1. Top of page
  2. Abstract
  8. Acknowledgements
  9. References

Helicobacter pylori causes persistent inflammation in the human stomach, yet only a minority of persons harbouring this organism develop peptic ulcer disease or gastric malignancy. An important question is why such variation exists among colonized individuals. Recent evidence has demonstrated that H. pylori isolates possess substantial phenotypic and genotypic diversity, which may engender differential host inflammatory responses that influence clinical outcome. For example, H. pylori strains that possess the cag pathogenicity island induce more severe gastritis and augment the risk for developing peptic ulcer disease and distal gastric cancer. An alternative, but not exclusive, hypothesis is that enhanced inflammation and injury is a consequence of an inappropriate host immune response to the chronic presence of H. pylori within the gastric niche. Investigations that precisely delineate the mechanisms responsible for induction of gastritis will ultimately help to define which H. pylori-colonized persons bear the highest risk for subsequent development of clinical disease, and thus, enable physicians to focus eradication therapy.


  1. Top of page
  2. Abstract
  8. Acknowledgements
  9. References

The isolation of curved bacilli from patients with chronic gastritis nearly two decades ago sparked revolutions in the fields of gastroenterology, microbiology and molecular biology.1 Based on unique ultrastructural and genetic characteristics, these organisms are now classified as Helicobacter pylori.2H. pylori colonizes the stomachs of humans and primates for years, rather than days or weeks as might be expected for gastrointestinal pathogens. However, unlike other organisms (i.e. Mycobacterium tuberculosis) which can persist in infected hosts for decades but exist mainly in a latent or dormant state, H. pylori causes continuous inflammation.3 Chronic gastritis induced by H. pylori increases the risk for a wide spectrum of clinical outcomes, ranging from peptic ulcer disease to distal gastric adenocarcinoma and gastric mucosal lymphoproliferative diseases such as non-Hodgkin’s lymphoma (Figure 1).4–7 Gastric inflammation nearly always precedes the development of peptic ulceration, and is a critical component in initiating the multi-step progression towards gastric carcinogenesis (Figure 2).8 Similarly, although less is known regarding the specific factors crucial to gastric lymphoma development, the ongoing local chronic inflammatory process induced by H. pylori is likely to be of major importance.


Figure 1.  Relationship of H. pylori-induced gastric inflammation with variable disease outcomes.

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Figure 2.  Sequential histological events in the progression to gastric adenocarcinoma.

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Virtually all persons carrying H. pylori have coexisting gastric inflammation. However, only a small percentage of colonized individuals develop clinically apparent sequelae. Enhanced risk may be related to differences in the expression of specific bacterial products, to variations in the host inflammatory response to the bacteria, or to specific interactions between host and microbe. Recent evidence has also indicated that in some populations, H. pylori-induced chronic gastritis, particularly when localized to the acid-secreting corpus, is associated with a reduced risk for developing gastro-oesophageal reflux disease, Barrett’s oesophagus, and Barrett’s-associated oesophageal adenocarcinoma (Figure 1).9–14 However, these reciprocal relationships have not been universally observed, findings that underscore the importance of understanding how H. pylori induces gastritis.15–18 Herein, this review will focus on specific mechanisms by which H. pylori colonization leads to gastric inflammation and injury.


  1. Top of page
  2. Abstract
  8. Acknowledgements
  9. References

Gastric acidity and peristalsis normally inhibit bacterial colonization of the human stomach. However, natural selection has provided H. pylori with several mechanisms to elude these primary defences and establish persistent infection, such as the ability to withstand acidic gastric pH and motility (Table 1). Acute ingestion of H. pylori in humans leads to transient hypochlorhydria, which may develop in response to gastric inflammation.19, 20 In untreated hosts, inhibition of acid secretion resolves within several months and intraluminal pH decreases to within the normal range.20 In some individuals, acid production continues to increase within the context of chronic gastritis, which is likely to result from the compensatory rise in serum gastrin and fall in mucosal somatostatin levels induced by gastric inflammation.21–23

Table 1. H. pylori constituents that may permit gastric colonization Thumbnail image of


Another pH-altering mechanism is the production of urease, a nickel-containing hexadimer composed of two different subunits (60 and 27 kDa).24–26 There are seven genes in the H. pylori urease gene cluster: ureA and ureB encode the structural subunits of urease, while ureE, ureF, ureG and ureH encode accessory proteins necessary for assembly and Ni2+ insertion required to form active urease.27 The majority of this enzyme is cytoplasmic, although a fraction is present on the bacterial surface after prolonged in vitro growth.28 Urease activity increases 10- to 20-fold as the pH falls from 6.0 to 5.0, and thereafter remains constant down to a pH of 2.5.29, 30 Acid activation of cytoplasmic urease is mediated by expression of the third gene in the urease gene cluster, ureI, which encodes a H+ gated urea channel. UreI increases the permeability of the bacterial membrane to urea by at least 300-fold as the pH of the surrounding medium becomes acidic, and the presence of this acid-activated urea channel within the H. pylori inner membrane is necessary for efficient utilization of urea present in gastric juice.31 These data explain the absolute requirement for both urease and UreI for survival of H. pylori at a medium pH of less than 4.0 as well as for successful colonization of animal models.32, 33 Urease enzymatic activity is conserved among all known Helicobacter species and the primary structure of urease shows little divergence among H. pylori strains;34 these characteristics are consistent with the hypothesis that urease is a necessary factor for the establishment of chronic infection.

