The novel cytokine interferon-gamma-inducing factor (IGIF) augments natural killer (NK) cell activity in cultures of human peripheral blood mononuclear cells (PBMC), similarly to the structurally unrelated cytokine interleukin (IL)-12. IGIF has been found to enhance the production of interferon-gamma (IFN-gamma) and granulocyte/macrophage colony-stimulating factor (GM-CSF) while inhibiting the production of IL-10 in concanavalin A (Con A)-stimulated PBMC. In this study, when anti-CD3 monoclonal antibody (mAb)-stimulated human enriched T cells were exposed to IGIF, the cytokine dose-dependently enhanced the proliferation of the cells and this could be completely inhibited by a neutralizing antibody against IL-2 at lower concentrations of IGIF. Neutralizing antibody against IFN-gamma had only insignificant inhibitory effects on T cell proliferation at higher concentrations of IGIF. Enzyme-linked immunosorbent assays (ELISA) revealed that, like PBMC, T cells exposed to IGIF produced large amounts of IFN-gamma; however, changes in the production of IL-4 and IL-10 were minimal. IGIF, but not IL-12, significantly enhanced IL-2 and GM-CSF production in T cell cultures, as determined by CTLL-2 bioassay and ELISA, respectively; however, both IGIF and IL-12 enhanced IFN-gamma production by the T cells. When T cells were exposed to a combination of IGIF and IL-12, a synergistic effect was observed on the production of IFN-gamma, but not on production of IL-2 and GM-CSF. In conclusion, IGIF enhances T cell proliferation apparently through an IL-2-dependent pathway and enhances Th1 cytokine production in vitro and exhibits synergism when combined with IL-12 in terms of enhanced IFN-gamma production but not IL-2 and GM-CSF production. Based on structural and functional differences from any known cytokines, it was recently proposed that this cytokine be designated interleukin-18.

Recently, human interleukin 18 (hIL-18) cDNA was cloned, and the recombinant protein with a tentatively assigned NH 2 -terminal amino acid sequence was generated. However, natural hIL-18 has not yet been isolated, and its cellular processing is therefore still unclear. To clarify this, we purified natural hIL-18 from the cytosolic extract of monocytic THP.1 cells. Natural hIL-18 exhibited a molecular mass of 18.2 kDa, and the NH 2 -terminal amino acid was Tyr 37 . Biological activities of the purified protein were identical to those of recombinant hIL-18 with respect to the enhancement of natural killer cell cytotoxicity and interferon-␥ production by human peripheral blood mononuclear cells. We also found two precursor hIL-18 (prohIL-18)-processing activities in the cytosol of THP.1 cells. These activities were blocked separately by the caspase inhibitors Ac-YVAD-CHO and Ac-DEVD-CHO. Further analyses of the partially purified enzymes revealed that one is caspase-1, which cleaves prohIL-18 at the Asp 36 -Tyr 37 site to generate the mature hIL-18, and the other is caspase-3, which cleaves both precursor and mature hIL-18 at Asp 71 -Ser 72 and Asp 76 -Asn 77 to generate biologically inactive products. These results suggest that the production and processing of natural hIL-18 are regulated by two processing enzymes, caspase-1 and caspase-3, in THP.1 cells. Interleukin (IL)1 -18 (originally called IGIF, interferon-␥-inducing factor) is a novel cytokine with multiple biological functions. In 1995 we purified murine IL-18 from the liver extracts of mice sensitized with Propionibacterium acnes followed by elicitation with lipopolysaccaride (1). The cDNA of murine IL-18 was cloned from cDNA libraries prepared from the livers of mice with endotoxin shock (2). Using this as a probe, human IL-18 cDNA was also cloned from a human normal liver cDNA library (3). The recombinant human IL-18 with a tentatively assigned NH 2 -terminal amino acid based on its homology with the natural murine IL-18 sequence was expressed in Escherichia coli, and its biological activities were examined (3).IL-18 has an interleukin 1 (IL-1) signature-like sequence (3) as reported and is similar to the IL-1 family and fibroblast growth factor in terms of their trefoil structures (4, 5). Despite their similarities, IL-18 and IL-1␤ exhibit different biological activities (2, 3, 6), transmitted through their specific receptors.2 Genetic information suggested that IL-18 is synthesized as an inactive precursor form (prohIL-18) and that this prohIL-18 has no known signal peptide sequence. Therefore, proteolytic cleavage is required for its maturation like IL-1␤ (2, 3, 7, 8). Gu et al. (7) reported that IL-1␤-converting enzyme (ICE)/ caspase-1 cleaved murine proIL-18 at the authentic processing site, Asp 35 -Asn 36 , to generate biologically active mature murine IL-18. However, natural hIL-18 had not yet been isolated, and its maturation site remained unclear.In this report, we screened for hIL-18 mRNA-expressing cell lines and purified natural hIL-18 from ...

-S433). According to a homology analysis, no amino acid sequence homology was observed between Cryj II and Cryj I, another major allergen. But Cryj II showed homology with polygalacturonase (PG) derived from tomato (40% identity) at the amino acid level. The sequence information can potentially be used to devise an effective course of immunotherapy for Japanese cedar pollinosis.

We have recently reported that a novel molecule, murine IFN-gamma-inducing factor (IGIF) produced by mouse liver cells, possesses potent biologic activities, including the induction of IFN-gamma production by spleen cells and the enhancement of NK cell cytotoxicity. In this paper, we report on the isolation of human IGIF cDNA clones from normal human liver cDNA libraries using murine IGIF cDNA as a probe. The amino acid sequence deduced from the human cDNA clones indicated a 193-amino acid precursor peptide and revealed 65% homology with that of murine IGIF. The amino acid sequence of IGIF also included an IL-1 signature-like sequence. Subsequently, the cloned cDNA was expressed in Escherichia coli, and preliminary studies on the biologic activities of the recombinant protein were performed. The recombinant human IGIF induced IFN-gamma production by mitogen-stimulated PBMC and enhanced NK cell cytotoxicity, in a manner similar to murine IGIF. In addition, recombinant human IGIF also augmented granulocyte-macrophage-CSF production and decreased IL-10 production, but had no effect on IL-4 production by Con A-stimulated PBMC. Based on these pleiotropic effects of IGIF, we propose that this novel cytokine be designated as IL-18.

The effects of interferon-gamma (IFN-gamma) on platelet recovery were examined in mice with marrow aplasia induced by i.p. injection of 250 mg/kg of 5-fluorouracil (5-FU). The cytokine was administrated by microosmotic pump, with an ability to deliver a consistent intact dose of cytokine for 7 consecutive days. Administration of 250 IU/kg/day of IFN-gamma in combination with 10(3) U/kg/day of IL-3, which alone had no effect on platelet counts, diminished the nadir for platelet count and shortened the duration of thrombocytopenia. The effect was comparable to that of higher doses of IL-3 (10(5) U/kg/day). The administration of 250 IU/kg/day of IFN-gamma in combination with 10(3) U/kg/day of IL-3 also induced megakaryocyte proliferation in bone marrow cell cultures. Single administration of either 250 IU/kg/day of IFN-gamma or 10(3) U/kg/day of IL-3 had no significant effects. The effect of this combination was also comparable to that of a higher dose of IL-3 (10(5) U/kg/day). We suggest that IFN-gamma accelerates megakaryocyte development, which leads to platelet production in chemotherapy-induced marrow aplasia. The administration of IFN-gamma in combination with IL-3 might be useful for the management of marrow aplasia.