The effect of alpha-melanocyte stimulating hormone on endotoxin-induced intestinal injury.

We investigated the effect of alpha-melanocyte stimulating hormone (alpha-MSH) on endotoxin-induced intestinal inflammation and the role of nitric oxide and prostaglandins in this response. alpha-MSH treatment (25 microg/rat, intraperitoneally (i.p.); twice daily) reduced the severity of the lesions macroscopically and microscopically. This protective effect was found to be confined mainly to the distal ileum. These lesions were reversed by pretreatment with the non-selective COX inhibitor indomethacin (10 mg/kg, subcutaneously (s.c.)) but not by the selective COX-2 inhibitor nimesulide (3 mg/kg, s.c.), the NO donor sodium nitroprusside (4 mg/kg, i.v.) or the iNOS inhibitor dexamethasone (3 mg./kg, i.p.) at macroscopic level and reversed by Indo or Dex at microscopic level. Increased peroxidase activity -index of tissue neutrophil infiltration- in the distal ileum of LPS-treated rats was decreased by alpha-MSH and this effect was reversed by pretreatment with Indo. In conclusion, the neuropeptide alpha-MSH has a beneficial effect on endotoxin-induced distal intestinal lesions by a mechanism which probably involves nitric oxide and COX-1 derived prostaglandins.     

The effect of α-melanocyte stimulating hormone on endotoxin-induced intestinal injury

We investigated the effect of α-melanocyte stimulating hormone (α-MSH) on endotoxin-induced intestinal inflammation and the role of nitric oxide and prostaglandins in this response. α-MSH treatment (25 μg/rat, intraperitoneally (i.p.); twice daily) reduced the severity of the lesions macroscopically and microscopically. This protective effect was found to be confined mainly to the distal ileum. These lesions were reversed by pretreatment with the non-selective COX inhibitor indomethacin (10 mg/kg, subcutaneously (s.c.)) but not by the selective COX-2 inhibitor nimesulide (3 mg/kg, s.c.), the NO donor sodium nitroprusside (4 mg/kg, i.v.) or the iNOS inhibitor dexamethasone (3 mg./kg, i.p.) at macroscopic level and reversed by Indo or Dex at microscopic level. Increased peroxidase activity -index of tissue neutrophil infiltration- in the distal ileum of LPS-treated rats was decreased by α-MSH and this effect was reversed by pretreatment with Indo. In conclusion, the neuropeptide α-MSH has a beneficial effect on endotoxin-induced distal intestinal lesions by a mechanism which probably involves nitric oxide and COX-1 derived prostaglandins.     

Effects of proanthocyanidins from grape seed on treatment of recurrent ulcerative colitis in rats

The aim of the present study was to investigate the therapeutic effect and mechanism of proanthocyanidins from grape seed (GSPE) in the treatment of recurrent ulcerative colitis (UC) in rats. To induce recurrent colitis, rats were instilled with 2,4,6-trinitrobenzenesulfonic acid (TNBS) (80?mg/kg) into the colon through the cannula in the first induced phase, and then the rats were instilled a second time with TNBS (30?mg/kg) into the colon on the sixteenth day after the first induction UC. Rats were intragastrically administered GSPE (200?mg/kg) per day for 7?days after twice-induced colitis by TNBS. Sulfasalazine at 500?mg/kg was used as a positive control drug. Rats were killed 7?days after GSPE treatment. The colonic injury and inflammation were assessed by macroscopic and macroscopic damage scores, colon weight/length ratio (mg/cm), and myeloperoxidase activity. Then, superoxide dismutase, glutathione peroxidase, inducible nitric oxide synthase (iNOS) activities, and the levels of malonyldialdehyde, glutathione, and nitric oxide in serum and colonic tissues were measured. Compared with the recurrent UC group, GSPE treatment facilitated recovery of pathologic changes in the colon after induction of recurrent colitis, as demonstrated by reduced colonic weight/length ratio and macroscopic and microscopic damage scores. The myeloperoxidase and iNOS activities with malonyldialdehyde and nitric oxide levels in serum and colon tissues of colitis rats were significantly decreased in the GSPE group compared with those in the recurrent UC group. In addition, GSPE treatment was associated with notably increased superoxide dismutase, glutathione peroxidase activities, and glutathione levels of colon tissues and serum of rats. GSPE exerted a protective effect on recurrent colitis in rats by modifying the inflammatory response, inhibiting inflammatory cell infiltration and antioxidation damage, promoting damaged tissue repair to improve colonic oxidative stress, and inhibiting colonic iNOS activity to reduce the production of nitric oxide.     

