Nitro-Arginine Methyl Ester,a Non-Selective Inhibitor of Nitric Oxide Synthase Reduces Ibuprofen-Induced Gastric Mucosal Injury in the Rat

Ibuprofen is a commonly used non-steroidal anti-inflammatory drug. Gastrointestinal adverse drug reactions from ibuprofen usage include gastric mucosal ulcers and bleeding. The mechanism by which ibuprofen induces gastric mucosal damage is not clear. The present study is an attempt to examine the role of nitric oxide in the pathogenesis of ibuprofen-induced gastric mucosal damage. Ibuprofen administered orally at the dose of 100 mg/kg body weight for 6 days to the rats resulted in gastric mucosal injury. Serum nitrite and nitrosothiol were increased significantly as compared with the controls, which were treated with the vehicle alone. In the gastric mucosa, lipid peroxidation and protein thiols were increased, and the activity of glyceraldehyde 3-phosphate dehydrogenase, a nitric oxide sensitive enzyme was decreased significantly. Pretreatment of the rats daily with nitric oxide synthase inhibitor, nitro-arginine methyl ester (30 mg/kg body weight) 1 hr before treatment with ibuprofen reduced the gastric mucosal injury. Biochemically, it prevented the rise in serum nitrite levels and the increase in lipid peroxidation and protein thiol levels and the loss of glyceraldehyde 3-phosphate dehydrogenase activity in the gastric mucosa. The results of the present study suggest that increased nitric oxide production may be one of the mechanisms by which ibuprofen produces gastric mucosal injury and that inhibition of nitric oxide synthase reduces gastric mucosal injury.     

Effect of Ibuprofen on Cyclooxygenase and Nitric Oxide Synthase of Gastric Mucosa: Correlation with Endoscopic Lesions and Adverse Reactions

The aim of this study was to evaluate the effect of ibuprofen on gastric mucosa and enzymes involved in gastroprotection in healthy volunteers. Twenty-four Helicobacter pylori-negative subjects were randomized to treatment with ibuprofen or ibuprofen-arginate (each 600 mg/6 hr during 3 days). Endoscopies were performed 1 week before and after treatment. Biopsies were taken from the gastric antrum and corpus for determination of prostaglandin E2 (PGE2) by ELISA and cyclooxygenase (COX-1 and COX-2) and nitric oxide synthase (eNOS and iNOS) by western blot. All subjects had at least one gastric lesion except for two individuals taking ibuprofen-arginate. Ibuprofen-arginate caused a lower rate of clinical adverse reactions than ibuprofen. Subjects with gastric lesions or adverse reactions had lower PGE2 levels. COX-1, COX-2, eNOS, and iNOS were detectable in all subjects. The constitutive enzymes (COX-1 and eNOS) did not change after treatment. COX-2 was higher in corpus than antrum and it increased after ibuprofen treatment. iNOS tended to increase mildly in the corpus in subjects with adverse reactions or endoscopic lesions. There were no significant differences between ibuprofen and ibuprofen-arginate in PGE2, or enzymes.     

Nitric Oxide and its Role in Blastocyst Implantation

Reviews in Endocrine and Metabolic Disorders -     

Gastric Mucosal Blood Flow and Neutrophil Activation in Aspirin-Induced Gastric Mucosal Damage in Man

Konturek JW, Dembinski A, Stoll R, Konturek M, Domschke W. Gastric mucosal blood flow and neutrophil activation in aspirin-induced gastric mucosal damage in man. Scand J Gastroenterol 1993; 28:767-771.

