• Microbiology and Biotechnology Letters
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• v.25 no.2
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• pp.129-136
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• 1997

Helicobacter pylori is a spiral-shaped, microaerophilic human gastric pathogen causing chronic-active gastritis in association with duodenal ulcer and gastric cancer. To investigate the possibility of H. pylori outer membrane proteins (OMPS) as the oral vaccine antigens, sarcosine-insoluble outer membrane fraction has been prepared from H. pylori NCTC 11637. The major OMPs having apparent molecular masses of 62 kDa, 54 kDa and 33 kDa were detected by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), which were identified as urease B subunit (UreB), heat shock protein (Hsp54 kDa) and urease A subunit (UreA), respectively. Minor protein bands of 57 kDa, 52 kDa, 40 kDa, 36 kDa and 31 kDa were also observed. The antigenicity of H. pylori OMPs and antigenic cross-reactivity among the strains were determined by immunoblot analysis using anti-H. pylori OMPs antisera or intestinal lavage solutions. The results showed that UreB, Hsp54 kDa, UreA and 40 kDa proteins vigorously stimulated mucosal immune response rather than systemic immunity. From this results, these proteins seemed to be useful as the antigen candidates for the oral vaccine. The immunoblotting results with surface proteins from eight isolated H. pylori strains were similar to that of H. pylori NCTC 11637. The IgA which had been arised from oral administration of H. pylori OMPs, was able to bind H. pylori whole-cells.

• Journal of Microbiology and Biotechnology
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• v.16 no.2
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• pp.318-324
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• 2006

In order to develop an oral vaccine to prevent H. pylori infection, we have expressed the hpaA gene of H. pylori K51 isolated from Korean patients, encoding 29-kDa HpaA that is known to be localized on the cell surface and flagella sheath, in a live delivery vector system, Lactococcus lactis. The hpaA gene, amplified by PCR using the genomic DNA of H. pylori K51, was cloned in the pGEX-2T vector, and the DNA sequence analysis revealed that the hpaA gene of H. pylori K51 had 99.7% and 94.8% identity with individual hpaA genes of the H. pylori 26695 strain (U.K) and the J99 strain (U.S.A). A polyclonal anti-HpaA antibody was raised in rats using GST-HpaA fusion protein as the antigen. The hpaA gene was inserted in an E. coli-L. lactis-shuttle vector (pMG36e) to express in L. lactis. Western blot analysis showed that the expression level of HpaA in the L. lactis transformant remained constant from the exponential phase to the stationary phase, without extracelluar secretion. These results indicate that the HpaA of H. pylori K51 was successfully expressed in L. lactis, and suggest that the recombinant L. lactis expressing HpaA may be applicable as an oral vaccine to induce a protective immune response against H. pylori.

• Journal of the Korean Society of Food Science and Nutrition
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• v.34 no.4
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• pp.484-488
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• 2005

This study has been to produce of anti-H. pylori antibody in milk produced from cows immunized with antigen of Helicobacter pylori and to search the relationship between vaccine dosage and antibody formation and impact of vaccine dosage on cows. The content of anti-H. pylori antibody in serum and whey increased in accordance with vaccine dosage volume. It has been confirmed that the formation of high-quantified antibody was produced in all groups with vaccine dosages of 10 mL, 20 mL and 30 mL. It has been turned out that the antibody was formed most in 20 mL dosage. It was inclined to $12\%$ reduce caused by vaccine injection, but recovered after about maximum 1 week. In measurement of body temperature of cows after vaccine injected, it was inclined to rise with the normal scope in comparison with the controlled conditions.

• Journal of Microbiology and Biotechnology
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• v.10 no.6
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• pp.865-872
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• 2000

