Age-related changes in the occurrence and characteristics of thymic CD4(+) CD25(+) T cells in mice

Abstract

Natural regulatory CD4(+) CD25(+) T cells play an important role in preventing autoimmunity by maintaining self-tolerance. They express CD25 constitutively and are produced in the thymus as a functionally mature T-cell population. Changes in the potential of these cells to regulate the activity of conventional effector lymphocytes may contribute to an increased susceptibility to infection, cancer and age-associated autoimmune diseases. In this study we demonstrated that the thymi of aged mice are populated by a higher percentage of CD4(+) CD25(+) thymocytes than in young animals. The expression of several surface markers (CD69, CD5, CD28, CTLA-4, CD122, FOXP3), usually used to characterize the phenotype of CD4(+) CD25(+) T regulatory cells, was compared between young and aged mice. We also examined the ability of sorted thymus-deriving regulatory T cells of young and aged BALB/c mice to inhibit the proliferation of lymph node lymphocytes activated in vitro. Natural regulatory T cells isolated from the thymi of young mice suppress the proliferation of responder lymph node cells. We demonstrated that thymus-deriving CD4(+) CD25(+) T cells of old mice maintain their potential to suppress the proliferation of activated responder lymphocytes of young mice. However, their potential to inhibit the proliferation of old responder T cells is abrogated. Differences in the occurrence and activity of CD4(+) CD25(+) thymocytes between young and old animals are discussed in relation to the expression of these surface markers.

Figure 1

The distribution of thymocyte populations in young and old mice. DN, double-negative thymocytes, CD4^– CD8^–; DP, double-positive thymocytes, CD4⁺ CD8⁺ SP CD4, single-positive thymocytes, CD4⁺ CD8^–; SP CD8, single-positive thymocytes, CD8⁺ CD4^–. Statistically significant difference in the percentage of positive cells between old and young mice.

Figure 2

CD4⁺ CD25⁺ T cells in the thymi of young and old mice. R2, SP CD4⁺ thymocytes (CD4⁺ CD8^–). Statistically significant difference in the percentage of CD4⁺ CD25⁺ in the thymus between old and young mice.

Figure 3

CD69 expression on CD4⁺ CD25⁺ thymocytes in young and old mice. Dotted line, isotype control; solid line, positively stained cells; ^{^}Statistically significant difference in the percentage of positively stained cells or fluorescence intensity (FI) between CD4⁺ CD25⁺ and CD4⁺ CD25^– thymocytes in the group of young and old mice. *Statistically significant difference in the percentage of positively stained cells or fluorescence intensity (FI) between old and young mice in the CD4⁺ CD25⁺ subset.

Figure 4

CD5 expression on CD4⁺ CD25⁺ thymocytes in young and old mice. See legend to Fig. 3 for key.

Figure 5

CTLA-4 expression on CD4⁺ CD25⁺ thymocytes in young and old mice. See legend to Fig. 3 for key.

Figure 6

CD28 expression on CD4⁺ CD25⁺ thymocytes in young and old mice. See legend to Fig. 3 for key.

Figure 7

CD122 expression on CD4⁺ CD25⁺ thymocytes in young and old mice. See legend to Fig. 3 for key.

Figure 8

FOXP3 expression in CD4⁺ CD25⁺ thymocytes in young and old mice. See legend to Fig. 3 for key.

Figure 9

The inhibition of proliferation of activated lymph node lymphocytes by sorted CD4⁺ CD25⁺ thymocytes (a) Coculture of lymph node cells of young mice and CD4⁺ CD25⁺ thymocytes of young mice. (b) Coculture of lymph node cells of old mice and CD4⁺ CD25⁺ thymocytes of old mice. (c) Coculture of lymph node cells of young mice and CD4⁺ CD25⁺ thymocytes of old mice. Solid line, CFSE fluorescence intensity of activated lymph node cells cultured without CD4⁺ CD25⁺ thymocyte; dotted line, CFSE fluorescence intensity of activated lymph node cells cocultured with CD4⁺ CD25⁺ thymocytes.