Antigen-Specific T Cell-Mediated Suppression: IV. Role of Macrophages in Generation of L-Glutamic Acid⁶⁰-L-Alanine³⁰-L-Tyrosine¹⁰ (GAT)-Specific Suppressor T Cells in Responder Mouse Strains¹

The role of adherent cells (MØ) in 1) primary in vitro PFC responses and 2) in vitro generation of suppressor T cells (T_s) to the antigen L-glutamic acid⁶⁰-L-alanine³⁰-L-tyrosine¹⁰ in responder strain mice has been investigated. Removal of MØ from spleen cell suspensions by either serial culture on plastic surfaces or passage over Sephadex G-10 beads yielded lymphocytes unable to give primary in vitro PFC responses under modified Mishell-Dutton conditions to either soluble GAT or the particulate antigens sheep red blood cells (SRBC) or GAT-complexed electrostatically to methylated bovine serum albumin (GAT-MBSA). However, the viability of such MØ-depleted cultures was significantly lower than control cultures of unseparated spleen cells. Addition of 5 × 10^-5 M 2-mercaptoethanol (2-ME) to the culture medium restored viability and permitted nonadherent cells to respond to SRBC and GAT-MBSA, but not to soluble GAT. The response of nonadherent cells to soluble GAT could be restored in the absence of 2-ME by the addition of 1 to 3 × 10⁵ x-irradiated, anti-Thy 1.2 + C-treated peritoneal exudate cells (MØ), or in the presence of 2-ME by 1 to 2 × 10⁴ MØ. This low number of MØ was identical to the number of GAT-bearing MØ necessary in the presence of 2-ME to give optimal GAT PFC responses. Taken together, these data indicate a crucial antigen-presenting role for MØ in primary in vitro anti-GAT responses. Removal of such antigen-presenting cells from responder spleen cells left a lymphocyte population, which, when cultured for 2 days with normally immunogenic (10 µg/ml) concentrations of GAT, gave rise to highly active Thy 1.2⁺ GAT-specific suppressor cells. This T_s induction was demonstrated in several responder strains. The GAT-T_s were unable to suppress the in vitro responses to SRBC or the copolymer L-glutamic acid⁵⁰-l-tyrosine⁵⁰ (GT) as GT-MBSA. Increasing the concentration of GAT to ≥ 30 µg/ml permitted the induction of GAT-T_s in unseparated spleen cells. Thus, in responder mice, MØ able to present antigen in an immunogenic form play a central role in regulating the balance of activated T_H and T_s, suggesting that an Ir gene defect in nonresponder mice at the MØ level for this presentation function may account for the predominant suppressor T cell responses of such mice.