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Background: Endothelin-1, a key mediator of inflammatory processes, is produced from its biologically inactive precursor, big ET- by the action of endothelin converting enzyme-1(ECE-1). In this study, we applied the animal model of H. pylori lipopolysaccharide-induced gastritis to assess the effect of three different types of antiulcer agents on the gastric mucosal expression of ECE-1 activity., Methods: Rats, pretreated twice daily for 3 days with proton pump inhibitor, omeprazole at 40 mg/kg, gastroprotective agent, sulglycotide at 200 mg/kg, H2-receptor antagonist, ebrotidine at 100 mg/kg or the vehicle, were subjected to intragastric application of H. pylori lipopolysaccharide at 50 microg/animal, and after 2, and 4 additional days on the drug or vehicle regimen their mucosal tissue used for histologic and biochemical assessment., Results: In the absence of antiulcer agents, H. pylori lipopolysaccharide elicited a pattern of mucosal inflammatory responses resembling that of acute gastritis which reached a maximum by the 4th day and were accompanied by a 4.1-fold increase in the mucosal expression of ECE-1 activity and an 8.8-fold enhancement in TNF-alpha. Treatment with sulglycotide led to a 56.7% reduction in the extent of mucosal inflammatory involvement, the mucosal expression of ECE-1 activity fell by a 40.5% and the level of TNF-alpha declined by a 69%. Ebrotidine produced a 50.9% decrease in the extent of mucosal inflammatory involvement, a 33.6% decrease in the expression of ECE-1 activity and a 64.1% decline in TNF-alpha, whereas omeprazole elicited a 37.6% reduction in the extent of mucosal inflammatory involvement and a 29.5% decrease in TNF-alpha, but had no effect on the lipoploysaccharide-induced increase in the mucosal expression of ECE-1 activity., Conclusions: The findings implicate up-regulation of ECE-1 in triggering the induction of TNF-alpha and propagation of gastric mucosal inflammatory responses to H. pylori. We also show that omeprazole, in contrast to sulglycotide and ebrotidine, fails to counter the enhancement in the mucosal expression of ECE-1 caused by H. pylori- lipopolysaccharide.

Background: Helicobacter pylori is recognized as a primary etiologic factor in the development of gastric disease and the product of particular significance to the virulent action of the bacterium is its cell wall lipopolysaccharide. We applied the animal model of H. pylori lipopolysaccharide-induced acute gastritis to study the effect of antiulcer agents, omeprazole and sucralfate, on the course of mucosal inflammatory responses by analyzing the interplay between the extent of epithelial cell apoptosis and the mucosal expression of endothelin-1 (ET-1), tumor necrosis factor-alpha (TNF-alpha), and the activity of constitutive (cNOS) and inducible (NOS-2) nitric oxide synthase., Methods: Rats pretreated twice daily for 3 consecutive days with omeprazole at 40 mg/kg, sucralfate at 100 mg/kg or the vehicle, were subjected to intragastric application of H. pylori lipopolysaccharide at 50 microg/animal, and after 2, 4, and 10 additional days on the antiulcer drug or vehicle regimen their mucosal tissue used for histologic and biochemical assessment., Results: In the absence of antiulcer agents, H. pylori lipopolysaccharide elicited within 2 days a pattern of acute mucosal inflammatory responses accompanied by a massive epithelial cell apoptosis, a 2.9-fold increase in the mucosal expression of ET-1, an 11.7-fold enhancement in TNF-alpha, and a 9.3-fold increase in NOS-2, while cNOS activity showed a 5.5-fold decrease. The extent of mucosal inflammatory involvement reached a maximum by the 4th day and showed a decline by the 10th day. This was reflected in a marked reduction in epithelial cell apoptosis, decrease in the mucosal expression of ET-1, TNF-alpha and NOS-2, and the recovery in cNOS activity. Comparing to the vehicle controls, treatment with proton pump inhibitor, omeprazole, led at the end of a 10 day period to a 48.3% reduction in the extent of mucosal inflammatory involvement elicited by H. pylori lipopolysaccharide, while a 74.2% reduction in the mucosal inflammatory involvement was achieved with gastroprotective agent, sucralfate. Moreover, this advantage of sucralfate over omeprazole in countering the lipopolysaccharide-induced changes was reflected at the end of 10 day treatment period in a 20.4% greater decrease in apoptosis, a 47.5% greater reduction in TNF-alpha and a 50.7% greater reduction in ET-1. However, both agents exerted similar influence on the restoration of gastric mucosal cNOS activity and showed a comparable effect at the end of a 10 day treatment in countering the lipopolysaccharide-induced increase in the expression of NOS-2., Conclusions: The findings suggest that an increase in the mucosal ET-1 level elicited by H. pylori lipopolysaccharide, combined with a decline in cNOS may be responsible for the induction of TNF-alpha and triggering the inflammatory process. We also show that sucralfate exhibits greater efficacy than omeprazole in suppressing the H. pylori-induced mucosal inflammatory responses. This property of sucralfate may well be due to its ability to suppress the mucosal rise in ET-1.

Unlabelled: Helicobacter pylori lipopolysaccharide is a primary virulence factor responsible for eliciting acute mucosal inflammatory responses associated with H. pylori infection. In this study, we applied the animal model of H. pylori lipopolysaccharide-induced acute gastritis to assess the effect of antiulcer agent, ebrotidine, on the gastric mucosal inflammatory responses by analyzing the interplay between the activity of a key apoptotic caspase, caspase-3, epithelial cell apoptosis, and the expression of inducible nitric oxide synthase (NOS-2)., Methods: Rats, pretreated twice daily with ebrotidine at 100 mg/kg, or the vehicle, were subjected to intragastric application of H. pylori lipopolysaccharide at 50 microg/animal, and after 4 additional days on the antiulcer drug or vehicle regimen their mucosal tissue used for histologic assessment, assays of epithelial cells apoptosis, and the measurements of caspase-3 and NOS-2 activities., Results: In the absence of antiulcer agent, H. pylori lipopolysaccharide induced acute reaction characterized by the inflammatory infiltration of the lamina propria, hyperemia, and epithelial hemorrhage. This was accompanied by an 11.2-fold increase in epithelial cell apoptosis, a 6.5-fold induction in mucosal expression of NOS-2, and a 5.4-fold increase in caspase-3 activity. Treatment with H2-receptor antagonist ebrotidine, also known for its gastroprotective effects, produced a 50.9% reduction in the extent of mucosal inflammatory changes elicited by H. pylori lipopolysaccharide and an 82.5% decrease in the epithelial cells apoptosis, while the activity of caspase-3 decreased by 33.7% and that of NOS-2 showed a 72.8% decline., Conclusions: The findings implicate caspase-3 involvement in gastric mucosal inflammatory responses to H. pylori lipopolysaccharide, and point towards participation of NOS-2 in the amplification of the cell death-signaling cascade. Our study also demonstrate that ebrotidine exerts modulatory effect on the H. pylori-induced mucosal inflammatory responses by interfering with the events propagated by NOS-2 and caspase-3.

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.

