WorldWideScience

Sample records for agglutinins

  1. Febrile/cold agglutinins

    Science.gov (United States)

    ... diagnose certain infections and find the cause of hemolytic anemia (a type of anemia that occurs when red ... or cold agglutinins can help explain why the hemolytic anemia is occurring and direct treatment.

  2. Salivary agglutinin/glycoprotein-340/DMBT1

    DEFF Research Database (Denmark)

    Ligtenberg, Antoon J M; Veerman, Enno C I; Nieuw Amerongen, Arie V;

    2007-01-01

    Salivary agglutinin (SAG), lung glycoprotein-340 (gp-340) and Deleted in Malignant Brain Tumours 1 (DMBT1) are three names for identical proteins encoded by the dmbt1 gene. DMBT1/SAG/gp-340 belongs to the scavenger receptor cysteine-rich (SRCR) superfamily of proteins, a superfamily of secreted o...

  3. Purification and Characterization of Abrus precatorius Agglutinin

    OpenAIRE

    Absar, Nural; Funatsu, Gunki

    1984-01-01

    Abrus pvecatorius agglutinin (APA) has been purified by a new purification procedure from the seeds of semen jequiriti produced in Bangladesh and Taiwan. The method was accomplished by 33-50% saturation ammonium sulfate fraction from 1% acetic acid extract of the seeds of semen jequiriti using gel filtration on Sephadex G-75 followed by DEAE-cellulose column chromatography. The molecular weight was estimated to be 126,000 and 122,000 by gel filtration on Sephadex G-150 for Bangladesh-APA and ...

  4. Occurrence of granulocyte cytotoxins and agglutinins.

    Science.gov (United States)

    Hasegawa, T; Bergh, O J; Terasaki, P I; Graw, R G

    1975-01-01

    Granulocyte cytotoxic activity in sera from over 257 patients was shown to be distinct from HL-A lymphocytoxic activity. Granulocyte cytotoxins occur in approximately 25 per cent of sera from patients having leukemia, 45 per cent with aplastic anemia, 22 per cent with kidney disease on hemodialysis, and 19 per cent of pregnant women. By testing sera on the same panel of cells, the granulocyte cytotoxic activity was shown not to be associated with granulocyte agglutination activity or lymphocytotoxic acitivty. It is likely that granulocyte cytotoxins and granulocyte agglutinins will be useful in transfusion and bone marrow transplantation as a separate tool from the more widely used lymphocyte cytotoxicity reaction. PMID:1129831

  5. Antibody interactions with Ricinus communis agglutinins studied by biolayer interferometry

    Science.gov (United States)

    Two related agglutinins are present in the seeds of Ricinus communis (castor): ricin, a dichain ribosome-inactivating protein and Ricinus communis agglutinin-1 (RCA-1), a much less toxic hemagglutinin. Because ricin has been used for experimental cancer chemotherapy as well as for intentional poison...

  6. Llama-Derived Single Domain Antibodies Specific for Abrus Agglutinin

    Directory of Open Access Journals (Sweden)

    James P. Carney

    2011-11-01

    Full Text Available Llama derived single domain antibodies (sdAb, the recombinantly expressed variable heavy domains from the unique heavy-chain only antibodies of camelids, were isolated from a library derived from llamas immunized with a commercial abrin toxoid preparation. Abrin is a potent toxin similar to ricin in structure, sequence and mechanism of action. The selected sdAb were evaluated for their ability to bind to commercial abrin as well as abrax (a recombinant abrin A-chain, purified abrin fractions, Abrus agglutinin (a protein related to abrin but with lower toxicity, ricin, and unrelated proteins. Isolated sdAb were also evaluated for their ability to refold after heat denaturation and ability to be used in sandwich assays as both capture and reporter elements. The best binders were specific for the Abrus agglutinin, showing minimal binding to purified abrin fractions or unrelated proteins. These binders had sub nM affinities and regained most of their secondary structure after heating to 95 °C. They functioned well in sandwich assays. Through gel analysis and the behavior of anti-abrin monoclonal antibodies, we determined that the commercial toxoid preparation used for the original immunizations contained a high percentage of Abrus agglutinin, explaining the selection of Abrus agglutinin binders. Used in conjunction with anti-abrin monoclonal and polyclonal antibodies, these reagents can fill a role to discriminate between the highly toxic abrin and the related, but much less toxic, Abrus agglutinin and distinguish between different crude preparations.

  7. Complement activation by salivary agglutinin is secretor status dependent

    NARCIS (Netherlands)

    S.T.G. Gunput; A.J.M. Ligtenberg; B. Terlouw; M. Brouwer; E.C.I. Veerman; D. Wouters

    2015-01-01

    After mucosal damage or gingival inflammation, complement proteins leak into the oral cavity and mix with salivary proteins such as salivary agglutinin (SAG/gp-340/DMBT1). This protein is encoded by the gene Deleted in Malignant Brain Tumors 1 (DMBT1), and it aggregates bacteria, viruses and fungi,

  8. Fatal cold agglutinin-induced haemolytic anaemia: a case report

    Directory of Open Access Journals (Sweden)

    Reverberi Roberto

    2010-08-01

    Full Text Available Abstract Introduction Cold agglutinin disease usually develops as a result of the production of a specific immunoglobulin M auto-antibody directed against the I/i and H antigens, precursors of the ABH and Lewis blood group substances, on red blood cells. Autoimmune and lymphoproliferative disorders, Mycoplasma pneumoniae and other infections can be associated with the production of cold agglutinins. In its classic presentation with haemolytic anaemia and Raynaud's syndrome, cold agglutinin disease is usually idiopathic. Several factors play a role in determining the ability of a cold agglutinin to induce a haemolytic anaemia such as antibody concentration and temperature range, in particular the highest temperature at which antibodies interact with red blood cells. Case presentation A 48-year-old Caucasian man presented to our hospital with symptoms of extreme asthenia caused by severe anaemia. The transfusion of red blood cells (O Rh-positive, started as prescribed by the emergency guidelines in force without pre-transfusion tests, induced fatal haemolysis because of the presence of high levels of anti-H antibodies in his blood, that reacted with the large amount of H antigen in universal (0 red blood cells. Conclusion Emergency transfusion of universal red blood cells (0 Rh-positive or negative is usually accepted by the international guidelines in force in emergency departments. In this report we describe a rare complication caused by the very high concentration in the recipient of cold agglutinins and the activation of the complement system, responsible for red blood cell lysis and consequent fatal cardiovascular shock. We conclude that emergency transfusion of universal red blood cells (0 Rh-positive or negative may be dangerous and its risk should be assessed against the risk of delaying transfusion until the pre-transfusion tests are completed.

  9. Inflammatory and anti-inflammatory effects of soybean agglutinin

    OpenAIRE

    Benjamin C.F.; Figueiredo R.C.; Henriques M.G.M.O.; Barja-Fidalgo C.

    1997-01-01

    Soybean agglutinin (SBA) lectin, a protein present in raw soybean meals, can bind to and be extensively endocytosed by intestinal epithelial cells, being nutritionally toxic for most animals. In the present study we show that SBA (5-200 µg/cavity) injected into different cavities of rats induced a typical inflammatory response characterized by dose-dependent exudation and neutrophil migration 4 h after injection. This effect was blocked by pretreatment with glucocorticoid (0.5 mg/kg) or by co...

  10. Biogenesis and fate of the cell-cell adhesion molecule, agglutinin, during gametogenesis and fertilization of Chlamydomonas reinhardtii

    International Nuclear Information System (INIS)

    Fertilization in Chlamydomonas begins with the species-specific recognition and adhesion between gametes of opposite mating types via agglutinin molecules on the flagellar surface. This adhesion generates a cAMP-mediated sexual signal that initiates the subsequent events of call wall release, mating structure activation, and cell fusion. Although flagella of paired gametes remain attached to each other until the zygote forms, the process is dynamic. Engaged agglutinins rapidly become inactivated and turnover, requiring the constant supply of new agglutinins to replace the lost molecules. A population of cell body associated agglutinins has been postulated to the pool of agglutinins recruited during this turnover. Cell body agglutinins, therefore were identified, purified, localized within the cells and compared to flagellar agglutinins. The relationship between these two agglutinin populations was also examined. Cell body agglutinins were biochemically indistinguishable from the flagellar form with respect to their Mr, sedimentation coefficient, and hydrophobicity elution properties. Functionally, however, these molecules were inactive in situ. The calculated surface density of agglutinins in the cell body and flagellar domains was similar and thus could not explain their functional difference, but two domains contiguous and yet distinctive suggested they may be separated by a functional barrier. To test this, a method was developed, using a monoclonal antibody and cycloheximide, that removed the flagellar agglutinins so movement between the domains could be monitored. Mobilization of agglutinins onto the flagella did not occur unless sexual signaling was induced with cAMP and papaverine

  11. Human salivary agglutinin binds to lung surfactant protein-D and is identical with scavenger receptor protein gp-340

    DEFF Research Database (Denmark)

    Ligtenberg, T J; Bikker, F J; Groenink, J;

    2001-01-01

    agglutinin bound in a similar way to Streptococcus mutans and surfactant protein-D. Histochemically, the distribution of gp-340 in the submandibular salivary glands was identical with the agglutinin distribution, as shown in a previous paper [Takano, Bogert, Malamud, Lally and Hand (1991) Anat. Rec. 230, 307......-318]. We conclude that agglutinin is identical with gp-340, and that this molecule interacts with S. mutans and surfactant protein-D....

  12. Cold agglutinin disease associated with mycoplasma infection in an individual with type 2 diabetes: An atypical case

    OpenAIRE

    Chandrama Shrestha; Liu Min; Zhaohui Mo

    2012-01-01

    Cold Agglutinin Disease is a hemolytic anemia associated with cold reactive autoantibodies. Although the acute form of cold agglutinin disease can be attributed to autoimmune or infectious diseases and lymphoproliferative diseases, it has, to the best of our knowledge, so far,never been reported as secondary to mycoplasma pneumonia in a type 2 diabetic individual. In this paper, we report a case of cold agglutinin disease following mycoplasma pneumonia in a 47-year-old female patient with typ...

  13. Inflammatory and anti-inflammatory effects of soybean agglutinin

    Directory of Open Access Journals (Sweden)

    Benjamin C.F.

    1997-01-01

    Full Text Available Soybean agglutinin (SBA lectin, a protein present in raw soybean meals, can bind to and be extensively endocytosed by intestinal epithelial cells, being nutritionally toxic for most animals. In the present study we show that SBA (5-200 µg/cavity injected into different cavities of rats induced a typical inflammatory response characterized by dose-dependent exudation and neutrophil migration 4 h after injection. This effect was blocked by pretreatment with glucocorticoid (0.5 mg/kg or by co-injection of N-acetyl-galactosamine (100 x [M] lectin, but not of other sugars (100 x [M] lectin, suggesting an inflammatory response related to the lectin activity. Neutrophil accumulation was not dependent on a direct effect of SBA on the macrophage population since the effect was not altered when the number of peritoneal cells was increased or decreased in vivo. On the other hand, SBA showed chemotactic activity for human neutrophils in vitro. A slight increase in mononuclear cells was observed 48 h after ip injection of SBA. Phenotypic analysis of these cells showed an increase in the CD4+/CD8- lymphocyte population that returned to control levels after 15 days, suggesting the development of an immune response. SBA-stimulated macrophages presented an increase in the expression of CD11/CD18 surface molecules and showed some characteristics of activated cells. After intravenous administration, SBA increased the number of circulating neutrophils and inhibited in a dose-dependent manner the neutrophil migration induced by ip injection of carrageenan into peritoneal cavities. The co-injection of N-acetyl-galactosamine or mannose, but not glucose or fucose, inhibited these effects. The data indicate that soybean lectin is able to induce a local inflammatory reaction but has an anti-inflammatory effect when present in circulating blood

  14. Platelet cold agglutinins and thrombocytopenia: A diagnostic dilemma in the intensive care unit

    Directory of Open Access Journals (Sweden)

    TV Bharath Kumar

    2014-01-01

    Full Text Available We report a case of pseudo-thrombocytopenia due to cold agglutinins against platelets. These cold agglutinins were the cause for diagnostic confusion and resulted in extensive workup and unnecessary therapeutic precautions. A thirty two year old female with Guillain-Barre syndrome was admitted in the ICU and serial work-up showed markedly low levels of platelets. The patient had no symptoms of bleeding and patient was investigated extensively for deciphering the etiology of low platelet count. In-vitro clumping of platelets was suspected and in-vitro studies showed marked clumping of platelets with ethylene-diamine-tetra-acetic acid, citrate and heparinized samples. The manual platelet count was found to be within normal limits. Thrombocytopenia as a result of platelet cold agglutinins is a rare cause of in-vitro low platelet counts. No clinical problems have been reported due to the same.

  15. Reduction of Brucella species and Francisella tularensis cross-reacting agglutinins by dithiothreitol.

    OpenAIRE

    Behan, K A; Klein, G C

    1982-01-01

    Cross-reaction agglutinin titers to Brucella abortus antigen were found in 42 of 128 tularemia serum specimens, and cross-reaction titers to Francisella tularensis antigen were found in 8 of 34 brucellosis serum specimens. The cross-reaction titers were reduced to 10 or less by dithiothreitol, suggesting that the titers are due to immunoglobulin M antibody.

  16. Apoptosis induction by Maackia amurensis agglutinin in childhood acute lymphoblastic leukemic cells

    DEFF Research Database (Denmark)

    Kapoor, Sarika; Marwaha, Ram; Majumdar, Siddhartha;

    2007-01-01

    Malignant transformation is known to be associated with changes in cell surface carbohydrate-architecture, which can be detected by lectins. In the present study, Maackia amurensis agglutinin (MAA), specific for NeuNAcalpha(2-->3)Gal/GalNAc showed strong binding with lymphoblasts of children havi...

  17. Effect of in vivo γ-irradiation on the binding of wheat germ agglutinin on lymphocyte plasma membranes

    International Nuclear Information System (INIS)

    Using quantitative fluorimetry with fluoresceinated wheat germ agglutinin, we have been able to investigate in vivo gamma radiation-induced damage at the outer membrane level of rat splenic lymphocytes, namely damage to the glucosidic moieties of membrane glycoproteins and glycolipids. This paper demonstrates that below an irradiation level of 1 gray (Gy), removal of sialic acid is the major feature leading to new exposed specific binding sites for wheat germ agglutinin, since this lectin is specific for sialic acid and N-acetyl-D-glucosamine. Our studies also suggest that above 1 Gy of irradiation more internal damage occurs, since we observed a striking decrease in wheat germ agglutinin binding sites. (orig.)

  18. The frequency of ABO blood group maternal-fetal incompatibility, maternal iso-agglutinins, and immune agglutinins quantitation in Osogbo, Osun State, South-West of Nigeria

    Directory of Open Access Journals (Sweden)

    Oseni Bashiru

    2011-01-01

    Full Text Available Background : ABO incompatibility in maternal-fetal relationship has been shown to cause hemolytic disease of the newborn (HDNB; a survey which is not yet done in this locality. Aim: Frequency of ABO blood group maternal-fetal incompatibility, maternal iso-agglutinins, and immune agglutinins quantitation was carried out in Osogbo, Osun State, South-West of Nigeria. Settings and Designs : A total of 260 subjects comprising 130 postpartum mothers within the age range of 22-35 years having good obstetrics history and normal delivery, with their 130 neonate babies were used for the study. Materials and Methods : ABO cell and serum groupings were carried out on the subjects using standard antisera and cells with appropriate controls. Direct Coomb′s Test was carried out on neonate red cells. Antibody quantitation by double dilution on the maternal serum using red cells containing corresponding antigen to the antibody was determined. A titer, which is the reciprocal of the highest dilution showing agglutination by Indirect Coombs Test, was determined. Another batch of sera was pretreated with 2-mecarptoethanol before determining the titer. Statistical Analysis: The distribution study results obtained were compared in percentages, whereas the antibodies quantitation was expressed as titers using the mode of the titers for compariso-agglutininsn. Results and Conclusions : Thirty-eight percent (50 mothers were ABO incompatible with their babies, whereas 62% (80 mothers were compatible. The distribution of blood groups in the compatible population showed blood group O (45%; A (30%; B (20%; and AB (5%. Mothers O, A, and B carrying incompatible babies had a frequency of 24% each, whereas mothers AB had 28%. Serologist differences occur in maternal ABO antibodies of corresponding incompatible baby ABO antigens. A high incidence of ABO maternal-fetal incompatibility observed without detection of immune agglutinins is indicative of a rare incidence of HDNB due

  19. Adhesivity and rigidity of erythrocyte membrane in relation to wheat germ agglutinin binding

    OpenAIRE

    1984-01-01

    Binding of the plant lectin wheat germ agglutinin (WGA) to erythrocyte membranes causes membrane rigidification. One of our objectives has been to directly measure the effects of WGA binding on membrane rigidity and to relate rigidification to the kinetics and levels of WGA binding. Our other objective has been to measure the strength of adhesion and mechanics of cell separation for erythrocytes bound together by WGA. The erythrocyte membrane rigidity was measured on single cells by micropipe...

  20. Nematotoxicity of Marasmius oreades Agglutinin (MOA) Depends on Glycolipid Binding and Cysteine Protease Activity*

    OpenAIRE

    Wohlschlager, Therese; Butschi, Alex; Zurfluh, Katrin; Vonesch, Sibylle C.; auf dem Keller, Ulrich; Gehrig, Peter; Bleuler-Martinez, Silvia; Hengartner, Michael O.; Aebi, Markus; Künzler, Markus

    2011-01-01

    Fruiting body lectins have been proposed to act as effector proteins in the defense of fungi against parasites and predators. The Marasmius oreades agglutinin (MOA) is a Galα1,3Gal/GalNAc-specific lectin from the fairy ring mushroom that consists of an N-terminal ricin B-type lectin domain and a C-terminal dimerization domain. The latter domain shows structural similarity to catalytically active proteins, suggesting that, in addition to its carbohydrate-binding activity, MOA has an enzymatic ...

  1. O-linked oligosaccharides from salivary agglutinin: Helicobacter pylori binding sialyl-Lewis x and Lewis b are terminating moieties on hyperfucosylated oligo-N-acetyllactosamine

    NARCIS (Netherlands)

    S. Issa; A.P. Moran; S.N. Ustinov; J.H. Lin; A.J. Ligtenberg; N.G. Karlsson

    2010-01-01

    Salivary agglutinin plays a vital biological role modulating the protective effect in the oral cavity by interacting with a broad range of oral pathogens. Here, we describe the first characterization of the O-linked oligosaccharides of salivary agglutinin identified by negative ion liquid chromatogr

  2. Enrichment for CFU-C from murine and human bone marrow using soybean agglutinin

    International Nuclear Information System (INIS)

    Mouse bone marrow and spleen cells agglutinated by soybean agglutinin (SBA) or peanut agglutinin (PNA) were previously shown to be enriched for spleen colony-forming cells (CFU-S) and sufficiently depleted of graft-versus-host reaction producing cells to allow hematologic reconstitution of lethally irradiated allogeneic recipient mice. A similar enrichment for cells capable of forming colonies in soft agar culture (CFU-C) has now been found in the SBA-agglutinated fraction of mouse bone marrow cells, in contrast to the finding that in human bone marrow the majority of the CFU-C are in the fraction not agglutinated by SBA. Cytofluorometric studies with fluorescein-labeled SBA (FITC-SBA) revealed that the majority of both mouse and human bone marrow cells bind the lectin. Experiments mixing the human marrow fractions separated by SBA reveal that true enrichment for CFU-C is achieved in the unagglutinated fraction, as opposed to a possible depletion of a suppressor cell population. Granulocytic, monocytic, and mixed cell colonies were all enriched in the SBA-unagglutinated cell fraction from human bone marrow

  3. Problems of Cold Agglutinins in Cardiac Surgery: How to Manage Cardiopulmonary Bypass and Myocardial Protection

    Directory of Open Access Journals (Sweden)

    Kambiz Alizadeh

    2014-02-01

    Full Text Available Cold agglutinins are of unique relevance in cardiac surgerybecause of the use of hypothermic cardiopulmonary bypass (CPB. Cold autoimmune diseases are defined by the presence of abnormal circulating proteins (usually IgM or IgA antibodies that agglutinate in response to a decrease in body temperature. These disorders include cryoglobulinemia and cold hemagglutinin disease.Immunoglobulin M autoantibodies to red blood cells, which activateat varying levels of hypothermia, can cause catastrophic hemagglutination,microvascular thrombosis, or hemolysis. Management of anesthesia in these patients includes strict maintenance of normothermia. Patients scheduled for the surgery requiring cardiopulmonary bypass present significant challenges. Use of systemic hypothermia may be contraindicated, and cold cardioplegia solutions may precipitate intracoronary hemagglutination with consequent thrombosis, ischemia, or infarction. Management of CPB andmyocardial protection requires individualized planning. We describea case of MV repair and CABG in a patient with high titercold agglutinins and high thermal amplitude for antibody activation.Normothermic CPB and continuous warm blood cardioplegia weresuccessfully used.

  4. QUANTIFICATION AND LOCALIZATION OF WHEAT GERM AGGLUTININ RECEPTOR ON HUMAN SPERMME MBRANE IN FERTILE AND INFERTILE MALES

    Institute of Scientific and Technical Information of China (English)

    PANZhi-Xing; WANGYi-Fei

    1989-01-01

    It has been proved/n our prcvious study that wheat germ agglutinin (WGA) receptor on human sperm membrane is closely related to male fertility and there exists a significant difference in WGA receptors betweea fertile and infertile mcn. In this report, enzyme linked

  5. Salivary agglutinin/DMBT1SAG expression is up-regulated in the presence of salivary gland tumors

    DEFF Research Database (Denmark)

    Bikker, F J; van der Wal, J E; Ligtenberg, A J M;

    2004-01-01

    Salivary agglutinin (SAG) is encoded by the gene Deleted in Malignant Brain Tumors 1 (DMBT1) and represents the salivary variant of DMBT1 (DMBT1(SAG)). While SAG is a bona fide anti-caries factor, DMBT1 was proposed as a candidate tumor-suppressor for brain, digestive tract, and lung cancer. Though...

  6. The Salivary Scavenger and Agglutinin in Early Life: Diverse Roles in Amniotic Fluid and in the Infant Intestine

    NARCIS (Netherlands)

    Reichhardt, M.P.; Jarva, H.; Been, de M.; Rodriguez, J.M.; Quintana, E.J.; Loimaranta, V.; Vos, de W.M.; Meri, S.

    2014-01-01

    The salivary scavenger and agglutinin (SALSA), also known as gp340 and dmbt1, is an antimicrobial and inflammation-regulating molecule located at the mucosal surfaces. The present study revealed that SALSA was present in the amniotic fluid (AF) and exceptionally enriched in both meconium and feces o

  7. Expression of Monstera deliciosa agglutinin gene (mda) in tobacco confers resistance to peach-potato aphids.

    Science.gov (United States)

    Kai, Guoyin; Ji, Qian; Lu, Yang; Qian, Zhongying; Cui, Lijie

    2012-08-01

    The aphid is one of the most serious pests that causes damage to crops worldwide. Lectins from Araceae plant had been proved useful to control the aphid. Herein, the full-length cDNA of Monstera deliciosa agglutinin (mda) gene was cloned and then introduced into tobacco and the influence of the expression of mda in transgenic tobacco against peach-potato aphids (Myzus persicae) was investigated. Among 92 regenerated plants, 59 positive tobacco lines were obtained. Real-time PCR assays and aphid bioassay test revealed that there is a positive correlation between the expression level of mda and the inhibitory effect on peach-potato aphids. The average anti-pests ability of mda transgenic tobacco was 74%, which was higher than that of other reported lectins from Araceae plant. These results indicated that MDA is one of promising insect resistance proteins selected for the control of peach-potato aphids. PMID:22660606

  8. Application of an Optical Biosensor to Study the Interaction between Porphyrins and Wheat Germ Agglutinin

    Institute of Scientific and Technical Information of China (English)

    2006-01-01

    An optical biosensor with a stirred cuvette has been used to monitor the interaction of immobilized wheat germ agglutinin (WGA) with two water-soluble cationic porphyrins. The association constants (Ka) of the free base porphyrin and its Zn(Ⅱ) complex form were 2.66 and 27.31×105 L/mol at 20 ℃ respectively. The interactions of the free base porphyrin were further investigated at temperatures between 15 ℃ and 37 ℃. The thermodynamics parameters, changes in free energy, enthalpy and entropy, were -31.23, 22.92, 54.15 kJ/mol respectively. The heat capacity change was -355.53 J·mol-1·K-1. The binding was driven by entropic contribution, and showed strong enthalpy-entropy compensation. It was governed primarily by hydrophobic forces.

  9. Dynamic light scattering study of peanut agglutinin: Size, shape and urea denaturation

    Indian Academy of Sciences (India)

    Sagarika Dev; Avadhesha Surolia

    2006-12-01

    Peanut agglutinin (PNA) is a homotetrameric protein with a unique open quaternary structure. PNA shows non-two state profile in chaotrope induced denaturation. It passes through a monomeric molten globule like state before complete denaturation (Reddy et al 1999). This denaturation profile is associated with the change in hydrodynamic radius of the native protein. Though the molten globule-like state is monomeric in nature it expands in size due to partial denaturation. The size and shape of the native PNA as well as the change in hydrodynamic radius of the protein during denaturation has been studied by dynamic light scattering (DLS). The generation of two species is evident from the profile of hydrodynamic radii. This study also reveals the extent of compactness of the intermediate state.

  10. Cloning and Functional Analysis of the Bifunctional Agglutinin/Trypsin Inhibitor from Helianthus tuberosus L.

    Institute of Scientific and Technical Information of China (English)

    Tuanjie Chang; Hongli Zhai; Songbiao Chen; Guisheng Song; Honglin Xu; Xiaoli Wei; Zhen Zhu

    2006-01-01

    In order to find new insect resistance genes, four homologous cDNAs, hta-a, hta-b, hta-c and hta-d with lengths of 775, 718, 784 and 752 bp, respectively (GenBank accession numbers AF477031-AF477034), were isolated from a tuber cDNA expression library of Helianthus tuberosus L. Sequence analysis revealed that all four cDNAs contain an open reading frame of 444 bp, coding a polypeptide of 147 amino acid residues, and that the sequences of the cDNAs are very similar to those of the mannose-binding agglutinin genes of the jacalin-related family. In hemagglutination reactions and hapten inhibition assays, affinity-purified HTA (Helianthus tuberosus agglutinin) from induced Escherichia coli BL21(DE3) expressing GST-HTA shows hemagglutination ability and a higher carbohydrate-binding ability for mannose than other tested sugars.Trypsin inhibitory activity was detected in the crude extracts of induced E. coli BL21(DE3)expressing HTA,and was further verified by trypsin inhibitory activity staining on native polyacrylamide gel. The mechanism of interaction between HTA and trypsin was studied by molecular modeling. We found that plenty of hydrogen bonds and electrostatic interactions can be formed between the supposed binding sites of HTA-b and the active site of trypsin, and that a stable HTA/trypsin complex can be formed. The results above imply that HTA might be a bifunctional protein with carbohydrate-binding activity and trypsin inhibitory activity. Moreover,Northern blotting analysis demonstrated that hta is predominantly expressed in tubers of H. tuberosus, very weakly expressed in stems, but not expressed at all in other tissues. Southern blotting analysis indicated that hta is encoded by a multi-gene family. The insect resistance traits have been described in another paper.

  11. THE PERSISTENCE OF LEPTOSPIRAL AGGLUTININS TITERS IN HUMAN SERA DIAGNOSED BY THE MICROSCOPIC AGGLUTINATION TEST

    Directory of Open Access Journals (Sweden)

    Eliete C. ROMERO

    1998-05-01

    Full Text Available The persistence of agglutinins detected by MAT has created some problems to the interpretation of the results. The aim of this study was to examine the data of serology from 70 patients with serologically confirmed diagnosis of leptospirosis by during 3-13 months after being affected with leptospires in order to elucidate the interpretation of the persistence of agglutinins detected by MAT. Sixty-one patients sera (87.14% had titers equal or greater than 800. Of these, two individuals maintained titers of 800 thirteen months after the onset. This study showed that only one sample of sera with high titers is not reliable to determine the time at which infection occurred.Persistência de títulos de aglutininas anti-leptospiras em soros humanos diagnosticados pelo teste de aglutinação microscópica A persistência de aglutininas detectadas por MAT tem criado problemas na interpretação dos resultados. O objetivo deste trabalho foi examinar os resultados da sorologia de 70 pacientes com confirmação sorológica de leptospirose durante 3-13 meses após terem sido infectados para se poder elucidar a interpretação da persistência de aglutininas detectadas por MAT. Sessenta e um soros de pacientes (87,14% apresentaram títulos iguais, ou maiores, que 800. Destes, 2 indivíduos mantiveram títulos de 800 treze meses após terem sido infectados. Este estudo mostra que apenas uma amostra de soro, mesmo com alto título de aglutininas, não pode ser considerada para determinar a fase da doença.

  12. Low Prevalence of Brucella Agglutinins in Blood Donors in Central Province of Iran

    Directory of Open Access Journals (Sweden)

    Masoomeh Sofian

    2013-03-01

    Full Text Available Background: Brucellosis is a zoonotic disease of worldwide distribution and has great economic importance. Despite its control in many countries, it remains endemic in Iran. Brucellosis was investigated in many high risk occupational groups; however, few studies on the prevalence of brucellosis among blood donors are available. To determine the seroprevalence of brucellosis antibodies in blood donors, a serological study was carried out in central province of Iran.Materials and Methods: A total of 897 healthy blood donors with mean age 37.23 ± 10.9 years were enrolled in the study. Laboratory tests including Standard Tube Agglutination Test (STA and 2-mercaptoethanol (2ME agglutination were checked in all samples. STA dilution ≥ 1:80, and in the presence of 2-mercaptoethanol (2ME agglutination ≥ 20 was considered positive.Results: Out of 897 cases, 11.9% were inhabitants of rural areas. 41.5% had history of consumption of unpasteurized dairy products and 9.3% had history of contact with domestic animals. A very low level of Brucella agglutinins was present in 3(0.33% of the samples and only one sample (0.11% was found to be truly positive for Brucella agglutinins. 2ME was negative in all samples. None of these 4 subjects showed signs and symptoms of brucellosis in 6 months follow-up.Conclusion: On the basis of our data, brucellosis has no epidemiological and clinical importance in our blood donors; therefore, it is not recommended to perform screening tests such as, STA and 2ME to identify brucellosis antibodies in the sera of blood donors.

  13. Agglutinins and cardiac surgery: a web based survey of cardiac anaesthetic practice; questions raised and possible solutions

    OpenAIRE

    Shah, S.; Gilliland, H; Benson, G

    2014-01-01

    Introduction Cardiac surgery involves cardiopulmonary bypass during which the core temperature is generally lowered to hypothermic levels. Patients presenting for cardiac surgery are sometimes reported to have cold or warm autoantibodies at the time of blood screening. It is known that cold agglutinins may cause potentially life-threatening haemolysis, intracoronary haemagglutination leading to inadequate cardioplegia distribution, thrombosis, embolism, ischaemia or infarction. The risk (if a...

