Suppression of Helicobacter pylori-induced interleukin-8 production in gastric cancer cell lines by an anti-ulcer drug,geranylgeranylacetone

BACKGROUND: Geranylgeranylacetone (GGA) is an antigastritis and anti-ulcer agent, with as yet an unknown mechanism of action. In this study, we investigated the effect of GGA on Helicobacter pylori-induced interleukin (IL)-8 production and IL-8 mRNA expression in KATOIII cells, an established gastric cell line. METHODS: Interleukin-8 production in H. pylori-infected KATOIII cells was measured by using enzyme-linked immunoassay. The cytotoxicity of H. pylori on KATOIII cells was measured by a 51Cr release assay. The effect of GGA on H. pylori-induced IL-8 mRNA expression was measured by using northern blotting. RESULTS: Interleukin-8 production increased with time and H. pylori dose; the most significant increase was seen within 6-24 h of coculture with H. pylori. A dose of 0.1 mmol GGA suppressed IL-8 production (P = 0.0077) and inhibited H. pylori-induced IL-8 mRNA expression (P = 0.0019). Furthermore, H. pylori-induced gastric mucosal cell injury associated with IL-8 and neutrophil activation was enhanced by NH3, and this enhancement was suppressed by GGA (P = 0.0043). CONCLUSIONS: Helicobacter pylori-infected gastric mucosal cells produce IL-8, which can promote neutrophil activation, thus contributing to mucosal tissue injury associated with H. pylori infection. Agents like GGA, which can suppress IL-8 production may have a protective role in the treatment of mucosal tissue damage seen in H. pylori infection.     

Inhibitory effect of polaprezinc on the inflammatory response to Helicobacter pylori.

Helicobacter pylori-infected gastrointestinal mucosa is frequently infiltrated by polymorphonuclear leukocytes (PMN) and monocytes, and these invading cells have been implicated in gastrointestinal mucosal inflammation. To clarify the efficacy of polaprezinc, a chelate compound consisting of zinc and L-carnosine, against H pylori-induced inflammation including PMN infiltration, the in vitro effects of this drug on interleukin (IL)-8 production by an established gastric cancer cell line (MKN 45 cells) and on PMN-endothelial cell adhesive interactions was investigated. Polaprezinc and zinc sulphate inhibited IL-8 production by MKN 45 cells in response to stimulation with H pylori water extract (HPE) in a dose-dependent manner from 10(-7) M to 10(-5) M. In addition, the expression of CD11b and CD18 on PMN and PMN-dependent adhesion to endothelial cells elicited by HPE was inhibited by polaprezinc and zinc sulphate in a concentration-dependent manner. L-carnosine did not have any effects on IL-8 production or PMN-endothelial cell interactions. These results suggest that polaprezinc, mainly the zinc component, may inhibit H pylori-induced PMN-mediated gastric inflammation by attenuating CD11b/CD18 expression on PMN and IL-8 production from gastric epithelial cells.     

Helicobacter pylori infection: mechanism of colonization and functional dyspepsia Reduced colonization of gastric mucosa by Helicobacter pylori in mice deficient in interleukin-10

BACKGROUND AND AIMS: Interleukin-10 (IL-10) is a potent anti-inflammatory and immunoregulatory cytokine. Mice deficient in IL-10 production (IL-10-/-mice) develop a spontaneous chronic enterocolitis, suggesting that IL-10 is an important regulator of the mucosal immune response in vivo. The objective of this study was to determine the role of endogenous IL-10 in the host defense against gastric colonization by Helicobacter pylori by using IL-10-deficient mice. METHODS: The IL-10-/-mice were inoculated intragastrically with a mouse-adapted H. pylori isolate (Sydney Strain 1). Gastric colonization by H. pylori (biopsy urease test and bacterial colony counts), serum levels of H. pylori-specific immunoglobulin (Ig) M, A, G, isotypes of IgG, and the gastric mucosal inflammatory scores were determined 6 weeks after inoculation. Results were compared with those obtained from H. pylori-infected control mice (IL-10+/-mice). RESULTS: The colonization of gastric mucosa by H. pylori was reduced approximately 100-fold (P < 0.0001) in IL-10-/-mice (log10 4.87 +/- 0.26CFU/g tissue) as compared to IL-10+/-mice (log10 6.64 +/- 0.22 CFU/g tissue). Furthermore, IL-10-/-mice infected with H. pylori had significantly higher H. pylori-specific IgA and IgG antibodies in serum (P < or = 0.01), and developed much more severe chronic active gastritis than infected IL-10+/-mice. The median scores of the infiltration of gastric mucosa by mononuclear cells and neutrophils were up to threefold higher in IL-10-/-mice than they were in IL-10+/-mice. CONCLUSION: Our studies suggest that endogenous IL-10 is an inhibitor of the protective immune response to H. pylori infection. Interleukin-10 participates in the downregulation of H. pylori-induced gastric inflammatory responses, which apparently confers a survival advantage to the organism promoting more effective colonization of gastric mucosa.     