The effect of the antiulcer agent sulglycotide on gastric epithelial cell apoptosis and the expression of tumor necrosis factor-alpha (TNF-alpha) and interleukin-4 (IL-4) during Helicobacter pylori lipopolysaccharide-induced acute gastritis was investigated. Rats, pretreated twice daily for 3 consecutive days with sulglycotide at 200 mg/kg or vehicle, were subjected to surface epithelial application of H. pylori lipopolysaccharide (50 microg/animal), and, after 4 additional days on the drug or vehicle regimen, their mucosal tissue was used for histologic assessment, quantitation of TNF-alpha and IL-4, and the assay of epithelial cell apoptosis. In the absence of sulglycotide, H. pylori lipopolysaccharide caused acute mucosal responses manifested by the inflammatory infiltration of the lamina propria with lymphocytes and plasma cells, edema, hyperemia, and epithelial hemorrhage. These responses were accompanied by an 11-fold increase in epithelial cell apoptosis and a 9-fold enhancement of the mucosal expression of proinflammatory cytokine TNF-alpha. However, the mucosal expression of regulatory cytokine IL-4 decreased by 15%. Treatment with sulglycotide produced significant (56.6%) reduction in the extent of acute mucosal inflammatory changes caused by H. pylori lipopolysaccharide. Moreover, the effect of sulglycotide was manifested in an 88.3% reduction in the epithelial cell apoptosis and a 69.1% decrease in the mucosal expression of TNF-alpha, whereas the expression of IL-4 showed only marginal (6%) enhancement. The results suggest that the cytoprotective agent sulglycotide suppresses the inflammatory and apoptotic events elicited in gastric mucosa by H. pylori lipopolysaccharide through stimulation of TNF-alpha expression.

Background: Helicobacter pylori lipopolysaccharide is emerging as a primary factor in the bacterium virulence, and its involvement in causing gastric mucosal responses typical of gastritis has recently been shown. In this study we investigated the effect of the antiulcer agent sucralfate on the expression of regulatory cytokines, tumor necrosis factor-alpha (TNF-alpha) and interleukin-4 (IL-4), and epithelial cell apoptosis during H. pylori lipopolysaccharide-induced acute gastritis., Methods: The experiments were conducted with rats pretreated intragastrically twice daily for 3 days with sucralfate at 100 mg/kg or the vehicle. The rats were then subjected to intragastric surface epithelial application of H. pylori lipopolysaccharide at 50 microg per animal and maintained on the sucralfate or vehicle regimen for an additional 4 days. The animals were killed 16 h after the last dose, and their gastric mucosal tissue used for histologic assessment, quantitation of TNF-alpha and IL-4 expression, and the assay of epithelial cell apoptosis., Results: In the absence of sucralfate, H. pylori lipopolysaccharide induced acute mucosal responses characterized by the inflammatory infiltration of the lamina propria, hyperemia, and epithelial hemorrhage. This was accompanied by an 11-fold increase in gastric epithelial cell apoptosis and a 9-fold enhancement of the mucosal expression of TNF-alpha, but the level of IL-4 fell by 15%. Intragastric administration of sucralfate produced a 62% reduction in the extent of mucosal damage caused by H. pylori lipopolysaccharide, a 51% decrease in the mucosal expression of TNF-alpha, and a 7-fold reduction in the extent of epithelial cell apoptosis, whereas the expression of IL-4 increased by 52%., Conclusions: Gastric mucosal inflammatory responses to H. pylori lipopolysaccharide are characterized by a massive enhancement of the proinflammatory cytokine TNF-alpha and epithelial cell apoptosis and repression of IL-4. Our data also show that sucralfate is capable of inducing expression of the regulatory cytokine IL-4 and the suppression of apoptotic events triggered in gastric mucosa by the increase in TNF-alpha that is elicited by H. pylori lipopolysaccharide.

1. Gastric mucus from Helicobacter pylori-positive patients with active chronic gastritis type B, before and after successful combination therapy consisting of the H2 receptor antagonist ebrotidine (400 mg) and amoxicillin (1,000 mg) administered b.i.d., in the morning and at bedtime for a period of 2 weeks followed by administration of ebrotidine alone for the subsequent 4 weeks, was assessed for the changes in physicochemical properties associated with the mucosal protective potential and anti-H. pylori activity. 2. The results of physicochemical measurements revealed that eradication of H. pylori associated with the successful therapy for active chronic gastritis type B with ebrotidine-amoxicillin was accompanied by a 36% increase in gastric mucus hydrogen ion retardation capacity, a 1.5-2.1-fold increase in mucus gel viscosity and a 2.4-fold increase in its hydrophobicity. 3. The beneficial changes brought about by the ebrotidine-amoxicillin therapy in the physical properties of gastric mucus were also manifest in a 2.7-fold enhancement in the proportion of the high-molecular-weight polymeric mucin form responsible for the maintenance of gastric mucus gel integrity. 4. Moreover, assays of H. pylori aggregating titer of gastric mucus revealed that the successful combination therapy with ebrotidine and amoxicillin led to a 3.8-fold increase in mucin anti-H. pylori activity. 5. The results demonstrate that combination therapy with ebrotidine and amoxicillin for H. pylori eradication leads to a marked improvement in the protective qualities of gastric mucus essential for the preservation of mucosal integrity and enhances the inherent mucosal defense against H. pylori infection.

Infection with H. pylori is now recognized as a major factor in the pathogenesis of gastric disease. Here, we examined the susceptibility of epidermal growth factor (EGF), basic fibroblast growth factor (bFGF), transforming growth factor-beta (TGF beta) and platelet derived growth factor (PDGF) to degradation by H. pylori protease, and assessed the effect of a cytoprotective agent, sulglycotide, on this process. The 125I-labeled EGF, bFGF, TGF beta and PDGF were incubatet with H. pylori protease, obtained from the filtrates of saline washes of the bacterium culture, in the presence of 0-100 micrograms sulglycotide. The results showed that, under the assay conditions, H. pylori protease caused only 5% degradation of EGF and 7% degradation of bFGF. However, the protease evoked a 61.7% degradation of PDGF and a 62.3% degradation of TGF beta. Introduction of sulglycotide to the reaction assay system caused a dose-dependent inhibition in PDGF and TGF beta proteolysis by the H. pylori enzyme. The maximal inhibitory effect was obtained with sulglycotide at 100 micrograms/ml, at which dose an 84.4% decrease in PDGF and 88.3% decrease in TGF beta degradation was achieved. The results provide a strong evidence for the effectiveness of sulglycotide in the protection of gastric mucosal growth factors against degradation by H. pylori.

Background: The progression of the events associated with gastric mucosal repair is controlled in an orderly manner by a plethora of the extracellular cues exerting their effect on the cell cycle regulatory proteins, cyclins, and cyclin-dependent kinases, the expression of which varies through the cycle stages. The purpose of this study was to evaluate the expression of cyclin-dependent kinase (Cdk2) and proliferating cell nuclear antigen (PCNA) during chronic ulcer healing with sulglycotide., Methods: Rats with experimentally induced gastric ulcers were treated twice daily for 14 days either with sulglycotide at 200 mg/kg or vehicle, and at different stages of the treatment their stomachs were used for macroscopic damage assessment and quantitation of gastric mucosal Cdk2 and PCNA expression., Results: The assays showed that the ulcer healing was accompanied by an increase in mucosal expression of Cdk2 and PCNA. The maximum increase in Cdk2 (2.3-fold) occurred by the 4th day of healing. An accelerated ulcer healing (10 days) with sulglycotide treatment was reflected in a marked enhancement in Cdk2 expression attained a maximum 2.1-fold increase by the 6th day of treatment and remained substantially increased for up to 10 days., Conclusions: The findings show a complex interplay between the extracellular cues and cell cycle regulatory proteins, an orderly progression that drives the mucosal repair process. We also show that the gastroprotective agent sulglycotide is capable of affecting the expression of the cell cycle regulatory proteins that control cell cycle progression through G1 and into S phase.