  14. Discovering the Secrets of the Candida albicans Agglutinin-Like Sequence (ALS) Gene Family—a Sticky Pursuit

    OpenAIRE

    HOYER, LOIS L.; GREEN, CLAYTON B.; Oh, Soon-Hwan; Zhao, Xiaomin

    2008-01-01

    The Agglutinin-Like Sequence (ALS) family of Candida albicans includes eight genes that encode large cell-surface glycoproteins. The high degree of sequence relatedness between the ALS genes and the tremendous allelic variability often present in the same C. albicans strain complicated definition and characterization of the gene family. The main hypothesis driving ALS family research is that the genes encode adhesins, primarily involved in host-pathogen interactions. Although adhesive functio...

  15. Purification and partial characterization of an agglutinin from Octopus maya serum.

    Science.gov (United States)

    Alpuche, Juan; Pereyra, Ali; Mendoza-Hernández, Guillermo; Agundis, Concepción; Rosas, Carlos; Zenteno, Edgar

    2010-05-01

    A 66-kDa lectin (OmA) was purified from the serum of the Yucatan peninsula endemic octopus (Octopus maya) by a single step affinity chromatography on glutaraldehyde-fixed stroma from rat erythrocytes. OmA corresponds to 0.8% of the total circulating protein in the hemolymph; it is composed of three equal subunits of 22kDa each, and 7.4% of linked carbohydrates. The amino acids' composition indicated that agglutinin contained mainly aspartic and glutamic acids, and cysteine and methionine were identified in minor proportion. OmA agglutinates mainly rat, guinea pig, and rabbit erythrocytes, and this activity is partially inhibited by galactosamine, melobiose, galacturonic acid, mannose, and methyl alpha and beta galactosides. Hemagglutinating activity is not dependent on divalent cations, such as Ca(2+), Mg(2+), or Mn(2+). The OmA subunits showed no identity for any lectin in databases but partial identity with the type A hemocyanin from Octopus dolfleini hemolymph; the main similarities are related to tyrosinase domains and copper A and B sites that conform to the oxygen-binding site of hemocyanin. PMID:20105460

  16. Transneuronally transported wheat germ agglutinin labels glia as well as neurons in the rat visual system

    International Nuclear Information System (INIS)

    Following intraocular injection in adult rats of 125I-labeled wheat germ agglutinin (I-WGA), the ultrastructural distribution of label in the superior colliculus and lateral geniculate was examined by electron microscope autoradiography. Three days after injection, 5.4% of the label in the lateral geniculate was associated with neuronal perikarya, and 3.6% was associated with glial perikarya. The corresponding figures for the superior colliculus were 5.1% and 0.8%. When the data were expressed as number of grains per micron 2 cytoplasm, there was no statistically significant difference between the grain density over neuronal or glial cytoplasm in either the lateral geniculate or the superior colliculus. A statistical analysis of the distance between the silver grains and the cell membranes showed that in both neurons and glia, the observed labeling was the product of internalized I-WGA and not the result of scatter from the neuropil or from label bound to the surface of the cells. These results indicate that much of the WGA released from axons and axon terminals is not confined to a specific ''transsynaptic'' pathway, but produces a generalized labeling of nearby cells, much like a microinjection of WGA into the region

  17. The prevalence of anti-leptospiral agglutinins in sera of wildlife in southeastern Australia.

    Science.gov (United States)

    Milner, A R; Wilks, C R; Spratt, D M; Presidente, P J

    1981-04-01

    Anti-leptospiral agglutinins were found in the serum from 18 (7 species) of 419 (25 species) animals sampled from various areas of southeastern Australia. Positive serologic reactions were observed in 5 of 25 (20%) brush-tailed possum (Trichosurus vulpecula), 1 of 26 (3.8%) tammar wallaby (Macropus eugenii), 2 of 12 (16.7%) swamp wallaby (Wallabia bicolor), 1 of 3 (33.3%) koala (Phascolarctos cinereus), 3 of 41 (7.3%) common wombat (Vombatus ursinus), 2 of 100 (2%) bush rat (Rattus fuscipes) and 4 of 12 (25%) rusa deer (Cervus timorensis). The majority (55.5%) of serologic reactions were to serovar hardjo. No serologic reactions were observed in samples from echidna (Tachyglossus aculeatus), brown antechinus (Antechinus stuartii), swainson's antechinus (Antechinus swaisonsii), long-nosed bandicoot (Perameles nasuta), brown bandicoot(Isoodon obesulus), common ringtail (Pseudocheirus peregrinus), greater glider (Schoinobates volans), eastern grey kangaroo (Macropus giganteus), red-necked wallaby (Macropus rufogriseus), rabbit (Oryctolagus cuniculus), water rat (Hydromys chrysogaster), black rat (Rattus rattus), eastern swamp rat (Rattus lutreolus), broad-toothed rat (Mastacomys fuscus), fox (Vulpes vulpes), sambar deer (Cervus unicolor), hog deer (Axis porcinus) and fallow deer (Dama dama). PMID:7241704

  18. Evolution of the rapidly mutating human salivary agglutinin gene (DMBT1) and population subsistence strategy.

    Science.gov (United States)

    Polley, Shamik; Louzada, Sandra; Forni, Diego; Sironi, Manuela; Balaskas, Theodosius; Hains, David S; Yang, Fengtang; Hollox, Edward J

    2015-04-21

    The dietary change resulting from the domestication of plant and animal species and development of agriculture at different locations across the world was one of the most significant changes in human evolution. An increase in dietary carbohydrates caused an increase in dental caries following the development of agriculture, mediated by the cariogenic oral bacterium Streptococcus mutans. Salivary agglutinin [SAG, encoded by the deleted in malignant brain tumors 1 (DMBT1) gene] is an innate immune receptor glycoprotein that binds a variety of bacteria and viruses, and mediates attachment of S. mutans to hydroxyapatite on the surface of the tooth. In this study we show that multiallelic copy number variation (CNV) within DMBT1 is extensive across all populations and is predicted to result in between 7-20 scavenger-receptor cysteine-rich (SRCR) domains within each SAG molecule. Direct observation of de novo mutation in multigeneration families suggests these CNVs have a very high mutation rate for a protein-coding locus, with a mutation rate of up to 5% per gamete. Given that the SRCR domains bind S. mutans and hydroxyapatite in the tooth, we investigated the association of sequence diversity at the SAG-binding gene of S. mutans, and DMBT1 CNV. Furthermore, we show that DMBT1 CNV is also associated with a history of agriculture across global populations, suggesting that dietary change as a result of agriculture has shaped the pattern of CNV at DMBT1, and that the DMBT1-S. mutans interaction is a promising model of host-pathogen-culture coevolution in humans. PMID:25848046

  19. A novel autotransporter of uropathogenic Proteus mirabilis is both a cytotoxin and an agglutinin.

    Science.gov (United States)

    Alamuri, Praveen; Mobley, Harry L T

    2008-05-01

    One of the six predicted Proteus mirabilis autotransporters (ATs), ORF c2341, is predicted to contain a serine protease motif and was earlier identified as an immunogenic outer membrane protein in P. mirabilis. The 3.2 kb gene encodes a 117 kDa protein with a 58-amino-acid-long signal peptide, a 75-kDa-long N-terminal passenger domain and a 30-kDa-long C-terminal translocator. Affinity-purified 110 kDa AT exhibited chymotrypsin-like activity and hydrolysed N-Suc-Ala-Ala-Pro-Phe-pNa and N-Suc-Ala-Ala-Pro-Leu-pNa with a K(M) of 22 muM and 31 muM, respectively, under optimal pH of 8.5-9.0 in a Ca(2+)-dependent manner. Activity was inhibited by subtilase-specific inhibitors leupeptin and chymostatin. Both the cell-associated and purified form elicited cytopathic effects on cultured kidney and bladder epithelial cells. Substrate hydrolysis as well as cytotoxicity was associated with the passenger domain and was compromised upon mutation of any of the catalytic residues (Ser366, His147 and Asp533). At alkaline pH and optimal cell density, the AT also promoted autoaggregation of P. mirabilis and this function was independent of its protease activity. Cytotoxicity, autoaggregation and virulence were significantly reduced in an isogenic pta mutant of P. mirabilis. Proteus toxic agglutinin (Pta) represents a novel autotransported cytotoxin with no bacterial homologues that works optimally in the alkalinized urinary tract, a characteristic of urease-mediated urea hydrolysis during P. mirabilis infection. PMID:18430084

  20. Effect of wheat germ agglutinin on the viscoelastic properties of erythrocyte membrane.

    Science.gov (United States)

    Smith, L; Hochmuth, R M

    1982-07-01

    The elasticity and viscosity of the human erythrocyte membrane were measured as a function of the concentration of wheat germ agglutinin (WGA) in a suspending solution containing 1 mg/ml albumin, approximately 5 X 10(5) cells/ml and between 0.0 and 0.2 microgram/ml WGA. Membrane elasticity was characterized by the elastic shear modulus, which provided a measure of the resistance of the membrane to constant-area elastic deformations that occurred in the membrane plane. The elastic shear modulus was determined by aspirating a portion of the membrane into a micropipette and measuring the extension of the membrane into the pipette as a function of the suction pressure. The results indicated no significant change in shear modulus for concentrations of WGA between 0.0 and 0.2 microgram/ml. Membrane viscosity was characterized by the coefficient of surface viscosity, which, in effect, was a measure of the membrane's resistance to rates of deformation. This coefficient was determined from the time required for an erythrocyte to recover its undeformed shape after it had been elongated by the application of an equal and opposite force applied at diametrically opposite points on the erythrocyte rim. The value for the coefficient of surface viscosity was found to increase by a factor of almost three when the WGA concentration was increased from 0.0 to 0.2 microgram/ml. These results indicated that, in the presence of albumin, WGA can increase membrane dissipation (viscosity) without altering the structural rigidity (elasticity) of the membrane. PMID:6896878

  1. Vicia villosa agglutinin separates freshly isolated Peyer's Patch T cells into interleukin 5- or interleukin 2-producing subsets

    OpenAIRE

    1989-01-01

    Murine L3T4 T cells freshly isolated from Peyer's Patch were fractionated based on differential adherence to Vicia villosa agglutinin (VVA). VVA adherent cells secreted IL-5, but not IL-2, after stimulation with Con A and IL-1. In striking contrast, VVA nonadherent PP L3T4 T cells secreted IL-2, but not IL-5, under the same conditions. In addition, supernatants from VVA adherent, but not from VVA nonadherent cells cultures, enhanced IgA secretion by LPS-stimulated splenic B cells to the same ...

  2. Evaluation of the Role of Candida albicans Agglutinin-Like Sequence (Als) Proteins in Human Oral Epithelial Cell Interactions

    OpenAIRE

    Murciano, Celia; Moyes, David L.; Runglall, Manohursingh; Tobouti, Priscila; Islam, Ayesha; HOYER, LOIS L.; Naglik, Julian R.

    2012-01-01

    The fungus C. albicans uses adhesins to interact with human epithelial surfaces in the processes of colonization and pathogenesis. The C. albicans ALS (agglutinin-like sequence) gene family encodes eight large cell-surface glycoproteins (Als1-Als7 and Als9) that have adhesive function. This study utilized C. albicans Δals mutant strains to investigate the role of the Als family in oral epithelial cell adhesion and damage, cytokine induction and activation of a MAPK-based (MKP1/c-Fos) signalin...

  3. Identification of the bacteria-binding peptide domain on salivary agglutinin (gp-340/DMBT1), a member of the scavenger receptor cysteine-rich superfamily

    DEFF Research Database (Denmark)

    Bikker, Floris J; Ligtenberg, Antoon J M; Nazmi, Kamran;

    2002-01-01

    Salivary agglutinin is encoded by DMBT1 and identical to gp-340, a member of the scavenger receptor cysteine-rich (SRCR) superfamily. Salivary agglutinin/DMBT1 is known for its Streptococcus mutans agglutinating properties. This 300-400 kDa glycoprotein is composed of conserved peptide motifs: 14...... containing exclusively SRCR and SID domains that binds to S. mutans. To define more closely the S. mutans-binding domain, consensus-based peptides of the SRCR domains and SIDs were designed and synthesized. Only one of the SRCR peptides, designated SRCRP2, and none of the SID peptides bound to S. mutans....... Strikingly, this peptide was also able to induce agglutination of S. mutans and a number of other bacteria. The repeated presence of this peptide in the native molecule endows agglutinin/DMBT1 with a general bacterial binding feature with a multivalent character. Moreover, our studies demonstrate for the...

  4. Studies on glycoproteins produced by wild type and wheat germ agglutinin-resistant B16 mouse melanoma cells

    International Nuclear Information System (INIS)

    Two variants of B16 mouse melanoma cells have been selected in serum-free medium for their resistance to toxic levels of wheat germ agglutinin isolation 1 (WGA). Chromosome analysis and characteristic melanin production showed that the variants are derived from the parent mouse melanoma cell lines. However, the two variants were less tumorigenic in mice compared to the parent B16 mouse melanoma cells. The variants showed a marked decrease in cell agglutination with WGA. Cell agglutination with recin and peanut lectin was not different between the three cell lines, but the two variants showed a slight increase in agglutination with concanavalin A. The binding of 125I-labeled wheat germ agglutinin to the two variant cells was reduced compared to that of the parent cell. Glycoproteins secreted or shed by the three lines were isolated after growth in serum-free medium in the presence of [3He]glucosamine and bovine serum albumin (1%). These metabolically labeled products were fractionated on the basis of their interaction with WGA-Sepharose (2 mg/ml). The WGA-Sepharose affinity chromatographic data suggested a decrease in WGA-binding glycoprotein(s) secreted to the medium by the two variants. The WGA-bound glycoproteins from the two variants upon SDS-PAGE revealed three bands of approximate molecular weights, 92,000, 56,000, and 42,000, none of which were present in the parent cell line (50,000 molecular weight)

  5. Prevalence and spatial analysis of antileptospiral agglutinins in dairy cattle - Microregion of Sete Lagoas, Minas Gerais, 2009/2010

    Directory of Open Access Journals (Sweden)

    R.R. Nicolino

    2014-06-01

    Full Text Available The aim of this study was to evaluate the seroprevalence of leptospirosis in the dairy herds from Minas Gerais, Brazil, during the years 2009 and 2010. A total of 2,915 serum samples were collected from the lactating cows of 151 properties in eleven municipalities located in the Sete Lagoas region. The Microscopic Agglutination Test was used to detect antileptospiral agglutinins. An individual animal prevalence of 20.7% (95% CI = 17.1% - 24.3% and a herd prevalence of 80.8% (95% CI = 73.8% = 87.7% were determined. The most prevalent serovars were hardjoprajitno at 19.4%; hardjoprajitno strain Norma at 17.4%; and hardjo-bovis at 17.4%. These results show the significance of the hardjo serovar in bovine leptospirosis cases in Minas Gerais.

  6. Transgenic tetraploid Isatis indigotica expressing Bt Cry1Ac and Pinellia ternata agglutinin showed enhanced resistance to moths and aphids.

    Science.gov (United States)

    Xiao, Ying; Wang, Kai; Ding, Ruxian; Zhang, Hanming; Di, Peng; Chen, Junfeng; Zhang, Lei; Chen, Wansheng

    2012-01-01

    Co-expression of multiple genes encoding different kinds of insect resistant proteins has been developed to confer a broader spectrum of pest control. Tetraploid Isatis indigotica Fort was transformed with a plasmid, p3300BP, containing Bacillus thuringiensis Cry1Ac gene (Bt) and Pinellia ternata agglutinin gene (Pta) and the selectable marker herbicide resistance gene (Bar) driven by the CaMV35S promoter via Agrobacterium tumefaciens-mediated transformation. The integration and expression of introduced genes in regenerated transgenic plants were confirmed by PCR and Western blot assays. Insect bioassay test demonstrated transgenic lines had significant inhibition to diamondback moths (Plutella xylostella L.) and peach potato aphids (Myzus persicae Sulzer) simultaneously. Our study reported here would be a great motivation for field culture of tetraploid I. indigotica, also providing an efficient molecular breeding strategy to provide insect tolerant plants. PMID:21559837

  7. An Unusual Member of the Papain Superfamily: Mapping the Catalytic Cleft of the Marasmius oreades agglutinin (MOA) with a Caspase Inhibitor

    OpenAIRE

    Gabriele Cordara; André van Eerde; Grahn, Elin M.; Harry C Winter; Goldstein, Irwin J.; Ute Krengel

    2016-01-01

    Papain-like cysteine proteases (PLCPs) constitute the largest group of thiol-based protein degrading enzymes and are characterized by a highly conserved fold. They are found in bacteria, viruses, plants and animals and involved in a number of physiological and pathological processes, parasitic infections and host defense, making them interesting targets for drug design. The Marasmius oreades agglutinin (MOA) is a blood group B-specific fungal chimerolectin with calcium-dependent proteolytic a...

  8. Measurement of the Fucosylated Sugar Chains of Serum α_1-Antitrypsin : The Relationship between Reactivity with Lens Culinaris Agglutinin and HPLC Analysis of the Pyridylaminated Sugar Chains

    OpenAIRE

    Saitoh, Atsushi; Aoyagi, Yutaka; Mori, Shigeki; Suda, Takeshi; Suzuki, Yasufumi; Sekine, Chuichi; Asakura, Hitoshi

    1997-01-01

    Twelve samples of α_1-anlitrypsin (AAT) were purified from six patients with hepatocellular carcinoma, one with pancreatic cancer and five healthy individuals, and subjected to the measurement of the fucosylated sugar chain. We compared different results from crossed immunoaffinoelectrophoresis (CIAE) with Lens culinaris agglutinin (LCA) and by the carbohydrate analysis of high-performance liquid chromatography (HPLC) with the pylidylaminated (PA・) oligosaccharides obtained from each AAT. A s...

  9. Allergenicity assessment of Allium sativum leaf agglutinin, a potential candidate protein for developing sap sucking insect resistant food crops.

    Directory of Open Access Journals (Sweden)

    Hossain Ali Mondal

    Full Text Available BACKGROUND: Mannose-binding Allium sativum leaf agglutinin (ASAL is highly antinutritional and toxic to various phloem-feeding hemipteran insects. ASAL has been expressed in a number of agriculturally important crops to develop resistance against those insects. Awareness of the safety aspect of ASAL is absolutely essential for developing ASAL transgenic plants. METHODOLOGY/PRINCIPAL FINDINGS: Following the guidelines framed by the Food and Agriculture Organization/World Health Organization, the source of the gene, its sequence homology with potent allergens, clinical tests on mammalian systems, and the pepsin resistance and thermostability of the protein were considered to address the issue. No significant homology to the ASAL sequence was detected when compared to known allergenic proteins. The ELISA of blood sera collected from known allergy patients also failed to show significant evidence of cross-reactivity. In vitro and in vivo assays both indicated the digestibility of ASAL in the presence of pepsin in a minimum time period. CONCLUSIONS/SIGNIFICANCE: With these experiments, we concluded that ASAL does not possess any apparent features of an allergen. This is the first report regarding the monitoring of the allergenicity of any mannose-binding monocot lectin having insecticidal efficacy against hemipteran insects.

  10. Abrus agglutinin is a potent anti-proliferative and anti-angiogenic agent in human breast cancer.

    Science.gov (United States)

    Bhutia, Sujit K; Behera, Birendra; Nandini Das, Durgesh; Mukhopadhyay, Subhadip; Sinha, Niharika; Panda, Prashanta Kumar; Naik, Prajna Paramita; Patra, Samir K; Mandal, Mahitosh; Sarkar, Siddik; Menezes, Mitchell E; Talukdar, Sarmistha; Maiti, Tapas K; Das, Swadesh K; Sarkar, Devanand; Fisher, Paul B

    2016-07-15

    Abrus agglutinin (AGG), a plant lectin isolated from the seeds of Abrus precatorius, has documented antitumor and immunostimulatory effects in murine models. To examine possible antitumor activity against breast cancer, we established human breast tumor xenografts in athymic nude mice and intraperitoneally administered AGG. AGG inhibited tumor growth and angiogenesis as confirmed by monitoring the expression of Ki-67 and CD-31, respectively. In addition, TUNEL positive cells increased in breast tumors treated with AGG suggesting that AGG mediates anti-tumorigenic activity through induction of apoptosis and inhibition of angiogenesis. On a molecular level, AGG caused extrinsic apoptosis through ROS generation that was AKT-dependent in breast cancer cells, without affecting primary mammary epithelial cells, suggesting potential cancer specificity of this natural compound. In addition, using HUVECs, AGG inhibited expression of the pro-angiogenic factor IGFBP-2 in an AKT-dependent manner, reducing angiogenic phenotypes both in vitro and in vivo. Overall, the present results establish that AGG promotes both apoptosis and anti-angiogenic activities in human breast tumor cells, which might be exploited for treatment of breast and other cancers. PMID:26914517

  11. Synthesis of tetravalent LacNAc-glycoclusters as high-affinity cross-linker against Erythrina cristagalli agglutinin.

    Science.gov (United States)

    Ogata, Makoto; Chuma, Yasushi; Yasumoto, Yoshinori; Onoda, Takashi; Umemura, Myco; Usui, Taichi; Park, Enoch Y

    2016-01-01

    Four kinds of tetravalent double-headed glycoclusters [(LacNAc)4-DHGs] were designed with linkers of varying lengths consisting of alkanedioic carboxyamido groups (C6, C12, C18 and C24) between two bi-antennary LacNAc-glycosides. These glycoclusters served as high-affinity cross-linking ligands for the LacNAc-binding lectin Erythrina cristagalli agglutinin (ECA). The binding activity and cross-linking between each ligand and ECA were characterized by a hemagglutination inhibition (HI) assay, isothermal titration calorimetry (ITC), a quantitative precipitation assay and dynamic light scattering (DLS). For the precipitation assay and DLS measurement, the synthesized (LacNAc)4-DHGs were found to be capable of binding and precipitating the ECA as multivalent ligands. ITC analysis indicated the binding of (LacNAc)4-DHGs was driven by a favorable enthalpy change. Furthermore, the entropy penalty from binding (LacNAc)4-DHGs clearly decreased in a spacer length-dependent manner. The binding affinities of flexible (LacNAc)4-DHGs (C18 and C24) with long spacers were found to be more favorable than those of the clusters having short spacers (C6 and C12). These results were supported by molecular dynamics simulations with explicit water molecules for the tetravalent glycoclusters with ECA. We concluded that the subtle modification in the epitope-presenting scaffolds exerts the significant effect in the recognition efficiency involved in the LacNAc moieties by ECA. PMID:26672510

  12. Effects of wheat germ agglutinin on human gastrointestinal epithelium: Insights from an experimental model of immune/epithelial cell interaction

    International Nuclear Information System (INIS)

    Wheat germ agglutinin (WGA) is a plant protein that binds specifically to sugars expressed, among many others, by human gastrointestinal epithelial and immune cells. WGA is a toxic compound and an anti-nutritional factor, but recent works have shown that it may have potential as an anti-tumor drug and as a carrier for oral drugs. To quantitate the toxicity threshold for WGA on normal epithelial cells we previously investigated the effects of the lectin on differentiated Caco2 cells, and showed that in the micromolar range of concentrations WGA could alter the integrity of the epithelium layer and increase its permeability to both mannitol and dextran. WGA was shown to be uptaken by Caco2 cells and only ∼ 0.1% molecules were observed to cross the epithelium layer by transcytosis. Here we show that at nanomolar concentrations WGA is unexpectedly bioactive on immune cells. The supernatants of WGA-stimulated peripheral blood mononuclear cells (PBMC) can alter the integrity of the epithelium layer when administered to the basolateral side of differentiated Caco2 cells and the effects can be partially inhibited by monoclonal antibodies against IL1, IL6 and IL8. At nanomolar concentrations WGA stimulates the synthesis of pro-inflammatory cytokines and thus the biological activity of WGA should be reconsidered by taking into account the effects of WGA on the immune system at the gastrointestinal interface. These results shed new light onto the molecular mechanisms underlying the onset of gastrointestinal disorders observed in vivo upon dietary intake of wheat-based foods.

  13. Development of transgenic cotton lines expressing Allium sativum agglutinin (ASAL for enhanced resistance against major sap-sucking pests.

    Directory of Open Access Journals (Sweden)

    Chakravarthy S K Vajhala

    Full Text Available Mannose-specific Allium sativum leaf agglutinin encoding gene (ASAL and herbicide tolerance gene (BAR were introduced into an elite cotton inbred line (NC-601 employing Agrobacterium-mediated genetic transformation. Cotton transformants were produced from the phosphinothricin (PPT-resistant shoots obtained after co-cultivation of mature embryos with the Agrobacterium strain EHA105 harbouring recombinant binary vector pCAMBIA3300-ASAL-BAR. PCR and Southern blot analysis confirmed the presence and stable integration of ASAL and BAR genes in various transformants of cotton. Basta leaf-dip assay, northern blot, western blot and ELISA analyses disclosed variable expression of BAR and ASAL transgenes in different transformants. Transgenes, ASAL and BAR, were stably inherited and showed co-segregation in T1 generation in a Mendelian fashion for both PPT tolerance and insect resistance. In planta insect bioassays on T2 and T3 homozygous ASAL-transgenic lines revealed potent entomotoxic effects of ASAL on jassid and whitefly insects, as evidenced by significant decreases in the survival, development and fecundity of the insects when compared to the untransformed controls. Furthermore, the transgenic cotton lines conferred higher levels of resistance (1-2 score with minimal plant damage against these major sucking pests when bioassays were carried out employing standard screening techniques. The developed transgenics could serve as a potential genetic resource in recombination breeding aimed at improving the pest resistance of cotton. This study represents the first report of its kind dealing with the development of transgenic cotton resistant to two major sap-sucking insects.

  14. An atypical IgM class platelet cold agglutinin induces GPVI-dependent aggregation of human platelets.

    Science.gov (United States)

    Sánchez Guiu, I M; Martínez-Martinez, I; Martínez, C; Navarro-Fernandez, J; García-Candel, F; Ferrer-Marín, F; Vicente, V; Watson, S P; Andrews, R K; Gardiner, E E; Lozano, M L; Rivera, J

    2015-08-01

    Platelet cold agglutinins (PCA) cause pseudothrombocytopenia, spurious thrombocytopenia due to ex vivo platelet clumping, complicating clinical diagnosis, but mechanisms and consequences of PCA are not well defined. Here, we characterised an atypical immunoglobulin (Ig)M PCA in a 37-year-old woman with lifelong bleeding and chronic moderate thrombocytopenia, that induces activation and aggregation of autologous or allogeneic platelets via interaction with platelet glycoprotein (GP)VI. Patient temperature-dependent pseudothrombocytopenia was EDTA-independent, but was prevented by integrin αIIbβ3 blockade. Unstimulated patient platelets revealed elevated levels of bound IgM, increased expression of activation markers (P-selectin and CD63), low GPVI levels and abnormally high thromboxane (TX)A2 production. Patient serum induced temperature- and αIIbβ3-dependent decrease of platelet count in allogeneic donor citrated platelet-rich plasma (PRP), but not in PRP from Glanzmann's thrombasthenia or afibrinogenaemia patients. In allogeneic platelets, patient plasma induced shape change, P-selectin and CD63 expression, (14)C-serotonin release, and TXA2 production. Activation was not inhibited by aspirin, cangrelor or blocking anti-Fc receptor (FcγRIIA) antibody, but was abrogated by inhibitors of Src and Syk, and by a soluble GPVI-Fc fusion protein. GPVI-deficient platelets were not activated by patient plasma. These data provide the first evidence for an IgM PCA causing platelet activation/aggregation via GPVI. The PCA activity persisted over a five-year follow-up period, supporting a causative role in patient chronic thrombocytopenia and bleeding. PMID:25994029

  15. Wheat germ agglutinin-conjugated PLGA nanoparticles for enhanced intracellular delivery of paclitaxel to colon cancer cells.

    Science.gov (United States)

    Wang, Chunxia; Ho, Paul C; Lim, Lee Yong

    2010-11-15

    The purpose of this study was to investigate the potentiation of the anticancer activity and enhanced cellular retention of paclitaxel-loaded PLGA nanoparticles after surface conjugation with wheat germ agglutinin (WGA) against colon cancer cells. Glycosylation patterns of representative colon cancer cells confirmed the higher expression levels of WGA-binding glycoproteins in the Caco-2 and HT-29 cells, than in the CCD-18Co cells. Cellular uptake and in vitro cytotoxicity of WNP (final formulation) against colon cell lines was evaluated alongside control formulations. Confocal microscopy and quantitative analysis of intracellular paclitaxel were used to monitor the endocytosis and retention of nanoparticles inside the cells. WNP showed enhanced anti-proliferative activity against Caco-2 and HT-29 cells compared to corresponding nanoparticles without WGA conjugation (PNP). The greater efficacy of WNP was associated with higher cellular uptake and sustained intracellular retention of paclitaxel, which in turn was attributed to the over-expression of N-acetyl-D-glucosamine-containing glycoprotein on the colon cell membrane. WNP also demonstrated increased intracellular retention in the Caco-2 (30% of uptake) and HT-29 (40% of uptake) cells, following post-uptake incubation with fresh medium, compared to the unconjugated PNP nanoparticles (18% in Caco-2) and (27% in HT-29), respectively. Cellular trafficking study of WNP showed endocytosed WNP could successful escape from the endo-lysosome compartment and release into the cytosol with increasing incubation time. It may be concluded that WNP has the potential to be applied as a targeted delivery platform for paclitaxel in the treatment of colon cancer. PMID:20804835

  16. Labelling of neurons in the rat superior cervical ganglion after injection of wheat-germ agglutinin-horseradish peroxidase into the contralateral ganglion: evidence of transneuronal labelling.

    OpenAIRE

    ATASEVER, A.; Palaoğlu, S; Erbengi, A; Celik, H H

    1994-01-01

    Recent studies have shown that injection of the tracer wheat-germ agglutinin-horseradish peroxidase (WGA-HRP) into the superior cervical ganglion (SCG) of one side results in labelling of neurons in the contralateral SCG and the stellate ganglion. This study was designed to verify whether or not bilateral projections from the superior cervical ganglion to the midline structures, particularly to the pineal gland, play a role in the transport of WGA-HRP to the contralateral SCG. One group of ra...

  17. Differences in the labeling of axons of passage by wheat germ agglutinin after uptake by cut peripheral nerve versus injections within the central nervous system

    International Nuclear Information System (INIS)

    Injections of a radiolabeled derivative of wheat germ agglutinin (WGA) in the subcortical white matter of the cerebral cortex in mice do not give rise to autoradiographic labeling of axon systems coursing through this fiber mass. Exposing the cut-end of mouse tibial nerve to WGA does, however, produce labeling within dorsal root ganglia and the spinal cord. These findings are discussed with consideration for dissimilarity in mode of central versus peripheral administration of the tracer, as well as in the light of potential relative differences in the uptake and transport of WGA and HRP. (Auth.)

