SUMMARYDespite the central role that dendritic cells (DC) play in immune regulation and antigen presentation, little is known about porcine DC. In this study, two sources of DC were employed. Bone marrow haematopoietic cell-derived DC (BM-DC) were generated using granulocyte± macrophage colony-stimulating factor (GM-CSF) in the presence or absence of tumour necrosis factor-a (TNF-a). Monocyte-derived DC (Mo-DC) were generated with GM-CSF and interleukin-4 (IL-4). In both systems, non-adherent cells developed with dendritic morphology, expressing high levels of major histocompatibility complex (MHC) class II. The presence of TNF-a increased the BM-DC yield, and enhanced T-cell stimulatory capacity. Both BM-DC and Mo-DC expressed the pan-myeloid marker SWC3, as well as CD1 and CD80/86, but were also CD14 + and CD16 + . The CD16 molecule was functional, acting as a low-af®nity Fc receptor. In contrast, the CD14 on DC appeared to differ functionally from monocyte CD14: attempts to block CD14, in terms of lipopolysaccharide (LPS)-induced procoagulant activity (PCA), failed. The use of TNF-a or LPS for DC maturation induced up-regulation of MHC class II and/or CD80/86, but also CD14. Allogeneic mixed leucocyte reactions and staphylococcal enterotoxin B antigen presentation assays demonstrated that these DC possessed potent T-cell stimulatory capacity. No T helper cell polarization was noted. Both the BM-DC and the Mo-DC induced a strong interferon-c and IL-4 response. Taken together, porcine DC generated in vitro possess certain characteristics relating them to DC from other species including humans, but the continued presence of CD14 and CD16 on mature and immature porcine DC was a notable difference.

Peripheral blood contains two major particular infrequent dendritic cells (DC) subsets linking the innate and specific immune system, the myeloid DC and plasmacytoid DC equivalent to the natural interferon-producing cells (NIPC). The functional characterization of these cells demands large volumes of blood, making a large animal model more appropriate and beneficial for certain studies. Here, two subsets of porcine blood mononuclear cells expressing swine workshop cluster 3 (SWC3, a SIRP family member), are described and compared to monocytes. The blood DC specialized in T-cell stimulation were major histocompatibility complex (MHC) class II+, CD80/86+, CD1+/-, CD4-, and in contrast to monocytes CD14-. A CD16- and a CD16+ subset could be discriminated. Granulocyte-macrophage colony-stimulating factor and interleukin-3 were survival factors for this DC subset, and culture induced an up-regulation of MHC class II and CD80/86. The second subset described, are porcine NIPC, typically CD4++, MHC class IIlow, CD80/86low, CD1-, CD8-/low, CD16-/low and CD45RA-/low. Porcine NIPC had high interleukin-3 binding capacity, and survived in response to this cytokine. Their unique function was strong interferon type I secretion after virus stimulation. Both subsets were endocytically active when freshly isolated, and down-regulated this activity after in vitro maturation. Taken together, the present report has delineated porcine blood DC and NIPC, permitting a more detailed understanding of innate immune defences, particularly in response to infections.

Dendritic cells (DCs) play crucial roles in innate and adaptive immune responses, rendering them critical targets for virus infections. Porcine circovirus type 2 (PCV2) is associated with the development of postweaning multisystemic wasting syndrome (PMWS) in piglets. We demonstrate here that 80 to 90% of monocyte-derived and bone marrow-derived DCs interact with PCV2 similar to the early stages of an infection. There was no evidence for virus replication, but the virus did persist in DCs without loss of infectivity nor the induction of cell death. This could reflect an abortive infection, but there was no evidence of virus uncoating-the infectivity remained intact for at least 5 days. Alternatively, the results may reflect DC endocytosis of antigenic material. However, there was no modulation of DC surface major histocompatibility complex class I and class II, CD80/86, CD25, CD16, or CD14. Furthermore, infected DC did not transmit virus to syngeneic T lymphocytes, even when the latter were activated. Such coculture did not induce PCV2 replication or death of the lymphocytes or DCs. These results demonstrate that PCV2 can persist in DCs in the absence of virus replication or degradation. Such a silent virus infection presents a novel mechanism of not only immune evasion but also escaping the DC degradation pathway. Because of their migratory capacity, infection of DCs thus provides a potent vehicle for transport of the virus throughout the host without the need for replication. In addition, the lymphopenia seen in PMWS is not a direct effect of the virus on lymphocytes but would require additional events, as proposed by others.