1. The effect of antiulcer agents, ebrotidine and sucralfate, on the expression of gastric mucosal proliferating cell nuclear antigen (PCNA) and cyclin-dependent kinase (p34Cdk2) during chronic ulcer healing was examined. 2. Rats with experimentally induced gastric ulcers were treated twice daily for 14 days with either ebrotidine at 100 mg/kg, sucralfate at 100 mg/kg or vehicle, and at different stages of treatment their stomachs were used for quantitization of gastric mucosal PCNA and Cdk2 expression. 3. The assays revealed that the ulcer healing was accompanied by a marked elevation in mucosal expression of PCNA and Cdk2. The maximum increase in PCNA (4.7-fold) occurred by the second day of healing, and the expression of Cdk2 reached a maximum increase (2.3-fold) by the fourth day. 4. Accelerated ulcer healing with ebrotidine (7 days) and sucralfate (8 days) treatments was reflected in a significant enhancement of PCNA and Cdk2 expression. By the second day of treatment, ebrotidine evoked a 15-fold increase in PCNA expression, and sucralfate produced an 11.8-fold enhancement. The mucosal expression of Cdk2 attained a maximum of 4.3-fold increase over that of the controls by the sixth day of healing with ebrotidine, and a fivefold increase in Cdk2 expression occurred by the fourth day of ulcer treatment with sucralfate. 5. The findings implicate cell cycle regulatory proteins in the processes to leading to mucosal repair and suggest that the two drugs exert a similar effect on the expression of proteins that control cell cycle progression.

In this study, we investigated the inhibitory effect of Helicobacter pylori lipopolysaccharide on the binding of somatostatin to gastric epithelial cell membrane receptor, and assessed the effect of antiulcer agent, sucralfate, on this process. The assays conducted with the somatostatin receptor, purified from the solubilized epithelial cell membranes by affinity chromatography on Affi-Gel-[D-Tryp8]SRIF-14, revealed a dose-dependent inhibition in receptor-somatostatin binding by the lipopolysaccharide which reached a maximum of 94.1%. This effect of H. pylori lipopolysaccharide was countered by sucralfate that produced a 92.5% restoration in receptor-somatostatin binding at 70 micrograms/ml of the drug. The findings demonstrate that sucralfate is capable of reversing the interference by H. pylori lipopolysaccharide with somatostatin-receptor binding on gastric mucosal G-cells.

Background: The gastric injury associated with nonsteroidal anti-inflammatory drug (NSAID) therapy has been linked to the detrimental effects of the agents on the processes of prostaglandin generation, leukocyte adherence, superoxides production, and mucosal cell proliferation. In the present study we investigated the expression of tumor necrosis factor-alpha (TNF-alpha) and epithelial cell apoptosis during indomethacin-induced gastric mucosal injury and evaluated the effect of two antiulcer agents on this process., Methods: The experiments were carried out with groups of rats subjected to intragastric pretreatment with 40 mg/kg omeprazole, 100 mg/kg ebrotidine, or vehicle, followed 30 min later by an intragastric dose of indomethacin at 60 mg/kg. After 2 h the animals were killed, and the gastric mucosal tissue used for macroscopic damage assessment, quantitation of TNF-alpha expression, and the assay of epithelial cell apoptosis., Results: In the absence of antiulcer drugs, indomethacin caused extensive multiple hemorrhagic lesions accompanied by a 47% increase in mucosal expression of TNF-alpha and a dramatic (> 300-fold) enhancement in gastric epithelial cell apoptosis. Pretreatment with a proton pump inhibitor, omeprazole, produced only marginal (6-8%) reduction in the extent of mucosal damage caused by indomethacin, whereas the mucosal expression of TNF-alpha decreased by 15% and the apoptotic DNA fragmentation by 10-13%. In contrast, the H2-receptor antagonist ebrotidine, also known for its gastroprotective effects, not only successfully prevented (98.3%) the enhancement in mucosal TNF-alpha expression caused by indomethacin but also caused a 54% reduction in the epithelial cell apoptosis. These effects of ebrotidine were, furthermore, reflected in a 90.2% prevention in the gastric mucosal damage., Conclusions: Our findings provide new insights into the mechanism of gastric injury caused by NSAIDs and show that ebrotidine protection against indomethacin-induced mucosal damage occurs through the inhibition of epithelial cell apoptosis triggered by the enhancement in the mucosal TNF-alpha expression. Our data also show that omeprazole does not possess antiapoptotic properties.

In this study, we investigated gastric epithelial cells' apoptosis and tumor necrosis factor-alpha (TNF-alpha) expression with ethanol-induced mucosal injury, and the effect of antiulcer agents on this process. Rats received intragastric pretreatment with the agent or vehicle followed 1h later by ethanol, and after 30 min the gastric mucosa was assessed for TNF-alpha and apoptosis. In the absence of antiulcer agents, ethanol caused extensive mucosal lesions accompanied by a 9.5-fold enhancement in apoptosis and a 2.5-fold increase in TNF-alpha. Pretreatment with omeprazole evoked a 54% reduction in TNF-alpha, but had no effect on ethanol-induced mucosal damage or apoptosis, the sucralfate reduced the extent of mucosal damage by 95%, apoptosis by 39% and TNF-alpha by 52%, while ebrotidine not only prevented mucosal injury and rise in TNF-alpha, but also caused a 70% reduction in epithelial cells' apoptosis. The results demonstrate that ethanol-induced gastric epithelial cells apoptosis triggered by the enhancement in mucosal TNF-alpha is efficiently counteracted by ebrotidine.

The effect of gastroprotective agents, sucralfate and sulglycotide, on the in vitro activity of H. pylori urease was investigated. The bacterium was subjected to sonication, centrifuged, and the supernatant used as an enzyme source. The assays revealed that the rate of urea degradation was proportional to enzyme protein up to 100 micrograms and remained constant with time for 10 min. Introduction of sucralfate or sulglycotide to the assay system led to the reduction in the rate of ammonia production. With both drugs the optimal inhibition was attained at 10 micrograms/ml, at which dose a 63.1% decrease in urease activity occurred with sucralfate and a 70.2% inhibition was obtained with sulglycotide. The findings demonstrate that the inhibitory action of sucralfate and sulglycotide on H. pylori urease activity may be of value in the treatment of gastric disease associated with H. pylori infection.

1. Among the consequences of H. pylori infection is an increase in gastric acid secretion due to the impairement in feedback inhibition by somatostatin. Here, we show that lipopolysaccharide from H. pylori inhibits the binding of somatostatin to gastric mucosal receptor, and that antiulcer agents, ebrotidine and sulglycotide, are capable of countering this effect. 2. The somatostatin receptor was prepared from the solubilized gastric mucosal epithelial cell membranes by affinity chromatography on Affi-Gel-bound [D-Tryp8] SRIF-14 and used in the binding assays for 125I-labeled somatostatin in the presence of H. pylori lipopolysaccharide and antiulcer agents. 3. The assays revealed a dose-dependent inhibition in the receptor-somatostatin binding by the lipopolysaccharide which reached a maximum of 94.1%. The effect of H. pylori lipopolysaccharide was countered by ebrotidine and sulglycotide, which at their optimal doses produced 94.9% and 84% restoration in somatostatin-receptor binding, respectively. 4. The results demonstrate that the antiulcer agents, ebrotidine and sulglycotide, possess the ability to counteract the H. pylori interference with somatostatin regulatory effect on gastric acid secretion.