H. pylori constituents required for motility and adherence

To facilitate locomotion within gastric mucus and to counteract peristalsis, H. pylori possesses five or six polar flagella consisting of two structural subunits: a major 53-kDa FlaA and a 54-kDa FlaB.35, 36 The genes encoding these two flagellins are located at distant sites on the H. pylori chromosome and are transcriptionally regulated by different promoters.35 The essential role for both flaA and flaB in the establishment of persistent colonization has been previously demonstrated by Eaton et al., who showed that aflagellate H. pylori strains only transiently colonized gnotobiotic piglets.37 Thus, similar to urease production, motility is necessary for the maintenance of gastric colonization (Table 1).

The vast majority of H. pylori in colonized hosts are free-living, but approximately 20% bind to gastric epithelial cells.38 Adherence properties are likely to be important for the acquisition of nutrients from the host and/or for resistance to shedding of the mucus gel layer. Adhesion by H. pylori to gastric epithelium is highly specific in vivo and when H. pylori is found in the duodenum, it only overlays islands of gastric metaplasia and not intestinal-type cells.39 Several H. pylori and host ligands involved in adherence have been described. These include a 20-kDa H. pylori haemagglutinin encoded by hpa that binds to sialic acid-containing components of erythrocyte membranes and a 63-kDa exoenzyme S-like protein that binds phosphatidylethanolamine and gangliotetraosylceramide in vitro.40–43H. pylori can also bind to non-sialylated ligands which include laminin, fibronectin, various collagens, heparin sulphate, and sulphatide.44–48

Additional H. pylori ligands include members of a family of well-conserved outer membrane proteins that share extensive sequence homology in the N- and C-terminal domains, an example of which is BabA, encoded by the gene babA2. BabA is a membrane-bound adhesin that binds the blood-group antigen Lewisb present on gastric epithelial cell membranes, yet unlike urease, only a subset of H. pylori strains possess babA2.49–52 A second gene, designated babA1, is essentially identical to babA2 except for the lack of a 10 base-pair insertion sequence that leads to expression of a truncated form of BabA, which cannot bind Lewisb. H. pylori babA2+ strains are associated with an increased risk for both duodenal ulceration and gastric adenocarcinoma, whereas strains that lack babA2 are more frequently associated with gastritis alone.49 The presence of babA2 is associated with the presence of additional H. pylori strain-specific, disease-related genes, such as vacA s1 and cagA; strains that possess all three of these genes together (babA2+, vacA s1, cagA+) further augment the risk for peptic ulcer disease and gastric cancer.49 These clinical relationships have been supported by experiments in an animal model of H. pylori-induced inflammation: transgenic mice that express Lewisb on surfaces of epithelial cells located within the gastric pit. Lewisb-expressing mice challenged with babA2+H. pylori strains develop similar bacterial colonization density levels when compared with infected wild-type mice. However, the pattern of bacterial distribution between the two populations is distinct.53 In wild-type mice, H. pylori are located only within the mucous gel layer, whereas in transgenic mice bacteria are also found to be adherent to epithelial cells.53 Further, the severity of gastric inflammation is significantly increased within infected transgenic mice compared to corresponding wild-type littermates, emphasizing a pivotal role for H. pylori adherence in the induction of gastritis.

Another adherence-related H. pylori product is HopZ (encoded by HP0009 and JHP0007 in sequenced strains 26695 and J99).54 Inactivation of the gene encoding this protein dramatically decreases the ability of H. pylori to bind gastric epithelial cells in vitro.54 Similarly, inactivation of genes encoding AlpA and AlpB attenuates binding of H. pylori to epithelial cells, indicating that these are also important adhesins.55 In contrast to the strain-specificity of babA2, the genes encoding HopZ, AlpA, and AlpB are present in most, if not all, H. pylori isolates, suggesting that they may have important roles in colonization.54, 55

H. pylori expression of Lewis antigens

The O-antigen of H. pylori LPS contains different human Lewis antigens, including Lewisx, Lewisy, Lewisa, and Lewisb; inactivation of the bacterial genes encoding Lewisx and Lewisy results in an inability of H. pylori to colonize mice.56–60 Because Lewis antigens are also expressed on gastric epithelial cell surfaces, a potential biological role for bacterial Lewis antigens is molecular mimicry (Table 1).61 This could allow H. pylori to escape host immunological defence mechanisms by preventing the formation of antibodies directed against shared bacterial and host epitopes, thus facilitating persistent infection. Initially, human and animal studies supported this paradigm by demonstrating a significant concordance between the Lewis phenotype of the host and infecting Helicobacter strain.62–65 However, two recent studies have challenged these earlier observations by failing to show a significant relationship between the human host Lewis phenotype and that of colonizing H. pylori isolates.66, 67 Moreover, strains that express both Lewisx and Lewisy can be isolated from a single human host.68 Thus, the role for H. pylori Lewis antigens in evasion of host responses has not been established conclusively.

Another potential consequence of H. pylori Lewis expression is induction of a humoral response that enhances gastric inflammation and injury.61H. pylori infection in mice induces anti-Lewisx antibodies with an affinity for parietal cells, and immunization of mice leads to the development of anti-Lewisx/y antibodies that cross-react with host Lewisx/y antigens on the H+,K+-ATPase located in parietal cell canaliculi.53, 69 In humans, H. pylori infection similarly induces auto-antibodies directed against the canaliculi of gastric parietal cells.70, 71 However, recent data have demonstrated that anti-Lewisx/y antibodies are not associated with H. pylori infection per se, but instead, occur naturally in sera from uninfected persons.72, 73 Thus, similar to host evasion, an unequivocal role for H. pylori Lewis antigens in the induction of pathogenic auto-antibodies has not been substantiated.

