Skip to main content

Evaluation of the effect of cagPAI genes of Helicobacter pylori on AGS epithelial cell morphology and IL-8 secretion

Antonie van Leeuwenhoek volume 105, pages 179–189 (2014)Cite this article

Abstract

Helicobacter pylori cagPAI genes play an important role in pathogenesis, however little is known about their functions in isolates from Turkish patients. We aimed to evaluate the intactness and the effect of the cagPAI genes (cagT, cagM, cagE, cagA) and cagA EPIYA motifs on the AGS morphological changes and IL-8 induction. Of 53 patients 38 were found infected with H. pylori. PCR amplification of the cagPAI genes showed 42.1 % intact, 39.5 % partially deleted and 18.4 % with complete deletions. Isolates from gastritis, duodenal and gastric ulcer patients with intact and partially deleted cagPAI genes induced higher IL-8 secretion than those with complete deletions. Isolates from gastritis patients had higher deletion frequencies of the cagT and cagM genes than the other two genes. Infection of AGS cells with isolates that possess intact cagPAI and EPIYA-ABC resulted in the formation of the hummingbird phenotype. The cagA positive isolates induced higher IL-8 secretion than cagA negative isolates. Isolates from DU patients with more than one EPIYA-C motif induced higher concentrations of IL-8 than those with EPIYA-ABC. In conclusion, the intactness of the cagPAI in our isolates from different patients was not conserved. An intact cagPAI was found to play an important role in the pathogenesis of DU but not GU or gastritis. The cagA gene, but not other cagPAI genes, was associated with the induction of IL-8 and the morphological changes of the AGS cells. An increase in the number of EPIYA-C motifs had noticeable effect on the formation of the hummingbird phenotype.

This is a preview of subscription content, access via your institution.

Access options

Buy single article

Instant access to the full article PDF.

43,34 €

Price includes VAT (Finland)
Tax calculation will be finalised during checkout.

Fig. 1
Fig. 2
Fig. 3

References

  • Alfizah H, Ramelah M (2012) Variant of Helicobacter pylori CagA proteins induce different magnitude of morphological changes in gastric epithelial cells. Malays J Pathol 34:29–34

    PubMed  Google Scholar 

  • Argent RH, Zhang Y, Atherton JC (2005) Simple method for determination of the number of Helicobacter pylori CagA variable-region EPIYA tyrosine phosphorylation motifs by PCR. J Clin Microbiol 43:791–795

    CAS  PubMed Central  PubMed  Article  Google Scholar 

  • Atherton JC, Blaser MJ (2009) Coadaptation of Helicobacter pylori and humans: ancient history, modern implications. J Clin Invest 119:2475–2487

    CAS  PubMed Central  PubMed  Article  Google Scholar 

  • Backert S, Schwarz T, Miehlke S, Kirsch C, Sommer C, Kwok T, Gerhard M, Goebel UB, Lehn N, Koenig W, Meyer TF (2004) Functional analysis of the cag pathogenicity island in Helicobacter pylori isolates from patients with gastritis, peptic ulcer, and gastric cancer. Infect Immun 72:1043–1056

    CAS  PubMed Central  PubMed  Article  Google Scholar 

  • Baghaei K, Shokrzadeh L, Jafari F, Dabiri H, Yamaoka Y, Bolfion M, Zojaji H, Aslani MM, Zali MR (2009) Determination of Helicobacter pylori virulence by analysis of the cag pathogenicity island isolated from Iranian patients. Dig Liver Dis 41:634–638

    CAS  PubMed Central  PubMed  Article  Google Scholar 

  • Bourzac KM, Botham CM, Guillemin K (2007) Helicobacter pylori CagA induces AGS cell elongation through a cell retraction defect that is independent of Cdc42, Rac1, and Arp2/3. Infect Immun 75:1203–1213

    CAS  PubMed Central  PubMed  Article  Google Scholar 

  • Brandt S, Kwok T, Hartig R, Konig W, Backert S (2005) NF-κB activation and potentiation of proinflammatory responses by the Helicobacter pylori CagA protein. Proc Natl Acad Sci USA 102:9300–9305

    CAS  PubMed  Article  Google Scholar 

  • Cendron L, Couturier M, Angelini A, Barison N, Stein M, Zanotti G (2009) The Helicobacter pylori CagD (HP0545, Cag24) protein is essential for CagA translocation and maximal induction of interleukin-8 secretion. J Mol Biol 386:204–217

