SUPPRESSION OF ALLOGRAFT RESPONSES BY COMBINING ALLOANTIGEN-SPECIFIC-IV PRE-SENSITIZATION WITH SUBOPTIMAL DOSES OF RAPAMYCIN

C57BL/6 (B6) mice were injected i.v. with class I H-P-disparate B1O.QBR spleen cells (10(7)/mouse). This regimen, termed donor alloantigen-specific i.v. pre-sensitization (DSP), induced almost complete reduction of the anti-B1O.QBR mixed lymphocyte reaction (MLR) that has been regarded to represent a cytotoxic T lymphocyte (CTL)-independent graft rejection pathway. Because the DSP regimen failed to affect the generation of CTL responses, it did not prolong graft survival. Repeated (four or 11 times) administration in vivo (during 5 or 18 days) of rapamycin at suboptimal doses (0.5-2.0 mg/kg/day) failed to eliminate the capacities to exhibit MLR as well as to generate CTL responses. The suppression of CTL responses was achieved only through the combination of these two treatments. It was also shown that prolongation of skin graft survival was not induced by either of a single DSP or rapamycin treatment alone, but by the combination of these. Potent suppression of CTL responses was also induced when a single DSP was combined with repeated injection of a suboptimal dose (0.75 mg/kg/day) of another immunosuppressive drug, FK506, instead of rapamycin. However, there was a substantial difference in cellular mechanisms underlying the suppression of CTL responses by the above two different combinations. Under conditions in which lymphoid cells from mice receiving the treatments with DSP plus FK506 failed to generate CTL responses, the addition of recombinant IL-2 (rlL-2) to cultures restored the CTL generation, suggesting that CTL precursors themselves are not attenuated by the combined treatment. In contrast, the addition of rlL-2 to cultures of lymphoid cells from mice receiving the treatments with DSP plus rapamycin failed to increase the generation of CTL responses. These results indicate that (i) DSP capable of eliminating a CTL-independent graft rejection pathway fails to suppress a CTL-involved pathway, because both CTL precursors and CTL helpers are DSP resistant; (ii) the administration of suboptimal doses of rapamycin alone is not sufficient for the suppression of CTL responses; and (iii) the combination of DSP with rapamycin treatment can reduce CTL responses through attenuation of CTL precursors, thus resulting in the prolongation of graft survival.