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A cDNA clone encoding a 380 a-a type 1 transmembrane protein with homology to human Siglec-3/CD33 was obtained from a swine small intestine library. An analysis of protein sequence identified two immunoglobulin-like domains, a transmembrane region, and a carboxi-terminal tail with two tyrosine-based signalling motifs. Binding assays of Siglec-3 transfected CHO cells to polyacrylamide glycoconjugates showed a preference for α2–6-linked sialic acids. Using mAbs raised against a fragment containing the two Ig-like domains, porcine Siglec-3 was found to be expressed on monocytes and granulocytes, and their bone marrow precursors. It was also detected in lymph node, splenic and alveolar macrophages. MAbs immunoprecipitated, from granulocyte lysates, a protein of 51–60 kDa under both non-reducing and reducing conditions. MAbs were also used to analyse functional activity of Siglec-3 on bone marrow and blood cells. Engagement of Siglec-3 by mAb had no apparent effect on cell proliferation or cytokine production. [ABSTRACT FROM AUTHOR]

Siglecs are sialic acid binding Ig-like proteins involved in the control of leukocyte responses. In this study we describe the characterization of a porcine orthologue of Siglec-10. A cDNA clone was obtained from a porcine library which encodes a protein with sequence homology to human Siglec-10. This cDNA codes for a type I transmembrane protein containing four Ig-like domains, a transmembrane region, and a cytoplasmic tail with three tyrosine-based motifs, including a membrane-proximal Grb2-binding motif, and two ITIM motifs. When expressed on transfected cells, porcine Siglec-10 was able to bind red blood cells in a sialic acid-dependent manner. Monoclonal antibodies were developed against this protein and used to examine its cell and tissue distribution in the pig. Siglec-10 was found to be expressed on blood B cells and B cell areas of the spleen and lymph nodes. A weak expression was also detected on monocytes. [ABSTRACT FROM AUTHOR]

Consumption of a Lactobacillus helveticus SBT2171 (LH2171)-containing cheese has been reported to exhibit immunoregulatory actions, including an increase in regulatory T cell population and reduction in proinflammatory cytokine production in mice. We examined the in vitro effects of LH2171 cells per se on immune cell function, specifically proliferation and cytokine production, which are primary reactions of the immune response. Immune cell fractions were prepared by mechanical disruption of mesenteric lymph nodes (MLN), Peyer's patches (PP), and spleens (SP) of mice. The cell fractions were dispensed into a culture plate and stimulated with anti-CD3/CD28 antibody beads in place of antigen-presenting cells or lipopolysaccharide (LPS) in the presence or absence of heat-treated LH2171 cells and other bacterial strains for comparison. After incubation, proliferation, cytokine production, and cell viability of the immune cells were determined. The LH2171 significantly inhibited the proliferation of MLN immune cells stimulated with anti-CD3/CD28 compared with other bacterial strains. The antiproliferative potency of LH2171 was effective not only on MLN but also on PP and SP stimulated with anti-CD3/CD28 or LPS. The LH2171 also decreased LPS-stimulated IL-6 production from MLN, PP, and SP, and IL-1β production from SP, but LH2171 did not affect the viability of immune cells. The LH2171 inhibited immune cell proliferation and proinflammatory cytokine (IL-6 and IL-1β) production. The inhibitory actions were not due to cytotoxicity to immune cells, suggesting that LH2171 is a dairy Lactobacillus strain with beneficial immunoregulatory properties. [ABSTRACT FROM AUTHOR]

Highlights: [•] Generation of cell lines expressing porcine Siglec-5. [•] Development of monoclonal antibodies to porcine Siglec-5. [•] Porcine Siglec-5 is expressed on myeloid cells and their precursors, and at low levels on plasmacytoid dendritic cells. [ABSTRACT FROM AUTHOR]

