Gastric Ulcer Area Research Articles

884 The Role of the Ferric Uptake Regulator (Fur) in Helicobacter pylori Resistance to Metronidazole

G A A b st ra ct s induced gastric lesions at 6, 12 and 24 h upon 3.5 h of WRS exposure (series B). In separate series, rats with chronic gastric ulcers induced by serosal application of acetic acid (ulcer area=28 mm2) were treated daily for 9 and 15 days with: 1) vehicle (saline); 2) Hp2-20 (0.5 25 mg/kg i.g.) or 3) control peptide Hp1 (10 mg/kg i.g.) with or without non-selective and selective inhibitors of cNOS and iNOS, L-NNA (20 mg/kg-d i.p.) and L-Nil (5 mg/kgd i.p.) without or with the combination with L-arginine (200 mg/kg-d s.c.). The gastric juice was collected for acid output assessment and the number and area of WRS lesions and ulcers were determined planimetrically, the gastric blood flow (GBF) was determined by H2-gas clearance technique and luminal NOx concentration, plasma VEGF levels, the expression of mRNAs for VEGF and FPR mRNA and protein were assessed by ELISA, RTPCR and Western Blot. Hp2-20 dose-dependently inhibited the gastric acid secretion and decreased the number of WRS-induced gastric lesions at 6h, 12 h and 24 h by 16%, 38% and 46%, respectively. Hp2-20 reduced the area of acetic acid ulcers; the dose inhibiting by 50% (ID50) of ulcer area at day 9 upon ulcer induction being 10 mg/kg and this was accompanied by the rise in GBF at ulcer margin, luminal NOx levels and VEGF concentration. Treatment with L-NNA and L-Nil which by themselves increased the area of gastric ulcer significantly reduced the healing effects of Hp2-20 and accompanying rise in the GBF and plasma NOx concentration. These increase in the area of gastric ulcers and accompanying fall in GBF and NOx content induced by L-NNA and L-Nil were reversed by L-arginine added to NOS inhibitors. Upregulation of mRNA and protein for VEGF and FPR at ulcer marginwas observed inHp2-20-treated animals and these effects were significantly attenuated by L-NNA or L-Nil. We conclude that Hp2-20 enhances the mucosal recovery from WRSinduced lesions and accelerates the healing of preexisting gastric ulcers due to upregulation of FRP receptors via mechanism involving inhibition of gastric acid secretion, an activation of NO/NOS pathway and an overexpression and release of VEGF at the ulcer margin responsible for new microvessels formation.

W1695 Effect of Allyl Isothiocyanate, a Pungent Ingredient of Wasabi and Mustard Oil, on Gastric Mucosal Blood Flow and Ulcerogenesis in Rats: Comparison With Capsaicin

arginine (200 mg/kg i.g.); 2) calcitonin gene related peptide (CGRP 5 μg/kg s.c.) and low dose of capsaicin (0.5 mg/kg i.g.) capable of stimulating sensory afferents on ischemia/ reperfusion (I/R, 30 min of ischemia to celiac artery followed by 3 hrs of reperfusion) injury (series A)and the healing of chronic acetic acid ulcers (ulcer area = 28 mm2) in rats with intact and capsaicin-deactivated sensory nerves (series B). The area of gastric lesions and ulcers was determined by planimetry, the gastric blood flow (GBF) was assessed by H2-gas clearance method, NOx gastric content, plasma MPO activity and the mucosal activity of antioxidazing enzymes SOD, GPx and MDA concentration as an index of oxygen radicallipid peroxidation were assessed. The gastric content of ADMA was determined by HPLC method and gastric mucosal expression of transcription factor NFκB, IL-1β and TNF-α mRNAs their plasma levels were analyzed by RT-PCR and ELISA. I/R produced gastric lesions (series A) accompanied by an increase in plasma ADMA, MDA, MPO contents, the fall in the GBF by 31% and the suppression of SOD and GPx activities. ADMA which significantly reduced the luminal NOx content, dose-dependently aggravated I/R damage and significantly delayed the healing of acetic acid ulcers (series B); both effects being worsened by capsaicin denervation. These effects were accompanied by further fall in the GBF and a significant decrease in the SOD and GPx activities and the significant increase in MDA content. The ADMA delay in ulcer healing was accompanied by the upregulation of NFκB, IL-1β and TNF-α mRNAs at ulcer margin followed by the rise in plasma IL-1β, TNF-α and ADMA levels and these effects were markedly diminished by addition of L-Arg, CGRP or capsaicin to ADMA. We conclude that: 1) ADMA plays an important role in the pathogenesis of acute gastric lesions induced by I/R and delays ulcer healing due to the inhibition of NO generation, NFκB activation, overexpression of proinflammatory cytokines IL-1β and TNF-α and enhancement in lipid peroxidation; 2) L-Arg, a donor of NO or sensory nerve vasodilatory neuropeptide CGRP counteract the ADMA-induced facilitation of mucosal lesions and the impairment of the healing of preexisting gastric ulcers.

