A dual altered peptide ligand down-regulates myasthenogenic T cell responses by up-regulating CD25- and CTLA-4-expressing CD4(+) T cells

Immunization of mice with two myasthenogenic peptides, p195–212 and p259–271, which are sequences of the human acetylcholine receptor, resulted in myasthenia gravis (MG)-associated immune responses. A dual altered peptide ligand (APL) composed of the two APLs of the myasthenogenic peptides inhibited, in vitro and in vivo, those responses. The aims of this study were to further elucidate the mechanism/s by which the dual APL down-regulates MG-associated responses in vivo and characterize the cell population/s involved in this immunomodulatory suppressive effect. We have shown here that s.c. administration of the dual APL activates CD4CD25-expressing cells in lymph nodes (LN) of SJL mice. Furthermore, depletion of these cells diminished significantly the inhibitory effect of the APL on p195–212-specific proliferative responses. Depletion of the CD4(+)CD25(+) cells was accompanied with a decrease in the secretion of the immunosuppressive cytokine, transforming growth factor (TGF)-β. Administration of the dual APL resulted also in the up-regulation of the expression of cytotoxic T lymphocyte antigen (CTLA)-4 and in a down-regulated expression of CD28 on LN cells. Blockade of the CTLA-4 function, in vitro, abrogated the inhibitory effect of the dual APL on the proliferative responses specific to p195–212. Thus, our results suggest that the active suppression exerted by the dual APL is mediated by the CD4(+)CD25(+) immunoregulatory cell population, either directly through the CTLA-4 molecule expressed on these cells, and/or indirectly by causing the differentiation of other regulatory T cell population/s that secrete immunosuppressive cytokines

The nature of the active suppression of responses associated with experimental autoimmune myasthenia gravis by a dual altered peptide ligand administered by different routes

Myasthenia gravis (MG) and experimental autoimmune MG (EAMG) are T-cell regulated, antibody-mediated diseases. Peptides p195–212 and p259–271 of the human acetylcholine receptor (AChR) α-subunit, were previously shown to be immunodominant T cell epitopes in MG patients as well as in SJL and BALB/c mice, respectively. A dual altered peptide ligand (APL) composed of the two single amino acid analogs of the myasthenogenic peptides was shown to inhibit, in vitro and in vivo, MG-associated autoimmune responses. Furthermore, the dual APL was shown to down-regulate the clinical manifestations of an established EAMG in C57BL/6 mice injected with Torpedo AChR (TAChR). In the present study we attempted the elucidation of the mechanism(s) by which the dual APL down-regulates EAMG-associated responses. It is shown here that the dual APL acts by actively suppressing, in a specific manner, myasthenogenic T cell responses. The active suppression is mediated, at least partially, by the up-regulation of the secretion of TGF-β following administration of the dual APL. The up-regulated secretion of TGF-β is accompanied by down-regulation of IFN-γ and IL-2 [T helper (Th) 1-type cytokine] secretion and by an up-regulation of IL-10 secretion (Th2-type cytokine). Furthermore, the inhibitory effect of the dual APL could be adoptively transferred to p195–212 or TAChR-immunized mice. The down-regulation of IL-2 secretion and the ability of recombinant IL-2 to rescue lymph node cells of mice treated with the dual APL from a state of unresponsiveness suggests that the dual APL acts also, at least partially, by causing the cells to undergo anergy