Maturation of bovine dendritic cells by lipopeptides

The response of DC, and the subsequent stimulation of T cells, is an essential part of the initiation of immune responses following microbial challenge. The response of human DC to bacterial lipopeptides is mediated by toll-like receptor 2, and is characterised by DC maturation and the enhanced capacity to stimulate of T cells. We report here that bovine DC are also induced to mature following lipopeptide stimulation. Exposure of DC to the model lipopeptide Pam3CSK4 was associated with increased expression of MHC, costimulatory molecules, and enhanced secretion of IL-12 and TNFalpha. Lipopeptide-matured DC were superior in their ability to induce T cell activation and IFNgamma secretion. In contrast, exposure of MPhi to lipopeptides induced down-regulation of MHC expression and much lower increases in IL-12 secretion. A lipopeptide derived from the sequence of a relevant mycobacterial lipoprotein, MPB83, also influenced bovine DC by stimulating increases in IL-12 and TNFalpha secretion. These different changes in bovine DC and MPhi may have important implications for immune responses induced following bacterial infection with uptake of microbes by DC resulting in potentiation of their immunostimulatory capacity and uptake by MPhi having a much less marked effect on immune responses.     

Functional characterization of equine dendritic cells propagated ex vivo using recombinant human GM-CSF and recombinant equine IL-4

Naive T cells can be activated both in vivo and in vitro by specialized antigen presenting cells, dendritic cells (DC), with potent antigen-specific, immunostimulatory activity. Indeed, DC can provide an extremely powerful and important immunological tool by which to potentiate the immune response for specific recognition of foreign antigens. Until recently, the direct isolation of DC from PBMC required laborious procedures with extremely poor yields (<0.1%). Methods have been developed for the human, lower primate, and murine model systems to propagate large numbers of DC from PBMC or bone marrow ex vivo with various cytokines. However, all other model systems, including equine, still require the laborious isolation procedures to obtain DC. In this study, we have adapted the methods developed for the human system to generate large numbers of equine DC from PBMC precursors using recombinant human GM-CSF and recombinant equine IL-4. Our report is the first documentation of ex vivo generated DC from PBMC in a domesticated animal model system. Equine DC derived from PBMC were rigorously characterized by analyzing morphological, phenotypic, and functional properties and were determined to have similar attributes as DC generated from human PBMC. Equine DC appeared stellate with large projectiles and veils and had cell surface antigens at similar levels as those defined on human and murine DC. Furthermore, functional attributes of the DC included rapidly capturing antigens by pinocytosis, receptor-mediated endocytosis, and phagocytosis, activating naive T cells in a mixed leukocyte reaction to a much greater extent than macrophage or lymphoblasts, presenting soluble and particulate antigen 10-100 fold more effectively to T cells on a per cell basis than macrophage or lymphoblasts, and presenting soluble and particulate antigen to both CD4+ and CD8+ T cells. Taken together, our study provides a framework by which equine DC can now be readily produced from PBMC precursors and presents an impetus for and model by which DC can be simply generated in other animal model systems.     

Comparison of dendritic cell-mediated immune responses among canine malignant cells

Dendritic cell (DC) vaccination is one of the most attractive immunotherapies for malignancies in dogs. To examine the differences in DC-mediated immune responses from different types of malignancies in dogs, we vaccinated dogs using autologous DCs pulsed with keyhole limpet hemocyanin (KLH) and cell lysate prepared from squamous cell carcinoma SCC2/88 (SCC-KLH-DC), histiocytic sarcoma CHS-5 (CHS-KLH-DC), or B cell leukemia GL-1 (GL-KLH-DC) in vitro. In vivo inductions of immune responses against these tumor cells were compared by the delayed-type hypersensitivity (DTH) skin test. The DTH response against SCC2/88 cells were observed in dogs vaccinated with autologous SCC-KLH-DC, while the response was undetectable against CHS-5 and GL-1 cells in dogs vaccinated with autologous CHS-KLH-DC and GL-KLH-DC. Skin biopsies taken from DTH challenge sites were then examined for immunohistochemistry, and recruitment of CD8 and CD4 T cells was detected at the site where SCC2/88 cells were inoculated in dogs vaccinated with SCC-KLH-DC. By contrast, neither CD8 nor CD4 T cell infiltration was found at the DTH challenge site in the dogs vaccinated with CHS-KLH-DC or GL-KLH-DC. These findings may reflect that the efficacy of immune induction by DC vaccination varies among tumor types and that immune responses could be inducible in squamous cell carcinoma. Our results encouraged further investigation of therapeutic vaccination for dogs with advanced squamous cell carcinoma in clinical trials.     

