The extent to which microbiota alterations define or influence the outcome of metabolic diseases is still unclear, but the byproducts of microbiota metabolism are known to have an important role in mediating the host-microbiota interaction. Here, we identify that in both pre-clinical and clinical settings, metabolic syndrome is associated with the reduced capacity of the microbiota to metabolize tryptophan into derivatives that are able to activate the aryl hydrocarbon receptor. This alteration is not merely an effect of the disease as supplementation with AhR agonist or a Lactobacillus strain, with a high AhR ligand-production capacity, leads to improvement of both dietary- and genetic-induced metabolic impairments, particularly glucose dysmetabolism and liver steatosis, through improvement of intestinal barrier function and secretion of the incretin hormone GLP-1. These results highlight the role of gut microbiota-derived metabolites as a biomarker and as a basis for novel preventative or therapeutic interventions for metabolic disorders.

A monoclonal antibody, HML-1, was produced by fusion of NSI myeloma cells with spleen cells of a mouse immunized with isolated human intestinal intraepithelial lymphocytes (IEL). Immunofluorescence studies of isolated cells, as well as immunoperoxidase staining of tissue sections, indicated that HML-1 labeled all the various subsets of human intestinal IEL, approximately 40% of lamina propria T cells, 30% mesenteric lymphoblasts and some lymphocytes in other mucosae, particularly IEL. Conversely, it revealed only rare cells in all other lymphoid compartments. Analysis by polyacrylamide gel gradient electrophoresis showed that HML-1 precipitated two major noncovalently bound components of approximate mol. masses of 105 and 150 kDa from human IEL. HML-1 thus defines a novel human membrane antigen present on a subpopulation of lymphocytes preferentially associated with epithelia, and particularly with the intestinal epithelium. The characteristics of this human antigen are very similar to those of an antigen we had previously described in the rat. The possible functional role of this novel class of lymphocyte membrane antigens as well as the nature of the mechanism that triggers their expression remain to be elucidated.

Classical growth factors for colon cancer cells have been extensively described including agonists of tyrosine kinase receptors such as epidermal growth factor and related proteins (1) or insulin-like growth factors (2). More recently, some G protein-coupled receptor (GPCR) 1 agonists such as peptide hormones (3-5), prostaglandins (6), or serine proteases (7, 8) have been shown also to promote colon cancer cell proliferation often through transactivation of the epidermal growth factor receptor (6, 8). These GPCRs are expressed in both normal colonic epithelium and colon tumors (9) or even ectopically expressed by cancer cells such as in the case of the neurotensin receptor NT1 (10) or the thrombin receptor protease-activated receptor 1 (7). Whatever their expression pattern, they probably all contribute to the growth of colon tumors because of the presence of abundant ligands in the neuroendocrine environment of colonic tumors and/or to the production of receptor ligands by the tumor itself (11, 12).Our knowledge of receptor agonist suppressing colon cancer cell growth is much more limited apart from a few observations regarding transforming growth factor-␤ (13) or Fas ligand (14). We reasoned that among the very rich environment of peptide hormones and neuropeptides in the gut, we should be able to find natural agonists behaving as suppressors of colon cancer growth. In order to test this hypothesis, we developed a very simple assay by using human colon adenocarcinoma cells HT29-D4 grown in 10% FCS and screened for various peptides by their ability to inhibit cell growth. We made two dramatic hits with orexin-A and orexin-B, which appear to be robust growth inhibitors as shown here.Orexin-A and orexin-B (15), also named hypocretin-1 and hypocretin-2 (16), were discovered in 1998 by orphan receptor technologies (15) or subtractive cDNA cloning (16). They are encoded by a single gene that drives the synthesis of preproorexin that is subsequently matured into the 33-amino acid orexin-A and the 28-amino acid orexin-B, sharing 46% amino acid identity in humans (reviewed in Ref. 17). Two orexin receptor subtypes OX 1 R and OX 2 R have been cloned (15). They are serpentine GPCRs that bind both orexins with poor selectivity and are coupled to Ca 2ϩ mobilization (15). Orexins were initially characterized as neuropeptides restricted to hypothalamic neurons that project in the brain to nuclei involved in the