Quebrachitol-induced gastroprotection against acute gastric lesions: Role of prostaglandins, nitric oxide and KATP channels

The effect of Quebrachitol (2-O-methyl-l-inositol), a bioactive component from Magonia glabrata fruit extract was investigated against gastric damage induced by absolute ethanol (96%, 0.2 ml/animal) and indomethacin (30 mg/kg, p.o.), in mice. Quebrachitol at oral doses of 12.5, 25, and 50 mg/kg markedly attenuated the gastric lesions induced by ethanol to the extent of 69%, 64%, and 53% and against indomethacin by 55%, 59%, and 26%, respectively. While pretreatment with TRPV1 antagonist capsazepine (5 mg/kg, i.p.) failed to block effectively the gastroprotective effect of quebrachitol (25 mg/kg) against ethanol damage, the non-selective cyclooxygenase inhibitor indomethacin (10 mg/kg, p.o.), almost abolished it. Furthermore, quebrachitol effect was significantly reduced in mice pretreated with l-NAME, or glibenclamide, the respective inhibitors of nitric oxide synthase and K⁺_ATP channel activation. Thus we provide the first evidence that quebrachitol reduces the gastric damage induced by ethanol and indomethacin, at least in part, by mechanisms that involve endogenous prostaglandins, nitric oxide release, and or the activation of K⁺_ATP channels.     

Indomethacin worsens and a leukotriene biosynthesis inhibitor accelerates mucosal healing in rat colitis.

The implication of leukotrienes as mediators of inflammation and recent evidence that prostaglandin analogues provide a beneficial effect during experimental colitis led to the speculation that (i) leukotrienes may be injurious and (ii) prostaglandins may be protective to colonic mucosa. Using a 2% acetic acid induced rat colitis model, we administered specific cyclooxygenase (indomethacin) and leukotriene biosynthesis inhibitors (MK-886) to examine the effect of endogenous prostaglandins and leukotrienes on colonic macroscopic injury, mucosal inflammation as measured by myeloperoxidase activity, net in vivo intestinal fluid absorption, and colonic PGE2 and LTB4 levels as measured by in vivo rectal dialysis. Indomethacin treatment prior to induction of colitis reduced endogenous mucosal PGE2 levels and exacerbated macroscopic ulceration and net fluid absorption. Addition of the exogenous PGE1 analogue misoprostol to the indomethacin-exacerbated colitis completely healed colonic macroscopic ulceration and inflammation but only partially improved fluid absorptive injury. The specific leukotriene biosynthesis inhibitor MK-886 administered prior to induction of colitis healed macroscopic ulceration and inflammation but not fluid absorptive injury. This mucosal reparative effect of MK-886 occurred at a dose that reduced colonic LTB4 synthesis while concomitantly enhancing PGE2 levels. Combining MK-886 with misoprostol treatment improved not only macroscopic ulceration and inflammation but also provided a synergistic effect that maintained net colonic fluid absorption at noncolitic control levels. These studies suggest that, during the induction of experimental colitis, endogenous prostaglandins play a pivotal role in providing a mucosal healing effect, and that leukotriene biosynthesis inhibitor may manifest part of its beneficial effect by shifting arachidonic acid metabolism towards production of prostaglandins.     

