Pylori In Mice Research Articles

An intranasal influenza virus vector vaccine protects against Helicobacter pylori in mice.

Helicobacter pylori is a human pathogen that infects almost half of the population. Antibiotic resistance in H. pylori threatens health and increases the demand for prophylactic and therapeutic vaccines. Traditional oral vaccine research faces considerable challenges because of the epithelial barrier, potential enterotoxicity of adjuvants, and the challenging conditions of the gastric environment. We developed an intranasal influenza A virus (IAV) vector vaccine based on two live attenuated influenza viruses with modified acidic polymerase protein (PA) genes encoding the A subunit of H. pylori neutrophil-activating protein (NapA), named IAV-NapA, including influenza virus A/WSN/33 (WSN)-NapA and A/Puerto Rico/8/34 (PR8)-NapA. These recombinant influenza viruses were highly attenuated and exhibited strong immunogenicity in mice. Vaccination with IAV-NapA induced antigen-specific humoral and mucosal immune responses while stimulating robust Th1 and Th17 cell immune responses in mice. Our findings suggest that prophylactic and therapeutic vaccination with influenza virus vector vaccines significantly reduces colonization of H. pylori and inflammation in the stomach of mice.IMPORTANCEHelicobacter pylori is the most common cause of chronic gastritis and leads to severe gastroduodenal pathology in some patients. Many studies have shown that Th1 and Th17 cellular and gastric mucosal immune responses are critical in reducing H. pylori load. IAV vector vaccines can stimulate these immune responses while overcoming potential adjuvant toxicity and antigen dosing issues. To date, no studies have demonstrated the role of live attenuated IAV vector vaccines in preventing and treating H. pylori infection. Our work indicates that vaccination with IAV-NapA induces antigen-specific humoral, cellular, and mucosal immunity, producing a protective and therapeutic effect against H. pylori infection in BALB/c mice. This undescribed H. pylori vaccination approach may provide valuable information for developing vaccines against H. pylori infection.

IL-8 Links NF-κB and Wnt/β-Catenin Pathways in Persistent Inflammatory Response Induced by Chronic <i>Helicobacter pylori</i> Infection

Helicobacter pylori (H. pylori) infection can cause persistent inflammatory response in human gastric mucosal epithelial cells, which may result in the occurrence of cancer. However, the underlying mechanism of carcinogenesis has not been elucidated yet. Herein, we established the models of chronic H. pylori infection in GES-1 cells and C57BL/6J mice. Interleukin 8 (IL-8) level was detected by ELISA. The expression of NF-κB p65, IL-8, Wnt2 and β-catenin mRNA and proteins was evaluated by real-time PCR, Western blotting, immunofluorescence staining, and immunohistochemistry. The infection of H. pylori in mice was evaluated by rapid urease test, HE staining and Warthin‒Starry silver staining. The morphological changes of gastric mucosa were observed by electron microscopy. Our results showed that in H. pylori infected gastric mucosal cells along with activation of NF-κB signaling pathway and increase of IL-8 level, the expression of Wnt2 was also increased significantly, which preliminarily indicates that IL-8 can positively regulate the expression of Wnt2. Studies in chronic H. pylori infected C57BL/6J mice models showed that there was an increased incidence of premalignant lesions in the gastric mucosa tissue. Through comparing changes of gastric mucosal cell ultrastructure and analyzing the relationship between NF-κB signaling pathway and Wnt2 expression, we found that H. pylori infection activated NF-κB signal pathways, and the massive release of IL-8 was positively correlated with the high expression of Wnt2 protein. Subsequently, the activated Wnt/β-catenin signal pathways may be involved in the malignant transformation of gastric mucosal cells. Collectively, H. pylori chronic infection may continuously lead to persistent inflammatory response: activate NF-κB pathway, promote IL-8 release and thereby activate Wnt/β-catenin pathway. IL-8 probably plays an important role of a linker in coupling these two signal pathways.

