Porcine interleukin-12 fusion protein and interleukin-18 in combination induce interferon-γ production in porcine natural killer and T cells

A pcDNA3 vector containing a gene encoding a porcine interleukin-12 (poIL-12) fusion protein was constructed, with the p40 chain and its signal peptide positioned first, followed by a linker and the p35 domain. When expressed in COS cells, secreted poIL-12 fusion protein showed high activity in terms of ability to induce interferon-gamma (IFN-gamma) production in porcine peripheral blood mononuclear cells (PBMCs) in vitro. The IFN-gamma production induced by poIL-12 fusion protein, as well as heterodimeric poIL-12 and human IL-12, was markedly dependent on the presence of human IL-18 (huIL-18). Furthermore, huIL-18 showed a dose-dependent induction of IFN-gamma production in PBMC in the presence of a constant concentration of huIL-12. A marked synergism between poIL-12 and IL-18 was consequently observed in poPBMC. The actual IFN-gamma producing cells were identified as probable NK cells (about 30%) and T lymphocytes (about 70%), using flow cytometry. Furthermore, a histidine-tagged poIL-12 fusion protein was expressed in Drosophila melanogaster Schneider 2 cells, using a modified pMT/V5-His vector lacking the V5 epitope, Such poIL-12 fusion protein was easily purified using Ni-NTA agarose and retained high biological activity. (C) 2002 Elsevier Science B.V. All rights reserved.