  18. Early Serologic Diagnosis of Mycoplasma pneumoniae Pneumonia: An Observational Study on Changes in Titers of Specific-IgM Antibodies and Cold Agglutinins

    OpenAIRE

    Lee, Sung-Churl; Youn, You-Sook; Rhim, Jung-Woo; Kang, Jin-Han; Lee, Kyung-Yil

    2016-01-01

    Abstract There have been some limitations on early diagnosis of Mycoplasma pneumoniae (MP) infection because of no immunoglobulin M (IgM) responses and variable detection rates of polymerase chain reaction in the early stage of the disease. We wanted to discuss regarding early diagnostic method using short-term paired titration of MP-specific IgM and cold agglutinins (CAs) in the early stage of MP pneumonia. The participants of this study were 418 children with MP pneumonia during 2 recent ep...

  19. Expression of peanut agglutinin-binding mucin-type glycoprotein in human esophageal squamous cell carcinoma as a marker

    Directory of Open Access Journals (Sweden)

    Balakrishnan Ramathilakam

    2003-11-01

    Full Text Available Abstract Background The TF (Thomson – Friedenreich blood group antigen behaves as an onco-foetal carcinoma-associated antigen, showing increased expression in malignancies and its detection and quantification can be used in serologic diagnosis mainly in adenocarcinomas. This study was undertaken to analyze the sera and tissue level detectable mucin-type glycoprotein (TF-antigen by Peanut agglutinin (PNA and its diagnostic index in serum as well tissues of human esophageal squamous cell carcinoma as marker. Results We examined 100 patients for serological analysis by Enzyme Linked Lectin Assay (ELISA and demonstrated a sensitivity of 87.5%, specificity of 90% and a positive predictive value of 95%. The immuno-histochemical localization of TF antigen by Fluorescence Antigen Technique (FAT in 25 specimens of normal esophageal squamous epithelium specimens and 92 specimens with different grades of, allowed a quicker and more precise identification of its increased expression and this did not correlate with gender and tumor size. There was a positive correlation between membrane bound TF antigen expression with different histological progression, from well differentiated to poorly differentiated, determined by PNA binding. Specimens showed morphological changes and a pronounced increase in PNA binding in Golgi apparatus, secretory granules of the cytosol of well differentiated and an increased cell membrane labeling in moderately and poorly differentiated, when compared with ESCC and normal tissues. Conclusion The authors propose that the expression of TF-antigen in human may play an important role during tumorigenesis establishing it as a chemically well-defined carcinoma-associated antigen. Identification of the circulating TF-antigen as a reactive form and as a cryptic form in the healthy individuals, using PNA-ELLA and Immunohistochemical analysis of TF antigen by FAT is positively correlated with the different histological grades as a simple

  20. Transgenic rice expressing Allium sativum leaf agglutinin (ASAL exhibits high-level resistance against major sap-sucking pests

    Directory of Open Access Journals (Sweden)

    Vudem Dasavantha

    2008-10-01

    Full Text Available Abstract Background Rice (Oryza sativa productivity is adversely impacted by numerous biotic and abiotic factors. An approximate 52% of the global production of rice is lost annually owing to the damage caused by biotic factors, of which ~21% is attributed to the attack of insect pests. In this paper we report the isolation, cloning and characterization of Allium sativum leaf agglutinin (asal gene, and its expression in elite indica rice cultivars using Agrobacterium-mediated genetic transformation method. The stable transgenic lines, expressing ASAL, showed explicit resistance against major sap-sucking pests. Results Allium sativum leaf lectin gene (asal, coding for mannose binding homodimeric protein (ASAL from garlic plants, has been isolated and introduced into elite indica rice cultivars susceptible to sap-sucking insects, viz., brown planthopper (BPH, green leafhopper (GLH and whitebacked planthopper (WBPH. Embryogenic calli of rice were co-cultivated with Agrobacterium harbouring pSB111 super-binary vector comprising garlic lectin gene asal along with the herbicide resistance gene bar, both under the control of CaMV35S promoter. PCR and Southern blot analyses confirmed stable integration of transgenes into the genomes of rice plants. Northern and western blot analyses revealed expression of ASAL in different transgenic rice lines. In primary transformants, the level of ASAL protein, as estimated by enzyme-linked immunosorbent assay, varied between 0.74% and 1.45% of the total soluble proteins. In planta insect bioassays on transgenic rice lines revealed potent entomotoxic effects of ASAL on BPH, GLH and WBPH insects, as evidenced by significant decreases in the survival, development and fecundity of the insects. Conclusion In planta insect bioassays were carried out on asal transgenic rice lines employing standard screening techniques followed in conventional breeding for selection of insect resistant plants. The ASAL expressing rice

  1. Distribution features of lymphocytes with peanut agglutinin positive receptors in gums epithelium of rats in norm and after intrauterine antigenic action

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    Burega Yu.

    2015-09-01

    Full Text Available Background. According to the conception “Lymphocyte is the main factor of morphogenesis” changes in lymphocyte receptor repertory, induced by antigenic action in the fetal period of development, influence on organs and tissues development after birth. Functional activity of immunological immature PNA+ lymphocytes inducing the change in functioning, imbalance in formation cells of microenvironment, synthesis of intracellular substance and the fibers of extracellular matrix leads to violation of morphological and functional condition of organs. Objective. Determine the features of distribution of lymphocytes with receptors to peanut agglutinin in gingival epithelium of rats in norm and after intrauterine antigenic action. Methods. The object of the research was: 224 jaws of 112 white laboratory rats. The rats divided into three groups. First group – intact rats. Second group –rats, which were introduced 0,05 ml solution of antigen in the amniotic fluid on the 18th day of pregnancy by the method of N. Voloshyn, the third group – control, the animals were introduced intrauterine 0,05 ml of physiological solution on the 18th day of pregnancy. The antigen was split vaccine Vaxigrip 2009. Results and conclusion. In newborn animals, after intrauterine antigen action it was determined significantly increased content of PNA+ lymphocytes in the epithelium of gingival mucous, compared with control group, where PNA+ lymphocytes number gradually decreases. On the 11 th day of life, in animals of second group, quantity of intraepithelial PNA+ lymphocytes remains higher. On 45th day of postnatal formation its share does not significantly differ from similar indicators in all groups and decreases compared with neonatal period. Citation: Burega Yu. Distribution features of lymphocytes with peanut agglutinin positive receptors in gums epithelium of rats in norm and after intrauterine antigenic action. Morphologia. 2015;9(3:8-11.

  2. AGLUTININAS ANTILEPTÓSPIRA EM EQÜINOS NO ESTADO DE GOIÁS ANTILEPTOSPIRA AGGLUTININS IN EQUINES IN THE GOIÁS STATE, BRAZIL

    Directory of Open Access Journals (Sweden)

    Marly Francisca Cândido

    2007-09-01

    Full Text Available

    Foram analisados 694 hemossoros de eqüinos, procedentes de sete rnunicípios goianos, encontrando-se uma positividade para aglutininas antileptóspira que variou de 8,8 a 25,0%, com o valor médio de 14,4%. Diferentes sorotipos foram pesquisados, pela técnica de GALTON et alii (1962, encontrando-se positividade para os sorotipos grippotyphosa, wolffi, ballum e australis. Os autores concluíram pela necessidade de prosseguimento de novas pesquisas, com a finalidade de averiguar-se a presença ou não de diferentes sorotipos observados em outros Estados e não encontrados na presente pesquisa, além da realização de estudo epidemiológico local e regional.

    694 equine serum blood tests were made from horses of 7 counties of the State of Goiás. Positive rates of Leptospira agglutinins varied from 8.8 to 25.0% with an average of 14.4%. Different serum types were studied by the technique developed by GALTON et alii (1962 finding positiveness for the following serum types: grippotyphosa, wolffi, ballum and australis.

  3. Enhanced Resistance of Snowdrop Lectin (Galanthus nivalis L. Agglutinin)-Expressing Maize to Asian Corn Borer (Ostrinia furnacalis Guenée)

    Institute of Scientific and Technical Information of China (English)

    Zhao-Yu WANG; Xiao-Fen SUN; Fei WANG; Ke-Xuan TANG; Ju-Ren ZHANG

    2005-01-01

    In order to enhance the resistance to pests, transgenic maize (Zea mays L.) plants from elite inbred lines containing the gene encoding snowdrop lectin (Galanthus nivalis L. agglutinin; GNA) under control of a phloem-specific promoter were generated through the Agrobacterium tumefaciens-mediated also studied. Thirty-six independently derived plants were subjected to molecular analyses. The level of GNA expression at 0.13%-0.28% of total soluble protein was observed in different transgenic plants. The progeny of three GNA-expressing independent transformants that were derived separately from the elite inbred lines DH4866, DH9942, and 8902, were selected for examination of resistance to ACB. These plants synthesized GNA at levels above 0.24% total soluble protein and enhanced resistance to ACB was demonstrated by exposing the plants to insects under greenhouse conditions. Semi-artificial diet bioassays also showed the toxic effect of GNA on ACB. Field evaluation of the transgenic plants supported the results from the artificial trial. In the present study, we have obtained new insect-resistant maize material for further breeding work and have found that GNA-expressing plants not only gained significant resistance to homopterans, but also showed toxicity to ACB, which is a type of Lepidoptera.

  4. An Unusual Member of the Papain Superfamily: Mapping the Catalytic Cleft of the Marasmius oreades agglutinin (MOA) with a Caspase Inhibitor.

    Science.gov (United States)

    Cordara, Gabriele; van Eerde, André; Grahn, Elin M; Winter, Harry C; Goldstein, Irwin J; Krengel, Ute

    2016-01-01

    Papain-like cysteine proteases (PLCPs) constitute the largest group of thiol-based protein degrading enzymes and are characterized by a highly conserved fold. They are found in bacteria, viruses, plants and animals and involved in a number of physiological and pathological processes, parasitic infections and host defense, making them interesting targets for drug design. The Marasmius oreades agglutinin (MOA) is a blood group B-specific fungal chimerolectin with calcium-dependent proteolytic activity. The proteolytic domain of MOA presents a unique structural arrangement, yet mimicking the main structural elements in known PLCPs. Here we present the X-ray crystal structure of MOA in complex with Z-VAD-fmk, an irreversible caspase inhibitor known to cross-react with PLCPs. The structural data allow modeling of the substrate binding geometry and mapping of the fundamental enzyme-substrate interactions. The new information consolidates MOA as a new, yet strongly atypical member of the papain superfamily. The reported complex is the first published structure of a PLCP in complex with the well characterized caspase inhibitor Z-VAD-fmk. PMID:26901797

  5. In vivo toxicity and immunogenicity of wheat germ agglutinin conjugated poly(ethylene glycol)-poly(lactic acid) nanoparticles for intranasal delivery to the brain

    International Nuclear Information System (INIS)

    Biodegradable polymer-based nanoparticles have been widely studied to deliver therapeutic agents to the brain after intranasal administration. However, knowledge as to the side effects of nanoparticle delivery system to the brain is limited. The aim of this study was to investigate the in vivo toxicity and immunogenicity of wheat germ agglutinin (WGA) conjugated poly(ethylene glycol)-poly(lactic acid) nanoparticles (WGA-NP) after intranasal instillation. Sprague-Dawley rats were intranasally given WGA-NP for 7 continuous days. Amino acid neurotransmitters, lactate dehydrogenase (LDH) activity, reduced glutathione (GSH), acetylcholine, acetylcholinesterase activity, tumor necrosis factor α (TNF-α) and interleukin-8 (IL-8) in rat olfactory bulb (OB) and brain were measured to estimate the in vivo toxicity of WGA-NP. Balb/C mice were intranasally immunized by WGA-NP and then WGA-specific antibodies in serum and nasal wash were detected by indirect ELISA. WGA-NP showed slight toxicity to brain tissue, as evidenced by increased glutamate level in rat brain and enhanced LDH activity in rat OB. No significant changes in acetylcholine level, acetylcholinesterase activity, GSH level, TNF-α level and IL-8 level were observed in rat OB and brain for the WGA-NP group. WGA-specific antibodies in mice serum and nasal wash were not increased after two intranasal immunizations of WGA-NP. These results demonstrate that WGA-NP is a safe carrier system for intranasal delivery of therapeutic agents to the brain.

  6. Topology of the polypeptide chain in the complex of agglutinin from castor bean seeds with β-D-galactose in the crystalline state

    International Nuclear Information System (INIS)

    The three-dimensional structure of the complex of agglutinin from Ricinus communis with β-D-galactose was established and refined at 2.5 A resolution by X-ray structure analysis. Biocrystals were obtained using dialysis through a semipermeable membrane. X-ray intensity data (Rmerge = 4.6%) were collected from one crystal at 100 K using synchrotron radiation at the DESY outstation [European Molecular Biology Laboratory (EMBL), Hamburg, Germany]. The initial phases were calculated by the molecular replacement method. The atoms of both protein and sugar molecules were localized. Unlike ricin, the ricinlike heterodimer RcA contains only one galactose-binding center in the region of the Asn46-Gly25-Trp37-Lys40 site in the first domain of the B subunit, whereas the second galactose-binding site of the B subunit is lost. One functionally important water molecule, which is bound to the residues Tyr123-Glu176-Arg179-Glu207, was revealed in the region of the active center in the A subunit

  7. Electrogenerated chemiluminescence biosensing for the detection of prostate PC-3 cancer cells incorporating antibody as capture probe and ruthenium complex-labelled wheat germ agglutinin as signal probe

    Energy Technology Data Exchange (ETDEWEB)

    Yang, Haiying [Key Laboratory of Applied Surface and Colloid Chemistry, Ministry of Education, School of Chemistry and Chemical Engineering, Shaanxi Normal University, Xi’an 710062 (China); Department of Chemistry, Yuncheng University, Yuncheng 044300 (China); Li, Zhejian; Shan, Meng; Li, Congcong; Qi, Honglan; Gao, Qiang [Key Laboratory of Applied Surface and Colloid Chemistry, Ministry of Education, School of Chemistry and Chemical Engineering, Shaanxi Normal University, Xi’an 710062 (China); Wang, Jinyi [College of Science and College of Veterinary Medicine, Northwest A& F University, Yangling 712100 (China); Zhang, Chengxiao, E-mail: cxzhang@snnu.edu.cn [Key Laboratory of Applied Surface and Colloid Chemistry, Ministry of Education, School of Chemistry and Chemical Engineering, Shaanxi Normal University, Xi’an 710062 (China)

    2015-03-10

    Highlights: • A novel biosensor was developed for the detection of prostate cancer cells. • The selectivity of the biosensor was improved using antibody as capture probe. • The biosensor showed the low extremely detection limit of 2.6 × 10{sup 2} cells mL{sup −1}. • The ruthenium complex-labelled WGA can be transported in the cell vesicles. - Abstract: A highly selective and sensitive electrogenerated chemiluminescence (ECL) biosensor for the detection of prostate PC-3 cancer cells was designed using a prostate specific antibody as a capture probe and ruthenium complex-labelled wheat germ agglutinin as a signal probe. The ECL biosensor was fabricated by covalently immobilising the capture probe on a graphene oxide-coated glassy carbon electrode. Target PC-3 cells were selectively captured on the surface of the biosensor, and then, the signal probe was bound with the captured PC-3 cells to form a sandwich. In the presence of tripropylamine, the ECL intensity of the sandwich biosensor was logarithmically directly proportion to the concentration of PC-3 cells over a range from 7.0 × 10{sup 2} to 3.0 × 10{sup 4} cells mL{sup −1}, with a detection limit of 2.6 × 10{sup 2} cells mL{sup −1}. The ECL biosensor was also applied to detect prostate specific antigen with a detection limit of 0.1 ng mL{sup −1}. The high selectivity of the biosensor was demonstrated in comparison with that of a lectin-based biosensor. The strategy developed in this study may be a promising approach and could be extended to the design of ECL biosensors for highly sensitive and selective detection of other cancer-related cells or cancer biomarkers using different probes.

  8. Electrogenerated chemiluminescence biosensing for the detection of prostate PC-3 cancer cells incorporating antibody as capture probe and ruthenium complex-labelled wheat germ agglutinin as signal probe

    International Nuclear Information System (INIS)

    Highlights: • A novel biosensor was developed for the detection of prostate cancer cells. • The selectivity of the biosensor was improved using antibody as capture probe. • The biosensor showed the low extremely detection limit of 2.6 × 102 cells mL−1. • The ruthenium complex-labelled WGA can be transported in the cell vesicles. - Abstract: A highly selective and sensitive electrogenerated chemiluminescence (ECL) biosensor for the detection of prostate PC-3 cancer cells was designed using a prostate specific antibody as a capture probe and ruthenium complex-labelled wheat germ agglutinin as a signal probe. The ECL biosensor was fabricated by covalently immobilising the capture probe on a graphene oxide-coated glassy carbon electrode. Target PC-3 cells were selectively captured on the surface of the biosensor, and then, the signal probe was bound with the captured PC-3 cells to form a sandwich. In the presence of tripropylamine, the ECL intensity of the sandwich biosensor was logarithmically directly proportion to the concentration of PC-3 cells over a range from 7.0 × 102 to 3.0 × 104 cells mL−1, with a detection limit of 2.6 × 102 cells mL−1. The ECL biosensor was also applied to detect prostate specific antigen with a detection limit of 0.1 ng mL−1. The high selectivity of the biosensor was demonstrated in comparison with that of a lectin-based biosensor. The strategy developed in this study may be a promising approach and could be extended to the design of ECL biosensors for highly sensitive and selective detection of other cancer-related cells or cancer biomarkers using different probes

  9. Crystal Structure of the C-terminal Region of Streptococcus mutans Antigen I/II and Characterization of Salivary Agglutinin Adherence Domains

    Energy Technology Data Exchange (ETDEWEB)

    Larson, Matthew R.; Rajashankar, Kanagalaghatta R.; Crowley, Paula J.; Kelly, Charles; Mitchell, Tim J.; Brady, L. Jeannine; Deivanayagam, Champion (King); (Cornell); (UAB); (Glasgow); (Florida)

    2012-05-29

    The Streptococcus mutans antigen I/II (AgI/II) is a cell surface-localized protein that adheres to salivary components and extracellular matrix molecules. Here we report the 2.5 {angstrom} resolution crystal structure of the complete C-terminal region of AgI/II. The C-terminal region is comprised of three major domains: C{sub 1}, C{sub 2}, and C{sub 3}. Each domain adopts a DE-variant IgG fold, with two {beta}-sheets whose A and F strands are linked through an intramolecular isopeptide bond. The adherence of the C-terminal AgI/II fragments to the putative tooth surface receptor salivary agglutinin (SAG), as monitored by surface plasmon resonance, indicated that the minimal region of binding was contained within the first and second DE-variant-IgG domains (C{sub 1} and C{sub 2}) of the C terminus. The minimal C-terminal region that could inhibit S. mutans adherence to SAG was also confirmed to be within the C{sub 1} and C{sub 2} domains. Competition experiments demonstrated that the C- and N-terminal regions of AgI/II adhere to distinct sites on SAG. A cleft formed at the intersection between these C{sub 1} and C{sub 2} domains bound glucose molecules from the cryo-protectant solution, revealing a putative binding site for its highly glycosylated receptor SAG. Finally, electron microscopy images confirmed the elongated structure of AgI/II and enabled building a composite tertiary model that encompasses its two distinct binding regions.

  10. Early Serologic Diagnosis of Mycoplasma pneumoniae Pneumonia: An Observational Study on Changes in Titers of Specific-IgM Antibodies and Cold Agglutinins.

    Science.gov (United States)

    Lee, Sung-Churl; Youn, You-Sook; Rhim, Jung-Woo; Kang, Jin-Han; Lee, Kyung-Yil

    2016-05-01

    There have been some limitations on early diagnosis of Mycoplasma pneumoniae (MP) infection because of no immunoglobulin M (IgM) responses and variable detection rates of polymerase chain reaction in the early stage of the disease. We wanted to discuss regarding early diagnostic method using short-term paired titration of MP-specific IgM and cold agglutinins (CAs) in the early stage of MP pneumonia.The participants of this study were 418 children with MP pneumonia during 2 recent epidemics (2006-2007 and 2011), and they were diagnosed by an anti-MP IgM antibody test (Serodia Myco II) examined twice during hospitalization at presentation and around discharge (mean of 3.4 ± 1.3 days apart). CA titers were simultaneously examined twice during study period. Anti-MP IgM antibody titer ≥1:40 and CA titer ≥1:4 were considered positive, respectively. The relationships between 2 IgM antibodies in the early stage were evaluated.Regarding MP-specific antibody titers, 148 patients showed a seroconversion, 245 patients exhibited increased titers, and 25 patients had unchanged higher titers (≥1:640) during hospitalization. The median MP-specific antibody titers at each examination time were 1:80 and 1:640, respectively; those of CAs were 1:8 and 1:32, respectively. Illness duration prior to admission showed a trend of association with both titers, and patients with shorter illness duration had a higher rate of negative titers or lower titers at each examination time. CAs and MP-specific antibody titers were correlated in the total patients at presentation and at 2nd examination (P < 0.001, respectively), and the diagnostic corresponding rates of CAs to IgM antibody test were 81% to 96% in patient subgroups.Short-term paired MP specific-IgM determinations in the acute stage may be used as a definitive diagnostic method for MP pneumonia. Paired CA titers showed a correlation with MP-specific antibody titers, suggesting they can be used as an adjuvant diagnostic

  11. Expressions of Proliferating Cell Nuclear Antigen and Wheat Germ Agglutinin Receptor in Human Bladder Carcinoma%膀胱癌增殖细胞核抗原与麦胚凝集素受体的相关关系

    Institute of Scientific and Technical Information of China (English)

    张士文; 葛根; 金伯涛

    2001-01-01

    [Purpose]To probe the relation of proliferating cell nuclear antigen (PCNA) and wheat germ agglutinin (WGA) receptors expressed in human bladder transitional cell carcinoma (TCC).[Methods]PCNA and WGA receptors were detected by immunohistochemical method (ABC method) in 63 specimens of TCC.[Results]We found that the distributions of PCNA and WGA receptors were increased with increase of histopathological grade in TCC (P<0.01).There was a higher expression in invasive tumors than that in superficial tumors (P<0.005),and there was a positive relation between PCNA and WGA receptors also.[Conclusion]It is shown that PCNA and WGA can be used as tumor markers for bladder cancer.%[目的 ]探讨增殖细胞核抗原 (proliferating cell nuclear antigen,PCNA)和麦胚凝集素 (wheat germ agglutinin,WGA)在膀胱移行细胞癌 (TCC)中表达的相关关系。 [方法 ]采用免疫组织化学 ABC法对 63例 TCC标本进行 PCNA和 WGA受体检测。 [结果 ]PCNA与 WGA的强阳性表达随着肿瘤的病理分级升高而增高;浸润性肿瘤中的 WGA受体的强阳性表达显著高于浅表性肿瘤 (P<0.05); PCNA与 WGA受体表达一致性良好,呈显著性相关 (P<0.005)。 [结论 ]我们认为 PCNA和 WGA受体均可作为 TCC的肿瘤标记物,证明了 TCC细胞的增殖活性增强将改变其细胞膜的抗原性。

  12. Expression of Vicia villosa agglutinin (VVA)-binding glycoprotein in primary breast cancer cells in relation to lymphatic metastasis: is atypical MUC1 bearing Tn antigen a receptor of VVA?

    Science.gov (United States)

    Kawaguchi, Takanori; Takazawa, Hiroshi; Imai, Shunsuke; Morimoto, Junji; Watanabe, Takanori; Kanno, Masahiko; Igarashi, Seiji

    2006-07-01

    Aberrant carbohydrate expression frequently occurs in breast cancer and may endow cells with metastatic potential. Here we first studied the relationship between expression of Vicia villosa agglutinin (lectin) (VVA)-binding carbohydrates and aggressive breast cancer. We then investigated the molecular characteristics of these glycoproteins and compared them with those of glycoproteins recognized by the mouse anti-Tn monoclonal antibody (MAb) HB-Tn1. Histochemical studies of samples from 322 cases of invasive ductal carcinoma demonstrated that VVA-binding carbohydrate expression correlated with tumor stage, lymphatic invasion, and lymph node metastasis (p=0.0385, p=0.0019, and p=0.0430. respectively). Western blotting analysis of frozen materials from 39 cases, under denaturing and reducing conditions, revealed that the major cancer cell-specific VVA-binding proteins were molecules of about 30, 33, and >200 kDa. Cases expressing the approximately 33 kDa molecule had significant lymphatic invasion more frequently than did cases not expressing this molecule (p=0.0076). Binding of VVA to the approximately 30 and approximately 33 kDa molecules was completely lost by preincubation of VVA with 1 mM Tn antigen (N-acetylgalactosamine alpha1-O-serine). The VVA-binding molecules appeared to react with VU-3C6 anti-MUC1 MAb. Expression of HB-Tn1 in breast cancer cells showed significant correlation with expression of VVA-binding carbohydrate(s) (p<0.0001) but HB-Tn1 reactivity was not clearly related to breast cancer aggressiveness. Because anti-Tn MAbs bound to Tn antigen clusters, we concluded that atypical MUC1 bearing the noncluster form of Tn antigen is implicated in aggressive growth of primary breast cancer cells, particularly in lymphatic metastasis. PMID:16752227

  13. Aglutininas antileptospíricas em búfalos do Vale do Ribeira, Estado de São Paulo Anti-leptospire agglutinins in buffaloes from Vale do Ribeira, São Paulo State, Brazil

    Directory of Open Access Journals (Sweden)

    Hélio Langoni

    1999-06-01

    Full Text Available Foram estudadas aglutininas antileptospíricas em 403 amostras de soro de búfalos, provenientes de sete Municípios do Vale do Ribeira, Estado de São Paulo, coletadas no período de janeiro de 1992 a junho de 1993. Utilizou-se o teste de soroaglutinação microscópica, considerando-se positivas as amostras cujo título fosse igual ou superior a 100. O maior título encontrado foi 1600 para o sorovar bratislava (1 amostra, seguido de 800 para wolffi (4 amostras. Do total, 152 (37,7% das amostras foram positivas, sendo que, dentre os sorovares testados, a prevalência em ordem decrescente foi: wolffi (68, 44,8%, icterohaemorrhagiae (51, 33,6%, hardjo (51, 33,6%, castellonis (25, 16,5%, djasiman (12, 7,9%, grippotyphosa (10, 6,6%, pomona (8, 5,2%, bratislava (6, 4,0%, copenhageni (5, 3,3% e tarassovi (4, 2,7%.A total of 403 buffaloes serum samples from seven counties of Vale do Ribeira, São Paulo State, Brazil, obtained between January 1992 and June 1993, were studied to determine the prevalence of anti-leptospire agglutinins using the microscopic serum agglutination test. A titre of 100 and above was considered positive. The highest titre found was 1600 to the sorovar bratislava (one sample, followed by 800 to wolffi (4 samples. 152 (37.7% samples from the total were positive to the serovars tested, and their prevalence, in decreasing order, was: wolffi (68, 44.8%, icterohaemorrhagiae (51, 33.6%, hardjo (51, 33.6%, castellonis (25, 16.5%, djasiman (12, 7.9%, grippotyphosa (10, 6.6%, pomona (8, 5.2%, bratislava (6, 4.0%, copenhageni (5, 3.3% and tarassovi (4, 2.7%.

  14. Cold agglutinin disease (CADwith autoimmune haemolytic anaemia: a case report of a coronary artery disease patient Doença por aglutininas a frio (DAC com anemia hemolítica auto-imune: relato de caso de um coronariopata

    Directory of Open Access Journals (Sweden)

    Leandro A. Barbosa

    2008-02-01

    Full Text Available Cold agglutinin disease (CAD with autoimmune haemolytic anemia is characterized by the production of harmful cold autoantibodies associated with increased red cell destruction during exposure to cold. The treatment of CAD is very difficult and a great effort is required to obtain therapeutic success. Cyclophosphamide is a potent immunosuppressive agent which is widely used in all bone marrow transplantation conditioning regimens for patients with acquired severe aplastic anemia. In this report, we describe the case of a coronary artery disease patient with severe CAD, but without lymphoproliferative disease, in which general measures and immunosuppressive therapies were adopted, there by avoiding blood transfusions.A doença por aglutininas a frio (CAD cursando com anemia hemolítica auto-imune (AHAI é decorrente da produção de autoanticorpos que reagem muito bem a baixas temperaturas, dirigidos contra hemácias autólogas. A habilidade desses anticorpos em destruir as hemácias encontra-se diretamente relacionada à sua capacidade em fixar complemento durante a exposição do paciente a baixas temperaturas. A AHAI por anticorpos frios pode ser idiopática - ausência de doença de base - ou secundária, geralmente associada a desordens linfoproliferativas de células B ou determinados processos infecciosos. A hemólise é intravascular, através de aglutininas da classe IgM, com teste direto da antiglobulina humana positivo para complemento. O tratamento da CAD é difícil, exigindo um esforço contínuo, necessário para se obter sucesso terapêutico. A ciclofosfamida é um agente imunossupressor potente, amplamente utilizado em transplantes de medula óssea, particularmente nos portadores de anemia aplástica. Descrevemos o caso de um coronariopata portador de CAD severa, cuja exploração diagnóstica excluiu doença linfoproliferativa. Adotamos medidas gerais de suporte e terapia imunossupressora, coibindo o uso de hemotransfusões.

