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Pre-pubertal murine models of acute colitis are lacking. Magnetic resonance colonography (MRC) is a promising minimally invasive tool to assess colitis. We aimed to: 1/ Adapt a model of acute experimental colitis to pre-pubertal rats and determine whether MRC characteristics correlate with histological inflammation. 2/ Test this model by administering a diet supplemented in transforming growth factor β2 to reverse inflammation. Twenty-four rats were randomized at weaning to one of 3 groups: Trinitrobenzene Sulfonic Acid (TNBS) group (n = 8) fed a standard diet, that received an intra-rectal 60 mg/kg dose of TNBS-ethanol; Control group (n = 8) fed standard diet, that received a dose of intra-rectal PBS; TNBS+MODULEN group (n = 8) that received a dose of TNBS and were exclusively fed MODULEN-IBD® after induction of colitis. One week after induction of colitis, rats were assessed by MRC, colon histopathology and inflammation markers (Interleukin 1β, Tumor necrosis factor α, Nitric Oxide Synthase 2 and Cyclooxygenase 2). TNBS induced typical features of acute colitis on histopathology and MRC (increased colon wall thickness, increased colon intensity on T2-weighted images, target sign, ulcers). Treatment with MODULEN-IBD® did not reduce signs of colitis on MRC. Inflammatory marker expression did not differ among study groups.

Undernutrition is a global health issue leading to 1 out 5 all deaths in children under 5 years. Undernutrition is often associated with environmental enteric dysfunction (EED), a syndrome associated with increased intestinal permeability and gut inflammation. We aimed to develop a novel murine model of undernutrition with these EED features. Post-weaning mice were fed with low-protein diet (LP) alone or combined with a gastrointestinal insult trigger (indomethacin or liposaccharides). Growth, intestinal permeability and inflammation were assessed. LP diet induced stunting and wasting in post-weaning mice but did not impact gut barrier. We therefore combined LP diet with a single administration of indomethacin or liposaccharides (LPS). Indomethacin increased fecal calprotectin production while LPS did not. To amplify indomethacin effects, we investigated its repeated administration in addition to LP diet and mice exhibited stunting and wasting with intestinal hyperpermeability and gut inflammation. The combination of 3-weeks LP diet with repeated oral indomethacin administration induced wasting, stunting and gut barrier dysfunction as observed in undernourished children with EED. As noninvasive methods for investigating gut function in undernourished children are scarce, the present pre-clinical model provides an affordable tool to attempt to elucidate pathophysiological processes involved in EED and to identify novel therapeutic strategies.

Ubiquitin proteasome system contributes to the regulation of intestinal inflammatory response as its inhibition is associated with tissue damage improvement. We aimed to evaluate whether glutamine is able to limit inflammation by targeting ubiquitin proteasome system in experimental colitis. Colitis was induced in male rats by intrarectal instillation of 2-4-6-trinitrobenzen sulfonic acid (TNBS) at day 1. From day 2 to day 6, rats daily received either an intrarectal instillation of PBS (TNBS/PBS group) or glutamine (TNBS/Gln). Rats were euthanized at day 7 and colonic samples were taken to evaluate ubiqutinated proteins by proteomic approach combining 2D electrophoresis and immunoblots directed against ubiquitin. Results were then confirmed by evaluating total expression of proteins and mRNA levels. Survival rate, TNFα, and IL-1β mRNA were improved in TNBS/Gln compared with TNBS/PBS (p < 0.05). Proteasome activities were affected by TNBS but not by glutamine. We identified eight proteins that were less ubiquitinated in TNBS/PBS compared with controls with no effect of glutamine. Four proteins were more ubiquitinated in TNBS/PBS group and restored in TNBS/Gln group. Finally, 12 ubiquitinated proteins were only affected by glutamine. Among proteins affected by glutamine, eight proteins (GFPT1, Gapdh, Pkm2, LDH, Bcat2, ATP5a1, Vdac1, and Vdac2) were involved in metabolic pathways. In conclusion, glutamine may regulate ubiquitination process during intestinal inflammation.