Coptisine inhibits Helicobacter pylori and reduces the expression of CagA to alleviate host inflammation in vitro and in vivo

Ethnopharmacological relevanceHelicobacter pylori (H. pylori) is a major pathogen colonized in the human stomach and is implicated in gastritis, peptic ulcer, and gastric carcinoma. Antibiotics are useful for eradicating H. pylori but failed for drug resistance, making it urgent to develop effective and safe drugs. Rhizoma Coptidis was reported as one of the most effective Chinese medicines to treat H. pylori-related gastrointestinal diseases, while the precise antimicrobial mechanism remains unclear. Thus, it is of great significance to study the antimicrobial ingredients and corresponding mechanisms of Rhizoma Coptidis. Aim of the studyTo search for the most effective alkaloid against H. pylori in Rhizoma Coptidis and illustrate the probable mechanisms. Materials and methodsFive main alkaloids in Rhizoma Coptidis were isolated. Minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) were tested to determine the most effective one. Bacterial growth experiments, Annexin V-FITC/PI staining, TUNEL staining, and transmission electron microscopy (TEM) were performed to further study the anti-H. pylori activity of coptisine (Cop). The in vivo effect of Cop on H. pylori eradication rate and H. pylori-induced inflammation was investigated in mice. Transcriptomics was used to understand the underlying mechanism of eradicating H. pylori and reducing host inflammation. Western blot, RT-PCR, and ELISA experiments were utilized and confirmed that cagA was one of the targets of Cop. ResultsAccording to the MIC and MBC, Cop was the most effective alkaloid against H. pylori, especially with no drug resistance developed. In vitro experiments showed that Cop inhibited H. pylori by inducing DNA fragmentation, phosphatidylserine exposure, and membrane damage. Cop (150 mg/kg/day) effectively eradicated H. pylori in mice and reduced the levels of IL-2 and IL-6 to relieve gastric inflammation. Transcriptomic analysis revealed that virulence factor cagA was one of the hub genes associated with the inflammation-improving effect of Cop. That is, Cop could decrease the expression of CagA and subsequently reduce the translocation of CagA to gastric epithelial cells, thereby improving the morphology of hummingbird-like phenotype induced by CagA and alleviating inflammation. ConclusionsCop is the most effective alkaloid in Rhizoma Coptidis and might act through multiple mechanisms for H. pylori eradication along with reducing the expression of CagA to alleviate inflammation.

IL-8 Links NF-κB and Wnt/β-Catenin Pathways in Persistent Inflammatory Response Induced by Chronic Helicobacter pylori Infection

Helicobacter pylori (H. pylori) infection can cause persistent inflammatory response in human gastric mucosal epithelial cells, which may result in the occurrence of cancer. However, the underlying mechanism of carcinogenesis has not been elucidated yet. Herein, we established the models of chronic H. pylori infection in GES-1 cells and C57BL/6J mice. Interleukin 8 (IL-8) level was detected by ELISA. The expression of NF-κB p65, IL-8, Wnt2 and β-catenin mRNA and proteins was evaluated by real-time PCR, Western blotting, immunofluorescence staining, and immunohistochemistry. The infection of H. pylori in mice was evaluated by rapid urease test, H&E staining and Warthin-Starry silver staining. The morphological changes of gastric mucosa were observed by electron microscopy. Our results showed that in H. pylori infected gastric mucosal cells along with activation of NF-κB signaling pathway and increase of IL-8 level, the expression of Wnt2 was also increased significantly, which preliminarily indicates that IL-8 can positively regulate the expression of Wnt2. Studies in chronic H. pylori infected C57BL/6J mice models showed that there was an increased incidence of premalignant lesions in the gastric mucosa tissue. Through comparing changes of gastric mucosal cell ultrastructure and analyzing the relationship between NF-κB signaling pathway and Wnt2 expression, we found that H. pylori infection activated NF-κB signal pathways, and the massive release of IL-8 was positively correlated with the high expression of Wnt2 protein. Subsequently, the activated Wnt/β-catenin signal pathways may be involved in the malignant transformation of gastric mucosal cells. Collectively, H. pylori chronic infection may continuously lead to persistent inflammatory response: activate NF-κB pathway, promote IL-8 release and thereby activate Wnt/β-catenin pathway. IL-8 probably plays an important role of a linker in coupling these two signal pathways.

Helicobacter pylori, Protected from Antibiotics and Stresses Inside Candida albicans Vacuoles, Cause Gastritis in Mice.