Rather than contributing to host evasion or autoimmunity, recent data have suggested that H. pylori Lewis antigens may actually mediate adhesion (Table 1). In vitro, pre-incubation of H. pylori with anti-Lewisx monoclonal antibodies inhibits bacterial binding to gastric epithelial cells.74, 75H. pylori strains that strongly express Lewisx/y are associated with enhanced neutrophilic infiltration and higher colonization densities than strains that only weakly express Lewisx/y.66 Further, H. pylori Lewis antigens undergo phase variation (i.e. random and reversible high-frequency alteration of phenotype);76–80 therefore, a functional role for changes in Lewis phenotype may be detachment of H. pylori from host epithelial cells, which could subsequently facilitate transfer to a new host.61


  1. Top of page
  2. Abstract
  8. Acknowledgements
  9. References

The gastric inflammatory response induced by H. pylori consists of neutrophils, lymphocytes (T and B cells), plasma cells, and macrophages, along with varying degrees of epithelial cell degeneration and injury.81 Invasion of the gastric mucosa is rarely if at all identified in vivo, and therefore other potential mechanisms for induction of inflammation must be postulated. One possibility is that H. pylori secretes substances that stimulate mucosal inflammation from afar. For example, urease has been detected within the lamina propria and the urease complex of H. pylori stimulates chemotaxis by both monocytes and neutrophils and activates mononuclear cells as well.82 Similarly, H. pylori porins and low-molecular-weight molecules, such as Paf-acether, possess chemotactic properties.83, 84 Finally, H. pylori water extracts promote neutrophil–endothelial cell interactions in vitro and increase leucocyte adherance via CD11a/CD18 and CD11b/CD18 interactions with intercellular adhesion molecule type-1 (ICAM-1).85

Contact-mediated induction of cytokine release and development of acute inflammation

Another mechanism by which H. pylori may induce inflammation is through direct contact with gastric epithelial cells and stimulation of cytokine release. Gastric epithelium from H. pylori-infected persons demonstrates enhanced levels of interleukin-1β (IL-1β), IL-2, IL-6, IL-8 and TNF-α.86–90 This model is consistent with previous data demonstrating that interaction of Salmonella typhimurium with epithelial cells results in chemotactic cytokine (chemokine)-mediated signalling that directs polymorphonuclear cell migration towards the epithelial cell surface.91, 92 Interleukin-8 (IL-8) is a potent neutrophil-activating chemokine and expression of this peptide is localized to gastric epithelial cells in vivo.93 As might be predicted, levels of IL-8 are also directly related to the severity of gastritis.89In vitro, H. pylori stimulates IL-8 release from gastric epithelial cells and these events require an active interplay between live bacteria and epithelial cells.94–96 Thus, a paradigm for the acute component of H. pylori-induced gastric inflammation is that contact between bacteria and epithelial cells stimulates IL-8 secretion, which then regulates neutrophilic infiltration into the gastric mucosa.

The human IL-8 gene contains several binding sites within its promoter region, including a NF-κB binding motif and a binding site for c-fos and c-jun, which together comprise the transcription factor AP-1.97 NF-κB is a transcription factor sequestered in the cytoplasm, whose activation and regulation is tightly controlled by a class of inhibitory proteins termed IκB’s that mask the nuclear localization signals of NF-κB, thereby preventing movement of NF-κB to the nucleus. Upon stimulation, phosphorylation of IκB leads to its ubiquitination and proteosome-mediated degradation, thereby liberating NF-κB to enter the nucleus where it regulates transcription of a variety of genes.98 Stimulation and activation of NF-κB does not require protein synthesis, thereby allowing efficient activation of target genes, such as IL-8. This system is particularly utilized in immune and inflammatory responses where rapid activation of defense genes following exposure to pathogens such as bacteria is critical for survival of an organism. Several studies have demonstrated that contact between H. pylori and gastric epithelial cells results in brisk activation of NF-κB, which is followed by increased IL-8 expression.99–101 The ability of H. pylori to activate NF-κB in vitro has also been corroborated in vivo as activated NF-κB is present within gastric epithelial cells of infected, but not uninfected patients, which mirrors the location of increased IL-8 protein within colonized mucosa.93, 99

In addition to NF-κB, mitogen-activated protein kinases (MAPK) have been implicated as mediators of H. pylori-induced IL-8 expression. Mitogen-activated protein kinases cascades are signal transduction networks that target transcription factors and thus participate in a diverse array of cellular functions including cytokine production.102–104 The cascades are organized in three-kinase tiers consisting of a mitogen-activated protein kinases, a MAPK kinase (MKK), and a MKK kinase (MKKK). Transmission of signals occurs by sequential phosphorylation and activation of the components specific to a respective cascade. In mammalian systems, five mitogen-activated protein kinases modules have been identified and characterized to date; these include extracellular signal-regulated kinase 1 and 2 (ERK 1/2), p38, and c-Jun N-terminal kinase (JNK).102–104 In addition to regulating NF-κB, mitogen-activated protein kinases can activate other transcription factors, such as AP-1, that regulate cytokine gene expression. H. pylori rapidly induces activation of ERK, p38, and JNK mitogen-activated protein kinases in cell culture systems, and similar to NF-κB activation, these effects are dependent upon the presence of live bacteria, raising the question as to whether H. pylori-induced IL-8 production is dependent upon activation of NF-κB, mitogen-activated protein kinases, or both.105–107In vitro studies utilizing IL-8 reporter constructs have now revealed that H. pylori-induced IL-8 gene expression is dependent upon activation of both NF-κB and AP-1 (via mitogen-activated protein kinases activation), indicating that synergistic interactions between AP-1 and NF-κB within the IL-8 promoter are required for maximal H. pylori-induced IL-8 production.97

