Lyt phenotypes of alloreactive precursor and effector cytotoxic T lymphocytes.

Lyt 1.2 and Lyt 2.2 antisera were produced by alloimmunization of mice with use of established procedures. Specific depletion of either Lyt 1+ or Lyt 2+ C57BL/6 responder spleen cells by negative selection with antibody and complement (C) before allogeneic stimulation in MLC resulted in almost complete abrogation of the ability of the residual cells to generate cell-mediated cytotoxicity, as measured in a 51Cr-release cell-mediated lympholysis (CML) assay. Further, when the responder leukocytes pretreated with anti-Lyt-1.2 + C were mixed with anti-Lyt-2.2 + C treated cells, the generation of cytotoxicity in allogeneic mixed leukocyte culture (MLC) was not significantly restored. This result suggests an obligate role for Lyt 1+2+ T cells in the generation of cell-mediated cytotoxicity in allogeneic MLC. Significant synergistic interactions in the generation of cytotoxicity were observed in these mixtures when negative selection with anti-Lyt-1.2 was suboptimal, presumably leaving some Lyt 1+2+ cells. One possible interpretation of our results is that the generation of allogeneic cell-mediated cytotoxicity in MLC results from collaboration between Lyt 1+2- amplifier cells and Lyt 1+2+ precursor cytotoxic T cells. The Lyt phenotype of effector cytotoxic T lymphocytes was also analyzed. C57BL/6 spleen cells presensitized in vitro with allogeneic leukocytes were treated with normal mouse serum (NMS), anti-Lyt-1.2, or anti-Lyt-2.2, plus C and assayed in CML at equal viable cell concentrations. Specific depletion of Lyt 2+ alloactivated cells abrogated cytotoxicity, whereas a significant reduction but not abrogation of CML resulted from depletion of Lyt 1+ cells. Thus, after allogeneic sensitization, effector CTL were present in both the operationally defined Lyt 1+2+ and Lyt 1-2+ subsets.