Role of modulation of vascular endothelial growth factor and tumor necrosis factor-alpha in gastric ulcer healing in diabetic rats

The aim of the present study was to assess the effect of drugs that increase gastric vascular endothelial growth factor (VEGF) and suppress gastric tumor necrosis factor-alpha (TNF-α) in gastric ulcer healing in streptozotocin-induced diabetic rats. Sixty male albino rats were made diabetic by intraperitoneal (i.p.) streptozotocin injection and ten rats were injected i.p. by a single dose of saline. Six weeks following streptozotocin or saline injection, gastric ulcers were induced by serosal application of acetic acid. Three days after acetic acid application, rats were divided into: group I (non-diabetic control), group II (streptozotocin-injected), groups III–VII (streptozotocin-injected rats treated with insulin, insulin and pentoxifylline, insulin and simvastatin, pentoxifylline as well as simvastatin, respectively, for 7 days following acetic acid application. The use of insulin, combinations of insulin and pentoxifylline or simvastatin resulted in a significant decrease in gastric ulcer area, significant increase in epithelial regeneration assessed histologically, significant increase in gastric VEGF concentration, and gastric von Willebrand factor (vWF) as well as significant decrease in gastric TNF-α. A significant difference in gastric ulcer area as well as in gastric TNF-α, VEGF and vWF levels could be observed between rats that received combinations of insulin and pentoxifylline or simvastatin compared to rats that received either drug alone. Our results suggest the feasibility of a novel treatment strategy, namely pentoxifylline and simvastatin, for patients in whom impairment of ulcer healing constitutes a secondary complication of diabetes mellitus.

Anti-Ulcer (Ulcer-Preventive) Activity of Ficus arnottiana Miq. (Moraceae) Leaf Methanolic Extract

Problem statement: In spite of being one of the well-known medicinal p lants used in Indian traditional medicine to treat several ailmen ts, studies pertaining to the pharmacological properties of Ficus arnottiana are very scarce. We studied the anti-ulcer activity and acute toxicity of Ficus arnottiana leaf methanolic extract for the first time . Approach: Freshly collected F. arnottiana leaves were dried, powdered and extracted in methanol. To study the anti-ulcer activity, Wistar rats were orally administered with different doses of th e extract (0, 250 and 500 mg kg -1 body weight day -1 ) or with the reference drug omeprazole (8 mg kg -1 ) for 10 days. After induction of ulcer using 5 mL kg -1 ethanol, stomachs of these animals were analyzed f or gastric volume, ulcer area and gross pathological changes. Results: Our results showed that F. arnottiana methanolic extract could prevent ulcer in rats in a dose-dependent manner. Histologi cal studies revealed that the extract had muco- protective activity. The extract did not show any a cute toxicity even at the dose of 5000 mg kg -1 indicating that the extract has no lethal effect. P reliminary phytochemical screening of this extract identified the presence of important secondary meta bolites like flavonoids and tannins. Conclusion/Recommendations: From this study, it is clear that F. arnottiana leaf extract had significant anti-ulcer activity in animal models. I t had muco-protective activity and gastric anti- secretary activity. The extract is non-toxic even a t relatively high concentrations.

Effects of progesterone on FSH-stimulated indomethacin ulcers in rats

To investigate whether the balance between progesterone and FSH levels has a role in ulcer formation/inhibition. Animal study. Ataturk University, Faculty of Medicine, Laboratory of Pharmacology Department. One hundred thirty-two female Albino Wistar rats. Ovariectomy. Chronic administration of progesterone and estrogen to ovariectomized rats. Acute administration of progesterone to ovariectomized rats. Acute administration of progesterone and FSH to intact rats. Combined administration of mifepristone with progesterone or FSH to intact rats. Indomethacin administration to all rats. Gastric ulcer areas as mm(2). Indomethacin-induced ulcers were significantly higher in ovariectomized rats than in intact rats. Chronic progesterone (1 mg/kg) inhibited gastrointestinal system ulcers by 51.2%, whereas 2 and 5 mg/kg chronic progesterone and 1, 2, and 5 mg/kg chronic estrogen were not effective. Acute progesterone (5 mg/kg) increased indomethacin ulcers significantly in ovariectomized rats. FSH increased indomethacin ulcers significantly. In addition, progesterone increased indomethacin ulcers significantly in intact rats. Mifepristone antagonized the ulcerogenic effects of FSH and progesterone. Our data suggest that progesterone is not an antiulcer hormone, and produces ulcers via its own receptor. In addition, FSH may produce ulcerogenic effects via progesterone receptors. The low doses of progesterone (inhibits endogenous FSH) cannot stimulate its own receptors sufficiently for ulcer formation and prevent the ulcerogenic effects of FSH by decreasing FSH concentration.

