OKT3 monoclonal antibody induces production of colony-stimulating factor(s) for granulocytes and macrophages in cultures of human T lymphocytes and adherent cells.
OKT3 monoclonal antibody (mab) recognizes a membrane antigen associated with the T cell antigen recognition receptor, and is known to be mitogenic and to induce lymphokine production. Our studies demonstrate the ability of OKT3 mab to induce from cultures of human T lymphocytes supplemented with adherent cells the production of colony-stimulating factor(s) for granulocytes and macrophages (GM-CSF) and interferon-gamma (IFN-gamma), an inhibitor of clonal growth of hematopoietic progenitor cells. As has been shown for the mitogenic and IFN-gamma-inducing activity of OKT3 mab, the induction of GM-CSF release in cultures of T cells is strictly dependent on the presence of adherent cells. However, the concentrations of OKT3 mab required for optimal GM-CSF production (50 ng/ml) were found to be 80-fold higher than those sufficient for maximal IFN-gamma production, proliferation, and interleukin 2 production. IFN-gamma activity induced by OKT3 mab partially inhibited colony and cluster formation from progenitor cells of granulocytes and macrophages in vitro. Therefore, neutralization of the IFN-gamma by monoclonal anti-human-IFN-gamma antibody before assay of conditioned medium in bone marrow cultures significantly enhanced the detection of GM-CSF. Kinetic studies demonstrated maximal cumulative GM-CSF production in response to optimal OKT3 mab concentrations on days 4 through 6 in cultures of T cells supplemented with 15% adherent cells. Highly enriched OKT4+ and OKT8+ T cell subsets co-cultured with adherent cells in the presence of OKT3 mab both produced GM-CSF and IFN-gamma and showed similar dose-response curves to OKT3 mab. The requirement for the presence of adherent cells could not be overcome by the addition of purified interleukin 1 or macrophage supernatants. Studies using irreversible inhibitors of DNA (mitomycin C) or protein biosynthesis (emetine-HCl) revealed the necessity of intact DNA synthesis and translation in mononuclear cells to produce GM-CSF in response to OKT3 mab. Loss of GM-CSF production was observed when either adherent cells or T lymphocytes were treated with emetine before co-culture with untreated cells of the other population in the presence of OKT3 mab. In contrast, mitomycin C reduced GM-CSF production significantly when T cells, but not adherent cells, were pretreated. These results suggest that T lymphocytes and adherent cells closely cooperate in the production of GM-CSF induced by OKT3 mab.
D000903:Antibiotics, Antineoplastic / Q000494:pharmacology; D000911:Antibodies, Monoclonal; D002478:Cells, Cultured; D003114:Colony-Forming Units Assay; D003115:Colony-Stimulating Factors / Q000096:biosynthesis*; D004640:Emetine / Q000494:pharmacology; D005434:Flow Cytometry; D006098:Granulocytes / Q000166:cytology*; D006801:Humans; D007371:Interferon-gamma / Q000096:biosynthesis*; D008222:Lymphokines / Q000032:analysis; D008264:Macrophages / Q000166:cytology*; D016685:Mitomycin; D008937:Mitomycins / Q000494:pharmacology; D009000:Monocytes / Q000276:immunology; D013601:T-Lymphocytes / Q000166:cytology / Q000187:drug effects / Q000276:immunology*
D000903:Antibiotics, Antineoplastic; D000911:Antibodies, Monoclonal; D003115:Colony-Stimulating Factors; D008222:Lymphokines; D008937:Mitomycins; D016685:Mitomycin; D007371:Interferon-gamma; D004640:Emetine