Facilitation of beta selection and modification of positive selection in the thymus of PD-1-deficient mice

Abstract

PD-1 is an immunoglobulin superfamily member bearing an immunoreceptor tyrosine-based inhibitory motif, and disruption of the PD-1 gene results in the development of lupus-like autoimmune diseases. In this study, we examined effects of the PD-1 deficiency on the thymocyte differentiation at the clonal level using T cell receptor (TCR)-beta (Vbeta8) and TCR-alpha/beta (H-Y and 2C) transgenic mice. In these TCR transgenic lines, PD-1 expression in the thymus was variably augmented, but as in the normal mice, confined largely to the CD4(-)CD8(-) thymocytes. The transgenic mice crossed with PD-1(-/)- mice in the neutral genetic backgrounds exhibited selective increase in the CD4(+)CD8(+) (DP) population with little effect on other thymocytes subsets. Similarly, the absence of PD-1 facilitated expansion of DP thymocytes in recombination activating gene (RAG)-2(-/)- mice by anti-CD3epsilon antibody injection. On the other hand, H-Y or 2C transgenic PD-1(-/)- mice with the positively selecting background showed significantly reduced efficiency for the generation of CD8(+) single positive cells bearing the transgenic TCR-alpha/beta in spite of the increased DP population. These results collectively indicate that PD-1 negatively regulates the beta selection and modulates the positive selection, and suggest that PD-1 deficiency may lead to the significant alteration of mature T cell repertoire.

Figure 1

Augmented expression of PD-1 in CD4⁻CD8⁻ compartment in TCR transgenic lines. (a) Thymocytes from wild-type (WT), Vβ8, H-Y (H-2^d), female H-Y (H-2^b), and 2C (H-2^b) mice were examined for PD-1 expression. Top, CD4/CD8 contour plots. Bottom, histograms for PD-1 expression in total, CD4⁺CD8⁺, and CD4⁻ CD8⁻ thymocytes. (b) Thymocytes from H-Y (H-2^b) and 2C (H-2^b) mice were examined for the expression of PD-1. The contours of CD4/CD8 profiles gated in PD-1⁺ (left) or PD-1⁻ (right) fractions are shown. (c) CD69 and PD-1 expressions are shown in total thymocytes from wild-type, female H-Y (H-2^b/d), and 2C (H-2^b) mice. The numbers indicate percentages in gated regions. Tg, transgenic.

Figure 2

Effects of PD-1 deficiency on T cell development in the TCR-α/β transgenic mice in positively selecting background. (a) Thymocytes from female H-Y (H-2^b) mice with or without the PD-1 mutation were analyzed for expression of CD4, CD8a, CD24, and transgenic TCR-α (T3.70). (b) Thymocytes from 2C (H-2^b) mice with or without the PD-1 mutation were analyzed for expression of CD4, CD8a, transgenic TCR-α/β (1B2), CD24, CD69, and CD8β for total thymocytes (top), CD4⁻CD8⁺ (middle), and CD4⁺CD8⁺ and CD4⁺CD8⁻ (bottom) subsets. Numbers indicate percentages of subsets in gated regions.