Ebrotidine (N-[(E)-[[2-[[[2-[(diaminomethylene)amino]-4-thiazolyl] methyl]thio]ethyl]amino]methylene]-4-bromo-benzenesulfonamide, CAS 100981-43-9, FI-3542) is a new antiulcer drug which combines the properties of an H2-receptor antagonist with those of a cytoprotective agent. The cytoprotective properties of ebrotidine are not dependent upon endogenous prostaglandin generation, but stem from the ability of the drug to induce mucosal responses manifested in the enhanced physicochemical characteristics of mucus gel. These include the increase in mucus gel dimension, viscosity, hydrophobicity and hydrogen ion retardation capacity. Improvements in mucus gel protective qualities with ebrotidine are directly related to the ability of the drug to enhance the synthesis and secretion of sulfo- and sialomucins and phospholipids of gastric mucus and to promote mucin macromolecular assembly. An equally important property of ebrotidine in promotion of ulcer healing is its capability to enhance the gastric mucosal expression of integrin receptors for the interaction with proteins of the extracellular matrix such as laminin. Furthermore, the accelerated ulcer healing with ebrotidine is reflected in a marked increase in the mucosal expression of EGF and PDGF receptors. The drug has also been shown to modulate the processes associated with cell cycle progression during ulcer healing, and is known to protect the gastric epithelial integrity from calcium imbalance. Thus, ebrotidine, unlike other H2-blockers, has a unique ability to promote the event essential for mucosal repair and the maintenance of mucosal integrity. These features make ebrotidine a drug of great potential in the treatment of ulcer disease.

1. H. pylori infection causes the loss of mucus coat continuity and its patchy appearance. Here, we present evidence that the bacterium, through its cell wall lipopolysaccharide, disrupts the interaction between mucin and its mucosal receptor, and that ebrotidine is capable of counteracting this process. 2. The receptor for mucin, isolated from the solubilized gastric epithelial cell membrane by affinity chromatography on Sepharose-bound wheat germ agglutinin, displayed a molecular weight of 97 kDa and exhibited specific affinity towards mucin-coated surface. 3. The mucin binding to the receptor was susceptible to the inhibitin by H. pylori lipopolysaccharide and reached a maximum of 91%. This effect of the lipopolysaccharide was counteracted by ebrotidine, which caused a dose-dependent relief of the lipopolysaccharide inhibitory effect with maximum restoration in mucin-receptor binding of 51% at 60 microliters/ml ebrotidine. 4. The results show that H. pylori, through its lipopolysaccharide, is capable of disrupting the integrity of mucus perimeter of gastric mucosal defense and that antiulcer agent, ebrotidine, counteracts this untoward effect.

Helicobacter pylori is considered a primary factor in the pathogenesis of gastric disease, and the successful mucosal colonization is linked to its urease activity. In this study, we assessed the effect of antiulcer agent, ebrotidine, on the in vitro activity of H. pylori urease. The results of assays showed a dose-dependent inhibition of the urease activity. A maximum inhibition (77%) in H. pylori urease activity occurred at 2.1 microM ebrotidine. A known H2-blocker, ranitidine, in a parallel experiment gave a maximal inhibition of 73% at a considerably higher concentration (6.4 microM). The results demonstrate that ebrotidine with its combined acid suppressant and anti-H. pylori activities offers an excellent choice in the treatment of H. pylori associated gastric disease.

1. Gastric mucus from duodenal ulcer patients before and following therapy with a new antiulcer agent, ebrotidine, at 400, 600 and 800 mg dose was examined for changes in the physicochemical qualities and anti-H. pylori activity. 2. The results of physical measurements revealed that successful therapy with ebrotidine was accompanied by a 25% increase in gastric mucus viscosity, and a 20% increase in H+ retardation capacity, while its hydrophobicity increased by 11%. 3. The enhancement in the physical properties of mucus with ebrotidine therapy were also reflected in a marked (2.6-2.9-fold) increase in the proportion of the high molecular weight form of mucin. Furthermore, following therapy with ebrotidine, the gastric mucins showed a 36% higher content of sulfate as compared to that before the therapy. 4. Assays on the H. pylori aggregating titer of gastric mucin revealed that ebrotidine therapy at all three doses evoked a 4-fold increase in mucin anti-H. pylori activity. 5. The data demonstrate that duodenal ulcer therapy with ebrotidine leads to a marked improvement in the protective qualities of gastric mucus essential for the maintenance of mucosal integrity and enhances the inherent mucosal defense against H. pylori infection.

1. The effect of an antiulcer agent, sulglycotide, on mucosal expression of EGF and PDGF receptors with chronic ulcer healing was investigated. 2. Rats with experimentally-induced gastric ulcers were treated twice daily for 14 consecutive days, either with sulglycotide at 200 mg/kg or vehicle, and at different stages of treatment used for quantitation of gastric mucosal EGF and PDGF receptors. 3. The ulcer healing was accompanied by an increase in mucosal expression of both types of receptors. A 1.8-fold increase in EGF and 3.1-fold increase in PDGF receptors occurred by the 4th day following the development of ulcer and reached a maximum of 2.4-3.9-fold increase by the 10-14th day. 4. Treatment with sulglycotide caused accelerated ulcer healing accompanied by a significant enhancement in the receptors expression. A 2.3- and 3.6-fold increase in EGF and PDGF receptor expression occurred by the 4th day of sulglycotide treatment, reaching a 5.5- and 5.6-fold respective increase by the 10th day when the ulcer essentially healed. 5. The results attest to the ability of sulglycotide to stimulate the gastric mucosal proliferative activities associated with ulcer healing.

Background: Infection with Helicobacter pylori is regarded as a primary factor in the pathogenesis of gastric disease, and successful therapies now include a combination of antiulcer drugs with antimicrobial agents. In this study, we investigated the effect of sucralfate and omeprazole on the vitro anti-H. pylori activity of metronidazole, erythromycin, tetracycline, and amoxycillin., Methods: Aliquots of H. pylori culture were transferred to the wells containing different concentrations of antibiotics either alone or in the presence of various doses of sucralfate and omeprazole and incubated for 3 days for MIC evaluation., Results: The assays in the absence of sucralfate and omeprazole gave MIC value 0.10 mg/l for erythromycin, 0.12 mg/l for amoxycillin, 0.15 mg/l for tetracycline, and 14 mg/l for metronidazole. Inclusion of sucralfate evoked a 28% enhancement in the MIC of metronidazole, 2.5-fold in tetracycline, 8-fold in erythromycin, and 2-fold in amoxycillin. In the presence of omeprazole, the MIC of erythromycin improved 4-fold, tetracycline 1.6-fold, and amoxycillin, 2-fold., Conclusions: The results demonstrate that sucralfate enhances the anti-H. pylori activity of antibiotics and that this effect is comparable to that of omeprazole.

Background: Among the disturbances in gastric mucosal defense associated with Helicobacter pylori infection is the loss of mucus coat continuity, which results in a severe disturbance in the ability of mucus coat to maintain its functions as the pre-epithelial element of gastric mucosal defense. Here, we show that H. pylori, through its cell-wall lipopolysaccharide, disrupts the interaction between gastric mucin and its mucosal receptor, and that sucralfate is capable of counteracting this untoward effect of the bacterium., Methods: The receptor was isolated from octylglucoside-solubilized gastric mucosal epithelial cell membranes by affinity chromatography on Sepharose-bound wheat germ agglutinin and following iodination with 125I, used in the binding assays for mucin in the presence of H. pylori lipopolysaccharide and sucralfate., Results: Preincubation of the receptor protein with H. pylori lipopolysaccharide led to a decrease in mucin binding. The inhibitory effect was proportional to the concentration of lipopolysaccharide and reached a maximum of 91% at 30 micrograms/ml. The effect of H. pylori lipopolysaccharide was countered by sucralfate, which caused a dose-dependent relief of the inhibitory effect. The maximum (75%) restoration in mucin-receptor binding occurred at 60 micrograms/ml sucralfate., Conclusions: The results provide strong evidence for the effectiveness of sucralfate in preventing the loss of gastric mucus coat continuity caused by H. pylori.

