Lymphocyte responses and cytokines

Elimination of CD25+ T cells, which constitute 5-10% of peripheral CD4+ T cells in normal naive mice, leads to spontaneous development of various autoimmune diseases. These immunoregulatory CD25+CD4+ T cells are naturally unresponsive (anergic) in vitro to TCR stimulation, and, upon stimulation, suppress proliferation of CD25-CD4+ T cells and CD8+ T cells. The antigen concentration required for stimulating CD25+CD4+ T cells to exert suppression is much lower than that required for stimulating CD25-CD4+ T cells to proliferate. The suppression, which results in reduced IL-2 production by CD25-CD4+ T cells, is dependent on cellular interactions on antigen-presenting cells (and not mediated by far-reaching or long-lasting humoral factors or apoptosis-inducing signals) and antigen non-specific in its effector phase. Addition of high doses of IL-2 or anti-CD28 antibody to the in vitro T cell stimulation culture not only breaks the anergic state of CD25+CD4+ T cells, but also abrogates their suppressive activity simultaneously. Importantly, the anergic/suppressive state of CD25+CD4+ T cells appeared to be their basal default condition, since removal of IL-2 or anti-CD28 antibody from the culture milieu allows them to revert to the original anergic/suppressive state. Furthermore, transfer of such anergy/suppression-broken T cells from normal mice produces various autoimmune diseases in syngeneic athymic nude mice. These results taken together indicate that one aspect of immunologic self-tolerance is maintained by this unique CD25+CD4+ naturally anergic/suppressive T cell population and its functional abnormality directly leads to the development of autoimmune disease.

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Immunologic self-tolerance maintained by CD25+CD4+ naturally anergic and suppressive T cells: induction of autoimmune disease by breaking their anergic/suppressive state.

This study shows that the normal thymus produces immunoregulatory CD25+4+8- thymocytes capable of controlling self-reactive T cells. Transfer of thymocyte suspensions depleted of CD25+4+8- thymocytes, which constitute approximately 5% of steroid-resistant mature CD4+8- thymocytes in normal naive mice, produces various autoimmune diseases in syngeneic athymic nude mice. These CD25+4+8- thymocytes are nonproliferative (anergic) to TCR stimulation in vitro, but potently suppress the proliferation of other CD4+8- or CD4-8+ thymocytes; breakage of their anergic state in vitro by high doses of IL-2 or anti-CD28 Ab simultaneously abrogates their suppressive activity; and transfer of such suppression-abrogated thymocyte suspensions produces autoimmune disease in nude mice. These immunoregulatory CD25+4+8- thymocytes/T cells are functionally distinct from activated CD25+4+ T cells derived from CD25-4+ thymocytes/T cells in that the latter scarcely exhibits suppressive activity in vitro, although both CD25+4+ populations express a similar profile of cell surface markers. Furthermore, the CD25+4+8- thymocytes appear to acquire their anergic and suppressive property through the thymic selection process, since TCR transgenic mice develop similar anergic/suppressive CD25+4+8- thymocytes and CD25+4+ T cells that predominantly express TCRs utilizing endogenous alpha-chains, but RAG-2-deficient TCR transgenic mice do not. These results taken together indicate that anergic/suppressive CD25+4+8- thymocytes and peripheral T cells in normal naive mice may constitute a common T cell lineage functionally and developmentally distinct from other T cells, and that production of this unique immunoregulatory T cell population can be another key function of the thymus in maintaining immunologic self-tolerance.

Thymus and Autoimmunity: Production of CD25+CD4+ Naturally Anergic and Suppressive T Cells as a Key Function of the Thymus in Maintaining Immunologic Self-Tolerance

Insulin-Dependent Diabetes Mellitus

Th1 and Th2 CD4+ T cells in the pathogenesis of organ-specific autoimmune diseases

Beginning at the time of insulitis (7 wk of age), CD4+ and CD8+ mature thymocytes from nonobese diabetic (NOD) mice exhibit a proliferative unresponsiveness in vitro after T cell receptor (TCR) crosslinking. This unresponsiveness does not result from either insulitis or thymic involution and is long lasting, i.e., persists until diabetes onset (24 wk of age). We previously proposed that it represents a form of thymic T cell anergy that predisposes to diabetes onset. This hypothesis was tested in the present study by further investigating the mechanism responsible for NOD thymic T cell proliferative unresponsiveness and determining whether reversal of this unresponsiveness protects NOD mice from diabetes. Interleukin 4 (IL-4) secretion by thymocytes from > 7-wk-old NOD mice was virtually undetectable after treatment with either anti-TCR alpha/beta, anti-CD3, or Concanavalin A (Con A) compared with those by thymocytes from age- and sex-matched control BALB/c mice stimulated under identical conditions. NOD thymocytes stimulated by anti-TCR alpha/beta or anti-CD3 secreted less IL-2 than did similarly activated BALB/c thymocytes. However, since equivalent levels of IL-3 were secreted by Con A-activated NOD and BALB/c thymocytes, the unresponsiveness of NOD thymic T cells does not appear to be dependent on reduced IL-2 secretion. The surface density and dissociation constant of the high affinity IL-2 receptor of Con A-activated thymocytes from both strains are also similar. The patterns of unresponsiveness and lymphokine secretion seen in anti-TCR/CD3-activated NOD thymic T cells were also observed in activated NOD peripheral spleen T cells. Exogenous recombinant (r)IL-2 only partially reverses NOD thymocyte proliferative unresponsiveness to anti-CD3, and this is mediated by the inability of IL-2 to stimulate a complete IL-4 secretion response. In contrast, exogenous IL-4 reverses the unresponsiveness of both NOD thymic and peripheral T cells completely, and this is associated with the complete restoration of an IL-2 secretion response. Furthermore, the in vivo administration of rIL-4 to prediabetic NOD mice protects them from diabetes. Thus, the ability of rIL-4 to reverse completely the NOD thymic and peripheral T cell proliferative defect in vitro and protect against diabetes in vivo provides further support for a causal relationship between this T cell proliferative unresponsiveness and susceptibility to diabetes in NOD mice.

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E H Leiter

Papers

Interleukin 4 reverses T cell proliferative unresponsiveness and prevents the onset of diabetes in nonobese diabetic mice.