    CAS  PubMed  Article  Google Scholar 

  • Censini S, Lange C, Xiang Z, Crabtree JE, Ghiara P, Borodovsky M, Rappuoli R, Covacci A (1996) cag, a pathogenicity island of Helicobacter pylori, encodes type I-specific and disease-associated virulence factors. Proc Natl Acad Sci USA 93:14648–14653

    CAS  PubMed  Article  Google Scholar 

  • Eaton KA, Kersulyte D, Mefford M, Danon SJ, Krakowka S, Berg DE (2001) Role of Helicobacter pylori cag region genes in colonization and gastritis in two animal models. Infect Immun 69:2902–2908

    CAS  PubMed Central  PubMed  Article  Google Scholar 

  • El-Etr SH, Mueller A, Tompkins LS, Falkow S, Merrell DS (2004) Phosphorylation-independent effects of CagA during interaction between Helicobacter pylori and T84 polarized monolayers. J Infect Dis 190:1516–1523

    CAS  PubMed  Article  Google Scholar 

  • Fischer W, Puls J, Buhrdorf R, Gebert B, Odenbreit S, Haas R (2001) Systematic mutagenesis of the Helicobacter pylori cag pathogenicity island: essential genes for CagA translocation in host cells and induction of interleukin-8. Mol Microbiol 42:1337–1348

    CAS  PubMed  Article  Google Scholar 

  • Guillemin K, Salama NR, Tompkins LS, Falkow S (2002) Cag pathogenicity island-specific responses of gastric epithelial cells to Helicobacter pylori infection. Proc Natl Acad Sci USA 99:15136–15141

    CAS  PubMed  Article  Google Scholar 

  • Ishikawa S, Ohta T, Hatakeyama M (2009) Stability of Helicobacter pylori CagA oncoprotein in human gastric epithelial cells. FEBS Lett 583:2414–2418

    CAS  PubMed  Article  Google Scholar 

  • Jafari F, Shokrzadeh L, Dabiri H, Baghaei K, Yamaoka Y, Zojaji H, Haghazali M, Molaei M, Zali MR (2008) vacA genotypes of Helicobacter pylori in relation to cagA status and clinical outcomes in Iranian populations. Jpn J Infect Dis 61:290–293

    CAS  PubMed Central  PubMed  Google Scholar 

  • Jones KR, Joo YM, Jang S, Yoo YJ, Lee HS, Chung IS, Olsen CH, Whitmire JM, Merrell DS, Cha JH (2009) Polymorphism in the CagA EPIYA motif impacts development of gastric cancer. J Clin Microbiol 47:959–968

    CAS  PubMed Central  PubMed  Article  Google Scholar 

  • Kauser F, Khan AA, Hussain MA, Carroll IM, Ahmad N, Tiwari S, Shouche Y, Das B, Alam M, Ali SM, Habibullah CM, Sierra R, Megraud F, Sechi LA, Ahmed N (2004) The cag pathogenicity island of Helicobacter pylori is disrupted in the majority of patient isolates from different human populations. J Clin Microbiol 42:5302–5308

    CAS  PubMed Central  PubMed  Article  Google Scholar 

  • Kawamura O, Murakami M, Araki O, Yamada T, Tomizawa S, Shimoyama Y, Minashi K, Maeda M, Kusano M, Mori M (2003) Relationship between gastric disease and deletion of cag pathogenicity island genes of Helicobacter pylori in gastric juice. Dig Dis Sci 48:47–53

    CAS  PubMed  Article  Google Scholar 

  • Kidd M, Lastovica AJ, Atherton JC, Louw JA (2001) Conservation of the cag pathogenicity island is associated with vacA alleles and gastroduodenal disease in South African Helicobacter pylori isolates. Gut 49:11–17

    CAS  PubMed  Article  Google Scholar 

  • Kim SY, Lee YC, Kim HK, Blaser MJ (2006) Helicobacter pylori CagA transfection of gastric epithelial cells induces interleukin-8. Cell Microbiol 8:97–106

    CAS  PubMed  Article  Google Scholar 

  • Kumar S, Kumar A, Dixit VK (2010) Diversity in the cag pathogenicity island of Helicobacter pylori isolates in populations from North and South India. J Med Microbiol 59:32–40

    CAS  PubMed  Article  Google Scholar 

  • Lage AP, Godfroid E, Fauconnier A, Burette A, Butzler JP, Bollen A, Glupczynski Y (1995) Diagnosis of Helicobacter pylori infection by PCR: comparison with other invasive techniques and detection of cagA gene in gastric biopsy specimens. J Clin Microbiol 33:2752–2756

