SummaryWe have developed a model of peripheral in vivo T cell tolerance that is induced by administration of the protein superantigen staphylococcal enterotoxin B (SEB). Rather than activating V08+ T cells, in vivo administration of SEB induced in them a profound state of anergy. This was shown by their failure to proliferate to subsequent in vitro restimulation with SEB or to antiV08 antibodies. This unresponsiveness was V08 specific since T cells from SEB-immunized mice responded normally to other antigens. 8 d after SEB administration, there was no reduction in the number of Va8+ T cells or in the intensity of V08 T cell receptor (TCR) expression . Although a portion of the V08+ T cells from SEB-primed mice were able to express interleukin 2 receptors (IIr2Rs), they failed to proliferate in response to exogenous IL2, indicating they were defective in their IL-2 responsiveness. 2-4 wk after SEB administration, there was a reduction of -50% in the number of V08+ cells in immunized compared with control animals . There was, however, no reduction in the level of TCR expression on the remaining V08+ cells. These data demonstrate that proteins that activate T cells in vitro in a V/3-specific manner can induce a state of anergy in peripheral T cells in vivo and may possibly further mediate clonal deletion in a portion of the tolerized cells.

A major mechanism for generating tolerance in developing T cells is the intrathymic clonal deletion of T cells that have receptors for those self antigens that are presented on hematopoietic cells. The mechanisms of tolerance induction to antigens not expressed in the thymus remain unclear. Tolerance to self antigens can be generated extrathymically through the induction of clonal nonresponsiveness in T cells with self-reactive receptors. A second mechanism of extrathymic tolerance was identified: clonal elimination of mature T cells with self-reactive receptors that had previously displayed functional reactivity.

To elucidate the parameters that lead to superantigen induced non-responsiveness, an in vitro model for studying primary and secondary responses to the bacterial superantigen staphylococcal enterotoxin B (SEB) was established. Upon re-activation with SEB, in vitro SEB primed T cells show an early proliferative response that 'quenches' in time and is severely impaired 3 days after re-stimulation. Despite their overall impaired proliferative capacity and IL-2 production, these T cells are able to produce IFN-gamma and to up-regulate activation markers CD69 and IL-2R alpha upon re-stimulation with SEB, demonstrating that SEB non-responsiveness is not absolute. Rather, it reflects the inability to mount an ongoing proliferative response upon re-stimulation with SEB. Our results also demonstrate that SEB-induced non-responsiveness is not simply the result of presentation in the absence of co-stimulation, since presentation of SEB on highly purified dendritic cells during the primary response did not prevent the induction of non-responsiveness. As previously shown, SEB induces a Th1 phenotype in responding CD4+ T cells. Skewing towards a Th2 phenotype by adding IL-4 and antibodies to IFN-gamma did not prevent the induction of non-responsiveness by SEB. Interestingly, T cells pretreated with plate-bound anti-CD3 epsilon and anti-V beta 8 were also non-responsive to SEB re-stimulation. Thus, non-responsiveness to SEB (defined here as inability to produce IL-2 and proliferate) seems to reflect an intrinsic inability of previously activated T cells to respond to SEB, probably reflecting differences in signal transduction pathways used by naive versus previously activated T cells.

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