apyrase and Schistosomiasis-mansoni

Granulomas are inflammatory tissue responses directed to a set of antigens. Trapped Schistosoma mansoni eggs promote productive granulomas in the tissues, and they are the main damage caused by schistosomiasis. Some S. mansoni antigenic proteins may have a direct involvement in the resolution of the granulomatous response. The ATP diphosphohydrolases isoforms of this parasite are immunogenic, expressed in all phases of the parasite life cycle and secreted by eggs and adult worms. Potato apyrase is a vegetable protein that cross-reactive with parasite ATP diphosphohydrolases isoforms. In this study, the vegetable protein was purified, before being inoculated in C57BL/6 mice that were later infected with cercariae. Sixty days after infection, adult worms were recovered, antibodies and cytokines were measured, and morphological granuloma alterations evaluated. Immunization of the animals induced significant levels of IgG and IgG1 antibodies and IFN-γ, IL-10 and IL-5 cytokines, but not IL-13, suggesting that potato apyrase is an immunoregulatory protein. Supporting this hypothesis, it was found that liver damage associated with schistosomiasis was mitigated, reducing the size of the areas affected by granuloma to 35% and increasing the presence of multinucleated giant cells in this environment. In conclusion, potato apyrase was found to be effective immunomodulatory antigen for murine schistosomiasis.

Topics: Animals; Apyrase; Female; Giant Cells; Mice; Mice, Inbred C57BL; Rodent Diseases; Schistosoma mansoni; Schistosomiasis mansoni; Solanum tuberosum

High percentages of structural identity and cross-immunoreactivity have been reported between potato apyrase and Schistosoma mansoni ATP diphosphohydrolase (SmATPDases) isoforms, showing the existence of particular epitopes shared between these proteins.. Potato apyrase was employed using ELISA, western blot, and mouse immunization methods to verify IgE reactivity.. Most of the schistosomiasis patient's (75%) serum was seropositive for potato apyrase and this protein was recognized using western blotting, suggesting that parasite and plant proteins share IgE-binding epitopes. C57BL/6 mice immunized with potato apyrase showed increased IgE antibody production.. Potato apyrase and SmATPDases have IgE-binding epitopes.

Topics: Animals; Antibodies, Helminth; Apyrase; Blotting, Western; Cross Reactions; Enzyme-Linked Immunosorbent Assay; Epitopes; Female; Immunoglobulin E; Mice, Inbred C57BL; Schistosoma mansoni; Schistosomiasis mansoni; Solanum tuberosum

Schistosomes are intravascular parasitic platyhelminthes infecting > 200 million people globally and causing a debilitating disease, schistosomiasis. Despite the relatively large size of the adult worms and their disruption of blood flow, surprisingly, they do not appear to provoke thrombus formation around them in vivo. We hypothesize that proteins expressed at the host-parasite interface are key to this ability. Here, we functionally express an ectonucleotide pyrophosphatase/phosphodiesterase homologue, SmNPP5, that is expressed at the tegumental surface of intravascular

Topics: Adenosine Diphosphate; Animals; Apyrase; Blood Platelets; Blood Vessels; Female; Helminth Proteins; Humans; Male; Mice; Phosphoric Diester Hydrolases; Platelet Aggregation Inhibitors; Pyrophosphatases; Schistosoma mansoni; Schistosomiasis mansoni; Virulence Factors

In this study, we evaluated the in vitro and in vivo schistosomicidal activities of chalcones against Schistosoma mansoni worms. In vitro assays revealed that chalcones 1 and 3 were the most active compounds, without affecting significantly mammalian cells. Confocal laser scanning microscopy and scanning electron microscopy studies revealed reduction on the numbers of tubercles and morphological alterations in the tegument of S. mansoni worms after in vitro incubation with chalcones 1 and 3. In a mouse model of schistosomiasis, the oral treatment (400 mg/kg) with chalcone 1 or 3 significantly caused a total worm burden reduction in mice. Chalcone 1 showed significant inhibition of the S. mansoni ATP diphosphohydrolase activity, which was corroborated by molecular docking studies. The results suggested that chalcones could be explored as lead compounds with antischistosomal properties.

Topics: Administration, Oral; Animals; Anthelmintics; Apyrase; Binding Sites; Chalcones; Disease Models, Animal; Helminth Proteins; Mice; Microscopy, Confocal; Microscopy, Electron, Scanning; Molecular Docking Simulation; Protein Structure, Tertiary; Schistosoma mansoni; Schistosomiasis mansoni; Structure-Activity Relationship

Topics: Amino Acid Sequence; Animals; Apyrase; Cross Reactions; Epitopes; Host-Parasite Interactions; Immunoglobulin E; Mice, Inbred BALB C; Peptides; Schistosoma mansoni; Schistosomiasis mansoni; Sequence Homology, Amino Acid

