Development of gut immunoglobulin A production in piglet in response to innate and environmental factors

International audienceThe current review focuses on pre- and post-natal development of intestinal immunoglobulin A (IgA) production in pig. IgA production is influenced by intrinsic genetic factors in the foetus as well as extrinsic environmental factors during the post-natal period. At birth, piglets are exposed to new antigens through maternal colostrums/milk as well as exogenous microbiota. This exposure to new antigens is critical for the proper development of the gut mucosal immune system and is characterized mainly by the establishment of IgA response. A second critical period for neonatal intestinal immune system development occurs at weaning time when the gut environment is exposed to new dietary antigens. Neonate needs to establish oral tolerance and in the absence of protective milk need to fight potential new pathogens. To improve knowledge about the immune response in the neonates, it is important to identify intrinsic and extrinsic factors which influence the intestinal immune system development and to elucidate their mechanism of action

Les récepteurs aux éléments microbiens dans l'intestin : analyse de leur expression dans les différents segments et organes lymphoïdes et le long de l'axe crypto-villositaire

Session 3 : Dialogue agresseurs-hôtes, adaptation réciproquesCommunication oraleabsen

Pattern recognition receptors are differentially expressed in the pig small intestine

International audiencePattern Recognition Receptors (PRR) include Toll-like receptors and NOD-like receptors. These molecules are involved in the innate immune response to infection, triggering inflammatory responses. The aim of this study was to determine if PRR and inflammatory cytokines are differentially expressed along the intestinal segments and relative lymphoid organs of healthy pigs. Duodenum, jejunum, ileum, jejunal Peyer patches, ileal Peyer patches and mesenteric lymph nodes were collected from 32 healthy pigs to study the expression of genes encoding for PRR (Tolllike receptors 1 to 10, NOD1 and NOD2) and inflammatory cytokines by quantitative PCR. Toll-like receptors 1, 2, 4, 5, 6, 8, 9, 10 were less expressed in the duodenum than in the other intestinal segments. Toll-like receptors 1, 2, 4, 7, 8, 9, 10, NOD1, 2 were more expressed in the mesenteric lymph nodes than in the intestinal segments. Toll-like receptors 1, 7, 8, 9, 10 showed a more pronounced expression in Peyer patches than in relative intestinal segment. A heatmap analysis of our data highlights two distinct clusters of organs. Intestinal segments belong to the first cluster. They displayed high expression levels of TLR 5, 6 and IL-1 beta. Among these segments, duodenum was characterized by a weak expression of PRRs and cytokines, which may be related to its local lumen environment, poor in microorganisms. Lymphoid organs belong to the second cluster. They showed weak expression levels of TLR3, 5 and IL-1 beta and a high expression of TLR 1, 2, 7, 9, 10, IL-6 and TNF-alpha. Among these organs, mesenteric lymph nodes presented a high expression of most PRR and inflammatory cytokines, which may be related to their antigen uptake function. Our findings demonstrate thus that PRR and cytokines are differentially expressed along the intestine, highlighting segment-specific mechanisms. Acknowledgment: The present study was supported by the ANR “Sus Flora” research grant (coordinator: Claire Rogel-Gaillard)

CCL28 involvement in mucosal tissues protection as a chemokine and as an antibacterial peptide.

International audienceCCL28 chemokine is expressed by epithelial cells of various mucosal tissues. This chemokine binds to CCR3 and CCR10 receptors and plays an essential role in the IgA antibody secreting cells (IgA-ASC) homing to mucosal surfaces and to lactating mammary gland as well. In addition, CCL28 has been shown to exert a potent antimicrobial activity against both Gram-negative and Gram-positive bacteria and fungi. Using the pig model, we investigated the expression of both CCR10 and CCR3 receptors in a large panel of mucosal tissues. RT-PCR analysis revealed the expression of CCR3 and CCR10 mRNA in salivary glands, nasal mucosae, Peyer's patches, small and large intestine, suggesting the presence of leucocytes expressing these receptors within these tissues. CCR10 mRNA was observed in sow mammary gland at late gestation with an increasing level during lactation. Recombinant porcine CCL28 protein was produced and mass spectrometry analysis revealed antimicrobial chemokines features such as a high pI value (10.2) and a C-terminal highly positively-charged region. Using a viable count assay, we showed that CCL28 displayed antimicrobial activity against enteric pathogens and was effective in killing Salmonella serotypes Dublin and Choleraesuis, enteroinvasive Escherichia coli K88 and non-pathogenic E. Coli K12. The potent antimicrobial function of CCL28 combined with its wide distribution in mucosal tissues and secretions suggest that this protein plays an important role in innate immune protection of the epithelial surfaces