Gastric and intestinal injury induced by nonsteroidal anti-inflammatory agents (NSAIDs) such as aspirin (ASA) is a common side effect of this class of drugs, but the mechanism by which these drugs act is not fully explained. In this study the effects of 3 days of continuous oral ASA administration (1 g twice daily) to eight healthy male volunteers were studied. To estimate the extent of mucosal damage, gastroscopy was performed before and after 3 days of ASA treatment, during which the mucosal blood flow was measured by means of laser-Doppler flowmetry. Before each endoscopy gastric microbleeding was measured. Since neutrophil activation has recently been suggested to be involved in the pathogenesis of ASA-induced gastric mucosal damage, we examined the influence of ASA treatment on the activation of leukocytes by determining their association with platelets in the blood. Aspirin-induced acute gastric damage reached about 3.5 in the endoscopic Lanza score. Mucosal blood flow increased significantly after ASA treatment, by about 50% in the oxyntic gland area and by 87% in the antral area. Gastric microbleeding rose from about 0.38 ml/day in the intact stomach to about 7.7 ml/day after ASA treatment. The platelet/neutrophil adherence increased significantly in both thrombin-unstimulated and thrombin-stimulated platelets. We conclude that acute 3 days' administration of ASA in man produces well-defined areas of gastric damage accompanied by a significant increase in gastric microbleeding and gastric blood flow and that ASA promotes platelet/neutrophil adhesion that may resemble the neutrophil/ endothelium interaction in the gastric mucosa.     

Studies of Nitric Oxide Generation from Salivary Nitrite in Human Gastric Juice

Background: Saliva contains substantial concentrations of nitrite derived from the enterosalivary recirculation of dietary nitrate. Methods: We have investigated factors in gastric juice influencing the fate of nitrite in swallowed saliva. When nitrite (100 μM) is added to human gastric juice pH 1.5 or pH 2.5 at 37 °C containing physiological concentrations of thiocyanate (1 mM) and ascorbic acid (200 μM), it is converted to nitric oxide within a few seconds. Results: The reduction of nitrite to nitric oxide is slower at pH 3.5 and very little is generated at pH 4.5. The rate of nitric oxide generation at acid pH increases with increasing thiocyanate concentration. The concentration of nitric oxide generated in the above way is maintained until the ascorbic acid is depleted by the recycling of nitric oxide to nitrite. In gastric juice depleted of ascorbic acid, very little nitrite is reduced to nitric oxide at any pH. Conclusion: These studies indicate that in the healthy acid-secreting stomach most salivary nitrite will be reduced to nitric oxide at the gastro-oesophageal junction and gastric cardia where it first encounters gastric juice.     

The Role of Nitric Oxide in the Failing Heart

Nitric oxide (NO) has effects on contractility, energetics and gene expression of failing myocardium. Initial studies on isolated cardiomyocytes showed NO to reduce systolic shortening but intracoronary infusions of NO-donors or of NO synthase (NOS) inhibitors failed to elicit changes in baseline LV contractility indices such as LVdP/dt(max). Intracoronary infusions of NO-donors or of substance P, which releases NO from the coronary endothelium, however demonstrated NO to induce a downward displacement of the left ventricular (LV) diastolic pressure-volume relation, consistent with increased LV diastolic distensibility. In end-stage failing myocardium, the increased oxygen consumption is related to reduced NO production and in isolated cardiomyocytes, NO blunts the norepinephrine-induced expression of the fetal gene programme thereby preserving myocardial calcium homeostasis.In dilated cardiomyopathy, changed endomyocardial NOS gene expression has been reported. Because of lower endomyocardial NOS gene expression in patients with higher functional class and lower LV stroke work, increased endomyocardial NOS gene expression seems to be beneficial rather than detrimental for the failing heart. A beneficial effect of increased NOS gene expression could result from NO's ability to increase LV diastolic distensibility, to augment LV preload reserve, to reduce myocardial oxygen consumption and to prevent downregulation of calcium ATPase. Upregulated endomyocardial NOS gene expression has also been reported in athlete's heart and could therefore play a role in physiological LV remodeling. Reduced endomyocardial NO content because of decreased NO or increased superoxide production could lower LV diastolic distensibility and contribute to diastolic heart failure. In many conditions such as aging, hypertension, diabetes or posttransplantation, the increased incidence of diastolic heart failure is indeed paralleled by reduced endothelium-dependent vasodilation.     

