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S.W. McPherson, N. Huess, T. Sam, J. Yang, D.S. Gregerson; A T Cell Receptor Transgenic Model for Analysis of CD4 T Cell Recognition of the Neo-Self Antigen b-Galactosidase in the Normal Eye . Invest. Ophthalmol. Vis. Sci. 2003;44(13):1053.
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© ARVO (1962-2015); The Authors (2016-present)
Purpose: To determine if naïve, MHC class II restricted T cells are capable of antigen recognition in the immunologically quiescent eye. Methods: A T cell receptor (TCR) transgenic (Tg) mouse was made using the TCR-alpha and TCR-beta chains from 3E9 cells. Previously, we demonstrated that CD4+, CD44hi 3E9 cells respond in vitro to both ß-galactosidase (ß-gal) and a MHC class II restricted peptide epitope of ß-gal (YVVDEANIETHGMV) and, when activated by antigen, were capable of mediating autoimmune disease in mice expressing ß-gal in the retina (Gregerson, et al. J. Immunol. 163:1073). The number of 3E9-Tg mouse T cells expressing bona fide 3E9 alpha and beta TCR chains was analyzed by flow cytometry (FACS). In vitro T cell proliferation assays were done to test the response of 3E9-Tg T cells to ß-gal and ß-gal peptide with cytokine production measured by ELISA on the supernatants. In vivo T cell response was assayed by transfer of CFSE-labeled 3E9-Tg cells into ß-gal Tg mice expressing ß-gal in the retina (hi-arr-ß-gal), eye and brain (GFAP-ß-gal), and systemically (ROSA26) followed by analysis of recovered T cells for CFSE content and cell surface activation markers by FACS. Results: FACS analysis using an antibody to the 3E9 TCR-beta chain (anti-Vß10.1) combined with in vivo proliferation data indicate at least 80% of the T cells in 3E9-Tg mice express the 3E9 TCR. Unprimed, naïve 3E9-Tg T cells proliferate in response to ß-gal and ß-gal peptide in a dose dependent manner and produce IL-2 and INF-γ but not IL-4 or IL-10. In vivo, 3E9-Tg T cells responded similarly to endogenous ß-gal (ROSA26) and exogenous ß-gal (B10.A + ß-gal peptide) as evidenced by dilution of CFSE content and an increase in T cells with the activation phenotype (CD45RBlo, CD62Llo, CD44hi, CD69hi) compared to controls (B10.A). However, 3E9-Tg T cells recovered from hi-arr-ß-gal and GFAP-ß-gal mice show no difference in CFSE content and activation markers compared to cells recovered from B10.A controls. Analysis of 3E9-Tg T cells recovered from hi-arr-ß-gal submandibular and cervical lymph nodes showed no difference compared to same or distal nodes from B10.A controls. Conclusions: Naïve, MHC class II restricted T cells give no evidence of recognition of antigen originating in the normal retina.
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