Influence of dl-alpha-tocopherol acetate on indomethacin-induced gastric mucosal injury in rats

1 There is an increasing body of evidence supporting the hypothesis that antioxidants are able to reduce gastric mucosal damage induced by stressors of different origin. The aim of the present study was to investigate the influence of dl-alpha-tocopherol acetate (TA) on indomethacin-induced gastric mucosal injury in rats and a possible role for an anti-oxidative mechanism in the response. 2 TA (25, 50 and 100 mg kg(-1)) was applied intraperitoneally as a pretreatment 1 h before the subcutaneous administration of indomethacin (30 mg kg(-1)). 3 TA reduced the area of indomethacin-induced gastric ulcers, the effect being significant (P < 0.05) at the highest dose of 100 mg kg(-1). 4 Histopathological examination of rat stomach samples demonstrated that TA caused an increase in gastric mucus production and a reduction of the severity of mucosal lesions. 5 The three doses of TA prevented indomethacin-induced elevation of plasma and mucosal malondialdehyde (MDA) levels, which in TA-pretreated rats were not significantly different from the control values. Neither indomethacin treatment nor TA pretreatment had a significant influence on the gastric mucosal levels of reduced glutathione or oxidized glutathione. 6 Our results suggest that the gastroprotective effect of TA is likely to be due to increased mucus production and interference with oxidative stress development as evidenced by the decreased plasma and gastric mucosal MDA.

Probiotic Lactobacillus rhamnosus GG enhances gastric ulcer healing in rats

Probiotics are widely used as functional foods which have been advocated for the maintenance of gastrointestinal microflora equilibrium and treatment of gastrointestinal disorders. However, studying the role of probiotics in peptic ulcer disease is limited. The aim of the present study is to investigate the effect of a probiotic strain Lactobacillus rhamnosus GG on gastric ulcer and to elucidate the mechanisms involved. Gastric kissing ulcers were induced in rats by acetic acid (60% v/v). L. rhamnosus GG was given intragastrically at 10 8 cfu/day or 10 9 cfu/day for three consecutive days after ulcer induction. L. rhamnosus GG successfully colonized in the gastric mucosa especially at the ulcer margin. It also significantly and dose-dependently reduced gastric ulcer area. Cell apoptosis to cell proliferation ratio was strongly decreased and accompanied by significant up-regulation of ornithine decarboxylase (ODC) and B-cell lymphoma 2 (Bcl-2) protein expression at the ulcer margin. Angiogenesis was also significantly stimulated together with the induction of vascular endothelial growth factor (VEGF) expression. Furthermore, L. rhamnosus GG up-regulated the phosphorylation level of epidermal growth factor receptor (EGF receptor) without altering the total EGF receptor expression. These findings suggested that L. rhamnosus GG enhanced gastric ulcer healing via the attenuation of cell apoptosis to cell proliferation ratio and increase in angiogenesis. Regulators of these processes such as ODC, Bcl-2, VEGF and EGF receptor are likely to be involved in the healing action of L. rhamnosus GG for gastric ulcer.

Selank and its metabolites maintain homeostasis in the gastric mucosa

Antiulcer properties of a synthetic anxiolytic Selank and in vivo formed metabolites of this compound were studied on 3 experimental models of ulceration. The test peptides decreased the area of experimental gastric ulcers.

Agonist of peroxisome proliferator-activated receptor gamma (PPAR-γ: A new compound with potent gastroprotective and ulcer healing properties

Pioglitazone, a specific ligand for peroxisome proliferator-activated receptor gamma (PPAR-gamma), was recently implicated in the control of inflammatory processes and in the modulation of the expression of various cytokines such as tumor necrosis factor alpha (TNF-alpha), but its role in the mechanism of gastric mucosal integrity has not been studied extensively. This study was designed to determine the effect of pioglitazone on gastric mucosal lesions induced in rats by topical application of 100% ethanol and by 3.5 h of water immersion and restraint stress (WRS) with or without pretreatment with indomethacin (5 mg/kg i.p.) to inhibit cyclooxygenase-1 (COX-1) and COX-2 enzyme activities and L-NNA (20 mg/kg i.p.) to suppress nitric oxide (NO)-synthase. In addition, the effect of pioglitazone on ulcer healing in rats with chronic acetic acid ulcers (ulcer area 28 mm2) was determined. Rats were killed 1 h and 3.5 h after ethanol administration or WRS exposure or at day 9 upon ulcer induction, and the number and area of gastric lesions were measured by planimetry, the gastric blood flow (GBF) was determined by H2-gas clearance technique and the mucosal PGE2 generation and gene expression and plasma concentration of TNF-alpha and IL-1beta were also evaluated. Pre-treatment with pioglitazone dose-dependently attenuated gastric lesions induced by 100% ethanol and WRS; the dose reducing these lesions by 50% (ID50) being 10 mg/kg and 7 mg/kg, respectively. The protective effect of pioglitazone was accompanied by the significant rise in the GBF, an increase in PGE2 generation and the significant fall in the plasma TNF-alpha and IL-1beta levels. Strong signals for IL-1beta- and TNF-alpha mRNA were recorded in gastric mucosa exposed to ethanol or WRS, and these effects were significantly decreased by pioglitazone. Indomethacin which suppressed PG generation by about 90%, while augmenting WRS damage, and L-NNA, that suppressed NO-synthase activity, significantly attenuated the protective and hyperaemic activity of this PPAR-gamma ligand. In the chronic study, pioglitazone significantly reduced the area of gastric ulcers on day 9 and significantly raised the GBF at the ulcer margin. The acceleration of ulcer healing by PPAR-gamma ligand was accompanied by a significant increase in the expression of PECAM-1 protein, a marker of angiogenesis. We conclude that (1) pioglitazone exerts a potent gastroprotective and hyperaemic actions on the stomach involving endogenous PG and NO and attenuation of the expression and release of proinflammatory cytokines TNF-alpha and IL-1beta, and (2) PPAR-gamma ligand accelerates ulcer healing, possibly due to the enhancement in angiogenesis at ulcer margin.

