March 2012
Volume 53, Issue 14
Free
ARVO Annual Meeting Abstract  |   March 2012
Inhibitory Effect Of Regulatory T Cells Expanded In Vitro By Human Retinal Pigment Epithelial (RPE) Cells
Author Affiliations & Notes
  • Ayano Imai
    Department of Ophthalmology & Visual Science, Tokyo Medical and Dental University, Tokyo, Japan
  • Sunao Sugita
    Department of Ophthalmology & Visual Science, Tokyo Medical and Dental University, Tokyo, Japan
  • Yuko Kawazoe
    Department of Ophthalmology & Visual Science, Tokyo Medical and Dental University, Tokyo, Japan
  • Shintaro Horie
    Department of Ophthalmology & Visual Science, Tokyo Medical and Dental University, Tokyo, Japan
  • Yukiko Yamada
    Department of Ophthalmology & Visual Science, Tokyo Medical and Dental University, Tokyo, Japan
  • Manabu Mochizuki
    Department of Ophthalmology & Visual Science, Tokyo Medical and Dental University, Tokyo, Japan
  • Footnotes
    Commercial Relationships  Ayano Imai, None; Sunao Sugita, None; Yuko Kawazoe, None; Shintaro Horie, None; Yukiko Yamada, None; Manabu Mochizuki, None
  • Footnotes
    Support  Scientific Research (C) 20592073 and Grants-in-Aid for Young Scientists (B) 21791671 & 21791672 of the Ministry of Education, Culture, Sports, Science and Technology, Japan
Investigative Ophthalmology & Visual Science March 2012, Vol.53, 4269. doi:
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      Ayano Imai, Sunao Sugita, Yuko Kawazoe, Shintaro Horie, Yukiko Yamada, Manabu Mochizuki; Inhibitory Effect Of Regulatory T Cells Expanded In Vitro By Human Retinal Pigment Epithelial (RPE) Cells. Invest. Ophthalmol. Vis. Sci. 2012;53(14):4269.

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      © ARVO (1962-2015); The Authors (2016-present)

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Abstract

Purpose: : Retinal pigment epithelium (RPE) cells are able to convert CD4+ T cells into T regulatory cells (Tregs) that greatly suppress bystander target T cells. In this study we designed to determine whether human RPE-induced Tregs can be expanded under Treg induction conditions and then, as RPE-induced Tregs, suppress activation of effector T cells.

Methods: : Peripheral blood mononuclear cells from healthy donors were co-cultured with supernatants from TGFβ2-pretreated human RPE cells on anti-human CD3-coated plate. Cells were then separated with a CD4+CD25+ regulatory T-cell isolation kit, and cultured with the supernatants from RPE, anti-CD3, anti-CD28, high-dose recombinant IL-2, and TGFβ2. By flow cytometry sorting, CD25+CD45RA- Tregs were separated. Expression of CD25high, Foxp3, CD152 (CTLA-4), and TNFRSF 18 (GITR) on RPE-induced Tregs was analyzed by flow cytometry. Production of cytokines (TGFβ1, IL-10, IFNγ, IL-17, and TNFα) by RPE-induced Tregs was measured by ELISA. In addition, proliferation of target T cells (Th1/Th17 cell clines) was assessed by [3H]-thymidine incorporation or CFSE incorporation.

Results: : RPE cells in the presence of TGFβ produce anti-inflammatory effects, and are able to induce Tregs in vitro. Human RPE-induced Tregs expressed higher levels of the Treg cell markers CD25high, Foxp3, CD152, and TNFRSF 18. In addition, human RPE-induced Tregs included significant numbers of CD4+CD25highFoxp3highCD45RA- active effector Tregs that significantly suppressed activation of Th1/Th17 cell lines, indicating that they have immunosuppressive properties. Furthermore, we are able to remove CD4+CD25lowFoxp3lowCD45RA- non-suppressing cytokine-secreting T cells from the in vitro-manipulated Treg population.

Conclusions: : Present data provide hope that Tregs might become useful as individualized therapeutic agents for various ocular autoimmune diseases.

Keywords: retinal pigment epithelium • immune tolerance/privilege • inflammation 
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