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Human intestinal tissue samples are barely accessible to study potential health benefits of nutritional compounds. Numbers of animals used in animal trials, however, need to be minimalized. Therefore, we explored the applicability of in vitro (human Caco-2 cells) and ex vivo intestine models (rat precision cut intestine slices and the pig in-situ small intestinal segment perfusion (SISP) technique) to study the effect of food compounds. In vitro digested yellow (YOd) and white onion extracts (WOd) were used as model food compounds and transcriptomics was applied to obtain more insight into which extent mode of actions depend on the model. The three intestine models shared 9,140 genes which were used to compare the responses to digested onions between the models. Unsupervised clustering analysis showed that genes up- or down-regulated by WOd in human Caco-2 cells and rat intestine slices were similarly regulated by YOd, indicating comparable modes of action for the two onion species. Highly variable responses to onion were found in the pig SISP model. By focussing only on genes with significant differential expression, in combination with a fold change > 1.5, 15 genes showed similar onion-induced expression in human Caco-2 cells and rat intestine slices and 2 overlapping genes were found between the human Caco-2 and pig SISP model. Pathway analyses revealed that mainly processes related to oxidative stress, and especially the Keap1-Nrf2 pathway, were affected by onions in all three models. Our data fit with previous in vivo studies showing that the beneficial effects of onions are mostly linked to their antioxidant properties. Taken together, our data indicate that each of the in vitro and ex vivo intestine models used in this study, taking into account their limitations, can be used to determine modes of action of nutritional compounds and can thereby reduce the number of animals used in conventional nutritional intervention studies., Competing Interests: The authors have declared that no competing interests exist.

Background: Gene expression profiles of intestinal mucosa of chickens and pigs fed over long-term periods (days/ weeks) with a diet rich in rye and a diet supplemented with zinc, respectively, or of chickens after a one-day amoxicillin treatment of chickens, were recorded recently. Such dietary interventions are frequently used to modulate animal performance or therapeutically for monogastric livestock. In this study, changes in gene expression induced by these three interventions in cultured “Intestinal Porcine Epithelial Cells” (IPEC-J2) recorded after a short-term period of 2 and 6 hours, were compared to the in vivo gene expression profiles in order to evaluate the capability of this in vitro bioassay in predicting in vivo responses. Methods: Lists of response genes were analysed with bioinformatics programs to identify common biological pathways induced in vivo as well as in vitro. Furthermore, overlapping genes and pathways were evaluated for possible involvement in the biological processes induced in vivo by datamining and consulting literature. Results: For all three interventions, only a limited number of identical genes and a few common biological processes/ pathways were found to be affected by the respective interventions. However, several enterocyte-specific regulatory and secreted effector proteins that responded in vitro could be related to processes regulated in vivo, i.e. processes related to mineral absorption, (epithelial) cell adherence and tight junction formation for zinc, microtubule and cytoskeleton integrity for amoxicillin, and cell-cycle progression and mucus production for rye. Conclusions: Short-term gene expression responses to dietary interventions as measured in the in vitro bioassay have a low predictability for long-term responses as measured in the intestinal mucosa in vivo. The short-term responses of a set regulatory and effector genes, as measured in this bioassay, however, provided additional insight into how specific processes in piglets and broilers may be modulated by “early” signalling molecules produced by enterocytes. The relevance of this set of regulatory/effector genes and cognate biological processes for zinc deficiency and supplementation, gluten allergy (rye), and amoxicillin administration in humans is discussed. [ABSTRACT FROM AUTHOR]