Role of Endogenous Nitric Oxide and Prostaglandin in Duodenal Bicarbonate Response Induced by Mucosal Acidification in Rats

Duodenal HCO₃ ^– secretion increases in response to mucosal acidification by luminal acid. Although this process is known to be mediated by endogenous prostaglandins (PGs), the role of nitric oxide (NO) in this response has been little studied. We examined the effects of indomethacin and N_G-nitro-l-arginine methyl ester (l-NAME) on the acid-induced HCO₃ ^– secretion in the rat duodenum, together with those on PGE₂ generation as well as luminal release of NO metabolites (NO_x). A proximal duodenal loop was perfused with saline, and the HCO₃ ^– secretion was measured at pH 7.0 using a pH-stat method and by adding 10 mM HCl. Mucosal acidification was performed by exposing the loop to 10 or 100 mM HCl for 10 min. Acidification of the duodenal mucosa stimulated the HCO₃ ^– secretion, with concomitant increase of mucosal PGE₂ contents and luminal release of NO_x, the response being much greater in case of 100 mM HCl. Indomethacin significantly inhibited the acid-induced HCO₃ ^– secretion as well as the PGE₂ biosynthetic response, without influence on the NO_x release. Pretreatment of the animals with l-NAME attenuated both the increase of mucosal PGE₂ contents and luminal release of NO_x following the acidification, resulting in a marked inhibition of the acid-induced HCO³> ^– response, and these effects were significantly antagonized by coadministration of l-arginine. Duodenal HCO₃ ^– secretion was also increased by mucosal exposure to NOR-3 (a NO donor), with concomitant increase of PGE₂ generation, but these effects were mitigated in the presence of indomethacin. In addition, the duodenal damage caused by mucosal perfusion with 100 mM HCl for 4 hr was markedly aggravated by pretreatment with l-NAME as well as indomethacin. These results suggest that both endogenous NO and PGs are involved in the mechanism for the acid-induced duodenal HCO₃ ^– secretion, and that NO may increase the HCO₃ ^– secretion by stimulating PG generation.     

脉络宁对新生大鼠缺氧缺血性脑损伤后nNOS和c-Fos表达的影响

目的:检测新生大鼠缺血缺氧后神经元型一氧化氮合酶(nNOS)和c-Fos免疫活性表达改变及脉络宁对其的影响。方法:结扎7d龄大鼠右侧颈总动脉1h,然后暴露在8%氧和92%氮的混合气体中2h,建立缺氧缺血脑损伤模型。用免疫组织化学方法检测并比较缺血缺氧后和脉络宁处理后c-Fos和nNOS的免疫活性。结果:缺血缺氧6h后c-Fos表达达高峰;与缺氧缺血组相比,脉络宁处理组c-Fos阳性神经元数量增加,表达c-Fos的时间延长;与假手术组相比,缺氧缺血组nNOS表达水平增高,脉络宁处理组则下降。结论:脉络宁可抑制nNOS表达,增强c-Fos表达,可能对缺血缺氧性脑损伤有保护作用。     