The mucin isolated from gastric secretion of duodenal ulcer patients before and after therapy with a new antiulcer agent, ebrotidine, was assessed for H. pylori aggregating activity and macromolecular organization. Analyses of mucin molecular forms revealed that successful therapy with ebrotidine was accompanied by a 2.6-2.9-fold increase in the high molecular weight mucin form. The H. pylori aggregation inhibition assays showed that therapy with ebrotidine evoked a 4-fold increase in mucin anti-H. pylori titer. The changes in the functional properties of mucin following ebrotidine therapy were also accompanied by a 36% increase in the content of sulfomucin. The results demonstrate that ulcer therapy with ebrotidine lead to a marked enhancement in mucin's qualities associated with maintenance of gastric mucosal integrity and strengthening the indigenous defenses against H. pylori.

1. A receptor for mucin was isolated from the solubilized gastric epithelial cell membrane by affinity chromatography on Sepharose-bound wheat germ agglutinin. 2. The receptor protein displayed a molecular weight of 97 kDa and exhibited specific affinity towards mucin-coated surfaces. The optimum for mucin binding occurred at 60-100 micrograms/ml, while the values for the receptor were 2.0-3.1 micrograms/ml. 3. The mucin binding to the receptor was susceptible to Helicobacter pylori lipopolysaccharide which caused maximum inhibition of 91% at 30 mu/ml. This inhibitory effect of the lipopolysaccharide was abolished by a gastroprotective agent, sulglycotide. 4. The effect of sulglycotide was dose dependent and at 50 micrograms/ml produced a 94% restoration in receptor-mucin binding. Furthermore, sulglycotide was also capable of enhancing (97%) the mucin binding to its receptor in the absence of the lipopolysaccharide. 5. The results demonstrate that H. pylori through its lipopolysaccharide interferes in the interaction of mucin with gastric epithelial surfaces and that a gastroprotective agent, sulglycotide, counteracts this effect, and hence is capable of preventing the loss of mucin coat continuity occurring with H. pylori infection.

1. Rat gastric mucosal cells, subjected to phospholipid labeling by incubating the cell suspension in DMEM with [3H]choline, were exposed to different concentrations (0-150 microM) of H2-receptor antagonists, ebrotidine and ranitidine, and the phospholipid secretory responses were evaluated. 2. In the absence of the drugs, the secretion of choline-containing phospholipids over a 1 hr period averaged 3.97% of the total cellular labeled phospholipids. Ebrotidine caused a dose-dependent increase in the rate of phospholipid secretion which was most pronounced at 1 hr and persisted for at least 2 hr. The maximal effect was attained at 120 microM ebrotidine giving a 36% increase in phospholipid secretion. 3. The phospholipid secretory response to ebrotidine was accompanied by an increase in gastric mucosal cell cAMP level which reached a maximum value of 2.1-fold over that of controls at 1 hr. Ranitidine, in contrast, neither evoked increase in cAMP level nor caused any stimulation in phospholipid secretion. 4. The results indicate that the gastroprotective properties of ebrotidine are associated with the ability of the drug to elicit a rapid stimulation in gastric mucus phospholipid secretion, and that ranitidine does not possess such property.

1. Gastric mucus glycoproteins are actively involved in the maintenance of mucosal integrity and the impairment in their elaboration is often a prominent feature in gastric pathology. In this study, we investigated the effect of an antiulcer drug, sofalcone, on the activity of tyrosylprotein sulfotransferase enzyme involved in the secretion of proteins and glycoproteins in male 8 week old rats. 2. Using poly-Glu6, Ala3, Tyr1 (EAY) as sulfate acceptor, and 3'phosphoadenosine 5'-phosphosulfate (PAPS) as sulfate donor the optimum Golgi TPST activity was obtained at pH 6.8, in presence of 0.5% Triton X-100, 20 mM MnCl2, 50 mM NaF and 2 mM 5'-AMP. 3. Introduction of sofalcone to the reaction mixture led to the enhancement in TPST activity. The rate of stimulation was proportional to the drug concentration up to 30 micrograms, at which concentration, a 55% increase in TPST activity was attained. 4. The results attest further to the value of sofalcone as a potent mucosal strengthening agent, and suggest that the agent may promote mucin secretion via activation of tyrosine sulfation.

Objectives: Epidermal growth factor (EGF) and platelet-derived growth factor (PDGF) play important roles in the process of mucosal repair and restitution, and their biological effects are mediated by receptors located on the target cell surfaces. The purpose of this study was to assess the effect of the antiulcer agent, ebrotidine, on the expression of EGF and PDGF receptors with chronic ulcer healing., Methods: Chronic gastric ulcers were developed in the rat by acetic acid technique. The animal were divided into two groups and were treated twice daily for 14 consecutive days, either with ebrotidine at 100 mg/kg, or placebo. At different stages of treatment, the animals were sacrificed and used for the isolation and quantification of gastric mucosal EGF and PDGF receptors., Results: The binding assays revealed that ulcer healing was accompanied by an increase in mucosal expression of both types of receptors. A 1.7-1.8-fold increase in PDGF and EGF receptors occurred by the 4th day after the development of ulcer and reached a maximum of 3-fold increase by the 14th day, when the ulcer was essentially healed. Treatment with ebrotidine caused accelerated ulcer healing (7 days) which was accompanied by a significant enhancement in receptor expression. Compared to the controls, a 1.5-fold increase in EGF and 1.7-fold increase in PDGF receptor expression occurred by the 7th day of ebrotidine treatment, and a 1.4- to 1.5-fold increase was still observed at the 14th day of treatment., Conclusions: The results suggest that ebrotidine is capable of enhancement of gastric mucosal proliferative activities associated with ulcer healing through the stimulation of EGF and PDGF receptor expression.

The effect of intragastric administration of sulglycotide, a cytoprotective sulfated glycopeptide, on the expression of gastric mucosal epidermal growth factor receptor was investigated. The experiments were conducted with groups of rats, one receiving twice daily for 5 consecutive days a dose of 200mg/kg sulglycotide, and the other only vehicle. Mucosal cell membranes were isolated from the stomachs at 16, 40 and 88h after the last dose, and used for EGF receptor assays. The binding assays revealed a marked increase in mucosal EGF receptor expression with sulglycotide. Compared to the controls, the sulglycotide-treated group showed a 4-fold increase in the EGF receptor expression at 16h after the last dose of sulglycotide, a 4.7-fold increase in the EGF receptor was observed by the 40h, and a 4.2-fold increase was still evident at 88h following the treatment. The results demonstrate that sulglycotide exhibits remarkable ability to enhance the gastric mucosal expression of EGF receptor.

1. The effect of antiulcer agent, ebrotidine, on the expression of gastric mucosal laminin receptor during ulcer healing was investigated. 2. Rats with acetic acid-induced chronic gastric ulcers were treated twice daily for 14 consecutive days either with ebrotidine at 100 mg/kg or placebo, and then at different stages of treatment used for the isolation and quantitation of gastric mucosal laminin receptor. 3. The binding assays revealed that the ulcer healing was accompanied by an increase in mucosal expression of laminin receptor. A 2.7-fold increase in the receptor expression occurred by 4th day following the development of ulcer and reached a maximum of 8.6-fold increase by the 14th day when the ulcer was essentially healed. 4. Treatment with ebrotidine caused accelerated ulcer healing (7 days), which was accompanied by a remarkable enhancement in the laminin receptor expression. A 2.5-fold increase in the receptor expression over that of controls occurred by the 4th day of ebrotidine treatment, and a 1.7-fold increase was still observed at the 14th day of treatment. 5. The results suggest that ebrotidine, by evoking enhanced mucosal cell laminin receptor expression, promotes reepithelization and, hence, hastens the ulcer healing.

