Melatonin reduces dinitrobenzene sulfonic acid-induced colitis

Inflammatory bowel disease (IBD) is characterized by oxidative and nitrosative stress, leukocyte infiltration, and up-regulation of intercellular adhesion molecule 1 (ICAM-1) expression in the colon. The aim of this study was to examine the effects of the pineal secretory product melatonin in rats subjected to experimental colitis. Colitis was induced in rats by intracolonic instillation of dinitrobenzene sulfonic acid (DNBS). Rats experienced bloody diarrhea and a significant loss of body weight. Four days after DNBS administration, the colon damage was characterized by areas of mucosal necrosis. Neutrophil infiltration (indicated by myeloperoxidase [MPO] activity in the mucosa) was associated with up-regulation of ICAM-1, expression of P-selectin, and high levels of malondialdehyde (MDA). Immunohistochemistry for nitrotyrosine and poly (ADP-ribose) synthetase (PARS) showed an intense staining in the inflamed colon. Staining of colon tissue sections obtained from DNBS-treated rats with an anti-cycloxygenase-2 (COX-2) antibody showed a diffuse staining of the inflamed tissue. Furthermore, expression of inducible nitric oxide synthase (iNOS) was found mainly in the macrophages of the inflamed colons from DNBS-treated rats. Treatment with melatonin (15 mg/kg daily, intraperitoneally) significantly reduced the appearance of diarrhea and the loss of body weight. This was associated with a remarkable amelioration of the disruption of the colonic architecture, as well as a significant reduction of colonic MPO activity and MDA levels. Melatonin also reduced the appearance of nitrotyrosine and PARS immunoreactivity in the colon, as well as reducing the up-regulation of ICAM-1 and the expression of P-selectin. The intensity and degree of the stainings for COX-2 and iNOS were markedly reduced in tissue sections obtained from melatonin-treated rats. The results of the this study suggest that the administration of melatonin might be beneficial for the treatment of IBD.     

Ameliorative effects of sodium ferulate on experimental colitis and their mechanisms in rats

AIM: To investigate the ameliorative effects of sodium ferulate (SF) on acetic acid-induced colitis and their mechanisms in rats.METHODS: The colitis model of Sprague-Dawley rats was induced by intracolon enema with 8 % (WV) of acetic acid.The experimental animals were randomly divided into model control, 5-aminosalicylic acid therapy group and three dose of SF therapy groups. The 5 groups were treated intracolonically and daily (8:00 am) for 7 days 24 h following the induction of colitis. A normal control group of rats clystered with normal saline instead of acetic acid was also included in the study.Pathological changes of the colonic mucosa were evaluated by the colon mucosa damage index (CMDI) and the histopathological score (HS). The insulted colonic mucosa was sampled for a variety of determinations at the end of experiment when the animals were sacrificed by decapitation.Colonic activities of myeloperoxidase (MPO) and superoxide dismutase (SOD), and levels of malondialdehyde (MDA)and nitric oxide (NO) were assayed with ultraviolet spectrophotometry. Colonic contents of prostaglandin E2 (PGE2) and thromboxane B2 (TXB2)were determined by radioimmunoassay. The expressions of inducible nitric oxide synthase (iNOS), cyclo-oxygenase-2 (COX-2) and nuclear factor kappa B (NF-κB) p65 proteins in the colonic tissue were detected with immunohistochemistry.RESULTS: Enhanced colonic mucosal injury, inflammatory response and oxidative stress were observed in the animals clystered with acetic acid, which manifested as the significant increase of CMDI, HS, MPO activities, MDA and NO levels,PGE2 and TXB2 contents, as well as the expressions of iNOS,COX-2 and NF-κB p65 proteins in the colonic mucosa,although the colonic SOD activity was significantly decreased compared with the normal control (CMDI: 2.9±0.6 vs0.0±0.0;HS: 4.3±0.9 vs0.7±1.1; MPO: 98.1±26.9 vs24.8±11.5; MDA:57.53±12.36 vs9.21±3.85; NO: 0.331±0.092 vs0.176±0.045;PGE2: 186.2±96.2 vs 42.8±32.8; TXB2: 34.26±13.51 vs 8.83±3.75; iNOS: 0.365±0.026 vs0.053±0.015; COX-2:0.296±0.028 vs0.034±0.013; NF-κB p65:0.314±0.026 vs 0.039±0.012; SOD: 28.33±1.17 vs36.14±1.91; P<0.01).However, these parameters were found to be significantly ameliorated in rats treated locally with SF at the given dose (CMDI: 1.8±0.8, 1.6±0.9; HS: 3.3±0.9, 3.1±1.0; MPO:63.8±30.5, 36.2±14.2; MDA: 41.84±10.62, 37.34±8.58; NO:0.247±0.042; 0.216±0.033; PGE2: 77.2±26.9, 58.4±23.9;TXB2:18.07±14.83; 15.52±8.62; iNOS:0.175±0.018, 0.106±0.019;COX-2: 0.064±0.018, 0.056±0.014; NF-κBp65: 0.215±0.019,0.189±0.016; SOD: 32.15±4.26, 33.24±3.69; P<0.05-0.01).amelioration of colonic mucosal injury as evaluated by CMDI and HS.CONCLUSION: Administration of SF intracolonically may have significant therapeutic effects on the rat model of colitis induced by acetic acid enema, which was probably due to the mechanism of antioxidation, inhibition of arachidonic acid metabolism and NF-κB expression.     

