Requirement of CD4⁺ T cells and antigen-presenting cells for primary in vitro generation of CD8⁺ cytotoxic T cells against L(d)-binding self-peptide p2Ca

We investigated the cellular requirement for primary in vitro generation of cytotoxic T-lymphocytes (CTL) in BALB/c spleen cells against L(d)-binding self-peptide p2Ca. Depletion of CD4⁺ T-cells in vitro by pretreatment with anti-CD4 monoclonal antibody (mAb) and complement or in vivo by administration of anti-CD4 mAb abrogated generation of CTL. Depletion of adherent cells by passing spleen cells through a nylon wool (NW) column also abrogated generation of CTL. Addition of peritoneal exudate cells (PEG) to spleen cells passed through the NW column restored CTL generation. These findings indicate that both CD4⁺ T-cells and antigen-presenting cells (APC) were necessary for CTL generation. Treatment of PEC with paraformaldehyde (PFA), but not mitomycin-C (MMC) abrogated their ability to restore CTL generation when mixed with spleen cells from the NW column, suggesting that an endocytic pathway could be involved in presentation of p2Ca on APC.