Helicobacter pylori: Molecular and Cellular Biology Chapter Abstracts

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Abstract

For more than 200 years, generations of pathologists, clinicians and epidemiologists have tried to explain the cause of peptic ulcer disease. Beginning with Robert Koch and Louis Pasteur in the 19^th century, one century was necessary to establish the connection between Helicobacter pylori and peptic ulcer disease. The isolation of H. pylori by Marshall and Warren 15 years ago initiated one of the major medical revolutions of the 20^th century (Marshall and Warren, 1984). However, it is becoming clear that the relationship between H. pylori and human disease is highly complex. We all seek simple answers but, as in the case of H. pylori, "truth is rarely pure and never simple" (Oscar Wilde, The Importance of Being Earnest, Act.1). With this in mind, we present a historical overview of H. pylori infection.

Chapter 2: Taxonomy and Phylogeny of Helicobacter

James Versalovic and James G. Fox

Abstract

The genus Helicobacter was formally proposed in 1989 and represents a rapidly expanding taxon with considerable medical interest. The importance of Helicobacter pylori stimulated investigations of multiple related species isolated from the gastrointestinal tracts of animals and humans. Currently, twenty formally named species have been designated and proposed names such as "Helicobacter rappini" likely include multiple taxa. Helicobacters have been grouped as gastric or enteric (enterohepatic), depending on their preferred site of colonization. These organisms are uniformly Gram-negative non-sporeforming bacilli which are motile and usually possess multiple or single bipolar sheathed flagella. Biochemical, genetic, and physiologic features distinguish Helicobacters from the related genera, Campylobacter and Arcobacter, and emphasize their unique contributions to the gastrointestinal microbiota.

Chapter 3: Epidemiologic Observations and Open Questions About Disease and Infection Caused by Helicobacter pylori

Roger A. Feldman

Abstract

This book documents the variety of virulence factors, geographic variations, and variation in response to environmental factors that exist within H. pylori. As a result, earlier epidemiologic analyses of prevalence rates and disease associations, which generally considered H. pylori as a single entity, benefit from reinterpretation and newer studies will deal separately and comparatively with the various genetic clusters, geographic variants and environmental factors identified in H. pylori. This chapter reviews the most recently available data dealing with several specific epidemiologic questions. Data are reviewed concerning mode of spread, risk factors, and acquisition of infection, both in children and in adults. Studies in children are analyzed concerning the frequency of both stable and transient infection. With adults, the data summarized relate both to the frequency and source of adult infection and to rates of reinfection after successful treatment. The questions addressed also relate to multiple infections and the association of persistent H. pylori infection with any of a multitude of diseases, ranging from iron deficiency anamia, heart disease and other non-gastroenterologic problems to gastritis, peptic ulcer, gastric cancer, and MALT lymphoma.

Chapter 4: Antigastric Autoimmunity and Pathology in Helicobacter pylori Gastritis

Gerhard Faller and Thomas Kirchner

Abstract

Chronic and active Helicobacter pylori gastritis represents the common platform for several gastroduodenal complications, such as duodenal or gastric ulcer, gastric mucosa atrophy, gastric carcinoma or gastric MALT-type lymphoma. These clinical outcomes are reflected by the histological pattern and topographical distribution of the inflammatory infiltrates in the gastric mucosa. Classification of chronic gastritis describes the different patterns of mucosa alterations and can also provide information on the probable clinical course and the aetiology of the disease. So far, autoimmune gastritis and H. pylori gastritis were considered strictly distinct. However, autoimmune gastritis and corpus predominant, atrophic H. pylori gastritis show many histological similarities. Furthermore, recent immunological studies revealed a close pathogenic relationship between autoimmune gastritis and one subset of H. pylori gastritis. These new findings are summarized in this chapter and lead to the speculation whether the strict distinction between autoimmune gastritis and H. pylori gastritis should be maintained in the future.