Role of Nitric Oxide in Cocaine-Induced Acute Hypertension

Cocaine causes acute hypertension by blocking catecholamine reuptake. There is evidence that it also impairs the peripheral endothelial nitric oxide system, which is normally vasodilatory. We further explored the role of nitric oxide in cocaine-induced vasoconstriction in anesthetized rats, and in vitro by using isolated carotid artery segments. Cocaine administered intravenously in rats increased mean arterial pressure by 30 to 40 mm Hg within 1 min. This effect was dose dependent and the maximum effect was observed at a dose of 1.25 mg/kg. The prototype catecholamine norepinephrine induced a similar increase in blood pressure. When rats were pretreated with N^G-monomethyl-l-arginine (L-NMMA, a blocker of nitric oxide) and challenged with cocaine, the increase in blood pressure was blocked by 80%, whereas pretreatment with L-NMMA did not block norepinephrine-induced vasoconstriction. Both cocaine and norepinephrine also induced an immediate vasoconstriction in isolated carotid artery preparations. The in vitro vasoconstriction induced by cocaine was blocked by pretreatment with L-NMMA, whereas L-NMMA did not block the norepinephrine-induced vasoconstriction in vitro. Furthermore, carotid artery stripped of endothelium responded to norepinephrine but failed to respond to L-NMMA or cocaine. S-nitroso-N-acetyl-d,l-penicillamine (SNAP)—a precursor of nitric oxide— stimulated nitric oxide production in control coronary artery fragments. When these fragments were incubated with cocaine there was a 20% reduction in the production of nitrite oxide. These results suggest that cocaine exerts its peripheral vasoconstriction at least in part by inhibiting local vasodilator nitric oxide.     

Role of Nitric Oxide During Swallow-Induced Esophageal Shortening in Cats

Swallowing induces esophageal shortening due to contraction of the longitudinal muscle (LM) layer. Experiments in the opossum have shown an excitatory effect of nitric oxide (NO) on esophageal LM strips. We evaluated the role of NO in swallow-induced esophageal shortening and assessed the effect of NO in vitro on feline LM strips. Swallow-induced esophageal shortening was studied before and after NO synthase blockade with l-NAME. In five cats esophageal shortening was measured using two endoscopically affixed mucosal clips. In another five cats LM contraction was measured by a strain gauge sutured on the serosal side at 2 cm above the LES; muscle strips from that region were obtained for in vitro studies. Swallowing induced esophageal shortening of 48.3 ± 8.3% and LM contraction of 4.4 ± 0.8 g in the control period and 32.1 ± 8% and 3.0 ± 0.4 g after l-NAME (P < 0.05). Nitric oxide and SNP did not change the basal tone of esophageal LM strips but provoked inhibition of metacholine-induced tonic and phasic activity. Electrical field stimulation induced frequency-dependent contractions that were reduced by atropine without further reduction after l-NAME. In conclusion, the reduction of esophageal shortening after l-NAME during the in vivo experiments suggested an excitatory effect of NO on the feline esophagus. The in vitro experiments, however, showed no contractile effect of NO or SNP on LM strips, but an inhibitory effect on the precontracted tissue. The influence of NO synthase blockade on in vivo esophageal LM shortening might be secondary to its effect on circular muscle contractility.     

肝硬化大鼠胃壁一氧化氮合酶的组化研究

目的：门脉高压性胃病在组织学上以胃臂粘膜及粘膜下血管扩张为特征，一氧化氮作为扩血管物质可能参与了此异常的发生。方法：使用ＮＡＤＰＨ黄递酶组化法显示肝硬化大鼠胃臂一氧化氮合酶（ＮＯＳ）活性。结果：肝硬化大鼠胃粘膜上皮、胃壁内小动脉及小静脉ＮＯＳ染色均增强，胃粘膜上皮脱落、变性、不规整，胃壁内小动脉、小静脉扭曲变形。结论：肝硬化大鼠胃壁一氧化氮合酶产生增加，可能参与了肝硬化门脉高压性胃病的发生。     

GABAergic Mechanisms of Gastroprotection in the Rat: Role of Sensory Neurons, Prostaglandins, and Nitric Oxide

gamma-Aminobutyric acid (GABA) is a neurotransmitter found in both the central and the peripheral nervous systems including the gastrointestinal tract. The aims of the present studies were to examine mechanisms by which GABA exerts gastroprotective effects against ethanol- and water-restraint stress (WRS)-induced gastric mucosal injury in the rat. GABA, administered intragastrically (400 mg/kg), induced gastroprotection against ethanol and WRS by activating gastric sensory neurons to release calcitonin gene-related peptide (CGRP) and promote nitric oxide (NO) synthesis and release. Furthermore, these protective effects of GABA were associated with an increase in gastric mucosal blood flow (GMBF) that was dependent on sensory neuron and NO systems. GABA-mediated protection involved GABAA receptor activation and prostaglandin generation. In conclusion, intraluminal GABA protects the stomach against ethanol- and WRS-induced injury by mechanisms which involve sensory neuron/CGRP/NO pathways and increases in GMBF and prostaglandin generation.     