To study host–probiotic interactions in parts of the intestine only accessible in humans by surgery (jejunum, ileum and colon), pigs were used as model for humans. Groups of eight 6-week-old pigs were repeatedly orally administered with 5 x 1012 CFU Lactobacillus plantarum 299v (L. plantarum 299v) or PBS, starting with a single dose followed by three consecutive daily dosings 10 days later. Gene expression was assessed with pooled RNA samples isolated from jejunum, ileum and colon scrapings of the eight pigs per group using Affymetrix porcine microarrays. Comparison of gene expression profiles recorded from L. plantarum 299v-treated pigs with PBS-treated pigs indicated that L. plantarum 299v affected metabolic and immunological processes, particularly in the ileum. A higher expression level of several B cell-specific transcription factors/regulators was observed, suggesting that an influx of B cells from the periphery to the ileum and/or the proliferation of progenitor B cells to IgA-committed plasma cells in the Peyer’s patches of the ileum was stimulated. Genes coding for enzymes that metabolize leukotriene B4, 1,25-dihydroxyvitamin D3 and steroids were regulated in the ileum. Bioinformatics analysis predicted that these metabolites may play a role in the crosstalk between intestinal immune cells and sub-mucosal adipocytes. Together with regulation of genes that repress NFKB- and PPARG-mediated transcription, this crosstalk may contribute to tempering of inflammatory reactions. Furthermore, the enzyme adenosine deaminase, responsible for the breakdown of the anti-inflammatory mediator adenosine, was strongly down-regulated in response to L. plantarum 299v. This suggested that L. plantarum 299vregulated production of adenosine by immune cells like regulatory T cells may also be a mechanism that tempers inflammation in the ileum, and perhaps also in other parts of the pig’s body. [ABSTRACT FROM AUTHOR]

Host–microorganism interactions in the intestinal tract are complex, and little is known about specific nonpathogenic microbial factors triggering host responses in the gut. In this study, mannose-specific interactions of Lactobacillus plantarum 299v with jejunal epithelium were investigated using an in situ pig Small Intestinal Segment Perfusion model. The effects of L. plantarum 299v wild-type strain were compared with those of two corresponding mutant strains either lacking the gene encoding for the mannose-specific adhesin ( msa) or sortase ( srtA; responsible for anchoring of cell surface proteins like Msa to the cell wall). A slight enrichment of the wild-type strain associated with the intestinal surface could be observed after 8 h of perfusion when a mixture of wild-type and msa-mutant strain had been applied. In contrast to the mutant strains, the L. plantarum wild-type strain tended to induce a decrease in jejunal net fluid absorption compared with control conditions. Furthermore, after 8 h of perfusion expression of the host gene encoding pancreatitis-associated protein, a protein with proposed bactericidal properties, was found to be upregulated by the wild-type strain only. These observations suggest a role of Msa in the induction of host responses in the pig intestine. [ABSTRACT FROM AUTHOR]

Abstract: The present study aimed to investigate whether long-lasting, recurrent tethering of sows leads to enduring effects on measures that may be indicative of chronic stress. Sows that had experienced tethering for about 1.5 or 4.5years and age-matched sows kept in a social housing system (loose sows) were compared. Immediately after slaughter, blood samples were taken to measure plasma cortisol levels, and the brain, spleen, and adrenals were dissected and weighed. Gene expression in the frontal cortex and hippocampus was analyzed. Plasma cortisol levels were higher in the tethered sows than in the loose sows. The older, but not the younger, tethered sows had heavier adrenal glands than their loose counterparts. The weight of the spleen was not affected by the housing conditions, but the pituitary gland was lighter in tethered sows than in loose sows. Microarray analyses revealed an increased expression of β-globin mRNA in the hippocampus and to a lesser extent in the frontal cortex of the older tethered sows, compared with the older loose sows. Taken together, the findings indicate that chronically stressed pigs develop depression-like symptoms. However, it can be questioned whether the pig subjected to repeated, long-term stress can be regarded an animal model of major depression. [Copyright &y& Elsevier]

Abstract: Acute secretory diarrhea is a major cause of morbidity and mortality in young animals and humans. Deaths result from excessive fluid and electrolyte losses. The disease is caused by non-invasive bacteria such as Vibrio cholerae and Escherichia coli which produce enterotoxins, however, much less is known about the role of individual host responses. Here we report the response of intact porcine small intestinal mucosa to infection with enterotoxigenic E. coli (ETEC). Jejunal segments in four piglets were infused with or without ETEC, and perfused for 8h, and net absorption measured. Microarray analysis at 8h post-infection showed significant differential regulation of on average fifteen transcripts in mucosa, with considerable individual variation. Differential net absorption varied between animals, and correlated negatively with the number of up regulated genes, and with one individual gene (THO complex 4). This shows that quantitative differences in gene regulation can be functionally linked to the physiological response in these four animals. [Copyright &y& Elsevier]