Immune Evasion by Helicobacter pylori: Gastric Spiral Bacteria Lack Surface Immunoglobulin Deposition and Reactivity with Homologous Antibodies

Background. Helicobacter pylori infection persists in the presence of potent serum and gastric mucosal anti-body responses against bacterial antigens. The aim of this article is to report on a study determine whether there is antibody deposition on H. pylori in vivo in the stomach of infected patients and whether gastric and cultured forms of H. pylori differ in their antibody reactivity.
Materials and Methods. Serum, gastric biopsies, and antral brushings were obtained from 10 patients having endoscopy. H. pylori was cultured from gastric biopsies. Bacterial samples were stained directly for immunoglobulin deposition and indirectly using rabbit antiurease serum or patient serum. Samples were examined by immunofluorescence microscopy and flow cytometry.
Results. Although spiral bacteria could be identified easily by acridine orange staining and antiurease staining of gastric brushings from H. pylori infected patients, gastric bacteria did not have detectable IgG or IgA present, and only one of five samples could be stained for IgG and IgA indirectly using patient serum. In contrast, cultured bacteria could be stained readily with homologous serum for IgG and IgA in the majority of cases. Low pH inhibited immunoglobulin reactivity with cultured H. pylori.
Conclusions. Gastric H. pylori may evade humoral defense owing to poor deposition of immunoglobulin in the gastric environment or failure to express surface antigens that are present on cultured forms of H. pylori.     

Mixed Infection with cagA-Positive and cagA-Negative Strains of Helicobacter pylori

Background Helicobacter pylori infection has been implicated strongly in the pathogenesis of gastritis, peptic ulcer disease, gastric adenocarcinoma, and gastric lymphoma, but the reasons for these widely different clinical outcomes are unknown. The aim of this study was to determine whether these differences could be due in part to mixed infection in the same individual, with bacteria having differences in pathogenic factors associated with ulcers.
Materials and Methods. The cagA gene of H. pylori was used to test for mixed infection because it is present in only some strains, and its presence has been associated with ulcers. Polymerase chain reaction (PCR) assays for the cagA gene were applied to H. pylori culture isolates and endoscopic gastric aspirates. Individual bacterial clones were tested for genetic similarity by random primer amplification and restriction endonuclease digestion of urease gene PCR products.
Results. The majority of H. pylori -positive patients had strongly cagA -positive culture isolates and endoscopic samples (62.5% and 69.6%, respectively). However, many of these patients had evidence of mixed infection with cagA negative and cagA positive strais in cultures isolates and endoscopic samples (25% and 17.4%, respectively). Mixed infection was found to be due to genetically unrelated strains in two patients in whom genetic analysis was performed.
Conclusion. Mixed infection with differences in substrain pathogenic factors might occur in H. pylori infection and might contribute to differences in clinical outcome.     

Limits to Catalysis by Ribonuclease A

Bovine pancreatic ribonuclease A (RNase A) catalyzes the cleavage of the P-O(5') bond in RNA. Although this enzyme has been the object of much landmark work in bioorganic chemistry, the nature of its rate-limiting transition state and its catalytic rate enhancement had been unknown. Here, the value of k(cat)/K(m) for the cleavage of UpA by wild-type RNase A was found to be inversely related to the concentration of added glycerol. In contrast, the values of k(cat)/K(m) for the cleavage of UpA by a sluggish mutant of RNase A and the cleavage of the poor substrate UpOC(6)H(4)-p-NO(2) by wild-type RNase A were found to be independent of glycerol concentration. Yet, UpA cleavage by the wild-type and mutant enzymes was found to have the same dependence on sucrose concentration, indicating that catalysis of UpA cleavage by RNase A is limited by desolvation. The rate of UpA cleavage by RNase A is maximal at pH 6.0, where k(cat) = 1.4 × 10(3) s(-1) and k(cat)/K(m) = 2.3 × 10(6) M(-1)s(-1) at 25°C. At pH 6.0 and 25°C, the uncatalyzed rate of [5,6-(3)H]Up[3,5,8-(3)H]A cleavage was found to be k(uncat) = 5 × 10(-9) s(-1) (t(1/2) = 4 years). Thus, RNase A enhances the rate of UpA cleavage by 3 × 10(11)-fold by binding to the transition state for P-O(5') bond cleavage with a dissociation constant of <2 × 10(-15) M.     