Another mechanism through which H. pylori may regulate phagocyte activation is via production of H. pylori neutrophil-activating protein. H. pylori neutrophil-activating protein is a 150-kDa dodecameric iron-binding protein originally identified as a component of the H. pylori outer membrane that promotes adhesion of neutrophils to endothelial cells.108, 109H. pylori neutrophil-activating protein is immunogenic in humans and mice, and stimulates phagocyte chemotaxis, NADPH oxidase assembly, and production of reactive oxygen species in vitro.110 All H. pylori strains analysed to date possess the gene encoding H. pylori neutrophil-activating protein (napA), although protein expression levels are variable.108 However, the relationship between H. pylori neutrophil-activating protein expression and clinical outcome, as well as the effects of isogenic disruption of napA on pathogenic responses, remain to be determined.

Humoral responses

H. pylori colonization induces an exuberant systemic and mucosal humoral response directed at multiple antigens.111–116 However, antibody production does not result in eradication even though H. pylori is susceptible in vitro to antibody-dependent complement-mediated phagocytosis and killing.117, 118 These observations suggest that gastric mucus may be a protective niche in which H. pylori exist and are relatively inaccessible to specific antibodies or their effector functions. The ineffective humoral response generated towards H. pylori and its components (i.e. Lewis antigens) may instead actually contribute to pathogenesis. Monoclonal antibodies directed against H. pylori cross-react with gastric epithelium of both mice and humans and delivery of these antibodies alone to mice can induce gastritis.70 In colonized human patients, H. pylori induces the formation of antibodies that recognize the H+,-K+ ATPase epitope on the luminal surface of acid-secreting parietal cells.70, 71 IgM antibodies generated by immortalized B cells obtained from H. pylori-colonized gastric mucosa also recognize gastric epithelium.119 These findings indicate that the mucosal humoral response to H. pylori, in addition to bacterial factors, may contribute to the development of distinct histological lesions. For example, an autoimmune reaction against parietal cells may lead to gastric atrophy with a concomitant reduction in gastric acidity; conversely, immunoglobulin-mediated destruction of epithelial cells may initiate and/or maintain mucosal inflammation and epithelial cell injury.

T lymphocyte-mediated responses

The gastrointestinal tract represents an important barrier between human hosts and microbial populations; the ability to distinguish pathogenic bacteria from commensuals is regulated through T-cell-dependent responses. CD4+ T-cells can be divided into two functional subsets: type 1 (Th1) and type 2 (Th2) T-helper cells, which are defined by distinct patterns of cytokine secretion. Th1 cells produce IL-2 and IFN-γ and promote cell-mediated immune responses, while Th2 cells secrete IL-4, IL-5, IL-6, and IL-10 and induce B-cell activation and differentiation.120 The type of immune response to a particular microbial agent is governed by preferential expansion of one T helper cell subset accompanied by a corresponding down-regulation of the other.120 Most intracellular bacteria induce Th1 responses, while extracellular pathogens stimulate Th2 type responses. Based on the fact that H. pylori is non-invasive and that infection is accompanied by an exuberant humoral response, one might predict that a Th2 response would be predominant within H. pylori-colonized gastric mucosa. Paradoxically, the majority of H. pylori antigen-specific T-cell clones isolated from infected gastric mucosa produce higher levels of IFN-γ than IL-4, which is reflective of a Th1 type response.121H. pylori also stimulates production of IL-12 in vitro, a cytokine that promotes Th1 differentiation.122 These findings raise the hypothesis that an aberrant host response (Th1) to an organism predicted to induce secretory immune responses (Th2) may influence and perpetuate gastric inflammation (Figure 3); animal models of H. pylori-induced gastritis have supported these conclusions. H. pylori infection of IFN-γ-deficient mice that cannot mount an appropriate Th1 response leads to decreased levels of gastric inflammation compared to wild-type mice, and in vivo neutralization of IFN-γ in mice infected with a related Helicobacter species (H. felis) similarly reduces the severity of gastritis.123–125 Certain strains of mice (C57/BL6) infected with H. felis that develop a Th1-type response exhibit extensive gastric inflammation, while other genetically distinct strains (BALB/c) that respond to infection with a Th2-like response develop only minimal gastritis.126 Adoptive transfer of Th2 lymphocytes from infected mice into infected recipients reduces bacterial colonization density, while transfer of Th1 cells increases the severity of gastritis.127 Recently, elegant studies have extended these observations and demonstrated that preceding challenge with a helminth (Heligmosomoides polygyrus) that induces a Th2-type mucosal reaction significantly attenuates the development of Th1-mediated gastritis and atrophy in response to H. felis infection.128 These data are consistent with a model in which an inappropriate host T-cell response towards H. pylori facilitates the development of gastric inflammation and injury (Figure 3).


Figure 3.  Working model for pathogenesis of H. pylori-induced diseases. Acquisition of H. pylori leads to the development of a Th1-mediated response, which facilitates induction and maintenance of gastric inflammation. Persons harbouring strains containing particular disease-related alleles, such as cagA, vacA s1 and iceA1, develop more severe mucosal inflammation, injury and atrophy.

