Type‐A CpG oligonucleotides activate exclusively porcine natural interferon‐producing cells to secrete interferon‐α, tumour necrosis factor‐α and interleukin‐12

Abstract

Natural interferon-producing cells (NIPC), also referred to as immature plasmacytoid dendritic cells (PDC), constitute a small population of leucocytes secreting high levels of type I interferons in response to certain danger signals. Amongst these signals are those from DNA containing unmethylated CpG motifs. The present work demonstrated that the CpG oligonucleotides (CpG-ODN) 2216, D32 and D19 induce high amounts of interferon-alpha (IFN-alpha), tumour-necrosis factor-alpha (TNF-alpha) and interleukin (IL)-12 in porcine peripheral blood mononuclear cells (PBMCs). Swine workshop cluster 3 (SWC3)1ow CD4high cells, with high IL-3-binding activity, representing NIPC, were the exclusive cytokine-producing cells responding to the CpG-ODN. These cells did not express CD6, CD8 or CD45RA. Importantly, monocyte-derived DC did not respond to CpG-ODN by secretion of IFN-alpha or TNF-alpha or by the up-regulation of costimulatory molecule expression. CpG-ODN up-regulated MHC class II and CD80\86 expression on the NIPC, but were unable to promote NIPC survival. Interestingly, certain CpG-ODN, incapable of inducing NIPC to secrete IFN-alpha or up-regulate MHC class II and CD80\86, did promote NIPC viability. Taken together, the influence of CpG-ODN on porcine NIPC, monocytes and myeloid DCs relates to that observed with their human equivalents. These results represent an important basis for the application of CpG-ODN as adjuvants for the formulation of novel vaccines and demonstrate the importance of the pig as an alternative animal model for this approach.

Classical swine fever virus induces activation of plasmacytoid and conventional dendritic cells in tonsil, blood, and spleen of infected pigs

“…These results were in agreement with cytokine production by in vitro CSFV-infected NIPC [4,42]. Moreover, porcine blood-isolated NIPC secreted IFN-α, TNF-α and IL-12, but not IL-10, after CpGoligonucleotide stimulation [20].…”

Regulation of cytokine expression in human plasmacytoid dendritic cells by prostaglandin I2 analogues

“…In all cases, the purity of isolated pDCs was .92% (see online supplementary material), as judged by their expression of CD123 but not CD14 markers on their surface by flow cytometry (data not shown). Purified pDCs (2610 5 cells?condition -1 ) were treated with varying doses of IP receptor agonists, iloprost or treprostinil, or a cAMP activator, forskolin, for 2 h and then stimulated with 10 mg?mL -1 of CpG oligodeoxynucleotide (ODN)-2216 or imiquimod as previously described [11,12], for 48 or 72 h. For analysis of IFN-a, pDCs were pre-treated with IL-3 (10 ng?mL -1 ) [13] for 30 min prior to the treatment with iloprost, followed by the stimulation of the cells with CpG or imiquimod. To examine the involvement of the iloprost-IP receptor axis, pDCs were pre-treated with an IP receptor antagonist, CAY10449, 1 h before the treatment of the cells with iloprost.…”

Porcine circovirus type 2-induced interleukin-10 modulates recall antigen responses

“…The adherent PBMC fraction was separated from the non-adherent subpopulation by adhesion to plastic cell culture flasks. The SwC3 + fraction of PBMCs (namely monocytes, macrophages and DCs) was purified from healthy pigs using an anti-SwC3 porcine pan-myeloid cell marker (CD172; monoclonal antibody BL1H7) and magnetic sorting (MACS; Miltenyi Biotech) as described previously (Guzylack-Piriou et al, 2004). Purity achieved for that cell subpopulation was ¢95 % as determined by flow cytometry.…”

“…The reagents for IFN-c, IL-10 and IL-2 ELISAs were from Biosource and for IL-12 from R&D Systems. Anti-IFN-a monoclonal antibodies (K9 and K17) and an IFN-a recombinant protein (PBL Biomedical laboratories) were used in ELISA as described previously (Guzylack-Piriou et al, 2004). The role of IL-10 was assessed by adding a neutralizing IL-10 antibody (2 mg ml 21 , clone 148801; R&D systems).…”