Interaction of antigen presenting cells with mycobacteria

The interaction of mycobacteria with antigen presenting cells is a key feature in the pathogenesis of tuberculosis and the outcome of this interaction is pivotal in determining whether immunity or disease ensues. Human and mouse macrophages and dendritic cells (DC) have been shown to become infected with mycobacteria and to produce a response to infection that reflects their suggested role in immunity. Thus, macrophages elicit anti-microbial mechanisms for elimination of mycobacteria and DC up-regulate expression of molecules that aid their stimulation of T lymphocytes. We have examined the effects of infection with the avirulent strain Mycobacterium bovis BCG and with virulent M. bovis on bovine antigen presenting cells. Differences in the intracellular survival of bacteria within DC and macrophages were observed with higher numbers of bacteria maintained within DC following infection compared to macrophages. BCG was killed more effectively than M. bovis. Alterations in the expression of cell surface molecules involved in antigen presentation and the stimulation of T cells, including MHC II and CD40, were observed following infection of bovine antigen presenting cells. In addition infected DC secreted IL-12, TNF and IL-10 whereas macrophages produced TNF, IL-10 and little IL-12. Generally responses were more marked when virulent M. bovis was used compared to BCG. These studies indicate that infection of bovine antigen presenting cells by mycobacterial species results in the induction of both innate and adaptive immune responses that are critical for the outcome of infection.     

Porcine gammadelta T cells: possible roles on the innate and adaptive immune responses following virus infection

gammadelta T cells recognise different types of antigen in alternative ways to alphabeta T cells, and thus appear to play a complementary role in the immune response. However, unlike alphabeta T cells, the role or function of gammadelta T cells is still unclear. As pigs possess a high proportion of circulating gammadelta T cells, they are suitable large animal model to study gammadelta T cell functions. This as yet has not been fully exploited, leaving porcine gammadelta T cell biology and its role in immunity in its infancy. Foot-and-mouth disease (FMD) high potency "emergency" vaccines are able to induce early protection from challenge and it has been suggested that, in part, there is some involvement of innate immune responses. The antigen component of the vaccine is able to stimulate purified naive pig gammadelta T cells and induce the mRNA of various cytokines and chemokines. This observation suggests that gammadelta T cells probably contribute to the early phase of the immune responses to FMD vaccination, and perhaps infection. A subset of these circulating gammadelta T cells display a phenotype similar to professional antigen presenting cells and are able to take up and present soluble antigen to CD4(+) T cells in a direct cell-cell interaction via MHC class II. This direct interaction between gammadelta T cells and CD4(+) T cells is likely to have a significant influence on the out come of the adaptive immune response.     

Induction of dendritic cell-mediated immune responses against canine malignant melanoma cells

To establish the basis for the use of dendritic cells (DC) in the treatment of canine melanoma, dogs were vaccinated using autologous DC pulsed with canine melanoma CMM2 cell lysate in the presence of keyhole limpet haemocyanin (KLH) in vitro (CMM2-KLH-DC), and the induction of immune responses against CMM2 cells in vivo was examined using the delayed-type hypersensitivity (DTH) skin test. The DTH responses against CMM2 cells and KLH were observed in dogs vaccinated with CMM2-KLH-DC, while the responses against KLH but not CMM2 cells were detected with DC pulsed with KLH alone (KLH-DC). Recruitment of CD8 and CD4 T cells was detected in the positively responding sites, suggested that vaccination with CMM2-KLH-DC efficiently elicits T cell-mediated immunity against CMM-2 cells in vivo. These findings demonstrate the potential utility of DC-based tumour vaccination in the treatment of canine malignant melanoma.     