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Epithelial cell injury

H. pylori-induced inflammation may also lead to other forms of epithelial cell injury and damage. Activated neutrophils generate reactive oxygen or nitrogen species that can induce oxidative DNA damage via formation of DNA adducts. Upon contact with H. pylori in vitro, an oxidative burst occurs within polymorphonuclear cells.129, 130 The specific types of cellular damage resulting from generation of reactive oxygen species include lipid peroxidation, protein oxidation, and oxidation of DNA. H. pylori increases oxidative damage of DNA in gastric epithelial cells.131, 132 Another consequence of persistent inflammation may be alterations in cellular turnover. Gastric epithelial cell proliferation rates within colonized mucosa are significantly increased compared to those in uninfected controls.133–135 In chronic H. pylori infection, there is a notable lack of epithelial cell necrosis, suggesting that other forms of cellular demise, such as apoptosis, may be induced.136 Apoptosis is a form of programmed cell death that consists of a tightly regulated series of energy-dependent molecular events, and the ability of H. pylori to alter apoptosis may influence clinical manifestations of infection.137 For example, enhanced rates of apoptosis could potentially accelerate the progression to atrophic gastritis, with a concomitant increase in the risk for distal gastric adenocarcinoma. In contrast, reduced rates of cell loss, especially when accompanied by hyperproliferation, could lead to a heightened retention of mutagenized cells, which may also predispose certain colonized individuals towards gastric carcinogenesis.

Several studies have reported increased levels of gastric epithelial cell apoptosis among H. pylori-infected persons.136, 138–140 However, substantial variation exists among apoptosis scores between infected and uninfected persons within these clinical populations. An important question raised by these observations is whether differing levels of apoptosis are a result of particular bacterial products or the induced host response. Recent data have suggested that epithelial cell apoptosis in vivo is likely to be regulated by host mediators present within inflamed mucosa. For example, IFN-γ, a Th1 lymphocyte-derived cytokine that is increased within colonized mucosa, is synergistic with H. pylori in inducing Fas–Fas ligand (FasL) regulated apoptosis of gastric epithelial cells in vitro.141, 142 Fas, a member of the TNF/nerve growth factor receptor superfamily, mediates apoptosis following activation by its cognate oligomerizing ligand, FasL. FasL is expressed by a variety of cells, including epithelial cells and activated T lymphocytes, and H. pylori can induce apoptosis in certain gastric epithelial and T-cell lines by activating Fas.140, 141, 143–146 Gastric T-cell lines that display a Th1 phenotype specifically recognize H. pylori antigens, and these same cells express FasL and produce TNF-α and IFN-γ, cytokines that are synergistic with H. pylori in inducing apoptosis and enhancing Fas expression on gastric epithelial cells in vitro141, 142, 147H. felis infection of Fas-deficient mice is also associated with reduced levels of inflammation, and mucosal apoptosis scores are decreased in parallel compared to infected wild-type mice.148 A paradigm invoked by these findings is that cytokines released by Th1 lymphocytes within H. pylori-inflamed mucosa may increase apoptosis by regulating Fas–FasL interactions between T-cells and gastric epithelial cells.121, 142

However, there are additional host genetic factors that may also contribute to the ability of H. pylori to alter apoptosis in vivo. Mice lacking secretory phospholipase A2 that are infected by H. felis, develop increased epithelial cell apoptosis.149 Apoptosis in response to H. pylori may reflect heterogeneity of class II MHC host genotypes. Gastric epithelial cells express class II MHC antigens on their surfaces and expression is higher in H. pylori-infected compared to uninfected tissue.150 Although class II MHC molecules are well-recognized for their role in regulating the immune response through binding and presentation of foreign antigens to CD4+ T-cells, ligation of class II MHC molecules also results in apoptosis, and binding of H. pylori to these IFN-γ-inducible molecules can directly stimulate apoptosis in vitro.151–153 El-Omar et al. have recently identified particular polymorphisms of the human interleukin-1β gene promoter that are genetic risk factors for both precursor lesions of gastric cancer (atrophic gastritis) as well as gastric adenocarcinoma among H. pylori-infected persons.154 IL–1β can stimulate multiple intracellular signalling pathways involved in apoptosis, including Fas, NF-κB, and MAP kinases, and thus differing levels of IL-1β expression within gastric mucosa may alter the ability of H. pylori to induce apoptosis.99, 105, 141, 142

In addition to differences in host genotype, particular bacterial constituents may influence the ability of H. pylori to alter cellular turnover. Two independent studies have demonstrated that persons infected with H. pylori isolates containing the strain-specific gene cagA (see below) have significantly higher proliferation rates, but lower apoptotic indices, than either cagA or uninfected persons.135, 155 Increases in proliferation that are not balanced by concordant increases in apoptosis over years of colonization may therefore contribute to the ability of cagA+ strains to augment the risk for gastric cancer in certain populations. This model does not, however, explain the enhanced risk of duodenal ulcer disease associated with cagA+ strains, as duodenal ulceration and distal gastric cancer appear to be mutually exclusively outcomes of chronic H. pylori infection. Further, a recent study in an ethnically distinct population has demonstrated increased apoptotic indices within cagA+-colonized mucosa.156 Therefore, the precise relationship between H. pylori cagA+ strains and cellular turnover in vivo remains to be clarified.