Cyclo-oxygenase-2 inhibitors ameliorate the severity of experimental colitis in rats

BACKGROUND: Both in experimental colitis and in inflammatory bowel disease, colonic eicosanoid generation is enhanced and may contribute to the pathogenesis of the inflammatory response. AIMS: To evaluate the effect of selective cyclo-oxygenase-2 (COX-2) inhibitors on the extent and severity of two models of experimental colitis. METHODS: Colitis was induced by intra-caecal administration of 2 ml 5% acetic acid or intra-colonic administration of 0.1 ml 3% iodoacetamide. Rats were treated intra-gastrically with nimesulide 2 x 10 mg/kg/day, or once with SC-236 6 mg/kg, and killed 1 or 3 days after damage induction. The colon was isolated, weighed, macroscopic damage was measured, and mucosal samples were obtained for histology and for determination of myeloperoxidase (MPO) and nitric oxide synthase (NOS) activities and eicosanoid generation. The serum levels of thromboxane B2 (TXB2), tumour necrosis factor-alpha (TNF-alpha) and interleukin-1beta (IL-1beta) were determined. RESULTS: Nimesulide significantly decreased the extent of colitis induced by acetic acid. Both nimesulide and SC-236 significantly decreased the extent of iodoacetamide-induced colonic damage. The decrease in the extent of colitis induced by nimesulide was accompanied by a significant decrease in mucosal MPO and NOS activities. Nimesulide and SC-236 decreased the enhanced colonic eicosanoid generation in acetic acid and iodoacetamide-induced colitis, and, in iodoacetamide-treated rats, nimesulide also decreased the elevated serum TNF-alpha and IL-1beta levels. CONCLUSIONS: The effective nimesulide and SC-236-induced amelioration of the severity of the colitis in acetic acid and iodoacetamide-treated rats confirms the role of eicosanoids in their pathogenesis and suggests that COX-2 inhibitors may be of value in the treatment of inflammatory bowel disease.     

Evaluation of the Role of Neutrophils in the Pathogenesis of Acetic Acid-Induced Colitis in Mice

Background: Neutrophils are thought to play a role in the pathogenesis of inflammatory bowel diseases (IBD) such as ulcerative colitis and Crohn's disease, since prominent neutrophil infiltration has been observed in the inflamed colonic mucosa of patients with IBD. However, the role of neutrophils in the pathogenesis of IBD and experimental colitis remains equivocal. The aim of the present study is to clarify the possible role of neutrophils in the progression of acetic acid-induced colitis in mice. Methods: Using neutropenic mice treated with cyclophosphamide or with an LTB₄ receptor antagonist, ONO-4057, the relationship between the severity of macroscopic colonic damage, the extent of myeloperoxidase (MPO) activities in the colonic tissues, and the number of neutrophils in the blood were examined after induction of colitis in mice. Results: Changes of MPO activity in the colonic tissues paralleled well with the severity of the mucosal damage. In spite of a significant reduction in the number of neutrophils in the blood in cyclophosphamide-treated mice, neither the severity of mucosal damage in the colon nor the increase in MPO activities in the colonic tissues was affected 24 h after induction of colitis. Treatment with ONO-4057 significantly suppressed both the severity of mucosal damage in the colon and MPO activities in the colonic tissues in acetic acid-induced colitis in mice. Conclusions: The present results, obtained using treatment with cyclophosphamide and ONO-4057, show that the severity or the progression of acetic acid-induced colitis in mice was not influenced by a reduction of circulating neutrophils to about 25% of base line.     