Schistosomes are parasitic worms that can live in the bloodstream of their vertebrate hosts for many years. It has been proposed that the worms impinge on host purinergic signalling by degrading proinflammatory molecules like ATP as well as prothrombotic mediators like ADP. This capability may help explain the apparent refractoriness of the worms to both immune elimination and thrombus formation. Three distinct ectoenzymes, expressed at the host-exposed surface of the worm's tegument, are proposed to be involved in the catabolism of ATP and ADP. These are alkaline phosphatase (SmAP), phosphodiesterase (SmNPP-5), and ATP diphosphohydrolase (SmATPDase1). It has recently been shown that only one of these enzymes-SmATPDase1-actually degrades exogenous ATP and ADP. However, a second ATP diphosphohydrolase homolog (SmATPDase2) is located in the tegument and has been reported to be released by the worms. It is possible that this enzyme too participates in the cleavage of exogenous nucleotide tri- and di-phosphates. To test this hypothesis, we employed RNA interference (RNAi) to suppress the expression of the schistosome SmATPDase1 and SmATPDase2 genes. We find that only SmATPDase1-suppressed parasites are significantly impaired in their ability to degrade exogenously added ATP or ADP. Suppression of SmATPDase2 does not appreciably affect the worms' ability to catabolize ATP or ADP. Furthermore, we detect no evidence for the secretion or release of an ATP-hydrolyzing activity by cultured parasites. The results confirm the role of tegumental SmATPDase1, but not SmADTPDase2, in the degradation of the exogenous proinflammatory and prothrombotic nucleotides ATP and ADP by live intravascular stages of the parasite.

Topics: Adenosine Diphosphate; Adenosine Triphosphate; Animals; Apyrase; Host-Parasite Interactions; Isoenzymes; Molecular Sequence Data; Reverse Transcriptase Polymerase Chain Reaction; RNA, Small Interfering; Schistosoma mansoni; Schistosomiasis mansoni; Transcriptome; Transfection

In this paper, we showed for the first time that the conserved domains within Schistosoma mansoni ATP diphosphohydrolase isoforms, shared with potato apyrase, possess epitopes for the IgG1 and IgG4 subtypes, as 24 (80%) of the 30 schistosomiasis patients were seropositive for this vegetable protein. The analyses for each patient cured (n = 14) after treatment (AT) with praziquantel revealed variable IgG1 and IgG4 reactivity against potato apyrase. Different antigenic epitopes shared between the vegetable and parasite proteins could be involved in susceptibility or resistance to S. mansoni AT with praziquantel and these possibilities should be explored.

Topics: Adolescent; Adult; Aged; Aged, 80 and over; Animals; Anthelmintics; Antibodies, Helminth; Apyrase; Child; Child, Preschool; Cross Reactions; Humans; Immunoglobulin G; Middle Aged; Praziquantel; Schistosoma mansoni; Schistosomiasis mansoni; Solanum tuberosum; Young Adult

Schistosoma mansoni ATP diphosphohydrolase isoforms and potato apyrase share conserved epitopes. By enzyme-linked immunosorbent assays, elevated levels of IgM, IgG2a and IgG1 antibody reactivity against potato apyrase were observed in S. mansoni-infected BALB/c mice during the acute phase of infection, while only IgM and IgG1 antibody reactivity levels maintained elevated during the chronic phase of infection. Antibody reactivity against potato apyrase was monitored over an 11-month period in chronically-infected mice treated with oxamniquine. Eleven months later, the level of seropositive IgM decreased significantly (approximately 30%) compared to the level found in untreated, infected mice. The level of seropositive IgG1 decreased significantly four months after treatment (MAT) (61%) and remained at this level even after 11 months. The IgG2a reactivity against potato apyrase, although unchanged during chronic phase to 11 MAT, appeared elevated again in re-infected mice suggesting a response similar to that found during the acute phase. BALB/c mouse polyclonal anti-potato apyrase IgG reacted with soluble egg antigens probably due to the recognition of parasite ATP diphosphohydrolase. This study, for the first time, showed that the IgG2a antibody from S. mansoni-infected BALB mice cross-reacts with potato apyrase and the level of IgG2a in infected mice differentiates disease phases. The results also suggest that different conserved-epitopes contribute to the immune response in schistosomiasis.