Background and aims-The Tage4 gene (tumour associated glycoprotein E4) is overexpressed in rat colon tumours and Min mouse intestinal adenomas. The rat Tage4 protein has approximately 40% identity with human CD155, a member of the immunoglobulin superfamily coding for a transmembrane protein capable of serving as an entry receptor for poliovirus, porcine pseudorabies virus, and bovine herpesvirus 1. Analysis of the rat Tage4 gene has revealed structural and functional similarities with the human CD155 gene. We therefore investigated expression of the CD155 gene in human colorectal carcinomas. Methods-Overall CD155 expression was assessed by semiquantitative reverse transcription-polymerase chain reaction (RT-PCR) and immunohistochemical analysis using tissue specimens from patients with colorectal adenomas and adenocarcinomas. We also used a qualitative RT-PCR assay to determine relative expression of diVerent splicing variants in each sample. Results-mRNA levels of CD155 were increased in six of six colorectal cancer tissues compared with the tumour free colon mucosa. Immunohistochemical analysis revealed an increased level of CD155 protein in 12 of 12 samples. The qualitative RT-PCR assay revealed that relative expression of the diVerent CD155 variant transcripts was similar in the diVerent normal and cancer samples tested, indicating that this overexpression is not associated with a particular mRNA variant generated by alternative splicing of the CD155 gene. Conclusion-We have shown for the first time that the CD155 gene is overexpressed in colorectal carcinoma and that this overexpression begins at an early stage in tumorigenesis and continues to late stages. (Gut 2001;49:236-240) Keywords: CD155; colorectal cancer; immunoglobulin superfamily; poliovirus receptor As our understanding of the biology of colorectal cancer progresses, new knowledge about tumorigenesis and tumour biology can be used to diagnose, treat, and prevent this type of cancer. In an initial study of an animal model for immunotherapy of colon tumours, monoclonal antibodies directed against rat colon carcinoma cells were raised.

The mucus layer covering the gastrointestinal mucosa is considered the first line of defense against aggressions arising from the luminal content. It is mainly composed of high molecular weight glycoproteins called mucins. Butyrate, a shortchain fatty acid produced during carbohydrate fermentation, has been shown to increase mucin secretion. The aim of this study was to test 1) whether butyrate regulates the expression of various MUC genes, which are coding for protein backbones of mucins, and 2) whether this effect depends on butyrate status as the major energy source of colonocytes. Butyrate was provided at the apical side of human polarized colonic goblet cell line HT29-Cl.16E in glucose-rich or glucose-deprived medium. In glucose-rich medium, butyrate significantly increased MUC3 and MUC5B expression (1.6-fold basal level for both genes), tended to decrease MUC5AC expression, and had no effect on MUC2 expression. In glucose-deprived medium, i.e., when butyrate was the only energy source available, MUC3 and MUC5B increase persisted, whereas MUC5AC expression was significantly enhanced (3.7-fold basal level) and MUC2 expression was strikingly increased (23-fold basal level). Together, our findings show that butyrate is able to upregulate colonic mucins at the transcriptional level and even better when it is the major energy source of the cells. Thus the metabolism of butyrate in colonocytes is closely linked to some of its gene-regulating effects. The distinct effects of butyrate according to the different MUC genes could influence the composition and properties of the mucus gel and thus its protective function. mucin; short-chain fatty acids; energy source; human colonic cell line THE MUCUS LAYER, COVERING the gastrointestinal mucosa, is considered the first line of defense against mechanical, chemical, or microbiological aggressions arising from the luminal contents (14). Mucus is mostly composed of mucins, i.e., glycoproteins of high molecular weight, whose protein backbones are encoded by MUC genes. So far, at least 15 different MUC genes have been identified in humans (15,32). In the large intestinal mucosa, the main MUC genes are MUC2, and to a lesser extent MUC1, MUC3, and MUC4. MUC2 codes for the main secreted mucin in the colon, whereas MUC1, MUC3, and MUC4 mainly code for membrane-located mucins but also present splicing variants coding for secreted mucins (50). Apart from their gel-forming protective function, some membrane-linked mucins, such as MUC1 (22) and MUC4 (12), exhibit specific functions in adhesion and cell signaling.MUC gene expression is altered in many colonic diseases. MUC2 is overexpressed in mucinous colorectal carcinoma, whereas its expression is particularly low in nonmucinous carcinoma (17, 43). MUC5AC and MUC6 expressions are abnormally induced in colon adenoma (6, 9). Aberrant expression of MUC genes (8) as well as modifications of their transcription (34, 45) have also been observed in inflammatory bowel disease. In addition, the thickness of the mucus layer is reduced in ulce...