Acupuncture therapy for experimental stomach ulcer and c-Fos expression in rats

To determine the role of acupuncture therapy in treating experimental gastric ulcer in rats. Twenty-eight adult male Sprague-Dawley rats were randomly divided into four groups (pre-acupuncture group; acupuncture group; paradistance-acupuncture group; and control group), and pre-acupuncture, paradistance-acupuncture, and control groups received 5 muL acetic acid (200 mL/L HAc) injection after a same course of electroacupuncture (EA) treatment (4 Hz, 0.6 mA, 0.45 ms, 45 min for 4 d). The rats in these three groups recovered within 4 d. The acupuncture group received EA therapy for 4 d, after HAc injection. The stomach was dissected to compare the pathological structures of ulcer. Also c-Fos activation in the nuclei of solitary tract (NTS) was observed under microscope after regular immunohistochemistry staining of brain stem sections. The number of ulcers was different among the four groups, especially between control group and paradistance-acupuncture group or pre-acupuncture group. In the latter group, the number of ulcers was much less. The gastric ulcer area was consistent with the histopathological results, indicating that pre-acupuncture had an obvious therapeutic effect on gastric ulcers. Acupuncture had a very modest effect and paradistance-acupuncture had no effect on gastric ulcers. No therapeutic effect was found in the control group. Fos-Li neurons in NTS induced by noxious gastric ulcer showed a significant difference between pre-acupuncture and control groups. Acupuncture before ulceration can obviously alleviate ulcer. The production of c-Fos proves that the vagus nerve mediates the induction of c-Fos in nuclei of solitary tract following experimental ulceration, suggesting that parasympathetic afferents promote the process of noxious visceral stimulation.

Role of capsaicin-sensitive nerves and histamine H 1, H 2, and H 3 receptors in the gastroprotective effect of histamine against stress ulcers in rats

It is assumed that an overproduction of gastric acid is the most important factor in the development of peptic ulcer. However, it has been also demonstrated that gastric defense mechanisms, which prevent mucosal injury, are enhanced by the same factors that increase acid secretion. The aim of this study was to examine the role of capsaicin-sensitive sensory nerves and histamine H 1, H 2, and H 3 receptors in histamine-induced gastroprotection against stress ulcers. Studies were performed on rats with intact or ablated sensory nerves. Ablation of sensory nerves was induced by neurotoxic doses of capsaicin. Gastric ulcers were induced by water immersion and restrain stress. Before exposure to stress, rats were pretreated with saline (control), histamine (10 μmol/kg), histamine H 1 receptor antagonist pyrilamine (100 μmol/kg), histamine H 2 receptor antagonist ranitidine (100 μmol/kg), histamine H 3 receptor antagonist thioperamide (100 μmol/kg), or a combination of histamine with these histamine receptor antagonists. Results Histamine alone reduced ulcer area evoked by stress and this effect was accompanied by an increase in gastric mucosal blood flow and mucosal DNA synthesis, as well as a decrease in serum pro-inflammatory interleukin-1β concentration. Treatment with combination of pyrilamine plus histamine caused an increase in gastric ulcer area and serum interleukin-1β above the value observed in animals treated with saline, and this effect was accompanied by a decrease in gastric mucosal DNA synthesis. Ranitidine, in combination with histamine, reduced the ulcer area and serum interleukin-1β to a minimal value, whereas gastric mucosal blood flow and DNA synthesis reached a maximal value. Pretreatment with thioperamide before histamine administration abolished the histamine-evoked reduction in gastric ulcer area. Ablation of sensory nerves increased the ulcer area in animals treated with saline or histamine, or histamine in combination with pyrilamine or ranitidine. In animals with sensory nerves ablation combined with administration of thioperamide plus histamine, the ulcer area was similar to that in saline-treated animals with intact sensory nerves. We conclude that: (1) histamine exhibits protective effect against stress-induced gastric ulcer and that this gastroprotection is related to stimulation of histamine H 1 and H 3 receptors; (2) blockade of histamine H 2 receptors exhibited beneficial effect on gastric mucosa against stress-induced gastric ulcers; and (3) ablation of sensory nerves aggravates stress-induced gastric ulcer and reduces histamine-evoked gastroprotection related to stimulation of histamine H 3 receptors.

