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Articles |
Department of Microbiology and Immunology and The Howard Hughes Medical Institute, Stanford University School of Medicine, Stanford, California 94305-5402
The mechanism of self-tolerance in the CD4+ T cell compartment was examined in a double transgenic (Tg) model in which T cell receptor (TCR)-
/β Tg mice with specificity for the COOH-terminal peptide of moth cytochrome c in association with I-Ek were crossed with antigen Tg mice. Partial deletion of cytochrome-reactive T cells in the thymus allowed some self-specific CD4+ T cells to be selected into the peripheral T cell pool. Upon restimulation with peptide in vitro, these cells upregulated interleukin (IL)-2 receptor but showed substantially lower cytokine production and proliferation than cells from TCR Tg controls. Proliferation and cytokine production were restored to control levels by addition of saturating concentrations of IL-2, consistent with the original in vitro definition of T cell anergy. However, the response of double Tg cells to superantigen stimulation in the absence of exogenous IL-2 was indistinguishable from that of TCR Tg controls, indicating that these self-reactive cells were not intrinsically hyporesponsive. Measurement of surface expression of Tg-encoded TCR and β chains revealed that cells from double Tg mice expressed the same amount of TCR-β as cells from TCR Tg controls, but only 50% of TCR-, implying expression of more than one chain. Naive CD4+ T cells expressing both Tg-encoded and endogenous chains also manifested an anergic phenotype upon primary stimulation with cytochrome c in vitro, suggesting that low avidity for antigen can produce an anergic phenotype in naive cells. The carboxyfluorescein diacetate succinimidyl ester cell division profiles in response to titered peptide ± IL-2 indicated that expression of IL-2 receptor correlated with peptide concentration but not TCR level, whereas IL-2 production was profoundly affected by the twofold decrease in specific TCR expression. Addition of exogenous IL-2 recruited double Tg cells into division, resulting in a pattern of cell division indistinguishable from that of controls. Thus, in this experimental model, cells expressing more than one chain escaped negative selection to a soluble self-protein in the thymus and had an anergic phenotype indistinguishable from that of low avidity naive cells. The data are consistent with the notion that avidity-mediated selection for self-reactivity in the thymus may lead to the appearance of anergy within the peripheral, self-reactive T cell repertoire, without invoking the induction of hyporesponsiveness to TCR-mediated signals.
Key Words: T cell receptor • double transgenic model • thymic deletion • T cell anergy • carboxyfluorescein diacetate succinimidyl ester labeling
Laila Girgis was supported by a National Health and Medical Research Council Medical Postgraduate Research Scholarship and Barbara Fazekas de St. Groth was supported by a Wellcome Trust Senior Research Fellowship. This work was supported by grants from the National Health and Medical Research Council of Australia, the Wellcome Trust, and the Medical Foundation of the University of Sydney.
Abbreviations used: CFSE, carboxyfluorescein diacetate succinimidyl ester; HEL, hen egg lysozyme; MCC, moth cytochrome c; PI, propidium iodide; rhIL-2, recombinant human IL-2; SEA, Staphylococcal enterotoxin A; TCM, tissue culture medium; Tg, transgenic.
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