Due to (i) the simultaneous presence of Helicobacter pylori (ulcer-induced bacteria) and Candida albicans in the stomach and (ii) the possibility of prokaryotic–eukaryotic endosymbiosis (intravacuolar H. pylori in the yeast cells) under stresses, we tested this symbiosis in vitro and in vivo. To that end, intravacuolar H. pylori were induced by the co-incubation of C. albicans with H. pylori under several stresses (acidic pH, non-H. pylori-enrichment media, and aerobic environments); the results were detectable by direct microscopy (wet mount) and real-time polymerase chain reaction (PCR). Indeed, intravacuolar H. pylori were predominant under all stresses, especially the lower pH level (pH 2–3). Interestingly, the H. pylori (an amoxicillin-sensitive strain) inside C. albicans were protected from the antibiotic (amoxicillin), while extracellular H. pylori were neutralizable, as indicated by the culture. In parallel, the oral administration of intravacuolar H. pylori in mice caused H. pylori colonization in the stomach resulting in gastritis, as indicated by gastric histopathology and tissue cytokines, similar to the administration of free H. pylori (extra-Candida bacteria). In conclusion, Candida protected H. pylori from stresses and antibiotics, and the intravacuolar H. pylori were able to be released from the yeast cells, causing gastric inflammation with neutrophil accumulations.

CagA+Helicobacter pylori, Not CagA–Helicobacter pylori, Infection Impairs Endothelial Function Through Exosomes-Mediated ROS Formation

BackgroundHelicobacter pylori (H. pylori) infection increases the risk for atherosclerosis, and ROS are critical to endothelial dysfunction and atherosclerosis. CagA is a major H. pylori virulence factor associated with atherosclerosis. The present study aimed to test the hypothesis that CagA+ H. pylori effectively colonizes gastric mucosa, and CagA+ H. pylori, but not CagA– H. pylori, infection impairs endothelial function through exosomes-mediated ROS formation.MethodsC57BL/6 were used to determine the colonization ability of CagA+ H. pylori and CagA– H. pylori. ROS production, endothelial function of thoracic aorta and atherosclerosis were measured in CagA+ H. pylori and CagA– H. pylori infected mice. Exosomes from CagA+ H. pylori and CagA– H. pylori or without H. pylori infected mouse serum or GES-1 were isolated and co-cultured with bEND.3 and HUVECs to determine how CagA+ H. pylori infection impairs endothelial function. Further, GW4869 was used to determine if CagA+H. pylori infection could lead to endothelial dysfunction and atherosclerosis through an exosomes-mediated mechanism.ResultsCagA+ H. pylori colonized gastric mucosa more effectively than CagA– H. pylori in mice. CagA+ H. pylori, not CagA– H. pylori, infection significantly increased aortic ROS production, decreased ACh-induced aortic relaxation, and enhanced early atherosclerosis formation, which were prevented with N-acetylcysteine treatment. Treatment with CagA-containing exosomes significantly increased intracellular ROS production in endothelial cells and impaired their function. Inhibition of exosomes secretion with GW4869 effectively prevented excessive aortic ROS production, endothelial dysfunction, and atherosclerosis in mice with CagA+ H. pylori infection.ConclusionThese data suggest that CagA+ H. pylori effectively colonizes gastric mucosa, impairs endothelial function, and enhances atherosclerosis via exosomes-mediated ROS formation in mice.

Outer Membrane Vesicles of Helicobacter pylori 7.13 as Adjuvants Promote Protective Efficacy Against Helicobacter pylori Infection.

Helicobacter pylori(H. pylori), a gram-negative bacterium in the human stomach with global prevalence, is relevant to chronic gastrointestinal diseases. Due to its increasing drug resistance and the low protective efficacy of some anti-H. pylori vaccines, it is necessary to find a suitable adjuvant to improve antigen efficiency. In our previous study, we determined that outer membrane vesicles (OMVs), a multicomponent secretion generated by gram-negative bacteria, of H. pylori were safe and could induce long-term and robust immune responses against H. pylori in mice. In this study, we employed two common vaccines, outer membrane proteins (OMPs) and whole cell vaccine (WCV) to assess the adjuvanticity of OMVs in mice. A standard adjuvant, cholera toxin (CT), was used as a control. Purified H. pylori OMVs used as adjuvants generated lasting anti-H. pylori resistance for 12 weeks. Additionally, both systematic and gastric mucosal immunity, as well as humoral immunity, of mice immunized with vaccine and OMVs combinations were significantly enhanced. Moreover, OMVs efficiently promoted Th1 immune response, but the response was skewed toward Th2 and Th17 immunity when compared with that induced by the CT adjuvant. Most importantly, OMVs as adjuvants enhanced the eradication of H. pylori. Thus, OMVs have potential applications as adjuvants in the development of a new generation of vaccines to treat H. pylori infection.