Anorexia nervosa is a severe eating disorder often leading to malnutrition and cachexia, but its pathophysiology is still poorly defined. Chronic food restriction during anorexia nervosa may induce gut barrier dysfunction, which may contribute to disease development and its complications. Here we have characterized intestinal barrier function in mice with activity-based anorexia (ABA), an animal model of anorexia nervosa.Male C57Bl/6 ABA or limited food access (LFA) mice were placed respectively in cages with or without activity wheel. After 5 days of acclimatization, both ABA and LFA mice had progressively limited access to food from 6 h/d at day 6 to 3 h/d at day 9 and until the end of experiment at day 17. A group of pair-fed mice (PF) was also compared to ABA.On day 17, food intake was lower in ABA than LFA mice (2.0 ± 0.18 g vs. 3.0 ± 0.14 g, p 0.001) and weight loss was more pronounced in ABA and PF compared to LFA mice (23.6 ± 1.6% and 24.7 ± 0.7% vs. 16.5 ± 1.2%; p 0.05). Colonic histology showed decreased thickness of the muscularis layer in ABA compared to LFA mice (p 0.05). Colonic permeability was increased in both ABA and PF compared to LFA mice (p 0.05) but jejunal paracellular permeability was not affected. Expression of claudin-1 in the colon was lower in the ABA than the LFA group (p 0.05), whereas occludin expression remained unaffected.Increased colonic permeability and histological alterations found in ABA mice suggest that intestinal barrier dysfunction may also occur in anorexia nervosa. The role of these alterations in the pathophysiology of anorexia nervosa should be further evaluated.

AIM To assess the feasibility of SPECT-computed tomography (CT) in rats with trinitrobenzene sulfonic acid (TNBS)-induced acute colitis and confront it with model inflammatory characteristics. METHODS Colitis was induced in Sprague-Dawley rats by intrarectal injection of TNBS (n = 10) while controls received vehicle (n = 10). SPECT-CT with intravenous injection of 10 MBq of 67Ga-Citrate was performed at day 2. SPECT-CT criteria were colon wall thickness and maximal wall signal intensity. Laboratory parameters were assessed: colon weight:length ratio, colon cyclooxygenase-2 expression by western blot and histological inflammatory score. RESULTS Colon weight/length ratio, colon COX-2 expression and histological inflammatory score were significantly higher in the TNBS group than in the control group (P = 0.0296, P < 0.0001, P = 0.0007 respectively). Pixel max tend to be higher in the TNBS group than in the control group but did not reach statistical significance (P = 0.0662). Maximal thickness is significantly increased in the TNBS group compared to the control group (P = 0.0016) while colon diameter is not (P = 0.1904). Maximal thickness and colon diameter were correlated to colon COX-2 expression (P = 0.0093, P = 0.009 respectively) while pixel max was not (P = 0.22). Maximal thickness was significantly increased when inflammation was histologically observed (P = 0.0043) while pixel max and colon diameter did not (P = 0.2452, P = 0.3541, respectively). CONCLUSION SPECT-CT is feasible and easily distinguished control from colitic rats.

Objective The ω-3 polyunsaturated fatty acid therapy in inflammatory bowel disease is focused on the effects on fish oil–derived polyunsaturated fatty acids. We speculated that a vegetal oil rich in α-linolenic acid (ALA) might also inhibit colitis. Therefore, we evaluated whether dietary ALA would decrease the expression of adhesion molecules by inducing the protective enzyme heme oxygenase-1 (HO-1) in a rat colitis model. Methods Colitis was induced at day 0 by an intrarectal injection of 2-4-6-trinitrobenzen sulfonic acid (TNBS), whereas control rats received the vehicle. Rats were fed an ALA-rich formula 450 mg · kg −1 · d −1 , whereas the other colitic group (TNBS) and the control group were fed an isocaloric corn oil formula for 14 d (from day −7 to day 7). The colonic expressions of intercellular adhesion molecule-1 (ICAM-1), vascular cell adhesion molecule-1 (VCAM-1), vascular endothelial growth factor A receptor-2 (VEGFR2), and HO-1 were studied by immunohistochemistry. Results The ALA-rich diet significantly decreased the expression of ICAM-1, VCAM-1, and VEGFR-2 compared the TNBS group, but it did not affect the expression of HO-1. Conclusion A vegetal ALA-rich formula decreases the expression of ICAM-1, VCAM-1, and VEGFR-2 and independently of HO-1 in rats with TNBS-induced colitis. Further studies are required to evaluate its therapeutic potential in inflammatory bowel disease as an alternative to fish oil.