Effects of dietary yeast strains on immunoglobulin in colostrum and milk of sows

International audienceThe ban of antibiotic growth promoters in pig diet required the development of alternative strategies and reinforced the importance of maternal immunity to protect neonates from intestinal disorders. Milk from sows fed active dry yeasts during gestation and lactation exhibited higher immunoglobulin (Ig) and protein content in milk at day 21 of lactation. In this study, we investigated whether the administration of Saccharomyces cerevisiae strains of various origins (Sc01, Sc02, Sb03) to sows during late gestation and lactation could induce higher Ig content in colostrum and milk. Results show that yeast supplementation did not increase significantly sow body weight at days 112 of gestation and 18 of lactation as well as piglet body weight gain from birth to weaning. In contrast, the IgG level in colostrum was increased in comparison with the control group when sows were supplemented with Sc01 at both 0.05 and 0.5% (p<0.05) and Sb03 at 0.5% (p<0.01). During the lactation, the level of milk IgG remained significantly higher in comparison with the control group when sows were supplemented with Sc02 at 0.05% and 0.5% and with Sb03 at 0.5%. Furthermore, in comparison with the control sows, the level of milk IgA was significantly maintained in sows supplemented with the 3 yeast strains at 0.05%. The incidence of piglet diarrhoea was decreased in groups Sc01 at both 0.05% and 0.5% and Sc02 at 0.05%. Thus, these results show that the 3 yeast strains display immunostimulatory effects on maternal immunity, but only Sc01 supplementation at 0.05% allowed jointly the increase of IgG level in colostrum, the maintenance of IgA level in milk and the decrease of piglet diarrhoea incidence. This stimulation of maternal immunity could be associated with a better systemic (colostrum IgG) and local (milk IgA) protection of neonates and suggests that dietary yeasts may have stimulated the local gut immune system of sows

Pattern recognition receptors in the gut: analysis of their expression along the intestinal tract and the crypt/villus axis

Pattern recognition receptors (PRRs) play a critical role in the detection of microorganisms and the induction of inflammatory and immune responses. Using PCR and Western-blot analysis, this study investigated the differential expression in the intestine of 14 PRRs and nine associated cytokines. Thirty-two pigs were used to determine the expression of these markers (1) along the proximal/distal axis of the small intestine (duodenum, jejunum, and ileum) and (2) between the intestinal segments and their respective lymphoid organs (Peyer's patches [PP] and mesenteric lymph nodes [MLN]). Six additional animals were used to quantify the expression of these genes along the crypt/villus axis of jejunum, using microdissected samples. Most genes showed increased expression (1) in the distal than in the proximal parts of the small intestine (TLR3, 5, RIG-I, IL-1β, IL-8, and IFN-γ); (2) in lymphoid organs (TLR1, 2, 6, 9, 10, IL-10, TNF-α), especially the MLN (TLR4, 7, 8, NOD1, NOD2, NALP3, IFN-α, IL-6, IL-12, and TGF-β), than in intestinal segments. The analysis along the crypt/villus identified: (1) genes with higher expression in lamina propria (TLR1, 2, 4, 9, NOD1, NOD2, IL-1β, IL-10, TGF-β, TNF-α) and (2) genes with higher expression in the villus (TLR3, 5, 6, RIG-I, IL-6). These results highlight the differential expression of PRRs and cytokines along the proximal/distal and the crypt/villus axis of the intestine, contributing to a fine analysis of the complex functional architecture of the small intestine and should be related to the gut microbiota

Gut microbiota in swine: composition, genetic parameters, and links with immunity and production traits

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Extensive comparison of genome wide transcription along pig small intestine and in Peyer’s patces by RNA-seq

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L'activité antibactérienne et immunomodulatrice d'un extrait d'algue verte riche en polysaccharides sulfatés

Session Santé AnimaleAntibiotics have been used for a long time in pig production to protect animals against pathogens. However, EU policy has been adopted to implement a sustainable production without adding antibiotics as growth promoters. Marine algae contain in their cell wall water-soluble sulfated polysaccharides with potential biological activities such as anticoagulant, antiviral, antibacterial and immunomodulating activities that are being explored to be used as an effective alternative to antibiotics. A crude extract containing sulfated polysaccharides was prepared from the green algae Ulva armoricana harvested in Brittany region (France) and tested for its antibacterial activity against five strains of bacterial pathogens: Salmonella Typhimurium, Staphylococcus aureus, Listeria monocytogenes, E. coli O78 and E. coli K88. The obtained results showed that this extract was more effective in inhibiting the growth of S. aureus than those of L. monocytogenes, E. coli K88 and E. coli O78. Furthermore, the ability of the extract to stimulate the expression of the immune response mediators was evaluated using an in vitro system of porcine differentiated intestinal epithelial cells IPEC-1. Analysis by RT-qPCR showed increased expression of several cytokines including TNF, IL-1, IL-6, IL-8 and CCL20. This stimulation of immune response factor expression involved the activation of TLR4 receptor. These results suggest that this extract could be used as a new prophylactic strategy to stimulate the immune response of animals and to protect mucosal tissues against pathogens

Extensive expression differences along porcine small intestine evidenced by transcriptome sequencing

The aim of this study was to analyse gene expression along the small intestine (duodenum, jejunum, ileum) and in the ileal Peyer's patches in four young pigs with no clinical signs of disease by transcriptome sequencing. Multidimensional scaling evidenced that samples clustered by tissue type rather than by individual, thus prefiguring a relevant scenario to draw tissue-specific gene expression profiles. Accordingly, 1,349 genes were found differentially expressed between duodenum and jejunum, and up to 3,455 genes between duodenum and ileum. Additionally, a considerable number of differentially expressed genes were found by comparing duodenum (7,027 genes), jejunum (6,122 genes), and ileum (6,991 genes) with ileal Peyer's patches tissue. Functional analyses revealed that most of the significant differentially expressed genes along small intestinal tissues were involved in the regulation of general biological processes such as cell development, signalling, growth and proliferation, death and survival or cell function and maintenance. These results suggest that the intrinsic large turnover of intestinal tissues would have local specificities at duodenum, ileum and jejunum. In addition, in concordance with their biological function, enteric innate immune pathways were overrepresented in ileal Peyer's patches. The reported data provide an expression map of the cell pathway variation in the different small intestinal tissues. Furthermore, expression levels measured in healthy individuals could help to understand changes in gene expression that occur in dysbiosis or pathological states