Barbara Metzler

Mechanisms of central and peripehral T‐cell tolerance: lessons from experimental models of multiple sclerosis

Summary: Studies of experimental autoimmune encephalomyelitis in conventional and transgenic mouse models have clarified mechanisms of central and peripheral tolerance to myelin antigens. It is now clear that myelin antigens are expressed in the thymus and their expression can influence generation of the potential autoimmune T‐cell repertoire. How autoreactive T cells escape tolerance in the thymus is largely unclear. One mechanism has been revealed through the use of a transgenic mouse expressing a T‐cell receptor specific for a myelin antigen. T cells specific for the N‐terminal epitope of myelin basic protein escape tolerance through low avidity interaction. The affinity of antigen binding to MHC also proves to be important for induction of peripheral tolerance. Peptides may be administered in solution to adults in order to reinstate peripheral tolerance and suppress disease. Induction of antigen‐specific suppression with synthetic peptides can result in either linked or bystander suppression and this appears to involve the generation of T cells secreting suppressive cytokines. The use of altered peptide ligands fur induction of peripheral tolerance has been investigated. This can be achieved but the complexity of the approach argues against its use for treatment of human autoimmune diseases.

Mucosal Tolerance in a Murine Model of Experimental Autoimmune Encephalomyelitisa

Attempts to induce oral tolerance in a murine model of EAE with either the dominant T-cell epitope or whole protein have failed. These results may, in part, be due to the extraordinarily low affinity for class II MHC displayed by the dominant T-cell epitope. This belief is supported by experiments using a high-affinity analogue of the peptide that was capable of inducing tolerance at a high dose. By contrast, peptide inhalation has proven an effective route for induction of mucosal tolerance in this model. Most importantly, the inhalation of a single peptide could inhibit disease induced by the complex mixture of antigens found in whole myelin. Peptide inhalation was effective both before and after disease induction, and there was a positive correlation between affinity of class II binding and tolerogenicity of a panel of analogues of the N-terminal peptide of myelin basic protein.

Modulation of T cell homeostasis and alloreactivity under continuous FTY720 exposure

The immunomodulator FTY720 inhibits lymph node (LN) and thymic egress, thereby constraining T cell circulation and reducing peripheral T cell numbers. Here, we analyzed in mouse models the as yet scarcely characterized impact of long-term (up to 6 months) FTY720 exposure on T cell homeostasis and possible consequences for alloreactivity. In green fluorescent protein (GFP) hemopoietic chimeras, the turnover of (initially GFP(-)) peripheral T cell pools was markedly delayed under FTY720, while normal homeostatic differences between CD4 and CD8 T cell sub-populations were retained or amplified further. Homeostatic proliferation was enhanced, and within shrinking T cell pools, the proportions of effector memory phenotype CD4 T cells (CD4T(PEM)) increased in spleens and LNs and of central memory phenotype CD8 T cells (CD8T(PCM)) in LNs. By contrast, the fractions of CD8T(PEM) and CD4T(PCM) remained stably small under FTY720. The enrichment for CD4T(PEM) and CD8T(PCM) correlated with larger proportions of IFNgamma-producing T cells upon nonspecific but not allospecific stimulation. Splenic CD4 T cells from FTY720-treated mice proliferated more strongly upon transfer to semi-allogeneic hosts. However, heart allograft survival was not compromised in FTY720 pre-treated recipients. It correlated with reduced intra-graft CD8 T cells, and the longest surviving transplants contained the highest numbers of CD4 T cells. Thus, continuous FTY720 exposure reveals differential homeostatic responses by memory phenotype CD4 and CD8 T cell sub-populations, and it may enhance alloreactive CD4 T cell proliferation and tissue infiltration without accelerating allograft rejection.

Kinetics of Peptide Uptake and Tissue Distribution Following a Single Intranasal Dose of Peptide

Stimulation with the combination of PDB plus ionomycin induced LFA-l/ICAM-1-dependent homotypic adhesion of tonsil B cells. Adhesion of tonsil B cells in our system induced tyrosine phosphorylation of Pyk-2. Disruption of homotypic adhesion and concomitant inhibition of induction of protein tyrosine phosphorylation was achieved by physical separation of the cells and by treatment with methyl-2,5-dihydroxycinnamate (MDHC), an inhibitor of protein tyrosine phosphorylation. These results suggest that protein tyrosine phosphorylation that is associated with homotypic adhesion is mediated by LFA-l/ICAM-1 interactions.

Antigen-Specific Tolerance Induction and the Immunotherapy of Experimental Autoimmune Disease

Antigen-specific tolerance induction is the ultimate goal for specific immunotherapy of autoimmune diseases. Here we will discuss recent experiments designed to induce tolerance following mucosal administration of antigens in a mouse model of experimental autoimmune encephalomyelitis (EAE). We were unable to induce oral tolerance either with whole myelin, myelin basic protein (MBP) or the immunodominant peptide antigen. Oral tolerance was possible, however, with an analogue of the immunodominant peptide modified to increase its affinity for the restricting major histocompatibility complex (MHC) antigen. By contrast, intranasal deposition of peptide antigen proved highly effective for both prevention and treatment of EAE. Prevention of disease was directly related to the antigenic property of the peptide which, in itself, was related to affinity for MHC. Notably, administration of a single peptide was shown to inhibit disease involving multiple epitopes. We investigated the resulting bystander regulation by studying the cellular basis of peripheral tolerance in a transgenic model. These studies indicate that bystander regulation may be the consequence of selective cytokine secretion.