Comparative gastrointestinal toxicity of selective cyclooxygenase (COX-2) inhibitors

Cyclooxygenase (COX-2) inhibitors were developed with the hope that they will cause fewer gastrointestinal adverse effects. Ability of selective as well as nonselective COX inhibitors to alter ischemia-reperfusion induced damage of gastric mucosa and hapten-induced colitis in rats has been compared. Celecoxib (10, 20 and 40 mg/kg(-l)) was significantly more potent at aggravating ischemia-reperfusion injury as compared to nimesulide. Similarly, celecoxib was found to maximally potentiate TNBS-induced colitis, followed by nimesulide and indomethacin. Celecoxib at its highest dose produced maximum deep histological injury. This paradoxic ulcer and colitis aggravating effect of selective COX-2 inhibitors may be explained by suppression of protective prostaglandins generated as a consequence of COX-2 induction in inflammatory states.     

The anti-inflammatory effect of leptin on experimental colitis: involvement of endogenous glucocorticoids

The present study was designed to compare the effect of leptin on acute colonic inflammation with that of acute stress exposure, which acts via the hypothalamic-pituitary-adrenal (HPA) axis. Sprague-Dawley rats of both sexes were administered intrarectally with acetic acid. Either leptin (10 microg/kg; i.p.) or saline was injected immediately before and 6 h after the induction of colitis. A group of rats was exposed to water avoidance stress (WAS) for 30 min at the 6th h of colitis induction. RU-486 (2 mg/kg; i.p.), a glucocorticoid receptor antagonist, was injected intraperitoneally, at 12 and 1 h before the initial leptin injection, and at 1 h before the second leptin injection or exposure to WAS. Rats were decapitated at 24 h and the distal 8 cm of the colon were removed for macroscopic and microscopic scoring, determination of tissue wet weight index (WI) and tissue myeloperoxidase activity (MPO). Acetic acid-induced colitis significantly increased macroscopic and microscopic damage scores, WI and MPO, compared to control group. Exposure to acute WAS or treatment with leptin reduced the elevations in damage scores, WI and MPO induced by colitis, but no additive inhibitory effect was observed when WAS and leptin were applied together. RU-486 treatment reversed the inhibitory effects of leptin or WAS on colonic inflammation. Our results demonstrate that exogenous leptin mimics the effects of HPA axis activation on colitis-induced inflammatory process. The results also suggest that the anti-inflammatory effect of leptin involves a tissue neutrophil-dependent mechanism and is dependent on the release of glucocorticoids.     

Cytoprotective effects of met-enkephalin and alpha-MSH on ethanol induced gastric lesions in rats.

We used the rat model of ethanol induced gastric lesions to measure cytoprotective effects of neuropeptides met-enkephalin and alpha-melanocyte stimulating hormone (alpha-MSH). The lesions were induced with i.g. application of 1 ml 96% ethanol. The peptides were given i.p. 1 h before the ethanol. Sacrifice was made 1 h after ethanol application and hemorrhagic gastric area was assessed in mm(2). alpha-MSH and met-enkephalin exhibited significant and additive cytoprotective effects. The protective effects of alpha-MSH were significantly stronger than of met-enkephalin. Almost total absence of lesions was obtained with met-enkephalin and alpha-MSH mixture 10:1 (10 mg/kg met-enkephalin and 1 mg/kg alpha-MSH). The addition of indomethacin (5 mg/kg s.c.) almost completely abolished the effect of met-enkaphalin, while alpha-MSH mediated cytoprotection was weakened but still present. Interestingly, indomethacin also blocked almost completely the cytoprotective effects of met-enkephalin and alpha-MSH mixture. The latter result may have a practical consequence for the clinical trials in which met-enkephalin and alpha-MSH could be used in combination with non-steroidal anti-inflammatory drugs.     

