Document Type : Original Article
Helicobacter pylori Research Group, Biotechnology Research Center, Pasteur Institute of Iran, Tehran, Iran
National Cell Bank of Iran, Pasteur Institute of Iran, Tehran, Iran
Department of Pathobiology, School of Public Health, Tehran University of Medical Sciences, Tehran, Iran
Cancer Research Center, Tehran University of Medical Sciences, Tehran, Iran
Objective: Helicobacter pylori (H. pylori) cytotoxin and its heterogeneity amongst strains has been closely linked to the varying infection-associated clinical outcomes. In order to determine the decisive role of the vacuolating cytotoxin (vacA) gene mosaicism in its corresponding gene expression and phenotype, we aimed to characterize vacA alleles of different H. pylori strains in addition to the resulting protein and its vacuolating activity in epithelial cell culture.
Materials and Methods: vacA gene polymorphism was determined for 80 H. pylori strains isolated from dyspeptic patients, using multiplex gene-specific polymerase chain reaction (PCR). VacA protein was detected by immuno-blotting assay using a polyclonal anti-VacA antibody. In vitro cytotoxicity assay was conducted on HeLa cells in order to evaluate the vacuolating cytotoxin activity.
Results: Genotyping revealed the following strain distribution: 26 (32.5%) s1m1, 35 (43.8%) s1m2, and 19 (23.8%) s2m2 subtypes. Infection with s1m1 type strain was significantly associated with gastric cancer as compared to non-ulcer dyspepsia (p=0.005) and peptic ulcer disease (p=0.008). A 95-kDa immuno-reactive band that represented the vacuolating toxin was demonstrated in SDS-PAGE analysis of concentrated culture filtrate (CCF) of H. pylori strains. H. pylori CCFs induced HeLa cell vacuolation which correlated with the strain genotype; s1m1 strains demonstrated higher levels of vacuolation as compared to s1m2 strains, whereas s2m2 strains showed no detectable cytotoxic activity.
Conclusion: The current study confirmed the relatively high cytotoxic activity of s1m1 type H. pylori strains which infect the majority of patients suffering from gastric cancer and may be partly responsible for the pathogenesis of this mortal disease.