Proteasome-mediated protein degradation may contribute to the regulation of intestinal inflammation. At the same time, low-grade inflammation and increased intestinal permeability seem to be involved in the pathophysiology of irritable bowel syndrome (IBS). Thus, we aimed to evaluate proteasome composition and activities in colonic mucosa of IBS patients and its putative pathogenic role.Proteasome activities and proteasome subunit expression were measured in colonic mucosa of IBS, Crohn's disease (CD), and control patients by fluorometric assays and western blot, respectively. Expression of inhibitor of kappa B factor (IkappaB alpha) and occludin, a tight junction protein, was also evaluated in colonic biopsies. The degradation of recombinant occludin incubated with protein extracts from colonic mucosa was evaluated in the presence or absence of proteasome inhibitor, MG132.Proteasome trypsin-like activity was increased in IBS patients compared with CD and controls, whereas chymotrypsin-like activity was upregulated in CD patients only. Caspase-like activity was reduced both in IBS and CD patients. IkappaB alpha expression was similar between IBS and controls. In contrast, occludin expression was lower in IBS than in controls, but occludin mRNA level was similar. Protein extracts from IBS patients but not from controls degraded recombinant occludin (20% over 160 min), which was blocked by MG132. Although mast cell number was increased in IBS patients, no correlation was found between this number and proteasome alterations.Our study shows that proteasome alterations are present in the colonic mucosa of IBS patients and may contribute to the pathophysiology of IBS by increasing occludin degradation.

Anticancer chemotherapy often induces side effects such as mucositis. Recent data suggest that a diet, Clinutren Protect (CP), containing whey proteins, glutamine, and transforming growth factor-b (TGFb)-rich casein limits intestinal mucositis and improves recovery after a single methotrexate (MTX) challenge in rats. Chemotherapy consists of alternating periods of treatment and rest. Thus, our study evaluated the effects of CP on nutritional outcome and intestinal mucositis in rats receiving repeated chemotherapeutic challenges. Thirty-six Sprague-Dawley rats received 3 cycles of MTX at 8-d intervals. Rats had free access to CP or control diet (Co) from 7 d before the first MTX injection until the end of the experiment at d 27. In Co, whey proteins and TGFb-rich casein were replaced by TGFb-free casein. L-Glutamine was replaced by L-alanine. Body composition was assessed by dual energy X-ray absorptiometry. Before MTX challenges, food intake and body weight were similar in both groups but became higher during MTX challenges in CP (P , 0.05). Fat mass decreased similarly in both groups. In contrast, the decrease of fat free mass between d 21 and d 27 was less pronounced in the CP group (29.5 g) than in the Co group (257.2 g) (P , 0.05). The intestinal damage score was lower in the CP group (0.6 6 0.3 vs. 2.1 6 0.6; P , 0.05). Fecal IgA increased over time in the CP group (P , 0.05) but not in the Co group. A diet containing whey proteins, glutamine, and TGFb improves nutritional outcome by limiting the reduction of fat free mass and reduces intestinal mucositis during repeated chemotherapeutic challenges in rats. J. Nutr. 140: 799‐805, 2010.