Cholera toxin promotes the generation of semi-mature porcine monocyte-derived dendritic cells that are unable to stimulate T cells

Cholera toxin (Ctx) is a powerful mucosal adjuvant with potential applications for oral vaccination of swine. Dendritic cells (DC) play a key role in the decision between immunity and tolerance, and are likely target cells for mediating Ctx functions in vivo. Therefore, we examined the capacity of Ctx to enhance stimulatory activity of porcine monocyte-derived DC (MoDC). Ctx promoted the development of a semi-mature DC phenotype, with decreased levels of MHC class II and CD40, but increased CD80/86 expression. These changes were associated with activation of extracellular signal-regulated kinase (ERK), but not NFkappaB or c-Jun N-terminal kinase (JNK). Functionally, Ctx-priming greatly diminished T cell stimulatory capacity both in antigen-specific and superantigen-induced proliferation assays. The lower proliferation rate was not due to increased apoptosis of either DC or T cells. Ctx suppressed TNFalpha secretion by MoDC, but induced IL-10 production. The observed effects on T cell proliferation could only be partially mimicked by IL-10 alone. However, addition of recombinant TNFalpha to co-cultures of Ctx-primed MoDC and lymphocytes restored lymphocyte proliferation in a concentration-dependent manner. Ctx-primed DC were not actively tolerogenic, since they could not suppress proliferative T cell reactions induced by untreated DC.     

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

Classical swine fever virus (CSFV) compromises the host immune system, causing indirect leucopoenia and disruption of in vitro T cell stimulation capacity. In order to explore the potential role of dendritic cells (DC) in such phenomena, the activation of conventional DC (cDC) and plasmacytoid DC (pDC) in blood and secondary lymphoid organs of infected pigs was investigated in the early time course post-inoculation (pi), together with viral components dissemination and cytokine production in serum. Whereas CD11R1+CD172a+ cDC frequencies were markedly reduced in blood and spleen, analysis of CD4+CD172a+ pDC numbers revealed a rapid turn-over of this DC subset in tissues pi. Both subsets matured and were activated after infection, as demonstrated by down-regulation of CD1a, up-regulation of the co-stimulation molecule CD80/86 and expression of cytokines. cDC essentially expressed tumor necrosis factor alpha (TNF-alpha) and interleukin (IL)-10, whereas pDC produced alpha interferon (IFN-alpha) and IL-12. IFN-alpha and TNF-alpha productions revealed an enhancement of innate anti-viral immune responses. Detection of antigen activated B lymphocytes in tonsil T-cell areas at 72 h pi, subsequently to the transient translocation of the viral E2 protein within germinal centres at 48 h pi, indicates the initiation of humoral response. This response was also evidenced by an important IL-10 production in serum one week pi. IL-12 expression in organs, as well as transient detection of IL-18 and IFN-gamma in serum, reflected the initiation of cellular immune responses. However, the uncommonly high levels of TNF-alpha and IFN-alpha produced by DC and measured in serum early post-infection, together with IL-10 expression in spleen, could play a role in the disruption of immune system cells, either inducing apoptosis or impairing DC functionalities themselves.     

Bovine NK cells acquire cytotoxic activity and produce IFN-gamma after stimulation by Mycobacterium bovis BCG- or Babesia bovis-exposed splenic dendritic cells

Early interactions of innate immune cell populations, such as dendritic cells (DC) and natural killer (NK) cells, can affect the ability of the acquired immune response to control infection of intracellular microorganisms. In this study, we investigated the activation of bovine NK cells by CD13(+) splenic DC stimulated with either Mycobacterium bovis BCG or Babesia bovis merozoites. Splenic DC were used either immediately after selection (cytokine(-)) or after exposure to GM-CSF, IL-4 and Flt3L for 72 h (cytokine(+)). Phenotypic analyses showed up-regulation of MHCII, CD80 and CD86 on cytokine(+) DC when compared to cytokine(-) DC. Purified NK cells (CD335(+)CD3(-)CD2(+/-)CD8alpha(+/-)) were co-cultured with microbial-exposed cytokine(-) DC or cytokine(+) DC in either transwell or cell-to-cell format and NK cell IFN-gamma production and cytotoxicity were assessed. NK cell IFN-gamma production was dependent on cell-to-cell contact. Microbial-stimulated cytokine(+) DC induced significantly more IFN-gamma production from NK cells than cytokine(-) cells. In contrast, cytotoxicity and perforin up-regulation were more pronounced in NK cells cultured with cytokine(-) DC than cytokine(+) DC. Therefore, activation of bovine NK cells by microbial-stimulated CD13(+) splenic DC is influenced by the maturation state of the DC suggesting different roles for the splenic DC during disease-induced maturation.     