    CAS  PubMed Central  PubMed  Google Scholar 

  • Lai CH, Wang HJ, Chang YC, Hsieh WC, Lin HJ, Tang CH, Sheu JJ, Lin CJ, Yang MS, Tseng SF, Wang WC (2011) Helicobacter pylori CagA-mediated IL-8 induction in gastric epithelial cells is cholesterol-dependent and requires the C-terminal tyrosine phosphorylation-containing domain. FEMS Microbiol Lett 323:155–163

    CAS  PubMed  Article  Google Scholar 

  • Lima VP, de Lima MA, Ferreira MV, Barros MA, Rabenhorst SH (2010) The relationship between Helicobacter pylori genes cagE and virB11 and gastric cancer. Int J Infect Dis 14:613–617

    Article  Google Scholar 

  • Lima VP, Silva-Fernandes IJ, Alves MK, Rabenhorst SH (2011) Prevalence of Helicobacter pylori genotypes (vacA, cagA, cagE and virB11) in gastric cancer in Brazilian’s patients: an association with histopathological parameters. Cancer Epidemiol 35:32–37

    Article  Google Scholar 

  • Maeda S, Yoshida H, Ikenoue T, Ogura K, Kanai F, Kato N, Shiratori Y, Omata M (1999) Structure of cag pathogenicity island in Japanese Helicobacter pylori isolates. Gut 44:336–341

    CAS  PubMed  Article  Google Scholar 

  • Matteo MJ, Granados G, Pérez CV, Olmos M, Sanchez C, Catalano M (2007) Helicobacter pylori cag pathogenicity island genotype diversity within the gastric niche of a single host. J Med Microbiol 56:664–669

    CAS  PubMed  Article  Google Scholar 

  • Mizushima T, Sugiyama T, Kobayashi T, Komatsu Y, Ishizuka J, Kato M, Asaka M (2002) Decreased adherence of cagG-deleted Helicobacter pylori to gastric epithelial cells in Japanese clinical isolates. Helicobacter 7:22–29

    CAS  PubMed  Article  Google Scholar 

  • Nguyen LT, Uchida T, Tsukamoto Y, Trinh TD, Ta L, Mai HB, Le HS, Ho DQ, Hoang HH, Matsuhisa T, Okimoto T, Kodama M, Murakami K, Fujioka T, Yamaoka Y, Moriyama M (2010) Clinical relevance of cagPAI intactness in Helicobacter pylori isolates from Vietnam. Eur J Clin Microbiol Infect Dis 29:651–660

    CAS  PubMed Central  PubMed  Article  Google Scholar 

  • Nilsson C, Sillén A, Eriksson L, Strand ML, Enroth H, Normark S, Falk P, Engstrand L (2003) Correlation between cag pathogenicity island composition and Helicobacter pylori-associated gastroduodenal disease. Infect Immun 71:6573–6581

    CAS  PubMed Central  PubMed  Article  Google Scholar 

  • Olbermann P, Josenhans C, Moodley Y, Uhr M, Stamer C, Vauterin M, Suerbaum S, Achtman M, Linz B (2010) A global overview of the genetic and functional diversity in the Helicobacter pylori cag pathogenicity island. PLoS Genet 6(8):e1001069

    PubMed Central  PubMed  Article  Google Scholar 

  • Pacheco AR, Proença-Módena JL, Sales AI, Fukuhara Y, da Silveira WD, Pimenta-Módena JL, de Oliveira RB, Brocchi M (2008) Involvement of the Helicobacter pylori plasticity region and cag pathogenicity island genes in the development of gastroduodenal diseases. Eur J Clin Microbiol Infect Dis 27:1053–1059

    CAS  PubMed  Article  Google Scholar 

  • Patra R, Chattopadhyay S, De R, Datta S, Chowdhury A, Ramamurthy T, Nair GB, Berg DE, Mukhopadhyay AK (2010) Intact cag pathogenicity island of Helicobacter pylori without disease association in Kolkata, India. Int J Med Microbiol 301:293–302

    PubMed Central  PubMed  Article  Google Scholar 

  • Peters TM, Owen RJ, Slater E, Varea R, Teare EL, Saverymuttu S (2001) Genetic diversity in the Helicobacter pylori cag pathogenicity island and effect on expression of anti-CagA serum antibody in UK patients with dyspepsia. J Clin Pathol 54:219–223