Impaired gastric ulcer healing in diabetic rats: role of heat shock protein, growth factors, prostaglandins and proinflammatory cytokines

Gastric mucosa of diabetic rats is highly vulnerable to acute injury, but little is known about the influence of diabetic conditions on the healing of gastric ulcers. In this study, streptozotocin (70 mg/kg injected intraperitoneally) was used to induce diabetes mellitus in rats. Four weeks after streptozotocin injection, gastric ulcers were induced using the acetic acid method, and 10 days later, the healing rate and the gastric blood flow (GBF) were measured by planimetry and hydrogen (H 2)-gas clearance method, respectively. Six major groups of rats with gastric ulcers were used: (1) vehicle (saline); (2) streptozotocin alone; (3) insulin (4 IU/day intraperitoneally); (4) streptozotocin plus insulin; (5) pentoxifylline, an inhibitor of synthesis and release of tumor necrosis factor-α (TNFα); and (6) aspirin, a non-selective inhibitor of cyclooxygenase-1 (COX-1) and cyclooxygenase-2 (COX-2), and rofecoxib, the highly selective COX-2. In the diabetic rats, a significant delay in ulcer healing (∼by 300%), accompanied by a decrease in the gastric mucosal blood flow was observed. The prolongation of the healing in diabetic animals was associated with an increase in gastric mucosal expression and release of TNFα, interleukin-1β (IL-1β), suppression of the vascular endothelial growth factor (VEGF), platelet endothelial cell adhesion molecule-1 (PECAM-1) and the mucosal overexpression of heat shock protein 70 (HSP 70). Administration of insulin reversed the delay in ulcer healing and significantly decreased the expression of IL-1β and TNF-α, while producing the rise in the expression of VEGF and PECAM. Pentoxifylline, an inhibitor of TNF-α, which by itself accelerated ulcer healing in non-diabetic rats, counteracted the increase in the area of gastric ulcer induced by streptozotocin, raised significantly gastric blood flow and suppressed the plasma TNF-α levels. Aspirin and rofecoxib, that significantly suppressed the mucosal prostaglandin E 2 generation in ulcer area, delayed significantly the rate of ulcer healing and decreased the GBF at ulcer margin in non-diabetic rats, and these changes were significantly augmented in diabetic animals. We conclude that: (1) Experimental diabetes dramatically impairs ulcer healing, depending upon the increased release of proinflammatory cytokines and the attenuation of angiogenesis that can limit the ulcer healing effects of locally produced HSP 70 and TNF-α. (2) Insulin reversed this impairment of ulcer healing in diabetic rats, mainly due to the enhancement of angiogenesis and reduction in expression of cytokines in the ulcer area. (3) Classic non-steroidal anti-inflammatory drugs such as aspirin prolonged ulcer healing under diabetic conditions due to suppression of endogenous prostaglandins and the fall in the microcirculation at the ulcer margin and these effects were mimicked by selective, so called “safe” COX-2 inhibitor, rofecoxib, suggesting that both COX isoforms are important sources of prostaglandins that are essential in the ulcer healing in diabetes.

Pioglitazone, a specific ligand of peroxisome proliferator-activated receptor-gamma, accelerates gastric ulcer healing in rat

The peroxisome proliferator-activated receptor-gamma (PPAR-γ) is a member of the nuclear hormone receptor superfamily that is involved in the control of inflammation and carcinogenesis. We determined the effect of the specific PPAR-γ ligand, pioglitazone (5–40 mg/kg intragastrically), on the healing of acetic-acid gastric ulcers in rats. At day 8 after ulcer induction, the ulcer area, the gastric blood flow and mucosal expression of proinflammatory cytokines such as interleukin-1β, tumour necrosis factor alpha (TNF-α) and cyclooxygenase-1, cyclooxygenase-2, constitutive nitric oxide synthase (cNOS), inducible nitric oxide synthase (iNOS) and heat shock protein 70 (HSP70) was determined. Pioglitazone reduced the area of gastric ulcers and raised significantly the gastric blood flow at the ulcer margin and downregulated the mRNA for interleukin-1β, TNF-α, cyclooxygenase-2 and iNOS while cyclooxygenase-1 mRNA was not affected. The expression of PPAR-γ mRNA was increased in the ulcerated gastric mucosa. We conclude that pioglitazone accelerates the healing of preexisting gastric ulcers due to the hyperemia at ulcer margin and the anti-inflammatory action including suppression of interleukin-1β, TNF-α, cyclooxygenase-2 and iNOS and by an overexpression of HSP70.

Role of prostaglandins, nitric oxide, sensory nerves and gastrin in acceleration of ulcer healing by melatonin and its precursor, L‐tryptophan

