Andre Dubois

Natural gastric infection with Helicobacter pylori in monkeys: A model for spiral bacteria infection in humans ★

BACKGROUND/AIMS There is no generally accepted model for Helicobacter pylori infection in humans. The aim of this study was to examine the natural history and effect of treatment in rhesus monkeys and sequentially define the immune response to H. pylori in relation to treatment. METHODS Infection and gastritis were graded blindly by histological analysis and culture of biopsy specimens harvested during gastroduodenoscopies in 26 anesthetized colony-bred monkeys. Plasma H. pylori-specific immunoglobulin (Ig) G levels were determined by enzyme-linked immunosorbent assay. RESULTS H. pylori and Gastrospirilum hominis-like organisms were present in 13 and 9 monkeys, respectively; 3 animals harbored both organisms, whereas 4 monkeys were not infected. Gastritis score was < or = 1.5 in animals uninfected or infected only with G. hominis-like organisms and > or = 2.0 in all H. pylori-infected animals. IgG ratios were > or = 0.5 in 12 of 13 H. pylori-infected animals and in 2 of 13 H. pylori-negative animals (P < 0.001). One monkey became infected with H. pylori during the observation period, with concurrent increase of gastritis and plasma IgG levels. In untreated animals, infection, gastritis, and plasma IgG levels remained unchanged over 7-15 months. Triple therapy eradicated H. pylori at 6 months in 4 of 6 animals while suppressing gastritis and plasma IgG levels. CONCLUSIONS Rhesus monkeys harboring H. pylori are persistently infected and have gastritis and elevated specific IgG levels, all of which may respond to appropriate therapy, whereas G. hominis infection is associated with little inflammation.

SabA is the H. pylori hemagglutinin and is polymorphic in binding to sialylated glycans

Adherence of Helicobacter pylori to inflamed gastric mucosa is dependent on the sialic acid–binding adhesin (SabA) and cognate sialylated/fucosylated glycans on the host cell surface. By in situ hybridization, H. pylori bacteria were observed in close association with erythrocytes in capillaries and post-capillary venules of the lamina propria of gastric mucosa in both infected humans and Rhesus monkeys. In vivo adherence of H. pylori to erythrocytes may require molecular mechanisms similar to the sialic acid–dependent in vitro agglutination of erythrocytes (i.e., sialic acid–dependent hemagglutination). In this context, the SabA adhesin was identified as the sialic acid–dependent hemagglutinin based on sialidase-sensitive hemagglutination, binding assays with sialylated glycoconjugates, and analysis of a series of isogenic sabA deletion mutants. The topographic presentation of binding sites for SabA on the erythrocyte membrane was mapped to gangliosides with extended core chains. However, receptor mapping revealed that the NeuAcα2–3Gal-disaccharide constitutes the minimal sialylated binding epitope required for SabA binding. Furthermore, clinical isolates demonstrated polymorphism in sialyl binding and complementation analysis of sabA mutants demonstrated that polymorphism in sialyl binding is an inherent property of the SabA protein itself. Gastric inflammation is associated with periodic changes in the composition of mucosal sialylation patterns. We suggest that dynamic adaptation in sialyl-binding properties during persistent infection specializes H. pylori both for individual variation in mucosal glycosylation and tropism for local areas of inflamed and/or dysplastic tissue.

Role of ABO Secretor Status in Mucosal Innate Immunity and H. pylori Infection

The fucosylated ABH antigens, which constitute the molecular basis for the ABO blood group system, are also expressed in salivary secretions and gastrointestinal epithelia in individuals of positive secretor status; however, the biological function of the ABO blood group system is unknown. Gastric mucosa biopsies of 41 Rhesus monkeys originating from Southern Asia were analyzed by immunohistochemistry. A majority of these animals were found to be of blood group B and weak-secretor phenotype (i.e., expressing both Lewis a and Lewis b antigens), which are also common in South Asian human populations. A selected group of ten monkeys was inoculated with Helicobacter pylori and studied for changes in gastric mucosal glycosylation during a 10-month period. We observed a loss in mucosal fucosylation and concurrent induction and time-dependent dynamics in gastric mucosal sialylation (carbohydrate marker of inflammation), which affect H. pylori adhesion targets and thus modulate host–bacterial interactions. Of particular relevance, gastric mucosal density of H. pylori, gastritis, and sialylation were all higher in secretor individuals compared to weak-secretors, the latter being apparently “protected.” These results demonstrate that the secretor status plays an intrinsic role in resistance to H. pylori infection and suggest that the fucosylated secretor ABH antigens constitute interactive members of the human and primate mucosal innate immune system.