CD200R1 and CD200R1-like are paired receptors which modulate activation of immune cells. Here, we describe the characterisation of their porcine homologues. Analysis of database porcine sequences shows an exceptionally high homology between the extracellular Ig-like domains of these receptors, being the rest more dissimilar. We have obtained two mAbs, PCT1 and PCT3, against a CD200R1-Fc recombinant protein, that bind on CHO cells expressing GFP-tagged CD200R1. The specificity of these mAbs was analysed on CD200R1 L, and also on a CD200R1 splicing variant that lacks the V-type Ig domain. PCT1 bound to both CD200R1 and CD200R1L, but not to the splicing variant, what suggests that recognises an epitope in the V-type Ig domain. PCT3 reacted with both CD200R1 variants, but not CD200R1L, probably binding to an epitope in the N-terminal sequence of CD200R1. Analysis of porcine cells with these mAbs showed expression of CD200R1/CD200R1L on B cells, monocytes and alveolar macrophages. • Porcine CD200R1 and CD200R1L show a single amino-acid difference in their Ig-like domains. • PCT1 and PCT3 mAbs discriminate between porcine CD200R1 and CD200R1L • PCT1 mAb binds both CD200R1 and CD200R1L while PCT3 mAb only reacts with CD200R1. • These mAbs stain monocytes, a subset of B lymphocytes and alveolar macrophages. [ABSTRACT FROM AUTHOR]

Abstract: The chemokine receptor CCR7 has been a useful marker for the characterization of human and mouse T cell subsets. We have produced the porcine CCR7 ligand CCL19 fused to the human IgG1 Fc fragment, and used it to analyse CCR7 expression in swine. CCL19-Fc bound to and induced the migration of cells expressing porcine CCR7 but not of untransfected cells, corroborating its specificity. On blood lymphocytes, CCL19-Fc labelled the majority of CD4+ T cells expressing the 2E3 marker, associated with a naïve phenotype, whereas the 2E3− cells were mostly negative. Among CD8+ T cells CCL19-Fc labelled two subsets: one, CD8βhi CD11alo CD45RA+, perforin−/lo , which produced low amounts of IFN-γ after stimulation, which might correspond to naïve cells; and a second small population of CD8βlo cells which expressed high levels of CD11a, and were mostly CD45RA−, a phenotype which resembles that of human central memory T cells. [Copyright &y& Elsevier]

Abstract: The genes encoding swine leukocyte antigen (SLA) and Toll-like receptor (TLR) are highly polymorphic in pig populations, and likely have influences on infection and the effects of vaccination. We explored the associations of different genotypes of SLA class II and of the genes TLR1, TLR4, TLR5, and TLR6 with antibody responses after vaccination against Erysipelothrix rhusiopathiae (ER) and Actinobacillus pleuropneumoniae (APP) serotypes 1, 2, and 5 in 191 Duroc pigs maintained under specific pathogen-free conditions. We demonstrated close relationships between SLA class II and ER antibody response and between TLR genes other than TLR4 and APP antibody responses. Pigs with specific haplotypes in SLA class II or TLR5 showed decreased antibody response to ER vaccination or increased responses to APP2 and APP5 vaccination, respectively. It might be possible to breed for responsiveness to vaccination and to implement new vaccine development strategies unaffected by genetic backgrounds of pigs. [Copyright &y& Elsevier]

The chicken MHC-B locus affects the response to several strains of Rous sarcoma virus (RSV). We evaluated the association between haplotypes of the MHC-B locus and responses to the J strain of RSV by using an F2 experimental resource family constructed with tumor-regressive (White Leghorn) and tumorprogressive (Rhode Island Red) chickens. The MHCB haplotypes were determined by genotyping of the microsatellite marker LE10258 and MHC-B locus class 1 α chain 2 (BF2). Two haplotypes in the resource family, one associated with tumor regression and one with progression, were defined by these 2 markers. To discrirninate more precisely the regressive haplotype in this family, we further developed 35 SNP markers at the MHC-B locus. Information on the haplotypes revealed here should be useful for identifying chickens with regression and progression phenotypes of J-strain RSV-induced tumors. [ABSTRACT FROM AUTHOR]