Ebrotidine is a new H2-receptor antagonist also known for its gastroprotective effect against ethanol-induced mucosal injury. In this study, we investigated the effect of ebrotidine on the activity of the gastric mucosal calcium channels. The channel complex was isolated from the solubilized gastric epithelial cell membranes by affinity chromatography on wheat germ agglutinin. The complex following labeling with [3H] PN200-110 was reconstituted into phosphatidylcholine vesicles which exhibited active 45Ca2+ uptake into intravesicular space and responded in a concentration-dependent manner to calcium channel activator, BAY K8644, as well as to calcium channel antagonist, PN200-100. The 45Ca2+ uptake was inhibited by ebrotidine which caused maximum inhibitory effect of 54.9% at 50 micrograms/ml. The gastric mucosal calcium channels on epidermal growth factor binding (EGF) in the presence of ATP responded by an increase in tyrosine phosphorylation of 55 and 170 kDa proteins, and the vesicles containing the phosphorylated channels displayed a 48% greater 45Ca2+ uptake. This phosphorylation process was inhibited by ebrotidine which also interfered with the binding of EGF to calcium channel protein. The results point towards the importance of EGF in the maintenance of gastric mucosal calcium homeostasis, and suggest that ebrotidine has the ability to protect the cellular integrity from calcium imbalance by modulating the EGF-stimulated gastic mucosal calcium channel phosphorylation.

Objectives: Infection with Helicobacter pylori is now recognized as a major factor in the etiology of gastric disease, and among the detrimental effects this bacterium exerts on the mucosal integrity is the elaboration of extracellular protease and lipase enzymes capable of mucus protein and lipids degradation. We present here evidence that the activities of these enzymes are inhibited by an gastroprotective agent, sulglycotide., Methods: The grown colonies of bacterium were washed with saline, filtered through sterilization filter, and the filtrate used as the enzyme source., Results: In the absence of sulglycotide, the H. pylori protease caused extensive degradation of human gastric mucus, while free fatty acids, glycerol monooleate and lysophosphatidylcholine were produced by the action of H. pylori lipase and phospholipase A enzymes. Introduction of sulglycotide to the incubation systems led to the reduction in the rate of mucus protein and lipid degradation. The rate of proteolysis inhibition was proportional to sulglycotide concentration up to 45 micrograms/ml, at which point a 43% reduction in mucus degradation was attained, whereas the maximum inhibition of lipase (39%) and phospholipase A (98%) activities occurred at a sulglycotide concentration of 100 micrograms/ml., Conclusions: This study indicates that sulglycotide is capable of counteracting the mucolytic activity of H. pylori, and thus may be of value in the therapy of H. pylori-associated gastric diseases.

1. The effect of cell-wall lipopolysaccharide from Helicobacter pylori, a bacterium implicated in the etiology of gastric disease, on the gastric mucosal laminin-receptor was investigated. 2. The receptor, isolated from gastric epithelial cell membrane by affinity chromatography on laminin-coupled Sepharose, was radioiodinated and incorporated into liposomes which exhibited specific affinity towards laminin-coated surface. 3. The binding of liposomal receptor to laminin-coated surface was inhibited by H. pylori lipopolysaccharide, which at 50 micrograms/ml caused a nearly complete (97%) inhibition in binding. 4. The inhibitory effect of the lipopolysaccharide was prevented by a cytoprotective agent, sulglycotide, that evoked a 92% restoration in binding at 40 micrograms/ml. 5. The results demonstrate that through its lipopolysaccharide H. pylori is capable of disrupting the gastric mucosal integrity and that this detrimental effect could be successfully countered by sulglycotide.

1. A glycosulfatase activity towards human gastric sulfomucin was identified in the extracellular material elaborated by Helicobacter pylori, a pathogen implicated in the etiology of gastric disease. 2. The purified enzyme displayed an apparent molecular weight of 30 kDa, and exhibited maximum activity at pH 5.7 in the presence of 0.3% Triton X-100 and 100 mM CaCl2. 3. The H. pylori glycosulfatase activity towards human gastric sulfomucin was inhibited by a gastroprotective agent, sulglycotide. The inhibitory effect was proportional to the concentration of sulglycotide up to 20 micrograms/ml, at which a 98% decrease in mucin desulfation occurred. However, the drug lost the inhibitory effect following its chemical desulfation. 4. The results demonstrate that sulglycotide is a potent inhibitor of H. pylori glycosulfatase and, hence, may be of value in the treatment of gastric disease associated with this bacterial infection.

1. The effect of a new antiulcer agent, ebrotidine, on the synthesis and secretion of sulfomucin in gastric mucosa was investigated. Rat gastric mucosal segments were incubated in DMEM containing [3H]proline, [3H]glucosamine and [35S]Na2SO4 as markers for mucin synthesis, glycosylation and sulfation, in the presence of 0-150 microM ebrotidine. 2. The drug, while showing no discernible effect on the apomucin synthesis, evoked a dose-dependent increase in mucin glycosylation and sulfation, which at 100 microM ebrotidine, attained its maximum of 2.4 and 2.7-fold stimulation, respectively. 3. The analysis of mucin secretory responses revealed that ebrotidine caused a concentration-dependent enhancement in sulfomucin secretion which attained its maximum increase of 3.3-fold at 100-120 microM ebrotidine. Furthermore, the sulfomucin elaborated in the presence of ebrotidine exhibited a higher content of a large molecular-weight mucus glycoprotein form, the assembly of which is intimately associated with the sulfation event. 4. The results suggest that the ability of ebrotidine to enhance gastric sulfomucin synthesis and secretion may play an important role in the gastroprotective mechanism of action of this agent.

Sulglycotide, a potent antiulcer agent derived from duodenal mucus glycopeptide through sulfation of the carbohydrate moieties, was evaluated with respect to its ability to interfere with H. pylori mucosal attachment. H. pylori cells were incubated with sulglycotide or human gastric mucin and then examined for their inhibitory capacity of H. pylori attachment to erythrocytes. Titration data revealed that the mucin inhibitory activity was confined to its sulfomucin fraction, the titer of which was found to be 16-fold higher than that of intact mucin. The data with sulglycotide showed that the inhibitory titer of this agent against H. pylori attachment was at least 30-fold higher than that of the sulfated gastric mucin fraction. The results point towards the involvement of sulfomucins in the protection of gastric mucosa from H. pylori colonization and demonstrate that sulglycotide, because of structural similarities, is ideally suited to augment the inherent mucosal defenses against this pathogen.

A glycosulfatase activity towards human gastric sulfomucin was identified in the extracellular material elaborated by H. pylori, a pathogen implicated in the etiology of gastric disease. The purified enzyme exhibited maximum activity at pH 5.7 in the presence of 0.3% Triton X-100 and 100mM CaCl2, and displayed on SDS-PAGE an apparent molecular weight of 30kDa. The H. pylori glycosulfatase effectively caused desulfation of N-acetylglucosamine-6-sulfate and galactose-6-sulfate of the carbohydrate chains of mucins, as well as that of glucose-6-sulfate of glyceroglucolipids, but was ineffective towards galactosyl -and lactosylceramide sulfates which contain galactose-3-sulfate. The glycosulfatase activity towards human gastric sulfomucin was inhibited by an antiulcer agent, nitecapone, which at its optimal concentration (100 micrograms/ml) caused a 61% decrease in mucin desulfation. The results show that H. pylori through its glycosulfatase activity causes desulfation of sulfated mucins and glyceroglucolipids of the protective mucus layer, and that nitecapone is able to interfere with this detrimental action.