  15. Determination of trace alkaline phosphatase by affinity adsorption solid substrate room temperature phosphorimetry based on wheat germ agglutinin labeled with 8-quinolineboronic acid phosphorescent molecular switch and prediction of diseases

    Science.gov (United States)

    Liu, Jia-Ming; Gao, Hui; Li, Fei-Ming; Shi, Xiu-Mei; Lin, Chang-Qing; Lin, Li-Ping; Wang, Xin-Xing; Li, Zhi-Ming

    2010-09-01

    The 8-quinolineboronic acid phosphorescent molecular switch (abbreviated as PMS-8-QBA. Thereinto, 8-QBA is 8-quinolineboronic acid, and PMS is phosphorescent molecular switch) was found for the first time. PMS-8-QBA, which was in the "off" state, could only emit weak room temperature phosphorescence (RTP) on the acetyl cellulose membrane (ACM). However, PMS-8-QBA turned "on" automatically for its changed structure, causing that the RTP of 8-QBA in the system increased, after PMS-8-QBA-WGA (WGA is wheat germ agglutinin) was formed by reaction between -OH of PMS-8-QBA and -COOH of WGA. More interesting is that the -NH 2 of PMS-8-QBA-WGA could react with the -COOH of alkaline phosphatase (AP) to form the affinity adsorption (AA) product WGA-AP-WGA-8-QBA-PMS (containing -NH-CO- bond), which caused RTP of the system to greatly increase. Thus, affinity adsorption solid substrate room temperature phosphorimetry using PMS-8-QBA as labelling reagent (PMS-8-QBA-AA-SSRTP) for the determination of trace AP was established. The method had many advantages, such as high sensitivity (the detection limit (LD) was 2.5 zg spot -1. For sample volume of 0.40 μl spot -1, corresponding concentration was 6.2 × 10 -18 g ml -1), good selectivity (the allowed concentration of coexisting material was higher, when the relative error was ±5%), high accuracy (applied to detection of AP content in serum samples, the result was coincided with those obtained by enzyme-linked immunoassay), which was suitable for the detection of trace AP content in serum samples and the forecast of human diseases. Meanwhile, the mechanism of PMS-8-QBA-AASSRTP was discussed. The new field of analytical application and clinic diagnosis technique of molecule switch are exploited, based on the phosphorescence characteristic of PMS-8-QBA, the AA reaction between WGA and AP, as well as the relation between AP content and human diseases. The research results promote the development and interpenetrate among molecule

  16. FREQÜÊNCIA DE AGLUTININAS ANTI-Brucella abortus EM CAPRINOS E OVINOS DO SERTÃO DO ESTADO DE PERNAMBUCO, BRASIL FREQUENCY OF ANTI-Brucella abortus AGGLUTININS IN GOATS AND sheep OF THE “SERTÃO” (BACKLANDS OF THE STATE OF PERNAMBUCO, BRAZIL

    Directory of Open Access Journals (Sweden)

    Vânia Lúcia de Assis Santana

    2008-12-01

    Full Text Available Objetivou-se investigar a freqüência de aglutininas anti-Brucella abortus em caprinos e ovinos do Sertão do Estado de Pernambuco, Brasil. Foram processadas 700 amostras de soros sangüíneos, das quais 340 eram da espécie caprina (115 machos e 225 fêmeas e 360 (136 machos e 224 fêmeas ovina. Empregou-se a técnica do antígeno acidificado tamponado (AAT corado com rosa bengala (RB. Das 340 amostras de caprinos avaliadas, duas (0,6% foram reagentes ao AAT. Não se observaram associações significativas para as variáveis faixa etária (p= 0,430, raça (p= 0,936 e sexo (p= 0,562. Das 360 amostras de ovinos, nove (2,5% foram reagentes. Também não houve associação significativa entre as variáveis analisadas e a soropositividade para brucelose: faixa etária (p= 0,522; raça (p= 0,576 e sexo (p= 0,461. Verificou-se associação significativa (p= 0,042 entre as espécies estudadas e soropositividade para brucelose nos animais investigados. A soropositividade para Brucella abortus em caprinos e ovinos foi descrita pela primeira vez no Sertão de Pernambuco, fato que pode dificultar o sucesso do Programa Nacional de Controle e Erradicação da Brucelose, tendo em vista que nessa região é comum a criação consorciada de pequenos ruminantes com bovinos, além de representar riscos à Saúde Pública.

    PALAVRAS-CHAVES: Brucelose, ovinos, caprinos, pequenos ruminantes, sorodiagnóstico. The objective was to investigate the frequency of anti-Brucella abortus agglutinins in goats and sheep of the backlands of the State of Pernambuco, Brazil. 700 samples of sanguine serums were processed, of which 340 were of the goat (115 males and 225 females and 360 (136 males and 224 females sheep. The technique of the Tamponed Acidified Antigen (AAT dyed with Bengalese Rose (BR was used. Of the 340 samples of goat evaluated two (0.6% were reactive to AAT. Significant associations were not observed for the variable age group (p = 0.430; race (p = 0

  17. Diversidade antigênica entre amostras de Arcobacter spp isoladas de suínos no Rio Grande do Sul e presença de anticorpos aglutinantes em amostras de soro de porcas com problemas reprodutivos Antigenic diversity among strains of Arcobacter spp isolated from pigs in Rio Grande do Sul, Brazil and presence of agglutinin titers in serum samples of sows with reproductive problems

    Directory of Open Access Journals (Sweden)

    Sérgio José de Oliveira

    1999-12-01

    there was presence of agglutinins in serum samples.

  18. Overexpression of glycosylated proteins in cervical cancer recognized by the Machaerocereus eruca agglutinin Overexpression of glycosylated proteins in cervical cancer recognized by the Machaerocereus eruca agglutinin

    Directory of Open Access Journals (Sweden)

    Carlos Solórzano

    2012-10-01

    Full Text Available In cervical cancer, glycosylation has been suggested as being involved in both its carcinogenesis and
    invasive capacity. In this work, we analyzed mucin type O-glycosylation in biopsies of invasive cervical cancer in
    FIGO stage II B through histochemistry using lectins specific for O-glycosidically linked glycans. Our results
    reveal that the lectin Machaerocereus eruca (MeA, specific for Gal in a Fuca1,2 (GalNAca1,3 Galb1,4 showed
    increased recognition of tumoral cells and tumoral stroma tissue compared to other lectins with similar specificity;
    healthy cervical tissue was negative for MeA. Trypsin treatment of recognized tissues abolished MeA’s recognition;
    moreover, interaction of MeA was inhibited with oligosaccharides from mucin. As demonstrated by
    Western blot of 2-D electrophoresis, MeA recognized ten glycoproteins in the range from 122 to 42 kDa in
    cervical cancer lysates. The LC-ESI-MS/MS analysis of the MeAs’ recognized peptides revealed that the latter
    matched mainly with the amino acid sequences of lamin A/C, vimentin, elongation factor 2, keratin 1, and beta
    actin. Our results suggest that MeA recognizes a complex of over-expressed O-glycosidically-linked proteins that
    play a relevant role in cervical cancer’s invasive capacity. O-glycosylation participates in the disassembly of intercellular
    junctions favoring cancer progression.In cervical cancer, glycosylation has been suggested as being involved in both its carcinogenesis and
    invasive capacity. In this work, we analyzed mucin type O-glycosylation in biopsies of invasive cervical cancer in
    FIGO stage II B through histochemistry using lectins specific for O-glycosidically linked glycans. Our results
    reveal that the lectin Machaerocereus eruca (MeA, specific for Gal in a Fuca1,2 (GalNAca1,3 Galb1,4 showed
    increased recognition of tumoral cells and tumoral stroma tissue compared to other lectins with similar specificity;
    healthy cervical tissue was negative for MeA. Trypsin treatment of recognized tissues abolished MeA’s recognition;
    moreover, interaction of MeA was inhibited with oligosaccharides from mucin. As demonstrated by
    Western blot of 2-D electrophoresis, MeA recognized ten glycoproteins in the range from 122 to 42 kDa in
    cervical cancer lysates. The LC-ESI-MS/MS analysis of the MeAs’ recognized peptides revealed that the latter
    matched mainly with the amino acid sequences of lamin A/C, vimentin, elongation factor 2, keratin 1, and beta
    actin. Our results suggest that MeA recognizes a complex of over-expressed O-glycosidically-linked proteins that
    play a relevant role in cervical cancer’s invasive capacity. O-glycosylation participates in the disassembly of intercellular
    junctions favoring cancer progression.

  19. Neuroglial response to neuron injury. A study using intraneural injection of ricinus communis agglutinin-60.

    OpenAIRE

    Ling, E A; Wen, C Y; J.Y. Shieh; Yick, T Y; Leong, S K

    1989-01-01

    The present study has shown the selective destruction of large ventral horn neurons in the lumbosacral cord segments following a single injection of RCA-60 into the sciatic nerve. The neurons appeared to undergo structural alteration beginning 3 days after the RCA application. In the postoperative period extending from 1 to 60 days, degeneration of neurons was progressive and irreversible and this elicited a rapid increase in the number of microglial cells. They were most numerous in the 7 da...

  20. Neuroglial response to neuron injury. A study using intraneural injection of ricinus communis agglutinin-60.

    Science.gov (United States)

    Ling, E A; Wen, C Y; Shieh, J Y; Yick, T Y; Leong, S K

    1989-06-01

    The present study has shown the selective destruction of large ventral horn neurons in the lumbosacral cord segments following a single injection of RCA-60 into the sciatic nerve. The neurons appeared to undergo structural alteration beginning 3 days after the RCA application. In the postoperative period extending from 1 to 60 days, degeneration of neurons was progressive and irreversible and this elicited a rapid increase in the number of microglial cells. They were most numerous in the 7 days postoperative animals. The massive microglial cells penetrated the neuropil and appeared to strip off the axon terminals from the postsynaptic somata. Occasional axon terminals were phagocytosed by microglia. The numerous microglial cells often formed a multilayered 'barrier' encircling the somata of the RCA-poisoned neurons which eventually became totally disorganised. It is postulated that in the course of neuronal degeneration induced by RCA, microglial cells serve to prevent the leakage or diffusion of the toxic lectin from the neuronal somata into the neighbouring neuropil. They also function as scavenger cells in the removal of degenerating myelinated axons in the longer surviving rats. Oligodendrocytes do not appear to react actively to the degeneration process. However, astrocytes showed a significant increase in the 7 and 15 day postoperative rats and this coincided with the presence of mitotic astrocytes in the same period. PMID:2606792

  1. Ulex europaeus agglutinin II (UEA-II) is a novel, potent inhibitor of complement activation

    OpenAIRE

    Lekowski, Robert; Collard, Charles D.; Reenstra, Wende R.; Stahl, Gregory L.

    2001-01-01

    Complement is an important mediator of vascular injury following oxidative stress. We recently demonstrated that complement activation following endothelial oxidative stress is mediated by mannose-binding lectin (MBL) and activation of the lectin complement pathway. Here, we investigated whether nine plant lectins which have a binding profile similar to that of MBL competitively inhibit MBL deposition and subsequent complement activation following human umbilical vein endothelial cell (HUVEC)...

  2. Anti-leptospiral agglutinins in marmosets (Saguinus oedipus and Saguinus leucopus from illegal trade

    Directory of Open Access Journals (Sweden)

    Viviana Gonzalez-Astudillo

    2015-09-01

    Full Text Available Objective. Determine the infection status with pathogenic Leptospira of one Saguinus oedipus and nine Saguinus leucopus at the Cali Zoo that had been confiscated in Colombia from illegal trade. Materials and methods. A full physical examination, blood work, urinalysis were conducted in all individuals during the reception health check-up, in addition to running the microagglutination test with a pool of 19 serovars, with a starting dilution of 1:50. Results. A high positive titer (≥1:3200 to Leptospira alexanderi serovar manhao in an asymptomatic S. oedipus was detected. All S. leucopus tested negative or less than 1:50. Conclusions. Captive locations have been documented to artificially enhance opportunities to come into contact with contaminated bodily fluids from peridomestic rodents. However, infectious diseases acquired during the illegal transport of wildlife to major metropolitan centers are rarely considered a wildlife conservation or public health threat. Infection with zoonotic pathogens should also be considered an additional threat to endangered wild primates involved in illegal trade, which could hamper reintroduction efforts or other population management procedures for primate species with restricted and fragmented distributions.

  3. Dynamics of Agglutinin-Like Sequence (ALS) Protein Localization on the Surface of Candida Albicans

    Science.gov (United States)

    Coleman, David Andrew

    2009-01-01

    The ALS gene family encodes large cell-surface glycoproteins associated with "C. albicans" pathogenesis. Als proteins are thought to act as adhesin molecules binding to host tissues. Wide variation in expression levels among the ALS genes exists and is related to cell morphology and environmental conditions. "ALS1," "ALS3," and "ALS4" are three of…

  4. Leptospiral agglutinins in captive and free ranging non-human primates in Sarawak, Malaysia

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    S. Thayaparan

    2014-06-01

    Full Text Available Aim: The proposed study was carried out to determine the extent of exposure to leptospirosis in non-human primates. Materials and Methods: Trapping of non-human primates was carried out opportunistically around the Bako National Park and the Matang Wildlife Center in the vicinity of human settlements and tourism areas of Sarawak. Blood samples were obtained from the saphenous vein to determine the presence of antibodies by the Microscopic Agglutination Test (MAT to 17 serovars of Leptospira commonly found in Malaysia. Results: This study reports the screening of twelve primates (eight captive and four free ranging for leptospirosis. Eight of the 12 monkeys (66.6%; 95% CI 34.9-90.1 reacted against one or two serovars of Leptospira (Lai and Leptospira Lepto175. The serovar Lai is considered pathogenic for different mammals, including humans. Leptospira Lepto 175 has been identified as an intermediate strain and further studies are being undertaken on this serovar. Conclusion: These results are important as primates may act as reservoirs of Leptospira spp. for humans, which may potentially affect tourism (economic loss, conservation efforts and public health.

  5. Postmitotic basal cells in squamous cell epithelia are identified with Dolichos biflorus agglutinin - functional consequences

    Czech Academy of Sciences Publication Activity Database

    Hrdličková-Celá, E.; Plzák, J.; Holíková, Z.; Dvořánková, B.; Smetana, Karel

    2001-01-01

    Roč. 109, č. 10 (2001), s. 714-720. ISSN 0903-4641 R&D Projects: GA AV ČR KSK4055109 Institutional research plan: CEZ:AV0Z4050913 Keywords : squamous cell epithelia * carcinoma * lectin Subject RIV: CE - Biochemistry Impact factor: 1.924, year: 2001

  6. Wisteria Floribunda Agglutinin-Labeled Perineuronal Nets in the Mouse Inferior Colliculus, Thalamic Reticular Nucleus and Auditory Cortex

    OpenAIRE

    Sarah M. Fader; Kazuo Imaizumi; Yuchio Yanagawa; Lee, Charles C.

    2016-01-01

    Perineuronal nets (PNNs) are specialized extracellular matrix molecules that are associated with the closing of the critical period, among other functions. In the adult brain, PNNs surround specific types of neurons, however the expression of PNNs in the auditory system of the mouse, particularly at the level of the midbrain and forebrain, has not been fully described. In addition, the association of PNNs with excitatory and inhibitory cell types in these structures remains unknown. Therefore...

  7. Transsynaptic transport of wheat germ agglutinin expressed in a subset of type II taste cells of transgenic mice

    Directory of Open Access Journals (Sweden)

    Mosinger Bedrich

    2008-10-01

    Full Text Available Abstract Background Anatomical tracing of neural circuits originating from specific subsets of taste receptor cells may shed light on interactions between taste cells within the taste bud and taste cell-to nerve interactions. It is unclear for example, if activation of type II cells leads to direct activation of the gustatory nerves, or whether the information is relayed through type III cells. To determine how WGA produced in T1r3-expressing taste cells is transported into gustatory neurons, transgenic mice expressing WGA-IRES-GFP driven by the T1r3 promoter were generated. Results Immunohistochemistry showed co-expression of WGA, GFP and endogenous T1r3 in the taste bud cells of transgenic mice: the only taste cells immunoreactive for WGA were the T1r3-expressing cells. The WGA antibody also stained intragemmal nerves. WGA, but not GFP immunoreactivity was found in the geniculate and petrosal ganglia of transgenic mice, indicating that WGA was transported across synapses. WGA immunoreactivity was also found in the trigeminal ganglion, suggesting that T1r3-expressing cells make synapses with trigeminal neurons. In the medulla, WGA was detected in the nucleus of the solitary tract but also in the nucleus ambiguus, the vestibular nucleus, the trigeminal nucleus and in the gigantocellular reticular nucleus. WGA was not detected in the parabrachial nucleus, or the gustatory cortex. Conclusion These results show the usefulness of genetically encoded WGA as a tracer for the first and second order neurons that innervate a subset of taste cells, but not for higher order neurons, and demonstrate that the main route of output from type II taste cells is the gustatory neuron, not the type III cells.

  8. Immune haemolytic anaemia associated with ampicillin dependent warm antibodies and high titre cold agglutinins in a patient with Mycoplasma pneumonia

    DEFF Research Database (Denmark)

    Mickley, H; Sørensen, P G

    1984-01-01

    A case of severe immune haemolytic anaemia in a 54-year-old man suffering from Mycoplasma pneumonia is presented. A strongly positive direct Coombs test with erythrocyte bound IgG, C3d and C4 was demonstrated during the haemolytic process. Further, serologic investigations revealed ampicillin-dep...

  9. Endogenous lectins from cultured soybean cells: isolation of a protein immunologically cross-reactive with seed soybean agglutinin and analysis of its role in binding of Rhizobium japonicum

    OpenAIRE

    1986-01-01

    Incubation of Rhizobium japonicum with the cultured soybean cell line SB-1, originally derived from the roots of Glycine max, resulted in specific adhesion of the bacteria to the plant cells. This binding interaction appears to be mediated via carbohydrate recognition, since galactose can inhibit the heterotypic adhesion but glucose cannot. Affinity chromatography, on a Sepharose column derivatized with N- caproyl-galactosamine, of the supernatant fraction of a SB-1 cell suspension after enzy...

  10. Ultrastructural changes of the nodose ganglion cells following an intraneural injection of Ricinus communis agglutinin-60 into the vagus nerve in hamsters.

    OpenAIRE

    Ling, E A; Wen, C Y; Shieh, J Y; Yick, T Y; Wong, W C

    1991-01-01

    Virtually all the ganglion cells in the nodose ganglion in hamsters underwent rapid degeneration following an intraneural injection of RCA-60 into the vagus nerve in the cervical region. The earliest signs of neuronal degeneration were evident in animals which survived 5 days after the ricin application. A remarkable feature was the appearance of a variable number of granular dense bodies measuring 1-4 microns in diameter in the cytoplasm. They were composed of closely stacked cisternae which...

  11. Luffa acutangula agglutinin: Primary structure determination and identification of a tryptophan residue involved in its carbohydrate-binding activity using mass spectrometry.

    Science.gov (United States)

    Kumar, Gnanesh; Mishra, Padmanabh; Anantharam, Vellareddy; Surolia, Avadhesha

    2015-12-01

    A lectin from phloem exudates of Luffa acutangula (ridge gourd) was purified on chitin affinity chromatography and characterized for its amino acid sequence and to study the role of tryptophan in its activity. The purified lectin was subjected to various proteolytic digestions, and the resulting peptides were analyzed by liquid chromatography coupled electrospray ionization ion trap mass spectrometer. The peptide precursor ions were fragmented by collision-induced dissociation or electron transfer dissociation experiments, and a manual interpretation of MS/MS was performed to deduce amino acid sequence. This gave rise to almost complete sequence coverage of the lectin which showed high-sequence similarity with deduced sequences of phloem lectins present in the database. Chemical modification of lysine, tyrosine, histidine, arginine, aspartic acid, and glutamic acid residues did not inhibit the hemagglutinating activity. However, the modification of tryptophan residues using N-bromosuccinimide showed the loss of hemagglutinating activity. Additionally, the mapping of tryptophan residues was performed to determine the extent and number of residues modified, which revealed that six residues per molecule were oxidized suggesting their accessibility. The retention of the lectin activity was seen when the modifications were performed in the presence of chitooligosaccharides due to protection of a tryptophan residue (W102) in the protein. These studies taken together have led to the identification of a particular tryptophan residue (W102) in the activity of the lectin. PMID:26597132

  12. Cell surface display of highly pathogenic avian influenza hemagglutinin on the surface of Pichia pastoris cells using alpha-agglutinin for production of oral vaccines

    Science.gov (United States)

    Yeast are an ideal organism to express viral antigens because yeast glycosylate proteins are more similar to mammals than bacteria, and expression of proteins in yeast is relatively fast and inexpensive. In addition to the convenience of production, for purposes of vaccination, yeast have been show...

  13. Anti-leptospirose agglutinins in equine sera, from São Paulo, Goias, and Mato Grosso do Sul, Brazil, 1996-2001

    Directory of Open Access Journals (Sweden)

    H. Langoni

    2004-01-01

    Full Text Available Equine leptospirosis can present a non-symptomatic form, an acute clinical form, or even develop chronically, causing reproductive alterations, such as abortion and recurrent uveitis. Since the prevalence of leptospirosis in several countries and regions is widely reported, the objective of this study was to verify the prevailing equine leptospirosis in different regions of Brazil. Sera from 1402 blood samples from horses of different age, sex, breed, and purpose were examined. These samples came from southeastern and central west states of Brazil. The method utilized was the Microscopic Agglutination Test (MAT, with 12 different Leptospira serovars. From the sera tested, 754 (54% were positive for one (385 or more (372 serovars. These results were higher when compared to national and international levels. The most commonly found serovars were icterohaemorrhagiae (37.01%, suggesting exposure to rodents, castellonis (16.97%, and djasiman (15.19%. There were significant differences of reagents between sexes, and a tendency toward higher positivity with age. Distribution of sera-reagents related to aptitude showed a markedly higher value for work animals than for sporting ones. Higher rates were found for animals with undefined breed. There were no significant differences related to regional origin. As an indication of multiple exposure, significant associations were observed between the following serovars: castellonis and djasiman; castellonis and grippotyphosa; castellonis and copenhageni; castellonis and icterohaemorrhagiae; castellonis and pomona; canicola and pomona; canicola and djasiman; djasiman and copenhageni; icterohaemorrhagiae and djasiman; icterohaemorrhagiae and pyrogenes; copenhageni and pomona. These results showed the necessity of further studies on the epidemiology of this disease in equines and its relationship to human illness.

  14. Main: 2CWG [RPSD[Archive

    Lifescience Database Archive (English)

    Full Text Available 2CWG 小麦 Bread Wheat ... Triticum aestivum Agglutinin Isolectin 1 Precursor Triticum Aestivum Wheat ... efinement And Structure Analysis Of The Complex Of Wheat ... Germ Agglutinin With A Bivalent Sialoglycopeptide ... GGRVCTNNYCCSKWGSCGIGPGYCGAGCQSGGCDGVFAEAITANSTLLQE wheat _2CWG.jpg ...

  15. How Is Waldenstrom Macroglobulinemia Diagnosed?

    Science.gov (United States)

    ... in cool temperatures and can block blood vessels). Cold agglutinins Cold agglutinins are antibodies that attack and ... attach to cells only if they contain specific molecules. These antibodies cause color changes, which can be ...

  16. Salivary agglutinin and lung scavenger receptor cysteine-rich glycoprotein 340 have broad anti-influenza activities and interactions with surfactant protein D that vary according to donor source and sialylation

    DEFF Research Database (Denmark)

    Hartshorn, Kevan L.; Ligtenberg, Antoon; White, Mitchell R.; van Eijk, Martin; Hartshorn, Max; Pemberton, Lily; Holmskov, Uffe; Crouch, Erika

    2006-01-01

    salivary gp-340 are identical in protein sequence, salivary gp-340 from one donor had significantly greater antiviral activity against avian-like IAV strains which preferentially bind sialic acids in alpha(2,3) linkage. A greater density of alpha(2,3)-linked sialic acids was present on the salivary gp-340...... from this donor as compared with salivary gp-340 from another donor or several preparations of lung gp-340. Hence, the specificity of sialic acid linkages on gp-340 is an important determinant of anti-IAV activity. Gp-340 binds to SP-D (surfactant protein D), and we previously showed that lung gp-340...

  17. Pesquisa de aglutininas anti Brucella canis em soros humanos na cidade de São Paulo, Brasil Research on agglutinins for Brucella canis in human sera in the city of S. Paulo, Brazil

    Directory of Open Access Journals (Sweden)

    Maria Helena Matiko Akao Larsson

    1980-09-01

    Full Text Available De 330 soros humanos examinados pela prova de soroaglutinação lenta em tubos, 4(1,21% apresentaram aglutininas anti Brucella canis em diluição 1:100 (1 reagente com título 100, 2 reagentes com título 200 e 1 reagente com título 400.Of the 330 human sera tested by tube agglutination test, 4 (1.21% were positive for Brucella canis antibodies with tilers 1:100 or higher (1 reagent with titer of 1:100, 2 reagents with titer of 1:200, and 1 reagent with tiler of 1:400.

  18. 小扁豆凝集素结合型甲胎异质体在肝癌诊断中的意义%Significance of Lens culinaris agglutinin-reactive alpha-fetoprotein in the diagnosis of hepatocellular carcinoma

    Institute of Scientific and Technical Information of China (English)

    卓传尚; 柳丽娟; 吴秋芳

    2009-01-01

    目的 探讨小扁豆凝集素结合型甲胎蛋白异质体(AFP-L3)在良恶性肝病鉴别诊断的临床价值.方法 应用装有耦联了小扁豆凝集素(LCA)的微量离心柱分离185例肝病患者的AFP-L3,用时间分辨荧光免疫检测血清AFP和AFP-L3含量,计算AFP-L3%.结果 肝细胞癌患者的AFP-L3%明显高于其他肝病患者(χ2=29.329,P<0.001);AFP、AFP-L3%检测肝细胞癌在ROC曲线下的面积AUC分别为0.407和0.841;以AFP-L3%≥12.6%作为诊断标准,AFP-L3诊断肝细胞癌敏感性和特异性分别为83.3%和86.3%.结论 AFP-L3对肝细胞癌诊断准确度明显高于AFP,微量离心柱法检测AFP-L3在良恶性肝脏病变鉴别诊断中具有重要临床价值.

  19. 阴道或直肠注入白芸豆凝集素的长期毒性研究%Study on long-term toxicity test of Yunnan white kidney bean agglutinin in mice

    Institute of Scientific and Technical Information of China (English)

    赵琎; 朱轩轩; 李阔; 杨明洁; 张田; 王敏康

    2012-01-01

    目的 观察长期阴道或直肠注入白芸豆植物凝集素( WKBA)对小鼠的影响,为人用安全剂量提供参考.方法 将昆明种小鼠分成白芸豆凝集素高、中、低剂量组、壬苯醇醚(乐乐迷,Nonoxynol -9)组、辅料组、生理盐水组,每组雌雄各15只.分别采用阴道/肛门给药.1周给药5次.3个月后进行检查.结果 连续3个月阴道或直肠注入白芸豆凝集素( ≤4 mg/d),小鼠的日常生活没有异常,对小鼠的尿液没有影响,而小鼠的血液学和血液生化指标没有变化规律.结论 连续3个月阴道或直肠注入白芸豆凝集素(≤4 mg/d),小鼠的日常生活没有异常,对小鼠的尿液指标和脏器系数没有影响,而对小鼠血液的生化指标影响不明确.

  20. Correlation between carbohydrate structures on the envelope glycoprotein gp120 of HIV-1 and HIV-2 and syncytium inhibition with lectins

    DEFF Research Database (Denmark)

    Hansen, J E; Nielsen, C M; Nielsen, C;

    1989-01-01

    The binding of 13 different lectins to gp120 partially purified from two HIV-1 isolates and one HIV-2 isolate was studied by in situ staining on electrophoretically separated and electroblotted HIV antigens. The lectins concanavalin A, wheat germ agglutinin, Lens culinaris agglutinin, Vicia faba...... agglutinin, Pisum sativum agglutinin and phytohaem(erythro)agglutinin bound to gp120 of all three isolates. The carbohydrate of gp120 recognized by lectins was thus arranged in at least four types of glycans: a high mannose type glycan, a bisected hybrid or complex type glycan, a biantennary fucosylated...

  1. Glycoproteins from the cuticle of the Atlantic shore crab Carcinus maenas: I. Electrophoresis and Western-blot analysis by use of lectins

    OpenAIRE

    Compère, P.; Jaspar-Versali, M.-F.; Goffinet, G.

    2002-01-01

    The protein and glycoprotein content of four different neutral or acidic solvent extracts (0.5 M KCl, 10% EDTA, 0.1 N HCI, or 2% acetic acid) from the mineralized exoskeleton of a decapod crustacean, the Atlantic shore crab Carcinus maenas, were characterized by quantitative analysis of proteins, SDS-PAGE analysis, and probing with lectins on blots. The lectins used were Conconavalin A, Jacalin, soybean agglutinin, Maackia amurensis agglutinin II, and Sambucus nigra agglutinin. The results sh...

  2. NCBI nr-aa BLAST: CBRC-MLUC-01-0257 [SEVENS

    Lifescience Database Archive (English)

    Full Text Available Complexes pdb|2CWG|A Chain A, Crystallographic Refinement And Structure Analysis ...inement And Structure Analysis Of The Complex Of Wheat Germ Agglutinin With A Bival...Of The Complex Of Wheat Germ Agglutinin With A Bivalent Sialoglycopeptide From Glycophorin A pdb|2CWG|B Chain B, Crystallographic Ref

  3. NCBI nr-aa BLAST: CBRC-MLUC-01-0257 [SEVENS

    Lifescience Database Archive (English)

    Full Text Available CBRC-MLUC-01-0257 sp|P10969|AGI3_WHEAT ... RecName: Full=Agglutinin isolectin 3; AltName: Full=WGA3; ... Flags: Precursor gb|AAA34257.1| wheat ... germ agglutinin [Triticum turgidum subsp. durum] p ...

  4. NCBI nr-aa BLAST: CBRC-MLUC-01-0257 [SEVENS

    Lifescience Database Archive (English)

    Full Text Available CBRC-MLUC-01-0257 pdb|2UWZ|A Chain A, Crystal Structure Of Wheat ... Germ Agglutinin Isolectin 3 In ... te Ligand pdb|2UWZ|B Chain B, Crystal Structure Of Wheat ... Germ Agglutinin Isolectin 3 In Complex With A Synt ...

  5. Main: 1WGC [RPSD[Archive

    Lifescience Database Archive (English)

    Full Text Available 1WGC 小麦 Bread Wheat ... Triticum aestivum Agglutinin Isolectin 1 Precursor Triticum Aestivum Wheat ... Analysis Of Two Refined N-Acetylneuraminyllactose-*Wheat ... Germ Agglutinin Isolectin Complexes J.Mol.Biol. V. ... GGRVCTNNYCCSKWGSCGIGPGYCGAGCQSGGCDGVFAEAITANSTLLQE wheat _1WGC.jpg ...

  6. Main: 2WGC [RPSD[Archive

    Lifescience Database Archive (English)

    Full Text Available 2WGC 小麦 Bread Wheat ... Triticum aestivum Agglutinin Isolectin 2 Precursor Triticum Aestivum Wheat ... Analysis Of Two Refined N-Acetylneuraminyllactose-*Wheat ... Germ Agglutinin Isolectin Complexes J.Mol.Biol. V. ... GGRVCTNNYCCSKWGSCGIGPGYCGAGCQSGGCDAVFAGAITANSTLLAE wheat _2WGC.jpg ...

  7. Lectin staining shows no evidence of involvement of glycocalyx/mucous layer carbohydrate structures in development of celiac disease

    DEFF Research Database (Denmark)

    Toft-Hansen, Henrik; Nielsen, Christian; Biagini, Matteo;

    2013-01-01

    CD and T1D were examined before and after remission following a gluten-free diet. We performed lectin histochemistry using peanut agglutinin (PNA) and Ulex europaeus agglutinin (UEA) staining for Gal-β(1,3)-GalNAc and Fucα1-2Gal-R, respectively, of the glycocalyx/mucous layer. The staining was scored...