Although the enteric nervous system (ENS) has been shown to regulate various mucosal functions, its role in the physiological control of the human intestinal epithelial barrier is unknown. The aim of this study was to investigate whether the ENS is able to modulate epithelial barrier permeability and a key tight junction-associated protein, zonula occludens-1 (ZO-1). Therefore, we developed a co-culture model, consisting of human submucosa containing the submucosal neuronal network and human polarized colonic epithelial monolayers (HT29-Cl.16E or Caco-2). Submucosal neurons were activated by electrical field stimulation (EFS). Permeability was assessed by measuring the flux of paracellular permeability markers (FITC-dextran or FITC-inulin) across epithelial monolayers. Expression of ZO-1 was determined by immunofluorescence, quantitative immunoblot analysis, and real time RT-PCR. Using the coculture model, we showed that EFS of submucosal neurons resulted in a reduction in FITC-dextran or FITC-inulin fluxes, which was blocked by TTX. In HT29-Cl.16E, the effect of submucosal neuron activation was blocked by a VIP receptor antagonist (VIPra) and reproduced by VIP. Furthermore, ZO-1 expression (mRNA, protein) assessed in HT29-Cl.16E, was significantly increased after submucosal neuron activation by EFS. These effects on ZO-1 expression were blocked by TTX and VIPra and reproduced by VIP. In conclusion, our results strongly suggest a modulatory role of VIPergic submucosal neuronal pathways on intestinal epithelial barrier permeability and ZO-1 expression.

IL-10 is an immunomodulatory cytokine that plays an obligate role in preventing spontaneous enterocolitis in mice. However, little is known about IL-10 function in the human intestinal mucosa. We showed here that IL-10 was constitutively expressed and secreted by the human normal colonic mucosa, including epithelial cells. Depletion of IL-10 in mucosal explants induced both downregulation of the IL-10-inducible, immunosuppressive gene BCL3 and upregulation of IFN-γ, TNF-α, and IL-17. Interestingly, TGF-β blockade also strongly induced IFN-γ production. In addition, the high levels of IFN-γ produced upon IL-10 depletion were responsible for surface epithelium damage and crypt loss, mainly by apoptosis. Polymyxin B, used as a scavenger of endogenous LPS, abolished both IFN-γ production and epithelial barrier disruption. Finally, adding a commensal bacteria strain to mucosa explant cultures depleted of both IL-10 and LPS reproduced the ability of endogenous LPS to induce IFN-γ secretion. These findings demonstrate that IL-10 ablation leads to an endogenous IFN-γ-mediated inflammatory response via LPS from commensal bacteria in the human colonic mucosa. We also found that both IL-10 and TGF-β play crucial roles in maintaining human colonic mucosa homeostasis.

Intestinal intraepithelial lymphocytes (IEL) were studied, after isolation in humans, for their surface antigens with a large variety of monoclonal antibodies. They show peculiar characteristics when compared with peripheral blood lymphocytes and intestinal lamina propria lymphocytes. Although a majority of human intraepithelial lymphocytes (IEL) express an alpha/beta type of T cell receptor (TcR), 13% express a gamma/delta TcR, a percentage which was significantly higher than that found in blood and in lamina propria. In contrast to observations in mice, there was no evidence that normal human TcR gamma/delta+ intestinal IEL might use preferential variable segments of gamma genes. About 10% of human intestinal IEL expressed the alpha chain but not the beta chain of CD8, thus resembling a subset of CD8 alpha+beta- IEL, which was recently described in mice and found to be of thymoindependent origin. In addition, 10% of human IEL had a unique phenotype of immature T cells, as they bore only CD7, but no other T cell or natural killer cell markers. Finally, even the major population of IEL which expressed the usual markers of the T cell lineage (CD3, TcR alpha/beta, CD2, CD4 or CD8 alpha/beta) differed from peripheral blood T lymphocytes by their peculiar expression of surface antigens associated with activation. Indeed, 80% of IEL were CD45R0+, CD45A-, but co-expression of CD11a, CD29 and LFA-3 was inconstant. In addition, 90% of IEL expressed HML-1.