Spermine causes loss of innate immune response to Helicobacter pylori by inhibition of inducible nitric-oxide synthase translation

Helicobacter pylori infection of the stomach elicits a vigorous but ineffective host immune and inflammatory response, resulting in persistence of the bacterium for the life of the host. We have reported that in macrophages, H. pylori up-regulates inducible NO synthase (iNOS) and antimicrobial NO production, but in parallel there is induction of arginase II, generating ornithine, and of ornithine decarboxylase (ODC), generating polyamines. Spermine, in particular, has been shown to restrain immune response in activated macrophages by inhibiting proinflammatory gene expression. We hypothesized that spermine could prevent the antimicrobial effects of NO by inhibiting iNOS in macrophages activated by H. pylori. Spermine did not affect the up-regulation of iNOS mRNA levels but in a concentration-dependent manner significantly attenuated iNOS protein levels and NO production. Reduction in iNOS protein was due to inhibition of iNOS translation and not due to iNOS degradation. ODC knockdown with small interfering (si) RNA resulted in increased H. pylori-stimulated iNOS protein expression and NO production without altering iNOS mRNA levels. When macrophages were cocultured with H. pylori, killing of bacteria was enhanced by transfection of ODC siRNA and prevented by addition of spermine. These results identify a mechanism of immune dysregulation induced by H. pylori in which stimulated spermine synthesis by the arginase-ODC pathway inhibits iNOS translation and NO production, leading to persistence of the bacterium and risk for peptic ulcer disease and gastric cancer.     

Identification of tyrosine phosphatases that dephosphorylate the insulin receptor. A brute force approach based on "substrate-trapping" mutants

Many pharmacologically important receptors, including all cytokine receptors, signal via tyrosine (auto)phosphorylation, followed by resetting to their original state through the action of protein tyrosine phosphatases (PTPs). Establishing the specificity of PTPs for receptor substrates is critical both for understanding how signaling is regulated and for the development of specific PTP inhibitors that act as ligand mimetics. We have set up a systematic approach for finding PTPs that are specific for a receptor and have validated this approach with the insulin receptor kinase. We have tested nearly all known human PTPs (45) in a membrane binding assay, using "substrate-trapping" PTP mutants. These results, combined with secondary dephosphorylation tests, confirm and extend earlier findings that PTP-1b and T-cell PTP are physiological enzymes for the insulin receptor kinase. We demonstrate that this approach can rapidly reduce the number of PTPs that have a particular receptor or other phosphoprotein as their substrate.     

Update on paramyosin in parasitic worms

Paramyosin was first identified as a structural component of invertebrate muscle. Analysis of crude, native, adult schistosome worm preparations identified a highly immunogenic protein which was later identified as paramyosin. Early vaccination/challenge studies with native paramyosin produced encouraging levels of protective efficacy against schistosomes, which led to the question as to how a sub-tegumental (muscular) protein could provide a target for vaccine-mediated immunological attack. Immunolocalisation studies of schistosomes confirmed the presence of paramyosin within the post-acetabular glands of cercariae and on the tegumental surface of lung schistosomula. Here we present an update on the more recent research on paramyosin in parasitic worms that has focused primarily in two directions: (i) further testing of the vaccine potency of paramyosin against schistosomes and other parasitic worms; and (ii) characterisation of the protein at the molecular and biochemical levels.     

Selective blockade of neurokinin (NK)(1) receptors facilitates the activity of adrenergic pathways projecting to frontal cortex and dorsal hippocampus in rats

The selective NK(1) receptor antagonist, GR205,171 (2.5-40.0 mg/kg, i.p.), dose-dependently elevated dialysate levels of noradrenaline (NA), but not serotonin (5-HT), in the frontal cortex of freely moving rats. This action was exerted stereospecifically inasmuch as its less active isomer, GR226,206, was ineffective. In the dorsal hippocampus, GR205,171 (but not GR226,206) also significantly increased dialysate levels of NA, whereas levels of 5-HT were unaffected. Further, in anaesthetized rats, GR205,171 dose-dependently (1.0-4.0 mg/kg, i.v.) increased the firing rate of adrenergic perikarya in the locus coeruleus. In contrast, their activity was not modified by GR226,206. These findings indicate that selective blockade of NK(1) receptors enhances the activity of ascending adrenergic pathways in rats. Adrenergic mechanisms may, thus, be involved in the potential antidepressant and other functional properties of NK(1) receptor antagonists.     