Melatonin, a major hormone of pineal gland, was recently shown to attenuate acute gastric lesions induced by strong irritants because of the scavenging of free radicals but its role in ulcer healing has been little investigated. In this study we compared the effects of intragastric (i.g.) administration of melatonin and its precursor, L-tryptophan, with or without concurrent treatment with luzindole, a selective antagonist of melatonin MT2 receptors, on healing of chronic gastric ulcers induced by serosal application of acetic acid (ulcer area 28 mm2). The involvement of endogenous prostaglandins (PG), nitric oxide (NO) and sensory nerves in ulcer healing action of melatonin and L-tryptophan was studied in rats treated with indomethacin and NG-nitro-L-arginine (L-NNA) to suppress, respectively, cyclo-oxygenases (COX) and NO synthases or in those with functionally deactivated sensory nerves with capsaicin. The influence of melatonin on gastric secretion during ulcer healing was tested in separate group of rats with gastric ulcer equipped with gastric fistulas (GF). At day 8 and 15 upon the ulcer induction, the area of gastric ulcers was measured by planimetry, the mucosal blood flow (GBF) was determined by H2-gas clearance technique and gastric luminal NO2-/NO3- levels was assessed by Griess reaction. Plasma melatonin and gastrin levels were measured by specific radioimmunoassay (RIA). Biopsy mucosal samples were taken for expression of constitutive NO-synthase (cNOS) and inducible NOS (iNOS) by reverse transcriptase-polymerase chain reaction (RT-PCR). Melatonin (2.5-20 mg/kg-d i.g.) and L-tryptophan (25-100 mg/kg-d i.g.) dose-dependently accelerated ulcer healing, the dose inhibiting by 50% (ED50) of ulcer area being 10 and 115 mg/kg, respectively. This inhibitory effect of melatonin (10 mg/kg-d i.g.) and L-tryptophan (100 mg/kg-d i.g.) on ulcer healing was accompanied by a significant rise in the GBF at ulcer margin and an increase of plasma melatonin. luminal NO2-/NO3- and plasma gastrin levels. Gastric acid and pepsin outputs were significantly inhibited during the ulcer healing in melatonin-treated gastric mucosa as compared with those in vehicle-treated animals. Luzindole abolished completely the healing effects of melatonin and L-tryptophan and attenuated significantly the rise in plasma gastrin evoked by the hormone and its precursor. Indomethacin (5 mg/kg-d i.p). that blocked PG biosynthesis by 90% or L-NAME (20 mg/kg i.v), inhibitor of NOS. that suppressed luminal NO release, attenuated significantly melatonin and L-tryptophan-induced acceleration of ulcer healing and accompanying rise in GBF at ulcer margin and luminal NO release. The melatonin-induced acceleration of ulcer healing, hyperemia at ulcer margin and increase in the release of NO were enhanced when L-arginine but not D-arginine was added to L-NAME. The ulcer healing and the GBF effects of melatonin and L-tryptophan were significantly impaired in rats with capsaicin-induced denervation of sensory nerves and both, ulcer healing and the hyperemia at ulcer margin were restored in these rats by addition of exogenous CGRP to melatonin and L-tryptophan. Expression of cNOS mRNA was detected by RT-PCR in the intact gastric mucosa as well as at the edge of gastric ulcers treated with both, vehicle and melatonin, while iNOS mRNA that was undetectable in the intact gastric mucosa, appeared during ulcer healing and especially this was strongly up-regulated in the melatonin-treated gastric mucosa. We conclude that (1) exogenous melatonin and that derived from its precursor, L-tryptophan, accelerate ulcer healing probably via interaction with MT2 receptors; (2) this ulcer healing action is caused by an enhancement by melatonin of the microcirculation at the ulcer margin possibly mediated by COX-derived PG and NO because of overexpression of iNOS and (3) gastrin, which exhibits trophic activity in the gastric mucosa and calcitonin gene related peptide (CGRP), released from sensory nerves, may also contribute to the ulcer healing action of melatonin.

Classic NSAID and selective cyclooxygenase (COX)-1 and COX-2 inhibitors in healing of chronic gastric ulcers.

Prostaglandins (PG) derived from COX-1 are essential for the maintenance of mucosal integrity but COX-2 isoform synthesizes PG at a site of inflammation. Recently, COX-2 mRNA expression was demonstrated at the ulcer edge during healing of chronic gastric ulcers but the role for expression of COX-2 and its products such as PGE(2) and cytokines including interleukin (IL-1beta) and tumor necrosis factor alpha (TNFalpha) in ulcer healing remains unknown. In this study, Wistar rats with gastric ulcers produced by serosal application of acetic acid (ulcer area 28 mm(2)) received daily treatment either with: (1) vehicle (saline); (2) NS-398 (10 mg/kg-d i.g.) and Vioxx (5 mg/kg-d i.g.), both, highly specific COX-2 inhibitors; (3) meloxicam (5 mg/kg-d i.g.), a preferential inhibitor of COX-2; (4) resveratrol (10 mg/kg-d i.g.), a specific COX-1 inhibitor; (5) indomethacin (5 mg/kg-d i.g); and (6) aspirin (ASA; 50 mg/kg-d i.g.), non-selective inhibitors of both COX-1 and COX-2. At day 3, 7, and 14 after ulcer induction, the animals were sacrificed and the area of gastric ulcers was determined by planimetry and histology, gastric blood flow (GBF) at ulcer base and margin was measured by H(2) clearance technique, and blood was withdrawn for measurement of plasma IL-1beta and TNFalpha levels. The mucosal biopsy samples were taken for the determination of PGE(2) generation by RIA and expression of COX-1, COX-2, IL-1beta, and TNFalpha mRNA by RT-PCR. In vehicle-treated rats, gastric ulcers healed progressively and at day 14 the healing was completed, accompanied by a significant rise in the GBF at ulcer margin. The IL-1beta, TNFalpha, and COX-1 mRNA were detected in intact and ulcerated gastric mucosa, whereas COX-2 mRNA were upregulated only in ulcerated mucosa with peak observed at day 3 after ulcer induction. The plasma IL-1beta level was significantly increased at day 3 and 7 but then declined at day 14 to that measured in vehicle-controls. Indomethacin and ASA, which suppressed PGE(2) generation both in the non-ulcerated and ulcerated gastric mucosa, significantly delayed the rate of ulcer healing and this was accompanied by the fall in GBF at ulcer margin and further elevation of plasma IL-1beta and TNFalpha levels, which was sustained up to the end of the study. Treatment with NS-398 and Vioxx, which caused only a moderate decrease in the PGE(2) generation in the non-ulcerated gastric mucosa, delayed ulcer healing and attenuated significantly the GBF at ulcer margin and PGE(2) generation in the ulcerated tissue, while raising the plasma IL-1beta and TNFalpha similarly to those observed in indomethacin- and ASA-treated rats. Resveratrol, which suppressed the PGE(2) generation in both non-ulcerated and ulcerated gastric mucosa, prolonged ulcer healing and this was accompanied by the fall in the GBF at the ulcer margin and a significant increase in plasma IL-1beta and TNFalpha levels. We conclude that (1) classic NSAID delay ulcer healing due to suppression of endogenous PG, impairment in GBF at ulcer area, and excessive cytokine expression and release, and (2) this deleterious effect of classic NSAID on the healing of pre-existing ulcers can be reproduced by selective COX-1 and COX-2 inhibitors, suggesting that both COX isoforms are important sources of PG that appear to contribute to ulcer healing.