Effect of quercitrin on the early stages of hapten induced colonic inflammation in the rat

Quercitrin is a flavonoid with antiinflammatory activity in experimental colitis, associated with an antioxidative action and amelioration of water absorption in vivo. However, its mechanism of action is unclear. This study focuses on the effect of quercitrin (1 and 5 mg/kg) in the early stages (24 h) of trinitrobenzene sulfonic acid colitis. Treatment with the flavonoid prevented the increase in colonic malondialdehyde and inhibited nitric oxide synthase and alkaline phosphatase activity but had no significant effects on observable damage. No effect on neutrophil infiltration (myeloperoxidase) was observed. On the other hand, quercitrin exerted complex effects on colonic hydroelectrolytic transport, showing a slight potentiation of water absorption in vivo (5 mg/kg) as well as a normalization of carbachol stimulated electrogenic ionic transport in the proximal colon in vitro (5 mg/kg). It is concluded that the beneficial effects of quercitrin on trinitrobenzene sulfonic acid chronic colitis arise from an early downregulation of the inflammatory cascade that is associated with amelioration of the disturbances in hydroelectrolytic transport.     

Prophylactic potential of montelukast against mild colitis induced by dextran sulphate sodium in rats.

Cysteinyl leukotrienes play a part in inflammatory processes such as inflammatory bowel diseases. The present study aimed to evaluate the effects of the cys-LT-1 receptor antagonist montelukast on a mild colitis model in rats. Colitis was induced by administrating 4% dextran sulphate sodium (DSS, MW 45,000) in drinking water for 9 days. Montelukast (10 mg/kg/day) or vehicle was given by gastric gavage once daily simultaneously with DSS administration. A healthy control group receiving water as drinking fluid and vehicle by gastric gavage was included. Body weight loss, consistency of faeces (loose/diarrhoea) and occult blood in the faeces/ gross bleeding were assessed on days 6 - 9. After sacrifice, the following were assessed: colonic histology, the expression of inducible nitric oxide synthase, macrophage/monocyte marker ED1, cyclooxygenase-1 and cyclooxygenase-2, as well as the production of leukotriene B(4) and E(4), prostaglandin E(2), its metabolite bicyclic-prostaglandin E(2) and thromboxane B(2) in the colonic tissue incubation in vitro. Rats receiving DSS exhibited bloody diarrhoea from day 6 onwards. Montelukast significantly reduced the occult blood in the faeces/ gross bleeding, maintained normal body weight gain and tended to decrease the ratio of leukotriene B(4)/ prostaglandin E(2) production in the colon in vitro. The results indicate that montelukast has some potential to ameliorate mild experimental colitis induced by DSS.     

Roles of capsaicin-sensitive sensory nerves, endogenous nitric oxide, sulfhydryls, and prostaglandins in gastroprotection by momordin Ic, an oleanolic acid oligoglycoside, on ethanol-induced gastric mucosal lesions in rats.

The roles of capsaicin-sensitive sensory nerves (CPSN), endogenous nitric oxide (NO), sulfhydryls (SHs), prostaglandins (PGs) in the gastroprotection by momordin Ic, an oleanolic acid oligoglycoside isolated from the fruit of Kochia scoparia (L.) SCHRAD., on ethanol-induced gastric mucosal lesions were investigated in rats. Momordin Ic (10 mg/kg, p.o.) potentially inhibited ethanol-induced gastric mucosal lesions. The effect of momordin Ic was markedly attenuated by the pretreatment with capsaicin (125 mg/kg in total, s.c., an ablater of CPSN), N(G)-nitro-L-arginine methyl ester (L-NAME, 70 mg/kg, i.p., an inhibitor of NO synthase), N-ethylmaleimide (NEM, 10 mg/kg, s.c., a blocker of SHs), or indomethacin (10 mg/kg, s.c., an inhibitor of PGs biosynthesis). The attenuation of L-NAME was abolished by L-arginine (300 mg/kg, i.v., a substrate of NO synthase), but not by D-arginine (300 mg/kg, i.v., the enatiomer of L-arginine). The effect of the combination of capsaicin with indomethacin, NEM, or L-NAME was not more potent than that of capsaicin alone. The combination of indomethacin and NEM, indomethacin and L-NAME, or indomethacin and NEM and L-NAME increased the attenuation of each alone. These results suggest that CPSN play an important role in the gastroprotection by momordin Ic on ethanol-induced gastric mucosal lesions, and endogenous PGs, NO, and SHs interactively participate, in rats.     