In vitro, urease can induce apoptosis by binding to class II MHC molecules expressed on epithelial cell surfaces.153 VacA has also been shown to induce apoptosis in cell culture systems by inserting into the mitochondrial membrane, inducing the release of cytochrome c, and activating caspase-3 (see below).140, 157, 158 Collectively, these data suggest that heterogeneity in degrees of epithelial cell apoptosis among infected clinical populations is probably dependent upon the tremendous genetic diversity that exists both between isolates of H. pylori and between human hosts.


  1. Top of page
  2. Abstract
  8. Acknowledgements
  9. References

Although the majority of persons colonized with H. pylori remain asymptomatic despite the presence of gastritis, serious sequelae of infection can occur. One hypothesis to explain why disease develops in only a minority of persons is that H. pylori strains differ in degrees of virulence. For this to be correct, a critical requirement is that variation must exist among H. pylori strains. Certain H. pylori components such as urease are ubiquitous and necessary for colonization and survival (Table 1). Therefore, variation in clinical outcome is unlikely to develop as a result of such highly conserved traits. Genes that are heterogeneously represented among H. pylori strains, however, may encode candidate virulence factors that influence pathologic course. Genetic studies indicate that H. pylori are tremendously diverse, with virtually each isolate from a different patient being unique.159–163 In addition, an individual may carry more than one H. pylori strain, and isolates present within an individual may gradually change over time due to recombination between strains, gene rearrangements, or deletions. Although this diversity has hindered characterization of bacterial factors associated with various disease outcomes, several different genetic loci have been identified. These include, among others, cagA, vacA, and iceA, for which persons harbouring particular alleles have different risks of disease (Table 2). These markers are not completely independent of each other and, importantly, are not absolutes but reflect degrees of risk.63, 164, 165

Table 2.   Comparison of several H. pylori genes linked to disease outcome Thumbnail image of

The cag pathogenicity island

One specific phenotype shown to differ among H. pylori isolates is expression of an immunodominant protein (CagA) encoded by the gene cagA, which is present in approximately 60–70% of US strains.166–169H. pylori cagA+ strains are associated with a significantly increased risk for severe gastritis, atrophic gastritis, peptic ulcer disease, and distal gastric cancer, compared to cagA strains.89, 114, 115, 170–179cagA is the terminal open reading frame of a ≈ 40 kb locus containing 31 genes (the cag pathogenicity island) that was probably acquired from another bacterial species.51, 52, 161cagA is commonly used as a marker for the entire cag locus, although it is becoming apparent that the presence of cagA does not always signify the presence of an intact cag island.168, 180, 181 Several cag island genes possess homology to components of a type IV bacterial secretion system which, in other prokaryotic species, functions as a conduit for export of multimeric proteins and nucleoproteins across both the inner and outer bacterial membrane. For H. pylori, several cag island genes are required, both for translocation of bacterial proteins into host cells, and induction of pro-inflammatory cytokine release.168, 181–186

Following H. pylori adherence to host cells, CagA is translocated into and phosphorylated within the epithelial cell, where it induces cytoskeletal changes, including cell elongation, cell spreading, and production of filapodia and lamellipodia.182–184, 187 Deconvolution immunofluorescence studies have revealed that phosphorylated CagA localizes in a cylindrical form directly beneath an attached H. pylori bacterium, and within the host cell cytoplasm.187 Because CagA tyrosine phosphorylation triggers host cell morphological changes, pathways that control organization of the actin cytoskeleton probably represent targets of intracellular CagA modification.

The ability of H. pylori to induce epithelial cell responses related to pathogenesis, such as IL-8 production, is not uniform across strain populations. Clinical observations that the cag island represented a genetic locus related to disease focused subsequent molecular investigations and, not surprisingly, one of the first H. pylori strain-specific constituents identified as being necessary for IL-8 production was a component of the cag island, cagE (picB).185 Inactivation of cagE not only attenuates IL-8 expression but also decreases NF-κB and mitogen-activated protein kinases activation in vitro.100, 105–107, 186 Numerous cag island genes (cagG, cagH, cagI, cagL and cagM), but not cagA, have now been demonstrated to be required for NF-κB and mitogen-activated protein kinases-mediated IL-8 production.105–107, 186 Clinical cagA+ strains are more potent in stimulating IL-8 production in vitro than cagA strains.96, 188 Similar to findings in vitro, carriage of cagA+ strains augments mucosal IL-8 expression in vivo and such increases are directly related to the more severe inflammatory response induced by these strains.89, 90 Our group and another have recently extended these findings by demonstrating that loss of cagE significantly attenuates the severity of H. pylori-induced gastritis in a rodent model of inflammation, Mongolian gerbils.181, 189 Thus, cagA+ strains appear to be disproportionately represented among persons who develop serious sequelae of H. pylori infection, and genes within the cag island are necessary for induction of epithelial cell responses relevant to pathogenesis (Table 2).

H. pylori vacA and the vacuolating cytotoxin

A second locus of heterogeneity is the gene vacA, which encodes the vacuolating cytotoxin. Approximately 50% of H. pylori strains produce a toxin that induces vacuole formation in epithelial cells.190 There is a strong correlation between vacuolating cytotoxin activity and the presence of cagA. However, vacA and cagA map to two distinct loci on the H. pylori chromosome, and mutation of cagA, resulting in loss of expression of the CagA protein, does not affect toxin production, indicating that expression of the two proteins is independent.191vacA encodes a precursor protein of approximately 140 kDa, which is processed to yield a mature 87 kDa protein (VacA).192–196 Mature VacA monomers then assemble into oligomeric structures that are composed of 12–14 sub-units.197, 198 When subjected to acid or alkaline conditions, the VacA oligomer disassembles but retains the capacity to re-anneal after pH neutralization.198 Acid activation also enhances the interaction of VacA with cellular membranes in vitro by increasing its insertion into artificial lipid bilayers where it forms anion channels.199–202 Cytoplasmic internalization of VacA is required for cytotoxicity and vacuole formation and is mediated by active cellular processes.203 Vacuoles have been suggested to originate from an intermediate intracellular compartment juxtaposed between the lysosomal and late endosomal stage, because vacuoles express the late endosomal marker Rab7, and their formation depends on the presence of sub-vacuolating concentrations of weak bases such as ammonia.204–208 However, VacA may also exert additional activities other than vacuolation; for example, several studies have recently reported that VacA induces gastric epithelial cell apoptosis.140, 157, 158