Intervention effects of lotus leaf flavonoids on gastric mucosal lesions in mice infected with Helicobacter pylori.

Helicobacter pylori (H. pylori) is one of the main factors that cause gastric lesions. The lotus leaf is an edible plant used in traditional Eastern medicine. This study evaluates the intervention effects of lotus leaf flavonoids (LLF) on gastric mucosal lesions in mice infected with H. pylori and explores their mechanism of action. High-performance liquid chromatography analysis reveals that LLF contain kaempferitrin (kaempferol-3,7-dirhamnoside), hypericin, astragalin (kaempferol-3-glucoside), phlorizin, and quercetin. LLF can reduce the number of gastric mucosal lesions and tissue lesions in mice with H. pylori-induced gastric lesions. LLF can increase the levels of somatostatin and vasoactive intestinal peptide in the serum of mice with gastric lesions and decrease the levels of substance P and endothelin-1 to inhibit gastric lesions. LLF can also reduce the levels of interleukin (IL)-6, IL-12, tumor necrosis factor (TNF)-α, and interferon-gamma cytokines in the serum of mice with gastric lesions. Using a quantitative polymerase chain reaction assay it can be seen that LLF can downregulate mRNA expressions of TNF-α, IL-1β, myeloperoxidase, keratin (KRT) 16, KRT6b, and transglutaminase 3 epidermal in the gastric tissues of mice with gastric lesions. Western blot analysis indicates that LLF can downregulate the protein expressions of caspase-1, Nod-like receptor protein 3, IL-1β, TNF-α, and Toll-like receptor 4 in the gastric tissues of mice with gastric lesions. LLF have beneficial effects on gastric lesions induced by H. pylori. Meanwhile LLF is more active in competition with ranitidine. LLF represent an active substance that can inhibit H. pylori-induced gastric lesions. The flavones of LLF may enhance the inhibition of gastric mucosal lesions by promoting the interaction between the compounds.

IL-22-induced antimicrobial peptides are key determinants of mucosal vaccine-induced protection against H. pylori in mice.

Despite the recent description of the mucosal vaccine-induced reduction of Helicobacter pylori natural infection in a phase 3 clinical trial, the absence of immune correlates of protection slows the final development of the vaccine. In this study, we evaluated the role of interleukin (IL)-22 in mucosal vaccine-induced protection. Gastric IL-22 levels were increased in mice intranasally immunized with urease+cholera toxin and challenged with H. felis, as compared with controls. Flow cytometry analysis showed that a peak of CD4+IL-22+IL-17+ T cells infiltrating the gastric mucosa occurred in immunized mice in contrast to control mice. The inhibition of the IL-22 biological activity prevented the vaccine-induced reduction of H. pylori infection. Remarkably, anti-microbial peptides (AMPs) extracted from the stomachs of vaccinated mice, but not from the stomachs of non-immunized or immunized mice, injected with anti-IL-22 antibodies efficiently killed H. pylori in vitro. Finally, H. pylori infection in vaccinated RegIIIβ-deficient mice was not reduced as efficiently as in wild-type mice. These results demonstrate that IL-22 has a critical role in vaccine-induced protection, by promoting the expression of AMPs, such as RegIIIβ, capable of killing Helicobacter. Therefore, it can be concluded that urease-specific memory Th17/Th22 cells could constitute immune correlates of vaccine protection in humans.

Elucidating the function and tolerance mechanism of gamma delta (γ δ) T cells in a Helicobacter pylori infection model.

In this study, the functions and mechanisms of γ δ T cells were analyzed in patients infected with Helicobacter pylori. Peripheral blood was collected from gastritis patients in the Gastroenterology Department of Ningbo No. 2 Hospital. Preliminary analyses revealed 24 H. pylori-positive and 17 H. pylori-negative patients. The wild-type and γ δ T knockout mice were infected with cultured H. pylori cells (obtained from the H. pylori-positive patients). H. pylori in mice was quantified by polymerase chain reaction; gastritis was confirmed by hematoxylin and eosin staining. The TCR-δ(-/-) mice were treated with vein adoptive immunotherapy 24 h prior to H. pylori inoculation; the same method was used to detect the extent of gastritis and bacterial colonization. The γ δ T knockout mice showed high levels of H. pylori infection than the wild-type mice; in addition, the knockout mice showed severe disease pathology. γ δ T knockout mice also displayed increased matrix metalloproteinase-9 (MMP-9) and decreased MMP-7 expression in the gastric mucosa. γ δ T cells play a protective role in patients infected with H. pylori. γ δ T cell [responsible for the production of interleukin-17 (IL-17) and IL-22] expression was increased in H. pylori-positive patients, indicating statistical significance. However, there was no significant difference in interferon-gamma + γ δ T expression between the positive and negative patients. This study demonstrated the probable involvement of γ δ T cells in the immune response of an organism, via the secretion of IL-17 and IL-22.