Effects of melatonin on the expression of iNOS and COX－2 in rat models of colitis

AIM: To investigate the effects of melatonin (MT) on the expression of inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) in rat models of colitis. METHODS: Healthy adult Sprague-Dawlay (SD) rats of both sexes, weighing 280+/-30 g, were employed in the present study. The rat models of colitis were induced by either acetic acid or 2,4,6-trinitrobenzene sulfonic acid (TNBS) enemas. The experimental animals were randomly divided into melatonin treatment and model control group that were intracolicly treated daily with melatonin at doses of 2.5, 5.0, 10.0 mg/kg(-1) and equal amount of saline respectively from 24 h following induction of colitis in rats inflicted with acetic acid enema and the seventh day in rats with TNBS to the end of study. A normal control group of rats treated with neither acetic acid nor TNBS but saline enema was also included in the study. On the 28(th) day of the experiment, the rat colon mucosal damage index (CDMI) was calculated, and the colonic prostaglandin E(2) (PGE(2)), nitric oxide (NO), as well as the iNOS and COX-2 expression were also determined biochemically or immunohistochemically. RESULTS: CDMI increased to 2.87+/-0.64 and 3.12+/-1.12 respectively in rats treated with acetic acid and TNBS enema, which was in accordance with the significantly elevated colonic NO and PGE(2) contents, as well as the up-regulated colonic iNOS and COX-2 expression in both of the two rat models of colitis. With treatment by melatonin at the doses of 5.0 and 10.0 mg/kg(-1), CDMI in both models of rat colitis was significantly decreased (P<0.05-0.01), which accorded synchronously and unanimously with the reduced colonic NO and PGE(2) content, as well as the down-regulated expression of colonic iNOS and COX-2. CONCLUSION: Melatonin has a protective effect on colonic injury induced by both acetic acid and TNBS enemas, which is probably via a mechanism of local inhibition of iNOS and COX-2 expression in colonic mucosa.     

Transforming growth factor-alpha (TGF-alpha) is not needed for malignant transformation in experimental colitis

BACKGROUND: Transforming growth factor-alpha (TGF-alpha) is a key mediator of colonic mucosal protection and/or repair mechanisms in orally induced acute dextran sodium sulphate (DSS) colitis. However, it also has been suggested that TGF-alpha may contribute to malignant transformation in the colon. The aim of the studies was to determine whether TGF-alpha is needed for malignant transformation in orally induced chronic DSS colitis using TGF-alpha deficient mice (wa-1) and Balb/c mice, a strain competent in TGF-alpha. METHODS: Chronic colitis was induced by oral administration of DSS (5%) for 7 days followed by drinking water for 10 days in wa-1 and Balb/c mice (n = 20, per group). In the two subsequent cycles (7 days DSS, 10 days water) 3% DSS-water was utilized due to a high mortality in the wa-1 group. Mucosal injury severity was assessed histologically and graded (three grades). A crypt damage score (CDS) reflecting all three grades of mucosal pathology was calculated. Mucosal dysplasia and cancerous lesions were noted. RESULTS: Seven per cent of the entire colonic mucosa was completely destroyed in wa-1 animals compared to 3% in Balb/c mice (P < 0.05). The CDS was 10.2 +/- 0.4 and 4.8 +/- 0.3 in wa-1 and Balb/c mice, respectively (P < 0.05). Fifteen incidences of mucosal dysplasia were found in the 10 surviving wa-1 animals and 31 incidences were found in 20 Balb/c animals. In both groups, one fully developed adenomatous cancerous lesion was present. CONCLUSIONS: The markedly increased severity of mucosal injury in chronic induced DSS colitis in TGF-alpha deficient wa-1 mice compared to Balb/c mice further substantiates that endogenous TGF-alpha is a pivotal mediator of protection and/or healing mechanisms in the colon. The appearance of dysplastic and cancerous lesions in TGF-alpha deficient animals suggests that TGF-alpha per se is not essential for malignant mucosal cell transformation in colitis.     

Healing impairment effect of cyclooxygenase inhibitors on dextran sulfate sodium-induced colitis in rats

We examined the effects of various cyclooxygenase (COX) inhibitors on the healing of colonic lesions induced by dextran sulfate sodium (DSS) in the rat. Colonic lesions were induced by 2.5% DSS in the drinking water for 7 days, and then the animals were fed with tap water for subsequent 7 days. Indomethacin (a nonselective COX inhibitor), SC-560 (a selective COX-1 inhibitor), or rofecoxib (a selective COX-2 inhibitor) was given orally twice daily after termination of the DSS treatment. DSS treatment caused severe colonic lesions with a decrease in body weight gain and colon length as well as an increase in myeloperoxidase activity and thiobarbituric acid reactant levels. The severity of colitis gradually reduced, with an improvement of morphological and histological alterations. Daily administration of indomethacin and rofecoxib significantly delayed the healing of colitis with deleterious influences on histological restitution as well as mucosal inflammation, while SC-560 had no effect. Although COX-1 mRNA was expressed in the colon without much alteration during the test period, the expression of COX-2 was upregulated with a peak on day 3 and decreased thereafter. The mucosal prostaglandin E2 content in the colon showed a biphasic change, in parallel with that of the COX-2 expression. The increased prostaglandin E2 production in the injured mucosa was attenuated by indomethacin and rofecoxib, but not by SC-560. These results suggest that endogenous prostaglandins produced by COX-2 play an important role in the healing of DSS-induced colonic lesions. Caution should be paid to the use of selective COX-2 inhibitors as well as nonsteroidal anti-inflammatory drugs in patients with colitis.     

