INHIBITION OF ENDOGENOUS NITRIC OXIDE ACTIVITY IMPAIRS THE COLONIC SPARING EFFECT OF ROFECOXIB, THE CYCLOOXYGENASE-2 INHIBITOR AND RESVERATROL, THE PREFERENTIAL CYCLOOXYGENASE-1 INHIBITOR IN THE COURSE OF EXPERIMENTAL COLITIS. ROLE OF OXIDATIVE STRESS BIOMARKERS AND PROINFLAMMATORY CYTOKINES

The gut mucosal barrier plays a key role in the physiology of gastrointestinal (GI) tract, preventing under homeostatic conditions, the epithelial cells of the gastric mucosa from hydrochloric acid and intestinal mucosa from alkaline secretion, food toxins and pathogenic microbiota. Previous studies have documented that blockade of both isoforms of cyclooxygenase (COX): constitutive (COX-1) and inducible (COX-2), as well NO synthase in the stomach exacerbated the gastric damage induced by various ulcerogens, however, such as effects of non-selective and selective inhibition of COX-1, COX-2 and NOS enzymes on colonic damage have been little studied. The supplementation of NO by intragastric (i.g.) treatment with NO-releasing compound NO-aspirin (NO-ASA) or substrate for NO synthase L-arginine ameliorated the damage of upper GI-tract, but whether similar effect can be observed in colonic mucosa associated with the experimental colitis, and if above mentioned compounds can be effective in aggravation or protection of experimental colitis remains less recognized. In this study rats with experimental colitis induced by intrarectal administration of 2,4,6-trinitrobenzosulphonic acid (TNBS) were daily treated for 7 days with: 1) vehicle (i.g.), 2) ASA 40 mg/kg (i.g.), 3) rofecoxib 10 mg/kg (i.g.), 4) resveratrol 10 mg/kg (i.g.), 5) NO-ASA 40 mg/kg (i.g.), 6) L-arginine 200 mg/kg (i.g.) with or without of L-NNA 20 mg/kg (i.p.). The macroscopic and microscopic area of colonic damage was determined planimetrically, the colonic blood flow (CBF) was assessed by Laser flowmetry, and the oxidative stress biomarkers malondialdehyde and 4-hydroxynonenal (MDA+4-HNE), the antioxidative factors superoxide dismutase (SOD) and glutathione (GSH), as well as proinflammatory cytokines in the colonic mucosa (tumor necrosis factor alpha (TNF-a) and interleukin-1beta (IL-1β)) were measured. We have documented that administration of TNBS produced gross and microscopic colonic damage and significantly decreased CBF (p<0.05). Treatment with ASA significantly increased the area of colonic damage (p<0.05), an effect accompanied by a significant decrease in the CBF, the significant increment of MDA+4- HNE, and the attenuation of the antioxidative properties in colonic mucosa, documented by a significant decrease of SOD activity and GSH concentration, and elevation of the colonic tissue levels of TNF-α and IL-1β comparing to control Veh-treated TNBS rats. Administration of rofecoxib or resveratrol also significantly increased the colonic damage and significantly decreased the CBF, causing an increase in MDA+4-HNE and mucosal content of TNF-α and IL-1α and a significant decrease of the SOD activity and GSH content (p<0.05), however, these changes were significantly less pronounced as compared with ASA. On the contrary, the treatment with NO-ASA, or L-arginine, significantly diminished the area of colonic lesions, the MDA+4-HNE concentration, attenuated the TNF-α and IL- 1β levels, while increasing the CBF, SOD activity and GSH content (p<0.05). The concomitant treatment of L-NNA with rofecoxib or resveratrol reversed an increase in area of colonic damage and accompanying changes in CBF, colonic mucosa TNF-α and IL-1β levels, the MDA+4-HNE concentration, and SOD activity and GSH content comparing to those observed in TNBS rats treated with these COX-inhibitors alone (p<0.05). In contrast, cotreatment with L-NNA and NO-ASA or L-arginine failed to significantly affect the decrease of colonic lesions accompanied by the rise in CBF, the attenuation of MDA+4-HNE concentration, TNF-α and IL-1β levels, SOD activity and GSH content exerted by NO-ASA- or L-arginine treatment of the respective control TNBS-rats without L-NNA administration. These observations suggest that 1) the increase of NO availability either from NO-releasing donors such as NO-ASA or NO precursors such as L-arginine, can inhibit the inflammatory and microvasculature alterations, as well as increase in lipid peroxidation due to the enhanced efficacy of these compounds to increase the antioxidative properties of colonic mucosa, 2) unlike ASA which exacerbated the severity of colitis, the treatment with rofecoxib, the specific 'safer' COX-2 inhibitor or resveratrol, the polyphenolic compound known to act as the dual COX-1 and COX-2 inhibitor, can attenuate the colonic damage during course of TNBS colitis possibly via antiinflammatory and antioxidative properties, and 3) the blockade of endogenous NO activity by L-NNA which also exacerbated the severity of mucosal damage in colitis, can abolish the sparing effect of rofecoxib and resveratrol indicating the NO bioavailability plays an important role in enhanced efficacy of both specific and dual COX inhibitors to ameliorate the experimental colitis. [ABSTRACT FROM AUTHOR]