Prevention by aluminium phosphate of gastric lesions induced by ethanol in the rat: role of endogenous prostaglandins and sulfhydryls

This study was designed to demonstrate the cytoprotective effect of an antacid containing aluminium phosphate (Phosphalugel) against ethanol-induced gastric injury in the rat and to determine whether this cytoprotective effect is mediated by endogenous prostaglandins and sulfhydryls. We have quantitatively evaluated gastric mucosal lesions using macroscopic and histological techniques one hour after ethanol administration. Two ml of aluminium phosphate given orally one hour before administration of 2 ml of 100% ethanol significantly (p less than 0.01) reduced the area of macroscopic lesions induced by ethanol (3.3 +/- 0.9%) when compared to distilled water (20 +/- 4.8%). The histological study showed that aluminium phosphate prevented deep tissue necrosis. However, it did not protect surface epithelial cells against ethanol injury. Pretreatment with indomethacin, 5 mg/kg sc one hour before aluminium phosphate, slightly but significantly (p less than 0.05) reduced the cytoprotective effect of aluminium phosphate. Macroscopic lesions occupied 4.3 +/- 0.94% and 1.88 +/- 0.41% of total mucosal area in indomethacin group and in vehicle group, respectively. On the other hand, the sulfhydryl blocker, N-ethyl-maleimide, 10 mg/kg sc, given one hour before aluminium phosphate, completely abolished the cytoprotective effect of aluminium phosphate (32.92 +/- 4.85% in N-ethyl-maleimide group versus 3.78 +/- 1.41% in vehicle group; p less than 0.01). These results show that aluminium phosphate has a cytoprotective effect against ethanol injury in the rat. This property appears to be mediated by both endogenous prostaglandins and sulfhydryls.     

Gastroprotective potential of risperidone, an atypical antipsychotic, against stress and pyloric ligation induced gastric lesions

Risperidone has been used in some stress disorders and may be potentially protective against stress-induced gastric lesions. Thus, the aim of the present study is to investigate, whether risperidone, a D(2) receptor and 5-HT(2A) receptor antagonist, would be able to result in gastroprotective effect in stress-induced lesions and also explore the possible mechanism of action behind its gastroprotective activity. Gastroprotective activity of risperidone was evaluated both by single treatment and 21 days repeated (0.03, 0.1, 0.3 and 1mg/kg, p.o.) treatment in the cold restraint stress (CRS) model and 21 days repeated treatment in the pyloric ligation (PL) model and compared with that of sulpiride (D(2) receptor antagonist) and ketanserin (5-HT(2) receptor antagonist) as standard. Histopathological assessment was done to evaluate the gastroprotective activity of risperidone in CRS model. The roles of nitric oxide (NO), sulfhydryl (SH) group, ATP-sensitive K(+) channels (K(ATP) channels) and prostaglandins (PGs) in the gastroprotective effect of risperidone against CRS were also investigated. PGE(2), hexosamine as a marker of mucus barrier and microvascular permeability were also estimated. Results show that repeated treatment of risperidone, sulpiride and ketanserin exhibited a gastroprotective effect against CRS-induced lesions while single administration of risperidone was found to be ineffective. Moreover, repeated treatment of risperidone and ketanserin was found to be ineffective in case of PL in contrast to sulpiride. Risperidone pretreatment reverses the stress induced alteration in hexosamine, PGE(2) and microvascular permeability. Pretreatment with l-NAME, NEM, glibenclamide and indomethacin reversed the gastroprotective effect of risperidone. The results suggest that risperidone has significant gastroprotective effects in CRS-induced gastric lesions models, which appears to be mediated by endogenous NO, SH, PGs and K(ATP) channel opening.     

Dual role of endogenous nitric oxide in development of dextran sodium sulfate-induced colitis in rats.

