Phylogeographic origin of Helicobacter pylori determines host-adaptive responses upon coculture with gastric epithelial cells

Abstract

While Helicobacter pylori infects over 50% of the world's population, the mechanisms involved in the development of gastric disease are not fully understood. Bacterial, host, and environmental factors play a role in disease outcome. To investigate the role of bacterial factors in H. pylori pathogenesis, global gene expression of six H. pylori isolates was analyzed during coculture with gastric epithelial cells. Clustering analysis of six Colombian clinical isolates from a region with low gastric cancer risk and a region with high gastric cancer risk segregated strains based on their phylogeographic origin. One hundred forty-six genes had increased expression in European strains, while 350 genes had increased expression in African strains. Differential expression was observed in genes associated with motility, pathogenicity, and other adaptations to the host environment. European strains had greater expression of the virulence factors cagA, vacA, and babB and were associated with increased gastric histologic lesions in patients. In AGS cells, European strains promoted significantly higher interleukin-8 (IL-8) expression than did African strains. African strains significantly induced apoptosis, whereas only one European strain significantly induced apoptosis. Our data suggest that gene expression profiles of clinical isolates can discriminate strains by phylogeographic origin and that these profiles are associated with changes in expression of the proinflammatory and protumorigenic cytokine IL-8 and levels of apoptosis in host epithelial cells. These findings support the hypothesis that bacterial factors determined by the phylogeographic origin of H. pylori strains may promote increased gastric disease.

Fig 1

Clustering analysis of 6 Colombian isolates. Microarray data from three strains from a low gastric cancer risk area (PZ5004, PZ5024, and PZ5026) and three strains from a high gastric cancer risk area (PZ5056, PZ5080, and PZ5086) were assessed in triplicate. The dendrogram's edges are represented by numbers, and letters denote the P values for the edge (au/bp). Approximately unbiased P value (au) is based on multiscale bootstrap resampling, and bootstrap probability P value (bp) is based on normal bootstrap sampling. a, au & bp P value, <0.001; b, au & bp P value, <0.01; c, au & bp P value, <0.1.

Fig 2

Functional analysis of genes with increased expression by strain origin: 89 of 146 genes with increased expression in European strains (gray) and 163 of 350 genes with increased expression in African strains (black) had assigned functions.

Fig 3

Histopathology in patients from which PZ5004 (A, B) (nonatrophic gastritis), PZ5056 (C, D) (intestinal metaplasia), and PZ5086 (E, F) (intestinal metaplasia) were isolated. PZ5004 shows diffuse mononuclear leukocytic infiltration throughout and well-preserved glands (arrow). PZ5056 and PZ5086 both show irregular Goblet cells (arrow). Images are at magnifications of ×100 (left) and ×200 (right). Images courtesy of Maria Blanca Piazuelo.

Fig 4

Levels of IL-8 mRNA expression (A) and apoptosis (B) in AGS cells stimulated with H. pylori clinical isolates. Cells were cocultured with H. pylori strains at a multiplicity of infection of 200 for 6 h (A) and for 24 h (B). *, P < 0.05; **, P < 0.01; ***, P < 0.001 versus unstimulated control cells; §, P < 0.05; §§§, P < 0.001 compared to both low-risk African strains (PZ5004 and PZ5024); #, P < 0.05 versus African strain PZ5004.