Characterization of conventional and plasmacytoid dendritic cells in swine secondary lymphoid organs and blood

Dendritic cells (DCs) act as antigen presenting cells that bridge innate and adaptive immune systems with the unique capacity to initiate primary T-cell responses and efficiently stimulate memory responses. In pig, little information is available about these cells in secondary lymphoid organs, the place where T cell activation usually occurs. As increased knowledge on DC is a necessary prerequisite to further understand their role in response to microbial infection or in protection after vaccination, we investigated the DC types that would be present in tonsil, spleen and non-subcutaneous lymph nodes in the steady state. One population was composed of CD172a(+)CD11R1(+)CD1(+/-)CD80/86(+/-) cells and would correspond to conventional DCs (cDC), while the other one was composed of CD172a(+)CD4(+)CD1(+/-)CD80/86(+/-) cells and would correspond to plasmacytoid DCs (pDC). These subsets were also detected in blood but spleen was the tissue with the higher frequency of such DCs. In lymphoid organs, most of cDC and pDC were in an immature status, as revealed by the low percentage of cells expressing the co-stimulatory molecule CD80/86. However, expression of that marker by 5% of DCs in organs and up to 15% in blood, together with lower expression of CD1a and expression of CD208, would indicate a partial activation and/or semi-maturation. Interestingly, 8% of tonsil pDC and 15% of blood pDC were shown to secrete IFN-alpha, while 18-20% of cDC expressed TNF-alpha in these tissues. Both cell types also expressed IL-12 and IL-10 in the steady state. Measurements of IFN-alpha, TNF-alpha, IL-12 and IL-10 levels in serum confirmed their production within immune homeostasis, whereas IL-6, IL-18 and IFN-gamma could not be detected. Altogether, these data complete knowledge on porcine immune system cells and will be a useful tool for further in vivo studies on porcine DC role in peripheral tolerance induction and in immune responses to pathogens.     

Limited phenotypic and functional maturation of bovine monocyte-derived dendritic cells following Mycobacterium avium subspecies paratuberculosis infection in vitro

After encountering antigen, dendritic cells (DC) must differentiate into a fully mature phenotype to induce a protective, lasting T cell immunity. Paratuberculosis is a disease caused by the intracellular pathogen Mycobacterium avium subspecies paratuberculosis (M. paratuberculosis) and is characterized by a transient cell mediated immune response, that when dissipates correlates to the onset of clinical disease. In order to study the mechanism of early cellular immunity associated with M. paratuberculosis infection, we tested the hypothesis that M. paratuberculosis infected bovine DC have impaired activation and maturation thus are defective in the initiation of a sustainable and protective Th1 immune response locally. Our results demonstrate that M. paratuberculosis infected DC showed decreased endocytosis of ovalbumin, indicating some functional maturation. Co-stimulatory molecules CD40 and CD80 mRNA expression from M. paratuberculosis infected DC was increased over untreated immature DC. M. paratuberculosis infection induced chemokine receptor CCR7 increase in DC, yet CCR5 remained high. MHC II surface expression remained low on M. paratuberculosis infected DC. M. paratuberculosis infection inhibited pro-inflammatory cytokine IL-12 production and promoted IL-10 secretion by bovine DC. Together, our findings showed evidence of phenotypic and functional maturation of DC. However, we did not see the expected antigen presentation via MHC II and cytokine responses as a fully mature DC. This may suggest semi-mature DC phenotype induced by M. paratuberculosis infection.     

Species-specific properties and translational aspects of canine dendritic cells

Dogs are affected by spontaneously occurring neoplastic and inflammatory diseases which often share many similarities with pathological conditions in humans and are thus appreciated as important translational animal models. Dendritic cells (DCs) represent the most potent antigen presenting cell population. Besides their physiological function in the initiation of primary T cell responses and B cell immunity, a deregulation of DC function is involved in immune-mediated tissue damage, immunosuppression and transplantation complication in human and veterinary medicine. DCs represent a promising new target for cancer immunotherapy in dogs. However, the therapeutic use of canine DCs is restricted because of a lack of standardized isolation techniques and limited information about dog-specific properties of this cell type. This article reviews current protocols for the isolation and in vitro generation of canine monocyte- and bone marrow-derived DCs. DCs of dogs are characterized by unique morphological features, such as the presence of cytoplasmic projections and periodic microstructures. Canine DCs can be discriminated from other hematopoietic cells also based on phenotypic properties and their high T cell stimulatory capability in mixed leukocyte reactions. Furthermore, the classification of canine DC-derived neoplasms and the role of DCs in the pathogeneses of selected infectious, allergic and autoimmune diseases, which share similarities with human disorders, are discussed. Future research is needed to expand the existing knowledge about DC function in canine diseases as a prerequisite for the development of future therapies interfering with the immune response.     

