In milk-fed calves, the effects of sodium-butyrate (Na-butyrate) to replace flavomycin on growth performance and some mechanisms involved were studied. Pancreatic and intestinal morphology, digestive enzyme activities, plasma gut regulatory peptide concentrations, and expression of their receptors in the gastrointestinal tract were measured. Gastrointestinal tract defense systems were examined by measuring protein levels of 2 heat-shock proteins (HSP27 and HSP70). The calves were randomly allocated into 2 groups fed the same basic diet with flavomycin as an antimicrobial growth promoter or with Na-butyrate (3 g/kg of dry matter). Sodium-butyrate disappeared quickly in the upper gut and was not found in circulating blood. Supplementation with Na-butyrate enhanced growth rate and improved feed conversion into body weight gain compared with the flavomycin group. Supplementation with Na-butyrate was likely associated with an improvement in efficacy of the gastrointestinal tract digestive capacities expressed by enhanced production of digestive enzymes and increased absorptive capacities in the upper small intestine. The effects could have been controlled by insulin-like growth factor-1 but probably not by any of the cholecystokinin/gastrin peptide family. Concentrations of HSP27 and HSP70 were increased in stomach and colon of calves receiving Na-butyrate, thereby assuring protection of cells with intensive metabolism (chaperone function). In conclusion, beneficial effects of Na-butyrate on maturation of gastrointestinal functions were shown in milk-fed calves and may be applied to young mammals of other species.

Background & Aims Klotho deficiency in hypomorphic KL mice leads to premature senescence and phenotype consistent with impaired mineral homeostasis. Klotho has anti-inflammatory properties protecting from NO-induced endothelial dysfunction, reduces the expression of endothelial adhesion molecules, and may contribute to T-cell dysfunction. Since defective Ca2+/Pi homeostasis leading to osteopenia/osteoporosis is frequently associated with human IBD, we investigated the changes in Klotho gene expression as a consequence of experimental colitis. Methods We utilized three murine IBD models: TNBS colitis, microflora-induced colitis in gnotobiotic IL-10−/− mice, and adoptive CD4+CD45RBhigh T-cell transfer colitis. These studies were followed by in vitro approaches using renal epithelial cells (mpkDCT4 and mIMCD3), and the cloned murine KL gene promoter. Results Renal expression of Klotho mRNA and protein was significantly inhibited in all three models of human IBD. This degree of inhibition was correlated with the severity of colitis, and was reversed by neutralizing anti-TNF antibodies. In vitro, TNF resulted in a significant inhibition of KL expression and was further potentiated by IFN-γ. TNF/IFN-γ combination resulted in increased iNOS expression and significantly elevated the concentration of NO in medium. The effect of IFN-γ could be reproduced by cell exposure to SNAP (NO donor), and reversed by iNOS inhibitor, L-NIL. The cytokine effects were transcriptionally mediated since Klotho mRNA stability remained unaffected, while reporter constructs with the mKL gene promoter displayed significant downregulation in transiently transfected renal epithelial cells. Conclusions These novel findings could help explain several extraintestinal complications including abnormalities in bone homeostasis in patients with chronic colitis.

Paneth cells are the primary source of C-type lysozyme, a b-1,4-N-acetylmuramoylhydrolase that enzymatically processes bacterial cell walls. Paneth cells are normally present in human cecum and ascending colon, but are rarely found in descending colon and rectum; Paneth cell metaplasia in this region and aberrant lysozyme production are hallmarks of inflammatory bowel disease (IBD) pathology. Here, we examined the impact of aberrant lysozyme production in colonic inflammation. Targeted disruption of Paneth cell lysozyme (Lyz1) protected mice from experimental colitis. Lyz1-deficiency diminished intestinal immune responses to bacterial molecular patterns and resulted in the expansion of lysozyme-sensitive mucolytic bacteria, including Ruminococcus gnavus, a Crohn's disease-associated pathobiont. Ectopic lysozyme production in colonic epithelium suppressed lysozyme-sensitive bacteria and exacerbated colitis. Transfer of R. gnavus into Lyz1 À/À hosts elicited a type 2 immune response, causing epithelial reprograming and enhanced anti-colitogenic capacity. In contrast, in lysozyme-intact hosts, processed R. gnavus drove pro-inflammatory responses. Thus, Paneth cell lysozyme balances intestinal anti-and pro-inflammatory responses, with implications for IBD.