Gastric mucosal cytokine levels in relation to host interleukin-1 polymorphisms and Helicobacter pyloricagA genotype

OBJECTIVE: The outcome of a Helicobacter pylori infection is related in part to interrelationships among H. pylori virulence factors and the H. pylori-induced mucosal response. The host inflammatory response is partly governed by polymorphisms in pro-inflammatory genes. MATERIAL AND METHODS: Cytokine levels (interleukin (IL)-1beta, IL-6 and IL-8) were examined in H. pylori-infected and uninfected normal-appearing mucosa from patients with non-ulcer dyspepsia (NUD), margins of gastric ulcers and cancer tissues. Cytokine levels were compared with cagA genotypes and host interleukin (IL)-1 polymorphisms. RESULTS: The study comprised 168 Thai patients. All infected patients possessed anti-CagA antibody. Gastric mucosal IL-8 levels were significantly higher in H.pylori-positive cases than in -negative cases in all three tissue types (e.g. 1115 versus 217 pg/mg protein for NUD) (p<0.001). Normal-appearing but H. pylori-infected antral mucosa of patients with cagA type 1a strains had higher IL-8 levels than those with type 2a strains (2632 versus 1036 pg/mg protein) (p<0.005). IL-1B-511T/T carriers had higher antral mucosal IL-1ss levels versus non-carriers (pg/mg protein) (T/T=221, T/C=178, C/C=70) (p=0.005). IL-1B-511T/T carriers also had higher IL-1beta levels versus non-carriers in H. pylori-negative patients. CONCLUSIONS: It was found that both the host factors (IL-1 polymorphisms) and bacterial factors (cagA type 1a versus type 2a) influenced gastric mucosal cytokine levels. Future studies should concentrate on interactions among host factors (e.g. genetics and tissue responses) and bacterial and environmental factors.     

Expression of cytokeratins in Helicobacter pylori -associated chronic gastritis of adult patients infected with cagA ＋ strains： An immunohistochemical study

AIM:To investigate the expression of differentcytokeratins(CKs)in gastric epithelium of adult patientswith chronic gastritis infected with Helicobacter pylori(Hpylori)cagA strains.METHODS:The expression of CK 7,8,18,19 and 20was studied immunohistochemically in antral gastricbiopsies of 84 patients.All the CKs were immunostainedin cagA H pylori gastritis(57 cases),non-H pylori gastritis(17 cases)and normal gastric mucosa(10 cases).RESULTS:In cagA H pylori gastritis,CK8 wasexpressed comparably to the normal antral mucosafrom surface epithelium to deep glands.Distributionof CK18 and CK 19 was unchanged,i.e.transmucosal,but intensity of the expression was different in foveolarregion in comparison to normal gastric mucosa.Cytokeratin 18 immunoreactivity was significantly higherin the foveolar epithelium of H pylori-positive gastritiscompared to both Hpylori-negative gastritis and controls.On the contrary,decrease in CK19 immunoreactivityoccurred in foveolar epithelium of H pylori-positive gastritis.In both normal and inflamed antral mucosawithout Hpylori infection,CK20 was expressed strongly/moderately and homogenously in surface epithelium andupper foveolar region,but in H pylori-induced gastritissignificant decrease of expression in foveolar regionwas noted.Generally,in both normal antral mucosa andH pylori-negative gastritis,expression of CK7 was notobserved,while in about half cagA H pylori-infectedpatients,moderate focal CK7 immunoreactivity of theneck and coiled gland areas was registered,especially inareas with more severe inflammatory infiltrate.CONCLUSION:Alterations in expression of CK 7,18,19 and 20 together with normal expression of CK8 occurin antral mucosa of H pylori-associated chronic gastritisin adult patients infected with cagA strains.Alterationsin different cytokeratins expression might contribute toweakening of epithelial tight junctions observed in Hpylori-infected gastric mucosa.     

