The optimal activation of cytotoxic T lymphocytes requires metabolically intact stimulator cells not only for the activation of the interleukin 2-producing helper cells.

Primary cytotoxic responses in macrocultures against UV-treated stimulator cells or glutaraldehyde-fixed stimulator cells in the presence of third party stimulator cells were studied to investigate whether metabolically active stimulator cells are only required for the activation of interleukin 2-producing helper cells. Cultures containing splenic responder cells in a mixture with allogeneic UVtreated stimulator cells of a mouse strain X plus conventional (i.e., only ?-irradiated) stimulator cells of a third strain Y were found to generate strong cytotoxic activity against cells of strain Y but not strain X. Macrocultures with conventional stimulator cells of strain Y plus UVtreated stimulator cells of (X x Y)Fl hybrid mice also failed to generate substantial CTL activity against cells of strain X. The response against antigens of strain X was not reconstituted by interleukin 2(IL 2) containing factors. UV-treated stimulator cells did not suppress the response against conventional stimulator cells of the same H-2 haplotype. These results provide suggestive evidence that CTL precursor cells are optimally activated in primary cytotoxic responses when their receptors interact with antigen on metabolically active stimulator cells. The experiments excluded the possibility that the metabolically active stimulator cells were on/y required for the activation of an unlinked helper effect (i.e., for the stimulation of interleukin 2 production) because the response against the third party stimulator cell proceeded in the same culture at normal magnitude. Our experiments with the (X x Y)F1 stimulator cells also excluded the possibility that the metabolically active (i.e., UV-sensitive) stimulator cells were only required for the stimulation of a type of helper cell (or for the production of an antigen-specific helper factor) that interacts with the CTL precursor cell through a cellular antigen bridge (antigenically linked helper effect). A antigenically linked helper effect through a molecular antigen bridge was not formally excluded. Cold target competition experiments revealed that the antigenic structures were not detectably destroyed by the UV-irradiation procedure. Cultures that received UV-treated stimulator cells on day 0 and an optimal dose of corresponding conventional stimulator cells on day 2 generated still weaker cytotoxicity than control cultures with conventional stimulator cells on day 0, indicating that the metabolically active (UVsensitive) stimulator cells were required in the early phase of the culture to achieve optimal cytotoxic responses.