Colonic production of nitric oxide gas in ulcerative colitis, collagenous colitis and uninflamed bowel

BACKGROUND: Nitric oxide (NO) produced in excess by the inflamed human colon is generally considered a pathway of mucosal damage. In an attempt to quantify colonic mucosal production of NO in various forms of colitis we performed 'steady-state' gas perfusion of whole colon in 11 patients with ulcerative colitis, 10 patients with collagenous colitis and 20 controls with uninflamed mucosa. METHODS: The tip of a Teflon tube was placed in the caecum during colonoscopy. Subsequently, argon was infused at a constant rate for 70-180 min. Argon and NO in gas sampled from the rectum were measured by neutron activation analysis and the chemiluminescence technique, respectively. RESULTS: The use of argon as a marker of colonic NO output was justified by complete recovery (96%+/-2; mean +/- s(x); n = 5) of argon in gas collected from the rectum and a constant output of NO at varying perfusion rates (25, 50 and 75 ml/min coefficient of variation 21%; n = 6). In patients with ulcerative colitis, colonic output of NO was 10-fold higher (P < 0.001) than in controls and positively correlated (P < 0.01) to indices of disease activity. In patients with collagenous colitis, colonic output of NO was 50-fold higher (P < 0.01) than in controls during periods with watery diarrhoea (n = 6), but within the range observed in ulcerative colitis in the absence of diarrhoea (n = 4). CONCLUSIONS: Argon gas perfusion of whole colon using chemiluminescence technique for measurement of NO is a reliable method for quantification of colonic mucosal NO production. Increased colonic production of NO in collagenous colitis, which exceeds the output observed even in extensive ulcerative colitis, militates against the theory that NO per se is a cause of mucosal injury.     

Protective Effect of Lafutidine, a Novel Histamine H2-Receptor Antagonist, on Dextran Sulfate Sodium-Induced Colonic Inflammation Through Capsaicin-Sensitive Afferent Neurons in Rats

Lafutidine, a histamine H2-receptor antagonist, exhibits gastric mucosal protective action mediated by capsaicin-sensitive afferent neurons, in addition to a potent antisecretory effect. In this study we examined the effect of lafutidine on dextran sulfate Na (DSS)-induced ulcerative colitis in rats, in relation to capsaicin-sensitive afferent neurons. Experimental colitis was induced in rats by daily treatment with 3% DSS in drinking water for 7 days. Lafutidine, capsaicin, and cimetidine were administered per os twice daily for 6 days. The ulceration area, colon length, and myeloperoxidase (MPO) activity were measured on day 7 after the onset of DSS treatment. DSS caused severe mucosal lesions in the colon, accompanied by an increase in MPO activity as well as a decrease in body weight gain and colon length. Daily administration of lafutidine dose-dependently reduced the severity of DSS-induced colitis and significantly mitigated changes in the colon length and MPO activity. The effects of lafutidine were mimicked by daily administration of capsaicin but not cimetidine and were totally abolished by chemical ablation of capsaicin-sensitive afferent neurons. In contrast, desensitization of afferent neurons significantly worsened the colonic inflammation induced by DSS. It was also found that both lafutidine and capsaicin increased the secretion of mucus in the colonic mucosa. These results suggest that lafutidine is effective against the ulcerative colitis induced by DSS through capsaicin-sensitive afferent neurons. This action might be attributable at least partly to the enhancement of colonic mucus secretion.     

Expression and significance of nuclear factor κB p65 in colon tissues of rats with TNBS-induced colitis