Lafutidine inhibits Helicobacter pylori-induced interleukin-8 production in human gastric epithelial cells

BACKGROUND AND AIM: Attachment of Helicobacter pylori to gastric epithelial cells leads to the production of chemokines, such as interleukin-8 (IL-8), which in turn activate and recruit neutrophils to the site of infection. Lafutidine [(+/-)-2-(furfurylsulfinyl)-N-(4-(4-(piperidinomethyl)-2-pyridyl)oxy-(Z)-2-butenyl)acetamide] is a new type of antiulcer drug that possesses an antisecretory action as well as gastroprotective activity, independent of its antisecretory action. In the present study, we examined the effects of lafutidine on H. pylori-induced IL-8 release and H. pylori adhesion to MKN45 cells. METHODS: MKN45 cells were stimulated with H. pylori, tumor necrosis factor (TNF)-alpha, or IL-1beta, then IL-6 and IL-8 levels in the culture supernatants were determined with a specific enzyme-linked immunosorbent assay kit. RESULTS: Lafutidine significantly inhibited both the release of IL-8 induced by H. pylori and the adhesion of H. pylori to cells in a dose-dependent manner. These properties of lafutidine are unrelated to the blockade of histamine H(2)-receptors, because the same effects have not been observed with other H(2)-receptor antagonists, such as cimetidine and famotidine. Lafutidine also significantly inhibited H. pylori-induced IL-6 release. Both TNF-alpha and IL-1beta-induced IL-8 releases, conversely, were little affected by lafutidine up to a concentration of 10(-5) M. CONCLUSIONS: These results suggest that lafutidine inhibits IL-8 release by inhibiting H. pylori adherence to gastric epithelial cells, indicating a novel mechanism by which lafutidine protects against the mucosal inflammation associated with H. pylori infection.     

High concentrations of human β-defensin 2 in gastric juice of patients with Helicobacter pyloriinfection

AIM: Human beta-defensin (HBD)-1 and HBD-2 are endogenous antimicrobial peptides. Unlike HBD-1, the HBD-2 expression is augmented by Helicobacter pylori (H pylori). We sought to determine HBD-1 and HBD-2 concentrations in gastric juice during H pylori infection. METHODS: HBD-1 and HBD-2 concentrations were measured by radioimmunoassay in plasma and gastric juice of 49 H pylori-infected and 33 uninfected subjects and before and after anti-H pylori treatment in 13 patients with H pylori-associated gastritis. Interleukin (IL)-1beta and IL-8 concentrations in gastric juice were measured by enzyme-linked immunosorbent assay (ELISA). Histological grades of gastritis were determined using two biopsy specimens taken from the antrum and corpus. Reverse phase high performance liquid chromatography (RP-HPLC) was used to identify HBD-2. RESULTS: HBD-2 concentrations in gastric juice, but not in plasma, were significantly higher in H pylori-positive than -negative subjects, albeit the post-treatment levels were unchanged. Immunoreactivity for HBD-2 was exclusively identified in H pylori-infected mucosa by RP-HPLC. HBD-2 concentrations in gastric juice correlated with histological degree of neutrophil and mononuclear cell infiltration in the corpus. IL-1beta levels correlated with those of IL-8, but not HBD-2. Plasma and gastric juice HBD-1 concentrations were similar in H pylori-infected and uninfected subjects. CONCLUSION: Our results place the beta-defensins, especially HBD-2, in the front line of innate immune defence. Moreover, HBD-2 may be involved in the pathogenesis of H pylori-associated gastritis, possibly through its function as immune and inflammatory mediator.     