幽门螺杆菌感染者血清一氧化氮水平及胃粘膜细胞凋亡的变化

目的探讨幽门螺杆菌(Hp)感染者血清一氧化氮(NO)含量变化及其对胃粘膜细胞凋亡的影响.方法Hp阳性者36例为研究对象,Hp阴性者31例作为对照.用酶联免疫法测定血清NO含量,TUNEL法检测胃粘膜细胞凋亡.结果Hp用性组血清NO含量(85.6士21.4)μmoI/L,Hp阴性组为(69.2士19.3)μmol/L,两者比较P＜0.01;Hp阳性组胃粘膜细胞凋亡指数为9.7士2.8,而Hp阴性组为8.1士2.2,两者比较P<0.01.结论Hp感染患者NO明显高于非Hp感染者,并有显著的细胞凋亡现象,这可能与Hp可诱导N0合成酶有关.     

Gastric Mucosal Protection by Aegle Marmelos Against Gastric Mucosal Damage: Role of Enterochromaffin Cell and Serotonin

Background/Aims:

Serotonin (5-hydroxytryptamine; 5-HT) released from enterochromaffin (EC) cells in gastric mucosa inhibits gastric acidity by increasing the gastric mucus secretion. In the present study, we evaluated the effect of aqueous extract of Aegle marmelos (AM) ripe fruit pulp (250 mg/kg body weight) on mean ulcer index (MUI), EC cells, 5-HT content, and adherent mucosal thickness of ulcerated gastric tissue in adult albino rats.

Material and Methods:

Ulceration was induced by using aspirin (500 mg/kg, p.o.), cerebellar nodular lesion and applying cold-restraint stress.

Results:

In all cases increased MUI in gastric tissue along with decreased EC cell count was observed with concomitant decrease of 5-HT content and adherent mucosal thickness (P < 0.05). Pretreatment with AM for 14 days decreased MUI, increased EC cell count, and 5-HT content as well as adherent mucosal thickness in all ulcerated group (P < 0.05).

Conclusion:

AM produces gastric mucosal protection mediated by increased EC cell count and 5-HT levels.     

Role of Nitric Oxide in the Regulation of Substrate Metabolism in Heart Failure

Solid experimental evidence indicates that nitric oxide (NO) inhibits oxygen utilization in vitro and in vivo. The role played by NO in cellular metabolism is likely extended to the control of substrate utilization. Studies performed in normal hearts show that NO inhibits glucose uptake and that a reduced synthesis of NO impairs free fatty acid consumption. Interestingly, we found also that myocardial free fatty acid utilization decreases while glucose consumption is enhanced in end stage heart failure, when cardiac NO production falls dramatically. This phenomenon led us to the hypothesis that the reduced synthesis of NO could be at least in part responsible for myocardial metabolic alterations occurring in severe heart failure. The present review mentions some of the seminal studies that defined the function of NO as metabolic modulator. A particular emphasis is put on available data suggesting a role for NO in the control of cardiac substrate utilization in normal and failing hearts.     

