Transient depletion of CD4+ CD25+ regulatory T cells results in multiple autoimmune diseases in wild-type and B-cell-deficient NOD mice

Abstract

Approximately 80% of female wild-type non-obese diabetic (WT NOD) mice spontaneously develop diabetes, whereas B-cell-deficient (B(-/-)) NOD mice are resistant to diabetes. B(-/-) mice are also resistant to other spontaneous and experimentally induced autoimmune diseases, including arthritis, systemic lupus erythematosus, Sjögren syndrome and thyroiditis. Under normal conditions, activation of self-reactive T cells in the periphery is limited by CD4(+) CD25(+) natural regulatory T (Treg) cells. B(-/-) NOD.H-2h4 mice, normally resistant to spontaneous autoimmune thyroiditis (SAT), develop SAT when Treg cells are depleted, suggesting that Treg cells are preferentially activated when autoantigen is initially presented by non-B-cell antigen-presenting cells. To test the hypothesis that increased Treg cell activity in B(-/-) mice contributes to their resistance to other autoimmune diseases, WT and B(-/-) NOD mice were given anti-CD25 to transiently deplete CD4(+) CD25(+) Treg cells. The WT and B(-/-) NOD mice given anti-CD25 developed diabetes much earlier than WT mice given rat IgG, whereas rat IgG-treated B(-/-) mice did not develop diabetes. Treg-cell-depleted mice had increased lymphocyte infiltration of the pancreas, salivary glands and thyroid compared with controls given rat IgG. These results are consistent with the hypothesis that resistance of B-cell-deficient NOD mice to several autoimmune diseases is due to the activity of Treg cells.

Figure 1

Depletion of CD25⁺ regulatory T (Treg) cells results in early onset and increased incidence of diabetes. Blood glucose levels were determined in unmanipulated wild-type (WT) or B-cell-deficient (B^−/−) non-obese diabetic (NOD) mice starting at week 10 of age. Graph represents percentage of diabetic mice over time; n = 15 (a). B^−/− (b) or WT (c) NOD mice were given two injections of 0·5 mg anti-CD25 antibody 1 week apart to deplete CD4⁺ CD25⁺ Treg cells, or rat IgG as control. Blood glucose levels were determined weekly starting at 8 weeks of age and continued until 16 weeks of age. Experiments were terminated at 16 weeks because of health issues in diabetic animals; 14–15 mice per group.

Figure 2

Depletion of CD25⁺ regulatory T (Treg) cells results in increased lymphocytic infiltration of multiple tissues. B-cell deficient (B^−/−) or wild-type (WT) non-obese diabetic (NOD) mice were given two injections of 0·5 mg anti-CD25 antibody 1 week apart to deplete CD4⁺ CD25⁺ Treg cells. Control mice were given rat IgG. At 16 weeks of age the pancreas, salivary gland and thyroid were harvested and stained with haematoxylin and eosin. Three representative mice are shown for each group. Arrows indicate lymphocytic infiltration. Pancreas sections show intra-insulitis in anti-CD25 groups, and peri-insulitis in control groups. Magnification: 100 ×.

Figure 3

Pancreatic infiltration after regulatory T (Treg) cell depletion in wild-type (WT) or B-cell-deficient mice. At 16 weeks of age, pancreas was harvested from mice treated with two, weekly injections of anti-CD25 or rat IgG starting 10–11 days after birth. The islets were scored as no insulitis, peri-insulitis, mild intra-insulitis or severe intra-insulitis as described in the Materials and methods: (a) depicts the percentage of total islets from each group that were given each score. (b) represents the number of islets of each score and the overall total number of islets per group.

Figure 4

Depletion of CD25⁺ regulatory T (Treg) cells results in significant increase in salivary gland infiltrate and thyroiditis in B-cell-deficient (B^−/−) mice. B^−/− or wild-type (WT) non-obese diabetic (NOD) mice were given two injections of 0·5 mg anti-CD25 antibody 1 week apart to deplete CD4⁺ CD25⁺ Treg cells. At 16 weeks of age the salivary gland (a) and thyroid (b) were harvested and stained with haematoxylin and eosin. (a) The number of foci of lymphocytic infiltrate in the salivary gland were counted and (b) thyroid infiltration and follicular destruction were scored as described in Materials and methods. Depletion of regulatory T cells resulted in significant increases in salivary and thyroid infiltrates in both B^−/− and WT mice. *P < 0·05, ***P < 0·001.