Partial protection and intrathecal invasion of CD8(+) T cells in acute canine distemper virus infection

Initial non-inflammatory demyelination in canine distemper virus infection (CDV) develops against a background of severe immunosuppression and is therefore, thought to be virus-induced. However, recently we found a marked invasion of T cells throughout the central nervous system (CNS) in dogs with acute distemper despite drastic damage to the immune system. In the present study, this apparent paradox was further investigated by immunophenotyping of lymphocytes, following experimental CDV challenge in vaccinated and non-vaccinated dogs. In contrast to CDV infected, unprotected dogs, vaccinated dogs did not become immunosuppressed and exhibited a strong antiviral immune response following challenge with virulent CDV. In unprotected dogs rapid and drastic lymphopenia was initially due to depletion of T cells. In peripheral blood, CD4(+) T cells were more sensitive and depleted earlier and for a longer time than CD8(+) cells which recovered soon. In the cerebrospinal fluid (CSF) we could observe an increase in the T cell to B cell and CD8(+) to CD4(+) ratios. Thus, partial protection of the CD8(+) cell population could explain why part of the immune function in acute distemper is preserved. As found earlier, T cells invaded the CNS parenchyma in these dogs but also in the protected challenged dogs, which did not develop any CNS disease at all. Since markers of T cell activation were upregulated in both groups of animals, this phenomenon could in part be related to non-specific penetration of activated T cells through the blood brain barrier. However, in diseased animals much larger numbers of T cells were found in the CNS than in the protected dogs, suggesting that massive invasion of T cells in the brain requires CDV expression in the CNS.     

Target recognition by bovine mononuclear, MHC-unrestricted cytotoxic cells.

A population of bovine non B/non T, cytotoxic lymphocytes with natural killer activity against virus-infected and non-infected embryonic kidney cells was functionally characterized. The data obtained in experiments of flow cytometry and immuno-peroxidase staining show that a CD2-, CD4-, CD8-, TcR gamma delta-, CD3+, CD45+, FcR+ lymphoid killer cell does exist within bovine peripheral blood leucocytes. This population can detect the down-regulation of class I MHC antigens or the expression of embryonic forms thereof, as shown by experiments of 17-hour 51Cr release and binding to target cells. This model was tested in vitro in experiments on virus-infected bovine kidney cells. The emerging picture was substantially in agreement with the "missing self" theory as a possible option for target cell recognition. In this respect, the profound alteration of MHC Class I expression could represent a major early event, recognized on virus-infected cells by the immune system.     

New understanding of immunological mechanisms

The understanding and importance of antigen-specific immune responses after vaccination has completely changed in recent years. In the past, the focus for monitoring a vaccine-specific immune reaction was principally on the humoral branch of the immune system. The efficacy of vaccines, as assessed by the induction of protective immunity was mainly correlated with antibodies and antibody-titers. However, this correlation often failed and other parts of the immune system had also to be considered: namely, the innate immune system and the cellular branch of the antigen-specific immune system. With regard to vaccines, the innate immune system plays its main role in the effective activation of the antigen-specific immune response, in antigen-uptake and antigen-presentation. The dendritic cells (DCs) are the most important antigen presenting cells which present processed protein antigens (peptides) through MHC-molecules: MHC-class I, for the presentation of endogenous synthesised antigen; MHC-class II for exogenous antigen. Activation of DC leads to an enhanced production of cytokines and chemokines, to an up-regulation of co-stimulatory and activation molecules and also molecules for cell-cell interactions, e.g. interactions with cells of the antigen-specific immune system. T lymphocytes are the effector cells of the cellular branch of the antigen-specific immune system. They act either as MHC-class I-restricted cytolytic T lymphocytes (CTL) or as MHC-class II-restricted T-helper cells providing support for B lymphocytes (T(H)2) and the cellular part of the antigen-specific immune system (T(H)1). In order to achieve effective vaccination, the activation of all T-cell subpopulations is of advantage, but more important is the generation of antigen-specific memory T and B lymphocytes. In addition to these 'generic' immunological factors which are essential for the design of more efficacious vaccines, our detailed knowledge about feline and canine immune reactions after vaccination, which is still poor, has to be improved.     

