charles
authors.initials: JC
authors.lastname: Grivel
authors.affiliation: Laboratory of Immunology, National Eye Institute, National Institutes of Health, Bethesda, Maryland, United States.
authors.forename: Benjamin
authors.initials: B
authors.lastname: Chaigne-Delalande
authors.affiliation: Laboratory of Immunology, National Eye Institute, National Institutes of Health, Bethesda, Maryland, United States.
authors.forename: H Nida
authors.initials: HN
authors.lastname: Sen
authors.affiliation: Section of Intercellular Interactions, Eunice Kennedy-Shriver National Institute of Child Health and Human Development, Bethesda, Maryland, United States.
authors.forename: Leonid
authors.initials: L
authors.lastname: Margolis
authors.affiliation: Laboratory of Immunology, National Eye Institute, National Institutes of Health, Bethesda, Maryland, United States.
authors.forename: Robert B
authors.initials: RB
authors.lastname: Nussenblatt
mesh_terms: D015153:Blotting, Western; D016923:Cell Death; D002478:Cells, Cultured; D016207:Cytokines / Q000378:metabolism*; D000067128:Extracellular Vesicles / Q000276:immunology* / Q000378:metabolism; D005434:Flow Cytometry; D006801:Humans; D007111:Immunity, Cellular; D009000:Monocytes / Q000378:metabolism / Q000473:pathology; D055213:Retinal Pigment Epithelium / Q000276:immunology / Q000378:metabolism* / Q000473:pathology; D015854:Up-Regulation; D014605:Uveitis / Q000276:immunology / Q000378:metabolism / Q000473:pathology
chemical_list: D016207:Cytokines