Identification of IFN-γ-Producing Cells in IL-12/IL-18-Treated Mice

Both IL-12 and IL-18 have been characterized as effective IFN-γ-inducing cytokines. Concomitant treatment with IL-12 and IL-18 has been shown to synergistically induce IFN-γ and may be an effective therapy for treating cancer, allergy, and infectious diseases. To understand the mechanisms underlying the strong induction of IFN-γ by IL-12/IL-18 in mice, we focused our studies on the IFN-γ-producing cells in various lymphoid organs and tissues and utilized the intracellular cytokine staining method to detect such cells in situ. After combined treatment with IL-12 and IL-18, IFN-γ-positive cells in C57BL/6 mice were detected in the liver (12.18%), spleen (0.68%), bone marrow (1.80%), and peritoneum (2.12%), but not in the thymus or lymph nodes (<0.05 and <0.08%, respectively). A two-color staining method revealed that the majority of IFN-γ-producing cells in the liver were NK1.1+ cells, while those in the spleen were mostly CD3+ cells, and to a lesser degree NK1.1+ cells. Both CD4+ and CD8+ cells in the liver and in the spleen produced IFN-γ. The CD19+ B cell population was not definitely shown to produce IFN-γ in our induction experiments. NKT cells, which are a subpopulation of NK1.1+ CD3+ cells, were diminished in the liver and did not seem to contribute to IFN-γ production arising from IL-12/IL-18 treatment. Further in vitro experiments confirmed the responsiveness of hepatic mononuclear cells to IL-12/IL-18 stimulation. This study is the first to show the IFN-γ-producing mechanisms of IL-12/IL-18 treatment at the phenotypic level.