Na+/H+ exchanger 3 (NHE3) provides a major route for intestinal Na+ absorption. NHE3 has been considered a target of proinflammatory cytokines and enteropathogenic bacteria, and impaired NHE3 expression and/or activity may be responsible for inflammation-associated diarrhea. However, the possibility of loss of NHE3 function reciprocally affecting gut immune homeostasis has not been investigated. In this report, we describe that NHE3-deficient mice spontaneously develop colitis restricted to distal colonic mucosa. NHE3(-/-) mice housed in a conventional facility exhibited phenotypic features such as mild diarrhea, occasional rectal prolapse, and reduced body weight. Genomewide microarray analysis identified not only a large group of transport genes that potentially represent an adaptive response, but also a considerable number of genes consistent with an inflammatory response. Histological examination demonstrated changes in the distal colon consistent with active inflammation, including crypt hyperplasia with an increased number of 5-bromo-2'-deoxyuridine-positive cells, diffuse neutrophilic infiltrate with concomitant 15-fold increase in matrix metalloproteinase 8 expression, an increased number of pSer276-RelA-positive cells, and a significant decrease in periodic acid-Schiff-positive goblet cells. Real-time PCR demonstrated elevated expression of inducible nitric oxide synthase (38-fold), TNF-alpha (6-fold), macrophage inflammatory protein-2 (48-fold), and IL-18 (3-fold) in the distal colon of NHE3(-/-) mice. NHE3(-/-) mice showed enhanced bacterial adhesion and translocation in the distal colon. Colitis was ameliorated by oral administration of broad-spectrum antibiotics. In conclusion, NHE3 deficiency leads to an exacerbated innate immune response, an observation suggesting a potentially novel role of NHE3 as a modifier gene, which when downregulated during infectious or chronic colitis may modulate the extent and severity of colonic inflammation.

. Reduced colonic microbial diversity is associated with colitis in NHE3-deficient mice.

Competition of commensal and probiotic bacteria with pathogens for adhesion and colonization is one of the important protective mechanisms of gastrointestinal tract. In this study, we examined the ability of Lactobacillus paracasei to inhibit the adhesion of pathogenic Salmonella enterica to human colon adenocarcinoma Caco-2 cells. Caco-2 cells were grown for 6 or 21 days to obtain nondifferentiated or well-differentiated cells, respectively. In adhesion experiments, bacteria were added to the cells for 2 or 4 hours. The number of attached bacteria was expressed as colony-forming units (CFUs), Caco-2 cells were counted in hematocytometer. Both bacterial strains used adhered better to well-differentiated than to nondifferentiated Caco-2 cells, however, the amount of Salmonella adhered to Caco-2 after 2 hours of contact was 12-fold higher in comparison to L. paracasei and almost 27-fold higher after 4 hours of contact. Two types of experiments were done: coincubation (both bacteria were added to Caco-2 cells simultaneously), and preincubation (L. paracasei was incubated with Caco-2 cells first, and then S. enterica was added). In coincubation experiment, the presence of L. paracasei decreased S. enterica adhesion by 4-fold and in preincubation experiment even 7-fold. Generally, Lactobacillus spent culture supernatants (SCSs) acted weaker as inhibitors of Salmonella adhesion in comparison to the whole L. paracasei culture in coincubation experiment. In conclusion, the displacement of pathogens by lactic acid bacteria and its secretions showed here depends on the time of bacteria-epithelial cell contact, and also on the stage of Caco-2 differentiation.

Background & Aims NHE3 is a target of inhibition by proinflammatory cytokines and pathogenic bacteria, an event contributing to diarrhea in infectious and idiopathic colitis. In mice, NHE3 deficiency leads to mild diarrhea, increased intestinal expression of IFN-γ, and distal colitis, suggesting its role in epithelial barrier homeostasis. Aim To investigate the role of NHE3 in maintaining mucosal integrity. Methods Control or DSS-treated, 6–8 wk wild-type (WT) and NHE3−/− mice were used for the experiments. Small intestines were dissected for further analysis. Results NHE3−/− mice have elevated numbers of CD8α+ T and NK cells in the IEL and LPL compartments, representing the source of IFN-γ. NHE3−/− mice display alterations in epithelial gene and protein expression patterns which predispose them to a high susceptibility to DSS, with accelerated mortality resulting from intestinal bleeding, hypovolemic shock, and sepsis, even at a very low DSS concentration. Microarray analysis and intestinal hemorrhage indicate that NHE3 deficiency predisposes mice to DSS-induced small intestinal injury, a segment never reported as affected by DSS, and demonstrate major differences in the colonic response to DSS challenge in WT and NHE3−/− mice. In NHE3−/− mice, broad spectrum oral antibiotics or anti-asialo GM1 antibodies reduce the expression of IFN-γ and iNOS to basal levels and delay, but do not prevent, severe mortality in response to DSS treatment. Conclusion These results suggest that NHE3 participates in mucosal responses to epithelial damage acting as a modifier gene determining the extent of the gut inflammatory responses in the face of intestinal injury.

Larmonier CB, Uno JK, Lee KM, Karrasch T, Laubitz D, Thurston R, Midura-Kiela MT, Ghishan FK, Sartor RB, Jobin C, Kiela PR. Limited effects of dietary curcumin on Th-1 driven colitis in IL-10 deficient mice suggest an IL-10-dependent mechanism of protection.