AIM: To investigate the role of NF-κB in the pathogenesis of TNBS-induced colitis in rats. METHODS: Thirty-two healthy adult Sprague-Dawley (SD) rats were randomly divided into four groups of eight each: normal, NS, model Ⅰ, model Ⅱ groups in our study. Rat colitis model was established through 2-,4-,6-trinitrobenzene sulfonic acid (TNBS) enema. At the end of four weeks, the macroscopical and histological changes of the colon were examined and mucosa myeloperoxidase (MPO)activities assayed. NF-~B p65 expression was determined by Western blot assessment in cytoplasmic and nuclear extracts of colon tissue, and the expressions of TNF-α (and ICAM-1 protein in colon tissue were examined by immunohistochemistry. The relativities between expression of NF-κB p65 and other parameters were analyzed. RESULTS: TNBS enema resulted in pronounced pathological changes of colonic mucosa in model Ⅱ group (macroscopic and histological injury indices 6.25&#177;1.39 and 6.24&#177;1.04, respectively), which were in accordance with the significantly elevated MPO activity (1.69+0.11). And the nuclear level of NF-κB and expression of TNF-α, ICAM-1 in rats of model Ⅱ group were higher than that of normal control(9.7&#177;1.96 vs 1.7&#177;0.15, 84.09&#177;14.52 vs 16.03&#177;6.21,77.69&#177;8.09 vs 13.41&#177;4.91 P&lt;0.01), Linear correlation analysis revealed that there were strong correlations between the nuclear level of NF-κB and the tissue positive expression of TNF-α and ICAM-1, MPO activities, macroscopical and histological indices in TNBS-induced colitis, respectively (r = 0.8235, 0.8780, 0.8572, 0.9152,0.8247; P&lt;0.05). CONCLUSION: NF-κB plays a pivotal role in the pathogenesis of ulcerative colitis, which might account for the up-regulation the expression of TNF-α and ICAM-1.     

The effects of colloidal bismuth tartrate on colitis induced by immune-complex in rabbits

AIM To observe the therapeutic effect of colloidal bismuth tartrate in an animal colitis model.METHODS Immune-complex colitis was induced in groups of rabbits by formalin, and two hours later0.85 mL heat-aggregated rabbit IgG was given intravenously through the ear cannula. Animals wereintracolonically treated with colloidal bismuth tartrate (BITNAL), and its effect was compared withsulfasalazine (SASP), indomethacin (IND) and bifidobiogen (BIFG). Animals were killed, the mucosalappearance was scored (0-4), and tissue saved for histological studies, the number of neutrophils present ininflamed colonic tissue was quantitated by the myeloperoxidase (MPO) activity assay, the production oflipoxygenase and cyclo-oxygenase products was monitored and eicosanoid production were assayed byincubation colonic specimens and the media for prostaglandin E2(PGE2), leukotriene (LTB4), thromboxaneB2(TXPe) were examined by radiommunoassay.RESULTS Immune-complex colitis was induced by formalin and IgG, colonic damage persisted for at least1 wk by macrography. Histologically, the inflammatory response included mucosal and submucosalinfiltration by polymorphonuclear leukocytes, macrophages, lymphocytes and fibroblasts, the macroscopic,persent 2 wk after IgG, was correlated with greatly increased PGE2, LTB4 and TXB2 compared with levels incontrols. Treatment with BITNAL (500 mg/kg) resulted in a lowered inflammation index, lowered MPOactivity and inhibited the increased formation of PGF-2, LTB4 and TXB2 by the inflamed colon, and IND(500 mg/kg) markedly inhibited prostanoid formation in both inflamed and control colon but did not reducetissue damage, SASP (500 mg/kg) also inhibited the formation of PGE2, LTB4 and TXB2 but the effectswere less marked. BIFG (400 mg/kg) did not significantly reduce the colonic injury and the media sythesizedby the rabbit colon.CONCLUSION BITAL provides better therapeutic effects in experimental colitis than anti-inflammatorydrug IND or SASP.     

COX-1 and COX-2 conversely promote and suppress ischemia-reperfusion gastric injury in mice

OBJECTIVE: Neutrophil activation followed by free radical production is a feature that is common to the various forms of gastric injury. However, the roles of cyclooxygenase (COX)-1 and -2 in neutrophil activation have yet to be clarified in the gastric mucosa. We examined the roles of both COX-1 and COX-2 in neutrophil activation and free radical production in ischemia-reperfusion (IR) injury in the gastric mucosa of mice. MATERIAL AND METHODS: Ischemia was induced by clamping the celiac artery for 30 min, then removing the clamp for 90 min. SC-560, a selective COX-1 inhibitor; NS-398, a selective COX-2 inhibitor; or rebamipide, a mucoprotective agent, was administered to mice 60 min before ischemia. Gastric damage was evaluated histologically and by measuring myeloperoxidase (MPO) activity. Expressions of COX protein and intercellular adhesion molecule (ICAM)-1 were evaluated by Western blot analysis and ELISA, respectively. Effects of these drugs on thiobarbituric acid reactive substances (TBARS) and gastric blood flow were also evaluated. RESULTS: COX-2 expression was induced in gastric mucosa 60 min after reperfusion, whereas COX-1 expression remained unaltered. Localization of COX-1 and ICAM-1 in IR-injured mucosa was observed mainly in endothelial cells, while COX-2 expression was detected in mesenchymal cells such as mononuclear cells, spindle-like cells and endothelial cells. SC-560 significantly decreased gastric blood flow at the reperfusion point and reduced gastric mucosal injury in IR mice. Furthermore, SC-560 pretreatment significantly reduced MPO activity, TBARS levels and ICAM-1 expression. In contrast, NS-398 significantly increased ICAM-1 expression, MPO activity and TBARS levels, and aggravated gastric damage in IR mice. Rebamipide pretreatment reduced both COX-2 expression and IR injury. CONCLUSIONS: In IR mice, COX-2 protects the gastric mucosa by down-regulating ICAM-1 expression, whereas COX-1 is involved in up-regulating reperfusion flow, thereby aggravating the mucosa.     

