Studies on the induction and expression of T cell-mediated immunity. XI. Inhibition of the "lethal hit" in T cell-mediated cytotoxicity by heterologous rat antiserum made against alloimmune cytotoxic T lymphocytes.

Xenogeneic antisera were prepared in rats by i.p. injection of alloimmune thymus-derived cytotoxic T lymphocytes (CTL) that were “activated” on monolayers of PHA-P-coated L929 fibroblasts. The Rat Anti-T* serum (termed RAT*) effectively inhibits both specific and nonspecific (lectin-induced) T cell mediated cytotoxicity. Analysis of the mechanism of RAT* antiserum inhibition revealed the following: 1) The antiserum was not cytotoxic by itself to CTL nor did it reduce their functional activity after preincubation at 37°C for 4 hr. 2) The inhibition was not due to interaction of the antiserum with target cell antigens. 3) Suspension of CTL-target cell conjugates in RAT* antiserum did not alter conjugate frequencies as determined microscopically in a single-cell cytotoxicity assay yet still completely blocked lysis. However, the serum could inhibit conjugation when performed directly in the presence of high concentrations of serum. 4) The antiserum could inhibit cytotoxicity when added to ongoing CTL reactions at a time when EDTA or EGTA could not inhibit lysis. Thus, the RAT* serum could inhibit the “lethal hit” after “programming for lysis” had occurred. 5) As an appropriate negative control, the serum could not inhibit the lysis of antibody-coated P815 cells by human effector K cells. Exhaustive absorption on P815, normal C57 spleen cells, C57 thymocytes, or alloimmune CTL did not reduce the inhibitory activity of the RAT* serum. However, absorption on “activated” cells, i.e., Con A blasts, CTL-P815 conjugates, or PHA-P “activated” alloimmune T lymphocytes, substantially removed the blocking activity. These data collectively suggest that CTL possess a lytic mechanism that is cryptic (not reactive with RAT* antiserum) but becomes exposed when activated by interaction with specific antigen or lectins. We propose that RAT* serum recognizes structures involved in the lytic step of cytolysis independent of antigen recognition or lymphocyte-target binding.