Helocobacter phylori is the major cause of gastritis, peptic ulcer, and a principal risk factor for gastric cancer. As the firs step towards a vaccine against H. pylori infection, Hy.pylori urease was expressed and purified as a recombinant apoenzyme (rUrease) in E. coli. In order to develop an effective immunization protocol using rUrease, the host immune responses were evaluated after the oral immunization of mice with rUrease preparations plus cholera toxin relative to various conditions, such as the physical nature of the antigen, the frequency of the booster immunization, the dose of the antigen, and the route of administration. The protective efficacy was assessed using a quantitative culture following an H. pylori SS1 challenge. It was demonstrated that rUrease, due to its particulated nature, was more superior than the UreB subunit as a vaccine antigen. The oral immunization of rUrease elicited significant systemic and secretory antibody responses, and activated predominantly Th2-type cellular responses. The bacterial colonization was significantly reduced (~100-fold) in those mice immunized with three or four weekly oran doses of rUrease plus cholera toxin (p<0.05), when compared to the non-immunized/challenged controls. The protection correlated well with the elicited secretory IgA level against rUrease, and these secretory antibody responses were highly dependent on the frequency of the booster immunization, yet unaffected by the dose of the antigen (25-200$\mu\textrm{g}$). These results demonstrate the remarkable potential of rUrease as a vaccine antigen, thereby strengthening the possibility of developing an H. pylori vaccine for humans.

• 한국생물공학회:학술대회논문집
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• 2003.10a
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• pp.186-189
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• 2003

Helicobacter pylori is the etiologic agent of human gastritis and peptic ulceration and produces urease as the major protein component on its surface. H. pylori urease is known to serve as a potent immunogen as well as major virulence factor. In order to express the recombinant urease in tobacco plants, a DNA fragment containing the minimal H. pylori urease gene cluster was subcloned into a plant expression vector. The recombinant vector was transformed to tobacco plants. The integration of the recombinant plasmids into tobacco chromosomal genome was verified by genomic PCR. Expression to mRNA was confirmed by Northern blot analysis, and expression to recombinant urease protein was observed by Western blot analysis. These results showed that the recombinant urease can be produced in tobacco plants and will be tested for immune response to use as an edible vaccine.

• Korean Journal of Plant Tissue Culture
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• v.27 no.3
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• pp.239-243
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• 2000

Helicobacter pylori, an etiologic agent of gastritis and peptic ulceration, produces urease which elicits a powerful immunoglobulin response in H. pylori-infected individuals. To establish a model plant vaccine agains H. pylori, 750 bp -ureA DNA amplified by polymerase chain reaction from pH 808 plasmid harboring urease gene cluster was cloned and manipulated to be expressed in tobacco plants. From the regenerated transgenic tobacco plants, ureA DNA integration,m its mRNA expression and protein synthesis were analyzed and confirmed by standard molecular techniques. The CaMV 35S promoter-driving ureA construct was expressed to produce a 30 kDa protein which was identical with bacterial UreA in size when detected on immunoblot of SDS polyacrylamide gel electrophoresis.

• Journal of the Korean Society for Industrial and Applied Mathematics
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• v.13 no.1
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• pp.1-11
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• 2009

Current therapies against ulcers caused by H. pylori infection consist of antibiotics, an acid reducer, and some clinical trials underway to develop a H. pylori vaccine. We develop a structured model with age-dependent mortality of peptic ulcers and H. pylori infection. Our main goal is to analyze our structured model mathematically and to compare it to our previously unstructured model to examine the disease transmission dynamics in terms of annual prevalence and annual incidence of the disease.

• Korean Journal of Veterinary Research
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• v.46 no.4
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• pp.327-335
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• 2006

A stable and reliable Helicobacter pylori (H. pylori) infection animal model would be necessary for evaluating vaccine efficacy and helpful for understanding the pathological mechanism of the organism. The aim of the present study is to investigate the effect of ethanol treatment prior to H. pylori inoculation on associated gastric mucosal injury and to establish ethanol-pretreating animal model to study H. pylori infection. Male Mongolian gerbils were used for the study. H. pylori was orally inoculated after 12 h fasting. 3 h prior to H. pylori inoculation, a group of gerbils was orally treated with absolute ethanol, 60% and 40% ethanol respectively. Another group of animals was treated either with H. pylori culture media alone or with different concentrations of ethanol plus culture media. Gerbils were killed 4 or 8 weeks after H. pylori inoculation. The colonization of H. pylori was confirmed by both histological examination and rapid urease test. Mucosal damage was evaluated grossly and histologically according to the criteria. The colonization of H. pylori and pathological changes in gastric mucosa of the animals were also observed. Although no significant change to the gastric mucose was observed in the animals treated either with H. pylori culture media alone or with different concentrations of ethanol plus culture media, persistent H. pylori infection was seen in the mucosa and mucosal leucocyte infiltration and severe epithelial damage was observed in the Helicobacter and ethanol + Helicobacter groups after 4 weeks. The gross and histological scores were higher in the ethanol + Helicobacter than in the Helicobacter alone group. As the results, ethanol-pretreatment with 60% concentration induced severe pathogenic changes by H. pylori infection in 5 weeks-old Mongolian gerbils. These results suggested that ethanol-pretreatment before H. pylori inoculation could increase the severity of gastric mucosal inflammation and enhance the colonization of H. pylori. The established ethanol-pretreating animal model would contribute to screen new drugs against H. pylori and be used as an useful tool for various animal experiments with H. pylori strains.