  8. Histological and lectin histochemical studies on the olfactory and respiratory mucosae of the sheep.

    Science.gov (United States)

    Ibrahim, Dalia; Nakamuta, Nobuaki; Taniguchi, Kazumi; Yamamoto, Yoshio; Taniguchi, Kazuyuki

    2014-03-01

    The olfactory and respiratory mucosae of the Corriedale sheep were examined using lectin histochemistry in order to clarify the histochemical and glycohistochemical differences between these two tissues. The olfactory epithelium was stained with 13 lectins out of 21 lectins examined, while the respiratory epithelium was positive to 16 lectins. The free border of both of the olfactory and respiratory epithelia was stained with 12 lectins: Wheat germ agglutinin (WGA), succinylated-wheat germ agglutinin (s-WGA), Lycopersicon esculentum lectin (LEL), Solanum tuberosum lectin (STL), Datura stramonium lectin (DSL), Soybean agglutinin (SBA), Bandeiraea simplicifolia lectin-I (BSL-I), Ricinus communis agglutinin-I (RCA-120), Erythrina cristagalli lectin (ECL), Concanavalin A (Con A), Phaseolus vulgaris agglutinin-E (PHA-E) and Phaseolus vulgaris agglutinin-L (PHA-L). The associated glands of the olfactory mucosa, Bowman's glands, were stained with 13 lectins. While both the goblet cells and mucous nasal glands were stained with 8 lectins; five of them (WGA, s-WGA, STL, Vicia villosa agglutinin (VVA) and ECL) were mutually positive among the Bowman's glands, mucous nasal glands and the goblet cells. These findings indicate that the glycohistochemical characteristics of the free borders of both olfactory and respiratory epithelia are similar to each other, suggesting that secretions from the Bowman's glands and those of the goblet cells and mucous nasal glands are partially exchanged between the surface of two epithelia to contribute the functions of the respiratory epithelium and the olfactory receptor cells, respectively. PMID:24200894

  9. NCBI nr-aa BLAST: CBRC-OCUN-01-0528 [SEVENS

    Lifescience Database Archive (English)

    Full Text Available CBRC-OCUN-01-0528 ref|XP_795774.2| PREDICTED: similar to plus agglutinin [Strongylo...centrotus purpuratus] ref|XP_001192339.1| PREDICTED: similar to plus agglutinin [Strongylocentrotus purpuratus] XP_795774.2 0.062 32% ...

  10. Main: 1WGT [RPSD[Archive

    Lifescience Database Archive (English)

    Full Text Available tivum Wheat Germ Agglutinin (Isolectin 3) Lectin (Agglutinin) K.Harata, H.Nagahora, Y.Jigami K.Harata, H.Nagahora, Y.Jigami X-Ray...0026; CHIT_BIND_I_1; 4.|PROSITE; PS50941; CHIT_BIND_I_2; 4. X-Ray Diffraction Len

  11. Cold autoimmune haemolytic anaemia secondary to Epstein Barr virus infection presenting with peripheral gangrene; case report

    Directory of Open Access Journals (Sweden)

    Karunarathne Suneth

    2012-04-01

    Full Text Available Abstract A sixty year old male presented with dark urine, symptomatic anaemia and peripheral gangrene following cold exposure. Investigations revealed that he had haemolysis and serological evidence of recent Epstein Barr virus infection. Although acrocyanosis is commonly associated with cold agglutinin disease, gangrene is a rare complication. Management of secondary cold agglutinin disease is mainly supportive.

  12. Cold autoimmune haemolytic anaemia secondary to Epstein Barr virus infection presenting with peripheral gangrene; case report

    OpenAIRE

    Karunarathne Suneth; Weerasinghe Sajitha; Govindapala Dumitha; Fernando Harshini; Jayaratne Bhaddika

    2012-01-01

    Abstract A sixty year old male presented with dark urine, symptomatic anaemia and peripheral gangrene following cold exposure. Investigations revealed that he had haemolysis and serological evidence of recent Epstein Barr virus infection. Although acrocyanosis is commonly associated with cold agglutinin disease, gangrene is a rare complication. Management of secondary cold agglutinin disease is mainly supportive.

  13. A lectin histochemical study on carbohydrate moieties of the gonadotropin-like substance in the epithelial cells of Hatschek's pit of Branchiostoma belcheri

    Science.gov (United States)

    Fang, Y. Q.; Welsch, U.

    1997-03-01

    The present light microscopic lectin, histochemical study suggests for the first time that the vertebrate gonadotropin-like substance in the basal part of the epithelial cells of Hatschek's pit is a sialic acid-containing glycoprotein. The binding intensity of the epithelial cells in Hatschek's pit to 6 lectins ( Limulus polyphemus agglutinin (LPA), Wheat germ agglutinin (WGA), Helix pomatia agglutinin (HPA), Concanavalin A (Con A), Ulex europaeus agglutinin I (UEA I) and Ricinus communis agglutinin I (RCA I)) indicate that the carbohydrate composition of the gonadotrophic glycoprotein is similar to that of mammals and fish, and that N-acetyl-D-galactosamine, sialic acid, glucosamine, D-mannose and L-fucose are components of the carbohydrate portion.

  14. Genetic labeling of both the axons of transduced, glutamatergic neurons in rat postrhinal cortex and their postsynaptic neurons in other neocortical areas by Herpes Simplex Virus vectors that coexpress an axon-targeted ß-galactosidase and wheat germ agglutinin from a vesicular glutamate transporter-1 promoter

    OpenAIRE

    Zhang, Guo-rong; Cao, Haiyan; Li, Xu; Zhao, Hua; Geller, Alfred I.

    2010-01-01

    Neuronal circuits comprise the foundation for neuronal physiology and synaptic plasticity, and thus for consequent behaviors and learning, but our knowledge of neocortical circuits is incomplete. Mapping neocortical circuits is a challenging problem because these circuits contain large numbers of neurons, a high density of synapses, and numerous classes and subclasses of neurons that form many different types of synapses. Expression of specific genetic tracers in small numbers of specific sub...

  15. Distribution of lectin-bindings in the testis of the lesser mouse deer, Tragulus javanicus.

    Science.gov (United States)

    Agungpriyono, S; Kurohmaru, M; Kimura, J; Wahid, A H; Sasaki, M; Kitamura, N; Yamada, J; Fukuta, K; Zuki, A B

    2009-06-01

    The distribution of lectin bindings in the testis of the smallest ruminant, lesser mouse deer (Tragulus javanicus), was studied using 12 biotinylated lectins specific for d-galactose (peanut agglutinin PNA, Ricinus communis agglutinin RCA I), N-acetyl-d-galactosamine (Dolichos biflorus agglutinin DBA, Vicia villosa agglutinin VVA, Soybean agglutinin SBA), N-acetyl-D-glucosamine and sialic acid (wheat germ agglutinin WGA, s-WGA), D-mannose and d-glucose (Lens culinaris agglutinin LCA, Pisum sativum agglutinin PSA, Concanavalin A Con A), L-fucose (Ulex europaeus agglutinin UEA I), and oligosaccharide (Phaseolus vulgaris agglutinin PHA-E) sugar residues. In Golgi-, cap-, and acrosome-phase spermatids, lectin-bindings were found in the acrosome (PNA, RCA I, VVA, SBA, WGA and s-WGA), and in the cytoplasm (PNA, RCA I, VVA, SBA, WGA, LCA, PSA, Con A and PHA-E). s-WGA binding was confined to the spermatid acrosome, but other lectins were also observed in spermatocytes. In spermatogonia, VVA, WGA, Con A, and PHA-E bindings were observed. Sertoli cells were intensely stained with DBA and Con A, and weakly with PHA-E. In interstitial Leydig cells, RCA I, DBA, VVA, Con A, PSA, LCA, WGA and PHA-E were positive. UEA I was negative in all cell types including spermatogenic cells. Unusual distribution of lectin-bindings noted in the testis of lesser mouse deer included the limited distribution of s-WGA only in the spermatid acrosome, the distribution of DBA in Sertoli cells, Leydig cells and lamina propria, and the absence of UEA I in all type cells. The present results were discussed in comparison with those of other animals and their possible functional implications. PMID:19245668

  16. A lectin histochemical study on the testis of the babirusa, Babyroussa babyrussa (Suidae).

    Science.gov (United States)

    Agungpriyono, S; Kurohmaru, M; Prasetyaningtyas, W E; Kaspe, L; Leus, K Y G; Sasaki, M; Kitamura, N; Yamada, J; Macdonald, A A

    2007-10-01

    The distribution of lectin bindings in the testis of babirusa, Babyrousa babyrussa (Suidae) was studied histochemically using 10 biotinylated lectins, Peanut agglutinin (PNA), Ricinus communis agglutinin (RCA I), Dolichos biflorus agglutinin (DBA), Vicia villosa agglutinin (VVA), Soybean agglutinin (SBA), Wheat germ agglutinin (WGA), Lens culinaris agglutinin (LCA), Pisum sativum agglutinin (PSA), Concanavalin A(Con A) and Ulex europaeus agglutinin (UEA I). Nine of 10 lectins showed a variety of staining patterns in the seminiferous epithelium and interstitial cells. The acrosome of Golgi-, cap- and acrosome-phase spermatids displayed various PNA, RCA I, VVA, SBA and WGA bindings, indicating the presence of glycoconjugates with D-galactose, N-acetyl-D-galactosamine and N-acetyl-D-glucosamine sugar residues respectively. No affinity was detected in the acrosome of late spermatids. LCA, PSA and Con A which have affinity for D-mannose and D-glucose sugar residues were positive in the cytoplasm of spermatids and spermatocytes. DBA was positive only in spermatogonia. In addition to DBA, positive binding in spermatogonia was found for VVA, WGA and Con A, suggesting the distribution of glycoconjugates with N-acetyl-D-galactosamine, N-acetyl-D-glucosamine, D-mannose and D-glucose sugar residues. Sertoli cells were stained intensely with RCA I, WGA and Con A. In Leydig cells, RCA I and Con A were strongly positive, while WGA, LCA and PSA reactions were weak to moderate. The present findings showed that the distribution pattern of lectin binding in the testis of babirusa is somewhat different from that of pig or other mammals reported previously. PMID:17845223

  17. Salmonella enterocolitis

    Science.gov (United States)

    ... infection if you have: Eaten foods such as turkey, turkey dressing, chicken, or eggs that have not been ... skin. Tests that may be done include: Blood culture Complete blood count with differential Febrile/cold agglutinins ( ...

  18. Cellular carbohydrate components in human, rabbit and rat lacrimal gland. Studies using fluorescein and peroxidase labelled lectins.

    Science.gov (United States)

    Ahmed, A; Grierson, I

    1989-01-01

    Orbital lacrimal glands from adult male and female rabbits, rats and humans were examined for the presence of intracellular receptors of four lectins: concanavalin-A agglutinin, lutus tetragonolobus agglutinin, ricinus comunis-60 agglutinin and wheat-germ agglutinin using fluorescein-conjugated lectin and peroxidase labelling methods for fluorescence and electron microscopy, respectively. Lectins were used as specific probes to detect carbohydrate moiety of the lacrimal gland. The pattern of labelling with the lectins suggests that N-acetyl-glucosamine, N-acetyl-D-galactosamine, D-galactose, D-mannose, sialic acid and L-fucose are contained in the lacrimal gland of the three species. The significance of these findings is discussed. PMID:2920911

  19. Use of an Intravascular Warming Catheter during Off-Pump Coronary Artery Bypass Surgery in a Patient with Severe Cold Hemagglutinin Disease.

    Science.gov (United States)

    Tholpady, Ashok; Bracey, Arthur W; Baker, Kelty R; Reul, Ross M; Chen, Alice J

    2016-08-01

    Cold hemagglutinin disease with broad thermal amplitude and high titers presents challenges in treating cardiac-surgery patients. Careful planning is needed to prevent the activation of cold agglutinins and the agglutination of red blood cells as the patient's temperature drops during surgery. We describe our approach to mitigating cold agglutinin formation in a 77-year-old man with severe cold hemagglutinin disease who underwent off-pump coronary artery bypass surgery without the use of preoperative plasmapheresis. This experience shows that the use of an intravascular warming catheter can maintain normothermia and prevent the activation and subsequent formation of cold agglutinins. To our knowledge, this is the first reported use of this technique in a patient with cold hemagglutinin disease. The chief feature in this approach is the use of optimal thermal maintenance-rather than the more usual decrease in cold-agglutinin content by means of therapeutic plasma exchange. PMID:27547154

  20. Evaluation of glycophenotype in prostatic neoplasm by chemiluminescent assay

    OpenAIRE

    da Silva, Lúcia Patrícia Bezerra Gomes; Almeida, Sinara Mônica Vitalino; Lima, Luiza Rayanna Amorim; Cavalcanti, Carmelita de Lima Bezerra; Lira, Mariana Montenegro de Melo; da Silva, Maria da Paz Carvalho; Beltrão, Eduardo Isidoro Carneiro; Júnior, Luiz Bezerra de Carvalho

    2014-01-01

    This work aimed to evaluate the glycophenotype in normal prostate, bening prostatic hyperplasia (BPH) and prostatic adenocarcinoma (PCa) tissues by a chemiluminescent method. Concanavalin A (Con A), Ulex europaeus agglutinin (UEA-I) and Peanut agglutinin (PNA) lectins were conjugated to acridinium ester (lectins-AE). These conjugates remained capable to recognize their specific carbohydrates. Tissue samples were incubated with lectins-AE. The chemiluminescence of the tissue-lectin-AE complex ...

  1. Main: 1K7T [RPSD[Archive

    Lifescience Database Archive (English)

    Full Text Available 1K7T 小麦 Bread Wheat ... Triticum aestivum Agglutinin Isolectin 3 Precursor Triticum Aestivum Molec ... ata M.Muraki, M.Ishimura, K.Harata Interactions Of Wheat -Germ Agglutinin With Glcnacbeta1, 6gal Sequence Bi ... phys.Acta V.1569 10 2002 Hevein-Type Fold SWS:AGI3_WHEAT ,P10969|EMBL; J02961; AAA34257.1; -.|PIR; A28401; A ...

  2. Main: 1K7V [RPSD[Archive

    Lifescience Database Archive (English)

    Full Text Available 1K7V 小麦 Bread Wheat ... Triticum aestivum Agglutinin Isolectin 3 Precursor Triticum Aestivum Molec ... ata M.Muraki, M.Ishimura, K.Harata Interactions Of Wheat -Germ Agglutinin With Glcnacbeta1, 6gal Sequence Bi ... phys.Acta V.1569 10 2002 Hevein-Type Fold SWS:AGI3_WHEAT ,P10969|EMBL; J02961; AAA34257.1; -.|PIR; A28401; A ...

  3. Main: 1K7U [RPSD[Archive

    Lifescience Database Archive (English)

    Full Text Available 1K7U 小麦 Bread Wheat ... Triticum aestivum Agglutinin Isolectin 3 Precursor Triticum Aestivum Molec ... ata M.Muraki, M.Ishimura, K.Harata Interactions Of Wheat -Germ Agglutinin With Glcnacbeta1, 6gal Sequence Bi ... phys.Acta V.1569 10 2002 Hevein-Type Fold SWS:AGI3_WHEAT ,P10969|EMBL; J02961; AAA34257.1; -.|PIR; A28401; A ...

  4. Lectins with Anti-HIV Activity: A Review

    OpenAIRE

    Ouafae Akkouh; Tzi Bun Ng; Senjam Sunil Singh; Cuiming Yin; Xiuli Dan; Yau Sang Chan; Wenliang Pan; Randy Chi Fai Cheung

    2015-01-01

    Lectins including flowering plant lectins, algal lectins, cyanobacterial lectins, actinomycete lectin, worm lectins, and the nonpeptidic lectin mimics pradimicins and benanomicins, exhibit anti-HIV activity. The anti-HIV plant lectins include Artocarpus heterophyllus (jacalin) lectin, concanavalin A, Galanthus nivalis (snowdrop) agglutinin-related lectins, Musa acuminata (banana) lectin, Myrianthus holstii lectin, Narcissus pseudonarcissus lectin, and Urtica diocia agglutinin. The anti-HIV al...

  5. Lectin histochemistry of palatine glands in the developing rat.

    Science.gov (United States)

    Hakami, Zaki; Kitaura, Hideki; Honma, Shiho; Wakisaka, Satoshi; Takano-Yamamoto, Teruko

    2014-05-01

    This study examined the binding pattern of lectins, soybean agglutinin (SBA), Dolichos biflorus agglutinin (DBA), Vicia villosa agglutinin (VVA), Ulex europaeus agglutinin-I (UEA-I), peanut agglutinin (PNA), wheat germ agglutinin (WGA), and succinylated WGA (sucWGA) in the developing rat palatine glands. In adult rats, heterogeneous lectin binding patterns were revealed between the anterior and posterior portions of palatine glands, as DBA, VVA, and WGA were bound more intensely and broadly in the posterior portion. SBA, PNA, and sucWGA showed far less reactivity in the anterior than in the posterior portion. At embryonic day 18 (E18), weak labeling was observed with UEA-I and WGA at the basal membrane of terminal buds, UEA-I and PNA labeled the epithelial cord, and there was no apparent binding for SBA, DBA, VVA, and sucWGA. At E20, after acinar lumenization, all lectins were detected at the acinar cell basal membranes. After birth, all lectins detectably labeled at the mucous cell apical membranes and progressively, with maturation, extended from the apical to basal portions of the cytoplasm. Apparent serous cells were observed around postnatal day 10 (PN10) and bound UEA-I. Lectins reached peak reactivity at PN21 and the binding patterns became identical to those of adults around PN28. PMID:24345684

  6. Diversidade antigênica entre amostras de Arcobacter spp isoladas de suínos no Rio Grande do Sul e presença de anticorpos aglutinantes em amostras de soro de porcas com problemas reprodutivos Antigenic diversity among strains of Arcobacter spp isolated from pigs in Rio Grande do Sul, Brazil and presence of agglutinin titers in serum samples of sows with reproductive problems

    OpenAIRE

    Sérgio José de Oliveira; David Emilio Santos Neves de Barcellos; Sandra Maria Borowski

    1999-01-01

    O teste de aglutinação microscópica, usando a técnica descrita para o diagnóstico de leptospirose, foi utilizado para verificar a antigenicidade de 47 amostras de Arcobacter cryaerophilus e duas amostras de Arcobacter butzleri isoladas de suínos no Rio Grande do Sul, Brasil, em frente a soros hiperimunes produzidos em coelhos a partir de amostras padrões das bactérias. Verificou-se grande heterogeneidade antigênica e apenas quatro amostras provocaram títulos acima de 1.600 com os anti-soros p...

  7. Flow cytometric analysis of lectin binding to in vitro-cultured Perkinsus marinus surface carbohydrates

    Science.gov (United States)

    Gauthier, J.D.; Jenkins, J.A.; La Peyre, Jerome F.

    2004-01-01

    Parasite surface glycoconjugates are frequently involved in cellular recognition and colonization of the host. This study reports on the identification of Perkinsus marinus surface carbohydrates by flow cytometric analyses of fluorescein isothiocyanate-conjugated lectin binding. Lectin-binding specificity was confirmed by sugar inhibition and Kolmogorov-Smirnov statistics. Clear, measurable fluorescence peaks were discriminated, and no parasite autofluorescence was observed. Parasites (GTLA-5 and Perkinsus-1 strains) harvested during log and stationary phases of growth in a protein-free medium reacted strongly with concanavalin A and wheat germ agglutinin, which bind to glucose-mannose and N-acetyl-D-glucosamine (GlcNAc) moieties, respectively. Both P. marinus strains bound with lower intensity to Maclura pomifera agglutinin, Bauhinia purpurea agglutinin, soybean agglutinin (N-acetyl-D-galactosamine-specific lectins), peanut agglutinin (PNA) (terminal galactose specific), and Griffonia simplicifolia II (GlcNAc specific). Only background fluorescence levels were detected with Ulex europaeus agglutinin I (L-fucose specific) and Limulus polyphemus agglutinin (sialic acid specific). The lectin-binding profiles were similar for the 2 strains except for a greater relative binding intensity of PNA for Perkinsus-1 and an overall greater lectin-binding capacity of Perkinsus-1 compared with GTLA-5. Growth stage comparisons revealed increased lectin-binding intensities during stationary phase compared with log phase of growth. This is the first report of the identification of surface glycoconjugates on a Perkinsus spp. by flow cytometry and the first to demonstrate that differential surface sugar expression is growth phase and strain dependent. ?? American Society of Parasitologists 2004.

  8. Different glycosylation of cadherins from human bladder non-malignant and cancer cell lines

    Directory of Open Access Journals (Sweden)

    Lityńska Anna

    2002-06-01

    Full Text Available Abstract Background The aim of the present study was to determine whether stage of invasiveness of bladder cancer cell lines contributes to alterations in glycan pattern of their cadherins. Results Human non-malignant epithelial cell of ureter HCV29, v-raf transfected HCV29 line (BC3726 and transitional cell cancers of urine bladder Hu456 and T24 were grown in cell culture. Equal amounts of protein from each cell extracts were separated by SDS-PAGE electrophoresis and were blotted on an Immobilon P membrane. Cadherins were immunodetected using anti-pan cadherin mAb and lectin blotting assays were performed, in parallel. N-oligosaccharides were analysed by specific reaction with Galanthus nivalis agglutinin (GNA, Sambucus nigra agglutinin (SNA, Maackia amurensis agglutinin (MAA, Datura stramonium agglutinin (DSA, Aleuria aurantia agglutinin (AAA, Phaseolus vulgaris agglutinin (PHA-L and wheat germ agglutinin (WGA. The cadherin from HCV29 cell line possessed bi- and/or 2,4-branched triantennary complex type glycans, some of which were α2,6-sialylated. The cadherin from BC3726 cell line exhibited exclusively high mannose type glycans. Cadherins from Hu456 and T24 cell lines expressed high mannose type glycans as well as β1,6-branched oligosaccharides with poly-N-acetyllactosamine structures and α2,3-linked sialic acid residues. Additionally, the presence of fucose and α2,6-sialic acid residues on the cadherin from T24 cell line was detected. Conclusions These results indicate that N-glycosylation pattern of cadherin from bladder cancer cell line undergoes modification during carcinogenesis.

  9. The typing of Staphylococcus epidermidis by a lectin-binding assay

    DEFF Research Database (Denmark)

    Jarløv, J O; Hansen, J E; Rosdahl, V T;

    1992-01-01

    A new typing method for Staphylococcus epidermidis was developed. Four biotinylated lectins--wheat germ agglutinin (WGA), soy bean agglutinin (SBA), lentil agglutinin (LCA) and Concanavalin A (ConA)--were added to immobilised whole cells of coagulase-negative staphylococci (CNS) in microtitration...... plates. The amount of bound lectin was measured by peroxidase-conjugated avidin followed by a peroxidase reaction. The method was compared to antibiotic-resistance analysis, phage typing, plasmid DNA profiles and slime production. A total of 113 isolates of CNS from 21 patients was investigated and 71...... strains of CNS, including 64 strains of S. epidermidis, were detected if all typing methods were taken into consideration. If only one typing method was used the highest discriminatory power among the S. epidermidis isolates was obtained with the lectin-binding assay which allowed 49 different strains to...

  10. Serine versus threonine glycosylation with α-O-GalNAc: unexpected selectivity in their molecular recognition with lectins.

    Science.gov (United States)

    Madariaga, David; Martínez-Sáez, Nuria; Somovilla, Víctor J; García-García, Laura; Berbis, M Álvaro; Valero-Gónzalez, Jessika; Martín-Santamaría, Sonsoles; Hurtado-Guerrero, Ramon; Asensio, Juan L; Jiménez-Barbero, Jesús; Avenoza, Alberto; Busto, Jesús H; Corzana, Francisco; Peregrina, Jesús M

    2014-09-22

    The molecular recognition of several glycopeptides bearing Tn antigen (α-O-GalNAc-Ser or α-O-GalNAc-Thr) in their structure by three lectins with affinity for this determinant has been analysed. The work yields remarkable results in terms of epitope recognition, showing that the underlying amino acid of Tn (serine or threonine) plays a key role in the molecular recognition. In fact, while Soybean agglutinin and Vicia villosa agglutinin lectins prefer Tn-threonine, Helix pomatia agglutinin shows a higher affinity for the glycopeptides carrying Tn-serine. The different conformational behaviour of the two Tn biological entities, the residues of the studied glycopeptides in the close proximity to the Tn antigen and the topology of the binding site of the lectins are at the origin of these differences. PMID:25111627

  11. [Studies on the location of eight lectins in breast carcinoma].

    Science.gov (United States)

    Zheng, Z; Ji, Z M

    1990-12-01

    100 cases of breast carcinoma were studied with lectin affinitive histochemistry technology. The result showed that Ricinus comunis agglutinin (RCA1) was located in almost all intraductal carcinomas but one, while the positive rates in the other types were obviously low (P less than 0.05). The positive rate of Ulex europaeus agglutinin-I (UEA1) in well-differentiated types was higher than that in poorly-differentiated ones (P less than 0.05). The location of Peanut agglutinin (PNA), Bandeiraea Simplicifolia (BSL) and UEA1 in breast carcinomas exhibited some regularity and it might be useful in understanding the differentiation of breast carcinomas. No relationship between changes of the eight lectins and metastases in axillary lymph nodes was observed, but the authors considered that PNA-affinitive histochemistry was beneficial to the detection of micrometastases in lymph nodes. PMID:1964401

  12. Characterization of colonic cellular glycoconjugates in colitis and cancer-prone tamarins versus colitis and cancer-resistant primates.

    Science.gov (United States)

    Moore, R; King, N; Alroy, J

    1988-06-01

    Differences in colonic secretory glycoconjugates (ie, mucin) between normal and ulcerative colitis-prone patients have been noted. Similar differences may occur in a corresponding primate model, the cotton-top tamarin (CTT), Saguinus oedipus, a New World monkey which suffers from spontaneous chronic colitis and colon cancer. Lectin reagents were used to characterize and compare colonic cell surface, cytoplasmic, and secretory glycoconjugates of 9 clinically healthy cotton-top tamarins, 7 colitis-susceptible, cancer-resistant tamarins (Callithrix jacchus, Saguinus fuscicollis), and 8 colitis and cancer-resistant primates (Aotus trivirgatus, Saimiri sciureus, Macaca fascicularis, and Macaca mulatta). Paraffin-embedded colonic sections were labeled with ten different biotinylated lectins and visualized by the avidin-biotin peroxidase (ABC) method. Significant differences were demonstrated in the pattern of lectin staining between the colitis-resistant and colitis-prone groups of primates. The differences were noted with Griffonia simplicifolia-I (GS-I), Dolichos biflorus agglutinin (DBA), peanut agglutinin (PNA) before and after neuraminidase, Ricinus communis agglutinin-I (RCA-I), soybean agglutinin (SBA), Ulex europaeus agglutinin-I (UEA-I), wheat germ agglutinin (WGA), and succinylated WGA (S-WGA). Significant differences between the CTT and phylogenetically related colitis-prone but cancer-resistant tamarins were demonstrated with SBA, UEA-I, and PNA after desialylation with neuraminidase. These results suggest that differences in colonic cellular glycoconjugates between colitis- and cancer-susceptible species versus colitis-susceptible, cancer-resistant species may be associated with risk of cancer. PMID:3132857

  13. Morphometric Measurements to Quantify the Cerulein Induced Hyperstimulatory Pancreatitis of Rats under the Protective Effect of Lectins

    OpenAIRE

    Jonas, Ludwig; Mikkat, Ulrike; Witte, Anke; Beckmann, Uta; Dölker, Katrin; Weber, Heike; Hahnel, Christian; Kundt, Günther; Nizze, Horst

    1998-01-01

    In preceding papers we demonstrated an inhibitory effect of wheat germ agglutinin (WGA) and Ulex europaeus agglutinin (UEA) on the cholecystokinin (CCK) binding to the CCK receptor of rat pancreatic cells and also on the CCK induced Ca2+ release and α-amylase secretion in vitro as well as on pancreatic secretion of intact rats in vivo. In the present study we show the same inhibitory effect of both lectins on the cerulein pancreatitis of rats. This acute pancreatitis was induced by supramaxim...

  14. Anopheles gambiae heat shock protein cognate 70B impedes o'nyong-nyong virus replication

    OpenAIRE

    Higgs Stephen; Vanlandingham Dana L; Tsetsarkin Konstantin A; Hong Young S; Sim Cheolho; Collins Frank H

    2007-01-01

    Abstract Background Phylogenetic and functional analysis was conducted on an Anopheles gambiae gene, ENSANGG00000017398. Based on phylogenetic analysis, this gene belongs to the same lineage as Heat shock protein cognate 70-4 (Hsc70-4) in Drosophila. Accordingly, we propose to name this gene Heat shock protein cognate 70B (HSC70B). We previously reported that expression of HSC70B and other genes including elongation factor-1α (EF-1α) and the agglutinin attachment subunit (agglutinin) were up-...

  15. Characterisation of N-glycans bound to IGFBP-3 in sera from healthy adults.

    Science.gov (United States)

    Masnikosa, Romana; Baricević, Ivona; Lagundzin, Dragana; Nedić, Olgica

    2010-01-01

    Human IGFBP-3 contains three potential N-linked glycosylation sites. Published data concerning the type and saccharide composition of the N-glycans is scarce. The aim of this study was to characterise N-glycans covalently attached to IGFBP-3 from sera of healthy adults (men and women). In order to do that a panel of eight lectins covering broad saccharide specificity was used: agarose-immobilised SNA (Sambucus nigra agglutinin), Con A (lectin from Canavalia ensiformis), RCA I (Ricinus communis agglutinin I), PHA-E (Phaseolus vulgaris erythroagglutinin), PHA-L (P. vulgaris leukoagglutinin), succinylated WGA (wheat germ agglutinin), ECL (Erythrina cristagalli lectin) and UEA (Ulex europaeus agglutinin). IGFBP-3 interacted with SNA, Con A, RCA I, PHA-E and, to a much lesser extent, with PHA-L. These results indicate that human IGFBP-3 bears mostly biantennary complex type N-glycans with a very high content of alpha-2,6-linked Sia at their termini. Hybrid type and high-mannose type N-glycans are present, as well as a bisecting GlcNAc residue, which may be core fucosylated. N-glycosylation of IGFBP-3 follows the N-glycosylation pattern of major serum proteins. This study represents a ground for the future research of glycosylation pattern of IGFBP-3 from the circulation of men and women diagnosed with different illnesses. PMID:19800385

  16. Deep trophoblast invasion and spiral artery remodelling in the placental bed of the lowland gorilla

    DEFF Research Database (Denmark)

    Pijnenborg, R; Vercruysse, L; Carter, Anthony Michael

    2011-01-01

    chimpanzee, we postulated the occurrence of deep invasion in gorilla pregnancy. Tissues were processed for histology (PAS, orcein), lectin staining (Ulex europaeus agglutinin 1) and immunohistochemistry (cytokeratin 7/17, α-actin). A specimen of young but undetermined gestational age included deep placental...

  17. Deep trophoblast invasion and spiral artery remodelling in the placental bed of the chimpanzee

    DEFF Research Database (Denmark)

    Pijnenborg, R; Vercruysse, L; Carter, Anthony Michael

    2011-01-01

    -actin, as well as Ulex europaeus agglutinin 1 (UEA1) lectin staining, in this archival material. In both specimens interstitial trophoblast invasion had occurred in both decidua and myometrium. Because of a lack of published data on fetal growth for this species, fetal sizes (7cm and 13cm) could not be...

  18. Haemangioblastoma, Histological and immunohistological study of an enigmatic cerebellar tumour

    OpenAIRE

    Cruz-Sánchez, F. F.; Rossi, M L; Rodríguez-Prados, S.; Nakamura, N; Hughes, J T; Coakham, H. B.