Summary Background & aims Increased intestinal permeability occurs during chemotherapy-induced intestinal mucositis. Previous data suggest that glutamine and arginine may have additive or synergic effects to limit intestinal damage. The present study aimed to evaluate the effects of glutamine and arginine, each alone or in combination, on gut barrier function during methotrexate (MTX)-induced mucositis in rats. Methods Eighty Sprague Dawley rats received during 7 days (d) standard chow supplemented with protein powder (PP), glutamine (G, 2%), arginine (A, 1.2%) or glutamine plus arginine (GA). All diets were isonitrogenous. Rats received subcutaneous injections of MTX (2.5 mg/kg) from d0 to d2. The intestinal permeability and tight junction proteins were assessed at d4 and d9 in the jejunum by FITC-dextran and by western blot and immunohistochemistry, respectively. Results At d4, intestinal permeability was increased in MTX-PP, MTX-A and MTX-GA rats compared with controls but not in MTX-G rats. The expression of claudin-1, occludin and ZO-1 was decreased in MTX-PP group compared with controls but was restored in MTX-G and MTX-A rats. In MTX-GA rats, occludin expression remained decreased. These effects could be explained by an increase of erk phosphorylation and a decrease of IκBα expression in MTX-PP and MTX-GA rats. At d9, Intestinal permeability remained higher only in MTX-GA rats. This was associated with a persistent decrease of occludin expression. Conclusions Glutamine prevents MTX-induced gut barrier disruption by regulating occludin and claudin-1 probably through erk and NF-κB pathways. In contrast, combined glutamine and arginine has no protective effect in this model.

Magnetic resonance colonography (MRC) has been recently developed to assess bowel inflammation in inflammatory bowel disease (IBD) patients. Evaluating animal models of inflammation with MRC may be important in new drug-screening processes. The aim of this study was to assess the feasibility of MRC in colitic rats and confront it with model characteristics.Colitis was induced by rectal injection of trinitrobenzene-sulfonic acid (TNBS) in 13 rats while six rats received the vehicle. MRC was performed at day 2. Colon inflammation and production of inflammatory mediators were evaluated. Image quality was assessed by wall and motion artifacts. MRC criteria were bowel wall thickness, wall signal intensity on T2-weighted (T2w) and T1w images, the appearance of a target sign pattern, and irregular patterns of mucosal surface.MRC quality was good or excellent in 16/21 examinations with no difference between groups. Colitis rats were significantly different from controls in terms of wall thickness (P = 0.004), the appearance of a target sign pattern (P = 0.02), irregular patterns of mucosal surface (P = 0.01), and hyperintensity on T1w images (P = 0.03). All MRC criteria except maximal bowel wall thickness were associated with colon weight:length ratio and inflammatory biomarkers (all P0.05). Minimal bowel wall thickness and wall signal intensity on T2w images were associated with histological score (P0.05).MRC is feasible and reliable in rats with TNBS-induced colitis. MRC criteria including colon wall thickness, wall signal intensity on T2w images, hyperintensity in T1w sequence, and the appearance of a target sign pattern may be potential targets for new IBD drugs.

5-aminosalicylic acid (5-ASA) is widely used for the treatment of inflammatory bowel disease (IBD). Recent studies have evaluated the potential of nutritional intervention as adjunct therapy to 5-ASA in IBD. N-3 polyunsaturated fatty acids (PUFA) have shown potent anti-inflammatory properties in gut inflammation. Therefore, we aimed to evaluate the efficacy of the dual therapy (n-3 PUFA plus 5-ASA) in rats with 2, 4, 6-trinitrobenzen sulfonic acid (TNBS)-induced colitis. Colitis was induced by intrarectal injection of TNBS while control rats received the vehicle. Rats received by gavage a fish oil-rich formula (n-3 groups) or an isocaloric and isolipidic oil formula supplemented with 5-ASA for 14 days. A dose response of 5-ASA (5-75 mg. suppression mg kg(-1) d(-1)) was tested. Colitis was evaluated and several inflammatory markers were quantified in the colon. COX-2 expression (P

