Maternal Supplementation of Food Ingredient (Prebiotic) or Food Contaminant (Mycotoxin) Influences Mucosal Immune System in Piglets.

The early life period is crucial for the maturation of the intestinal barrier, its immune system, and a life-long beneficial host-microbiota interaction. The study aims to assess the impact of a beneficial dietary (short-chain fructooligosaccharides, scFOS) supplementation vs. a detrimental dietary environment (such as mycotoxin deoxynivalenol, DON) on offspring intestinal immune system developmental profiles. Sows were given scFOS-supplemented or DON-contaminated diets during the last 4 weeks of gestation, whereas force-feeding piglets with DON was performed during the first week of offspring life. Intestinal antigen-presenting cell (APC) subset frequency was analyzed by flow cytometry in the Peyer's patches and in lamina propria and the responsiveness of intestinal explants to toll-like receptor (TLR) ligands was performed using ELISA and qRT-PCR from post-natal day (PND) 10 until PND90. Perinatal exposure with scFOS did not affect the ontogenesis of APC. While it early induced inflammatory responses in piglets, scFOS further promoted the T regulatory response after TLR activation. Sow and piglet DON contamination decreased CD16+ MHCII+ APC at PND10 in lamina propria associated with IFNγ inflammation and impairment of Treg response. Our study demonstrated that maternal prebiotic supplementation and mycotoxin contamination can modulate the mucosal immune system responsiveness of offspring through different pathways.

Lactobacillus amylovorus inhibits the TLR4 inflammatory signaling triggered by enterotoxigenic Escherichia coli via modulation of the negative regulators and involvement of TLR2 in intestinal Caco-2 cells and pig explants.

Inflammation derived from pathogen infection involves the activation of toll-like receptor (TLR) signaling. Despite the established immunomodulatory activities of probiotics, studies relating the ability of such bacteria to inhibit the TLR signaling pathways are limited or controversial. In a previous study we showed that Lactobacillus amylovorus DSM 16698T, a novel lactobacillus isolated from unweaned pigs, protects the intestinal cells from enterotoxigenic Escherichia coli (ETEC) K88 infection through cytokine regulation. In the present study we investigated whether the ability of L. amylovorus to counteract the inflammatory status triggered by ETEC in intestine is elicited through inhibition of the TLR4 signaling pathway. We used the human intestinal Caco-2/TC7 cells and intestinal explants isolated from 5 week-old crossbreed Pietrain/Duroc/Large-White piglets, treated with ETEC, L. amylovorus or L. amylovorus cell free supernatant, either alone or simultaneously with ETEC. Western blot analysis showed that L. amylovorus and its cell free supernatant suppress the activation of the different steps of TLR4 signaling in Caco-2/TC7 cells and pig explants, by inhibiting the ETEC induced increase in the level of TLR4 and MyD88, the phosphorylation of the IKKα, IKKß, IκBα and NF-κB subunit p65, as well as the over-production of inflammatory cytokines IL-8 and IL-1ß. The immunofluorescence analysis confirms the lack of phospho-p65 translocation into the nucleus. These anti-inflammatory effects are achieved through modulation of the negative regulators Tollip and IRAK-M. We also found that L. amylovorus blocks the up-regulation of the extracellular heat shock protein (Hsp)72 and Hsp90, that are critical for TLR4 function. By using anti-TLR2 antibody, we demonstrate that TLR2 is required for the suppression of TLR4 signaling activation. These results may contribute to develop therapeutic interventions using L. amylovorus in intestinal disorders of piglets and humans.

Deoxynivalenol as a new factor in the persistence of intestinal inflammatory diseases: an emerging hypothesis through possible modulation of Th17-mediated response.

BACKGROUND/AIMS: Deoxynivalenol (DON) is a mycotoxin produced by Fusarium species which is commonly found in temperate regions worldwide as a natural contaminant of cereals. It is of great concern not only in terms of economic losses but also in terms of animal and public health. The digestive tract is the first and main target of this food contaminant and it represents a major site of immune tolerance. A finely tuned cross-talk between the innate and the adaptive immune systems ensures the homeostatic equilibrium between the mucosal immune system and commensal microorganisms. The aim of this study was to analyze the impact of DON on the intestinal immune response. METHODOLOGY: Non-transformed intestinal porcine epithelial cells IPEC-1 and porcine jejunal explants were used to investigate the effect of DON on the intestinal immune response and the modulation of naive T cells differentiation. Transcriptomic proteomic and flow cytometry analysis were performed. RESULTS: DON induced a pro-inflammatory response with a significant increase of expression of mRNA encoding for IL-8, IL-1α and IL-1ß, TNF-α in all used models. Additionally, DON significantly induced the expression of genes involved in the differentiation of Th17 cells (STAT3, IL-17A, IL-6, IL-1ß) at the expenses of the pathway of regulatory T cells (Treg) (FoxP3, RALDH1). DON also induced genes related to the pathogenic Th17 cells subset such as IL-23A, IL-22 and IL-21 and not genes related to the regulatory Th17 cells (rTh17) such as TGF-ß and IL-10. CONCLUSION: DON triggered multiple immune modulatory effects which could be associated with an increased susceptibility to intestinal inflammatory diseases.

The fungal T-2 toxin alters the activation of primary macrophages induced by TLR-agonists resulting in a decrease of the inflammatory response in the pig.

T-2 toxin is known to be one of the most toxic trichothecene mycotoxins. Exposure to T-2 toxin induces many hematologic and immunotoxic disorders and is involved in immuno-modulation of the innate immune response. The objective of this work was to evaluate the effects of T-2 toxin on the activation of macrophages by different agonists of Toll-like receptors (TLR) using an in vitro model of primary porcine alveolar macrophages (PAM). Cytotoxic effects of T-2 toxin on PAM were first evaluated. An IC50 of 19.47 ± 0.9753 nM was determined for the cytotoxicity of T-2 toxin. A working concentration of 3 nM of T-2 toxin was chosen to test the effect of T-2 toxin on TLR activation; this dose was not cytotoxic and did not induce apoptosis as demonstrated by Annexin/PI staining. A pre-exposure of macrophages to 3 nM of T-2 toxin decreased the production of inflammatory mediators (IL-1 beta, TNF-alpha, nitric oxide) in response to LPS and FSL1, TLR4 and TLR2/6 agonists respectively. The decrease of the pro-inflammatory response is associated with a decrease of TLR mRNA expression. By contrast, the activation of TLR7 by ssRNA was not modulated by T-2 toxin pre-treatment. In conclusion, our results suggest that ingestion of low concentrations of T-2 toxin affects the TLR activation by decreasing pattern recognition of pathogens and thus interferes with initiation of inflammatory immune response against bacteria and viruses. Consequently, mycotoxins could increase the susceptibility of humans and animals to infectious diseases.