    1990-01-01

    Paraffin-embedded blocks of 36 cerebellar haemangioblastomas were reacted with a panel of antibodies including glial fibrillary acidic protein, vimentin, epithelial membrane antigen, cytokeratin, Factor VIII, a neuroendocrine marker and with Ulex europaeus. agglutinin The main histological features, apart from the characteristic large abnormal vessels, were a prominent reticulin network, a cystic architecture and cellular and nuclear polymorphism. Two cell type...

  19. A comparative analysis of the attachment of Leptospira interrogans and L. borgpetersenii to mammalian cells.

    Science.gov (United States)

    Andrade, Gabrielle I; Brown, Paul D

    2012-06-01

    Leptospirosis, the world's most ubiquitous zoonosis, is caused by pathogenic Leptospira. As microbe-host interactions are specific in pathogenesis, it is likely that there are several molecules mediating the attachment of the Leptospira to mammalian cells. In this study, we analysed the attachment of Leptospira interrogans serovar Portlandvere and Leptospira borgpetersenii serovar Jules to untreated HEp-2 cells or HEp-2 cells treated with the various enzymes, lectins or sugars and to integrins αVβ3 and α5β1, relative to control wells. We found that both serovars bound equally well to HEp-2 cells; however, serovar Jules showed a higher level of attachment to integrins. Both serovars showed an increase in attachment to HEp-2 cells coated with lectins peanut agglutinin, Ulex europaeus agglutinin, soybean agglutinin and Erythrina cristagalli agglutinin (p < 0.05); in the case of Concanavalin A, Jules showed an increase, while Portlandvere showed a significant decrease in attachment. Trypsinizing monolayers resulted in a decrease in attachment for both serovars, while when chondroitinase, neuraminidase and heparinase were used an increase in attachment was recorded. Leptospires coated with sugars showed a decrease in attachment. These results show that serovar Jules' general greater affinity for the mediators examined may suggest a greater potential for virulence over serovar Portlandvere. PMID:22409511

  20. The diagnosis of unilateral testicular obstruction in subfertile males.

    Science.gov (United States)

    Hendry, W F; Parslow, J M; Stedronska, J; Wallace, D M

    1982-12-01

    Thirty-two subfertile males with sperms in the ejaculate and unilateral testicular obstruction are reported: the diagnosis was established by exploration of scrotum in 26, clinically in 2, 3 had had previous partially successful epididymovasostomies, and 1 had had an epispadias repair. The past medical history gave relevant information in 27 (84%), and useful findings were made on clinical examination in a further 3 cases. Fifteen patients had sperm counts over 20 million per ml, and 15 were less than 10 million per ml. Twenty-six (81%) had serum antisperm antibodies detected by tray agglutination test (TAT), 21 (81%) of whom had evidence of head-to-head (HH) agglutinins in pure or mixed form. Comparison with 162 vasectomised males and 160 naturally infertile males with antisperm antibodies showed that 55% of the former and 24% of the latter had HH agglutinins on TAT, differences that were highly significant on statistical analysis. Evidence of obstruction was found in 14 (37%) of 38 naturally infertile males with antisperm antibodies and HH or mixed agglutination, but only in 12 (10%) of 122 with TT agglutinins: this difference was also highly significant. Clinical history, physical examination and serum antisperm antibodies, especially if HH agglutinins are present, can suggest the possibility of unilateral testicular obstruction, but confirmation of the diagnosis requires exploration of scrotum. PMID:7150940

  1. Early events of secretory granule formation in the rat parotid acinar cell under the influence of isoproterenol. An ultrastructural and lectin cytochemical study

    Directory of Open Access Journals (Sweden)

    F D’Amico

    2009-12-01

    Full Text Available The events involved in the maturation process of acinar secretory granules of rat parotid gland were investigated ultrastructurally and cytochemically by using a battery of four lectins [Triticum vulgaris agglutinin (WGA, Ulex europaeus agglutinin I (UEA-I, Glycine max agglutinin (SBA, Arachys hypogaea agglutinin (PNA]. In order to facilitate the study, parotid glands were chronically stimulated with isoproterenol to induce secretion. Specimens were embedded in the Lowicryl K4M resin. The trans-Golgi network (TGN derived secretory granules, which we refer to as immature secretory granules, were found to be intermediate structures in the biogenesis process of the secretory granules in the rat parotid acinar cell. These early structures do not seem to be the immediate precursor of the mature secretory granules: in fact, a subsequent interaction process between these early immature granule forms and TGN elements seems to occur, leading, finally, to the mature granules. These findings could explain the origin of the polymorphic subpopulations of the secretory granules in the normal acinar cells of the rat parotid gland. The lectin staining patterns were characteristic of each lectin. Immature and mature secretory gran- ules were labelled with WGA, SBA, PNA, and lightly with UEA-I. Cis and intermediate cisternae of the Golgi apparatus were labelled with WGA, and trans cisternae with WGA and SBA.

  2. Histological and lectin histochemical studies on the main and accessory olfactory bulbs in the Japanese striped snake, Elaphe quadrivirgata.

    Science.gov (United States)

    Kondoh, Daisuke; Wada, Akimi; Endo, Daisuke; Nakamuta, Nobuaki; Taniguchi, Kazuyuki

    2013-01-01

    The main and accessory olfactory bulbs were examined by histological methods and lectin histochemistry in the Japanese striped snake. As the results, the histological properties are similar between the main olfactory bulb and the accessory olfactory bulb. In lectin histochemistry, 21 lectins used in this study showed similar binding patterns in the main olfactory bulb and the accessory olfactory bulb. In detail, 15 lectins stained these olfactory bulbs with similar manner, and 6 lectins did not stain them at all. Two lectins, Lycopersicon esculentum lectin (LEL) and Solanum tuberosum lectin (STL), stained the nerve and glomerular layers and did not stain any other layers in both olfactory bulbs. Four lectins, Soybean agglutinin (SBA), Vicia villosa agglutinin (VVA), Peanut agglutinin (PNA) and Phaseolus vulgaris agglutinin-L (PHA-L) stained the nerve and glomerular layers more intensely than other layers in both olfactory bulbs. In addition, VVA showed the dot-like stainings in the glomeruli of both olfactory bulbs. These findings suggest that the degree of development and the properties of glycoconjugates are similar between the main olfactory bulb and the accessory olfactory bulb in the Japanese striped snake. PMID:23257605

  3. Ultrastructural and histochemical properties of the olfactory system in the japanese jungle crow, Corvus macrorhynchos.

    Science.gov (United States)

    Kondoh, Daisuke; Nashimoto, Mai; Kanayama, Shunsaku; Nakamuta, Nobuaki; Taniguchi, Kazuyuki

    2011-08-01

    Although it has been commonly believed that birds are more dependent on the vision and audition than the olfaction, recent studies indicate that the olfaction of birds is related to the reproductive, homing, and predatory behaviors. In an attempt to reveal the dependence on the olfactory system in crows, we examined the olfactory system of the Japanese jungle crow (Corvus macrorhynchos) by histological, ultrastructural, and lectin histochemical methods. The olfactory epithelium (OE) of the crow occupied remarkably a small area of the nasal cavity (NC) and had the histological and ultrastructural features like other birds. The olfactory bulb (OB) of the crow was remarkably small and did not possess the olfactory ventricle. The left and right halves of the OB were fused in many cases. In the lectin histochemistry, soybean agglutinin (SBA) and Vicia villosa agglutinin (VVA) stained a small number of the receptor cells (RCs) in the OE and the olfactory nerve layer (ONL) and glomerular layer (GL) on the dorsocaudal region of the OB. Phaseolus vulgaris agglutinin-E (PHA-E) stained several RCs in the OE and the ONL and GL on the ventral region of the OB. These results suggest that 1) the crow has less-developed olfactory system than other birds, and 2) the dedicated olfactory receptor cells project their axons to the specific regions of the OB in the crow. PMID:21478653

  4. Primary breast cancer tumours contain high amounts of IgA1 immunoglobulin

    DEFF Research Database (Denmark)

    Welinder, Charlotte; Baldetorp, Bo; Blixt, Klas Ola;

    2013-01-01

    The Tn antigen (GalNAc alpha-O-Ser/Thr) as defined by the binding of the lectin, helix pomatia agglutinin (HPA) or anti-Tn monoclonal antibodies, is known to be exposed in a majority of cancers, and it has also been shown to correlate positively with the metastatic capacity in breast carcinoma. T...

  5. Quantitative approach to lectin-based glycoprofiling of thymic tissues in the control- and the dexamethasone-treated mice.

    Science.gov (United States)

    Balcan, Erdal

    2016-06-01

    Dexamethasone (DEX) is the most commonly used synthetic glucocorticoid in treatment of various inflammatory conditions. Here we focused on evaluating the effect of DEX on apoptosis and glycan profile in the mouse thymic tissues. Histological examinations revealed that the DEX treatment cause severe alterations in thymus, such as disruption of thymic capsule, impaired epithelial cell-thymocyte contacts, cellular loss and increased apoptosis. The identification of thymic glycans in the control- and the DEX-treated mice was carried out by using a panel of five plant lectins, Maackia amurensis agglutinin (MAA), peanut agglutinin (PNA), Sambucus nigra agglutinin (SNA), Concanavalin A (ConA) and wheat germ agglutinin (WGA). Lectin histochemistry results showed that glycosylation pattern of thymus changes upon DEX treatment. For further detailed quantitative analyses of the binding intensities for each lectin, histochemical data were scored as high positive (HP), mild positive (MP) and low positive (LP) and differences among signaling densities were investigated. The staining patterns of thymic regions observed with lectin histochemistry suggest that DEX can affect the thymic glycan profile as well as thymocyte apoptosis. These results are consistent with the opinion that not only sialic acid, but also other sugar motifs may be responsible for thymocyte development. PMID:27067421

  6. Gastrointestinal nematodes and anthelmintic resistance in Danish goat herds

    DEFF Research Database (Denmark)

    Holm, Signe A.; Sørensen, Camilla; Thamsborg, Stig M.;

    2014-01-01

    stained with peanut agglutinin (PNA) for specific detection of Haemonchus contortus. Strongyle eggs were detected with an individual prevalence of 69%, including Nematodirus battus (3.6%) and other Nematodirus species (15.0%). Eimeria spp. were observed in 99.6% of the kids. H. contortus was found in 11...

  7. Partial Purification of the 5-Hydroxytryptamine-Reuptake System from Human Blood Platelets Using a Citalopram-Derived Affinity Resin

    NARCIS (Netherlands)

    Biessen, E.A.L.; Horn, A.S.; Robillard, G.T.

    1990-01-01

    This paper describes a procedure for the synthesis and application of a citalopram-derived affinity resin in purifying the 5HT-reuptake system from human blood platelets. A two-step scheme has been developed for partial purification, based on wheat germ agglutinin-lectin (WGA) affinity and citalopra

  8. NCBI nr-aa BLAST: CBRC-MDOM-02-0035 [SEVENS

    Lifescience Database Archive (English)

    Full Text Available protein, putative; cell-surface glycoprotein, putative [Candida dubliniensis CD36] emb|CAX43113.1| ALS fami...ly protein, putative; agglutinin-like protein, putative; cell-surface glycoprotein, putative [Candida dubliniensis CD36] XP_002419518.1 5e-46 22% ...

  9. A morphological study of the vomeronasal organ and the accessory olfactory bulb in the Korean roe deer, Capreolus pygargus.

    Science.gov (United States)

    Park, Changnam; Ahn, Meejung; Lee, Jae-Yuk; Lee, Sang; Yun, Youngmin; Lim, Yoon-Kyu; Taniguchi, Kazumi; Shin, Taekyun

    2014-01-01

    The vomeronasal organ (VNO) and accessory olfactory bulb (AOB) of the Korean roe deer (Capreolus pygargus) were studied histologically to evaluate their morphological characteristics. Grossly, the VNO, encased by cartilage, has a paired tubular structure with a caudal blind end and a rostral connection through incisive ducts on the hard palate. In the VNO, the vomeronasal sensory epithelium (VSE) consists of galectin-3-positive supporting cells, protein gene product (PGP) 9.5-positive receptor cells, and basal cells. The vomeronasal respiratory epithelium (VRE) consists of a pseudostratified epithelium. The AOB strata included a vomeronasal nerve layer (VNL), a glomerular layer (GL), a mitral/tufted cell layer, and a granular cell layer. All lectins used in this study, including Bandeiraea simplicifolia agglutinin isolectin B4 (BSI-B4), soybean agglutinin (SBA), Ulex europaeus agglutinin I (UEA-I), and Triticum vulgaris wheat germ agglutinin (WGA), labeled the VSE with varying intensity. In the AOB, both the VNL and the GL reacted with BSI-B4, SBA, and WGA with varying intensity, but not with UEA-I. This is the first morphological study of the VNO and AOB of the Korean roe deer, which are similar to those of goats. PMID:24055195

  10. Bacterial, fungal, and algal lectins: combatants in tug of war against HIV.

    Science.gov (United States)

    Feizi, Ten; Liu, Yan; Palma, Angelina S

    2011-08-10

    High-resolution X-ray crystallography and NMR studies by Koharudin and Gronenborn in this issue provide new information on the mode of N-glycan recognition by a cyanobacterial agglutinin, with anti-HIV activity pointing to the pentamannosyl core as a novel target for therapeutic intervention. PMID:21827940

  11. Main: 2SBA [RPSD[Archive

    Lifescience Database Archive (English)

    Full Text Available A.Dessen, D.Gupta, S.Sabesan, C.F.Brewer, J.C.Sacchettini X-Ray Crystal Structure Of The Soybean Agglutinin ...; PS00307; LECTIN_LEGUME_BETA; 1. X-Ray Diffraction, Resolution: 2.60 Angstrom, R-Factor:: 0.208, R-Free:: N

  12. Comparison of the binding properties of the mushroom Marasmius oreades lectin and Griffonia simplicifolia I-B isolectin to alphagalactosyl carbohydrate antigens in the surface phase

    DEFF Research Database (Denmark)

    Kirkeby, Svend; Winter, Harry C; Goldstein, Irwin J

    2004-01-01

    The binding of two alpha-galactophilic lectins, Marasmius oreades agglutinin (MOA), and Griffonia simplicifolia I isolectin B(4) (GS I-B(4)) to neoglycoproteins and natural glycoproteins were compared in a surface phase assay. Neoglycoproteins carrying various alpha-galactosylated glycans and lam...

  13. Residu Gula Glikokonjugat pada Lambung Depan Kerbau Rawa (Bubalus bubalis Kalimantan Selatan (SUGAR RESIDU OF GLYCOCONJUGATES IN FORESTOMACH OF SOUTH KALIMANTAN SWAMP BUFFALO (BUBALUS BUBALIS

    Directory of Open Access Journals (Sweden)

    Anni Nurliani

    2014-08-01

    Full Text Available The ability of swamp buffaloes to adapt with swamp environment was suggested to be supported bytheir digestive system efficiency. The research was done to obtain scientific explanation about digestiveefficiency of swamp buffalo by identification on kinds and distribution of glycoconjugates in swamp buffaloforestomach. Six male swamp buffaloes aged more than 2.5 year old and had body weight between 300-400kg were used in this study. Samples were obtained from Regency of Banjar slaughter house, SouthKalimantan. Every parts of the forestomach included rumen, reticulum, and omasum was taken andprocessed for microscopic observation with hematoxyline eosin (HE and alcian blue-periodic acid schiff(AB-PAS stainings. Sugar residues of glycoconjugates were localized with lectin histochemistry wheatgerm agglutinin (WGA, ulex europaeus agglutinin (UEA, ricinus communis agglutinin (RCA, concanavalinagglutinin (Con A, and soybean agglutinin (SBA. Every part of swamp buffalo forestomach had kinds ofspecific glycoconjugates with special distribution pattern which were different with other ruminant, andwere suitable for their functions in that part. The existence of D mannose/D glucose glycoconjugates thatwas dominant in forestomach estimated that had important role in supporting fermentative digestionfunction in swamp buffalo, through its function as receptor bacteria attachment. This is suggested as aspecial characteristic in digestive system of swamp buffalo which causes high digestive efficiency inswamp buffalo.

  14. Lectin binding profiles of SSEA-4 enriched, pluripotent human embryonic stem cell surfaces

    Directory of Open Access Journals (Sweden)

    Shin Soojung

    2005-07-01

    Full Text Available Abstract Background Pluripotent human embryonic stem cells (hESCs have the potential to form every cell type in the body. These cells must be appropriately characterized prior to differentiation studies or when defining characteristics of the pluripotent state. Some developmentally regulated cell surface antigens identified by monoclonal antibodies in a variety of species and stem cell types have proven to be side chains of membrane glycolipids and glycoproteins. Therefore, to examine hESC surfaces for other potential pluripotent markers, we used a panel of 14 lectins, which were chosen based on their specificity for a variety of carbohydrates and carbohydrate linkages, along with stage specific embryonic antigen-4 (SSEA-4, to determine binding quantitation by flow cytometry and binding localization in adherent colonies by immunocytochemistry. Results Enriching cells for SSEA-4 expression increased the percentage of SSEA-4 positive cells to 98–99%. Using enriched high SSEA-4-expressing hESCs, we then analyzed the binding percentages of selected lectins and found a large variation in binding percentages ranging from 4% to 99% binding. Lycopersicon (tomatoesculetum lectin (TL, Ricinus communis agglutinin (RCA, and Concanavalin A (Con A bound to SSEA-4 positive regions of hESCs and with similar binding percentages as SSEA-4. In contrast, we found Dolichos biflorus agglutinin (DBA and Lotus tetragonolobus lectin (LTL did not bind to hESCs while Phaseolus vulgaris leuco-agglutinin (PHA-L, Vicia villosa agglutinin (VVA, Ulex europaeus agglutinin (UEA, Phaseolus vulgaris erythro-agglutinin (PHA-E, and Maackia amurensis agglutinin (MAA bound partially to hESCs. These binding percentages correlated well with immunocytochemistry results. Conclusion Our results provide information about types of carbohydrates and carbohydrate linkages found on pluripotent hESC surfaces. We propose that TL, RCA and Con A may be used as markers that are associated with the

  15. The Glycoprofile Patterns of Endothelial Cells in Usual Interstitial Pneumonia

    Directory of Open Access Journals (Sweden)

    A Barkhordari

    2014-09-01

    Full Text Available [THIS ARTICLE HAS BEEN RETRACTED FOR DUPLICATE PUBLICATION] Background: The pathological classification of cryptogenic fibrosing alveolitis has been a matter of debate and controversy for histopathologists.Objective: To identify and specify the glycotypes of capillary endothelial cells in usual interstitial pneumonia (UIP compared to those found in normal tissue.Methods: Sections of formalin-fixed, paraffin-embedded blocks from 16 cases of UIP were studied by lectin histochemistry with a panel of 27 biotinylated lectins and an avidin-peroxidase revealing system.Results: High expression of several classes of glycan was seen de novo in capillary endothelial cells from patients with UIP including small complex and bi/tri-antennary bisected complex N-linked sequences bolund by Concanavalin A and erythro-phytohemagglutinin, respectively, GalNAca1 residues bound by Helix pomatia and Maclura pomifera agglutinins, and L-fucosylated derivatives of type II glycan chains recognized by Ulex europaeus agglutinin-I. Glycans bound by agglutinins from Lycopersicon esculentum (β1,4GlcNAc and Wisteria floribunda (GalNAc as well as GlcNAc oligomers bound by Phytolacca americana and succinylated Wheat Germ agglutinin were also seen in the capillary endothelial cells of UIP. In contrast, L-fucosylated derivatives of type I glycan chains were absent in cells from cases of UIP when Anguilla anguilla agglutinin was applied, unlike the situation in normal tissue.Conclusion: These results may indicate existence of two distinct populations of endothelial cell in UIP with markedly different patterns of glycosylation, reflecting a pattern of differentiation and angiogenesis, which is not detectable morphologically.

  16. Seroepidemiology of Toxoplasma gondii in dogs in Trinidad and Tobago.

    Science.gov (United States)

    Ali, C N; Harris, J A; Watkins, J D; Adesiyun, A A

    2003-05-01

    A cross-sectional study was conducted to determine the seroprevalence of Toxoplasma gondii agglutinins and to investigate the relationship between various risk factors and occurrence of toxoplasmosis in dogs in Trinidad. Of a total 250 dogs, comprising domestic, hunting and stray dogs, 80 (32.0%) were positive for T. gondii agglutinins at a titre of > or =1:32 using a latex agglutination test. Stray dogs (60.5%) had statistically significantly higher (P2-3 years age group compared with other age groups. Dogs that consumed home-cooked foods had a seroprevalence of 32.9% compared with those fed commercial dog foods (17.2%) and dogs fed both home-cooked and commercial foods (21.0%). However, the difference was not statistically significant (P>0.05; chi(2)). The rather high prevalence of toxoplasmosis in stray dogs is a good indication of the extent of the infection in the environment. PMID:12719132

  17. Physico-chemical characterization of bacterial and hemagglutinins from the serum of the mud crab Scylla serrata

    Directory of Open Access Journals (Sweden)

    SS Jayaraj

    2010-02-01

    Full Text Available A naturally occurring hemagglutinin (HA, with activity against bacteria and yeast cells were detected in the serum of Scylla serrata using mammalian erythrocytes (RBC, various bacteria and yeast as indicator cells. The serum gave highest HA titer with rabbit RBC, tripsinized yeast and Vibrio fluvialis. An analysis of the physico-chemical properties of the HA showed it to be specifically dependent on the presence of Ca2+ for its activity, stable between pH 7 to 9 and showed thermal stability between 10 to 30 °C. Further studies demonstrated that the HA is precipitable by ammonium sulphate and TCA. HA- inhibition assays performed with carbohydrates revealed that the serum HA was specific for non-reducing terminal glucose with 1-2 glucosidic linkages. Thus this agglutinin appears to be unique among all the known crustacean agglutinins.

  18. The immunological properties of Brucella ribosomal preparations.

    Science.gov (United States)

    Corbel, M J

    1976-01-01

    Ribosomes were isolated from Brucella abortus strains 19 and 45/20 by disruption of the cells followed by differential ultracentrifugation. The ribosome preparations contained 2-3 components reacting in immunodiffusion tests but were free of detectable lipopolysaccharide-protein agglutinogen. They crossreacted with antisera to Br. abortus, Br. melitensis, Br. suis and Br. ovis and elicited intradermal delayed hypersensitivity reactions in animals infected with Br. abortus, Br. melitensis or Br. suis. The ribosomes were antigenic in rabbits, guinea pigs and mice. Those from Br. abortus S19 induced agglutinins reaction with smooth brucella strains whereas those from Br. abortus 45/20 induced agglutinins reacting with rough brucella strains. Cattle vaccinated with S19 or 45/20 vaccines or infected with Br. abortus developed pricipitins to ribosomal components at an early stage in the immune response. PMID:816681

  19. PESQUISA DE AGLUTININAS ANTILEPTOSPIRA EM SOROS SANGUÍNEOS DE ASININOS (Equus Asinus E DE CONDUTORES DE VEÍCULOS DE TRAÇÃO ANIMAL NA CIDADE DE SÃO LUÍS, MA, BRASIL

    Directory of Open Access Journals (Sweden)

    Danilo Cutrim Bezerra

    2010-12-01

    Full Text Available With the objective of searching the presence of antileptospiras agglutinin in sanguine serum of donkeys used in animal traction vehicles and of the wagon’s conductors in the city of São Luis, MA, and of identifying serovars of higher occurrence, 60 serum samples of donkeys and 60 of conductors were tested regarding 30 serovars of Leptospira interrogans by using the microscopic seroagglutination test. Of the 60 blood serum samples of donkeys, 85% showed seropositivity, reacting to 21 serovars out of the 30 tested. Copenhageni, Pyrogenes, Autumnalis and Icterohaemorrhagiae were the most frequent ones, in decreasing sequence. Regarding the serum of the conductors, 38.34% presented seropositivity, reacting to 12 serovars of Leptospira interrogans and the most frequenty ones were Copenhageni, Pyrogenes, Icterohaemorrhagiae and Autumnalis. These results indicate a high incidence of anti-leptospira agglutinins in donkeys and humans, being serovars copenhageni and pyrogenes the most prevalent.

  20. AcEST: DK948418 [AcEST

    Lifescience Database Archive (English)

    Full Text Available or domain OS=St... 46 0.002 tr|A2DMT1|A2DMT1_TRIVA A-agglutinin attachment subunit, putative.....ESD 1725 >tr|A2DMT1|A2DMT1_TRIVA A-agglutinin attachment subunit, putative OS=Trichomonas vaginalis G3 GN=TV....3 E-value 2.0e-05 Report BLASTX 2.2.19 [Nov-02-2008] Reference: Altschul, Stephe... adhesin for platelets OS=Staphy... 46 2e-04 sp|Q99QY4|SRAP_STAAM Serine-rich adhesin for platelets... OS=Staphy... 46 2e-04 sp|Q8NUJ3|SRAP_STAAW Serine-rich adhesin for platelets OS=Staphy... 45 5e-04 sp|Q

  1. AcEST: DK961080 [AcEST

    Lifescience Database Archive (English)

    Full Text Available 6 2.0 tr|A2DMT1|A2DMT1_TRIVA A-agglutinin attachment subunit, putative... 35 3.5 tr|Q4SYS6|Q4SYS6_TETNG Chromosome undet...080 Tissue type prothallia with plantlets Developmental stage gametophytes with s... SYD ++P SN Y QT +P+Q Sbjct: 307 LTKFSVISDSTCFSSSK-FTTKSYDIESPFKQSNNYVNQTPQPSQ 350 >tr|A2DMT1|A2DMT1_TRIVA A-agglutinin attachme... 2.2.19 [Nov-02-2008] Reference: Altschul, Stephen F., Thomas L. Madden, Alejandro A. Schaffer, Jinghui Zhan...omonas vaginalis G3 Align length 97 Score (bit) 38.1 E-value 0.53 Report BLASTX 2.2.19 [Nov-02-2008] Reference: Altsch

  2. Increased fucosyl glycoconjugate by Mycoplasma hyopneumoniae enhances adherences of Pasteurella multocida type A in the ciliated epithelial cells of the respiratory tract

    OpenAIRE

    Park, Changhoon; Jeong, Jiwoon; KANG, Ikjae; Choi, Kyuhyung; Park, Su-Jin; Chae, Chanhee

    2016-01-01

    Background The objective of this study was to elucidate the pathogenic mechanisms of how Mycoplasma hyopneumoniae enhances secondary Pasteurella multocida type A infection which leads to porcine enzootic pneumonia in infected pigs. Sixteen pigs were experimentally infected with M. hyopneumoniae and then euthanized at 7, 14, 21 and 28 days post inoculation. In situ hybridization for M. hyopneumoniae DNA and Ulex europaeus agglutinin-I (UEA-I) lectin histochemistry for fucosyl glycoconjugate, w...

  3. Identification of lectin-binding proteins in Chlamydia species.

    OpenAIRE

    Swanson, A F; Kuo, C. C.

    1990-01-01

    Lectin-binding proteins of chlamydiae were detected by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and immunoblotting. All three Chlamydia species tested expressed two proteins when whole-elementary-body lysates were reacted with the biotinylated lectin Dolichos biflorus agglutinin. The protein with a molecular mass of 18 kilodaltons (kDa) responded strongly compared with a higher-molecular-mass protein that varied from 27 to 32 kDa with each chlamydia strain tested. Among six l...

  4. Glycoconjugates Distribution during Developing Mouse Spinal Cord Motor Organizers

    OpenAIRE

    Vojoudi, Elham; Ebrahimi, Vahid; Ebrahimzadeh-Bideskan, Alireza; Fazel, Alireza

    2015-01-01

    Background: The aim of this research was to study the distribution and changes of glycoconjugates particularly their terminal sugars by using lectin histochemistry during mouse spinal cord development. Methods: Formalin-fixed sections of mouse embryo (10-16 fetal days) were processed for lectin histochemical method. In this study, two groups of horseradish peroxidase -labeled specific lectins were used: N-acetylgalactosamine, including Dolichos biflorus, Wisteria floribunda agglutinin (WFA), ...

  5. Endothelial progenitor cells regenerate infracted myocardium with neovascularisation development ☆

    OpenAIRE

    M.T. Abd El Aziz; Abd El Nabi, E.A.; Abd El Hamid, M.; D. Sabry; Atta, H.M.; L.A. Rahed; A. Shamaa; Mahfouz, S.; Taha, F.M.; S. Elrefaay; Gharib, D.M.; Elsetohy, Khaled A

    2013-01-01

    We achieved possibility of isolation, characterization human umbilical cord blood endothelial progenitor cells (EPCs), examination potency of EPCs to form new blood vessels and differentiation into cardiomyoctes in canines with acute myocardial infarction (AMI). EPCs were separated and cultured from umbilical cord blood. Their phenotypes were confirmed by uptake of double stains dioctadecyl tetramethylindocarbocyanine-labeled acetylated LDL and FITC-labeled Ulex europaeus agglutinin 1 (DILDL-...

  6. M-Cell Targeting of Whole Killed Bacteria Induces Protective Immunity against Gastrointestinal Pathogens▿

    OpenAIRE

    Chionh, Yok-Teng; Wee, Janet L. K.; Every, Alison L.; Ng, Garrett Z.; Sutton, Philip

    2009-01-01

    As the majority of human pathogens infect via a mucosal surface, delivery of killed vaccines by mucosal routes could potentially improve protection against many such organisms. Our ability to develop effective killed mucosal vaccines is inhibited by a lack of adjuvants that are safe and effective in humans. The Ulex europaeus agglutinin I (UEA-I) lectin specifically binds M cells lining the murine gastrointestinal tract. We explored the potential for M-cell-targeted vaccination of whole, kill...

  7. Biochimical and Structural caracterization of a lectin from Platypodium elegans Vogel seeds

    OpenAIRE

    Leite, Raquel,

    2011-01-01

    Lectin activity with specificity for mannose and glucose has been detected in the seed of Platypodium elegans, a legume from the Dalbergiae tribe. The gene of the lectin PELa has been cloned and the resulting 261 amino acid protein belongs to the legume lectin family with similarity with Pterocarpus angolensis agglutinin (PAL) from the same tribe. The recombinant lectin has been expressed in E. coli and refolded from inclusion bodies. Analysis of specificity by Glycan Array evidenced a very u...

