Replenishment of Glutathione Levels Improves Mucosal Function in Experimental Acute Colitis

Abstract

SUMMARY:Because reactive oxygen species (ROS) have been implicated as mediators of inflammatory bowel disease (IBD), the purpose of the present work was to determine the functional role of mucosal GSH in the trinitrobenzenesulfonic acid in 50% ethanol (TNBSϩethanol)-induced colitis in rats. Mucosal samples were taken to evaluate the temporal relationship between the extent of injury, the levels of glutathione (GSH) during acute colitis induced by TNBSϩethanol, and the effect of N-acetylcysteine (NAC) administration. In vitro assays revealed the interaction of TNBS with GSH leading to the almost instantaneous disappearance of GSH, while the reductive metabolism of TNBS by GSSG reductase generated ROS. Mucosal samples from TNBSϩethanol-treated rats indicated a direct correlation between GSH depletion and injury detected as soon as 30 minutes after TNBSϩethanol administration that persisted 24 hours post treatment. Although, short term depletion of mucosal GSH per se by diethylmaleate did not result in mucosal injury, the oral administration of NAC (40 mM) 4 hours after TNBSϩethanol treatment increased GSH stores (2-fold), decreasing the extent of mucosal injury (60 -70%) examined at 24 hours post treatment. However, an equimolar dose of dithiothreitol failed to increase GSH levels and protect mucosa from TNBSϩethanol-induced injury. Interestingly, GSH levels in TNBSϩethanol-treated rats recovered by 1-2 weeks, an effect that was accounted for by an increase of ␥-glutamylcysteine synthetase (␥-GCS) activity due to an induction of ␥-GCS-heavy subunit chain mRNA. Thus, TNBS promotes two independent mechanisms of injury, GSH depletion and ROS generation, both being required for the manifestation of mucosal injury as GSH limitation renders intestine susceptible to the TNBS-induced ROS overgeneration. Accordingly, in vivo administration of NAC attenuates the acute colitis through increased mucosal GSH levels, suggesting that GSH precursors may be of relevance in the acute relapse of IBD. (Lab Invest 2000, 80:735-744).I nflammatory bowel disease (IBD) is a chronic inflammatory disorder whose etiology is not completely understood. Several factors are recognized to contribute to its development including an overgeneration of reactive oxygen species (ROS). Production of ROS from several sources initiate a cytotoxic cascade of events that culminate in cell death. Thus, stimulated inflammatory cells present in the inflamed mucosa are capable of producing superoxide anion and hydrogen peroxide (Harris et al, 1992;McKenzie et al, 1996;Simmonds and Rampton, 1993).The xantine oxidase pathway and the oxidation of arachidonic acid in colonocytes constitute additional sources of ROS (Biemond et al, 1988;Markowitz et al, 1988;Sedghi et al, 1993). The interaction of these species with specific vascular or interstitial components yield potent chemoattractants for inflammatory phagocytes, establishing a self-amplifying cycle of ROS production that may overwhelm defense strategies resulting in tissue damage (Lewis et al, 1988;Shingu and ...

Beneficial Effects of N-Acetylcysteine on Acetic Acid-Induced Colitis in Rats

“…Data in this study support the view that NAC has beneficial effect on GSH level in colonic injury (Ardite et al 2000;Nosal'ova et al 2000). Moreover, treated-NAC-IR was demonstrated to protect the levels of GSH more effectively than the other treatment groups against the harmful effect of AA.…”

Increased tumour necrosis factor‐α plasma levels during moderate-intensity exercise in COPD patients

“…Total vastus lateralis muscle ribonucleic acid (RNA) was extracted using the TriPure TM kit (BoehringerMannheim, Mannheim, Germany), a commercial modification of the acid guanidinium isothiocyanate/phenol/chloroform method described by CHOMCZYNSKI and SACCHI [14]. TNF-a mRNA was measured using a two-step reverse transcriptase (RT) polymerase chain reaction (PCR) method [15] using appropriate forward (59-AAGAGTTCCCCAGGGACCTCT-39) and reverse (59-CCTGGGAGTAGATGAGGTACA-39) primers (Gene Specific Relative RT-PCR kit; Ambion, Inc., Austin, TX, USA). The predicted size of the amplified product using these primers is 233 base pairs (bp).…”

Dipeptidyl Peptidase IV in Inflammatory Bowel Diseases (DPP IV/CD26)

“…Nadalje, pokazano je da interakcija -NO 2 skupine iz TNBS-a sa specifičnim flavoproteinima, uz prisutnost molekularnog kisika, dovodi do nastanka superoksidnog aniona te vodikova peroksida. Njihovom reakcijom s određenim faktorima, poput iona željeza, dolazi do stvaranja snažnih slobodnih radikala koji mogu izravno oštetiti makromolekule od vitalne važnosti za stanicu, poput DNA, proteina i lipida (130).…”