Topics: Acute Disease; Animals; Anthelmintics; Antibodies, Helminth; Antigens, Helminth; Apyrase; Chronic Disease; Cross Reactions; Enzyme-Linked Immunosorbent Assay; Female; Immunoglobulin G; Immunoglobulin M; Mice; Mice, Inbred BALB C; Oxamniquine; Schistosoma mansoni; Schistosomiasis mansoni; Solanum tuberosum

An immunosensor for detecting the antibody anti-apyrase of Schistosoma mansoni based on rigid composite materials, containing graphite powder and epoxy resins, developed in this work, is described. A surface modification strategy for the use of oxidized graphite in the detection of antibody-antigen interaction was developed. This modification strategy is based on silanization of conductive composite. First, the graphite powder-epoxy resin was treated with concentrated hydrogen peroxide to improve surface hydroxyl groups and to form a hydrophilic layer. Second, 3- aminopropyltriethoxysilane was subsequently used to functionalize the treated surface to form amino groups, which were further activated with glutaraldehyde to introduce a layer of aldehyde groups. Contact angle microscopy and scanning electron microscopy were used as a qualitative analysis of the deposition of silane on the surface of the sensor. The effectiveness of the modification strategy was validated by amperometric immunoassays of S. mansoni. Amperometric signals related to concentrations of this immobilized protein were observed, and the effects of pH and incubation times were analyzed. This surface modification strategy provides a platform on which proteins can be directly immobilized for immunosensor and protein array applications.

Topics: Animals; Antibodies, Protozoan; Apyrase; Buffers; Electrochemistry; Electrodes; Enzyme-Linked Immunosorbent Assay; Enzymes, Immobilized; Epoxy Compounds; Graphite; Hot Temperature; Humans; Hydrogen Peroxide; Hydrogen-Ion Concentration; Immunochemistry; Indicators and Reagents; Microscopy, Electron, Scanning; Schistosoma mansoni; Schistosomiasis mansoni; Serology; Temperature

We have previously showed that Schistosoma mansoni ATP-diphosphohydrolase and Solanum tuberosum potato apyrase share epitopes and the vegetable protein has immunostimulatory properties. Here, it was verified the in situ cross-immunoreactivity between mice NTPDases and anti-potato apyrase antibodies produced in rabbits, using confocal microscopy. Liver samples were taken from Swiss Webster mouse 8 weeks after infection with S. mansoni cercariae, and anti-potato apyrase and TRITC-conjugated anti-rabbit IgG antibody were tested on cryostat sections. The results showed that S. mansoni egg ATP diphosphohydrolase isoforms, developed by anti-potato apyrase, are expressed in miracidial and egg structures, and not in granulomatous cells and hepatic structures (hepatocytes, bile ducts, and blood vessels). Therefore, purified potato apyrase when inoculated in rabbit generates polyclonal sera containing anti-apyrase antibodies that are capable of recognizing specifically S. mansoni ATP diphosphohydrolase epitopes, but not proteins from mammalian tissues, suggesting that autoantibodies are not induced during potato apyrase immunization. A phylogenetic tree obtained for the NTPDase family showed that potato apyrase had lower homology with mammalian NTPDases 1-4, 7, and 8. Further analysis of potato apyrase epitopes could implement their potential use in schistosomiasis experimental models.

Topics: Adenosine Triphosphatases; Amino Acid Sequence; Animals; Antibodies, Helminth; Apyrase; Cross Reactions; Disease Models, Animal; Male; Mice; Microscopy, Confocal; Molecular Sequence Data; Rabbits; Schistosoma mansoni; Schistosomiasis mansoni; Solanum tuberosum

An ATP-diphosphohydrolase (EC 3.6.1.5) was identified in the tegumental fraction isolated from Schistosoma mansoni worms. Both ATP and ADP were hydrolyzed to AMP at similar rates by the enzyme. Other nucleotides were also degraded by the tegument enzyme, revealing a broad substrate specificity. Electrophoretic separation of tegumental proteins under non-denaturing conditions followed by addition of ATP or ADP as substrate revealed a single band of activity with similar mobility. In addition, similar heat-inactivation profiles were obtained for ATPase or ADPase activities, indicating that a single enzyme is responsible for degrading both nucleotides. The enzyme was not inhibited by vanadate, levamisole, tetramisole, ouabain or sodium azide. The ADPase activity was not affected by adenosine (5')-pentaphospho-(5')-adenosine (Ap5A) or by an excess of glucose and hexokinase used as an ATP-trapping system, thus excluding the presence of any significant adenylate kinase activity. The ATP-diphosphohydrolase displayed micromolar affinities for both Mg2+ and Ca2+, and the calcium-activated enzyme was inhibited by millimolar Mg2+. In intact live worms a calcium phosphate precipitate was formed on the outer tegumental surface upon incubation of the worms with either ATP or ADP, indicating the ectolocalization of this enzyme. In addition, ultrastructural histochemical localization of the enzyme was obtained. A distinct deposition of lead phosphate granules on the outer surface of the tegument was observed by electron microscopy, in the presence of either ATP or ADP as substrate. It is suggested that the ATP-diphosphohydrolase could regulate the concentration of purine nucleotides around the parasites and hence enable them to escape the host hemostasis by preventing ADP-induced platelet activation.

Topics: Adenosine Diphosphate; Adenosine Triphosphate; Animals; Apyrase; Blood Platelets; Cricetinae; Histocytochemistry; Humans; Mesocricetus; Microscopy, Electron; Platelet Activation; Schistosoma mansoni; Schistosomiasis mansoni; Substrate Specificity