Rapid development of colitis in NSAID-treated IL-10-deficient mice

BACKGROUND & AIMS: Interleukin (IL)-10 is an anti-inflammatory and immune regulatory cytokine. IL-10-deficient mice (IL-10(-/-)) develop chronic inflammatory bowel disease (IBD), indicating that endogenous IL-10 is a central regulator of the mucosal immune response. Prostaglandins are lipid mediators that may be important mediators of intestinal inflammation. In this study we assessed the role of prostaglandins in the regulation of mucosal inflammation in the IL-10(-/-) mouse model of IBD. METHODS: Prostaglandin (PG) synthesis was inhibited with nonselective or cyclooxygenase (COX)-isoform selective inhibitors. Severity of inflammation was assessed histologically. Cytokine production was assessed by ribonuclease protection analysis and enzyme-linked immunosorbent assay. PGE(2) levels were assessed by enzyme immunoassay. COX-1 and COX-2 expression was assessed by Western blot analysis. RESULTS: Nonsteroidal anti-inflammatory drug (NSAID) treatment of wild-type mice had minimal effect on the colon. In contrast, NSAID treatment of 4-week-old IL-10(-/-) mice resulted in rapid development of colitis characterized by infiltration of the lamina propria with macrophages and interferon gamma-producing CD4(+) T cells. Colitis persisted after withdrawal of the NSAID. NSAID treatment decreased colonic PGE(2) levels by 75%. Treatment of IL-10(-/-) mice with sulindac sulfone (which does not inhibit PG production) did not induce colitis whereas the NSAID sulindac induced severe colitis. COX-1- or COX-2-selective inhibitors used alone did not induce IBD in IL-10(-/-) mice. However, the combination of COX-1- and COX-2-selective inhibitors did induce colitis. CONCLUSIONS: NSAID treatment of IL-10(-/-) mice results in the rapid development of severe, chronic IBD. Endogenous PGs are important inhibitors of the development of intestinal inflammation in IL-10(-/-) mice.     

Chronic cyclooxygenase-2 inhibition blunts low sodium-stimulated renin without changing renal haemodynamics

BACKGROUND: Cyclooxygenase-2 (COX-2), the inducible isoform of cyclooxygenase, is found in the macula densa of the renal cortex and is upregulated by dietary sodium restriction. Because of this discrete cortical localization, we hypothesized that COX-2 plays a role in the chronic stimulation of renin via the macula densa pathway. METHODS: We examined the effect of the selective COX-2 inhibitor NS 398 in male Sprague-Dawley rats. RESULTS: A low sodium diet (0.02% NaCl) for 14 days elevated plasma-renin activity (PRA) nine-fold, from 6.1 +/- 2.0 to 54.9 +/- 6.5 ng angiotensin I (Ang I)/ml per h (P < 0.0001). Selective COX-2 inhibition with NS 398 had no effect on PRA in animals on normal sodium (5.1 +/- 1.3 ng Ang I/ml per h), but decreased PRA by 41% in sodium-restricted rats, to 33.3 +/- 3.6 ng Ang I/ml per h (P < 0.05). Chronic treatment with NS 398 did not decrease renal renin content (31.8 +/- 1.8 versus 33.5 +/- 2.6 ng Ang I/ mg per h, with NS 398 versus controls), nor did it influence systemic blood pressure or renal haemodynamics. Neither urinary sodium excretion nor prostaglandin (PG)E2 excretion was altered in rats given NS 398. Chronic treatment with the non-selective COX inhibitor indomethacin during sodium restriction over 5 days reduced PRA by 35%, from 29.36 +/- 4.81, to 19.13 +/- 2.88 ng Ang I/ml per h (P < 0.05). Indomethacin had no effect on blood pressure or renal blood flow but reduced urinary PGE2 excretion by 70%. CONCLUSIONS: One component of the chronic stimulation of PRA by dietary sodium restriction via the macula densa pathway appears to involve the induction of COX-2.     