Helicobacter pylori is invasive and it may be a facultative intracellular organism.

The pathogenicity of many bacteria colonizing the gastrointestinal tract often depends on their ability to gain access to cells that are normally non‐phagocytic. Helicobacter pylori colonizes the stomach of over half the world population and is the main cause of peptic ulcer disease and gastric cancer. It is generally considered to be a non‐invasive pathogen present only in the lumen of the stomach and attached to gastric epithelial cells although a number of in vivo and in vitro studies have demonstrated that H. pylori is in fact invasive. In addition, H. pylori can repopulate the extracellular environment after complete elimination of extracellular bacteria with gentamicin, suggesting it may be considered a facultative intracellular bacterium. This review examines the validity of these observations and describes the evidence suggesting that the intracellular presence of H. pylori plays a role in the induction of diseases, in immune evasion, and in life‐long persistence of the bacterium in the stomach of a majority of humans.

Gastrointestinal Dysfunction in Systemic Mastocytosis: A Prospective Study

In 16 consecutive patients with systemic mastocytosis, we prospectively evaluated a variety of gastrointestinal functions and examined how they relate to the occurrence of gastrointestinal symptoms. Nine patients had either a duodenal ulcer or duodenitis. Hypersecretion of gastric acid was present in 6 patients, and in these patients the mean basal acid output was 20.7 +/- 4.1 mEq/h (range 14-39 mEq/h). Impaired small intestinal absorption occurred in 5 patients, although this was usually mild. The mean fractional emptying rate of liquids for all patients (14.7% +/- 2.3% per minute) did not differ from that for controls (10.7% +/- 0.6% per minute). Mean mouth-to-cecum transit time measured by breath hydrogen testing was the same among patients (87.7 +/- 6.7 min) and controls (86.7 +/- 8.0 min). Plasma histamine concentrations were increased in all patients (mean 1886 pg/ml, range 480-7450) and correlated with the basal acid output (r = 0.64, p less than 0.02) but not maximal acid output or the presence or absence of pain or diarrhea. Mean fasting plasma concentrations of motilin, substance P, and neurotensin from 6 patients did not differ significantly from controls, whereas gastrin and vasoactive intestinal peptide were significantly less than in controls (p less than 0.01). Gastrointestinal symptoms, consisting of abdominal pain or diarrhea, occurred in 80% of patients. Abdominal pain classified as dyspeptic was usually associated with acid-peptic disease of the duodenum and hypersecretion of gastric acid, whereas abdominal pain of a nondyspeptic character was not. Only in those cases of diarrhea consisting of greater than 200 g stool/day was gastric acid hypersecretion frequently found. Neither fecal urgency nor nondyspeptic pain could be accounted for by alterations of gastrointestinal transit. These results demonstrate that gastrointestinal symptoms, peptic disease, and mild malabsorption are much more common than described previously in patients with systemic mastocytosis. Furthermore, the results provide no evidence for the contention that altered gastrointestinal transit is involved in the pathogenesis of these symptoms.

Host specificity of Helicobacter pylori strains and host responses in experimentally challenged nonhuman primates

BACKGROUND & AIMS The specificity of colonization by Helicobacter pylori and complex host-bacterium interactions cannot be readily examined in humans. The aim of this study was to perform such analyses in rhesus monkeys. METHODS Four animals that had been cured of natural H. pylori colonization were challenged with a mixture of 7 strains of human origin, and bacteria recovered during periodic videogastroscopy were DNA fingerprinted. RESULTS Three animals carried mixtures of several strains for 4 months, after which strain J166 predominated. In the fourth animal, only strain J238 was isolated from the earliest phase of colonization through 7 months, but strain J166 again became predominant by 10 months after the challenge. Gastritis scores and plasma gastrin and anti-H. pylori immunoglobulin G titers reached levels observed in naturally colonized animals by 4 months after the challenge; however, no plasma immunoglobulin A response was observed up to 10 months. CONCLUSIONS These results show that (1) natural colonization does not elicit protective immunity against subsequent H. pylori challenge; (2) individuals differ in susceptibility to different H. pylori strains during initial stages of colonization; and (3) certain strains are better suited than others for long-term survival in different hosts. These observations show the complexity of H. pylori-host interactions.