• Journal of Microbiology
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• v.45 no.1
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• pp.21-28
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• 2007

In order to search for specific genotypes related to this unique phenotype, we used whole genomic DNA microarray to characterize the genomic diversity of Helicobacter pylori (H. pylori) strains isolated from clinical patients in China. The open reading frame (ORF) fragments on our microarray were generated by PCR using gene-specific primers. Genomic DNA of H. pylori 26695 and J99 were used as templates. Thirty-four H. pylori isolates were obtained from patients in Shanghai. Results were judged based on In(x) transformed and normalized Cy3/Cy5 ratios. Our microarray included 1882 DNA fragments corresponding to 1636 ORFs of both sequenced H. pylori strains. Cluster analysis, revealed two diverse regions in the H. pylori genome that were not present in other isolates. Among the 1636 genes, 1091 (66.7%) were common to all H. pylori strains, representing the functional core of the genome. Most of the genes found in the H. pylori functional core were responsible for metabolism, cellular processes, transcription and biosynthesis of amino acids, functions that are essential to H. pylori's growth and colonization in its host. In contrast, 522 (31.9%) genes were strain-specific genes that were missing from at least one strain of H. pylori. Strain-specific genes primarily included restriction modification system components, transposase genes, hypothetical proteins and outer membrane proteins. These strain-specific genes may aid the bacteria under specific circumstances during their long-term infection in genetically diverse hosts. Our results suggest 34 H. pylori clinical strains have extensive genomic diversity. Core genes and strain-specific genes both play essential roles in H. pylori propagation and pathogenesis. Our microarray experiment may help select relatively significant genes for further research on the pathogenicity of H. pylori and development of a vaccine for H. pylori.

• Pediatric Infection and Vaccine
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• v.14 no.1
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• pp.97-103
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• 2007

Purpose : Lewis antigen has been known to have a role in the attachment of H. pylori to the gastric mucosa, but its expression pattern in children with H. pylori infection is still unclear. The recently described blood group antigen-binding adhesin BabA is known to mediate adherence of H. pylori to Lewis B receptors on gastric epithelium. We investigated the expression of Lewis antigen in gastric mucosa of Korean children with H. pylori infection. Methods : The expression of Lewis A ($Le^a$), B ($Le^b$), X ($Le^x$), and Y ($Le^y$) was evaluated by immunohistochemistry in H. pylori positive biopsy specimens from 35 children (antral gastritis in 30, peptic ulcer in 5) and in H. pylori negative specimens from 19 children. PCR assays for cagA and babA2 gene of H. pylori were performed. Results : We confirmed the expression of $Le^a$ in 60%, $Le^b$ in 97%, $Le^x$ in 100%, and $Le^y$ in 100% of the superficial epithelium of the 35 H. pylori positive children. In H. pylori negative patients, $Le^a$, $Le^b$, $Le^x$, and $Le^y$ expression was 52%, 100%, 89%, and 100%, respectively. The cagA gene was detected in 65% and babA2 gene in 25% of 35 patients. No differences in neutrophil activity and chronic inflammation were found according to the presence of cagA and babA2 genes in H. pylori. Conclusion : $Le^b$, $Le^x$ and $Le^y$ antigen were highly expressed in gastric mucosa of Korean children, but they were not associated with the status of H. pylori infection and the positivity of babA2 gene. Further studies for other mucosal receptors and toxins are needed to define the immune responses to H. pylori infection in gastric mucosa of Korean children.