Helicobacter hepaticus cholesterol-α-glucosyltransferase is essential for establishing colonization in male A/JCr mice.

Helicobacter pylori cholesterol-α-glucosyltransferase (cgt) is essential for survival of H. pylori in mice. Enterohepatic H. hepaticus, the cause of colonic and hepatocellular carcinoma in susceptible mouse strains, contains an ortholog of the H. pylori cgt. However, the role of cgt in the pathogenesis of H. hepaticus has not been investigated. Two cgt-deficient isogenic mutants of wild-type H. hepaticus (WT) 3B1 were generated and used to inoculate male A/JCr mice. Cecal and hepatic colonization levels of the mutants and WT 3B1 as well as select inflammation-associated cytokines were measured by qPCR at 4 months postinoculation. Both mutants were undetectable in the cecum of any inoculated mice (10 per mutant) but were detected in two livers (one for each mutant); by contrast, 9 and 7 of 10 mice inoculated with WT 3B1 were qPCR positive in the ceca and livers, respectively. The mice inoculated with the mutants developed significantly less severe hepatic inflammation (p < .05) and also produced significantly lower hepatic mRNA levels of proinflammatory cytokines Ifn-γ (p < .01) and Tnf-α (p ≤ .02) as well as anti-inflammatory factors Il10 and Foxp3 compared with the WT 3B1-inoculated mice. Additionally, the WT 3B1-inoculated mice developed significantly higher Th1-associated IgG2a (p < .0001) and Th2-associated IgG1 responses (p < .0001) to H. hepaticus infection than mice dosed with isogenic cgt mutants. Our data indicate that the cholesterol-α-glucosyltransferase is required for establishing colonization of the intestine and liver and therefore plays a critical role in the pathogenesis of H. hepaticus.

Combating Helicobacter Pylori Infections with Mucoadhesive Nanoparticles Loaded with Garcinia Mangostana Extract

To combat the resistance of Helicobacter pylori to antibiotics through the use of Garcinia mangostana extract (GME) in the form that can be localized at stomach mucosa. GME and its major active component, α-mangostin, are encapsulated into the moderately acid stable mucoadhesive nanocarriers, and tested for anti-H. pylori and antiadhesion activities in vitro and their ability to eradicate H. pylori in infected mice. The two in vitro activities are observed and are enhanced when the materials are encapsulated into nanocarriers. Preliminary in vivo tests revealed the ability to combat H. pylori in mice following oral administration of the encapsulated GME, but not the unencapsulated GME. Nanoencapsulated GME is a potential anti-H. pylori agent.

Sa1885 IL-23 Contributes to Vaccine-Induced Protective Immunity Against H. pylori in Mice

Sa1885 IL-23 Contributes to Vaccine-Induced Protective Immunity Against H. pylori in Mice

Systemic immunization with an epitope-based vaccine elicits a Th1-biased response and provides protection against Helicobacter pylori in mice

Vaccine-mediated Th1-biased CD4+ T cell responses have been shown to be crucial for protection against Helicobacter pylori (H. pylori). In this study, we investigated whether a vaccine composed of CD4+ T cell epitopes together with Th1 adjuvants could confer protection against H. pylori in a mouse model. We constructed an epitope-based vaccine, designated Epivac, which was composed of predicted immunodominant CD4+ T cell epitopes from H. pylori adhesin A (HpaA), urease B (UreB) and cytotoxin-associated gene A product (CagA). Together with four different Th1 adjuvants, Epivac was administered subcutaneously and the prophylactic potential was examined. Compared to non-immunized mice, immunization with Epivac alone or with a Th1 adjuvant significantly reduced H. pylori colonization, and better protection was observed when an adjuvant was used. Immunized mice exhibited a strong local and systemic Th1-biased immune response, which may contribute to the inhibition of H. pylori colonization. Though a significant specific antibody response was induced by the vaccine, no correlation was found between the intensity of the humoral response and the protective effect. Our results suggest that a vaccine containing CD4+ T cell epitopes is a promising candidate for protection against H. pylori infection.

