Effect of an aluminum hydroxide-magnesium hydroxide combination drug on adhesion, IL-8 inducibility, and expression of HSP60 by Helicobacter pylori.

BACKGROUND: Co-magaldrox (Maalox) is used world-wide as an antacid and as a cytoprotective agent for gastritis and peptic ulcer diseases. We examined the effects of co-magaldrox on Helicobacter pylori. METHODS: Adhesion of H. pylori to human gastric epithelial cells (MKN45) was evaluated by flow cytometry. Morphologic changes in H. pylori caused by co-magaldrox were determined by scanning electron microscopy. Induction of interleukin-8 (IL-8) from MKN45 cells was examined with enzyme-linked immunosorbent assay, and the intracellular and extracellular expression of heat-shock protein 60 (HSP60) was analyzed with sodium dodecyl sulphate-polyacrylamide gel electrophoresis and flow cytometry. RESULTS: Adhesion of H. pylori to MKN 45 cells was significantly inhibited by 1.25%-5% comagaldrox. H. pylori aggregated with co-magaldrox according to an electron microscopic examination. IL-8 secretion from MKN45 cells after H. pylori infection was also inhibited by co-magaldrox. Extracellular expression of HSP60 on the surface of H. pylori was decreased after treatment with comagaldrox, whereas the intracellular synthesis of HSP60 was not. HSP60-induced IL-8 secretion was significantly inhibited by co-magaldrox in a dose-dependent manner. CONCLUSIONS: These results show that co-magaldrox suppressed the expression of the following virulence factors: adhesion, IL-8 inducibility, and expression of extracellular HSP60. Therefore, co-magaldrox is a potent anti-H. pylori and cytoprotective drug.     

Effect of an Aluminum Hydroxide-Magnesium Hydroxide Combination Drug on Adhesion,IL-8 Inducibility,and Expression of HSP60 by Helicobacter pylori

Background: Co-magaldrox (Maalox®) is used world-wide as an antacid and as a cytoprotective agent for gastritis and peptic ulcer diseases. We examined the effects of co-magaldrox on Helicobacter pylori. Methods: Adhesion of H. pylori to human gastric epithelial cells (MKN45) was evaluated by flow cytometry. Morphologic changes in H. pylori caused by co-magaldrox were determined by scanning electron microscopy. Induction of interleukin-8 (IL-8) from MKN45 cells was examined with enzyme-linked immunosorbent assay, and the intracellular and extracellular expression of heat-shock protein 60 (HSP60) was analyzed with sodium dodecyl sulphate-polyacrylamide gel electrophoresis and flow cytometry. Results: Adhesion of H. pylori to MKN 45 cells was significantly inhibited by 1.25%-5% co-magaldrox. H. pylori aggregated with co-magaldrox according to an electron microscopic examination. IL-8 secretion from MKN45 cells after H. pylori infection was also inhibited by co-magaldrox. Extracellular expression of HSP60 on the surface of H. pylori was decreased after treatment with co-magaldrox, whereas the intracellular synthesis of HSP60 was not. HSP60-induced IL-8 secretion was significantly inhibited by co-magaldrox in a dose-dependent manner. Conclusions: These results show that co-magaldrox suppressed the expression of the following virulence factors: adhesion, IL-8 inducibility, and expression of extracellular HSP60. Therefore, co-magaldrox is a potent anti-H. pylori and cytoprotective drug.     

Plaunotol suppresses interleukin-8 secretion induced by Helicobacter pylori: therapeutic effect of plaunotol on H. pylori infection

BACKGROUND: It has been suggested that gastric mucosal injury induced by Helicobacter pylori infection is mediated by interleukin-8 (IL-8). METHODS: We studied the effect of plaunotol, a drug extracted from the Plau-noi tree of Thailand, and reported it to be effective in the treatment of ulcers, of IL-8 secretion induced by H. pylori and of the inhibitory adhesion activity of the bacterium to gastric epithelial cells. Moreover, the therapeutic effect of plaunotol on H. pylori infection was assessed by using the gnotobiotic murine model. RESULTS: Plaunotol inhibited the growth of H. pylori (1.5 x 10(4) c.f.u./mL) at high doses (24-48 microg/mL), but not at low doses (3-6 microg/mL). Interleukin-8 secretion induced by H. pylori was inhibited by coculture with plaunotol in a dose-dependent manner. The adhesion of H. pylori to MKN45 cells was also suppressed by coculture with plaunotol in a dose-dependent manner. An in vivo study showed that plaunotol improved histological gastritis and decreased the H. pylori antibody titre. CONCLUSIONS: These findings suggest that plaunotol has a therapeutic effect on gastritis induced by H. pylori.     