H. pylori strains that express vacuolating activity are more common among patients with peptic ulcer disease and distal gastric cancer than among infected patients with superficial gastritis alone.209–213 However, unlike cagA and the cag island, vacA is present in virtually all H. pylori strains examined.193, 209 The difference between strains with and without cytotoxic activity has been determined to be due to variations in vacA gene structure (Figure 4).193vacA sequences from cytotoxin-producing and non-cytotoxin-producing strains are distinct and represent families of mosaic alleles. Regions of diversity are localized to both the vacA signal sequence (s1a, s1b, s1c and s2) and the mid-region (m1 and m2) (Figure 4).160, 209, 214 In general, strains possessing an s1-type signal sequence allele produce a functional cytotoxin, while those with s2 signal sequences possess little if any cytotoxic activity, and s1/m1 strains are more toxigenic than s1/m2 strains.209 In the United States and Europe, H. pylori strains of the vacA s1a subtype are associated with enhanced gastric inflammation, peptic ulcer disease, and distal gastric cancer, compared to vacA s2 strains.164, 210 Although the precise mechanism by which VacA may lead to mucosal damage in humans is unknown, oral administration of H. pylori culture supernatants containing cytotoxin induces gastric epithelial cell damage and erosions in mice.215


Figure 4.  Schematic representation of vacA, the gene encoding the H. pylori vacuolating cytotoxin. vacA alleles are distinguished by mosaicism within the signal sequence (types s1a, s1b, s1c and s2) and mid-region (types m1 and m2).

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iceA (induced by contact with epithelium)

In addition to cagA and vacA, another strain-specific H. pylori locus that has been described is iceA. iceA was originally identified using a modified differential display technique as a gene whose transcription is up-regulated following adherence to gastric epithelial cells.216iceA exists in two major allelic sequence variants, iceA1 and iceA2, yet only iceA1 is induced following contact with epithelial cells. The deduced H. pylori iceA1 product demonstrates strong homology to a restriction endonuclease, NlaIII in Neisseria lactamica.217 However, mutations including insertions and deletions found in the majority of iceA1 sequences preclude translation of a full-length homologue. In contrast to iceA1, iceA2 has no homology to known proteins and its structure reveals patterns of repeated peptide cassettes (Figure 5). In its most common form, iceA2 can encode a protein of 59 amino acids with two conserved outer domains of 14 and 10 amino acids that flank three internal peptide domains of 13, 16, and six amino acids, respectively (Figure 5).218 Sequence analysis of iceA2 from several H. pylori strains has demonstrated that the internal 35 amino acid cassette (comprised of the 13, 16, and six amino acid domains) may be absent or repeated up to three times, resulting in deduced proteins of 24, 59, 94, or 129 amino acids (Figure 5).218 That iceA genes constitute two distinct families of alleles, significantly different from each other, suggests there may be a biologically relevant function for both types of iceA products.


Figure 5.  Schematic representation of the genetic organization of iceA2. The top schematic represents the prototype iceA2, encoding a protein of 5 9 amino acids (aa). The diagrams below represent subsequently identified iceA2 variants. Each of the five iceA2 peptide motifs of 14, 13, 16, six, and 10 amino acids is represented by a box. The existence of two distinct 16 amino acids domains is indicated by differently filled boxes. The total number of amino acids in each iceA2 ORF is shown for each variant.

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H. pylori iceA1 strains are significantly associated with the presence of peptic ulceration in certain populations, and levels of iceA1 expression within colonized human gastric mucosa are directly related to the severity of acute inflammation.164, 216, 219iceA1 is associated with the presence of cagA and the vacA s1 allele, but is only found in 25% of US H. pylori isolates, which approximates the percentage of infected persons who progress to peptic ulceration.216 The association of iceA1 with increased tissue damage and peptic ulcer disease, its allelic distribution in a minority of clinical strains, its linkage disequilibrium with cagA and vacA s1 alleles, and its induction by physiologic events that induce pathologic responses (adherence), collectively suggest that iceA1 may be a marker for strains that induce more severe gastric inflammation and injury.

H. pylori FldA and gastric mucosa-associated lymphoid tissue (MALT) lymphoma

Another H. pylori antigen that is significantly associated with a specific clinical sequelae of long-term colonization is FldA, a putative flavodoxin protein. Chang et al. recently reported that this 19-kDa protein is recognized by sera from H. pylori-infected patients with mucosa-associated lymphoid tissue lymphoma significantly more frequently than by sera from H. pylori-colonized patients without lymphoma.220 These investigators also found that an 11-amino acid truncation of the FldA protein was more common among H. pylori strains isolated from mucosa-associated lymphoid tissue patients (nine out of nine) than in control strains (six out of 17).220 However, FldA antibodies are not universally present among patients with mucosa-associated lymphoid tissue lymphomas, and not universally absent among controls, indicating that they are neither necessary nor sufficient for disease to occur. Further, it is unclear why a truncation of FldA would affect host production of serum antibodies. Therefore, it remains to be determined whether FldA plays an important causal role in mucosa-associated lymphoid tissue development or, alternatively, may simply be a marker for strains associated with this neoplasm.

