Effects of age and zinc supplementation on transport properties in the jejunum of piglets.

Zinc is effective in the prevention and treatment of post-weaning diarrhoea and in promoting piglet growth. Its effects on the absorption of nutrients and the secretory capacity of the intestinal epithelium are controversial. We investigated the effects of age, dietary pharmacological zinc supplementation and acute zinc exposure in vitro on small-intestinal transport properties of weaned piglets. We further examined whether the effect of zinc on secretory responses depended on the pathway by which chloride secretion is activated. A total of 96 piglets were weaned at 26 days of age and allocated to diets containing three different levels of zinc oxide (50, 150 and 2500 ppm). At the age of 32, 39, 46 and 53 days, piglets were killed, and isolated epithelia from the mid-jejunum were used for intestinal transport studies in conventional Ussing chambers, with 23 µm ZnSO4 being added to the serosal side for testing acute effects. Absorptive transport was stimulated by mucosal addition of d-glucose or l-glutamine. Secretion was activated by serosal addition of prostaglandin E2 , carbachol or by mucosal application of Escherichia coli heat-stable enterotoxin (Stp ). Jejunal transport properties showed significant age-dependent alterations (p < 0.03). Both absorptive and secretory responses were highest in the youngest piglets (32 d). The dietary zinc supplementation had no significant influence on jejunal absorptive and secretory responses. However, the pre-treatment of epithelia with ZnSO4 in vitro led to a small but significant decrease in both absorptive and secretory capacities (p < 0.05), with an exception for carbachol (p = 0.07). The results showed that, in piglets, chronic supplementation with zinc did not sustainably influence the jejunal transport properties in the post-weaning phase. Because transport properties are influenced by the addition of zinc in vitro, we suggest that possible epithelial effects of zinc depend on the acute presence of this ion.

Cross-talk Between Host, Microbiome and Probiotics: A Systems Biology Approach for Analyzing the Effects of Probiotic Enterococcus faecium NCIMB 10415 in Piglets.

A comprehensive data-set from a multidisciplinary feeding experiment with the probiotic Enterococcus faecium was analyzed to elucidate effects of the probiotic on growing piglets. Sixty-two piglets were randomly assigned to a control (no probiotic treatment) and a treatment group (E. faecium supplementation). Piglets were weaned at 26 d. Age-matched piglets were sacrificed for the collection of tissue samples at 12, 26, 34 and 54 d. In addition to zootechnical data, the composition and activity of intestinal microbiota, immune cell types, and intestinal responses were determined. Our systems analysis revealed clear effects on several measured variables in 26 and 34 days old animals, while response patterns varied between piglets from different age groups. Correlation analyses identified reduced associations between intestinal microbial communities and immune system reactions in the probiotic group. In conclusion, the developed model is useful for comparative analyses to unravel systems effects of dietary components and their time resolution. The model identified that effects of E. faecium supplementation most prominently affected the interplay between intestinal microbiota and the intestinal immune system. These effects, as well as effects in other subsystems, clustered around weaning, which is the age where piglets are most prone to diarrhea.

Porcine intestinal mast cells. Evaluation of different fixatives for histochemical staining techniques considering tissue shrinkage.

Staining of mast cells (MCs), including porcine ones, is critically dependent upon the fixation and staining technique. In the pig, mucosal and submucosal MCs do not stain or stain only faintly after formalin fixation. Some fixation methods are particularly recommended for MC staining, for example the fixation with Carnoy or lead salts. Zinc salt fixation (ZSF) has been reported to work excellently for the preservation of fixation-sensitive antigens. The aim of this study was to establish a reliable histological method for counting of MCs in the porcine intestinum. For this purpose, different tissue fixation and staining methods that also allow potential subsequent immunohistochemical investigations were evaluated in the porcine mucosa, as well as submucosa of small and large intestine. Tissues were fixed in Carnoy, lead acetate, lead nitrate, Zamboni and ZSF and stained subsequently with either polychromatic methylene blue, alcian blue or toluidine blue. For the first time our study reveals that ZSF, a heavy metal fixative, preserves metachromatic staining of porcine MCs. Zamboni fixation was not suitable for histochemical visualization of MCs in the pig intestine. All other tested fixatives were suitable. Alcian blue and toluidine blue co-stained intestinal goblet cells which made a prima facie identification of MCs difficult. The polychromatic methylene blue proved to be the optimal staining. In order to compare MC counting results of the different fixation methods, tissue shrinkage was taken into account. As even the same fixation caused shrinkage-differences between tissue from small and large intestine, different factors for each single fixation and intestinal localization had to be calculated. Tissue shrinkage varied between 19% and 57%, the highest tissue shrinkage was found after fixation with ZSF in the large intestine, the lowest one in the small intestine after lead acetate fixation. Our study emphasizes that MC counting results from data using different fixation techniques can only be compared if the respective study-immanent shrinkage factor has been determined and quantification results are adjusted accordingly.

Characterization of the effects of Enterococcus faecium on intestinal epithelial transport properties in piglets.

Probiotics have been shown to have positive effects on growth performance traits and the health of farm animals. The objective of the study was to examine whether the probiotic strain Enterococcus faecium NCIMB 10415 (E. faecium) changes the absorptive and secretory transport and barrier properties of piglet jejunum in vitro and thereby to verify tendencies observed in a former feeding trial with E. faecium. Further aims were to assess a potential mechanism of probiotics by testing effects of IL-α, which is upregulated in the peripheral blood mononuclear cells of E. faecium-supplemented piglets, and to test the hypothesis that IL-1α induces a change in ion transport. Sows and their piglets were randomly assigned to a control group and a probiotic group supplemented with E. faecium. The sows received the probiotic supplemented feed from d 28 before parturition and the piglets from d 12 after birth. Piglets were killed at the age of 12 ± 1, 26 ± 1, 34 ± 1, and 54 ± 1 d. Ussing chamber studies were conducted with isolated mucosae from the mid jejunum. Samples were taken for mRNA expression analysis of sodium-glucose-linked transporter 1 (SGLT1) and cystic fibrosis transmembrane conductance regulator (CFTR). The Na(+)/glucose cotransport was increased in the probiotic group compared with the control group at 26 (P = 0.04) and 54 d of age (P = 0.01). The PGE2-induced short circuit current (Isc) was greater at 54 d of age in the probiotic group compared with the control group (P = 0.03). In addition, effects of age on the absorptive (P < 0.01) and secretory (P < 0.01) capacities were observed. Neither SGLT1 nor CFTR mRNA expression was changed by probiotic supplementation. Mannitol flux rates as a marker of paracellular permeability decreased in both groups with increasing age and were less in the probiotic group at the 26 d of age (P = 0.04), indicating a tighter intestinal barrier. The ΔIsc induced by IL-1α was inhibited by bumetanide (P < 0.01), indicating an induction of Cl(-) secretion. Thus, in this experimental setup, E. faecium increased the absorptive and secretory capacity of jejunal mucosae and enhanced the intestinal barrier. Furthermore, the results indicated that IL-1α induces bumetanide-sensitive chloride secretion. The effects of cytokines as potential mediators of probiotic effects should, therefore, be the subject of further studies.