Protective effect of angelica sinensis polysaccharide on experimental immunological colon injury in rats

AIM: To study the effect of angelica sinensis polysaccharide (ASP) on immunological colon injury and its mechanisms in rats.METHODS: Immunological colitis model of rats was induced by intracolon enema with 2, 4, 6-trinitrobenzene sulfonic acid (TNBS) and ethanol. The experimental animals were randomly divided into normal control, model control, 5-aminosalicylic acid therapy groups and three doses of ASP therapy groups. The 6 groups were treated intracolonically with normal saline, normal saline, 5-aminosalicylic acid (100 mg.kg-1), and ASP daily (8:00 am) at the doses of 200, 400 and 800 mg.kg-1 respectively for 21 days 7 d following induction of colitis. The rat colon mucosa damage index (CMDI), the histopathological score (HS), the score of occult blood test (OBT), and the colonic MPO activity were evaluated. The levels of SOD, MDA, NO, TNF-α, IL-2 and IL-10 in colonic tissues were detected biochemically and immunoradiometrically. The expressions of TGF-β and EGF in colonic tissues were also determined immunochemically.RESULTS: Enhanced colonic mucosal injury, inflammatory response and oxidative stress were observed in colitis rats,which manifested as significant increases of CMDI, HS, OBT,MPO activity, MDA and NO contents, as well as the levels of TNF-α and IL-2 in colonic tissues, although colonic TGF-β protein expression, SOD activity and TL-10 content were significantly decreased compared with the normal control (P＜0.01). However, these parameters were found to be significantly ameliorated in colitis rats treated intracolicly with ASP at the doses of 400 and 800 mg@kg-1 (P＜0.05-0.01).Meantime, colonic EGF protein expression in colitis rats was remarkably up-regulated.CONCLUSION: ASP has a protective effect on immunological colon injury induced by TNBS and ethanol enema in rats,which was propably due to the mechanism of antioxidation,immunomodulation and promotion of wound repair.     

Excessive production of reactive oxygen metabolites by inflamed colon: analysis by chemiluminescence probe.

Reactive oxygen metabolites (ROMs) are involved in inflammatory diseases and are postulated to contribute to tissue injury in colitis. To determine whether excessive ROMs are generated by inflamed colonic mucosa and to identify possible sources and type of ROMs, mucosal ROMs were estimated in rats and humans using a chemiluminescence probe. Colitis was induced in rats by intracolonic injection of acetic acid or intraperitoneal injection of mitomycin C. Intact, inflamed colon in rats produced more ultraweak chemiluminescence than normal colon. Inflamed mucosal scrapings from both rat models produced significantly more luminol-enhanced chemiluminescence. Addition of catalase, an H2O2 scavenger, or azide, a myeloperoxidase inhibitor, into the media significantly decreased chemiluminescence from inflamed mucosal scrapings. Indomethacin, an antioxidant cyclo-oxygenase inhibitor, also decreased chemiluminescence, but MK-866, a 5-lipoxygenase inhibitor, had no effect. Colonic biopsy specimens obtained during colonoscopy from patients with ulcerative colitis also produced more catalase-inhibitable chemiluminescence than normal colonic mucosa. These data indicate that excessive ROMs are produced by inflamed colonic mucosa in both humans and rats, which may contribute to tissue injury.     

Aggravation by Selective COX-1 and COX-2 Inhibitors of Dextran Sulfate Sodium (DSS)-Induced Colon Lesions in Rats

We examined the effect of cyclooxygenase (COX) inhibitors on dextran sulfate sodium (DSS)-induced ulcerative colitis in rats and investigated the role of COX isozymes in the pathogenesis of this model. Experimental colitis was induced by treatment with 2.5% DSS in drinking water for 6 days. Indomethacin (a nonselective COX inhibitor), SC-560 (a selective COX-1 inhibitor), or celecoxib (a selective COX-2 inhibitor) was given PO twice daily for 6 days, during the first 3 or last 3 days of the experimental period. Daily treatment with 2.5% DSS for 6 days caused damage to the colon, with a decrease in body weight gain and colon length as well as an increase of myeloperoxidase (MPO) activity. All COX inhibitors given for 6 days significantly worsened the severity of DSS-induced colonic damage with increased MPO activity. The aggravation was also observed by SC-560 given for the first 3 days or by celecoxib given for the last 3 days. The expression of COX-2 mRNA in the colon was upregulated on day 3 during DSS treatment, with significant increase of prostaglandin E₂ PGE₂ production. The PGE₂ content on day 3 during DSS treatment was inhibited by both indomethacin and SC-560, but not by celecoxib; on day 6 it was suppressed by both indomethacin and celecoxib, but not SC-560. These results suggest that endogenous prostaglandins (PGs) afford protection against colonic ulceration, yet the COX isozyme responsible for the production of PGs differs depending on the stage of ulceration; COX-1 in the early stage and COX-2 in the late stage.     

Recombinant interleukin-1 receptor antagonist blocks the proinflammatory activity of endogenous interleukin-1 in rabbit immune colitis.