Suppression of canine myeloid cells by soluble factors from cultured canine tumor cells

BackgroundCancer profoundly affects immunity and causes immunosuppression that contributes to tumor escape, metastases and resistance to therapy. The mechanisms by which cancer cells influence immune cells are not fully known but both innate and adaptive immune cells can be altered by cancer. Myeloid cells are innate immune cells that comprise the mononuclear phagocytic system (MPS) and include monocytes, macrophages, dendritic cells (DCs) and their progenitors. Myeloid cells play important roles in both the promotion and regulation of immune responses. Dysregulated myeloid cells are increasingly being recognized as contributing to cancer-related immunosuppression. This study investigated whether soluble factors produced by canine tumor cells inhibited canine myeloid cell function.MethodsThese studies investigated the utility of using the canine DH82 cell line for assessment of canine myeloid responses to tumor-derived soluble factors (TDSFs). Phenotypic comparisons to canine bone marrow-derived DCs (BM-DCs) and bone marrow-derived macrophages (BM-MΦs) were performed and expression of myeloid cell markers CD11b, CD11c, CD80, and major histocompatibility complex (MHC) class II were evaluated by flow cytometry. Phenotypic and functional changes of DC populations were then determined following exposure to tumor-conditioned media (TCM) from canine osteosarcoma, melanoma and mammary carcinoma cell lines.ResultsWe found that the canine BM-DCs and the DH82 cell line shared similar CD11b, CD11c and MHC II expression and morphologic characteristics that were distinct from canine BM-MΦs. Myeloid cells exposed to TDSFs showed decreased expression of MHC class II and CD80, had reduced phagocytic activity and suppressed the proliferation of responder immune cells.ConclusionThese results show that soluble factors secreted from canine tumor cells suppress the activation and function of canine myeloid cells. Our results suggest that, similar to humans, dysregulated myeloid cells may contribute to immunosuppression in dogs with cancer.     

Comparative analysis of canine monocyte- and bone-marrow-derived dendritic cells

Dendritic cells (DC) represent a heterogeneous cell family of major importance for innate immune responses against pathogens and antigen presentation during infection, cancer, allergy and autoimmunity. The aim of the present study was to characterize canine DC generated in vitro with respect to their phenotype, responsiveness to toll-like receptor (TLR) ligands and T-cell stimulatory capacity. DC were derived from monocytes (MoDC) and from bone marrow hematopoietic cells cultured with either Flt3-ligand (FL-BMDC) or with GM-CSF (GM-BMDC). All three methods generated cells with typical DC morphology that expressed CD1c, CD11c and CD14, similar to macrophages. However, CD40 was only found on DC, CD206 on MΦ and BMDC, but not on monocytes and MoDC. CD1c was not found on monocytes but on all in vitro differentiated cells. FL-BMDC and GM-BMDC were partially positive for CD4 and CD8. CD45RA was expressed on a subset of FL-BMDC but not on MoDC and GM-BMDC. MoDC and FL-DC responded well to TLR ligands including poly-IC (TLR2), Pam3Cys (TLR3), LPS (TLR4) and imiquimod (TLR7) by up-regulating MHC II and CD86. The generated DC and MΦ showed a stimulatory capacity for lymphocytes, which increased upon maturation with LPS. Taken together, our results are the basis for further characterization of canine DC subsets with respect to their role in inflammation and immune responses.     

Antigen presenting cells in mucosal sites of veterinary species

The ability of antigen presenting cells, in particular dendritic cells, to integrate a variety of environmental signals, together with their ability to respond appropriately by initiating either tolerance or defensive immune responses make them cells of particular relevance and importance in the mucosal environment. They have been demonstrated in a variety of mucosal tissues in veterinary species and have been characterized to varying degrees, showing that fundamental immunological principles apply throughout all species, but also highlighting some species differences. A major advantage of carrying out immunological research in veterinary species is their size: it is possible to cannulate lymphatic ducts and obtain information about cell migration between different tissues. It is also possible to obtain pure populations of relatively rare cell types such as the plasmacytoid dendritic cells or mucosal dendritic cells ex vivo for the study of immune responses to diseases in their natural host and for other thorough functional studies. Two major myeloid antigen presenting cell (APC) (dendritic cells, DC) cell populations have been described in gut draining lymph and other mucosal sites in ruminants and pigs, characterised by the presence or absence of surface molecules, their enzyme profiles, their ability to phagocytose and their different potential as APC. There is evidence that one of these subsets has migrated from the diffuse mucosal tissue, where it is found as a phagocytic as well as stimulatory APC population, which in turn may be derived from blood macrophages. In addition, the presence and role in viral infection of the IFN-alpha producing plasmacytoid DC in mucosal tissue is discussed, based on studies in pigs.