Chapter 5: Cytokine Responses in Helicobacter pylori Infection

Jean E. Crabtree

Abstract

Cytokines play a critical role in the initiation and modulation of gastric mucosal inflammatory responses to Helicobacter pylori. The gastric epithelium which secretes chemokines in response to H. pylori has an important role in the initiation of the acute inflammatory response. NF-kB activation and secretion of C-X-C chemokines such as IL-8 and GRO-a are induced by strains with the cag pathogenicity island (PAI). Mutational studies show multiple genes in the cag PAI are required for induction of this signalling response. In vivo cag PAI positive infection is associated with increased mucosal C-X-C chemokine expression and enhanced inflammatory responses. Strain heterogeneity may contribute to the broad spectrum of clinical outcome of infection. Antigen-specific chronic inflammation is a predominant Th1 response characterized by gamma interferon secreting effector cells. The mucosal production of gamma interferon and Th1 inducing cytokines such as IL-12 and IL-18 may be important in the induction of mucosal damage and development of autoimmune responses. Cytokines also play an important role in the changes in host gastric physiological responses associated with infection.

Chapter 6: Role of Th1/Th2 Cells in H. pylori­induced Gastric Disease

Mario M. D'Elios and Gianfranco Del Prete

Abstract

H. pylori infection represents the major cause of gastroduodenal diseases such as chronic gastritis, peptic ulcer, gastric cancer or B-cell lymphoma of the mucosa-associated lymphoid tissue (MALT), but only a minority of infected patients ever develop these pathologies. The type of the host immune response against H. pylori seems to be crucial for the outcome of infection.

A model has been developed to investigate the gastric T-cell response to H. pylori occurring in infected patients. A predominant H. pylori -specific Th1 response, characterized by high IL-12, IFN-g and TNF-a production was found in the gastric antrum of patients with peptic ulcer. In uncomplicated chronic gastritis most gastric H. pylori -specific T cells secrete both Th1- and Th2-type cytokines, showing a Th0 profile. Th0 was also the predominant cytokine profile of T cells derived from gastric MALToma, but these gastric T cells exhibit abnormal help for autologous B-cell proliferation and reduced perforin- and Fas-Fas Ligand-mediated killing of B cells, unlike T cells from chronic gastritis with or without ulcer.

These data suggest that the host gastric immune response to H. pylori can influence the clinical picture. Peptic ulcer may be an immunopathological consequence of a Th1-polarized response to some H. pylori antigens, whereas deregulated H. pylori -induced T cell-dependent B-cell activation may support the onset of low-grade B-cell lymphoma.

Chapter 7: Helicobacter pylori Induced Apoptosis

Siegfried Wagner and Winfried Beil

Abstract

Helicobacter pylori gastritis is accompanied by a reversible induction of gastric epithelial cell apoptosis and proliferation which leads to an increased rate of gastric epithelial cell turnover. Enhanced apoptotic rates were found at the margin of peptic ulcers, while inadequately low apoptotic rates were documented in foci of intestinal metaplasia, suggesting a possible role of the apoptosis to proliferation ratio as a factor determining the clinical outcome of H. pylori infection. In vitro studies have demonstrated that H. pylori is capable of directly inducing apoptosis and a synergistic role of proinflammatory cytokines has also been shown. All clinical isolates of H. pylori, irrespective of their cagA and vacA status, induced DNA fragmentation in gastric epithelial cells, whereas Campylobacter jejuni failed to do so. H. pylori induced apoptosis was associated with an up-regulation of the cell death receptor CD95/APO-1/Fas and of the pro-apoptotic Bcl-2 homologues Bak and Bax in gastric epithelial cells. H. pylori induced DNA fragmentation was preceded by the sequential activation of caspase-9, caspase-8, and caspase-3. The molecular nature of the apoptosis inducing factor(s) and its interaction with the apoptotic signalling pathway are unknown and need to be addressed in future studies.

Chapter 8: Animal Models for Helicobacter pylori Gastritis

Kathryn A. Eaton

Abstract

Almost since the first successful culture of Helicobacter pylori and its association with human disease, animal models have been central in the progression of Helicobacter research. Studies in animals have illuminated numerous aspects of disease including the roles of bacterial virulence factors in colonization, the role of the host immune response in clearing infection as well as contributing to disease, and mechanisms of carcinogenesis. In addition, the development of effective treatment regimes, and the current progress in development of safe and effective vaccines are largely products of the use of animal models.