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.     

Mucosal in Vitro Permeability in the Intestinal Tract of the Pig,the Rat,and Man: Species- and Region-Related Differences

Background: Excessive upregulation of gastric epithelial cell apoptosis is speculated to be associated with ulcerogenesis in Helicobacter pylori -positive peptic ulcer disease. H. pylori may have an ulcerogenic effect through induction of gastric epithelial cell apoptosis mediated by infiltrating T cells and their soluble products. Methods: The contents of soluble Fas ligand (sFasL) and interferon- &;#110 (IFN- &;#110 ) in organ cultures and the degree of apoptosis and the expression of apoptosis-related proteins in the gastric epithelium were examined using the mucosal tissues obtained from the antrum and the ulcer site in patients with H. pylori -positive gastric ulcer (GU). The molecular mechanisms of gastric epithelial cell apoptosis induced by sFasL and IFN- &;#110 were analyzed using epithelial cell lines, MKN 45 and KATO III. Results: The mucosal tissues of the ulcer site had substantially higher contents of sFasL and IFN- &;#110 in organ cultures regardless of its healing stage in association with increased numbers of apoptotic cells and enhanced expression of proapoptotic proteins Bak and Bax in the surface foveolar epithelium as compared with the antral tissues in patients with H. pylori -positive GU. The addition of sFasL caused increases in cytotoxic cell death and caspase-3 activation in MKN 45 and KATO III cells in which IFN- &;#110 -treated cells had more prominent effects than untreated cells. The expression of Bak in MKN 45 cells increased when they were treated with IFN- &;#110. Conclusions: Upregulation of mucosal sFasL and IFN- &;#110 may be involved in ulcerogenesis in patients with H. pylori- positive GU through induction of gastric epithelial cell apoptosis.     

cag致病岛的差异及不同激酶抑制剂对幽门螺杆菌诱导胃上皮细胞IL-8分泌的影响

目的：分析中国临床分离的幽门螺杆菌的cag致病岛的差异和不同激酶抑制剂对幽门螺杆菌诱导中国人胃上皮细胞IL-8分泌的影响。方法：在体外分别用中国临床分离的cagA^ cagE^ 、cagA^ cagE^ 、cagA^ cagE^ 、cagA^ cagE^ 的幽门螺杆菌与中国人胃上皮细胞MGC-803共培养,IL-8分泌用ELISA进行检测,比较蛋白激酶A、C、G和蛋白酪氨酸激酶的抑制剂对幽门螺杆菌诱导胃上皮细胞IL-8分泌的影响。结果：cagA^ cagE^ 幽门螺杆菌显增加了胃上皮细胞IL-8的分泌,cagA^ cagE^ 、cagA^ cagE^ 幽门螺杆菌作用次之,而cagA^ cagE^ 幽门螺杆菌不能增加胃上皮细胞IL-8的分泌。蛋白激酶A、C、G的抑制剂不能阻断幽门螺杆菌增加胃上皮细胞IL-8的分泌,而蛋白酪氨酸激酶的抑制剂阻断了幽门螺杆菌增加胃上皮细胞IL-8的分泌。结论：cagA^ cagE^ 幽门螺杆菌显增加了胃上皮细胞IL-8的分泌并且依赖于蛋白酪氨酸激酶的活性。     

Helicobacter pylori induced transactivation of SRE and AP-1 through the ERK signalling pathway in gastric cancer cells

