Regulation of Intestinal Protein Metabolism by Amino Acids

Gut homeostasis plays a major role in health and may be regulated by quantitative and qualitative food intake. In the intestinal mucosa, an intense renewal of proteins occurs, at approximately 50% per day in humans. In some pathophysiological conditions, protein turnover is altered and may contribute to intestinal or systemic diseases. Amino acids are key effectors of gut protein turnover, both as constituents of proteins and as regulatory molecules limiting intestinal injury and maintaining intestinal functions. Many studies have focused on two amino acids: glutamine, known as the preferential substrate of rapidly dividing cells, and arginine, another conditionally essential amino acid. The effects of glutamine and arginine on protein synthesis appear to be model and condition dependent, as are the involved signaling pathways. The regulation of gut protein degradation by amino acids has been minimally documented until now. This review will examine recent data, helping to better understand how amino acids regulate intestinal protein metabolism, and will explore perspectives for future studies

Colonic Proteome Signature in Immunoproteasome-Deficient Stressed Mice and Its Relevance for Irritable Bowel Syndrome

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Dietary salt exacerbates intestinal fibrosis in chronic TNBS colitis via fibroblasts activation

International audienceAbstract Intestinal fibrosis is a frequent complication in inflammatory bowel diseases (IBD). It is a challenge to identify environmental factors such as diet that may be driving this risk. Intestinal fibrosis result from accumulation of extracellular matrix (ECM) proteins secreted by myofibroblasts. Factors promoting intestinal fibrosis are unknown, but diet appears to be a critical component in its development. Consumption of salt above nutritional recommendations can exacerbate chronic inflammation. So far, high salt diet (HSD) have not been thoroughly investigated in the context of intestinal fibrosis associated to IBD. In the present study, we analyze the role of dietary salt in TNBS chronic colitis induced in rat, an intestinal fibrosis model, or in human colon fibroblast cells. Here, we have shown that high-salt diet exacerbates undernutrition and promoted ECM-associated proteins in fibroblasts. Taken together, our results suggested that dietary salt can activate intestinal fibroblasts, thereby contributing to exacerbation of intestinal fibrosis. Dietary salt may be considered as a putative environmental factor that drives intestinal fibrosis risk

Evaluation of Ubiquitinated Proteins by Proteomics Reveals the Role of the Ubiquitin Proteasome System in the Regulation of Grp75 and Grp78 Chaperone Proteins during Intestinal Inflammation

The ubiquitin proteasome system (UPS) is the major pathway of intracellular protein degradation and may be involved in the pathophysiology of inflammatory bowel diseases or irritable bowel syndrome. UPS specifically degrades proteins tagged with an ubiquitin chain. We aimed to identify polyubiquitinated proteins during inflammatory response in intestinal epithelial HCT-8 cells by a proteomic approach. HCT-8 cells were incubated with interleukin 1β, tumor necrosis factor-α, and interferon-γ for 2 h. Total cellular protein extracts were separated by 2D gel electrophoresis and analyzed by an immunodetection using antiubiquitin antibody. Differential ubiquitinated proteins were then identified by LC-ESI MS/MS. Seven proteins were differentially ubiquitinated between control and inflammatory conditions. Three of them were chaperones: Grp75 and Hsc70 were more ubiquitinated (p < 0.05) and Grp78 was less ubiquitinated (p < 0.05) under inflammatory conditions. The results for Grp75 and Grp78 were then confirmed in HCT-8 cells and in 2-4-6-trinitrobenzen sulfonic acid induced colitis in rats mimicking inflammatory bowel disease by immunoprecipitation. No difference was observed in irritable bowel syndrome like model. In conclusion, we showed that a proteomic approach is suitable to identify ubiquitinated proteins and that UPS-regulated expression of Grp75 and Grp78 may be involved in inflammatory response. Further studies should lead to the identification of ubiquitin ligases responsible for Grp75 and Grp78 ubiquitination

An Enteral Leucine Supply Modulates Human Duodenal Mucosal Proteome and Decreases the Expression of Enzymes Involved in Fatty Acid Beta-Oxidation