In recent years, as our understanding of Helicobacters progressed, animal models have become increasingly refined. The introduction of robust mouse models of disease due to Helicobacter pylori has led to an explosion of studies using these models in recent years, and progress has been rapid in the elucidation of all aspects of disease. This review describes the history and use of animal models of H. pylori gastritis, with emphasis on recently developed mouse and other rodent models. Features of animal models are described as are their uses in and contributions to the investigation of bacterial virulence factors, carcinogenesis, and host responses and immunology.

Chapter 9: Transgenic Mouse Models for Studying the Relationship between Helicobacter pylori Infection and Gastric Cancer

Per G. Falk, Andrew J. Syder, Janaki L.Guruge, Lars G. Engstrand, and Jeffrey I. Gordon

Abstract

Sero-epidemiologic studies suggest that H. pylori infection may increase the risk of developing gastric adenocarcinoma. Nonetheless, the host and microbial factors that may produce this outcome are poorly understood. Transgenic mouse models offer one approach for developing and conducting direct tests of hypotheses about the mechanisms that underlie H. pylori-associated tumorigenesis.

Chapter 10: Helicobacter pylori Urease

Harry L.T. Mobley

Abstract

Urease, a protein expressed in high quantity by Helicobacter pylori, catalyzes the hydrolysis of urea to yield ammonia and carbamate. The K_m ranges from 0.17-0.48 mM urea and the specific activity ranges from 1100 to 1700 mmol urea/min/mg protein. The apoenzyme is comprised of six copies of each of two structural subunits, UreA and UreB and the metalloenzyme contains two nickel ions in each of the six active sites. Seven contiguous genes, ureABIEFGH, are transcribed in the same direction from a 6.13 kb sequence and all of them except ureI are necessary for synthesis of a catalytically active enzyme. Active urease is required for the colonization of the gastric mucosa.

Chapter 11: Helicobacter Motility and Chemotaxis

Christine Josenhans and Sebastian Suerbaum

Abstract

All gastric Helicobacter species are highly motile. Their characteristic sheathed flagellar filaments are composed of two copolymerized flagellins, FlaA and FlaB. Experiments in three different animal models with Helicobacter pylori, Helicobacter mustelae and Helicobacter felis showed that flagellar motility is essential for Helicobacter species to colonize the gastric mucus. H. pylori has homologs of all flagellar structural proteins known from Salmonella but the regulatory network of H. pylori motility genes lacks a typical flhCD master operon and a flgM sigma factor antagonist gene. Although H. pylori manifests chemotaxis towards urea and bicarbonate, it is still unknown which chemical gradients are used by H. pylori in vivo to maintain an optimal position in the gastric mucus layer. The chemotaxis system of H. pylori is genetically similar to the Salmonella system as based on genomic analysis but extensive functional analyses have not yet been performed.

Chapter 12: Helicobacter pylori, An Adherent Pain in the Stomach

Markus Gerhard, Siiri Hirmo, Torkel Wadström, Halina Miller-Podraza, Susann Teneberg, Karl-Anders Karlsson, Ben Appelmelk, Stefan Odenbreit, Rainer Haas, Anna Arnqvist and Thomas Borén

Abstract

Modern strains of Helicobacter pylori are the result of selection for life in the human gastric mucosa. This is a most demanding environment, with high acidity, peristalsis, gastric emptying and shedding of cells and mucus. In order to ensure efficient long-term survival and colonization, these microbes have developed arrays of adherence properties for homing to the optimal set of host cells. Survival demands an adequate balance between movement due to flagellar motility and attachment by adherence. This balance allows the microbes to take advantage of continuous supplies of nutrients without cutting off escape from the mucus layer to evade the cellular immune responses. H. pylori possesses a multitude of genetic mechanisms for the flexible shifting of adherence specificities to allow adaptation to the host and the local environment. The detailed characterization of the mechanisms that support and maintain bacterial adherence will identify key-elements for the persistence of infection that can be targeted for anti-microbial drug strategies.