H. pylori outer membrane proteins

In Western populations, persons colonized with cagA+vacA s1a iceA1 strains are more likely to develop peptic ulcer disease than persons harbouring cagAvacA s2 iceA2 strains (Figure 3). However, important geographical differences in susceptibility to disease exist; clear-cut markers for H. pylori strains that affect Western populations have little or no predictive power among East Asian populations.221, 222 Sequence analysis of the genomes from H. pylori strains 26695 and J99 has revealed that an unusually high proportion (1%) of identified open reading frames are predicted to encode outer membrane proteins.51 Consequently, recent attention has been directed toward a possible role of these outer membrane proteins in H. pylori pathogenesis.51, 52, 223 One such outer membrane protein is a 34-kDa proinflammatory protein encoded by oipA.224 Yamaoka et al. made the seminal observation that the vast majority of cagA+ strains isolated from patients in East Asia have an intact copy of oipA and when co-cultured with gastric epithelial cells in vitro, these strains were found to induce high levels of IL-8. Inactivation of oipA decreased IL-8 levels by approximately 40%, while inactivation of both oipA and cagE in the same strain completely attenuated IL-8 production, indicating a potential role of OipA in H. pylori virulence.224

The identification of additional strain-specific genes related to disease has been facilitated recently by the use of an H. pylori whole genome micro-array.180, 181 This particular micro-array contains representations of each gene present in the two H. pylori strains that have been completely sequenced.51, 52 Microarray-based comparisons between strains therefore allow each gene within the H. pylori genome to be interrogated simultaneously (Figure 6). Further, because evolutionary pressures tend to select for the co-inheritance of genes involved in common pathways, identification of genes that segregate with known H. pylori virulence-related loci (i.e. cag island) by micro-array may reveal functional relationships that are not evident from sequence data alone. In point of fact, Salama and colleagues recently hybridized DNA from a panel of H. pylori isolates to a whole genome microarray, and identified a subset of genes that co-vary with the cag pathogenicity island, including two that encode predicted outer membrane proteins: omp27 and babA.180 If these genes are found in future studies to be related to distinct disease outcomes, they may represent novel molecular epidemiologic markers which could identify colonized persons at increased risk for clinical sequelae, who then might be considered for eradication therapy.


Figure 6.  Identification of H. pylori genes that are markers for specific clinical outcomes using DNA microarray. Genomic DNA isolated from H. pylori strains associated with varying disease sequelae can be labelled by incorporating one of two fluoresent nucelotide analogues (Cy3 or Cy5) into the DNA. Differentially labelled DNA samples are then mixed and co-hybridized to a H. pylori whole-genome microarray. Resulting signal intensities from each fluorophore are then compared for each ORF represented on the array, thus allowing one to identify differences in gene content between H. pylori strains.

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  1. Top of page
  2. Abstract
  8. Acknowledgements
  9. References

Considerable efforts have focused on delineating the precise mechanisms by which H. pylori colonization leads to gastric inflammation. However, an intriguing characteristic of H. pylori-induced gastritis is its capacity to persist for decades without causing serious damage in most cases. This is in marked contrast to inflammatory reactions induced by other Gram-negative pathogens, which either resolve over a limited time-span or progress to eliminate the host. Clinical complications of H. pylori colonization, such as peptic ulcer disease and gastric cancer, are therefore likely to represent imbalances in gastric homeostasis that are disadvantageous for both microbe and host, particularly if death of the host ensues.

A hypothesis based on these micro-ecological perspectives is that H. pylori also possesses mechanisms in its repertoire to subdue the host inflammatory response, a requirement seemingly inherent for an infectious microbe that persists for the lifetime of its host. In support of this, Crabtree et al. have found that inactivation of a cag island gene (cag10) results in a paradoxical increase in IL-8 secretion compared to levels induced by wild-type H. pylori.225H. pylori infection is associated with increased mucosal levels of IL-10, an anti-inflammatory peptide that inhibits secretion of pro-inflammatory chemokines from macrophages and neutrophils.89, 90, 122, 226 When compared with LPS from the Enterobacteriaceae, H. pylori LPS is 1000-fold less active, and only weakly activates macrophages.227 The failure of H. pylori to invade the mucosa may contribute to its long-term persistence. It is tempting to speculate that H. pylori can also orchestrate the host response by negatively regulating intracellular eukaryotic signalling pathways, as has recently been described in Salmonella typhimurium.228 Certain non-pathogenic Salmonella attenuate IL-8 secretion induced by pathogenic bacteria by inhibiting the ubiquitination of IκBα, a novel mechanism for dampening the inflammatory response.228 However, it is becoming increasingly apparent that regulation of chronic gastric inflammation by H. pylori is governed by levels of host-bacteria equilibria that are not found during cellular interactions with acute pathogens. Clinically apparent disease probably results from the cumulative effect of multiple interactions between H. pylori and its host.


  1. Top of page
  2. Abstract
  8. Acknowledgements
  9. References

This study was supported in part by the National Institutes of Health (DK02381, CA77955), and by the Medical Research Service of the Department of Veterans Affairs.


  1. Top of page
  2. Abstract
  8. Acknowledgements
  9. References
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