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Background and Aims: Helicobacter pylori is a highly diverse pathogen, which encounters epithelial cells as the initial defense barrier during its lifelong infection. The structure of epithelial cells can be disrupted through cleavage of microfilaments. Cytokeratin 18 (CK18) is an intermediate filament, the cleavage of which is considered an early event during apoptosis following activation of effector caspases., Methods: Helicobacter pylori strains were isolated from 76 dyspeptic patients. cagA 3' variable region and CagA protein status were analyzed by PCR and western blotting, respectively. Eight hours post-co-culture of AGS cells with different H. pylori strains, flow cytometric analysis was performed using M30 monoclonal antibody specific to CK18 cleavage-induced neo-epitope., Results: Higher rates of CK18 cleavage were detected during co-culture of AGS cells with H. pylori strains bearing greater numbers of cagA EPIYA-C and multimerization (CM) motifs. On the other hand, H. pylori strains with greater numbers of EPIYA-B relative to EPIYA-C demonstrated a decrease in CK18 cleavage rate. Thus, H. pylori-mediated cleavage of CK18 appeared proportional to the number of CagA EPIYA-C and CM motifs, which seemed to be downplayed in the presence of EPIYA-B motifs., Conclusions: Our observation associating the heterogeneity of cagA variants with the potential of H. pylori strains in the induction of CK18 cleavage as an early indication of apoptosis in gastric epithelial cells supports the fact that apoptosis may be a type-specific trait. However, additional cagA-targeted experiments are required to clearly identify the role of EPIYA and CM motifs in apoptosis and/or the responsible effector molecules., (© 2012 Blackwell Publishing Ltd.)

Helicobacter pylori, a microaerophilic fastidious bacterium, has been cultured on various plating and broth media since its discovery. Although the agar media can be sufficient for the identification, typing, and antibiotic resistance studies, no secretory antigen of H. pylori can be evaluated in such media. Thus, satisfactory growth of H. pylori in liquid culture which is needed for analysis of secretory proteins without the presence of interfering agents is in demand. We assessed the impact of beta-cyclodextrin, Fetal Bovine Serum (FBS), and charcoal as supplements for H. pylori growth. Furthermore, we aimed to identify the most favorable supplement that supports the secretion of the dominant secretory protein, vacuolating cytotoxin (VacA). Five clinical strains were cultured on broth media and the growth, viability, morphology, and protein content of each strain were determined. Our results revealed that beta-cyclodextrin supports the growth rate, viability, and cell lysate protein content to the extent similar to FBS. Application of beta-cyclodextrin is found to postpone spiral to coccoid conversion up to 72 h of incubation. Although FBS supports a higher VacA protein content, presence of interfering macromolecules in FBS questions its utility particularly for purposes of studying extra cellular proteins such as VacA. This study recommends further application of beta-cyclodextrin as a culture supplement with the potential capacity in neutralizing toxic compounds and flourishing the secretion of H. pylori proteins without addition of interfering proteins.

Background/aim: The identification of the vacA intermediate region has provided new insights into the role of vacA heterogeneity in relation to gastro-duodenal pathogenesis. The aim of this study was to assess vacA polymorphism in Iranian Helicobacter pylori strains and its association with cagA as a major virulence determinant, gastric histopathology and disease., Methods: vacA polymorphism and serum antibody responses were studied in 207 H. pylori-infected (139 NUD, 34 PUD, and 34 GC) patients and correlated with gastric histopathology., Results: Multivariate logistic regression analysis found intermediate region typing superior to signal or mid region typing for screening high risk patients. vacA i1 allele was identified as an independent predictor of dysplasia (OR = 9.044; 95% CI: 1.11-73.33). Possession of s1/i1/cagA(+) strains was also identified as a predictor of intestinal metaplasia (OR = 3; 95% CI: 1.13-7.95), dysplasia (OR = 9.9; 95% CI: 1.23-80.86) and risk of GC (OR = 6.9; 95% CI: 2.5-18.66) as well as induction of anti-VacA sero-positivity (OR = 5.04; 95% CI: 1.8-13.6). Anti-VacA serology correctly detected 83.8% of s1/i1/cagA(+) strains carried by high-risk patients., Conclusions: The current study emphasizes the implication of vacA polymorphic structure, especially the s1/i1/cagA(+) genotype, in increasing the risk of GC by revealing their association with gastric pre-neoplastic changes and their reflection in VacA sero-positivity which encourages the application of noninvasive procedures in population screening., (Copyright 2009 S. Karger AG, Basel.)