BACKGROUND AND AIMS: Helicobacter pylori infection induces expression of proinflammatory cytokines such as interleukin (IL)-8 and tumour necrosis factor alpha (TNF-alpha) in gastric mucosa, and their genes have AP-1 binding sites in the promoter region. c-Fos is important for transactivation of AP-1 which has SRE in the promoter region. We conducted this study to confirm H pylori induced transactivation of these binding sites. METHODS: Transactivation of SRE and AP-1 was evaluated in human gastric cancer cells TMK1 and MKN45 by luciferase reporter assay in transient transfection. We compared the effects of coculture with four H pylori strains, a cag pathogenicity island (PAI) positive strain TN2, its isogenic vacA negative (TN2-DeltavacA) or cagE negative (TN2-DeltacagE) mutants, and a cag PAI negative clinical isolate T68. Phosphorylation of ERK1/2, JNK, and c-Jun was measured by immunoblot, induction of IL-8 secretion by ELISA, and the effects of MEK by inhibitor U0126. RESULTS: Both SRE and AP-1 were transactivated by coculture with TN2. Although TN2-DeltavacA induced comparable transactivation, TN2-DeltacagE and T68 showed decreased transactivation of SRE (65% and 51%) and AP-1 (71% and 54%, respectively, of TN2). Heat killed TN2 or indirect contact using a permeable membrane inhibited transactivation. Levels of phosphorylated ERK1/2, JNK, and c-Jun were increased by coculture with TN2. MEK inhibitor U0126 reduced TN2 induced transactivation of SRE and AP1, as well as secretion of IL-8, by 83%, 87%, and 53%, respectively, of TN2. CONCLUSIONS: Transactivation of SRE and AP-1, through ERK/MAPK and JNK/SAPK cascades, respectively, was found in gastric cancer cells cocultured with H pylori. Direct contact with viable bacteria possessing intact cag PAI is a prerequisite for the onset of intracellular signalling leading to AP-1 transactivation.     

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.     

Investigation of the sensitivities of distinct gastric cancer cells to parvovirus H‐1 induced cytotoxicity

OBJECTIVE: To investigate the sensitivities of distinct gastric cancer cells to parvovirus H‐1 induced cytotoxicity and the possible mechanism(s). METHODS: There were six distinct differentiated gastric cancer cell lines: HGC27 (undifferentiated), BGC823 (undifferentiated), MKN45 (poorly differentiated), AGS (poorly differentiated), SGC7901 (moderately differentiated) and MKN28 (well differentiated). The cell cycle distributions were measured by flow cytometry and the differential sensitivities of the six distinct gastric cancer cells after H‐1 virus infection were detected by MTT assay. RT‐PCR was used to detect viral NS1 gene expression in all six gastric cancer cell lines. RESULTS: The S phase ratios of HGC27, BGC823, MKN45, AGS, SGC7901 and MKN28 were 24.72%, 30.15%, 27.10%, 29.03%, 31.82% and 33.73%, respectively. HGC27 cells were sensitive to H‐1 virus induced cytotoxicity, followed by SGC7901 cells. MKN45 and AGS cells were moderately sensitive and MKN28 cells were insensitive. However, BGC823 cells were resistant to H‐1 virus induced cytotoxicity. The expressions of viral NS1 were higher in HGC27, BGC823, MKN45 and SGC7901 cells, and lower in AGS and MKN28 cells. CONCLUSIONS: The sensitivities of the distinct gastric cancer cells to H‐1 virus induced cytotoxicity were markedly different. In general, the poorly differentiated cells showed an enhanced sensitivity to H‐1 virus attack compared with well‐differentiated ones. The enhanced sensitivity of poorly versus well‐differentiated gastric cancer cells to H‐1 virus is related in part to the enhanced capacity of the former for NS1 protein production and accumulation. The undifferentiated BGC823 cells were resistant to H‐1 virus triggered cytotoxicity. It may further verify that not all tumor cells are sensitive to H‐1 virus lytic effects.     