Role of leptin in ulcer healing

Leptin was shown to exhibit similar to cholecystokinin (CCK) cytoprotective activity against acute gastric lesions, but its role in ulcer healing has not been examined. The aims of this study were: (1) to compare the effects of exogenous leptin to those of CCK on the course of healing of chronic gastric ulcers; (2) to study the gene and protein expression of leptin at the ulcer margin during ulcer healing; and (3) to assess the effects of leptin administration on the mucosal gene expression of main growth factor such as transforming growth factor alpha (TGFα). Gastric ulcers were produced in rats by the acetic acid method. Rats with ulcers were divided in following treatment groups: (1) vehicle; (2) leptin (10 μg/kg i.p.); (3) CCK (10 μg/kg s.c.); and (4) leptin or CCK with or without tyrphostin A46 (200 μg/kg i.p.), an inhibitor of epidermal growth factor (EGF)-receptor tyrosine kinase or N G-nitro- l-arginine (20 mg/kg i.g.), a blocker of nitric oxide synthase. Animals were euthanized 9 days after ulcer induction. The area of gastric ulcers and the gastric blood flow at the ulcer area were determined. In addition, mucosal biopsy samples were taken from the ulcer area for histological evaluation as well as for the determination of mRNA and protein expression for leptin and constitutive nitric oxide synthase (cNOS) and inducibile nitric oxide synthase (iNOS) by reverse-transcriptase polymerase chain reaction (RT-PCR) and Western blot, respectively. In addition, the gene expression for TGFα was analyzed by RT-PCR. Both leptin and CCK reduced significantly the ulcer area as compared to vehicle-treated group by ∼50%. The treatment with tyrphostin or N G-nitro- l-arginine reversed in part the acceleration of ulcer healing by leptin and CCK. The expression of leptin mRNA and protein was significantly increased at the ulcer edge. The leptin-induced acceleration of ulcer healing was associated with increased expression of transcripts for TGFα as well as increased mRNA and protein expression for cNOS and iNOS at the ulcer margin. We conclude that leptin accelerates ulcer healing by mechanisms involving the up-regulation of TGFα and increased production of nitric oxide due to up-regulation of cNOS and iNOS in the ulcer area.

In vivo selectivity of nonsteroidal antiinflammatory drugs and gastrointestinal ulcers in rats.

The aim of the present work was to study in vivo COX-2-COX-1 selectivity of 16 nonsteroidal anti-inflammatory drugs (NSAIDs) in equipotent ulcerogenic doses in two in vivo experimental models. Indomethacin, ibuprofen, nimesulide, aceclofenac, aspirin, sodium diclofenac, meloxicam, naproxene, paracetamol, piroxicam, tenoxicam, nabumetone, ketoprofen, mefenamic acid, etodolac, and ketorolac were administered to female Wistar rats (N = 10 each group). In experiment I, solid food plus subcutaneous NSAIDs were given. In experiment II, NSAIDs were given by oral gavage and in bolus. Macroscopic gastric antral ulcer area (30%) and intestinal erosiva area (295 mm2) in experiment I and necrotic gastric fundus area (65%) and erosive intestinal area (182 mm2), "in vivo" the NSAIDs COX-1 was showed. Neutrofilia assessed in gastric intestinal mucosa where also ibuprofen and paracetamol not given neotrophilic infiltration. In conclusion, COX-2-COX-1 selectivity was demonstrated in vivo with the drugs aceclofenac, nabumetone, meloxicam, nimesulide, and paracetamol.