The role of nitric oxide (NO) in the etiology of ulcerative colitis is controversial with reports of the improvement and aggravation of colonic lesions by inducible NO synthase (iNOS) inhibitors. In the present study, we compared the effect of the selective iNOS inhibitor aminoguanidine and the nonselective NOS inhibitor N(G)-nitro-L-arginine methyl ester (L-NAME) on a dextran sulfate sodium (DSS)-induced model of colitis in rats. Experimental colitis was induced by a 3% DSS-solution added to drinking water for 7 days. Aminoguanidine (5 approximately 20 mg/kg) and L-NAME (10 mg/kg) were administered p.o. twice daily for the first 3 days, the last 3 days or all 6 days of DSS treatment. Body weight and severity of colitis (diarrhea, bloody feces) were observed over a period of 7 days. DSS treatment resulted in severe colonic lesions, accompanied by diarrhea, bloody feces, decrease of body weight and colon shortening. All of the parameters investigated improved significantly with aminoguanidine treatment at 20 mg/kg for 6 days or the last 3 days of DSS-treatment, but L-NAME did not significantly affect the colitis during these periods. When L-NAME or aminoguanidine was given in the first 3 days of DSS treatment, the colonic lesions were slightly aggravated by L-NAME but not affected by aminoguanidine. The expression of iNOS mRNA was observed from the 3(rd) day of DSS treatment. These results suggested that endogenous NO exerts a biphasic influence on DSS-induced colitis, depending on the NOS isoenzyme; a beneficial effect of NO derived from constitutive NOS and a detrimental effect of NO produced by iNOS in the development of colitis.     

iNOS-Dependent Increase in Colonic Mucus Thickness in DSS-Colitic Rats

Aim

To investigate colonic mucus thickness in vivo in health and during experimental inflammatory bowel disease.

Methods

Colitis was induced with 5% DSS in drinking water for 8 days prior to experiment, when the descending colonic mucosa of anesthetized rats was studied using intravital microscopy. Mucus thickness was measured with micropipettes attached to a micromanipulator. To assess the contributions of NOS and prostaglandins in the regulation of colonic mucus thickness, the non-selective NOS-inhibitor L-NNA (10 mg/kg bolus followed by 3 mg/kg/h), the selective iNOS-inhibitor L-NIL (10 mg/kg bolus followed by 3 mg/kg/h) and the non-selective COX-inhibitor diclofenac (5 mg/kg) were administered intravenously prior to experiment. To further investigate the role of iNOS in the regulation of colonic mucus thickness, iNOS −/− mice were used.

Results

Colitic rats had a thicker firmly adherent mucus layer following 8 days of DSS treatment than untreated rats (88±2 µm vs 76±1 µm). During induction of colitis, the thickness of the colonic mucus layer initially decreased but was from day 3 significantly thicker than in untreated rats. Diclofenac reduced the mucus thickness similarly in colitic and untreated rats (−16±5 µm vs −14±2 µm). While L-NNA had no effect on colonic mucus thickness in DSS or untreated controls (+3±2 µm vs +3±1 µm), L-NIL reduced the mucus thickness significantly more in colitic rats than in controls (−33±4 µm vs −10±3 µm). The importance of iNOS in regulating the colonic mucus thickness was confirmed in iNOS−/− mice, which had thinner colonic mucus than wild-type mice (35±3 µm vs 50±2 µm, respectively). Furthermore, immunohistochemistry revealed increased levels of iNOS in the colonic surface epithelium following DSS treatment.

Conclusion

Both prostaglandins and nitric oxide regulate basal colonic mucus thickness. During onset of colitis, the thickness of the mucus layer is initially reduced followed by an iNOS mediated increase.     