Morphological responses by Bosmina longirostris and Eubosmina tubicen to changes in copepod predator populations during a whole-lake acidification experiment

Changes in zooplankton populations during the experimental acidilicationof Little Rock Lake provided an opportunity to examine specificmechanisms underlying the morphological responses of bosminidsto changing predation pressure. Two large copepods, Epischuralacustris and Mesocyclops edax, disappeared from the lake'sacidified basin in 1986 and 1989, respectively, while a smallercopepod predator, Tropocyclops extensus, increased during laterstages of acidification. The two bosminid species showed distinctlydifferent responses coinciding with the changes in copepod predation.Bosmina longirostris exhibited a significant decrease in mucrolength with the decline of M.edax and E.lacustris. Its meanbody and antennule length, however, did not change. We suggestthat the decoupling of B.longirostris mucro length and antennulelength may have been related to the persistence of the smallercopepod predator, T.extensus. Eubosmina tubicen showed no apparentresponse to declines in M.edax and E.lacustris abundance ineither mean mucro, antennule or body length. Allometric analysesindicated, however, that mucro length was related to size-dependentcopepod predation for both B.longirostris and E.tubicen.     

Isotopic fractionation during nitrate uptake by phytoplankton grown in continuous culture

The isotopic fractionation associated with uptake of NO₃^–by six species of phytoplankton (two diatoms, one cryptophyte,one chlorophyte and two haptophytes) was measured at a varietyof steady-state growth rates in nitrogen-limited continuousculture. The magnitude of the isotopic fractionation factor(     

Structure-function relationships of the alternative oxidase of plant mitochondria: A model of the active site

A major characteristic of plant mitochondria is the presence of a cyanide-insensitive alternative oxidase which catalyzes the reduction of oxygen to water. Current information on the properties of the oxidase is reviewed. Conserved amino acid motifs have been identified which suggest the presence of a hydroxo-bridged di-iron center in the active site of the alternative oxidase. On the basis of sequence comparison with other di-iron center proteins, a structural model for the active site of the alternative oxidase has been developed that has strong similarity to that of methane monoxygenase. Evidence is presented to suggest that the alternative oxidase of plant mitochondria is the newest member of the class II group of di-iron center proteins.     

URF13, a ligand-gated,pore-forming receptor for T-toxin in the inner membrane ofcms-T mitochondria

URF13 is the product of a mitochondrial-encoded gene (T-urfl3) found only in maize plants containing the Texas male-sterile cytoplasm (cms-T), and it is thought to be responsible for both cytoplasmic male sterility and the susceptibility ofcms-T maize to the fungal pathogensBipolaris maydis race T andPhyllosticta maydis. Mitochondria isolated fromcms-T maize are uniquely sensitive to pathotoxins (T-toxin) produced by these fungi and to methomyl (a commercial insecticide). URF13 acts as a receptor that specifically binds T-toxin to produce hydrophilic pores in the inner mitochondrial membrane. When expressed inEscherichia coli cells, URF13 also forms hydrophilic pores in the plasma membrane if exposed to T-toxin or methomyl. Topological studies established that URF13 contains three membrane-spanning -helices, two of which are amphipathic and can contribute to pore formation. Chemical crosslinking of URF13 was used to demonstrate the existence of URF13 oligomers incms-T mitochondria andE. coli cells. The ability of the carboxylate-specific reagent,N,N-dicyclohexycarbodiimide, to cross-link URF13 was used in conjunction with site-directed mutagenesis to establish that the URF13 tetramer has a central core consisting of a four--helical bundle which undergoes a conformational change after interaction with T-toxin or methomyl. Overall, the experimental evidence indicates that URF13 functions as a ligand-gated, pore-forming T-toxin receptor incms-T mitochondria.     

Nerve Growth Factor as a Mitogen for a Pancreatic Carcinoid Cell Line

Abstract: Carcinoid tumors are a group of neuroendocrine neoplasms distributed widely throughout the body but most commonly occurring in the gut. These tumors retain many characteristics of their neural crest origin, including secretion of neuroactive peptides and responsiveness to neurotrophic substances. Nerve growth factor (NGF), a neurotrophic protein involved in maintenance and differentiation of peripheral sympathetic and sensory neurons, regulates growth of several neural tumor cells by inducing a differentiated phenotype and subsequent inhibition of cell growth rate. We examined the actions of NGF in a functioning human pancreatic carcinoid cell line (termed BON). NGF has no effect on the cytoarchitecture or constitutive secretion of bioamines in this carcinoid cell line. NGF, however, stimulates the in vitro cellular proliferation of BON cells. BON cells possess mRNA for the NGF receptors (p75^LNGFR and p140^trkA) and membrane-associated tyrosine kinase activity is increased in response to NGF. Both the mitogenic activity of NGF, as well as the receptor-linked tyrosine kinase activity, can be abrogated in BON cells by the trkA inhibitor K-252a and specific anti-NGF antibody. Our studies demonstrate that NGF is a mitogen for this carcinoid cell line without effect on cellular phenotype or cytoarchitecture. NGF may play a role in the development and progression of human carcinoid tumors.