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Hp Infection (see also Chapter 51)
Hp is a Gram-negative microaerophilic bacterium that infects nearly half the world’s population and is recognized as the primary etiologic agent for gastric cancer. Indeed, Hp has been classified as a class I (or definite) carcinogen by the International Agency for Research on Cancer (IARC), a branch of the WHO. Infection with Hp has been found in every population studied, although the prevalence is higher in developing countries and much of east Asia.20,21
The natural history of chronic Hp infection includes 3 possible outcomes22 : (1) simple gastritis, where most patients remaining asymptomatic; (2) duodenal ulcer phenotype, which occurs in 10% to 15% of infected subjects; and (3) gastric ulcer/gastric cancer phenotype, which is the least common in the USA. The risk for gastric cancer related to the types of gastritis and, in general, an increased risk is associated with a low acid state. Hp-induced duodenal ulcer disease is associated with a high gastric acid output as well as a reduced risk for developing gastric cancer.23 Studies suggest that Hp-infected patients develop chronic atrophic gastritis at a rate of 1% to 3% per year of infection.14,24,25 Thus, those patients who are genetically predisposed to developing atrophic gastritis in response to Hp infection are predisposed to gastric cancer. Although Helicobacter infection is associated with both diffuse-type and intestinal-type adenocarcinomas, the mechanisms responsible for the formation of intestinal-type adenocarcinoma have been better studied and are focused on here. The association of Hp with mucosa-associated lymphoid tissue (MALT) lymphoma is discussed in Chapter 31.
The increased risk of development of gastric adenocarcinoma due to Hp infection depends on multiple factors including host genetic factors, the strain of bacteria, the duration of infection, and the presence or absence of other environmental risk factors (e.g., poor diet, smoking). In a Japanese cohort, only those infected with Hp developed gastric adenocarcinoma during follow-up (2.9% vs. 0%; P < 0.001).26 Additional cohort studies from China and Taiwan have reported similar findings.27,28 In Western countries, the association between Hp and gastric cancer appears to be confined to non-cardia tumors.29
A combination of a virulent bacterial strain, a genetically permissive host, and a favorable gastric environment may be necessary for cancer to occur. Currently, genetic susceptibility factors of the human host are studied on the basis of individual genes, but new technologies such as next-generation sequencing will enhance the identification of host genetic factors. Nevertheless, the most important factor appears to be the induction of chronic inflammation by Hp infection. Several aspects of the inflammatory milieu have been implicated as carcinogens; they include increased oxidative stress and the formation of oxygen free radicals, leading to DNA damage, increased CD4 + T cells and myeloid cells, and elevated proinflammatory cytokine production, all leading to accelerated cell turnover, reduced apoptosis, and the potential for faulty or incomplete DNA repair.30 Indeed, recent studies suggest that animals with deficient DNA repair mechanisms display more severe gastric dysplasia after chronic infection with Hp.31Thus, evidence to date clearly indicates that the most important cofactor in the induction of Helicobacter-related disease is the host immune response. Indeed, chronic inflammation has been linked to a large number of non-gastric cancers.
Chronic inflammation appears necessary for the progression through atrophy to gastric cancer. Disease mechanisms are difficult to study in human infection, and therefore, much of our understanding of the immune response to Helicobacter organisms comes from work performed in a mouse model. Different inbred strains of mice respond to infection with varying degrees of disease susceptibility, and several knockout models have helped to elucidate the roles of individual components of the immune response in disease.
Genetic manipulation of the C57BL/6 susceptible murine strain has facilitated detailed study and has thus led to a deeper understanding of genetic factors that promote murine gastric cancer, and in particular, the role of the adaptive immune response. For example, gastric Helicobacter infection in mice deficient in lymphocytes, including mice with recombinase-activating gene (RAG) deficiency, severe combined immunodeficiency, or T cell deficiency, does not result in tissue damage, cell lineage alterations, or the metaplasiadysplasia-carcinoma sequence.32,33 In contrast, infection in B cell–deficient mice (which retain a normal T cell response) results in severe atrophy and metaplasia identical to that seen in infected wild-type mice.33 Taken together, these studies underscore the crucial role of CD4 + T lymphocytes in orchestrating gastric neoplasia.
How do CD4 + T lymphocytes contribute to gastric cancer progression? Susceptible mouse strains, such as C57BL/6, mount a strong Th1 (T helper cell type 1, expressing interferon [IFN]-γ and interleukin [IL]-12) type of immune response, whereas resistant strains, such as the BALB/c, have a polarized Th2 response (expressing IL-4 and IL-5).34-36 A Th2 response is associated with protection from mucosal damage despite the inability to eliminate bacterial colonization and, in fact, is often associated with higher bacterial colonization rates. Mouse strains such as the C3H, which has a mixed Th1/ Th2 cytokine profile, show intermediate disease, suggesting that cytokines within an immune response interact to form a continuum of disease rather than discrete disease states. More recently, Th17 cells (expressing IL-17), have been shown to be an important component of Hp-induced gastritis.