THE ADVENT OF HIGHLY SELECTIVE INHIBITORS OF CYCLOOXYGENASE : A REVIEW

Cyclooxygenase (COX) exists in two isoforms, COX-1 and COX-2, COX-1 is present and is constitutively expressed in most cells and tissues, whereas COX-2 is felt to principally mediate inflammation. However, this distinction appears to be challenged by recent observations. This review addresses the roles of COX-1 and COX-2 isoforms in physiologic and pathophysiologic states and reviews potential therapeutic roles for selective COX inhibitors.

Inflammatory Cytokine mRNA Expression during Early and Persistent Helicobactev pylori Infection in Nonhuman Primates

The role of mononuclear phagocytes in orchestrating the host responses to Helicobacter pylori is inadequately understood. Therefore, gene expression for the monocyte/macrophage-derived cytokines interleukin (IL)-1beta, IL-6, and tumor necrosis factor (TNF)-alpha was determined before and during H. pylori infection of rhesus monkeys by use of a highly sensitive quantitative reverse transcriptase polymerase chain reaction. The numbers of molecules of IL-1beta, IL-6, and TNF-alpha mRNA in gastric tissue during early infection (7 weeks) significantly exceeded the preinfection numbers (P<.03). Moreover, the numbers of IL-1beta, IL-6, and TNF-alpha mRNA molecules in persistently infected animals (6 years) also were elevated compared with preinfection numbers (P<.02, P=.03, P=.16, respectively). Cytokine gene expression coincided with progressive H. pylori gastritis, confirmed by increased gastritis scores over preinfection scores (P<.005). These findings provide quantitative evidence that H. pylori induces local gene expression of monocyte/macrophage-derived inflammatory cytokines and evokes an innate response in gastric tissue of nonhuman primates.

Helicobacter pylori adhesion to carbohydrates.

Adherence of bacterial pathogens to host tissues contributes to colonization and virulence and typically involves specific interactions between bacterial proteins called adhesins and cognate oligosaccharide (glycan) or protein motifs in the host that are used as receptors. A given pathogen may have multiple adhesins, each specific for a different set of receptors and, potentially, with different roles in infection and disease. This chapter provides strategies for identifying and analyzing host glycan receptors and the bacterial adhesins that exploit them as receptors, with particular reference to adherence of the gastric pathogen Helicobacter pylori.

Rhesus monkey gastric mucins: oligomeric structure, glycoforms and Helicobacter pylori binding.

Mucins isolated from the stomach of Rhesus monkey are oligomeric glycoproteins with a similar mass, density, glycoform profile and tissue localization as human MUC5AC and MUC6. Antibodies raised against the human mucins recognize those from monkey, which thus appear to be orthologous to those from human beings. Rhesus monkey muc5ac and muc6 are produced by the gastric-surface epithelium and glands respectively, and occur as three distinct glycoforms. The mucins are substituted with the histo blood-group antigens B, Le(a) (Lewis a), Le(b), Le(x), Le(y), H-type-2, the Tn-antigen, the T-antigen, the sialyl-Le(x) and sialyl-Le(a) structures, and the expression of these determinants varies between individuals. At neutral pH, Helicobacter pylori strains expressing BabA (blood-group antigen-binding adhesin) bind Rhesus monkey gastric mucins via the Le(b) or H-type-1 structures, apparently on muc5ac, as well as on a smaller putative mucin, and binding is inhibited by Le(b) or H-type-1 conjugates. A SabA (sialic acid-binding adhesin)-positive H. pylori mutant binds to sialyl-Le(x)-positive mucins to a smaller extent compared with the BabA-positive strains. At acidic pH, the microbe binds to mucins substituted by sialylated structures such as sialyl-Le(x) and sialylated type-2 core, and this binding is inhibited by DNA and dextran sulphate. Thus mucin- H. pylori binding occurs via at least three different mechanisms: (1) BabA-dependent binding to Le(b) and related structures, (2) SabA-dependent binding to sialyl-Le(x) and (3) binding through a charge-mediated mechanism to sialylated structures at low pH values.