Factors Involved in Upregulation of Inducible Nitric Oxide Synthase in Rat Small Intestine Following Administration of Nonsteroidal Anti-inflammatory Drugs

We investigated the functional mechanisms underlying the expression of inducible nitric oxide (NO) synthase (iNOS) in the rat small intestine following the administration of nonsteroidal anti-inflammatory drugs (NSAIDs) and found a correlation with the intestinal ulcerogenic properties of NSAIDs. Conventional NSAIDs (indomethacin, dicrofenac, naproxen, and flurbiprophen), a selective cyclooxygenase (COX)-1 inhibitor (SC-560) and a selective COX-2 inhibitor (rofecoxib) were administered p.o., and the intestinal mucosa was examined 24 hours later. Indomethacin decreased prostaglandin E₂ (PGE₂) production in the intestinal mucosa and caused intestinal hypermotility and bacterial invasion as well as the upregulation of iNOS expression and NO production, resulting in hemorrhagic lesions. Other NSAIDs similarly inhibited PGE₂ production and caused hemorrhagic lesions with intestinal hypermotility as well as iNOS expression. Hypermotility in response to indomethacin was prevented by both PGE₂ and atropine but not ampicillin, yet all these agents inhibited not only bacterial invasion but also expression of iNOS as well, resulting in prevention of intestinal lesions. SC-560, but not rofecoxib, caused a decrease in PGE₂ production, intestinal hypermotility, bacterial invasion, and iNOS expression, yet this agent neither increased iNOS activity nor provoked intestinal damage because of the recovery of PGE₂ production owing to COX-2 expression. Food deprivation totally attenuated both iNOS expression and lesion formation in response to indomethacin. In conclusion, the expression of iNOS in the small intestine following administration of NSAIDs results from COX-1 inhibition and is functionally associated with intestinal hypermotility and bacterial invasion. This process plays a major pathogenic role in the intestinal ulcerogenic response to NSAIDs.     

Gastric mucosal injury and associated changes in mucosal blood flow and gastric fluid secretion caused by dimethyl sulfoxide (DMSO) in rats

Dimethyl sulfoxide applied intragastrically for 10 min in rats caused extensive mucosal damage. In concentrations of 5%, 10%, or 100%, dimethyl sulfoxide caused superficial damage to 33%, 36%, and 97%, respectively, of the corpus mucosa, and 28%, 44%, and 96%, respectively, of the antral mucosa. Concentrated dimethyl sulfoxide also caused damage to the pits and glands in some areas of the mucosa. The amount of fluid in the stomach increased by 0.24 ml, 0.48 ml, and 2.07 ml during application of 5%, 10%, and 100% dimethyl sulfoxide. The 10% dimethyl sulfoxide increased mucosal blood flow by 0.57 ml/min/g in the antrum, and 100% dimethyl sulfoxide increased mucosal blood flow by 2.21 ml/min/g in the antrum and by 1.17 ml/min/g in the corpus. We conclude that dimethyl sulfoxide is a gastric irritant, which should be considered when it is used as an oxygen radical scavenger, as a drug or carcinogen vehicle, or as oral medication in patients. The protective effect of intragastric dimethyl sulfoxide against stress and various drug-induced gastric injury may be due to adaptive cytoprotection rather than an oxyradical scavenger effect.This work was supported by grants from the Norwegian Cancer Society. Dr. Sørbye is a Research Fellow of the Norwegian Cancer Society.     

Limited Capacity for Ammonia Removal by Brain in Chronic Liver Failure: Potential Role of Nitric Oxide

Chronic liver failure leads to hyperammonemia and consequently increased brain ammonia concentrations, resulting in hepatic encephalopathy. When the liver fails to regulate ammonia concentrations, the brain, devoid of a urea cycle, relies solely on the amidation of glutamate to glutamine through glutamine synthetase, to efficiently clear ammonia. Surprisingly, under hyperammonemic conditions, the brain is not capable of increasing its capacity to remove ammonia, which even decreases in some regions of the brain. This non-induction of glutamine synthetase in astrocytes could result from possible limiting substrates or cofactors for the enzyme, or an indirect effect of ammonia on glutamine synthetase expression. In addition, there is evidence that nitration of the enzyme resulting from exposure to nitric oxide could also be implicated. The present review summarizes these possible factors involved in limiting the increase in capacity of glutamine synthetase in brain, in chronic liver failure.