1. The effect of a new antiulcer agent, nitecapone, on the synthesis and secretion of sulfomucin in gastric mucosa was investigated using mucosal segments incubated in the presence of [3H]proline, [3H]glucosamine and [35S]sulfate. 2. The drug, while showing no discernible effect on the apomucin synthesis, evoked a dose-dependent increase in mucin glycosylation and sulfation, which at 225 microM nitecapone, attained its maximum of 1.8 and 2.2-fold stimulation, respectively. 3. The analysis of mucin secretory responses revealed that nitecapone caused a concentration-dependent enhancement in sulfomucin secretion attaining maximum increase of 1.5-fold at 150 microM nitecapone. 4. The stimulatory effect of nitecapone on sulfomucin secretion was accompanied by 1.4-fold increase in mucosal cAMP level, and showed sensitivity to protein kinase A inhibitors, thus pointing towards the involvement of protein kinase A in mediation of gastric sulfomucin secretory responses to nitecapone. 5. The ability of nitecapone to enhance sulfomucin synthesis and secretion could be of importance to the gastroprotective action of this agent.

A gastric mucosal calcium channel-epidermal growth factor (EGF) receptor complex was isolated from solubilized epithelial cell membranes by means of a wheat germ agglutinin affinity. The complex, following reconstitution into phosphatidylcholine vesicles, exhibited active 45Ca2+ uptake as evidence by concentration-dependent response to the calcium channel activator BAY K8644, and the calcium channel antagonist PN200-110. The complex on the addition of EGF and ATP showed an increase in tyrosine phosphorylation of both a 55 and a 170kDa protein, while the vesicles containing the phosphorylated complex displayed a 48% greater 45Ca2+ uptake. The phosphorylation process was inhibited by an anti-ulcer agent, ebrotidine, which also interfered with the binding of EGF to calcium channel protein. The results suggest that ebrotidine protects the cellular integrity from calcium imbalance by modulating the EGF-stimulated gastric mucosal calcium channel activation.

The effect of a new antiulcer agent, ebrotidine, on the synthesis and secretion of gastric sulfomucin was investigated using mucosal segments incubated in the presence of [3H]proline, [3H]glucosamine and [35S]Na2SO4. The drug, while showing no discernible effect on the apomucin synthesis, evoked a dose-dependent increase in mucin glycosylation and sulfation, which at 100 microM ebrotidine, attained its maximum of 2.4 and 2.7-fold stimulation, respectively. The analysis of mucin secretory responses revealed that ebrotidine caused a concentration-dependent enhancement in sulfomucin secretion which attained its maximum increase of 3.3-fold at 100-120 microM ebrotidine. The sulfomucin elaborated in the presence of ebrotidine exhibited a higher content of a large molecular-weight mucus glycoprotein form, the assembly of which is intimately associated with the sulfation event. The results suggest that the ability of ebrotidine to enhance gastric sulfomucin synthesis and secretion may play an important role in the gastroprotective mechanism of action of this agent.

1. The effect of ebrotidine, a new antiulcer agent, on the activity of mucus degrading of protease and lipase enzymes elaborated by Helicobacter pylori was investigated. 2. In the absence of ebrotidine, the H. pylori protease caused extensive degradation of gastric mucus protein, while free fatty acids, glycerol mono-oleate and lysophosphatidylcholine were produced by the action of H. pylori lipase and phospholipase A enzymes. 3. Introduction of ebrotidine to the incubation system led to the reduction in the rate of mucus protein and lipid degradation. The rate of proteolysis inhibition was proportional to ebrotidine concentration up to 35 micrograms/ml at which point, a 57% reduction in mucus degradation was obtained, while the maximum inhibition of phospholipase A (96%) and lipase (93%) activities occurred at ebrotidine concentration of 60 micrograms/ml. 4. The results indicate that ebrotidine is capable of counteracting the mucolytic activity of H. pylori towards protein and lipid constituents of gastric mucus layer.

The effect of prolonged administration of an antiulcer drug, sofalcone, on the physicochemical properties of gastric mucus was investigated. The experiments were conducted with two groups of rats in which one group received a dose of 100 mg/kg of sofalcone twice daily for three consecutive days, and the control group received daily doses of vehicle. The rats were killed 16 h after the last dose, their stomachs dissected, and the mucosa subjected to physicochemical measurements. The results revealed that sofalcone evoked a 23% increase in mucus gel thickness, and its content of sulfo- and sialomucins increased by 54 and 25%, respectively. These changes in mucus with sofalcone were accompanied by a 16% increase in H+ retardation capacity, twofold increase in viscosity, and a 39% increase in the gel hydrophobicity. The mucus elaborated in the presence of sofalcone exhibited a 10% lower content of protein, 30% higher content of carbohydrate, and 18% higher content of total lipids, which were particularly enriched (20%) in phospholipids and contained 67% more covalently bound fatty acids. Furthermore, the high molecular weight mucus glycoprotein form accounted for about 30% of gel mucin in the control group, whereas its content in mucus gel of animals receiving sofalcone reached 50%. The results indicate that sofalcone enhances the protective qualities of the mucus component of the gastric mucosal barrier and hence strengthens the gastric mucosal integrity.

The effect of prolonged administration of an antiulcer drug, sofalcone, on the physicochemical properties of gastric mucus was investigated. The experiments were conducted with groups of rats receiving twice daily for three consecutive days a dose of 100 mg/kg sofalcone, while the control group received daily doses of vehicle. The rats were sacrificed 16 h after the last dose and gastric mucosa subjected to physicochemical measurements. The results revealed that sofalcone evoked a 23% increase in mucus gel dimension, while sulfo- and sialomucins content of the gel increased by 54 and 25%, respectively. These changes were accompanied by a 16% increase in mucus H+ retardation capacity, 2-fold increase in viscosity, and a 39% increase in the gel hydrophobicity. The mucus elaborated in the presence of sofalcone contained 67% more covalently bound fatty acids, exhibited 10% lower content of protein, 30% higher content of carbohydrate, and 18% higher content of lipids. The mucus of the sofalcone group also showed an increase in the proportion of the high molecular weight mucus glycoprotein form, which in the control group accounted for about 30% of gel mucin, while its content in mucus gel of animals receiving sofalcone reached the value of 50%. The results indicate that sofalcone enhances the protective qualities of mucus component of gastric mucosal barrier.

1. A gastric mucosal laminin receptor has been isolated from the epithelial cell membrane by affinity chromatography on Sepharose-bound laminin, and following radioiodination was incorporated into liposomes which displayed specific affinity towards the laminin-coated surface. 2. The binding of liposomal receptor to the laminin-coated surface was inhibited by Helicobacter pylori lipopolysaccharide and reached a maximum value of 96% at 50 micrograms/ml. 3. The inhibitory effect of H. pylori lipopolysaccharide on the receptor-laminin binding was prevented by an antiulcer agent, nitecapone. The effect was concentration dependent and produced a maximum response of 83% at 30 micrograms/ml of drug concentration. 4. The results demonstrate that H. pylori is capable of disrupting gastric mucosal integrity by interfering with epithelial cell-laminin binding, and that an antiucler agent, nitecapone, counteracts this effect.

1. The mechanism of gastric mucosal protection by an anticular agent, nitecapone, against injury was investigated in rats with and without indomethacin pretreatment. 2. Animals received intragastrically either a dose of nitecapone or vehicle alone, followed by ethanol given at various intervals up to 5 hr, and their gastric mucosa subjected to histologic and physicochemical assessment. 3. Ethanol caused extensive gastric hemorrhagic lesions which were essentially prevented by nitecapone at doses of 30 mg and higher per kg body weight. The maximal protection was achieved by 1.5 hr which persisted up to 4 hr and was not thwarted by indomethacin. 4. Physicochemical measurements revealed that nitecapone evoked 78% increase in mucus gel dimension, and showed 21% increase in phospholipids, and the content of sulfo-(22%) and sialomucins (72%). This was accompanied by 1.6-fold increase in mucus viscosity, 31% increase in H+ retardation capacity and 2.2-fold increase in hydrophobicity. 5. The results suggest that the gastroprotective action of nitecapone occurs through the enhancement of the physicochemical characteristics of mucus layer.