The effects of human recombinant interleukin-1 receptor antagonist (IL-1ra) on inflammation, tissue damage, and production of inflammatory mediators in rabbit formalin-immune complex colitis were examined. Treatment of rabbits with intravenous administration of IL-1ra before and periodically throughout the first 43 hours after the induction of colitis (total 7 doses) suppressed inflammation and tissue damage in a dose-related manner. Maximum inhibition of inflammatory index (from 3.0 +/- 0.3 to 0.8 +/- 0.3, P less than 0.001), edema (from 2.3 +/- 0.2 to 0.6 +/- 0.2, P less than 0.001), percent of mucosal necrosis (from 44% +/- 7% to 7% +/- 3%, P less than 0.02) and myeloperoxidase (MPO) activity (from 4.9 +/- 1.0 U/g to 1.0 +/- 0.3 U/g, P less than 0.001) occurred with the dose of 5 mg/kg. Colonic prostaglandin E2 (PGE2) and leukotriene B4 (LTB4) production, measured in rectal dialysates by specific radioimmunoassays, was also dose dependently suppressed (from 1124 +/- 319 pg/mL to 190 +/- 75 pg/mL, P less than 0.001 and 568 +/- 192 pg/mL to 92 +/- 51 pg/mL, P less than 0.001, respectively, at 5 mg/kg). In contrast, colonic IL-1 alpha tissue levels measured by a specific radioimmunoassay after tissue extraction were similar in all groups. When only two doses of IL-1ra, 10 minutes before and 3 hours after the induction of colitis were given, there was no longer an inhibitory effect on inflammation or production of inflammatory mediators. However, delaying the initial IL-1ra treatment 3 hours after the induction of colitis (total 6 doses) was effective in reducing inflammatory index (by 60%), MPO activity (by 79%), PGE2 (by 62%), and LTB4 (by 72%) whereas colonic IL-1 alpha levels were unchanged compared with vehicle-treated animals. These studies show the ability of human recombinant IL-1 receptor antagonist to suppress the proinflammatory activity of IL-1 produced in the colon during the induction and progression of rabbit immune complex colitis.     

Protective effect of melatonin in acetic acid induced colitis in rats

Possible protective effects of exogenous melatonin on colonic inflammation were studied in rats. Colitis was induced by intracolonic (i.c.) instillation of 4% acetic acid (AA) and the resulting injury was assessed after 1 and 48 hr. Diffuse hyperemia and bleeding with erosions and ulcerations were observed in the colons of vehicle-treated rats. Melatonin administered in doses of 5 and 10 mg/kg reduced significantly the extent of gross mucosal damage after intraperitoneal as well as i.c. dosing. The inflammation induced increase in colonic wet weight was also reduced by melatonin treatment. In the early phase of colonic inflammation (60 min), melatonin partly prevented the decrease of reduced glutathione (GSH) content and limited lysosomal enzyme, N-acetyl-glucosaminidase and cathepsin D, activities induced by AA, with no changes in proteins or acid phosphatase activity. Increase of myeloperoxidase activity (MPO) caused by colonic inflammation was prevented by melatonin given i.c. As observed 48 hr after AA exposure, there was no difference between the effect of vehicle and melatonin on the content of GSH. Colitis did not influence the melatonin content of the colon. After administration of exogenous melatonin, plasma, pineal and gut melatonin tended to increase. The results indicate that melatonin participates in various defense mechanisms against colonic inflammatory processes by preserving the important endogenous antioxidant reserve of GSH, by preventing lysosomal enzyme disruption, by inhibiting enhanced MPO activity, thus reducing the extent of colonic damage, mainly in the early phase of colitis.     

MUC Genes Are Differently Expressed During Onset and Maintenance of Inflammation in Dextran Sodium Sulfate-Treated Mice

Colonic mucosal protection is provided by mucous gel, mainly composed of secreted (Muc2) and membrane-bound (Muc1, Muc3, Muc4) mucins. Our aim was to determine the expression profile of secreted and membrane-bound mucins in experimental dextran sulfate sodium (DSS)-induced colitis. Acute colitis was induced in Balb/C mice by oral administration of 1.0% DSS (5 days) and chronic colitis was maintained by subsequent 0.15% DSS treatment (28 days). Clinical symptoms (mortality, weight gain), stool scores, and MPO activity confirmed the inflammatory state in the two phases of colitis. Muc2 gene expression was not modified by colitis, whereas Muc3 gene expression was increased (×2) only in the cecum and the distal colon of mice after acute colitis. Muc1 and Muc4 mRNA levels were more significantly increased in the cecum (×8–10) than in colonic segments (×4) after acute colitis. TFF3 involved in mucosal repair was up-regulated during colitis induction. These results indicate that Muc and TFF3 genes are regulated early in inflammation and suggest that their mRNA levels could be used as early markers of inflammation.