Modulation of immune responses by extracellular vesicles from retinal pigment epithelium

Jared E. Knickelbein, Baoying Liu, Anush Arakelyan, Sonia Zicari, Susan Hannes, Ping Chen, Zhiyu Li, Jean-Charles B. Grivel, Benjamin Chaigne-Delalande, H. Nida Sen, Leonid Margolis, Robert B. Nussenblatt

Research output: Contribution to journalArticle

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Abstract

Purpose. 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. Methods. Extracellular vesicles were isolated from ARPE-19 cultures stimulated or not with the inflammatory cytokines IL-1β, LFN-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. Results. 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 immunoregula-tory 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. Conclusions. 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 immuno-regulatory 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.

Original languageEnglish
Article number22
Pages (from-to)4101-4107
Number of pages7
JournalInvestigative Ophthalmology and Visual Science
Volume57
Issue number10
DOIs
Publication statusPublished - 1 Aug 2016

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Retinal Pigment Epithelium
Monocytes
Cytokines
Exosomes
T-Lymphocytes
Extracellular Vesicles
Phenotype
Up-Regulation
Cell Proliferation
Interleukin-1

Keywords

  • Extracellular vesicles
  • Immune privilege
  • Monocytes
  • Retinal pigment epithelium
  • T cells

ASJC Scopus subject areas

  • Ophthalmology
  • Sensory Systems
  • Cellular and Molecular Neuroscience

Cite this

Knickelbein, J. E., Liu, B., Arakelyan, A., Zicari, S., Hannes, S., Chen, P., ... Nussenblatt, R. B. (2016). Modulation of immune responses by extracellular vesicles from retinal pigment epithelium. Investigative Ophthalmology and Visual Science, 57(10), 4101-4107. [22]. https://doi.org/10.1167/iovs.15-18353

Modulation of immune responses by extracellular vesicles from retinal pigment epithelium. / Knickelbein, Jared E.; Liu, Baoying; Arakelyan, Anush; Zicari, Sonia; Hannes, Susan; Chen, Ping; Li, Zhiyu; Grivel, Jean-Charles B.; Chaigne-Delalande, Benjamin; Sen, H. Nida; Margolis, Leonid; Nussenblatt, Robert B.

In: Investigative Ophthalmology and Visual Science, Vol. 57, No. 10, 22, 01.08.2016, p. 4101-4107.

Research output: Contribution to journalArticle

Knickelbein, JE, Liu, B, Arakelyan, A, Zicari, S, Hannes, S, Chen, P, Li, Z, Grivel, J-CB, Chaigne-Delalande, B, Sen, HN, Margolis, L & Nussenblatt, RB 2016, 'Modulation of immune responses by extracellular vesicles from retinal pigment epithelium', Investigative Ophthalmology and Visual Science, vol. 57, no. 10, 22, pp. 4101-4107. https://doi.org/10.1167/iovs.15-18353
Knickelbein, Jared E. ; Liu, Baoying ; Arakelyan, Anush ; Zicari, Sonia ; Hannes, Susan ; Chen, Ping ; Li, Zhiyu ; Grivel, Jean-Charles B. ; Chaigne-Delalande, Benjamin ; Sen, H. Nida ; Margolis, Leonid ; Nussenblatt, Robert B. / Modulation of immune responses by extracellular vesicles from retinal pigment epithelium. In: Investigative Ophthalmology and Visual Science. 2016 ; Vol. 57, No. 10. pp. 4101-4107.
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abstract = "Purpose. 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. Methods. Extracellular vesicles were isolated from ARPE-19 cultures stimulated or not with the inflammatory cytokines IL-1β, LFN-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. Results. 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 immunoregula-tory 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. Conclusions. 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 immuno-regulatory 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.",
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AU - Hannes, Susan

AU - Chen, Ping

AU - Li, Zhiyu

AU - Grivel, Jean-Charles B.

AU - Chaigne-Delalande, Benjamin

AU - Sen, H. Nida

AU - Margolis, Leonid

AU - Nussenblatt, Robert B.

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N2 - Purpose. 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. Methods. Extracellular vesicles were isolated from ARPE-19 cultures stimulated or not with the inflammatory cytokines IL-1β, LFN-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. Results. 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 immunoregula-tory 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. Conclusions. 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 immuno-regulatory 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.

AB - Purpose. 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. Methods. Extracellular vesicles were isolated from ARPE-19 cultures stimulated or not with the inflammatory cytokines IL-1β, LFN-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. Results. 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 immunoregula-tory 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. Conclusions. 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 immuno-regulatory 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.

KW - Extracellular vesicles

KW - Immune privilege

KW - Monocytes

KW - Retinal pigment epithelium

KW - T cells

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