Analysis of interferon-gamma production in killed BCG-pretreated mice after stimulation with staphylococcal enterotoxin A

Abstract

Staphylococcal enterotoxin A (SEA), a T cell mitogen, was found to induce a high level of interferon-gamma (IFN-gamma) in mice which had been immunized with killed Bacillus Calmette-Guérin (BCG) in water-in-oil-in-water (W/O/W) emulsion. The phenomenon was analysed by in vivo and in vitro experiments, and the following results were obtained. 1. The SEA-induced IFN was inactivated by treatment with 0.2M glycine-HCl (pH 2.0) but not by heating at 56 degrees C for 30 min. nor by treatment with anti-IFN alpha/beta antibodies, and the fact suggest that the IFN belonged to the gamma type. 2. Treatment of the BCG-sensitized mice with silica and 2-chloroadenosine (2CA), specific lethal agents for macrophages, reduced the SEA-induced IFN production. 3. The SEA-induced IFN production occurred in mice immunized with BCG either intravenously or intraperitoneally, although they showed weak or no footpad reaction to purified protein derivatives (PPD). In contrast, mice sensitized subcutaneously with BCG showed strong foodpad reaction to PPD but not the SEA-induced IFN production. Thus, the mere presence of BCG-sensitized T cells does not appear to result in the SEA-induced IFN production. 4. In vitro experiments, in which SEA-induced IFN production was determined in the culture of BCG-sensitized spleen cells, showed that principal IFN-producing cells were Lyt-1+ T cells. 5. Deprivation of macrophages from BCG-sensitized spleen cell population by passing through Sephadex G-10 column reduced the SEA-induced IFN production in the culture. Addition of 2CA to the culture medium also reduced the SEA-induced IFN production by the BCG-sensitized spleen cells. 6. The SEA-induced IFN production in the culture of the BCG-sensitized spleen cells was suppressed in the presence of lipoxygenase inhibitor, i.e., caffeic acid or nordihydroguaiaretic acid. 7. The plastic adherent spleen cells (i.e. macrophages) from mice sensitized with BCG produced leukotriene C4 (LTC4). The suppression of the SEA-induced IFN production of BCG-sensitized spleen cells in the presence of the lipoxygenase inhibitor was prevented by addition of synthetic LTC4. These results suggest that LTC4 released from macrophages activated by BCG causes production of IFN-gamma by BCG-sensitized T cells responding to SEA.