In healthy individuals, T cells react against incoming pathogens, but remain tolerant to self-antigens, thereby preventing autoimmune reactions. CD4 regulatory T cells are major contributors in induction and maintenance of peripheral tolerance, but a regulatory role has been also reported for several subsets of CD8 T cells. To determine the molecular basis of peripheral CD8 T-cell tolerance, we exploited a double transgenic mouse model in which CD8 T cells are neonatally tolerized following interaction with a parenchymal self-antigen. These tolerant CD8 T cells have regulatory capacity and can suppress T cells in an antigen-specific manner during adulthood. Dickkopf-3 (DKK3) was found to be expressed in the tolerant CD8 T cells and to be essential for the observed CD8 T-cell tolerance. In vitro, genetic deletion of DKK3 or blocking with antibodies restored CD8 T-cell proliferation and IL-2 production in response to the tolerizing self-antigen. Moreover, exogenous DKK3 reduced CD8 T-cell reactivity. In vivo, abrogation of DKK3 function reversed tolerance, leading to eradication of tumors expressing the target antigen and to rejection of autologous skin grafts. Thus, our findings define DKK3 as a immune modulator with a crucial role for CD8 T-cell tolerance.immune tolerance | parenchymal cells | autoimmunity

Motifs of peptides naturally associated with H-2E k and E d molecules were determined by (I) pool sequencing of natural ligand mixtures and (II) sequencing of Individual natural ligands followed by their alignment to the basic motif suggested by pool sequencing. The data reveal nine amlno acid motifs with Interaction sites at relative positions P1, P4, P6 and P9, with specificities that are identical at some but different at other anchor positions between E d and E k motifs, Illustrating the different requirements for peptides to be presented by these two MHC molecules. The anchors with the most restricted specificity are P1 and P9.

Keratinocytes have a pivotal role in the regulation of immune responses, but the impact of antigen presentation by these cells is still poorly understood, particularly in a situation where the antigen will be presented only in adult life. Here, we generated a transgenic mouse model in which keratinocytes exclusively present a myelin basic protein (MBP) peptide covalently linked to the major histocompatibility complex class II β-chain, solely under inflammatory conditions. In these mice, inflammation caused by epicutaneous contact sensitizer treatment resulted in keratinocyte-mediated expansion of MBP-specific CD4(+) T cells in the skin. Moreover, repeated contact sensitizer application preceding a systemic MBP immunization reduced the reactivity of the respective CD4(+) T cells and lowered the symptoms of the resulting experimental autoimmune encephalomyelitis. This downregulation was CD4(+) T-cell-mediated and dependent on the presence of the immune modulator Dickkopf-3. Thus, presentation of a neo self-antigen by keratinocytes in the inflamed, adult skin can modulate CD4(+) T-cell auto-aggression at a distal organ.

Functional inactivation of self-reactive T lymphocytes contributes to the maintenance of immunologic self-tolerance. At the same time, tolerance induction limits immune responses against tumors expressing tolerizing self-antigens. Some cancer therapies include the adoptive transfer of tumor-reactive T lymphocytes into lymphopenic patients. Lymphopenia provides an activation signal to T lymphocytes, which undergo lymphopeniainduced proliferation (LIP), acquire effector functions, and reject tumors. However, it is so far unknown to which extent LIP may result in reversal of established antigen-specific CD8 T-cell tolerance. Here, we report that neonatally induced dominant CD8 T-cell tolerance remained stable under lymphopenic conditions also in the presence of systemic inflammation induced by Toll-like receptor ligands. However, when lymphopenic recipients were irradiated, the tolerant status was lost, because CD8 T cells acquired effector functions in an interleukin-15-dependent fashion and efficiently rejected tumors. In conclusion, we show that lymphopenia is not sufficient to break CD8 T-cell tolerance. Furthermore, we demonstrate that pretreatment regimens are crucial to circumvent this problem and to optimize adoptive T-cell therapy. (Blood. 2010;115: 2196-2202)