Helicobacter pylori outer membrane proteins and gastric inflammation

Approximately 4% of the Helicobacter pylori genome, significantly more than that of any other known bacterial species, is composed of genes encoding outer membrane proteins (OMPs). The best described H pylori OMP is the adhesin BabA, which is thought to mediate host/bacterial interactions and has been linked to microbial pathogenicity.¹,²,³,⁴ Dissection of OMP function helps our understanding of the complex gastric biology of infection and may offer a possibility of identifying those with an increased risk of disease development.

In an interesting and comprehensive study, Yamaoka et al (Gut 2005;55:775–81) analysed expression of several outer membrane proteins (BabA, BabB, SabA, OipA) in a large number of H pylori strains and described their association with gastroduodenal diseases. One important finding was the strong linkage of OipA expression with the cag pathogenicity island (cagPAI), a major H pylori virulence factor. The authors showed that in 94% of Columbian and US H pylori strains, CagA and OipA phenotypes were identical.

OipA expression is regulated by slipped strand mispairing based on the number of CT dinucleotide repeats in the 5′ region of the gene. We sequenced this genetic region of H pylori strains from 53 German patients with chronic gastritis and further determined the presence of the cagPAI marker gene cagA, as described previously.³,⁵ Similarly to Yamaoka et al, we found linked expression of CagA and OipA in our European strain population, with 83% concordance (fig 1A). The reason for this strong association is not understood. However, as there is no obvious common genetic mechanism controlling expression of the two genes, linkage of cagA and oipA seems to be based on selection in the host. Indeed, it has been demonstrated that H pylori is capable of regulating OMP expression in vivo by phase variation as a mechanism of adaptation to changing environmental conditions.² It is thus conceivable that OipA offers a selective advantage for cagPAI positive bacteria that may for example prefer a gastric microenvironment in which OipA expression is beneficial.

Figure 1 Influence of OipA and the cag pathogenicity island (cagPAI) on interleukin 8 (IL‐8) expression in vitro and in vivo. (A) Genetic characterisation of Helicobacter pylori strains from 53 patients according to cagA and oipA status. The cagA gene was detected by polymerase chain reaction, as described previously.³oipA status was determined after sequencing of the 5′ region of the gene. The “on” or “off” status depends on the number of CT dinucleotide repeats that render the gene in or out of frame. In 83% of strains, cagA and oipA status was identical. (B) IL‐8 expression in the gastric cancer cell line KATO‐III after incubation with the H pylori strain B128 and its isogenic oipA and cagE mutant strains at an MOI of 50. Bacterial mutagenesis was performed as described previously.¹⁰ Cytokine expression was analysed by real time reverse transcription‐polymerase chain reaction and normalised to glyceraldeyde‐3‐phosphate dehydrogenase (GAPDH) expression, as described previously.³ Bars indicate the arithmetic mean (SEM) from five samples per group. Shown is one of 10 similar experiments. (C) IL‐8 expression in the gastric mucosa of 53 patients infected with the H pylori strain types shown in (A). Statistically significant differences were found between any cagA⁻ and cagA⁺ group (p<0.05, Student's t test). In contrast, there was no significant independent association of oipA‐on status with increased IL‐8 expression.

The influence of OipA on proinflammatory signalling is under debate.⁵,⁶,⁷,⁸,⁹ To investigate whether OipA is directly linked to pathogenesis, we performed mutagenesis of the oipA and cagE genes for functional investigations in vitro. As shown in fig 1B, incubation of the cagA⁺/oipA⁺ strain B128 with KATO‐III gastric epithelial cells caused increased expression of interleukin 8 (IL‐8), a signature chemokine of gastric inflammation. Whereas the isogenic oipA knockout strain (ΔoipA) induces IL‐8 expression to a similar extent, the ΔcagE strain, which lacks a functional cagPAI, is severely impaired in its IL‐8 inducing capacity. Despite using varying concentrations of bacteria (MOI 1–100), different cell lines, and several independent oipA mutants of the H pylori strains B128 and G27, we could not detect an influence of OipA on IL‐8 expression/secretion (not shown). Similarly, we found no independent influence of OipA on gastric IL‐8 expression or inflammation in vivo. As shown in fig 1C, IL‐8 expression was higher in the mucosa of patients infected with cagA positive strains than in patients infected with cagA negative strains. However, in both groups chemokine expression was independent of oipA status.

Bacterial OMPs have multiple functions (for example, as adhesins, porins, or in complement resistance and immune regulation). Although several studies have investigated the physiological role of OipA,⁵,⁶,⁷,⁸,⁹ its precise function is still not clear. Strong linkage with cagPAI however suggests that differential regulation of OipA expression by phase variation may contribute to the fitness of cagPAI positive or negative strains in vivo. This is supported by the finding that oipA mutated cagPAI positive H pylori strains are impaired in their ability to colonise Mongolian gerbils.⁹

Acknowledgements

AD is a recipient of a grant from the DAAD (Deutscher Akademischer Austauschdienst). RR is a recipient of a grant from Bund der Freunde der Technischen Universität München.