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Glutamine treatment attenuates endoplasmic reticulum stress and apoptosis in TNBS-induced colitis.

ABSTRACT: Endoplasmic reticulum (ER) stress and apoptotic cell death play an important role in the pathogenesis and perpetuation of inflammatory bowel disease (IBD). We aimed to explore the potential of glutamine to reduce ER stress and apoptosis in a rat model of experimental IBD. Colitis was induced in male Wistar rats by intracolonic administration of 30 mg of 2,4,6-trinitrobenzene sulfonic acid (TNBS). Glutamine (25 mg/dL) was given by rectal route daily for 2 d or 7 d. Both oxidative stress (TBARS concentration and oxidised/reduced glutathione ratio) and ER stress markers (CHOP, BiP, calpain-1 and caspase-12 expression) increased significantly within 48 h of TNBS instillation, and glutamine attenuated the extent of the changes. Glutamine also inhibited the significant increases of ATF6, ATF4 and spliced XBP-1 mRNA levels induced by TNBS instillation. TNBS-colitis resulted in a significant increase in p53 and cytochrome c expression, and a reduced Bcl-xL expression and Bax/Bcl-2 ratio. These effects were significantly inhibited by glutamine. Treatment with the amino acid also resulted in significant decreases of caspase-9, caspase-8 and caspase-3 activities. Double immunofluorescence staining showed co-localization of CHOP and cleaved caspase-3 in colon sections. Phospho-JNK and PARP-1 expression was also significantly higher in TNBS-treated rats, and treatment with glutamine significantly decreased JNK phosphorylation and PARP-1 proteolysis. To directly address the effect of glutamine on ER stress and apoptosis in epithelial cells, the ER stress inducers brefeldin A and tunicamycin were added to Caco-2 cells that were treated with glutamine (5 mM and 10 mM). The significant enhancement in PERK, ATF6 phosphorylated IRE1, BiP and cleaved caspase-3 expression induced by brefeldin A and tunicamycin was partly prevented by glutamine. Data obtained indicated that modulation of ER stress signalling and anti-apoptotic effects contribute to protection by glutamine against damage in TNBS-induced colitis.

SUBMITTER: Crespo I

PROVIDER: S-EPMC3508929 | biostudies-literature | 2012

REPOSITORIES: biostudies-literature

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Glutamine treatment attenuates endoplasmic reticulum stress and apoptosis in TNBS-induced colitis.

Crespo Irene I San-Miguel Beatriz B Prause Carolina C Marroni Norma N Cuevas María J MJ González-Gallego Javier J Tuñón María J MJ

PloS one 20121128 11

Endoplasmic reticulum (ER) stress and apoptotic cell death play an important role in the pathogenesis and perpetuation of inflammatory bowel disease (IBD). We aimed to explore the potential of glutamine to reduce ER stress and apoptosis in a rat model of experimental IBD. Colitis was induced in male Wistar rats by intracolonic administration of 30 mg of 2,4,6-trinitrobenzene sulfonic acid (TNBS). Glutamine (25 mg/dL) was given by rectal route daily for 2 d or 7 d. Both oxidative stress (TBARS co ...[more]

PMID: 23209735

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Project description:BackgroundPalmitic acid (PA), the main component of dietary saturated fat, causes apoptosis in many cell types, including mouse granulosa cell. Melatonin, an important endogenous hormone, has beneficial effects on female reproductive processes. Since elevated PA levels are present in follicular fluid (FF) of patients with infertility and are shown to be toxic for granulosa cells, we investigated the molecular mechanisms of PA toxicity in mouse granulosa cells and explored the effects of melatonin on PA-induced apoptosis.MethodsGranulosa cells from immature female mice were cultured for 24 h in medium containing PA and/or melatonin. Then, the effects of PA alone or combined with melatonin on viability, apoptosis and endoplasmic reticulum (ER) stress in granulosa cells were detected by methyl thiazolyl tetrazolium (MTT) assay, flow cytometry assay and western blot. After 48 h of PA and/or melatonin treatment, the concentrations of estradiol (E2) and progesterone (P4) in the culture supernatants were measured with ELISA kits.ResultsIn this study, we explored the effects of melatonin on cell viability and apoptosis in PA-treated mouse granulosa cells and uncovered the signaling pathways involved in these processes. Our results showed that 200-800 μM PA treatment reduces cell viability, induces cell apoptosis, enhances the expression of apoptosis-related genes (Caspase 3 and B-cell lymphoma-2 (BCL-2) associated X protein (BAX)), and activates the expression of ER stress marker genes (glucose-regulated protein 78 (GRP78) and CCAAT/enhancer binding protein homologous protein (CHOP)). Melatonin treatment (1-10 μM) suppresses 400 μM PA-induced cell viability decrease, cell apoptosis, Caspase 3 activation, and BAX, CHOP, and GRP78 expression. In addition, we found that 10 μM melatonin successfully attenuated the 400 μM PA-induced estrogen (E2) and progesterone (P4) decreases.ConclusionsThis study suggests that PA triggers cell apoptosis via ER stress and that melatonin protects cells against apoptosis by inhibiting ER stress in mouse granulosa cells.

Project description:Glucosamine impairs hepatic apolipoprotein B100 (apoB100) production by inducing endoplasmic reticulum (ER) stress and enhancing cotranslational and posttranslational apoB100 degradation (Qiu, W., R. K. Avramoglu, A. C. Rutledge, J. Tsai, and K. Adeli. Mechanisms of glucosamine-induced suppression of the hepatic assembly and secretion of apolipoprotein B-100-containing lipoproteins. J. Lipid Res. 2006. 47: 1749-1761). Here, we report that glucosamine also regulates apoB100 protein synthesis via ER-stress-induced PERK activation. Short-term (4 h) glucosamine treatment of HepG2 cells reduced both cellular (by 62%) and secreted apoB100 (by 43%) without altering apoB100 mRNA. Treatment with proteasomal inhibitors only partially prevented the suppressive effects of glucosamine, suggesting that mechanisms other than proteasomal degradation may also be involved. Glucosamine-induced ER stress was associated with a significantly reduced apoB100 synthesis with no significant change in posttranslational decay rates, suggesting that glucosamine exerted its effect early during apoB biosynthesis. The role of PERK and its substrate, alpha-subunit of eukaryotic initiation factor 2 (eIF2alpha), in the suppressive effects of glucosamine on apoB synthesis was then investigated. Coexpression of apoB15 (normally resistant to intracellular degradation) with wild-type double stranded (ds) RNA activated protein kinase (PKR)-like endoplasmic reticulum kinase (PERK) in COS-7 cells resulted in a dramatic reduction in the levels of newly synthesized apoB15. Interestingly, cotransfection with apoB15 and a kinase inactive PERK mutant (K618A) increased apoB15 expression. In addition, short-term glucosamine treatment stimulated an increase in phosphorylation of PERK and eIF2alpha. Taken together, these data suggest that in addition to the induction of ER-associated degradation and other degradative pathways, ER stress is associated with suppression of apoB synthesis via a PERK-dependent mechanism.

Dataset Information

Glutamine treatment attenuates endoplasmic reticulum stress and apoptosis in TNBS-induced colitis.

Publications

Glutamine treatment attenuates endoplasmic reticulum stress and apoptosis in TNBS-induced colitis.

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