Anti-Helicobacter pylori therapy significantly reduces Helicobacter pylori -induced gastric mucosal damage in Mongolian gerbils

AIM: To investigate the effectiveness of 4 d‘anti-Helicobacter pyloritherapy on the H pylori-infected Mongolian gerbils based on physiological and pathological changes. METHODS: We used 6-wk-old male gerbils orally inoculated with H pylori (ATCC43504, 2&#215;10^8 CFU/mL). Seven weeks alter Hpyloriinoculation, the animals of study group received 4 d‘arti-H pyloritriple therapy (Hpylorieradicated group). Seven days later, all animals of the H pylori-eradicated and control groups (H pylori-infected &amp; H pylori-uninfected groups) were sacrificed. We examined gastric mucosal lesions macroscopically, studied gastritis microscopically and determined the stomach weight ratio, myeloperoxidase (MPO) activity and prostaglandin (PG) E2 level. RESULTS: The results showed that both macroscopic and histological gastric damages were significantly less in H pylori-eradicated group than H pylori-infected group. Stomach weight ratio, MPO activity and PGE2 levels were significantly higher in H pylori-infected group than those in the other two groups. CONCLUSION: Four days‘anti-H pylori therapy was effective in the improvement of H pylori-induced gastric lesions in Mongolian gerbils.     

Effect of acid-suppressive therapy on Helicobacter pylori production of interleukin-8 in the gastric mucosa.

BACKGROUND: Recent studies have shown that acid-suppressive therapy increases the severity of Helicobacter pylori- associated gastritis in the corpus. PURPOSE: To evaluate interleukin (IL)-8 production in the gastric corpus mucosa before and during acid-suppressive therapy in H pylori-infected patients. PATIENTS AND METHODS: Ten patients with reflux esophagitis (five H pylori-positive and five H pylori-negative) were treated with omeprazole 20 mg. Serum gastrin concentrations, H pylori colonization density and mucosal IL-8 levels in the corpus were investigated at entry and two weeks after starting therapy. IL-8 levels were measured by ELISA. The organism density was determined, and scored according to area occupied by the bacterial colonies. The presence of H pylori was assessed by rapid urease testing and histological finding of gastric biopsy specimens. RESULTS: In H pylori-positive patients, concentrations of IL-8 during therapy significantly exceeded those before therapy (36.2+/-6. 8 versus 18.3+/-3.8 pg/mg protein; P<0.05) without altering H pylori density. In H pylori-negative patients, IL-8 levels were similar before and during therapy (6.1+/-2.7 versus 6.3+/-3.0 pg/mg protein). Concentrations of gastrin during therapy were significantly higher than those before therapy in all patients. CONCLUSIONS: The results suggest that acid suppression increases mucosal IL-8 levels in H pylori-infected patients with reflux esophagitis.     

Elevated concentrations of alpha-defensins in gastric juice of patients with Helicobacter pylori infection

OBJECTIVE: Defensins (alpha- and beta-defensins) are endogenous antimicrobial peptides. Little is known about alpha-defensins during Helicobacter pylori infection. METHODS: The concentrations of human neutrophil peptides (HNP-1, -2, and -3), the major components of neutrophils-derived alpha-defensins, were measured by radioimmunoassay (RIA) in plasma and gastric juice of 61 H. pylori-infected and 33 uninfected subjects, and before and after anti-H. pylori treatment in 12 patients with H. pylori-associated gastritis. Interleukin (IL)-8 concentrations in gastric juice were measured by enzyme-linked immunosorbent assay. Histological grades of gastritis and neutrophil counts (/mm(2)) infiltrating in the gastric mucosa were determined using two biopsy specimens taken from the antrum and corpus. Immunohistochemistry and reverse-phase high performance liquid chromatography (RP-HPLC) were used to identify HNPs 1-3. RESULTS: HNP 1-3 concentrations in gastric juice were significantly higher in H. pylori-positive than in H. pylori-negative patients and significantly decreased after cure. HNP 1-3 concentrations in gastric juice correlated with IL-8 levels and neutrophil densities in the gastric mucosa and were associated with histological degree of gastritis, especially the grades of activity. Intense immunoreactivity for anti-HNPs 1-3 antiserum was noted in infiltrating neutrophils in H. pylori-infected mucosa. In RP-HPLC analysis, all of the HNP 1-3 molecules were identified as their mature forms. Plasma HNP 1-3 concentrations were similar in H. pylori-infected and non-infected groups and showed no correlations with other parameters. CONCLUSIONS: We demonstrated significantly elevated levels of HNPs 1-3 in gastric juice during H. pylori infection. The elevation of HNPs is presumably secondary to H.pylori-associated gastric inflammation involving neutrophil infiltration.     