Although the composite immune milieu most likely dictates disease manifestations, there may be a role for individual cytokines in both the predisposition to and protection from disease. During Helicobacter infection, the Th1 cytokine IFN-γ is able to promote or inhibit inflammation-driven cancer of the stomach, suggesting that a more specific immune response is responsible for cancer promotion or surveillance. While studies in the past have suggested that IFN-γ might promote the development of gastric preneoplasia,37 IFN-γ overexpression in the stomach at low levels was recently shown to be able to suppress gastric cancer in models of IL-1β and Helicobacter felis–dependent carcinogenesis.38 In addition, IFN-γ was shown to counteract the development of Th17 cells.38 Thus, different composition of the same cells and cytokines in the tumor microenvironment can contribute to a constellation that favors or inhibits carcinogenesis. On the other hand, mice lacking IL-10, a cytokine that acts to dampen an immune response, demonstrate severe atrophic gastritis in response to infection.32-36 More recently, genetic murine models have illustrated the importance of the IL-6/IL-11 family of cytokines in the development of gastric cancer.39
Manipulation of the immune response within wild-type strains confirms the central role of the Th1/Th2 response in producing disease. For example, infection with the intestinal helminth Heligmosomoides polygyrus skews the immune response toward Th2 polarization and protects the C57BL/6 host from Helicobacter-induced atrophy and metaplasia.40 This mouse model mimics both the parasitic infection status and the paradoxical low gastric cancer–high Hp infection rates seen in areas of Africa, potentially explaining this apparent inconsistency. These observations in mice led to human studies in Africa and Latin America that confirmed that geographic regions with low gastric cancer rates had much higher Th2/Th1 immune responses to Hp.41,42 In general, the increased Th2 type responses were found in areas where serum IgE levels were high and the prevalence of intestinal parasitism by helminthes is above 50%. These findings further stress the importance of the host response to infection and suggest the possibility that manipulation of the genetically predetermined host cytokine profile in response to environmental challenges may lessen or exacerbate the disease process.
Whereas Hp infection has been unequivocally linked to gastric cancer, the development of dysplasia and invasive cancer tends to occur at a time when Hp colonization has either dramatically declined or, in some cases, has disappeared from the stomach altogether. Gastric cancer almost always occurs in the setting of prolonged gastric atrophy and hypochlorhydria, a condition that predisposes to enteric bacterial overgrowth. While antibiotic eradication therapy targeting Hp delays and inhibits development of gastric cancer in mice,16,43 antibiotics eradicate not only Hp but also other microorganisms that colonize the atrophic, hypochlorhydric stomach. Indeed, infection of otherwise germ-free INS-GAS mice with Hp resulted in delayed progression to gastric cancer compared to Hp-infected INS-GAS mice colonized with conventional flora.44 Thus, Hp may represent simply the initial, or the most prevalent, microbial factor responsible for gastric cancer progression.
There is a great deal of genetic diversity between strains of Hp owing to point mutations, insertions, deletions, and base-pair substitutions within the genome. Several strains may infect a single individual, and existing strains can undergo mutations and change over time.45,46 Despite this genetic diversity, several genes are recognized as risk factors for gastric carcinoma, including the cag pathogenicity island, the vacA gene, and the babA2 gene.
The Hp genome is 1.65 million base pairs and codes for approximately 1500 genes, two thirds of which have been assigned biological roles.47 The function of the remaining one third of the genome remains obscure, but genome-wide analyses using DNA microarray or whole-genome sequencing technology will give a broad view of the genome of Hp in the near future. Factors that contribute to carcinogenesis include those that enable the bacteria to effectively colonize the gastric mucosa, those that incite a more aggressive host immune response, and those that affect host cell-growth signaling pathways.
Motility toward epithelial cells of the stomach is a vital feature of Hp survival tactics. This is ensured by several factors. Spiraling movement is mediated by the FlaA and FlaB proteins, which are designed to navigate the thick gastric mucus. Additionally, Hp produces HP1069. A putative collagenase, which modifies the extracellular matrix and mucus layer, thus decreasing viscosity and allowing bacterial penetration.48,49 In addition, Hp expresses a variety of genes that contribute to buffering of stomach acid in order to main a relatively neutral pH. This includes a urease gene cluster consisting of 7 genes, of which UreA/UreB complex (comprising the urease enzyme) codes for 10% of the protein of Hp and is vital for its survival.
The cag pathogenicity island is approximately 40 kb and contains 31 genes. The terminal gene of this island, cagA, is often used as a marker for the entire cag locus. Compared with cagA-negative (cagA−) strains, cag-positive (cagA+) strains are associated with more severe inflammation, higher degrees of atrophy, and a greater chance of progressing to gastric adenocarcinoma.50-53 The estimated relative risk has ranged from 2 to as high as 28.4.22 However, many of the genes adjacent to cagA code for a type 4 secretion system (TFSS), often viewed as a molecule needle that injects bacterial proteins (e.g., cagA) into host cells. The remarkable finding that CagA is injected into host cells where it is phosphorylated by Srcand c-Abl kinases, has raised the possibility that CagA could directly promote growth, migration, and transformation. Indeed, transgenic expression of Hp CagA induces both GI and hematopoietic neoplasms in mice.54 Other genes within the pathogenicity island are also believed to be important for disease (cagE or picB, cagG, cagH, cagI, cagL, cagM) because they appear to be required for in vitro epithelial cell cytokine release, although they do not seem to have as great an effect on immune cell cytokine activation.55-57 These findings may explain the attenuated inflammatory response and lower cancer risk with cagA− strains in vivo.58-61
All strains of Hp carry the vacA gene, which codes for a pore-forming, vacuolating toxin, but expression differs according to allelic variation. Approximately 50% of Hp strains express the vacA protein, which has been shown to be a very powerful inhibitor of T cell activation in vitro.62 Although vacA and cagA map to different loci within the Hp genome, the vacA protein is commonly expressed in cagA+ strains. There are various forms of vacA, and the s1m1 strains are highly toxigenic. Other bacterial virulence factors, such as cagE, may play a role in the modulation of apoptosis and the host inflammatory response, thereby contributing to disease manifestations. Indeed, “virulent strains” (cagA+, cagE+, and VacA+ s1m1) appear to be more potent inducers of proinflammatory mediators than “nonvirulent strains” (cagA−, cagE−, and VacA−), possibly explaining the higher association of cagA+ strains with gastric cancer.63