Efficacy of Setarud (IMod), a novel drug with potent anti-toxic stress potential in rat inflammatory bowel disease and comparison with dexamethasone and infliximab

The inflammatory bowel disease (IBD) is an idiopathic, immune-mediated and chronic intestinal condition. In the present study, the effect of Setarud (IMOD), a novel natural drug with known immunomodulatory, anti-inflammatory and antioxidant properties was investigated in experimental colitis in rats and compared with the dexamethasone and infliximab. Immunologic colitis was induced by intracolonic administration of a mixture of trinitrobenzene sulfonic acid (TNBS) and absolute ethanol in male Wistar rats. Animals were divided into 6 groups of sham (normal group), control (vehicle-treated), positive control (dexamethasone 1 mg/kg/day given orally and infliximab 5 mg/kg/day given subcutaneously) and 3 Setarud-treated groups (13.3, 20, 30 mg/kg/day given intraperitoneally). The treatment continued for 14 consecutive days and then animals were decapitated on the day 15 and distal colons were removed for macroscopic, microscopic, and biochemical assays. Biochemical markers, including TNF-alpha, IL-1beta, ferric reducing/antioxidant power (FRAP), myeloperoxidase (MPO) activity and thiobarbitoric acid-reactive substance (TBARS) were measured in the homogenate of colonic tissue. A remarkable reduction in macroscopic and histological damage scores was observed in the animals treated with Setarud. These findings were confirmed by decreased levels of TNF-alpha, interleukin-1beta, MPO activity and TBARS, and raised levels of FRAP in the colon tissue. These observations confirmed the immunomodulatory, anti-inflammatory and antioxidant properties of Setarud in experimental colitis, which was comparable to those of dexamethasone and infliximab.     

The role of cyclooxygenase inhibition in the effect of alpha-melanocyte-stimulating hormone on reactive oxygen species production by rat peritoneal neutrophils

The effect of alpha-MSH on reactive oxygen species (ROS) production by rat peritoneal neutrophils and the effect of cyclooxygenase (COX) inhibition were investigated using the chemiluminescence (CL) technique. Cells were obtained by peritoneal lavage 4h after administration of oyster glycogen to rats and were stimulated with lipopolysaccharide (LPS) from Salmonella enderitidis and phorbol 12-myristate 13-acetate (PMA). The increasing concentrations of alpha-MSH (10(-12)-10(-6) M) were added to stimulated cells alone or along with the COX inhibitors indomethacin, ketorolac or nimesulide (10(-8)-10(-5) M). Luminol and lucigenin CL levels were significantly increased in cells stimulated with LPS and PMA compared to unstimulated ones. alpha-MSH significantly reduced lucigenin CL values and this effect was completely reversed in the presence of indomethacin (10(-8) and 10(-7) M). In conclusion, alpha-MSH inhibits the production of superoxide radicals by activated rat peritoneal neutrophils and COX contributes to this effect.     

Endothelin-1, interleukin-4 and nitric oxide synthase modulators of gastric mucosal injury by indomethacin: effect of antiulcer agents.

