Basic Science S41

P079Effects of a mixture of amino acids on colonic mucosahealing after experimental colitis

X. Liu1 *, M. Baumont1, J.-C. Marie2, C. Chaumontet1,M. Andriamihaja1, F. Walker2, H. Matsumoto3, D. Tome1,A.-M. Davila1, F. Blachier1. 1Institut National de la RechercheAgronomique (INRA), AgroParisTech, Centre de Rechercheen Nutrition Humaine-Ile de France, UMR 914 Physiologiede la Nutrition et du Comportement Alimentaire, Paris,France, 2Institut National de la Sante et de la RechercheMedicale (INSERM) Universite Paris 7 Denis Diderot, U773,Centre de Recherche Biomedicale Bichat Beaujon CRB3, Paris,France, 3Institute for Innovation, Ajinomoto Co. Inc, FrontierResearch Laboratory, Kawasaki, Japan

Background: Mucosal healing (MH) is considered today asan important goal for the treatment of inflammatory boweldiseases as well as an end point for clinical trials. Duringthe process of healing, the colonic mucosa has specificrequirements in terms of energy, carbon and nitrogen sources.MH is believed to require increased supply in some amino acids.The objective of this study was to test the impact of specificamino acids supplementation (Thr, Met and Glu) on colonic MHusing DSS-induced colitis rat model.Methods: Experimental colitis was induced in Wistar malerats by the ingestion of 5% (w/v) DSS in drinking water for6 days (baseline data). The rats with colitis induced werethus randomly assigned to 2 different groups. One receivedthe amino acids mixture (AA) (Thr 0.5 g/j, Met 0.31 g/j andGlu 0.57 g/j) and the other received alanine (0.86 g/j) as iso-nitrogenous control in drinking water for 3, 7 or 10 days. Ateach time point, colonic luminal contents were collected forwater content measurement and distal segments of colon weresampled for histopathology studies, myeloperoxidase activityand glutathione content measurements, and for IL-1beta,TNF-alpha, IL-6, IL-10, MIP-2, TGF-beta, MUC-2 and MUC-3 geneexpression assessment.Results: At baseline (after 6d DSS treatment), the colonic lumi-nal water content, MPO activity, pro-inflammatory cytokines,regulatory cytokines and the percentage of inflammatorysurface of colonic mucosa were significantly higher in DSS-treated rats. After 7d of AA treatment, the colonic luminalwater content was significantly lower (71.5%+/-1.7%) comparingto the control group (77.4%+/-1.2%) (p < 0.05), indicating anamelioration of the overall absorptive function of the colon.After 10d of AA treatment, the percentage of regenerationand reepithelialization of the colonic mucosa was significantlyhigher (81%+/-7%) compared to the control group (51%+/-8%)(p < 0.05), indicating an improvement of the colonic MH.Conclusions: Our study points out a beneficial effect of specificamino acids supplementation in the colonic MH (structure andfunctionality) after experimental colitis in rats. Our resultsalso provide data on the evolution of all parameters for theestablishment of a set of criteria that could be utilized forthe monitoring of post-DSS healing phase in this experimentalmodel. Further work is needed to confirm the effects of Thr,Met and Glu on MH in clinical trials.

P080Effect of insulin like growth factor-1 on experimental colitis

D. Cano-Martínez1 *, L. Sebastian Monasor1, I.D. Roman1,M.D. Fernandez-Moreno1, B. Hernandez-Breijo1, M.V.T. Lobo2,P. Sanmartín-Salinas1, J.P. Gisbert3, L.G. Guijarro1.1Universidad de Alcala de Henares and CIBEREHD,Biochemistry and Molecular Biology, Alcala de Henares, Spain,2Universidad de Alcala de Henares IRYCIS, Cell Biology andGenetics, Alcala de Henares, Spain, 3Hospital Universitariode la Princesa-IP, Gastroenterology and CIBEREHD, Madrid,Spain

