Development of autoreactive L3T4+ T cells from double‐negative (L3T4−/Ly‐2−) Thy‐1+ spleen cells of normal mice

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Abstract
Thy-l+/L3T4−/Ly-2− spleen cells were purified from normal C57BL/6 (B6) and C.B-17 mice. Cells within this subset expressed the T cell receptor (TcR) for antigen: the majority of cells in this subset were CD3+; a fraction of the cells was stained with the monoclonal antibody (mAb) F23.1; and the TcR molecule was immunoprecipitable with mAb F23.1 from cells within this subset. In limiting dilution analyses, about 1/30 cells within this subset were growth inducible in vitro by stimulation with phorbol myristate acetate (PMA) plus ionomycin; conditioned media containing interleukin (IL)1, IL2, IL3 or IL4 activity neither triggered nor promoted in vitro growth of these cells. The in vitro generated T cells displayed the Thy-l+/L3T4+/Ly-2− surface phenotype and were self-reactive, i.e., proliferated preferentially in response to syngeneic stimulator cells, and secreted IL2 and IL3 only in response to syngeneic but not allogeneic stimulator cells. The proliferative response of these cells to syngeneic stimulator cells was blocked by anti-self Ia mAb. This autoreactive helper T cell subset was not inducible in purified Thy-1+ spleen cell subsets from athymic nude mice or scid mice. Autoreactive helper T cells did not express detectable levels of the IL2 receptor (IL2R), and their proliferative response was not blocked by anti-IL2R mAb. From PMA plus ionomycin-stimulated double-negative Thy-l+ spleen cells, 14 T cell clones were established in long-term culture which displayed the CD3+CD4+CD8− surface phenotype and were self-reactive.