    CAS  PubMed  Article  Google Scholar 

  • Quiroga AJ, Huertas A, Cómbita AL, Bravo MM (2010) Variation in the number of EPIYA-C repeats in CagA protein from Colombian Helicobacter pylori strains and its ability middle to induce hummingbird phenotype in gastric epithelial cells. Biomedica 30:251–258

    PubMed  Google Scholar 

  • Reyes-Leon A, Atherton JC, Argent RH, Puente JL, Torres J (2007) Heterogeneity in the activity of Mexican Helicobacter pylori strains in gastric epithelial cells and its association with diversity in the cagA gene. Infect Immun 75:3445–3454

    CAS  PubMed Central  PubMed  Article  Google Scholar 

  • Ritter B, Kilian P, Reboll MR, Resch K, DiStefano JK, Frank R, Beil W, Nourbakhsh M (2011) Differential effects of multiplicity of infection on Helicobacter pylori-induced signaling pathways and interleukin-8 gene transcription. J Clin Immunol 31:60–68

    CAS  PubMed  Article  Google Scholar 

  • Salih BA, Bolek BK, Arikan S (2010) DNA sequence analysis of cagA 3′ motifs of Helicobacter pylori strains from patients with peptic ulcer diseases. J Med Microbiol 59:144–148

    CAS  PubMed  Article  Google Scholar 

  • Schmidt HM, Andres S, Nilsson C, Kovach Z, Kaakoush NO, Engstrand L, Goh KL, Fock KM, Forman D, Mitchell H (2010) The cag PAI is intact and functional but HP0521 varies significantly in Helicobacter pylori isolates from Malaysia and Singapore. Eur J Clin Microbiol Infect Dis 29:439–451

    CAS  PubMed  Article  Google Scholar 

  • Selbach M, Moese S, Meyer TF, Backert S (2002) Functional analysis of the Helicobacter pylori cag pathogenicity island reveals both VirD4-CagA-dependent and VirD4-CagA-independent mechanisms. Infect Immun 70:665–671

    CAS  PubMed Central  PubMed  Article  Google Scholar 

  • Sgouras DN, Panayotopoulou EG, Papadakos K, Martinez-Gonzalez B, Roumbani A, Panayiotou J, van Vliet-Constantinidou C, Mentis AF, Roma-Giannikou E (2009) cagA and vacA polymorphisms do not correlate with severity of histopathological lesions in Helicobacter pylori-infected Greek children. J Clin Microbiol 47:2426–2434

    CAS  PubMed Central  PubMed  Article  Google Scholar 

  • Yamahashi Y, Hatakeyama M (2013) PAR1b takes the stage in the morphogenetic and motogenetic activity of Helicobacter pylori CagA oncoprotein. Cell Adhes Migr 7:11–18

    Article  Google Scholar 

  • Zaidi SF, Ahmed K, Yamamoto T, Kondo T, Usmanghani K, Kadowaki M, Sugiyama T (2009) Effect of resveratrol on Helicobacter pylori-induced interleukin-8 secretion, reactive oxygen species generation and morphological changes in human gastric epithelial cells. Biol Pharm Bull 32:1931–1935

    CAS  PubMed  Article  Google Scholar 

Download references

Acknowledgments

This study was supported by the grants of Fatih University (No: P50030903_2) and of the Scientific and Technological Research Council of Turkey (TÜBİTAK) (No: 111T370).

Conflict of interest

The authors declare that they have no conflict of interest.

Author information

Affiliations

  1. Department of Biology, Faculty of Science and Literature, Fatih University, B. Cekmece, Istanbul, Turkey

    Barik A. Salih, Ahmet Guner, Ahu Karademir, Merve Uslu, Mehmet Akif Ovali, Duygu Yazici & Bora Kazim Bolek

  2. Department of Surgery, Istanbul Teaching and Research Hospital, Istanbul, Turkey

    Soykan Arikan

Authors
  1. Barik A. Salih
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Ahmet Guner
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Ahu Karademir
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Merve Uslu
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Mehmet Akif Ovali
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Duygu Yazici
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Bora Kazim Bolek
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. Soykan Arikan
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Barik A. Salih.

Rights and permissions

Reprints and Permissions

About this article

Cite this article

Salih, B.A., Guner, A., Karademir, A. et al. Evaluation of the effect of cagPAI genes of Helicobacter pylori on AGS epithelial cell morphology and IL-8 secretion. Antonie van Leeuwenhoek 105, 179–189 (2014). https://doi.org/10.1007/s10482-013-0064-5

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s10482-013-0064-5

Keywords

  • Helicobacter pylori
  • cagPAI
  • cagA
  • AGS
  • IL-8
  • EPIYA motifs