Interleukin 2 is not sufficient as helper component for the activation of cytotoxic T lymphocytes but synergizes with a late helper effect that is provided by irradiated I-region-incompatible stimulator cells.

lnterleukin 2-containing supernatants from concanavalin A-activated spleen cells (CSCS) were found to provide strong helper activity for cytotoxic T lymphocyte (CTL) responses against allogeneic stimulator cells in microculture systems, but provided usually insufficient help for CTL responses against I-region compatible allogeneic or TNP-haptenated syngeneic stimulator cells. The interleukin 2-containing supernatant from HGG-activated AODH 7.1 hybridoma cells also mediated only relatively weak CTL responses against TNP-haptenated syngeneic cells in microcultures. Both types of supernatants, however, supported substantial responses against TNP-haptenated syngeneic stimulator cells if irradiated allogeneically activated syngeneic T cells or irradiated allogeneic spleen cells were added to the cultures. The allogeneic cells and the activated syngeneic T cells provided little helper activity if they were added in the absence of the interleukin 2-containing supernatants, thus demonstrating a synergistic effect between these 2 helper components. An I-region difference was sufficient for the helper effect of the allogeneic cells and control experiments showed that the presence of foreign I-region determinants could not be substituted for the TNP-haptenated stimulator cells. The AODH 7.1 supernatant was shown to provide a helper effect only if it was added at day 0, and had no effect at day 3 of the culture; but its helper effect was enhanced by irradiated allogeneic cells irrespective of whether these cells were added at day 0 or day 3. This indicated that the allogeneic cells provided a helper effect that was required during the late phase of the CTL response and therefore was qualitatively different from the early helper effect of interleukin 2. The late addition of allogeneic cells could not be substituted by the AODH 7.1 supernatant. Some of our CSCS batches and concentrated CSCS preparations were found to provide the late helper effect and to support CTL responses equally well in the presence or absence of I-region-incompatible cells. This indicated that the late helper effect was also mediated by a soluble mediator. Finally, our experiments showed that the helper cells that responded to the I-region differences and synergized with CSCS were provided by the irradiated allogeneic spleen cell populations and also by the responder cells