  8. Diagnosis of myocardial infarction based on lectin-induced erythrocyte agglutination: a feasibility study

    Science.gov (United States)

    Bocsi, József; Nieschke, Kathleen; Mittag, Anja; Reichert, Thomas; Laffers, Wiebke; Marecka, Monika; Pierzchalski, Arkadiusz; Piltz, Joachim; Esche, Hans-Jürgen; Wolf, Günther; Dähnert, Ingo; Baumgartner, Adolf; Tarnok, Attila

    2014-03-01

    Myocardial infarction (MI) is an acute life-threatening disease with a high incidence worldwide. Aim of this study was to test lectin-carbohydrate binding-induced red blood cell (RBC) agglutination as an innovative tool for fast, precise and cost effective diagnosis of MI. Five lectins (Ricinus communis agglutinin (RCA), Phaseolus vulgaris erythroagglutinin (PHA), Datura stramonium agglutinin (DSA), Artocarpus agglutinin (ArA), Triticum agglutinin (TA)) were tested for ability to differentiate between agglutination characteristics in patients with MI (n = 101) or angina pectoris without MI (AP) (n = 34) and healthy volunteers (HV) as control (n =68) . RBC agglutination was analyzed by light absorbance of a stirred RBC suspension in the green to red light spectrum in an agglutimeter (amtec, Leipzig, Germany) for 15 min after lectin addition. Mean cell count in aggregates was estimated from light absorbance by a mathematical model. Each lectin induced RBC agglutination. RCA led to the strongest RBC agglutination (~500 RBCs/aggregate), while the others induced substantially slower agglutination and lead to smaller aggregate sizes (5-150 RBCs/aggregate). For all analyzed lectins the lectin-induced RBC agglutination of MI or AP patients was generally higher than for HV. However, only PHA induced agglutination that clearly distinguished MI from HV. Variance analysis showed that aggregate size after 15 min. agglutination induced by PHA was significantly higher in the MI group (143 RBCs/ aggregate) than in the HV (29 RBC-s/aggregate, p = 0.000). We hypothesize that pathological changes during MI induce modification of the carbohydrate composition on the RBC membrane and thus modify RBC agglutination. Occurrence of carbohydrate-lectin binding sites on RBC membranes provides evidence about MI. Due to significant difference in the rate of agglutination between MI > HV the differentiation between these groups is possible based on PHA-induced RBC-agglutination. This novel assay

  9. Specificity of lectin-immobilized fluorescent nanospheres for colorectal tumors in a mouse model which more resembles the clinical disease

    OpenAIRE

    Kitamura, Tokio; Sakuma, Shinji; Shimosato, Moe; Higashino, Haruki; Masaoka, Yoshie; Kataoka, Makoto; Yamashita, Shinji; Hiwatari, Ken-Ichiro; Kumagai, Hironori; Morimoto, Naoki; Koike, Seiji; Tobita, Etsuo; Hoffman, Robert M.; John C Gore; Pham, Wellington

    2014-01-01

    We are investigating an imaging agent that enables real-time and accurate diagnosis of early colorectal cancer at the intestinal mucosa by colonoscopy. The imaging agent is peanut agglutinin-immobilized polystyrene nanospheres with surface poly(N-vinylacetamide) chains encapsulating coumarin 6. Intracolonically-administered lectin-immobilized fluorescent nanospheres detect tumor-derived changes through molecular recognition of lectin for the terminal sugar of cancer-specific antigens on the m...

  10. Histological and Lectin Histochemical Studies on the Olfactory and Respiratory Mucosae of the Sheep

    OpenAIRE

    Ibrahim, Dalia; NAKAMUTA, Nobuaki; TANIGUCHI, Kazumi; Yamamoto, Yoshio; TANIGUCHI, Kazuyuki

    2013-01-01

    ABSTRACT The olfactory and respiratory mucosae of the Corriedale sheep were examined using lectin histochemistry in order to clarify the histochemical and glycohistochemical differences between these two tissues. The olfactory epithelium was stained with 13 lectins out of 21 lectins examined, while the respiratory epithelium was positive to 16 lectins. The free border of both of the olfactory and respiratory epithelia was stained with 12 lectins: Wheat germ agglutinin (WGA), succinylated-whea...

  11. The A0 blood group genotype modifies the jejunal glycomic binding pattern profile of piglets early associated with a simple or complex microbiota.

    Science.gov (United States)

    Priori, D; Colombo, M; Koopmans, S-J; Jansman, A J M; van der Meulen, J; Trevisi, P; Bosi, P

    2016-02-01

    The intestinal epithelium glycocalyx sugar motif is an important determinant of the bacterial-host interaction and may be affected in pigs by gut microbiota and by blood group genotype. The aim was to study the effect of intestinal association with different microbiota and A0 blood group genotypes on the expressed glycomic pattern in the small intestine. Twelve caesarean-derived pigs previously associated with a simple association (SA) or complex association (CA) microbiota were selected at 26 to 37 d of age. In each subject, different jejunal loops were perfused for 8 h with enterotoxigenic K88 (ETEC), ETEC fimbriae (F4), (LAM), or a saline control. The piglets were genotyped for A0 blood group and the glycomic profile was evaluated by microscopic screening of lectin binding: peanut agglutinin (PNA), which is galactose specific; agglutinin I (UEA), which is fucose specific; lectin II (MALii), which is sialic acid specific; concavalin A, which is mannose specific; soybean agglutinin (SBA), which is -acetyl-galactosamine specific; and wheat germ agglutinin (WGA), which is -acetyl-glucosamine specific. A0 pigs had fewer UEA-positive cells, MALii-positive cells ( < 0.001), and SBA-positive cells ( < 0.10) than 00 pigs. Simple association pigs had more SBA positive cells ( < 0.01) than CA pigs. Enterotoxigenic K88-perfused intestinal loops had fewer UEA-positive cells ( < 0.01) and WGA positive cells ( < 0.001) cells and more PNA positive cells (only in SA pigs, < 0.01). No effects of introduction of F4 and LAM in the intestinal lumen were observed. The porcine A0 blood group genotype and the luminal presence of ETEC strongly affected the jejunal mucosa glycomic pattern profile whereas an early oral simple or complex microbial association had limited effects. Pig genetic background has relevance on the cross talk between intestinal epithelium glycocalyx sugar motif and ETEC and, ultimately, on the gut microbial colonization in later life. PMID:27065129

  12. A Novel Lectin with Antiproliferative and HIV-1 Reverse Transcriptase Inhibitory Activities from Dried Fruiting Bodies of the Monkey Head Mushroom Hericium erinaceum

    OpenAIRE

    Yanrui Li; Guoqing Zhang; Tzi Bun Ng; Hexiang Wang

    2010-01-01

    A lectin designated as Hericium erinaceum agglutinin (HEA) was isolated from dried fruiting bodies of the mushroom Hericium erinaceum with a chromatographic procedure which entailed DEAE-cellulose, CM-cellulose, Q-Sepharose, and FPLC Superdex 75. Its molecular mass was estimated to be 51 kDa and its N-terminal amino acid sequences was distinctly different from those of other isolated mushroom lectins. The hemagglutinating activity of HEA was inhibited at the minimum concentration of 12...

  13. Acetylcholinesterase-positive afferent axons in mucosa of urinary bladder of adult cats: retrograde tracing and degeneration studies

    OpenAIRE

    Wakabayashi, Y.; Kojima, Y.; Makiura, Y.; Tomoyoshi, T.; Maeda, T.

    1995-01-01

    Acetylcholinesterase (AchE)-positive afferent axons in the mucosa of the cat urinary bladder were examined in the present experiments. Smallsized dorsal root ganglion cells containing AchE enzyme activity were labelled by injection of retrograde tracer (wheat germ agglutinin conjugated to enzymatically inactive horseradish peroxidase gold complex) into the bladder mucosa of adult cats. Results show that 48.9% (901184) of the labelled ganglion cells possesse...

  14. Anti-M Antibody in Solid Tumors-Two Case Reports

    OpenAIRE

    Soni, Shiv Kumar; Goyal, Hari; Sood, S K; Setia, Rasika

    2013-01-01

    Anti-M antibodies are usually of IgM, appear as cold agglutinins and are clinically insignificant. We are reporting two cases of anti-M in cases of solid tumors where the anti-M caused discrepancy in blood grouping, reacted in coombs phase of crossmatching. Anti-M in first case showed dosage effect. These antibodies can be clinical significant when detected in coombs phase, making M antigen negative coombs compatible unit transfusion imperative.

  15. [Acute oliguric renal failure and haemolytic anaemia following infectious mononucleosis].

    Science.gov (United States)

    Brkovic, Natasa; Jørgensen, Kit Riegels; Rosenbæk, Jeppe Bakkestrøm; Pedersen, Erling Bjerregaard

    2015-11-01

    A 19-year-old man was admitted to hospital due to fatigue, nausea, abdominal pain and faint. He was pale and icteric, awake with sufficient respiration and circulation. He had infectious mononucleosis complicated with acute oliguric renal failure and severe haemolytic anaemia with a positive Coombs test. He had a cold agglutinin syndrome. The treatment comprised intermittent haemodialysis, plasmapheresis and heating. He recovered completely after two months. PMID:26573947

  16. Maintenance of biological activity of pertussis toxin radioiodinated while bound to fetuin-agarose.

    OpenAIRE

    Armstrong, G. D.; Peppler, M S

    1987-01-01

    We developed a method to produce radioiodinated pertussis toxin (PT) which was active in the goose erythrocyte agglutination and CHO cell assay systems. The procedure used fetuin coupled to agarose to prevent inactivation of the toxin during the iodination reaction. Analysis of the labeled PT by affinity chromatography on fetuin-agarose and wheat germ agglutinin-agarose and by sodium dodecyl sulfate-polyacrylamide gel electrophoresis indicated that there were minimal amounts of labeled fetuin...

  17. DBA-Lectin Reactivity Defines Mouse Uterine Natural Killer Cell Subsets with Biased Gene Expression 1

    OpenAIRE

    Chen, Zhilin; Zhang, Jianhong; Hatta, Kota; Lima, Patricia D.A.; Yadi, Hakim; Colucci, Francesco; Yamada, Aureo T.; Croy, B. Anne

    2012-01-01

    Endometrial decidualization, a process essential for blastocyst implantation in species with hemochorial placentation, is accompanied by an enormous but transient influx of Natural Killer (NK) cells. Mouse uterine (u)NK cell subsets have been defined by diameter and cytoplasmic granule number, reflecting stage of maturity and by histochemical reactivity with Periodic Acid Schiff’s (PAS) reagent, with or without co-reactivity with Dolichos biflorus agglutinin (DBA) lectin. We asked whether DBA...

  18. Optimising expression of the recombinant fusion protein biopesticide ω-hexatoxin-Hv1a /GNA in Pichia pastoris : sequence modifications and a simple method for the generation of multi-copy strains.

    OpenAIRE

    Pyati, P; Fitches, E.; Gatehouse, J A

    2014-01-01

    Production of recombinant protein bio-insecticides on a commercial scale can only be cost effective if host strains with very high expression levels are available. A recombinant fusion protein containing an arthropod toxin, ω-hexatoxin-Hv1a, (from funnel web spider Hadronyche versuta) linked to snowdrop lectin (Galanthus nivalis agglutinin; GNA) is an effective oral insecticide and candidate biopesticide. However, the fusion protein was vulnerable to proteolysis during production in the yeast...

  19. Accessory Olfactory Bulb Function is Modulated by Input from the Main Olfactory Epithelium

    OpenAIRE

    Slotnick, Burton; Restrepo, Diego; Schellinck, Heather; Archbold, Georgina; Price, Stephen; Lin, Weihong

    2010-01-01

    While it is now established that sensory neurons in both the main olfactory epithelium and the vomeronasal organ may be activated by both general and pheromonal odorants, it remains unclear what initiates sampling by the VNO. Anterograde transport of wheat germ agglutinin-horseradish peroxidase was used to determine that adequate intranasal syringing with zinc sulfate interrupted all inputs to the main olfactory bulb but left intact those to the accessory olfactory bulb. Adult male treated mi...

  20. Does Candida albicans Als5p Amyloid Play a Role in Commensalism in Caenorhabditis elegans?

    OpenAIRE

    Bois, Michael; Singh, Sean; Samlalsingh, Alyssa; Lipke, Peter N.; Garcia, Melissa C.

    2013-01-01

    Candida albicans, a dimorphic fungus and an opportunistic pathogen, possesses a myriad of adherence factors, including members of the agglutinin-like sequence (Als) family of mannoproteins. The adhesin Als5p mediates adhesion to many substrates and is upregulated during commensal interactions but is downregulated during active C. albicans infections. An amyloid-forming core sequence at residues 325 to 331 is important for Als5p function, because a single-amino-acid substitution at position 32...

  1. Interleukin 2-dependent and interleukin 2-independent pathways of regulation of thymocyte function by interleukin 6.

    OpenAIRE

    Le, J M; Fredrickson, G.; Reis, L F; Diamantstein, T; Hirano, T.; Kishimoto, T.; Vilcek, J

    1988-01-01

    Recombinant human interleukin 6 (IL-6), also termed B-cell-stimulatory factor 2 (BSF-2) or interferon-beta 2, was found to stimulate the proliferation of mouse thymocytes costimulated with phytohemagglutinin (PHA). In addition, IL-6 synergistically enhanced the stimulation of thymocyte proliferation by recombinant human interleukin 1 (IL-1) or interleukin 2 (IL-2). Mature thymocytes lacking peanut agglutinin receptor are the main target of IL-6 action. Incubation of thymocytes with IL-6 in th...

  2. Tyrosine-specific protein kinase activity is associated with the purified insulin receptor.

    OpenAIRE

    Kasuga, M.; Fujita-Yamaguchi, Y; Blithe, D L; Kahn, C. R.

    1983-01-01

    Highly purified human placental insulin receptors were obtained by sequential affinity chromatography on wheat germ agglutinin and insulin-agarose. The preparation had an insulin binding capacity of 4,700 pmol/mg of protein approaching theoretical purity. The purified receptor revealed three major bands of Mr 135,000, 95,000, and 52,000 in NaDodSO4/polyacrylamide gel electrophoresis after reduction by dithiothreitol. All three bands were immunoprecipitated by anti-insulin-receptor antibodies....

  3. Flow cytometry for age assessment of a yeast population and its application in beer fermentations

    OpenAIRE

    Kuřec, Michal; Baszczyňski, Martin; Lehnert, Radek; Mota, André; Teixeira, J.A.; Brányik, Tomáš

    2009-01-01

    An expeditious method of yeast age estimation was developed based on selective bud scar staining (Alexa Fluor 488-labelled wheat-germ agglutinin) and subsequent fluorescence intensity measurement by flow cytometry. The calibration curve resulting from the cytometric determination of average bud scar fluorescence intensities vs. microscopically counted average bud scar numbers of the same cell populations showed a good correlation and allowed routine cell age estimation by...

  4. Dietary Plant Lectins Appear to Be Transported from the Gut to Gain Access to and Alter Dopaminergic Neurons of Caenorhabditis elegans, a Potential Etiology of Parkinson's Disease.

    Science.gov (United States)

    Zheng, Jolene; Wang, Mingming; Wei, Wenqian; Keller, Jeffrey N; Adhikari, Binita; King, Jason F; King, Michael L; Peng, Nan; Laine, Roger A

    2016-01-01

    Lectins from dietary plants have been shown to enhance drug absorption in the gastrointestinal tract of rats, be transported trans-synaptically as shown by tracing of axonal and dendritic paths, and enhance gene delivery. Other carbohydrate-binding protein toxins are known to traverse the gut intact in dogs. Post-feeding rhodamine- or TRITC-tagged dietary lectins, the lectins were tracked from gut to dopaminergic neurons (DAergic-N) in transgenic Caenorhabditis elegans (C. elegans) [egIs1(Pdat-1:GFP)] where the mutant has the green fluorescent protein (GFP) gene fused to a dopamine transport protein gene labeling DAergic-N. The lectins were supplemented along with the food organism Escherichia coli (OP50). Among nine tested rhodamine/TRITC-tagged lectins, four, including Phaseolus vulgaris erythroagglutinin (PHA-E), Bandeiraea simplicifolia (BS-I), Dolichos biflorus agglutinin (DBA), and Arachis hypogaea agglutinin (PNA), appeared to be transported from gut to the GFP-DAergic-N. Griffonia Simplicifolia and PHA-E, reduced the number of GFP-DAergic-N, suggesting a toxic activity. PHA-E, BS-I, Pisum sativum (PSA), and Triticum vulgaris agglutinin (Succinylated) reduced fluorescent intensity of GFP-DAergic-N. PHA-E, PSA, Concanavalin A, and Triticum vulgaris agglutinin decreased the size of GFP-DAergic-N, while BS-I increased neuron size. These observations suggest that dietary plant lectins are transported to and affect DAergic-N in C. elegans, which support Braak and Hawkes' hypothesis, suggesting one alternate potential dietary etiology of Parkinson's disease (PD). A recent Danish study showed that vagotomy resulted in 40% lower incidence of PD over 20 years. Differences in inherited sugar structures of gut and neuronal cell surfaces may make some individuals more susceptible in this conceptual disease etiology model. PMID:27014695

  5. Differential lectin binding on walls of thoraco-cervical blood vessels and lymphatics in rats.

    Science.gov (United States)

    Kagami, H; Uryu, K; Okamoto, K; Sakai, H; Kaneda, T; Sakanaka, M

    1991-08-01

    Lectin binding in the walls of large to medium-sized blood vessels and lymphatics in the rat thoraco-cervical region was examined histochemically. The tunica intima of the aorta and superficial cervical artery showed positive reactions with wheat germ agglutinin (WGA) and Concanavalin A (ConA) but not with Dolichus biflorus agglutinin (DBA). The tunica media of the aorta exhibited intense WGA binding, especially on the smooth muscle cells, but the tunica media of the superficial cervical artery did not react with the lectin. Neither ConA nor DBA bound to the tunica media of the aorta and superficial cervical artery. The tunica adventitia of both arteries contained sites binding the three lectins, although DBA reactivity declined as the vascular diameter decreased. The tunica intima of the superior vena cava and azygos vein exhibited positive WGA and ConA binding, whereas DBA binding was noted on only part of the tunica intima of the superior vena cava and not on that of the azygos vein. The tunica media and tunica adventitia were reactive for all three lectins. The WGA and ConA binding sites in the tunica adventitia showed loose networks, suggesting lectin binding on connective tissue elements interlacing among smooth muscle bundles. Lectin binding sites in the walls of lymphatics exhibited an arrangement similar to those in the walls of the veins. Moreover valves protruding into the lumen showed intense WGA and ConA binding and scattered DBA binding. Three other lectins (Ulex europaeus agglutinin, peanut agglutinin, Maclura pomifera) were examined, but they showed no reactions with the vessels. Thus, the differential binding of lectins on the walls of blood vessels and lymphatics of various sizes suggests the functional complexity of monosaccharide residues in the vascular walls. PMID:1758681

  6. Streptococcus mutans Adherence: Presumptive Evidence for Protein-Mediated Attachment Followed by Glucan-Dependent Cellular Accumulation

    OpenAIRE

    Staat, Robert H.; Langley, Sharon D.; Doyle, R J

    1980-01-01

    Adherence of Streptococcus mutans to smooth surfaces has been attributed to the production of sucrose-derived d-glucans. However, several studies indicate that the bacterium will adhere in the absence of sucrose. The present data confirmed that S. mutans adherence to saliva-coated hydroxyapatite beads in the absence of sucrose is described by the Langmuir equation. The nature of the sucrose-independent adherence was studied with the Persea americana agglutinin as a selective adherence inhibit...

  7. Biochemical characterization, localization and immunostimulating properties of a soluble glycoprotein, Ag1, isolated from in vitro cultures of Plasmodium falciparum

    DEFF Research Database (Denmark)

    Jakobsen, P H; Jepsen, S; Riley, E M; Theander, T G; Grellier, P; Lihme, A; Hviid, L; Dziegiel, M; Schrevel, J

    1990-01-01

    Erythrina christagalli agglutinin, which is specific for carbohydrates bearing beta-D-galactose(1-4)-D-N-acetylglucosamine. Indirect immunofluorescence studies showed that Ag1 is located on the surface of trophozoites and schizonts but not on the surface of merozoites. Ag1 is recognized by human immune sera...... from six different malaria-endemic regions. Ag1 induces in vitro proliferation of lymphocytes from malaria-immune individuals in an antigen-specific manner....

  8. Biochemical characterization, localization and immunostimulating properties of a soluble glycoprotein, Ag1, isolated from in vitro cultures of Plasmodium falciparum

    DEFF Research Database (Denmark)

    Jakobsen, P H; Jepsen, S; Riley, E M; Theander, T G; Grellier, P; Lihme, A; Hviid, L; Dziegiel, M; Schrevel, J

    1990-01-01

    Erythrina christagalli agglutinin, which is specific for carbohydrates bearing beta-D-galactose(1-4)-D-N-acetylglucosamine. Indirect immunofluorescence studies showed that Ag1 is located on the surface of trophozoites and schizonts but not on the surface of merozoites. Ag1 is recognized by human immune sera...... from six different malaria-endemic regions. Ag1 induces in vitro proliferation of lymphocytes from malaria-immune individuals in an antigen-specific manner....

  9. Lectin-binding sites as markers of Golgi subcompartments: proximal-to- distal maturation of oligosaccharides

    OpenAIRE

    1983-01-01

    We investigated the subcellular sites of glycoprotein oligosaccharide maturation by using lectin conjugates to stain lightly-fixed, saponin- permeabilized myeloma cells. At the electron microscopic level, concanavalin A-peroxidase stains the cisternal space of the nuclear envelope, the rough endoplasmic reticulum, and cisternae along the proximal face of the Golgi stack. Conversely, wheat germ agglutinin- peroxidase stains cisternae along the distal face of the Golgi stack, associated vesicle...

  10. Development and Organization of Ocular Dominance Bands in Primary Visual Cortex of the Sable Ferret

    OpenAIRE

    RUTHAZER, E.S.; Baker, G. E.; Stryker, M.P.

    1999-01-01

    Thalamocortical afferents in the visual cortex of the adult sable ferret are segregated into eye-specific ocular dominance bands. The development of ocular dominance bands was studied by transneuronal labeling of the visual cortices of ferret kits between the ages of postnatal day 28 (P28) and P81 after intravitreous injections of either tritiated proline or wheat germ agglutinin-horseradish peroxidase. Laminar specificity was evident in the youngest animals studied and was similar to that in...

  11. Direct Production of Ethanol from Raw Corn Starch via Fermentation by Use of a Novel Surface-Engineered Yeast Strain Codisplaying Glucoamylase and α-Amylase

    OpenAIRE

    Shigechi, Hisayori; Koh, Jun; Fujita, Yasuya; Matsumoto, Takeshi; Bito, Yohei; Ueda, Mitsuyoshi; SATOH, Eiichi; Fukuda, Hideki; Kondo, Akihiko

    2004-01-01

    Direct and efficient production of ethanol by fermentation from raw corn starch was achieved by using the yeast Saccharomyces cerevisiae codisplaying Rhizopus oryzae glucoamylase and Streptococcus bovis α-amylase by using the C-terminal-half region of α-agglutinin and the flocculation functional domain of Flo1p as the respective anchor proteins. In 72-h fermentation, this strain produced 61.8 g of ethanol/liter, with 86.5% of theoretical yield from raw corn starch.

  12. Lectin histochemistry and ultrastructure of feline kidneys from six different storage diseases.

    Science.gov (United States)

    Castagnaro, M; Alroy, J; Ucci, A A; Glew, R H

    1987-01-01

    We have compared the pattern of lectin staining with the ultrastructural features of kidneys from normal cats and 19 cats with 6 different lysosomal storage diseases. The diseases studied include GM1 and GM2 gangliosidosis, mucopolysaccharidosis (MPS)-I and MPS-VI, sphingomyelin-lipidosis (i.e., Niemann-Pick disease) and mannosidosis. Ten different biotinylated lectins were used as histochemical probes for carbohydrate residues and avidin-biotin-peroxidase complex as visualant. Concanavalia ensiformis agglutinin (Con A) stained mesangial cells in all storage diseases but GM1, epithelial cells in sphingomyelin-lipidosis and mannosidosis, endothelial cells in GM1 and mannosidosis and Bowman's capsule cells in all but GM2. Griffonia simplicifolia agglutinin I (GS-I) stained the glomerular endothelium in all six diseases, but not in control kidneys. Ricinus communis agglutinin-I (RCA-I) stained the glomerular epithelium only in GM1 and MPS-I. Succinylated wheat germ agglutinin (SWGA) stained the glomerular endothelium and epithelium in mannosidosis, and the glomerular epithelium and Bowman's capsule in MPS-I. Ultrastructure studies demonstrated an accumulation of oligosaccharides in cases of mannosidosis and GM1 gangliosidosis, a mixture of oligosaccharides and lipids in MPS-I, MPS-VI and GM2 gangliosidosis and only lipid storage in sphingomyelin lipidosis. These studies show that morphologic and histochemical changes are manifested in some kidney cell types in lysosomal storage diseases, even though the enzyme deficiency occurs in all cell types. Furthermore, we show that the nature of the undegraded stored material is complex and that other factors, such as rate of membrane turn over, membrane composition, and cell function may influence the amount and nature of the "stored" material. PMID:2892300

  13. Glycophenotype Evaluation in Cutaneous Tumors Using Lectins Labeled with Acridinium Ester

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    Luiza Rayanna Amorim Lima

    2013-01-01

    Full Text Available Background. Tumor cells show alterations in their glycosylation patterns when compared to normal cells. Lectins can be used to evaluate these glycocode changes. Chemiluminescence assay is an effective technique for quantitative analysis of proteins, nucleic acids, and carbohydrates due to its high sensitivity, specificity, and rapid testing. Objective. To use histochemiluminescence based on lectin conjugated to acridinium ester (AE for the investigation of glycophenotype changes in cutaneous tumors. Methods. Concanavalin A (Con A, Peanut agglutinin (PNA, Ulex europaeus agglutinin-I (UEA-I, and Maackia amurensis agglutinin (MAA were conjugated to acridinium ester. Biopsies of cutaneous tumors and normal skin were incubated with the lectins-AE, and chemiluminescence was quantified and expressed as Relative Light Units (RLU. Results. Actinic keratosis (AK, keratoacanthoma (KA, squamous cell carcinoma (SCC, and basal cell carcinoma (BCC showed lower expression of α-D-glucose/mannose and α-L-fucose residues compared to normal tissue. Cutaneous tumors displayed higher expression of Gal-β(1-3-GalNAc residues than normal tissue. AK and SCC exhibited higher expression of Neu5Ac-α(2,3Gal residues than normal epidermis. KA and BCC showed equivalent RLU values compared to normal tissue. Conclusions. Lectin histochemiluminescence allowed quantitative assessment of the carbohydrate expression in cutaneous tissues, contributing to eliminate the subjectivity of conventional techniques used in the histopathological diagnosis.

  14. Fluorescence emission and polarization analyses for evaluating binding of ruthenium metalloglycocluster to lectin and tetanus toxin c-fragment

    Science.gov (United States)

    Okada, Tomoko; Minoura, Norihiko

    2010-02-01

    We have developed a fluorescent ruthenium metalloglycocluster as a powerful molecular probe for evaluating a binding event between carbohydrates and lectins by fluorescence emission (FE) and fluorescence polarization (FP) analysis. The fluorescent ruthenium metalloglycoclusters, [Ru(bpy-2Gal)3] and [Ru(bpy-2Glc)3], possess clustered galactose and glucose surrounding the ruthenium center. Changes in FE and FP of these metalloglycoclusters were measured by adding each lectin (Peanut agglutinin (PNA), Ricinus communis agglutinin 120 (RCA), Concanavalin A (ConA), or Wheat germ agglutinin (WGA)) or tetanus toxin c-fragment (TCF). Following the addition of PNA, the FE spectrum of [Ru(bpy- 2Gal)3] showed new emission peak and the FP value of [Ru(bpy-2Gal)3] increased. Similarly, the FE spectrum of [Ru(bpy-2Glc)3] showed new emission peak and the FP value increased following the addition of ConA. Since other combinations of the metalloglycoclusters and lectin caused little change, specific bindings of galactose to PNA and glucose to ConA were proved by the FE and FP measurement. From nonlinear least-squares fitting, dissociation constants (Kd) of [Ru(bpy-2Gal)3] to PNA was 6.1 μM, while the Kd values of [Ru(bpy)2(bpy-2Gal)] to PNA was ca. 10-4 M. Therefore, the clustered carbohydrates were proved to increase affinity to lectins. Furthermore, the FP measurements proved specific binding of [Ru(bpy-2Gal)3] to TCF.

  15. Lectin interactions with alpha-galactosylated xenoantigens

    DEFF Research Database (Denmark)

    Kirkeby, Svend; Moe, Dennis

    2002-01-01

    alpha-Galactosylated xenoantigens (Galalpha1-3Galbeta1-4GlcNAcbeta1 and Galalpha1-3Galbeta1-4GlcNAcbeta1-3Galbeta1-4Glc) are often detected with the alpha-Gal specific lectin Griffonia simplicifolia 1 isolectin B4 (GS1 B4). However, this lectin exhibits a broad and variable specificity for......-galactophilic lectins to alpha-galactosylated neoglycoproteins. The lectins were: Euonymus europaeus agglutinin (EEA), Griffonia simplicifolia 1 isolectin B4 (GS1 B4), Maclura pomifera agglutinin (MPA) and Pseudomonas aeruginosa agglutinin (PA-IL). Although both GS1 B4 and MPA strongly bound glycoconjugates terminating...... in Gal there seems to be some differentiation in their sugar binding preferences. MPA was the only lectin that showed high affinity for the pentasaccharide Galalpha1-3Galbeta1-4GlcNAcbeta1-3Galbeta1-4Glc and for the Galalpha-glycans on non-primate thyroglobulin. The length of the xenoantigenic...

  16. Fluorescence emission and polarization analyses for evaluating binding of ruthenium metalloglycoclusters to lectins and tetanus toxin C-fragment

    Science.gov (United States)

    Okada, Tomoko; Minoura, Norihiko

    2011-03-01

    We develop a fluorescent ruthenium metalloglycocluster for use as a powerful molecular probe in evaluating the binding between carbohydrates and lectins by fluorescence emission (FE) and fluorescence polarization (FP) analyses. Changes in the FE and FP of these metalloglycoclusters are measured following the addition of lectin [peanut agglutinin (PNA), Ricinus communis agglutinin 120, Concanavalin A (ConA), or wheat germ agglutinin] or tetanus toxin c-fragment (TCF). After the addition of PNA, the FE spectrum of [Ru(bpy-2Gal)3] shows a new emission peak and the FP value of [Ru(bpy-2Gal)3] increases. Similarly, the FE spectrum of [Ru(bpy-2Glc)3] shows a new emission peak and the FP value increases on addition of ConA. Because other combinations of metalloglycoclusters and lectins show little change, specific binding of galactose to PNA and that of glucose to ConA are confirmed by the FE and FP measurements. Resulting dissociation constants (Kd) prove that the metalloglycoclusters with highly clustered carbohydrates show higher affinity for the respective lectins than those with less clustered carbohydrates. Furthermore, specific binding of [Ru(bpy-2Gal)3] to TCF was confirmed by the FP measurement.