The role of polyamines in the regulation of macrophage polarization and function

Amino Acids - Naturally occurring polyamines are ubiquitously distributed and play important roles in cell development, amino acid and protein synthesis, oxidative DNA damage, proliferation, and...     

C, N, P concentrations and requirements of flowering Posidonia oceanica

The carbon, nitrogen and phosphorus contents in flowering and nonflowering shoots were compared after an important flowering event occurred in the Posidonia meadow of the Bay of Calvi. The flower formation caused a significant increase of C and a significant decrease of N concentrations in intermediate and adult leaves. Minimum daily requirements in mgshoot^-1day^-1 of 3.4 and 4.8 of C, 0.09 and 0.09 N, 0.01 and 0.02 of P respectively for nonflowering and flowering shoots were calculated. It shows that additional quantities of C and P are required for the inflorescence elaboration. The unchanged quantity of N required by the shoot for the inflorescence elaboration and the significant modification of N concentration in intermediate and adult leaves suggests that N is limited in the environment and that an efficient resorption of N occurs from leaves to ensure the inflorescence formation.     

Real-time PCR Demonstrates High Prevalence of Schistosoma japonicum in the Philippines: Implications for Surveillance and Control

Background

The Philippines has a population of approximately 103 million people, of which 6.7 million live in schistosomiasis-endemic areas with 1.8 million people being at risk of infection with Schistosoma japonicum. Although the country-wide prevalence of schistosomiasis japonica in the Philippines is relatively low, the prevalence of schistosomiasis can be high, approaching 65% in some endemic areas. Of the currently available microscopy-based diagnostic techniques for detecting schistosome infections in the Philippines and elsewhere, most exhibit varying diagnostic performances, with the Kato-Katz (KK) method having particularly poor sensitivity for detecting low intensity infections. This suggests that the actual prevalence of schistosomiasis japonica may be much higher than previous reports have indicated.

Methodology/Principal Findings

Six barangay (villages) were selected to determine the prevalence of S. japonicum in humans in the municipality of Palapag, Northern Samar. Fecal samples were collected from 560 humans and examined by the KK method and a validated real-time PCR (qPCR) assay. A high S. japonicum prevalence (90.2%) was revealed using qPCR whereas the KK method indicated a lower prevalence (22.9%). The geometric mean eggs per gram (GMEPG) determined by the qPCR was 36.5 and 11.5 by the KK. These results, particularly those obtained by the qPCR, indicate that the prevalence of schistosomiasis in this region of the Philippines is much higher than historically reported.

Conclusions/Significance

Despite being more expensive, qPCR can complement the KK procedure, particularly for surveillance and monitoring of areas where extensive schistosomiasis control has led to low prevalence and intensity infections and where schistosomiasis elimination is on the horizon, as for example in southern China.     

An Immunomics Approach to Schistosome Antigen Discovery: Antibody Signatures of Naturally Resistant and Chronically Infected Individuals from Endemic Areas

Schistosomiasis is a neglected tropical disease that is responsible for almost 300,000 deaths annually. Mass drug administration (MDA) is used worldwide for the control of schistosomiasis, but chemotherapy fails to prevent reinfection with schistosomes, so MDA alone is not sufficient to eliminate the disease, and a prophylactic vaccine is required. Herein, we take advantage of recent advances in systems biology and longitudinal studies in schistosomiasis endemic areas in Brazil to pilot an immunomics approach to the discovery of schistosomiasis vaccine antigens. We selected mostly surface-derived proteins, produced them using an in vitro rapid translation system and then printed them to generate the first protein microarray for a multi-cellular pathogen. Using well-established Brazilian cohorts of putatively resistant (PR) and chronically infected (CI) individuals stratified by the intensity of their S. mansoni infection, we probed arrays for IgG subclass and IgE responses to these antigens to detect antibody signatures that were reflective of protective vs. non-protective immune responses. Moreover, probing for IgE responses allowed us to identify antigens that might induce potentially deleterious hypersensitivity responses if used as subunit vaccines in endemic populations. Using multi-dimensional cluster analysis we showed that PR individuals mounted a distinct and robust IgG1 response to a small set of newly discovered and well-characterized surface (tegument) antigens in contrast to CI individuals who mounted strong IgE and IgG4 responses to many antigens. Herein, we show the utility of a vaccinomics approach that profiles antibody responses of resistant individuals in a high-throughput multiplex approach for the identification of several potentially protective and safe schistosomiasis vaccine antigens.