Epsilon Open Archive

Abstract

This thesis describes the possible adjuvant effects of bacterial DNA, in the form of a DNA vaccine plasmid (pcDNA3) or oligodeoxyribonucleotides (ODN), in the pig. The effects were assessed mainly as their ability to induce production of immunoregulatory cytokines. Furthermore, the cells producing type I or type II interferon (IFN) at various stimuli were studied. The plasmid pcDNA3 was shown to induce a local production of IFN-α in vivo, in subcutaneous tissue chambers. The in vivo effects of plasmids, expressing the cDNA for the cytokines GM-CSF or IL-6, were also tested. Such cytokine-expressing vectors could have adjuvant effects in DNA vaccines. The cDNA for porcine IL-12 (poIL-12) fusion protein was cloned into pcDNA3. The plasmid was shown to express poIL-12 with high in vitro biological activity in a bioassay. IL-12 and huIL-18 synergized in the induction of IFN-γ production in porcine peripheral blood mononuclear cells (poPBMC), and the IFN-γ producing cells were by flow cytometric analyses shown to be T-cells and NK cells. The antigen-presenting dendritic cells (DC) are important target cells in vaccination. Monocyte-derived dendritic cells (moDC) were differentiated from adherent monocytes in the presence of GM-CSF and IL-4 in vitro, and were able to produce different cytokines depending on stimuli. IL-12 was produced by the moDC mainly at stimulation with pcDNA3 (preincubated in lipofectin), Actinobacillus pleuropneumoniae and Sendai virus. IFN-α was only produced in response to Sendai virus, and IL-6 and IL-10 mainly in response to A. pleuropneumoniae. The presence of an unmethylated CpG dinucleotide in ODN was essential for the in vitro induction of IFN-α in poPBMC. The presence of a poly-guanine sequence in the ends of the ODN was shown to enhance its IFN-α inducing capacity. The IFN-α producing cells in response to CpG-ODN, lipofected pcDNA3 or Aujeszky’s disease virus were by flow cytometry and in situ hybridisation shown to be a rare cell population, resembling the human plasmacytoid DC, also referred to as natural IFN producing cells. The results show that certain forms of bacterial DNA have adjuvant effects in the pig, which could be of importance in the development of more efficient vaccines.