The effects of intragastric administration of an antiulcer drug, colloidal bismuth subcitrate, on the content, composition and physical properties of the mucus component of gastric mucosal barrier were investigated. The experiments were conducted with two groups of rats in which one group received twice daily for three consecutive days a dose of 100 mg/kg colloidal bismuth subcitrate, while the control group received saline. The animals were killed 16 h after the last dose, their stomachs dissected and the mucosa subjected to physicochemical measurements. The results revealed that colloidal bismuth subcitrate elicited a 49% increase in mucus gel dimension, while sulfo- and sialomucin content of the gel increased by 64 and 112%, respectively. The changes in mucus with colloidal bismuth subcitrate were accompanied by a 28% increase in H+ retardation capacity, 2.2-fold increase in viscosity, and a 26% increase in the gel hydrophobicity. The mucus elaborated in the presence of colloidal bismuth subcitrate exhibited 16% lower protein content and 68% higher content of carbohydrate than that of the control, displayed similar levels of total lipids and covalently bound fatty acids, but its phospholipid content was 32% higher. Furthermore, the mucus of the colloidal bismuth subcitrate group showed a marked increase in the proportion of the high molecular weight form of mucin. The results suggest that colloidal bismuth subcitrate is capable of the enhancement of mucus gel qualities associated with the maintenance of gastric mucosal integrity.

The effect of an antiulcer drug, colloidal bismuth subcitrate (De-Nol), on the proteolytic activity of pepsin toward gastric mucus and salivary epidermal growth factor was investigated. Samples of pig gastric mucus and mouse epidermal growth factor were incubated with pepsin in the absence and in the presence of De-Nol, and the released alpha-amino acid residues were quantified. Results of analysis revealed that, in the absence of De-Nol, the apparent Km value of pepsin toward gastric mucus was 1.4 g/L and that towards epidermal growth factor 120 microM. Introduction of De-Nol to the incubation mixtures led, in both cases, to reduction of the rates of proteolysis. With gastric mucus, the rate of proteolysis inhibition was proportional to the De-Nol concentration up to 1 x 10(-2) g/L, at which point a 54% reduction in mucus proteolysis occurred, whereas, with epidermal growth factor, this concentration of De-Nol caused nearly 52% inhibition in the rate of proteolysis. The apparent Ki value for peptic degradation of gastric mucus in the presence of De-Nol was 2.1 x 10(-4) g/L and that for peptic degradation of epidermal growth factor 1.8 x 10(-2) g/L. The results suggest that among the beneficial effects of colloidal bismuth subcitrate on ulcer healing is its ability to interfere with peptic digestion of the protective gastric mucus coat and of such important bioactive protein as epidermal growth factor.

Although the mechanism of gastric mucosal protection is multicomponential, the initial brunt of luminal insult falls on the layer of mucus weakening its physicochemical characteristics and is intimately associated with gastric disease. Hence, the agents capable of strengthening the qualities of mucus gel, associated with its protective function, are becoming increasingly popular in peptic ulcer therapy. Evaluation of the efficacy of these types of drugs requires delineation of their direct effect on the mucus layer constituents, from those evoked in the chemical composition of mucus as a result of prolonged drug administration. Studies with mucus layer strengthening agents indicate that such drugs as sucralfate and De-Nol interact tenaciously with the mucus gel and in essence seal the underlying mucosal surface from the noxious luminal contents. These drugs and other new antiulcer agents like sofalcone, geranylgeranylacetone and stable prostaglandin analogs are also capable of enhancing mucus gel viscosity and its hydrogen ion impedance ability. Furthermore, the majority of these drugs exert an inhibitory effect on peptic degradation of gastric mucus, and the colloidal bismuth preparation (De-Nol) is also capable of prolongation of the luminal availability of epidermal growth factor. The data on gastric mucus gel, following prolonged administration, show that these drugs enhance the lipid content of the mucus gel, and increase both the mucus coat dimension and the proportion of the high molecular weight form of its mucin component. These changes result in the improvement of viscoelastic, permselective and hydrophobic properties of gastric mucus. Hence, continuity of the function of mucus layer perimeter of mucosal defense is maintained.

There is increased awareness that infection with Campylobacter pylori could be a major factor in the pathogenesis of gastric disease. Here, we present evidence that the extracellular protease elaborated by this bacteria, which causes degradation of gastric mucus, is inhibited by an antiulcer agent, colloidal bismuth subcitrate (CBS; De-Nol). The study was conducted with C. pylori cultured from antral mucosal biopsy specimens of patients undergoing gastroscopy. The grown colonies of bacteria were washed with saline, filtered through sterilization filter, dialyzed, and lyophilized. The powder was used as the enzyme source for proteolytic activity assay employing pig gastric mucus as substrate. Optimum enzymatic activity was obtained at 37 degrees C and at pH 7.0. The apparent Km of C. pylori protease with gastric mucus was 0.71 g/L. Analyses of the degradation products indicated that the protease caused extensive proteolysis of mucus glycoprotein polymer. Introduction of CBS to the incubation mixtures led to a reduction of the rate of mucus degradation. The rate of proteolysis inhibition was proportional to CBS concentration up to 1 X 10(-1) g/L, at which point a 37% reduction in mucus proteolysis was obtained. The Km value for proteolytic degradation of mucus by C. pylori protease in the presence of CBS was 1.25 g/L. The results suggest that CBS is capable of counter-acting the proteolysis of the protective gastric mucus layer by C. pylori.

The effects of antiulcer drugs, sofalcone and sucralfate, on the activity of gastric mucosal mucus glycoprotein fatty acyltransferase were investigated. The acyltransferase enzyme, contained in the detergent extracts of the microsomal fraction of rat gastric mucosa, was incubated with the deacylated gastric mucin and palmitoyl-CoA substrates in the presence and absence of drugs, and the formed fatty acid acylated glycoprotein product was quantitated. In the absence of drugs, the enzymatic activity increased proportionally with increased concentrations of both substrates and of enzyme, and gave an apparent Km value of 5.6 X 10(-7) M. Introduction of sofalcone to the reaction mixtures led to an enhancement in the rate of mucus glycoprotein acylation. The rate of enhancement was proportional to sofalcone concentration up to 1.0 X 10(-5) M, with an apparent Km value of 3.7 X 10(-7) M. In contrast to sofalcone, the acyltransferase activity was inhibited by sucralfate. The rate of inhibition of mucus glycoprotein acylation by sucralfate was of the competitive type and at 1.0 X 10(-4) M reached a value of 25%. The apparent KI value calculated from the double-reciprocal plots for sucralfate was 9.1 X 10(-7) M. As the acylation of mucin with fatty acids plays an important role in the maintenance of gastric mucosal integrity, the results suggest that stimulation of the fatty acyltransferase enzyme by sofalcone may be one of the beneficial effects of this drug towards ulcer healing.

The effect of sucralfate on the viscosity of pig gastric mucus and on its ability to retard the diffusion of hydrogen ion was investigated. Using a cone/plate viscometer at shear rates between 1.15 and 230 s-1, it was found that preincubation of mucus with increasing concentrations of sucralfate led to a gradual enhancement of the mucus viscosity. This enhancement in viscosity was proportional to the sucralfate concentration up to 1.0 X 10(-4) M and increased about 18% for each 10-fold increment in its concentration. The permeability measurements, conducted in a specially designed two compartment chamber, revealed that addition of sucralfate to gastric mucus had a profound beneficiary effect on its ability to retard the diffusion of hydrogen ion. In the presence of 1.0 X 10(-6) M sucralfate the permeability of mucus to hydrogen ion decreased by 35%, while the 1.0 X 10(-3) M sucralfate reduced the mucus permeability by 68%. The results show that sucralfate increases the viscosity of gastric mucus and improves its ability to impede the hydrogen ion penetration.