Functional disruption of dendritic cells (DCs) is an important strategy for viral pathogens to evade host defences. Monocytotropic viruses such as classical swine fever virus (CSFV) could employ such a mechanism, since the virus can suppress immune responses and induce apoptosis without infecting lymphocytes. Here, CSFV was shown to infect and efficiently replicate in monocyte-and in bone marrow-derived DCs. Interestingly, the infected DCs displayed neither modulated MHC nor CD80/86 expression. Stimulation of DCs with IFN-a/TNF-a or polyinosinic-polycytidylic acid (pIC) induced phenotypic maturation with increased MHC and CD80/86 expression, both with mock-treated and infected DCs. In addition, the T cell stimulatory capacity of CSFV-infected DCs was maintained both in a polyclonal T cell stimulation and in specific antigen-presentation assays, requiring antigen uptake and processing. Interestingly, similar to macrophages, CSFV did not induce IFN-a responses in these DCs and even suppressed pIC-induced IFN-a induction. Other cytokines including interleukin (IL)-6, IL-10, IL-12 and TNF-a were not modulated. Taken together, these results demonstrated that CSFV can replicate in DCs and control IFN type I responses, without interfering with the immune reactivity. These results are interesting considering that DC infection with RNA viruses usually results in DC activation. INTRODUCTIONClassical swine fever (CSF) is a highly contagious disease of pigs caused by CSF virus (CSFV) and leads to important economic losses worldwide. CSFV together with bovine viral diarrhoea virus (BVDV) and border disease virus (BDV) form the genus Pestivirus within the family Flaviviridae.CSFV is a monocytotropic viral pathogen, which can efficiently evade and compromise the host's immune system. The virus has a high affinity for reticulo-endothelial cells (Cheville & Mengeling, 1969;Ressang, 1973;Susa et al., 1992) causing lymphopenia, thrombocytopenia, coagulation disorders and atrophy of the thymus and bone marrow Pauly et al., 1998; Sanchez-Cordon et al., 2002;Summerfield et al., 2000Summerfield et al., , 2001. Lymphopenia is caused, at least in part, by apoptosis detectable in uninfected lymphocytes Summerfield et al., 1998b). In addition, viable lymphocytes isolated from CSFV-infected pigs do not respond to mitogen stimulation (Pauly et al., 1998;Summerfield et al., 1998b;Van Oirschot et al., 1983). These modulated cells are not infected. Instead, it is the myeloid population, particularly monocytes (Mo) and macrophages (Mw), that contains the early target cell for infection and replication, both in vivo (Ressang, 1973;Gomez-Villamandos et al., 2001;Sanchez-Cordon et al., 2003;Summerfield et al., 2000;Trautwein, 1988) and in vitro (Knoetig et al., 1999). Despite this clear targeting and tropism, no direct evidence has been found of a role for infected Mo and Mw in the observed immunosuppression and death of T lymphocytes (Knoetig et al., 1999).Dendritic cells (DCs) are one of the primary immunological sentinels of the immune system ...

Summary Viral interactions with dendritic cells (DCs) have important consequences for immune defence function. Certain single‐stranded DNA viruses that associate with a number of species, including humans and pigs, exhibit interesting characteristics in this context. Porcine circovirus type 2 (PCV2) can persist within myeloid DCs in the absence of virus replication. Internalization was observed with both conventional blood DCs and plasmacytoid DCs [natural interferon‐producing cells (NIPCs)], as well as DC precursors. This PCV2–DC interaction neither induced nor inhibited DC differentiation. The maturation of myeloid DCs induced by a cocktail of interferon‐α/tumour necrosis factor‐α (IFN‐α/TNF‐α), and the ability to process and present antigen to T lymphocytes, remained intact in the presence of PCV2. The virus was clearly internalized by the DCs, a process noted with both mature and immature cells. This suggested a non‐macropinocytic uptake, confirmed by an insensitivity to wortmannin but sensitivity to cytochalasin D, chlorpromazine and bafilomycin. Nevertheless, PCV2 was immunomodulatory, being effected through the reaction of NIPC to danger signals. When NIPCs responded to the CpG‐oligonucleotide (CpG‐ODN), their costimulatory function which induces myeloid DC maturation was clearly impaired by the presence of PCV2. This was caused by a PCV2‐induced inhibition of the IFN‐α and TNF‐α normally produced following interaction with CpG‐ODN. Thus, the immunomodulatory activity of PCV2 is mediated through the disruption of NIPC function. This would impair the maturation of associated myeloid DC and have major implications for the efficient recognition of viral and bacterial danger signals, favouring the establishment of infections additional to that of PCV2.