March 2012
Volume 53, Issue 14
Free
ARVO Annual Meeting Abstract  |   March 2012
Thrombospondin Receptor CD47 On T Cells And Not On The Surface Of Antigen Presenting Cells Is Necessary For Treg Induction Associated With Ocular Immune Privilege
Author Affiliations & Notes
  • Fayaz Mir
    Harvard Medical School, Schepens Eye Research Inst, Boston, Massachusetts
  • Bruce Turpie
    Harvard Medical School, Schepens Eye Research Inst, Boston, Massachusetts
  • Sharmila Masli
    Harvard Medical School, Schepens Eye Research Inst, Boston, Massachusetts
  • Footnotes
    Commercial Relationships  Fayaz Mir, None; Bruce Turpie, None; Sharmila Masli, None
  • Footnotes
    Support  NEI EY015472-07
Investigative Ophthalmology & Visual Science March 2012, Vol.53, 6310. doi:
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      Fayaz Mir, Bruce Turpie, Sharmila Masli; Thrombospondin Receptor CD47 On T Cells And Not On The Surface Of Antigen Presenting Cells Is Necessary For Treg Induction Associated With Ocular Immune Privilege. Invest. Ophthalmol. Vis. Sci. 2012;53(14):6310.

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

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Abstract

Purpose: : Ocular antigen presenting cells (APCs), exposed to TGFβ in the eye, induce regulatory immune response associated with ocular immune privilege. Thrombospondin-1 expression by APCs is critical to this process. We now investigate the significance of autocrine vs. paracrine effects of TSP-1 via its receptor CD47 expressed both on APCs and on responding effectors during Treg induction

Methods: : Regulatory phenotype of CD4+CD25- OT-II T cells activated in vitro by untreated or TGFβ-treated (5 ng/ml) APCs from WT or CD47-/- mice was examined. Intracellular Foxp3 expression was determined by flow cytometry. Cytokines (IFN-γ and TGFβ) secreted in culture supernatant upon re-stimulation with anti-CD3 were measured by ELISA and the ability of Treg to suppress proliferation was determined by CFSE dilution assay. Expression of Foxp3 was determined in CD4+CD25- T cells from C57BL/6 (WT) and CD47-/- mice activated with anti-CD3 in the presence of CD47-binding TSP-derived peptide (4N1K) or control peptide. In some experiments blocking anti-CD47 antibody was used to prevent CD47 ligation by the 4N1K.

Results: : Upon co-culture with CD4+CD25- OT-II T cells these CD47-/- APCs induced similar numbers of Foxp3+ve cells as compared to the WT controls. Effectors activated by TGFβ-treated WT as well as CD47-/- APCs produced significantly reduced levels of IFN-γ and increased levels of TGFβ as compared to their corresponding untreated APC controls. The ability to suppress proliferation of T cells as determined by CFSE-dilution assay was comparable in OT-II T cells activated by TGFβ-treated either WT or CD47-/- APCs suggesting no contribution of CD47 receptor on APCs in TSP-1 dependent induction of Tregs. In the presence of CD47-binding peptide (4N1K) derived from TSP-1, activation of WT CD4+CD25- T cells significantly increased Foxp3+ve cells as compared to those activated in the presence of control peptide (4NGG). However, 4N1K mediated increase in Foxp3 expression was abrogated in the presence of anti-CD47 antibody or in CD4+CD25- T cells derived from CD47-/- mice.

Conclusions: : Thrombospondin receptor CD47 on the APCs does not play any direct role in inducing the Tregs associated with ocular immune privilege whereas this receptor on the T cells mediates the TSP-1 induced Treg generation.

Keywords: ACAID • immunomodulation/immunoregulation • flow cytometry 
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