Endothelin-1 (ET-1), nitric oxide, and cytokines are recognized mediators of the inflammatory processes associated with gastric mucosal injury. In this study, we investigated mucosal expression of ET-1, interleukin-4 (IL-4), and the activity of constitutive nitric oxide synthase (cNOS) during indomethacin-induced gastric mucosal injury, and evaluated the effect of antiulcer agents on this process. The experiments were conducted with groups of rats pretreated intragastrically with ranitidine (100 mg/kg), ebrotidine (100 mg/kg), sulglycotide (200 mg/kg) or vehicle, followed 30 min later by an intragastric dose of indomethacin (60 mg/kg). The animals were killed 2 h later and their mucosal tissue subjected to macroscopic damage assessment and the measurements of epithelial cell apoptosis, ET-1, IL-4, and cNOS. In the absence of antiulcer agents, indomethacin caused multiple hemorrhagic lesions and extensive epithelial cell apoptosis, accompanied by a 20.7% reduction in IL-4, a 3.1-fold increase in mucosal expression of ET-1 and a 4.2-fold decline in cNOS. Pretreatment with H2-receptor antagonist, ranitidine produced a 15.7% reduction in the mucosal damage caused by indomethacin, 29.5% decrease in epithelial cell apoptosis and a 19.6% reduction in ET-1, while the expression of IL-4 increased by 10.8% and that of cNOS showed a 2-fold increase. The H2-blocker, ebrotidine, also known for its gastroprotective effects, reduced the indomethacin-induced lesions by 90.2%, epithelial cell apoptosis decreased by 61% and ET-1 showed a 58.2% decline, while IL-4 increased by 30.6% and that of cNOS showed a 3.1-fold increase. Pretreatment with gastroprotective agent, sulglycotide, led to a 51.2% reduction in the extent of mucosal damage caused by indomethacin, a 43.9% decrease in apoptosis, and a 63.5% decrease in ET-1, while the expression of cNOS increased by 3.4-fold and the level of IL-4 showed a 32.2% increase. The results suggest that an increase in vasoconstrictive ET-1 level combined with a decrease in regulatory cytokine, IL-4, and a loss of compensatory action by cNOS may be responsible for gastric mucosal injury caused by indomethacin. Our findings also point to a value of ebrotidine and sulglycotide in countering the untoward gastrointestinal side effects of NSAID therapy.     

Beneficial haemodynamic effect of indomethacin during endotoxin shock in anaesthetized pigsputative involvement of nitric oxide?

Endotoxin shock was induced in 31 anaesthetized pigs by infusion of 5 mug/kg of Escbeicbia coli endotoxin (LPS) over 60 min into the superior mesenteric artery. Fifteen of these pigs died within 30 min of the start of LPS infusion whereas the remaining 16 survived the experimental period of 2 h. In a group of nine pigs indomethacin (2 mg/kg, i.v.)was inected 20-25 rain after the start of LPS infusion at which time mean arterial blood pressure (MABP) had decreased below 40 mmHg indicating imminent death. Indomethacin immediately reversed the hypotension. In another group of five pigs, N(G)-nitro L-arginine-methyl ester (L-NAME, 1 and 3 mg/kg)was iniected 10 and 5 min, respectively, before the expected death without any beneficial effect on the hypotension. Three rain after the last dose of L-NAME, indomethacin (2 mg/kg, i.v.) was iniected. In three animals the hypotension was reserved by indomethacin, although this beneficial effect was delayed in comparison with the LP-Streated group not receiving L-NAME. Four pigs were pretreated with L-NAME, 3 mg/kg, i.v., 10 min prior to LPS infusion. All pretreated animals tended to die within 30 min of the start of the LPS infusion. Five rain before the expected death (20-25 rain after the start of LPS infusion) indomethacin (2 mg/kg) was inected. In three of these animals indomethacin reversed hypotenston and prevented death. Interestingly, this rise in the MABP developed very slowly. These results suggest that the beneficial effect of indomethacin in endotoxin shock might be related partially to interference with nitric oxide, which is not the only factor determining blood pressure levels during endotoxic shock.     

Antiulcerogenic activity of the hydroalcoholic extract of leaves of Annona muricata Linnaeus in mice

Annona muricata Linnaeus, popularly known as “graviola” and also called soursop, is a species typical of countries with a tropical climate, and it is used in folk medicine as an anticancer, analgesic and antispasmodic agent. The aim of the present study was to validate the gastroprotective activity of the hydroalcoholic extract of the leaves of A. muricata (HEAM) and to investigate the underlying mechanisms of action for this effect. Gastric lesions were induced in mice by absolute ethanol, acidified ethanol or indomethacin. Before, the animals were pretreated with saline, omeprazole or HEAM orally at doses of 50–400 mg/kg. To determine the mechanism of action of the extract, we investigated, using specific inhibitors, the involvement of nitric oxide (NO), prostaglandins (PGEs), ATP-dependent K⁺ channels and α2-noradrenergic receptors. HEAM showed significant antiulcer activity against lesions induced by absolute ethanol, acidified ethanol or indomethacin, which was mediated by endogenous gastric prostaglandins.