The role of endothelial cells in inflammatory bowel disease has been recently emphasized. Endothelial activation and expression of adhesion molecules are critical for leukocytes recruitment into the inflammatory wall. Compelling evidence demonstrated anti-inflammatory effects of long chain n-3 PUFA in inflammatory models. We previously showed that long chain n-3 PUFA (EPA and DHA) inhibited inflammatory response in epithelial and dendritic cells. As long chain n-3 PUFA treatment led to a decreased expression of adhesion molecules in endothelial cells from other organs, we have now investigated their effect on intestinal endothelial cells in vitro and in colitic rats.In vitro study: Primary culture of human intestinal microvascular endothelial cells (HIMEC) were pre-treated with DHA and then incubated with IL-1β. In vivo study: Colitis was induced in 2 groups at day0 by intrarectal injection of 2-4-6-trinitrobenzen sulfonic acid (TNBS). Rats received by gavage either fish oil, rich in EPA and DHA (TNBS+n-3) or an isocaloric isolipidic oil formula for 14 days.DHA led to a decreased VCAM-1, TLR4, cyclooxygenase-2 and VEGFR2 expression and a decreased production of IL-6, IL-8 and GM-CSF and a reduced production of PGE(2) and LTB(4) (p 0.001) in IL-1β-induced HIMEC. Similarly, dietary intervention with fish oil rich in EPA and DHA significantly decreased colon production of PGE(2) and LTB(4,) endothelial VCAM-1 and VEGFR2 in rats with colitis.Data obtained from in vitro and in vivo studies reveal a potential anti-angiogenic role of long chain n-3 PUFA in intestinal endothelial cells. This protective effect of long chain n-3 PUFA may partly explain the observed benefit of dietary intake of long chain n-3 PUFA in IBD development.

BACKGROUND: Magnetic resonance colonography (MRC) has been developed to assess inflammatory bowel diseases. We aimed to assess the feasibility of MRC in rats with TNBS-induced chronic colitis and to confront imaging results with fibrosis and stenosing features of the model. MATERIALS AND METHODS: Chronic colitis was induced in 12 rats by weekly intra-rectal injection of increasing doses of TNBS for 6 weeks, while 8 control rats received the vehicle. At week 7, MRC was performed. Fibrosis scores were assessed and fibrosis mediators measured. RESULTS: Chronic colitis was associated with significant body weight loss (p

To investigate whether targeting proteasome might reverse intestinal fibrosis in rats.Chronic colitis was induced in rats by repeated administration of increasing dose of 2,4,6-trinitrobenzene sulfonic acid (TNBS, 15, 30, 45, 60, 60, 60 mg) by rectal injection for 6 wk (from day 0 to day 35), while control rats received the vehicle. TNBS + bortezomib (BTZ) rats received intraperitoneal injections of BTZ twice weekly (from day 37 to day 44) at a dose of 25 mg/kg, whereas the control and TNBS groups received the same amount of the vehicle. Histologic scoring of inflammation and fibrosis was performed. Colonic production of transforming growth factor (TGF)-β was measured by ELISA. Colon fibrosis-related proteins such as phospho-p38, phospho-SMAD2/3, Akt and peroxisome proliferator activated receptor γ (PPARγ) were studied by western blot. Expression of the tight junction proteins, occludin and claudin-1, were assessed by Western blot. Colon proteasome activities (chymotrypsin-like and trypsin-like activities) were assessed.TNBS-treated rats had a higher colon weight/length ratio compared to control rats (P0.01). Furthermore, fibrosis and inflammation scores were higher in TNBS-treated rats compared to control rats (P0.01 for both). Colonic production of TGF-β production tended to be higher in TNBS-treated rats (P0.06). Fibrosis-related proteins such as phospho-p38, phospho-SMAD2/3, and PPARγ were significantly higher in TNBS-treated rats compared to control rats (all P0.05). TNBS rats had a higher expression of Akt compared to control rats (P0.01). Tight junction proteins were modified by repeated TNBS challenge: colon occludin expression rose significantly (P0.01), whereas claudin-1 expression fell (P0.01). Bortezomib inhibition significantly decreased chymotrypsin-like activity (P0.05), but had no significant effect on trypsin-like activity (P0.05). In contrast, bortezomib had no effect on other studied parameters such as fibrosis score, TGF-β signaling, or tight junction expression (P0.05 for all).Rats with TNBS-induced chronic colitis exhibited colon fibrosis associated with higher TGF-β signaling. Proteasome inhibition by bortezomib had no effect on fibrosis in our experimental conditions.