Experimental study on prevention and treatment of Helicobacter pylori infection by Lactobacillus acidophilus in C57BL/6 mice

Objective To investigate the potential effects of Lactobacillus acidophilus strain L6 on prevention and treatment of Helicobacter pylori (H. pylori) infection in animal models. Methods A total of 200 hundred C57BL/6 mice were used in the study. ① Sixty mice were divided into control group (infected with H. pylori) and prevention group (previously treated with L6 and followed by infection with H. pylori) with 30 each. The incidence of H. pylori infection was compared between two groups.② Sixty mice were divided into control group (infected with H. pylori) and treatment group (infected with H. pylori for 4 weeks and followed by treatment with L6) with 30 each. The changes of H. pylori infection was compared between tow groups. ③ Eighty mice previously infected with H. pylori were orally administrated with L6 in the water supply over a period of 9 months. Of which, 40 H. pylori negative mice were either continuously or discontinuously treated with L6. The re-infection of H. pylori was compared between two groups. The H. pylori infection, gastric mucosal inflammatory and serum anti-H. pylori-lgG titer was measured by using urea breath test, histopathology and ELISA, respectively. Results The positive rate of H. pylori infection was 100% in control group and 20% in prevention group (pre-treated with L6 for 1 week). Whereas there was no H. pylori infection in the rest mice of prevention group pre-treated with L6 for 2 or 4 weeks. There was significant difference in H. pylori infection between two groups (P<0. 05). The serum anti-H. pylori IgG titer was lower in treatment group than in control group (P<0. 05). The reinfection of H. pylori in mice continuously treated with L6 was significantly lower than those discontiously treated with L6 (P<0. 05). Conclusions Preventively feeding L6 can significantly reduce the H. pylori infection in C57BL/6 mice, whereas the inhibition of H. pylori re-infection can also be achieved with long-term administration of L6. Key words: Lactobacillusacidophilus; Helicobacter pylori; Animal experimentation

Abstract 1476: Cell surface nucleolin acts as a specific receptor of Tipα, a carcinogenic factor released from H. pylori

Abstract TNF-α inducing protein (Tipα), a unique carcinogenic factor of H. pylori, has the potential to transform Bhas 42 (v-H-ras transfected BALB/3T3) cells. Tipα directly binds to gastric cancer cells and then induces expression of TNF-α gene, an endogenous tumor promoter, in gastric epithelial cells. H. pylori obtained from gastric cancer patients release significant amounts of Tipα into the culture medium compared with H. pylori of gastritis patients, indicating that Tipα plays important role in gastric cancer development. Recently, Tipα vaccination was reported to significantly reduce the colonization of H. pylori in mice associated with high levels of Tipα-specific antibody: All the results indicate that Tipα is a suitable molecular target for gastric cancer prevention. We recently demonstrated that nucleolin is a specific binding protein for Tipα in mouse gastric cancer cells MGT-40 using FLAG pull-down assay. Although nucleolin is known to a be non-ribosomal protein in nucleoli, we found that nucleolin is present on the cell surface of MGT-40 cells and various human gastric cancer cell lines, such as KATOIII and AGS, as determined by flow cytometry with anti-NUC295 antibody (recognizing cell surface nucleolin), and by cell fractionation. Pretreatment with anti-NUC295 antibody induced the enhancement of Tipα incorporation into the MGT-40 cells, but pretreatment with tunicamycin, an inhibitor of N-glycosylation decreased the amounts of cell surface nucleolin and inhibited internalization of Tipα, resulting in down-regulation of TNF-α gene expression. We here present evidence that cell surface localized nucleolin acts as a receptor of Tipα and shuttles Tipα from cell surface to cytosol and nuclei, and we discuss the significant roles of cell surface nucleolin in gastric cancer development by H. pylori infection. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 1476.

