IFN‐γ , a product of NK and T cells, is a key cytokine contributing innate and adaptive immunity. IFN‐γ  production is induced via direct cell‐cell contacts with APC and IFN‐γ ‐producing cells or by cytokines. During microbial infections macrophage‐derived IFN‐α , IL‐12, and IL‐18 enhance IFN‐γ  production and Th1 response. Here we show that IFN‐α  in combination with IL‐18 very efficiently induces IFN‐γ  expression also in primary, nonactivated NK cells and in NK‐92 cell line. Comparison of the kinetics of IFN‐γ  mRNA expression in nonactivated NK cells, NK‐92 cells and activated T cells stimulated with IFN‐α  or IL‐12 revealed that, although both of these cytokines directly up‐regulate IFN‐γ  mRNA expression, its levels remain elevated much longer with IL‐12 stimulation. In both NK cells and T cells, Stat4 is known to be critical in IL‐12 and IFN‐α  signaling. We show that Stat4 activation is transient in cells stimulated with IFN‐α , whereas IL‐12 induces more long‐lasting activation of the transcription factor. This prolonged activation of IFN‐γ  gene by IL‐12 may result in more efficient IFN‐γ  production compared to that of IFN‐α . Our results demonstrate that IFN‐α  and IL‐18 are important innate cytokines in inducing NK cell IFN‐γ  production.

SUMMARY The lipophilic toxin, cereulide, emitted by emetic food poisoning causing strains of Bacillus cereus, is a powerful mitochondria toxin. It is highly lipophilic and rapidly absorbed from the gut into the bloodstream. We tested how this toxin influences natural killer (NK) cells, which are important effectors in defence against infections and malignancy. Cereulide inhibited cytotoxicity and cytokine production of natural killer cells, caused swelling of natural killer cell mitochondria, and eventually induced natural killer cell apoptosis. The suppressive effect on cytotoxicity was fast and toxic concentration low, 20–30 μg/l. As the emesis causing concentration of cereulide is around 10 μg/kg of total body mass, our results suggest that emesis causing or even lower doses of cereulide may also have a systemic natural killer cell suppressive effect.

Streptomyces griseus strains isolated from indoor dust have been shown to synthesize valinomycin. In this report, we show that human peripheral blood lymphocytes treated with small doses (30 ng ml ؊1 ) of pure valinomycin or high-pressure liquid chromatography-pure valinomycin from S. griseus quickly show mitochondrial swelling and reduced NK cell activity. Larger doses (>100 ng/ml ؊1 ) induced NK cell apoptosis within 2 days. Within 2 h, the toxin at 100 ng ml ؊1 dramatically inhibited interleukin-15 (IL-15)-and IL-18-induced granulocyte-macrophage colony-stimulating factor and gamma interferon (IFN-␥) production by NK cells. However, IFN-␥ production induced by a combination of IL-15 and IL-18 was somewhat less sensitive to valinomycin, suggesting a protective effect of the cytokine combination against valinomycin. Thus, valinomycin in very small doses may profoundly alter the immune response by reducing NK cell cytotoxicity and cytokine production.Human peripheral blood lymphocytes (PBL) contain 5 to 20% NK cells, which can lyse certain tumor and normal cell lines without prior immunization. Upon activation, NK cells secrete cytokines such as gamma interferon (IFN-␥), tumor necrosis factor alpha, granulocyte-macrophage colony-stimulating factor (GM-CSF), and macrophage colony-stimulating factor (7,15,24). NK cells are thought to represent the first line defense in the immune system since they kill abnormal cells and simultaneously secrete cytokines to activate the other arms of the immune response.Interleukin-15 (IL-15) and IL-18 are cytokines which regulate NK cell function. IL-15 is required for NK cell maturation, and IL-18 is essential for NK cell activity (16,29). Valinomycin is an ionophore which is a cyclic-polypeptide-like dodecadepsipeptide whose folded conformation forms an inner cavity that can accommodate K ϩ but not other ions. For that reason, valinomycin is involved in a selective transport of K ϩ ions across the inner membrane of mitochondria (11,12). In rat ascites hepatoma cells, valinomycin induces apoptosis by disrupting the membrane potential of mitochondria and, when used at higher concentrations, causes apoptosis in many mammalian cell types (1, 10, 14, 21, 34). Andersson et al. (3) isolated Streptomyces griseus strains that produce valinomycin from an indoor environment. In schools and children's day care centers with dampness damage, such Streptomyces strains were frequently encountered (26).Here we report that pure commercial valinomycin and highpressure liquid chromatography (HPLC)-pure valinomycin from S. griseus inhibit human NK activity and cytokine production and induce apoptosis of NK cells at doses 10 to 500 times lower than those previously used. Thus, a toxin derived from bacteria that are abundant in the environment has the potential to cause immune suppression. MATERIALS AND METHODSCell isolation and culture. Leukocyte-rich buffy coats were obtained from healthy blood donors (Finnish Red Cross Blood Transfusion Service, Helsinki, Finland). PBL were isolated by Ficoll-Paque...

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IFN-gamma, a product of NK and T cells, is a key cytokine contributing innate and adaptive immunity. IFN-gamma production is induced via direct cell-cell contacts with APC and IFN-gamma -producing cells or by cytokines. During microbial infections macrophage-derived IFN-alpha, IL-12, and IL-18 enhance IFN-gamma production and Th1 response. Here we show that IFN-alpha in combination with IL-18 very efficiently induces IFN-gamma expression also in primary, nonactivated NK cells and in NK-92 cell line. Comparison of the kinetics of IFN-gamma mRNA expression in nonactivated NK cells, NK-92 cells and activated T cells stimulated with IFN-alpha or IL-12 revealed that, although both of these cytokines directly up-regulate IFN-gamma mRNA expression, its levels remain elevated much longer with IL-12 stimulation. In both NK cells and T cells, Stat4 is known to be critical in IL-12 and IFN-alpha signaling. We show that Stat4 activation is transient in cells stimulated with IFN-alpha, whereas IL-12 induces more long-lasting activation of the transcription factor. This prolonged activation of IFN-gamma gene by IL-12 may result in more efficient IFN-gamma production compared to that of IFN-alpha. Our results demonstrate that IFN-alpha and IL-18 are important innate cytokines in inducing NK cell IFN-gamma production.