Leucine is well known to regulate protein metabolism in muscle. We recently reported that enteral leucine infusion decreased proteasome activity in human duodenal mucosa and enhanced intestinal cell proliferation, but its effects on gut proteome remain unknown. Therefore, we aimed to assess the effects of an enteral leucine infusion on the whole proteome of duodenal mucosa. In this work, 5 healthy volunteers received for 5h, on 2 occasions and in random order, an enteral supply of maltodextrins (0.25 g kg(-1) h(-1)) or maltodextrins supplemented with leucine (0.035 g kg(-1) h(-1)). At the end of infusion, endoscopic duodenal biopsy samples were collected and analyzed by 2D-PAGE. Eleven protein spots were differentially and significantly (P<0.05) expressed in response to the leucine-supplemented maltodextrins compared with maltodextrins alone. Forty percent of identified proteins by mass spectrometry were located in mitochondria. Four proteins were involved in lipid metabolism: HADHA, ACADVL and CPT2 expressions were reduced, whereas FABP1 expression was increased. In addition, the expression of DHA kinase involved in glycerol metabolism was also downregulated. Finally, leucine supplementation altered the duodenal mucosal proteome by regulating the expression of several enzymes mainly involved in lipid metabolism. These results suggest that leucine supplementation may slowdown fatty acid beta-oxidation in human duodenal mucosa

Characterizing the metabolic perturbations induced by activity-based anorexia in the C57Bl/6 mouse using 1H NMR spectroscopy

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Influence of Glutamine and Branched-Chain Amino Acids Supplementation during Refeeding in Activity-Based Anorectic Mice.

International audienceOptimizing the refeeding of patients with anorexia nervosa remains important to limit somatic complications of malnutrition, as well as to avoid disease relapses by targeting persistent mood and intestinal disorders. We aimed to evaluate the effects of glutamine (Gln) and branched-chain amino acids (BCAA) supplementation during refeeding in activity-based anorectic (ABA) mice

Glutamine, but Not Branched-Chain Amino Acids, Restores Intestinal Barrier Function during Activity-Based Anorexia

International audienceBACKGROUND: During activity-based anorexia (ABA) in mice, enhanced paracellular permeability and reduced protein synthesis have been shown in the colon while the gut-brain axis has received increasing attention in the regulation of intestinal and mood disorders that frequently occur during anorexia nervosa, a severe eating disorder for which there is no specific treatment. In the present study, we assessed the effects of oral glutamine (Gln) or branched-chain amino acids (BCAA) supplementation during ABA to target intestinal functions, body composition and feeding behavior. METHODS: C57BL/6 male mice were randomized in Control (CTRL) and ABA groups. After ABA induction, mice received, or not, either 1% Gln or 2.5% BCAA (Leu, Ile, Val) for one week in drinking water. RESULTS: Neither Gln nor BCAA supplementation affected body weight and body composition, while only Gln supplementation slightly increased food intake. ABA mice exhibited increased paracellular permeability and reduced protein synthesis in the colonic mucosa. Oral Gln restored colonic paracellular permeability and protein synthesis and increased the mucin-2 mRNA level, whereas BCAA did not affect colonic parameters. CONCLUSION: In conclusion, oral Gln specifically improves colonic response during ABA. These data should be further confirmed in AN patients

Glutamine enema regulates colonic ubiquitinated proteins but not proteasome activities during TNBS-induced colitis leading to increased mitochondrial activity

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Sex differences in response to activity-based anorexia model in C57Bl/6 mice

International audienceAnorexia nervosa is a severe eating disorder often associated with physical hyperactivity and is more frequently observed in female sex. Activity-Based Anorexia (ABA) model combines physical activity (PA) and reduced food intake and thus allows a better understanding of the mechanisms underlying anorexia nervosa. We aimed to assess sex differences in response to ABA model in C57Bl/6 mice. Twenty four male and 16 female C57BL/6 mice were studied. ABA mice were placed in individual cages with a continuously recorded activity wheel. ABA mice had a progressive limited food access from 6 h/day (day 6) to 3 h/day (day 9) until the end of the protocol (day 17). Body weight and food intake were daily measured. We studied physical activity during 24 h, during the dark phase (D-PA) and the light phase (L-PA). We also evaluated the feeding anticipatory physical activity (A-PA), the physical activity during food intake period (FI-PA) and the post-prandial physical activity (PP-PA). We observed 16.7% of mortality in males (4 out of 24 mice) during ABA protocol while no female mice died (p = 0.09). At day 17, food intake was significantly higher in females than in males (p < 0.05) that was associated with a lower body weight loss than in females (p < 0.05). Before limited food access, no gender differences in wheel running activity were observed. From day 9, A-PA significantly increased over time in males (p < 0.05 vs females) while females exhibited higher FI-PA and PP-PA (p < 0.05 vs males). Correlations between wheel running activities and, respectively, food intake and body weight loss showed gender differences, in particularly for L-PA and A-PA. Our results suggest a greater susceptibility of male mice to develop ABA, males and females exhibit different patterns of physical activity after limitation of food access. Underlying mechanisms should be further investigated