Background: The involvement of insulin like growth factor-1-receptor (IGF-1R) in the ethiopathogenesis of inflammatorybowel disease IBD has been suggested, but there is no directevidence due to the fact that IGF-1R knockout mice die atbirth of respiratory failure. Conflicting results on the subjecthave been observed in IBD and in experimental models. Levelsof IGF-1 in blood are low in patients with ulcerative colitis andCrohn’s disease as well as in the experimental models of colitis.However the expression of IGF-1 receptor is upregulated in thecolon of patients with ulcerative colitis. All these data suggestthat during colitis there is a decrease in IGF-1 together withan increase in the sensitivity to the hormone. Therefore, thishormone could be useful for IBD treatment. Our aim was tostudy the effect of IGF-1 on body weight, mineral content, leanand fat mass, mucosal integrity, and the apoptosis of coloniccells in rats with colitis.Methods: Colitis was induced by oral administration of 5%DSS (dextran sulphate sodium) to rats (n = 12) and a group ofthese animals was also treated with IGF-1 (100 mg/kg/day/i.p.).We evaluated IGF-1 effect on: i) anatomical signs (totalweight, colon length, and body composition using DXA); ii)colonic mucosal integrity (by PAS-staining, Periodic Acid Schiff)and their apoptotic status (by Tunel and by western blotof caspase-3, PARP, bax and bcl-2); iii) serum biochemicalparameters.Results: Rats treated with DSS displayed pathologic signs ofcolitis, such as, body weight loss, diarrhea, blood in feces,shortening of colon length and decrease in serum glucose. Allthese changes correlated with the decrease of mineral contentand lean and fat mass. However, the treatment with IGF-1during 9 days produced a restoration of body weight and oflean mass without reversion of mineral content or fat mass.This indicates that the tissue that responds better and/or fasterto IGF-1 is the muscle. Moreover, in rats with colitis therewas a significant reduction of colon length that was partiallyreversed by IGF-1 treatment. The anatomical changes in colonduring colitis were accompanied by loss of mucus productionand cellular apoptosis which was partially reversed by IGF-1treatment. However, no effect of IGF-1 on diarrhea and bloodin feces was observed.Conclusions: The present results demonstrate that IGF-1ameliorates some symptoms of DSS-induced colitis, andtherefore could be useful in IBD treatment.

P081Effect of condroitin sulphate on pro-inflammatory mediatorsand disease activity in patients with inflammatory boweldisease (IBD)

P.M. Linares1 *, M. Chaparro1, A. Algaba2, M. Roman3, I. MorenoArza3, F. Abad Santos3, D. Ochoa3, F. Bermejo2, J.P. Gisbert1.1Hospital Universitario de la Princesa-IP, Gastroenterologyand CIBEREHD, Madrid, Spain, 2Hospital Universitario deFuenlabrada, Gastroenterology, Fuenlabrada, Spain, 3HospitalUniversitario de la Princesa-IP, Clinical Pharmacology andCIBEREHD, Madrid, Spain

Background: Condroitin sulphate (CS), a glycosaminogly-can that modulates NF-úB, might modulate several pro-inflammatory proteins involved in the pathogenesis of IBD. In

S42 Poster presentations

order to investigate the impact of CS on the clinical courseof IBD, its effect on the concentrations of these molecules inserum and urine was assessed.Methods: Prospective observational 12-month (m) follow-upstudy in patients with IBD in remission for at least 6 m, startingCS (Condrosan®, Bioiberica S.A., Barcelona, Spain) treatmentfor osteoarthritis (OA). Visits were as follows: Baseline, 3rd,6th, 9th and 12th m. CDAI and modified Truelove Witts clinicalindexes were calculated for Crohn’s disease (CD) and ulcerativecolitis (UC) respectively. Orosomucoid, C-reactive protein(CRP) and erythrocyte sedimentation rate (ESR) were alsodetermined. Levels of VEGFA, VEGFC, FGF2, HGF, Ang1, Ang2,TGFb, TNFa, IL1b, -6, -12, -17, -23, ICAM1, VCAM1, MMP3 andPGE2 were quantified by ELISA. OA joint pain was evaluated bya visual analogue scale. The study is still ongoing.Results: At present, 29 patients with IBD (16 UC, 13 CD) havebeen included. Mean age was 62 years, and 72% were women.The mean disease duration was 14 years. 69% of patients wereunder mesalazine, 3% with sulfasalazine, 14% with thipourines,and 3% with methotrexate. There was only one patient (withUC) that had a flare during follow-up (6th m visit). The incidencerate of relapse was 4.7% per patient-year of follow-up. Thatfigure seems lower than the relapse rate previously reportedfor IBD patients. 8 UC and 5 CD patients have completed the12 m follow-up. In CD, IL1b increased (from 1.24 to 2.07 pg/mL)and Ang2 levels decreased (from 3.19 to 2.68 ng/mL) betweenbaseline and the 6th m visit (P< 0.05). In UC patients, VEGFAdecreased between baseline (339 pg/mL) and the 6th m visit(236 pg/mL) (P< 0.05). Further differences regarding the otherstudied pro-inflammatory markers were not found. At 12th m,the OA joint pain had improved in all patients (P < 0.01). 30%of patients suffered adverse events, but only 6% were relatedto the drug.Conclusions: The incidence of IBD relapse in patients under CStreatment was lower than the generally reported. CS treatmentmight modulate VEGFA, IL1b and Ang2 serum levels, but it is notassociated with modifications in the concentrations of the otherstudied pro-inflammatory mediators. CS decreases pain relatedto OA in patients with IBD.

