Production of T cell differentiation factor in syngeneic lymphocyte macrophage culture for cytotoxic T lymphocyte generation.
This study demonstrated that T cell differentiation factor (TCDF) was produced in syngeneic lymphocyte-macrophage cultures. Conditioned medium containing TCDF and interleukin 2 (IL 2) induced the differentiation of leukoagglutinin (LA)-activated cytotoxic T lymphocyte precursors (CTLp) into cytotoxic T lymphocyte (CTL) effectors. The production of TCDF and IL 2 peaked at day 4 to 5 in cultures containing normal spleen cells, syngeneic peritoneal macrophages, and indomethacin. Macrophages and T cells with Thy-1+, L3T4+, and Lyt-2- phenotype were needed for TCDF production. There was no requirement for xenogeneic serum in the culture medium; thus, TCDF could be produced in a syngeneic system. Recognition of self Ia molecules appeared to be essential for TCDF production, which was completely abolished by the addition of monoclonal anti-Ia antibody. In our experiments, removal of IL 2 from conditioned medium containing TCDF abolished its ability to generate LA-activated CTL. However, the cytotoxic response could be restored by the addition of a small amount (5 U/ml) of purified human recombinant IL 2 (HRIL 2), which alone was unable to generate LA-activated CTL at this dose. The generation of LA-activated CTL by high dose HRIL 2 (greater than 50 U/ml) was likely due to the endogenous production of TCDF. The bulk of TCDF could be separated from IL 2 by gel filtration in a Sephadex G-100 column. The peak of TCDF activity was concentrated at a m.w. of 16K dalton, and there was very little IL 2 activity in these fractions. When added alone to the LA-activated lymphocyte cultures, these active fractions were unable to induce CTL; supplementation of exogenous IL 2 was needed to restore the cytotoxic responses. Our findings indicate that both IL 2 and TCDF, which are needed in CTL generation. are produced in syngeneic cultures in the absence of antigenic or mitogenic stimulation.
D000373:Agglutinins / Q000494:pharmacology; D000818:Animals; D003470:Culture Media; D005260:Female; D000949:Histocompatibility Antigens Class II / Q000235:genetics / Q000276:immunology; D007376:Interleukin-2 / Q000096:biosynthesis* / Q000302:isolation & purification; D007962:Leukocytes; D008213:Lymphocyte Activation; D007959:Lymphocyte Culture Test, Mixed; D008264:Macrophages / Q000378:metabolism*; D051379:Mice; D008810:Mice, Inbred C57BL; D008811:Mice, Inbred DBA; D010529:Peritoneal Cavity / Q000166:cytology; D010641:Phenotype; D011506:Proteins; D013154:Spleen / Q000166:cytology; D013602:T-Lymphocytes, Cytotoxic / Q000145:classification / Q000276:immunology*; D013997:Time Factors
D000373:Agglutinins; D003470:Culture Media; D000949:Histocompatibility Antigens Class II; D007376:Interleukin-2; D011506:Proteins; C100155:leukoagglutinins, leukocytes