Evaluation of ISCOMATRIX™ and ISCOM™ vaccines for immunisation against Helicobacter pylori

An important obstacle to development of an effective vaccine against Helicobacter pylori is the lack of a suitable adjuvant. This study evaluated the effectiveness of ISCOMATRIX™ and ISCOM™ vaccines at inducing protective immunity against H. pylori in mice. Immunisation with ISCOMATRIX™ and ISCOM™ vaccines resulted in a reduction in H. pylori colonisation equivalent to that induced by the gold standard cholera toxin (CT) adjuvant. Protection was induced in two mouse backgrounds, using two different bacterial antigens and following vaccine delivery via either the intranasal or subcutaneous route. This supports the potential of ISCOMATRIX™ and ISCOM™ technologies in H. pylori vaccine development.

The Adjuvant Effect of CpG Oligodeoxynucleotide Linked to the Non‐Toxic B Subunit of Cholera Toxin for Induction of Immunity Against H. pylori in Mice

The present study was carried out to test the immunostimulatory and adjuvant effects of the non-toxic B subunit of cholera toxin (CTB), CpG oligodeoxynucleotide (ODN) and CpG ODN linked to CTB (CTB-CpG) for generation of immunity against H. pylori in mice. Herein, we showed that CTB-CpG induces more potent proinflammatory cytokine and chemokine responses in the cervical and the mesenteric lymph nodes (CLN and MLN, respectively) cells in vitro compared with those of CTB and CpG ODN. The adjuvant effects of these agents were examined following intranasal immunization of C57Bl/6 mice with H. pylori lysate in combination with CpG ODN, CTB or CTB-CpG. All three immunization regimes resulted in high H. pylori-specific IgG antibody responses; however, only the CTB-CpG and, to some extent, the CpG ODN immunized mice mounted a sustainable IgG2c antibody response. Importantly, mice immunized with H. pylori antigen and CTB-CpG or CpG ODN, but not CTB, developed strong H. pylori-specific proliferative and IFN-gamma responses in their MLN CD4+ T cells upon recall antigen stimulation in vitro. These mice also had significantly lower bacterial load compared with the control-infected mice. Furthermore, the CTB-CpG and the CpG ODN immunized mice developed increased specific IgA antibody responses in their gastrointestinal tracts following H. pylori challenge. These results imply that CTB-CpG and CpG ODN, but not CTB, could serve as nasal adjuvants for induction of a H. pylori-specific Th1 type immunity in MLN and also a specific mucosal IgA antibody response in the gastrointestinal tract upon H. pylori challenge.

Gastric inflammatory markers and interleukins in patients with functional dyspepsia treated with astaxanthin

The chronic active inflammation caused by Helicobacter pylori is dominated by neutrophils, macrophages, lymphocytes and plasma cells. Several interleukins are involved in the inflammatory process. The aim of this study was to investigate the effect of astaxanthin on gastric inflammation in patients with functional dyspepsia. Forty-four consecutive patients were included, and biopsies were examined for IL-4, IL-6, IL-8, IL-10, interferon-gamma, CD4, CD8, CD14, CD19, CD25 and CD30. Patients were randomized: 21 patients were treated with 40 mg of astaxanthin daily, and 23 patients were treated with a placebo. There was a significant decrease in gastric inflammation in H. pylori-positive patients from both groups. There were no significant changes in the density of H. pylori or in any of the interleukins during or after treatment. There was a significant up-regulation of CD4 and down-regulation of CD8 in patients with H. pylori treated with astaxanthin. Astaxanthin had an effect on the inflammation and on the density of H. pylori in mice in a study where the diet could be standardized without antioxidants (Bennedsen et al., 1999). These dietary conditions are impossible in studies involving humans, and may be due to the minor effect when the host have access to antioxidants in their diet.

Considering Increased Mouse Stomach Mass When Calculating Prophylactic Vaccine Efficacy Against Helicobacter pylori

Calculating the level of protection follow vaccination against Helicobacter pylori in mice is conventionally performed based on stomach mass. However, prophylactic vaccination is associated with a post-immunisation gastritis which results in considerable thickening of the gastric mucosa. Here, an analysis of nine experiments demonstrates that this post-immunisation gastritis can actually induce a significant increase in stomach mass. To determine whether this mass increase impacts upon the perceived level of protection, calculations were performed comparing colony-forming units (CFU) either per gram of tissue or per whole stomach. A trend was apparent towards increased stomach mass influencing the degree of protection if CFU were calculated per gram. However, as this effect was not significant, it appears valid to express CFU either per gram or per stomach (if age and gender matched mice are used).