Expression of Epidermal Growth Factor and Transforming Growth Factor Alpha during Ulcer Healing: Time Sequence Study

Background: Growth factors such as epidermal growth factor (EGF) and transforming growth factor alpha (TGFα) have been shown to share common receptor (EGFR) and to accelerate ulcer healing due to stimulation of cell proliferation, but the time sequence of expression of EGF and TGFα during ulcer healing has not been investigated. In this study the rate of cell proliferation and the gastric secretion and gene expression of mRNA for EGF and TGFα were determined during ulcer healing. Methods: Gastric ulcers were induced in 150 Wistar rats by serosal application of 100% acetic acid (ulcer area, 14 mm). Some of these animals were also equipped with a gastric fistula for the assessment of gastric secretion during ulcer healing. The animals were killed 0, 2, 4, 6, or 8 days after ulcer induction, and the ulcer area was determined. The mucosal sections with gastric ulcer were immunostained for proliferating cell nuclear antigen (PCNA) and for immunoexpression of EGF, TGFα, and EGFR. The expression of mRNA EGF and mRNA TGFα was also determined in the ulcer margin by reverse transcriptase (RT) polymerase chain reaction (PCR) using specific primers. Results: Two, 4, 6, and 8 days after ulcer induction the gastric ulcer area was gradually reduced from the initial size (day 0) by 47%, 70%. 80%, and 87%. respectively, and this was accompanied by an increase in PCNA with its maximum on day 4. The gastric acid and pepsin secretion was significantly reduced by 75% and 79%, respectively, on day 2 after ulcer induction but then the secretion tended to return to normal value by day 8. The expression of EGF, TGFα, and EGFR was negligible on day 0 but increased significantly during the healing, reaching maximum on day 4. Expression of EGF mRNA was detected on days 2, 4, and 6, and that of TGFα mRNA on days 2, 4. 6. and 8 after ulcer induction, with the most intense signals for both transcripts observed on day 2. Conclusions: 1) The enhancement in cell proliferation during ulcer healing may be mediated by increased release of EGF and TGFα; 2) the expression of EGF and TGFα mRNA precedes the overexpression of these growth factors at the ulcer margin during ulcer healing; and 3) the overexpression of growth factors coincides with the inhibition of gastric secretion and increased blood flow at the ulcer margin, indicating that these factors affect gastric secretion and blood flow in the course of ulcer healing.

A randomized, double-blind comparison of placebo, etodolac, and naproxen on gastrointestinal injury and prostaglandin production

Background: NSAIDs frequently cause gastrointestinal injury and increase the risk of ulcer complications. We compared an NSAID suggested to cause less gastric injury (etodolac) with a standard NSAID (naproxen) and a placebo in a 4-week double-blind trial assessing the effects on gastroduodenal injury, symptoms, and prostaglandin production in healthy volunteers. Methods: Fifty-two healthy volunteers not taking NSAIDs, alcohol, antibiotics, bismuth, or anti-ulcer drugs and with a normal endoscopic examination were randomly assigned to identical drugs: placebo, etodolac 400 mg, or naproxen 500 mg b.i.d. for 4 weeks. Endoscopies with biopsies were repeated at weeks 1 and 4. The number and dimensions of ulcers and erosions were recorded to quantitate injury. Results: At week 1 the mean number and area of gastric ulcers per subject were greater with naproxen than placebo or etodolac (area: naproxen, 7.4 mm 2; placebo, 0.6 mm 2, p = 0.02 vs naproxen; etodolac, 2.1 mm 2, p = 0.06 vs naproxen). Ulcer scores at week 4 were low and comparable in the three groups. The mean number and area of gastric erosions per subject were greatest with naproxen at both weeks 1 and 4 (week 4 area: naproxen, 58.3 mm 2; placebo, 29.0 mm 2; etodolac, 13.9 mm 2, p < 0.02, naproxen vs placebo and vs etodolac). Placebo injury was presumably due to biopsies at prior endoscopy. Gastric mucosal prostaglandin E 2 production did not change significantly from baseline after 1 or 4 weeks of treatment with placebo or etodolac, but did decrease significantly with naproxen (week 0, 1689; week 1, 479; week 4, 577 pg/mg protein). Gastrointestinal symptoms were present in only 1 (5%) of 20 visits in which endoscopy showed no erosions or ulcers vs 21 (26%) of 82 visits in which a mucosal defect was identified ( p = 0.066). Conclusions: Gastric injury with 4 weeks of etodolac is comparable to that seen with placebo and significantly less than that occurring with naproxen, presumably due to the fact that etodolac does not suppress gastric mucosal prostaglandin production, whereas naproxen leads to a significant reduction. (Gastrointest Endosc 1995;42:428-33.)

Effects of quercetin on the gastrointestinal tract in rats and mice

AbstractIntraperitoneal administration of quercetin (6.25–50 mg/kg) significantly (p&lt;0.5‐0.01) reduced intestinal transit in mice and this effect was antagonized by yohimbine and phentolamine but not by atropine or naloxone. Quercetin (12.5–50 mg/kg) reduced also (p&lt;0.05‐0.01) intraluminal accumulation of fluid and diarrhoea induced by castor oil and these effects were antagonized by yohimbine. Finally quercetin (12.5–50 mg/kg) reduced the area of gastric ulcer but not the number. It is suggested that α2‐adrenergic receptors mediate the effect of quercetin on intestinal motility and secretion.