Diminished interferon-gamma production in gastric mucosa T lymphocytes after H. pylori eradication in duodenal ulcer patients.

BACKGROUND/AIMS: Helicobacter pylori (H. pylori) infects an estimated 50% of the world population; however, only a small proportion of individuals develop clinical symptoms of gastritis, peptic ulceration or gastric cancer. The variations in disease presentation may be due to differences in bacterial virulence and/or immune response to the pathogen. In a previous study we reported an increased expression of the IL-2 receptor in duodenal ulcer (DU) patients. The present study examines the expression of IL-2 receptor and intracellular lymphokine production in gastric mucosa infiltrating T lymphocytes in DU patients before and after H. pylori eradication. METHODOLOGY: T lymphocytes were isolated from gastric mucosa biopsies by using mechanical and enzymatic tissue desegregation. Ficoll-purified lymphocytes were incubated with monoclonal antibodies and analyzed by using 4-color flow cytometry analysis for the IL-2 receptor (CD25) and intracellular interferon-gamma (IFN-gamma) and IL-4 expression. Lymphocytes from 24 H. pylori-infected patients with severe gastric mucosa infiltration (G2 and G3 histological type in Sydney classification) were analyzed. RESULTS: We demonstrated a significant decrease in IL-2 receptor expression on gastric mucosa T cells 3 and 12 months after eradication of H. pylori. We also demonstrated a diminished IFN-gamma production 3 and 12 months after H. pylori eradication. CONCLUSIONS: Our results suggest that cellular immune activation in gastric mucosa is reversibly dependent on the presence of H. pylori.     

Impaired T-cell regulation of B-cell growth in Helicobacter pylori--related gastric low-grade MALT lymphoma.

BACKGROUND & AIMS: Neoplastic B cells of the Helicobacter pylori-related low-grade gastric mucosa-associated lymphoid tissue (MALT) lymphoma are responsive to T helper cells and sensitive to withdrawal of H. pylori-induced T-cell help. METHODS: The clonal progeny of T cells from the gastric mucosa of 5 patients with MALT lymphoma was compared with that of T-cell clones obtained from 5 H. pylori-infected patients with chronic gastritis. RESULTS: T-cell clones were assessed for specificity to H. pylori, cytokine profile, help for B-cell proliferation, and perforin- or Fas-mediated cytotoxic regulation of B-cell growth. Twenty-eight of 165 CD4(+) gastric clones from MALT lymphoma and 33 of 178 CD4(+) clones from chronic gastritis recognized H. pylori antigens. Cytokine production was similar in the 2 series of clones. All MALT lymphoma-derived clones dose-dependently increased their B-cell help, whereas clones from chronic gastritis lost helper activity at T-to-B-cell ratios greater than 1 because of concomitant cytolytic killing of B cells. T-cell clones from MALT lymphoma had both reduced perforin-mediated cytotoxicity and poor ability to induce Fas-mediated apoptosis. These defects were limited to gastric T cells. CONCLUSIONS: H. pylori-induced T cell-dependent B-cell activation and deficient cytotoxic control of B-cell growth may link H. pylori infection, local T-cell response, and genesis of low-grade gastric MALT lymphoma.     

Cultured human gastric explants: a model for studies of bacteria-host interaction during conditions of experimental Helicobacter pylori infection

The unique environment of the human stomach makes it difficult to establish representative in vitro models for Helicobacter pylori that mimic the natural infection. The in vitro explant culture (IVEC) technique is based on coculture of human gastric explants with H. pylori, where bacteria-host interaction is studied on the basis of interleukin (IL)-8 secretion of the explant tissue in response to infection. In this study, it was shown that H. pylori attachment to gastric epithelial tissue was required for induction of representative inflammatory responses, assessed here by IL-8 production. Furthermore, IL-8 production by the explant tissue in response to H. pylori infection demonstrated that the explants were adequately responsive. The IVEC technique for studies of the interplay between H. pylori and the human gastric mucosa during conditions of experimental infections in vitro could add knowledge to our understanding of the complex bacteria-host cross-talk in vivo.