P082Altered oligosaccharide structures reduce colitis inductionin mice defective in glycosyltransferases

S. Shinzaki1 *, H. Iijima1, T. Inoue1, M. Tsujii1, E. Miyoshi2,T. Takehara1. 1Osaka University, Gastroenterology andHepatology, Suita, Japan, 2Osaka University, MolecularBiochemistry and Clinical Investigation, Suita, Japan

Background: Oligosaccharide modifications induce variousfunctional changes in immune cells. We previously reportedthat the galactose-deficient fraction in fucosylated IgGoligosaccharides is increased with a decrease in beta-1,4-galactosyltransferase I (B4GalTI) in patients with Crohn’sdisease (CD) (Shinzaki S. et al. Am J Gastroenterol 2008).Recent reports demonstrate that B cells and immunoglobulinshave protective roles in inflammatory bowel diseases (IBD).The roles of oligosaccharide alterations in CD, however,have not been investigated yet. In the present study weinvestigated a possible role of oligosaccharide modificationin the pathophysiology of colitis using mice defective inglycosyltransferases.Methods: Colitis severity was compared between B4galt1+/

and B4galt1+/+ mice. B cells isolated from B4galt1+/ andB4galt1+/+ mice were adoptively transferred to recombinationactivating gene (Rag) 2 / mice, in which colitis was inducedby CD4+CD62L+ T cells. Cell-surface glycan profiles weredetermined by lectin microarray. Cytokine production wasdetermined in a co-culture of various types of cells isolatedfrom either B4galt1+/ or B4galt1+/+ mice.

Results: Dextran sodium sulfate (DSS) and trinitrobenzenesulfonic acid-induced colitis was significantly amelioratedin B4galt1+/ mice, which have galactose deficiency inIgG oligosaccharides (similar to that of patients with CD)compared with B4galt1+/+ mice. Amelioration of colitis wasassociated with an increased interleukin-10 (IL-10) productionfrom macrophages in B4galt1+/ mice. The amelioration ofcolitis in B4galt1+/ mice was lost in the absence of IL-10,indicating that IL-10 is indispensable for the protection ofcolitis in B4galt1+/ mice. Colitis in Rag2 / mice transferredwith CD4+CD62L+ T cells was ameliorated by co-transfer ofB cells isolated from B4galt1+/ , but not from B4galt1+/+

mice. Lectin microarray and flow cytometric analysis revealedincreased expression of polylactosamines on B4galt1+/ B cellsand macrophages in comparison with B4galt1+/+ cells. Theproduction of IL-10 from macrophages was induced via theirdirect interaction with B4galt1+/ B cells by cross-linking cell-surface polylactosamines by galectins. Moreover, alpha-1,6-fucosyltransferase (Fut8) deficient mice, which lack corefucose, also showed amelioration of DSS colitis.Conclusions: Altered oligosaccharide structures on immunecells modulate mucosal inflammation. Oligosaccharides inimmune cells can be a potential therapeutic target for IBD.

P083ADAMDEC1: a novel molecule linked to Crohn’s disease,is associated with an increased susceptibility to Citrobacterrodentium colitis in the knock out mouseN. O’Shea1 *, A. Smith1, J. Dunne1, T. Chew1, A. Segal1.1University College London, Centre of Molecular Medicine,Division of Medicine, London, United Kingdom

Background: Innate immunity is attenuated in patients withCrohn’s disease (CD), with impaired neutrophil recruitment,delayed clearance of E. coli, and defective secretion of pro-inflammatory cytokines from macrophages [1,2]. This primarymacrophage defect may result in failure to eradicate bacterialflora entering the tissues and lead to the chronic granulomatousinflammation characteristic of CD. To discover the moleculesresponsible, transcriptomic profiles were obtained fromcultured human macrophages from CD patients and controls.ADAMDEC1 a Disintegrin and Metalloprotease was under-expressed in ~10% of CD patients. This protein is almostexclusively found in macrophages and dendritic cells in thesmall and large bowel lamina propria. Here we describe theresponse of Adamdec1 / mice to an enteric bacterial infectionwith Citrobacter rodentium.Methods: Adamdec1 / and wild type mice were administered~108 or 109 C. rodentium by oral gavage and body weightmonitored for three weeks. At intervals mice were sacrificedand samples of serum, stool, colon and spleen were collected.Serum cytokine levels were measured and bacteria countedin stool and spleen. Bowel inflammation was assessedhistologically. Neutrophil and immune cell recruitment to thecolon were measured by MPO assay and qPCR respectively.Results: During infection, control mice experienced a mildself-limiting colitis, with minimal weight loss. Expression ofAdamdec1 was up-regulated in the colon and this normalisedwith resolution. Adamdec1 / mice were more susceptible toC. rodentium infection: they demonstrated dramatic weightloss (p < 0.001), a more severe colitis and a reduced survivalat the higher dose (67% vs 0%, p = 0.009). Serum levels ofTNF, IL-1 b and IL-6 were significantly lower in the knock-outmice (p < 0.05). Impaired survival was associated with positivecultures of the organisms from the spleen (p = 0.02).Conclusions: By analysing the transcriptome of macrophagesfrom CD patients we have identified a novel molecule involvedin mucosal immunity. Further work is underway to elucidatethe precise role of ADAMDEC1 in the immune response.Individuals with grossly attenuated expression levels may be