Chapter13 Helicobacter pylori Lipopolysaccharides

Anthony P. Moran

Abstract

Lipopolysaccharides (LPS), a family of toxic phosphorylated glycolipids in the outer membrane of Gram-negative bacteria are composed of a lipid moiety (termed lipid A), a core oligosaccharide, and a polymeric O-specific polysaccharide chain. Compared with LPS of other bacteria, H. pylori LPS has, in general, low immunological activity which may aid the persistence of infection in the gastric mucosa. The molecular basis for this lower activity resides in the under-phosphorylated and unusally acylated lipid A component of LPS. The core oligosaccharide of H. pylori LPS contributes to the binding of the bacterium to the host glycoprotein laminin, and hence interferes with gastric cell receptor-laminin interaction in the basement membrane. The core sugars of certain H. pylori strains, particularly those associated with gastric ulceration, have been implicated in pepsinogen induction, which may result also in reduced mucosal integrity. Of particular interest, the O-chains of most, but not all, H. pylori strains mimic Lewis (Le) antigens. Although investigations have focussed on the role of these antigens in H. pylori-associated autoimmunity, which remains to be unequivocally established, other pathogenic consequences of Lewis mimicry are becoming apparent. Expression of Lewis antigens may be crucial for H. pylori colonisation and adherence and, by aiding bacterial interaction with the gastric mucosa, potentiate delivery of secreted products, and hence influence the inflammatory response.

Chapter 14: The cag Type IV Secretion System of Helicobacter pylori and CagA Intracellular Translocation

Markus Stein, Rino Rappuoli and Antonello Covacci

Abstract

The evolution and dissemination of virulence traits between bacteria is based on mobilization of DNA by a) phage conversion, b) distant events of horizontal transfer that promoted the pathogenicity island assembly and c) plasmid mobilization. Type III and Type IV secretory apparatuses are often located on mobilized DNA and are associated with an increased virulence of the bacterial specialist. Type I strains of Helicobacter pylori possess the cag pathogenicity island that encodes a specialized Type IV secretion machinery activated during infection. The cag organelle is involved in cellular responses like induction of pedestals, secretion of interleukin-8 and phosphorylation of proteic targets. Co-cultivation of epithelial cells with Helicobacter pylori triggers signal transduction and tyrosine-phosphorylation of a 145 kDa putative host cell protein. We and others have recently demonstrated that this protein is not derived from the host but rather is the bacterial immunodominant antigen CagA, a virulence factor commonly expressed during infection in patients of peptic ulcer disease and thought to be orphan of a specific biological function. CagA is delivered into the epithelial cells by the cag type IV secretion system where it is phosphorylated on tyrosine residues by an as yet unidentified host cell kinase and wired to eukaryotic signal transduction pathways and cytoskeletal plasticity.

Chapter 15: Helicobacter pylori VacA Vacuolating Cytotoxin and HP-Nap Neutrophil Activating Protein

Cesare Montecucco, Emanuele Papini, Marina de Bernard, William G. Dundon, Mario Zoratti, Jean-Marc Reyrat, John L. Telford, Giuseppe del Giudice and Rino Rappuoli

Abstract

The vacuolating toxin (VacA) is a major determinant of Helicobacter pylori-associated gastric disease. Unlike former beliefs, this toxin is active in almost all strains of H. pylori. In non-polarised cells, VacA alters the endocytic pathway, resulting in the release of acid hydrolases and the reduction of both extracellular ligand degradation and antigen processing. In polarized monolayers, VacA reduces the transepithelial electrical resistance and forms trans-membrane anion-specific channels. Localization of the VacA channels in acidic intracellular compartments causes osmotic swelling which, together with membrane fusion, leads to vacuole formation. The neutrophil activating protein of H. pylori (HP-Nap) induces the production of oxygen radicals in human neutrophils via a cascade of intracellular activation events which may contribute to the damage of the stomach mucosa. The activities of both VacA and HP-Nap are discussed in relation to H. pylori's nutrient requirements in the hostile environment of the human stomach .