Helicobacter pylori infection induces duodenitis and superficial duodenal ulcer in Mongolian gerbils

BACKGROUND: There is no direct evidence for an animal model of Helicobacter pylori induced duodenal ulcer. AIM: In this study we evaluated the roles of bacterial strain and age of experimental animals in induction of duodenitis and duodenal ulcer in Mongolian gerbils after H pylori infection. METHODS: Specific pathogen free Mongolian gerbils were inoculated orally with three bacterial strains (H pylori ATCC 43504, TN2GF4, and K-6, a clinical isolate from a patient with gastric cancer in our clinic). These strains have both the cagA gene and VacA. Five week old gerbils were used to emulate prematurity infection and 14 week old animals were used as mature test subjects. Animals were observed for 12 weeks after inoculation. Interleukin 8 (IL-8) production in gastric epithelial cells (MKN74) after coculture with the H pylori strains was measured by ELISA. RESULTS: Gastritis and gastric ulcers were found in all gerbils infected with the three strains. However, duodenitis and gastric metaplasia were seen more frequently in gerbils infected with TN2GF4 and K-6 strains than in the ATCC 43504 infected or control groups (p<0.05). Superficial duodenal ulcers with severe duodenitis and gastric metaplasia were found in two gerbils inoculated at 14 weeks with the TN2GF4 strain but none at five weeks. The TN2GF4 strain stimulated significantly higher levels of IL-8 than ATCC 43504 and K6 strains (p=0.0039). CONCLUSIONS: When injected into adult Mongolian gerbils, a specific strain (TN2GF4) of H pylori can induce duodenitis with gastric metaplasia and superficial duodenal ulcers. Induction of duodenal ulcer in an animal model fulfills the requirements of Koch's postulates for establishing a role for H pylori as a causative agent.     

Use of SiRNA to investigate the role of CagA on H. pylori induced IL-8 production from gastric epithelial cells

BACKGROUND/AIMS: H. pylori infection can stimulate gastric epithelial cells to release IL-8. We studied the effect of small interference RNA (SiRNA) on CagA expression and the H. pylori-induced release of IL-8 by gastric epithelial cells. METHODOLOGY: SiRNA's were transferred into CagA positive strain H. pylori NCTC 11637 using electroporation. The CagA positive strain NCTC 11637 and the CagA negative strain NCTC 11639 were co-cultured with gastric epithelial cells and the level of IL-8 in the supernatant was measure by ELISA. RT-PCRs and Western blot were performed to detect CagA expression. RESULTS: The level of IL-8 induced by NCTC 11637 was higher than that produced by infection with NCTC 11639. Treatment with SiRNA iii or v resulted in a decrease in IL-8 production of more than 50% and a corresponding decrease in CagA mRNA. The levels of CagA mRNA at 6, 12, and 24 hours after electroporation were 31.3%, 43.5%, and 76.8% of the control levels, respectively (P = 0.0025). Western blot following SiRNA iii showed the level of CagA protein degraded to 30% of control by x hours. CONCLUSIONS: CagA is involved in H. pylori induced IL-8 release from gastric epithelial cells.     

Clinical presentation in relation to diversity within the Helicobacter pylori cag pathogenicity island

OBJECTIVE: This study investigated the genetic diversity of the cag pathogenicity island (PAI) in Helicobacter pylori (H. pylori) in relation to clinical outcome and interleukin (IL)-8 production. METHODS: Seven genes in the cag PAI (cagA, cagE, cagG, cagM, cagT, open reading frame 13 and 10) were examined by polymerase chain reaction and Southern blot hybridization using H. pylori from 120 patients with different presentations (duodenal ulcer, gastric cancer, gastritis alone). IL-8 production from AGS cells (gastric cancer cell line) cocultured with H. pylori was measured by ELISA. RESULTS: An intact cag PAI was present in 104 (87%) isolates, and five (4%) had deletions within the cag PAI; 11 (9%) lacked the entire cag PAI. Clinical isolates containing the complete cag PAI induced a greater secretion of IL-8 as compared with those without the cag PAI (3048 +/- 263 vs 480 +/- 28 pg/ml, p < 0.001). Deletion of only cagG reduced IL-8 secretion by two thirds. Deletions of more than one locus reduced IL-8 secretion to background. A similar proportion of H. pylori from patients with gastritis, duodenal ulcer, or gastric cancer had intact cag PAI (88%, 88%, and 85%, respectively). Although the presence of cagG was a better predictor of the presence of an intact cag PAI than cagA or cagE, the presence or absence of any of these genes had no association with clinical presentation. CONCLUSION: Although the cag PAI plays an important role in IL-8 production, clinical presentation cannot be predicted by the presence of an intact cag PAI or any of these seven cag PAI genes.