Chapter 16: Development of a Helicobacter pylori Vaccine

Subhas Banerjee and Pierre Michetti

Abstract

Novel strategies are needed to control H. pylori infection on a global scale and the potential value of a vaccine is increasingly recognized. Studies in rodents have demonstrated the feasibility of prophylactic and therapeutic mucosal immunization against Helicobacter infections. Initial human trials showed that oral immunization with H. pylori urease is safe, immunogenic, and can result in a decreased gastric bacterial load. However, more potent vaccines will be needed to protect against or cure H. pylori infection in humans. To achieve this goal, our knowledge of the mechanisms of immune protection in the stomach needs to be improved. In rodents, the MHC II-restricted CD4⁺ T cell response plays a prominent role whereas antibodies are not necessary for protection. In humans, mechanisms that mediate protection against noninvasive gastric pathogens are largely unknown. Multivalent vaccines, safe adjuvants, and improved vaccine delivery systems all need to be studied. Divalent combinations of antigens are superior to single antigen vaccines in rodents. The availability of two genomic sequences of H. pylori will certainly help identify additional candidate vaccine antigens. Detoxified bacterial toxin adjuvants, live vaccine vectors and alternate routes of immunization are currently being actively evaluated.

Chapter17: Comparative Analysis of the Helicobacter pylori Genomes

Richard A. Alm and Trevor J. Trust

Abstract

Helicobacter pylori is the first bacterial species to have two completed genomes sequences available for comparison. The availability of the genomic sequence from two unrelated H. pylori isolates, J99 and 26695, has enabled a detailed analysis on the overall level of genetic diversity that has been previously reported for this organism. The arrangement of the genes in the two chromosomes was more similar than predicted, being disrupted by a few organizational differences and the presence of genes that were unique to only one of the strains, which comprised between 6% and 7% of the total coding capability of each strain. Half of the unique genes to each strain were contained within a hypervariable region of the chromosome. Analysis of the orthologous genes and their respective proteins displayed a high degree of allelic differences, and these differences were greater in the H. pylori specific genes, assisting in the understanding of the high discriminatory power of nucleotide-dependent typing techniques. As with other bacterial genomes sequenced to date, only approximately 60% of the identified genes encoded a product that could be assigned a function. These results will promote analyses on the population genetics and the evolution of the association of H. pylori with the human host. The genome sequences also provide direct access to all of the potential therapeutic targets allowing for the development of either novel anti-bacterial drugs or vaccines against H. pylori for people suffering from H. pylori related diseases.

Chapter 18: Molecular Typing of Helicobacter pylori

Leen-Jan van Doorn, Céu Figueiredo, and Wim Quint

Abstract

The genome of Helicobacter pylori is heterogeneous, due to a high mutation rate and a high degree of recombination. Therefore, H. pylori should be considered as a population of genetically related but diverse bacteria. To analyze the epidemiological and clinical implications of H. pylori variability, adequate molecular tools should be used for molecular typing of isolates. A variety of available methods, including RFLP, PCR-fingerprinting, sequence analysis, and probe hybridization is described in this chapter. Also, typing of virulence-associated and antibiotic resistance-related genes is discussed. Finally, some general requirements for effective molecular typing methods are provided.

Chapter 19: Population Structure of Helicobacter pylori and Other Pathogenic Bacterial Species

Mark Achtman

Abstract

This articles reviews recent data on population genetics of Helicobacter pylori within the framework of more extensive knowledge about other bacterial species. DNA sequences from different natural isolates of H. pylori differ frequently at synonymous sites that encode the same amino acids. Unlike other bacterial species, the polymorphic synonymous sites in H. pylori are frequently shuffled among different strains due to recombination. Thus, the population structure of H. pylori is panmictic. Founder effects or geographical specialization have resulted in distinct non-overlapping populations in different continents.

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