  17. Effect of Bu-Zhong-Yi-Qi-Tang on deficiency of N-glycan/nitric oxide and islet damage induced by streptozotocin in diabetic rats

    Institute of Scientific and Technical Information of China (English)

    Xiao-Qiu Liu; Ling Wu; Xue-Jun Guo

    2009-01-01

    AIM: To investigate the effect of Bu-Zhong-Yi-Qi-Tang (Decoction for Reinforcing Middle Jiao and Replenishing Qi) on deficiency of N-glycan/nitric oxide (NO) and islet damage induced by injecting two medium doses of streptozotocin (STZ). METHODS: Diabetes was induced by intraperitoneal injection of STZ at 55 mg/kg on day 1 and day 8. Islet damage was evaluated using a scoring system. Nitrite, nitrate, α-mannosidase and amylase activities were measured by colorimetry. N-glycan patterns of amylase were determined with lectin [ConA, pisum sativum agglutinin (PSA), peanut agglutinin (PNA), and lens culinaris agglutinin (LCA)] affinity precipitation method. RESULTS: Severe islet necrosis and mild islet atrophy were observed in diabetic rats. The number and size of islets, the activities of α-mannosidase, amylase and nitrite were decreased, while the binding of PNA and LCA to amylase was increased. All of which were improved after treatment with Bu-Zhong-Yi-Qi-Tang. Islet damage was significantly correlated with nitrite, nitrate, α-mannosidase, amylase and the binding of LCA, PNA, and PSA to amylase. PNA, and PSA to amylase.CONCLUSION: STZ- induced i s let damage i s related to N-glycan def iciency in proteins by blocking α-mannosidase activity and no deficiency, accumulation of unfolded proteins, and endoplasmic reticulum stress and activation of cellular signals, all of which are improved after treatment with Bu-Zhong-Yi- Qi-Tang.

  18. Targeting the Cryptococcus neoformans var. grubii Cell Wall Using Lectins: Study of the Carbohydrate-Binding Domain

    Directory of Open Access Journals (Sweden)

    Pamella de Brito Ximenes

    2015-02-01

    Full Text Available Cryptococcus neoformans var. grubii is considered to be the major cause of cryptococcosis in immunosuppressed patients. Understanding cell wall glycoproteins using lectins is of medical interest and can contribute to specific therapy. The aim of this study was to evaluate the carbohydrates on the cell wall of Cryptococcus neoformans var. grubii clinical isolates, using a fluorescein isothiocyanate-lectin binding protocol. Thirty yeast strains stocked in the culture collection were cultivated for 2 days at 30 °C with shaking. Cells were obtained by centrifugation, washed in phosphate-buffered saline, and a suspension of 107 cells/mL was obtained. To determine the binding profile of lectins, concanavalin A (Con A, wheat germ agglutinin (WGA, Ulex europaeus agglutinin I (UEA-I, and peanut agglutinin (PNA conjugated to fluorescein were used. All the tested clinical isolates of Cryptococcus neoformans var. grubii were intensely stained by WGA, moderately stained by Con A, and weakly stained by PNA and UEA-I. Thus, Cryptococcus can be detected in clinical specimens such as blood and cerebrospinal fluid using the fluorescent lectin WGA, which may be considered as an option for detection in cases of suspected cryptococcosis with low laboratory sensitivity. Future applications may be developed using this basic tool.

  19. Prevalence of Salmonella typhi among Patients in Abeokuta, South-Western Nigeria

    Directory of Open Access Journals (Sweden)

    I.O. Okonko

    2010-06-01

    Full Text Available This study reports on the prevalence of typhoid fever between genders among patients in Abeokuta, Nigeria. Typhoid fever caused by Salmonella typhi is an endemic disease in the tropic and sub-tropic and has become a major public health problem in developing countries of the world with an estimated annual incidence of 540 per 100,000. The annual incidence of typhoid is estimated to be about 17 million cases worldwide. It is often encountered in tropical countries including Nigeria where they constitute serious sources of morbidities and mortalities. Blood samples were collected from 840 apparently healthy people; 460 (54.8% females and 380 (45.2% males. The samples were examined for the presence and levels of Salmonella typhi antibodies by Widal agglutination technique. The standard Samonella ‘O’ and ‘H’ suspension (ANTEC diagnostic products were used as antigens. Of the 840 sera tested, agglutinins to Salmonella typhi were most prevalent in female subjects accounting for [426(92.6%] of the ‘H’ antigens and [322(70.0%] of ‘O’ antigens at the various dilutions while in the male subjects, [351(92.4%] accounts for the ‘O��� and [327(86.1%] for the ‘H’ antigens. There was a female preponderance (F/M 2:1. The levels of agglutinin of Salmonella paratyphi C-H [93(24.5% ] and Salmonella typhi C-O [112(29.5%] in the males were however, low. In the females, the low significant agglutinin titres for Salmonella typhi O and Salmonella paratyphi A-O were observed in 27.6% and 36.1% of the sera respectively. The results of this study showed that more males had Salmonella agglutinin titres for S. typhi O [351(92.4%] and S. typhi H [327(86.1%]. More so, 132 (34.7% males had Salmonella agglutinin titres for S. paratyphi A-O, 119 (31.3% for S. paratyphi B-O, 112 (29.5% for S. paratyphi C-O, 117 (31.0% for S. paratyphi A-H, 125 (33.0% for S. paratyphi B-H, and 93 (24.5% for S. paratyphi C-H. It also showed that more females had Salmonella

  20. Real-time cytotoxicity assay for rapid and sensitive detection of ricin from complex matrices.

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    Diana Pauly

    Full Text Available BACKGROUND: In the context of a potential bioterrorist attack sensitive and fast detection of functionally active toxins such as ricin from complex matrices is necessary to be able to start timely countermeasures. One of the functional detection methods currently available for ricin is the endpoint cytotoxicity assay, which suffers from a number of technical deficits. METHODOLOGY/FINDINGS: This work describes a novel online cytotoxicity assay for the detection of active ricin and Ricinus communis agglutinin, that is based on a real-time cell electronic sensing system and impedance measurement. Characteristic growth parameters of Vero cells were monitored online and used as standardized viability control. Upon incubation with toxin the cell status and the cytotoxic effect were visualized using a characteristic cell index-time profile. For ricin, tested in concentrations of 0.06 ng/mL or above, a concentration-dependent decrease of cell index correlating with cytotoxicity was recorded between 3.5 h and 60 h. For ricin, sensitive detection was determined after 24 h, with an IC50 of 0.4 ng/mL (for agglutinin, an IC50 of 30 ng/mL was observed. Using functionally blocking antibodies, the specificity for ricin and agglutinin was shown. For detection from complex matrices, ricin was spiked into several food matrices, and an IC50 ranging from 5.6 to 200 ng/mL was observed. Additionally, the assay proved to be useful in detecting active ricin in environmental sample materials, as shown for organic fertilizer containing R. communis material. CONCLUSIONS/SIGNIFICANCE: The cell-electrode impedance measurement provides a sensitive online detection method for biologically active cytotoxins such as ricin. As the cell status is monitored online, the assay can be standardized more efficiently than previous approaches based on endpoint measurement. More importantly, the real-time cytotoxicity assay provides a fast and easy tool to detect active ricin in complex

  1. Adenovirus-mediated expression of pig α(1, 3) galactosyltransferase reconstructs Gal α(1, 3) Gal epitope on the surface of human tumor cells

    Institute of Scientific and Technical Information of China (English)

    2001-01-01

    Gal α(1,3)Gal(gal epitope)is a carbohydrate epitope and synthesized in large amount by α(1,3)galactosyltransferase [α(1,3)GT] enzyme on the cells of lower mammalian animals such as pigs and mice.Human has no gal epitope due to the inactivation of α(1,3)GT gene but produces a large amount of antibodies(anti-Gal)which recognize Gal α(1,3)Gal structures specifically.In this study,a replicationdeficient recombinant adenoviral vector Ad5sGT containing pig α(1,3)GT cDNA was constructed and characterized.Adenoviral vector-mediated transfer of pig α(1,3)GT gene into human tumor cells such as malignant melanoma A375,stomach cancer SGC-7901,and lung cancer SPC-A-1 was reported for the first time.Results showed that Gal epitope did not increase the sensitivity of human tumor cells to human complement-mediated lysis,although human complement activation and the binding of human IgG and IgM natural antibodies to human tumor cells were enhanced significantly after Ad5sGT transduction.Appearance of gal epitope on the human tumor cells changed the expression of cell surface carbohydrates reacting with Ulex europaeus I(UEA I)lectins,Vicia villosa agglutinin(VVA),Arachis hypogaea agglutinin(PNA),and Glycine max agglutinin(SBA)to different degrees.In addition,no effect of gal epitope on the growth in vitro of human tumor cells was observed in MTT assay.

  2. Dietary plant lectins appear to be transported from the gut to gain access to and alter dopaminergic neurons of Caenorhabditis elegans, a potential etiology of Parkinson’s disease

    Directory of Open Access Journals (Sweden)

    Jolene eZheng

    2016-03-01

    Full Text Available Lectins from dietary plants have been shown to enhance drug absorption in the gastrointestinal tract of rats, be transported trans-synaptically as shown by tracing of axonal and dendritic paths, and enhance gene delivery. Other carbohydrate-binding protein toxins are known to traverse the gut intact in dogs. Post-feeding rhodamine- or TRITC-tagged dietary lectins, the lectins were tracked from gut to dopaminergic neurons (DAergic-N in transgenic Caenorhabditis elegans (C. elegans (egIs1[Pdat-1::GFP] where the mutant has the Green Fluorescent Protein (GFP gene fused to a dopamine transport protein gene labeling dopaminergic neurons, The lectins were supplemented along with the food organism Escherichia coli (OP50. Among nine tested rhodamine/TRITC-tagged lectins, four, including Phaseolus vulgaris erythroagglutinin (PHA-E, Bandeiraea simplicifolia (BS-I, Dolichos biflorus agglutinin (DBA, and Arachis hypogaea (PNA, appeared to be transported from gut to the GFP-DAergic-N. Griffonia Simplicifolia (GSL-I and PHA-E, reduced the number of GFP-DAergic-N suggesting a toxic activity. PHA-E, BS-I, Pisum Sativum (PSA, and Triticum vulgaris agglutinin (Succinylated reduced fluorescent intensity of GFP-DAergic-N. PHA-E, PSA, Concanavalin A, and Triticum vulgaris agglutinin decreased the size of GFP-DAergic-N, while BS-I increased neuron size. These observations suggest that dietary plant lectins are transported to and affect DAergic-N in C. elegans, which support Braak and Hawkes’ hypothesis, suggesting one alternate potential dietary etiology of Parkinson’s disease (PD. A recent Danish study showed that vagotomy resulted in 40% lower incidence of PD over 20 years. Differences in inherited sugar structures of gut and neuronal cell surfaces may make some individuals more susceptible in this conceptual disease etiology model.

  3. Lectin binding and effects in culture on human cancer and non-cancer cell lines: examination of issues of interest in drug design strategies.

    Science.gov (United States)

    Petrossian, Karineh; Banner, Lisa R; Oppenheimer, Steven B

    2007-01-01

    By using a non-cancer and a cancer cell line originally from the same tissue (colon), coupled with testing lectins for cell binding and for their effects on these cell lines in culture, this study describes a simple multi-parameter approach that has revealed some interesting results that could be useful in drug development strategies. Two human cell lines, CCL-220/Colo320DM (human colon cancer cells, tumorigenic in nude mice) and CRL-1459/CCD-18Co (non-malignant human colon cells) were tested for their ability to bind to agarose microbeads derivatized with two lectins, peanut agglutinin (Arachis hypogaea agglutinin, PNA) and Dolichos biflorus agglutinin (DBA), and the effects of these lectins were assessed in culture using the MTT assay. Both cell lines bound to DBA-derivatized microbeads, and binding was inhibited by N-acetyl-D-galactosamine, but not by L-fucose. Neither cell line bound to PNA-derivatized microbeads. Despite the lack of lectin binding using the rapid microbead method, PNA was mitogenic in culture at some time points and its mitogenic effect displayed a reverse-dose response. This was also seen with effects of DBA on cells in culture. While this is a simple study, the results were statistically highly significant and suggest that: (1) agents may not need to bind strongly to cells to exert biological effects, (2) cell line pairs derived from diseased and non-diseased tissue can provide useful comparative data on potential drug effects and (3) very low concentrations of potential drugs might be initially tested experimentally because reverse-dose responses should be considered. PMID:17706752

  4. Isolation and partial characterization of a D-galactose-binding lectin from the latex of Synadenium carinatum

    OpenAIRE

    Maria Aparecida de Souza; Francielle Amâncio-Pereira; Cristina Ribeiro de Barros Cardoso; Adriano Gomes da Silva; Edmar Gomes Silva; Lívia Resende Andrade; Janethe Deolina Oliveira Pena; Henrique Lanza; Sandra Regina Afonso-Cardoso

    2005-01-01

    A lectin from the latex of Synadenium carinatum was purified by affinity chromatography on immobilized-D-galactose-agarose and shown to be a potent agglutinin of human erythrocytes. The haemagglutination of human red cells was inhibited by 3.0 mM N-acetyl-D-galactopyranoside, 6.3 mM methyl-beta-D-galactopyranoside, 50 mM methyl-alpha-D-galactopyranoside and 50 mM D-fucose but not by L-fucose, demonstrating an anomeric and a conformational specificity. According to SDS-PAGE analysis, the lecti...

  5. Effect of lectins on mouse peritoneal macrophage phagocytic activity.

    Science.gov (United States)

    Maldonado, G; Porras, F; Fernández, L; Vázquez, L; Zenteno, E

    1994-11-01

    We studied the in vitro ability of lectin-treated murine peritoneal macrophages to attach and phagocytize particulate antigens. Glucose and mannose specific lectins such as Con-A and lentil lectin, as well as complex lactosamine residues specific lectins, such as Phaseolus vulgaris var. cacahuate and Phaseolus coccineus var. alubia, increased the macrophage phagocytic activity towards heterologous erythrocytes, whereas peanut agglutinin, a galactose-specific lectin, diminished the macrophage phagocytic activity. These results suggest that a galactose-N-acetyl-D galactosamine-containing structure could participate as negative modulator of the phagocytic activity. PMID:7851961

  6. Display of wasp venom allergens on the cell surface of Saccharomyces cerevisiae

    DEFF Research Database (Denmark)

    Borodina, Irina; Jensen, B. M.; Søndergaard, Ib;

    2010-01-01

    Saccharomyces cerevisiae preserve their native allergenic properties and whether the yeast native surface glycoproteins interfere with IgE binding. We chose to use the major allergens from the common wasp Vespula vulgaris venom: phospholipase A1, hyaluronidase and antigen 5 as the model. Results: The proteins...... were expressed on the surface as fusions with a-agglutinin complex protein AGA2. The expression was confirmed by fluorescent cytometry (FACS) after staining the cells with antibody against a C-tag attached to the C-terminal end of the allergens. Phospholipase A1 and hyaluronidase retained their...

  7. Functional activities of the Tsh protein from avian pathogenic Escherichia coli (APEC) strains

    OpenAIRE

    Renata K. T. Kobayashi; Gaziri, Luis Carlos J.; Vidotto, Marilda C.

    2010-01-01

    The temperature-sensitive hemagglutinin (Tsh) expressed by strains of avian pathogenic Escherichia (E.) coli (APEC) has both agglutinin and protease activities. Tsh is synthesized as a 140 kDa precursor protein, whose processing results in a 106 kDa passenger domain (Tshs) and a 33 kDa β-domain (Tshβ). In this study, both recombinant Tsh (rTsh) and supernatants from APEC, which contain Tshs (106 kDa), caused proteolysis of chicken tracheal mucin. Both rTsh (140 kDa) and pellets from wild-type...

  8. INTERACTION BETWEEN THE SURFACE GLYCOSYLATED POLYPROPYLENE MEMBRANE AND LECTIN

    Institute of Scientific and Technical Information of China (English)

    Qian Yang; Ling-shu Wan; Zhi-kang Xu

    2008-01-01

    A glycopolymer bearing glucose residues was tethered onto the surface of polypropylene microporous membrane by UV-induced graft polymerization of α-allyl glucoside. Concanavalin A (Con A), a glucose recognizing lectin, could be specifically adsorbed to the membrane surface. On the other hand, the membrane surface showed no recognition ability to another lectin peanut agglutinin. Moreover, the recognition complex between the glycosylated membrane surface and Con Acould be inhibited by glucose and mannose solution. This surface glycosylated membrane could be used as affinity membrane for protein separation and purification.

  9. STUDY ON GLYCOCONJUGATE CHANGES ON CELL SURFACE IN PROGRESSIVE DEVELOPMENT OF PULMONARY TUMOR

    Institute of Scientific and Technical Information of China (English)

    WANG Xiao-mei; SHAN Jun; CHEN Zhuo-huai

    2002-01-01

    Aim: To investigate glycoconjugate changes on the cell surface of proliferative lesions and neoplasms of mice lungs at various stages of tumorigenesis, the relation between progressive development of mouse pulmonary tumors and expression of cell surface saccharide. Materials and methods: Thirty - one male A/J strain mice at 5 weeks of age were treated intraperitoneally with a single injection of 20 - methylcholanthrene (20 - MC), 292 pulmonary lesions including 31 hyperplasias, 145 alveolar adenomas, 61 papillary adenomas, 55 papillary adenocarcinomas and their combined type were obtained. The binding affinities of cells in normal respiratory epithelia and in proliferative lesions to four peroxidases - conjugated lectins, Maclura pomifera agglutinin (MPA), Arachis hypogea agglutinin (PNA), Ricinus communis agglutinin (RCA), and wheat germ agglutinin (WGA) were examined. Results: Cells of hyperplasia and alveolar adenoma showed fairly strong affinity to all the four lectins. However, part of papillary adenoma cells and greater part of papillary adenocarcinoma cells lost their binding affinity to MPA, PNA, and RCA, but not to WGA. The bindings of MPA, PNA and RNA were detected predominently on the luminal surfaces of benign tumors but not on the luminal surfaces of malignant tumors. WGA might bind to varied types of benign and malignant tumors. Pretreated with neuraminidase, the lesions enhanced the staining intensity for the four lectins, the binding sites of WGA to malignant tumor cells were numerous. A distinct difference in lectin binding affinity between hyperplasia / alveolar adenoma/papillary adenoma and papillary adenocarcinoma was clearly shown( x2 = 46.89, P < 0.01, x2 = 36.77, P < 0.01 and x2 = 52.87, P < 0.01 ) in this experiment. The complex glycoconjugates on the cell surface of malignant and benign lesions during the development of pulmonary tumor were changed,malignant tumor cells differed from the surface of benign tumor cells, the levels of

  10. Brain alpha-dystroglycan displays unique glycoepitopes and preferential binding to laminin-10/11.

    Science.gov (United States)

    McDearmon, Erin L; Combs, Ariana C; Sekiguchi, Kiyotoshi; Fujiwara, Hironobu; Ervasti, James M

    2006-06-12

    alpha-Dystroglycan was quantitatively enriched from mammalian brain based on its uniform reactivity with Vicia villosa agglutinin and resolved into sub-populations possessing or lacking the sulfated glucuronic acid epitope recognized by monoclonal antibody HNK-1. We generated a new monoclonal antibody specific for a glycoepitope on brain alpha-dystroglycan but absent from alpha-dystroglycan expressed in all other tissues examined. Finally, we found that laminin-10/11 preferentially bound to brain alpha-dystroglycan compared to skeletal muscle alpha-dystroglycan. Our results suggest that tissue-specific glycosylation modifies the laminin binding specificity of alpha-dystroglycan. PMID:16709410

  11. Synthesis of amides and sulfonamides of β-D- galactopyranosylamine and β-lactosylamine and evaluation of their interactions with the lectins from Erythrina cristagalli and Ricinus communis

    International Nuclear Information System (INIS)

    We report herein the synthesis of some β-D-galactopyranosylamine and β-lactosylamine amides and sulfonamides. The interactions of these compounds with lectins from the seeds of Erythrina cristagalli (LEC) and Ricinus communis (RCA120) were evaluated in a hemagglutination inhibitory activity assay. D-Galactose and lactose were used as reference compounds. The β-lactosylamine amides and sulfonamides were nearly as active as lactose in inhibiting LEC mediated hemagglutination and were less active against RCA120 agglutinin. The β-D-galactopyranosylamine amides and sulfonamides were, with one exception, considerably less active than D-galactose in the assay with both lectins. (author)

  12. [Seroepidemiologic survey of leptospirosis among environmental sanitation workers in an urban locality in the southern region of Brazil].

    Science.gov (United States)

    de Almeida, L P; Martins, L F; Brod, C S; Germano, P M

    1994-02-01

    Sera from 386 environmental sanitation workers, concerned with water supply, drains and drainage galleries, sewers, garbage collection and road sweepers, were examined for leptospiral agglutinins by the microscopic agglutination test. Altogether 40 of the 386 workers (10.4%) were positive to one or more serovars; however, the difference in seropositivity between the professional categories was not significant (p 0.05). Of the seropositive workers, 86.9% had agglutination titres > or = 100 and < or = 400; the rates for titres 100 and 400 were higher than 800, 1,600 and 3,200 (p < 0.05). PMID:7997826

  13. Cytostructural Localization of a Tumor-Associated Antigen

    Science.gov (United States)

    Howard, Donald R.; Batsakis, John G.

    1980-10-01

    Tumor cell membrane glycoproteins may be involved in the induction of tumor immunity or in the escape of tumors from immunologic defense mechanisms. Forty-four benign and malignant breast lesions were examined for the presence of a carbohydrate precursor antigen (T antigen) of the human blood group system MN. T antigen was demonstrated by means of an immunohistochemical technique to detect tissue binding of peanut agglutinin, a plant lectin, with affinity for T antigen. Malignant breast lesions showed a pattern of T antigen expression different from that of benign breast tissues. A possible role for T antigen in the modulation of the immune response to breast carcinoma is suggested.

  14. Enhancement of bone marrow allografts from nude mice into mismatched recipients by T cells void of graft-versus-host activity.

    OpenAIRE

    Lapidot, T; Lubin, I; Terenzi, A; Faktorowich, Y; Erlich, P; Reisner, Y

    1990-01-01

    Transplantation of 8 x 10(6) C57BL/6-Nu+/Nu+ (nude) bone marrow cells into C3H/HeJ recipients after conditioning with 8 Gy of total body irradiation has resulted in a markedly higher rate of graft rejection or graft failure compared to that found in recipients of normal C57BL/6 or C57BL/6-Bg+/Bg+ (beige) T-cell-depleted bone marrow. Mixing experiments using different numbers of nude bone marrow cells with or without mature thymocytes (unagglutinated by peanut agglutinin) revealed that engraft...

  15. Altered glycosylation, expression of serum haptoglobin and alpha-1-antitrypsin in chronic hepatitis C, hepatitis C induced liver cirrhosis and hepatocellular carcinoma patients.

    Science.gov (United States)

    Mondal, Gautam; Saroha, Ashish; Bose, Partha Pratim; Chatterjee, B P

    2016-04-01

    Liver cirrhosis with hepatitis C viral infection (HCV-LC) causes high risk to develop hepatocellular carcinoma (HCC). Besides diagnosis of liver cirrhosis by biochemical test, imaging techniques, assessment of structural liver damage by biopsy shows several disadvantages. Our aim was to monitor the changes in the expression level of serum proteins and their glycosylation pattern among chronic hepatitis C (HCV-CH), HCV-LC and HCC patients with respect to controls. 2D gel electrophoresis of HCV-CH, HCV-LC and HCC patients' sera showed several protein spots, which were identified by LC-MS. The change in the expression of two prominent protein spots, haptoglobin (Hp) and alpha 1-antitrypsin (AAT) was evaluated by western blot and ELISA. The changes in glycosylation pattern of these serum proteins were assayed using different lectins. Increased level of Hp and AAT was observed in HCV-LC and HCC patients' group whereas those were found to be present less in HCV-CH patient groups with respect to control as determined by ELISA using monoclonal antibodies. Decreased level of sialylation in both Hp and AAT was observed in HCV-LC and HCV-CH patients' group whereas increased level of sialylation was observed in HCC patient groups by ELISA using Sambucus nigra agglutinin. On the other hand increased level of fucosylation in two serum glycoproteins was observed in HCV-LC and HCC patients' group using Lens culinarris agglutinin. High glycan branching was found in HCV-LC and HCC patient groups in Hp but not in HCV-CH as determined by Datura stramonium agglutinin. However, there was no such change observed in glycan branching in AAT of HCV-CH and HCV-LC patients' groups, to the contrary high glycan branching was observed in HCC patients' group. Increased level of exposed galactose in both serum proteins was observed in both HCC patients' group as determined by Ricinus communis agglutinin. The present glycoproteomics study could predict the progression of HCV-CH, HCV-LC and HCC

  16. Mouse whey acidic protein is a novel member of the family of 'four-disulfide core' proteins.

    OpenAIRE

    Hennighausen, L G; Sippel, A E

    1982-01-01

    Unlike in other mammalian species, the major whey protein in mouse is not alpha-lactalbumin, but a cysteine rich, acidic protein with a molecular weight of 14.0 kDa. We have deduced the amino acid sequence of this mouse acidic of whey protein from the nucleotide sequence of cloned cDNA. The positions of the half cysteines suggest that mouse whey acidic protein (WAP) is a two domain protein, very similar in structure to the plant lectin wheat germ agglutinin and the hypothalamic carrier protei...

  17. Gum and Nose Bleeding as a Presentation of Pediatric Brucellosis

    Directory of Open Access Journals (Sweden)

    Hosseininasab

    2016-01-01

    Full Text Available Introduction Brucellosis symptoms are nonspecific; the most common complaints include fever, sweats, anorexia, headache, malaise, and arthralgia. Hematological manifestations of active brucellosis vary from mild anemia and leukopenia to thrombocytopenia and rarely pancytopenia. Case Presentation We report on an eight-year-old boy who presented epistaxis and gum bleeding. The physical examination revealed petechiae, purpura, ecchymosis, and cervical while inguinal lymphadenopathy and splenomegaly were noted. Brucella agglutinin titer was positive. After five days of specific therapy for brucellosis, fever was controlled, clinical signs and symptoms were improved and platelet count was dramatically increased. Conclusions Sever thrombocytopenia and bleeding may be the presentation of brucellosis.

  18. Investigation of the response to the enterobacterial common antigen after typhoid vaccination

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    Arlete M. Milhomem

    1987-03-01

    Full Text Available Antibodies against the Salmonella typhi enterobacterial common antigen (ECA and the O and H antigens were investigated in sera from healthy male subjects who had been previously vaccinated with the typhoid vaccine. No serological response to ECA was observed. Sera from subjects not previously vaccinated presented titers of ECA hemagglutinins which quantitatively were related to the presence ofH titers, but not to O agglutinins but with no statistical significance. The results are discussed in relation to the possible protective immunological mechanisms in typhoid fever.

  19. Langerhans Cell Histiocytosis of the Temporal Bone.

    Science.gov (United States)

    Ginat, Daniel Thomas; Johnson, Daniel N; Cipriani, Nicole A

    2016-06-01

    Langerhans cell histiocytosis involving the temporal bone region is uncommon and can resemble malignant neoplasms on imaging due to high cellularity. Although recognizing the presence of sharp margins with beveled-edges can be helpful, tissue sampling is often necessary for confirming the diagnosis. Cytology classically demonstrates kidney-bean shaped nuclei within the Langerhans cells and immunohistochemical staining is positive for S-100, peanut agglutinin (PNA), MHC class II, CD1a, and Langerin (CD 207). These features are exemplified in this sine qua non radiology-pathology correlation article. PMID:25903273

  20. Equestrian perniosis: a report of 2 cases and a review of the literature.

    Science.gov (United States)

    Stewart, Campbell L; Adler, Donald J; Jacobson, Abby; Brod, Bruce A; Shinohara, Michi M; Seykora, John T; Elenitsas, Rosalie; Rosenbach, Misha

    2013-04-01

    Equestrian perniosis (EP) is a rare condition in which patients develop tender burning nodular plaques on their bilateral thighs after riding in the cold. These lesions tend to resolve rapidly with minimal exposure to cold, and wearing loose, layered warm clothing. Unlike acral perniosis, EP has no known systemic disease associations, although 2 reported cases did have elevated cold agglutinins. The histology of this disease is similar to perniosis; however, EP is distinct in that the perivascular lymphocytic infiltrate prominently involves the fat. In this case report, we discuss the clinical and histological findings in 2 cases of EP, including the first documented in a man. PMID:22534636

  1. TOXIC SUBSTANCE PRESENT IN THE OIL FRACTION OF THE SAGABEAN

    Directory of Open Access Journals (Sweden)

    Oey Kam Nio

    2012-09-01

    Full Text Available Penelitian sebelumnya menunjukkan adanya suatu zat toksis pada biji saga yang tidak tergolong faktor "anti-nutritive" seperti "tripsin inhibitors", "phyto-agglutinins"dan "saponins". Ternyata bahwa zat toksis tersebut yang belum dapat diekstraksi dan diidentifikasi, berada di dalam fraksi minyak biji saga. Di samping adanya zat toksis, ditemukan pub bahwa kualitas protein biji saga lebih rendah dari­pada kacang kedele. Hasil analisa asam amino menunjukkan bahwa kadar methionine dan threonine ada­lah terbatas (limiting, apabila ditambah dengan kedua asam amino ini dalam jumlah yang cukup, kuali­tas proteinnya menjadi sama tingginya dengan kacang kedele rebus tanpa tambahan.

  2. Design of a covalently bonded glycosphingolipid microarray

    DEFF Research Database (Denmark)

    Arigi, Emma; Blixt, Klas Ola; Buschard, Karsten;

    2012-01-01

    agglutinin, a monoclonal antibody to sulfatide, Sulph 1; and a polyclonal antiserum reactive to asialo-G(M2)). Preliminary evaluation of the method indicated successful immobilization of the GSLs, and selective binding of test probes. The potential utility of this methodology for designing covalent...... 2-mercaptoethylamine, was also tested. Underivatized or linker-derivatized lyso-GSL were then immobilized on N-hydroxysuccinimide- or epoxide-activated glass microarray slides and probed with carbohydrate binding proteins of known or partially known specificities (i.e., cholera toxin B-chain; peanut...

  3. Rapid reconstitution of a transmembrane protein into supported planar lipid membranes.

    Science.gov (United States)

    Nakanishi, M

    1984-10-29

    A procedure for reconstituting a transmembrane protein by the freeze-thaw method into supported planar lipid layers has been developed. A solution containing human glycophorin A was introduced between an alkylated cover glass with lipid layers from soybean phospholipids and a bare glass slide, and was then put in a glass dish which was frozen outside by liquid nitrogen. The lipid layer membranes prepared in this manner have been examined by the binding of both macrophages and wheat germ agglutinin agarose. Macrophages bound more efficiently to the membranes bearing glycophorin A and spread more rapidly than those of the control membranes. PMID:6548452

  4. Direct production of ethanol from raw corn starch via fermentation by use of a novel surface-engineered yeast strain codisplaying glucoamylase and alpha-amylase.

    Science.gov (United States)

    Shigechi, Hisayori; Koh, Jun; Fujita, Yasuya; Matsumoto, Takeshi; Bito, Yohei; Ueda, Mitsuyoshi; Satoh, Eiichi; Fukuda, Hideki; Kondo, Akihiko

    2004-08-01

    Direct and efficient production of ethanol by fermentation from raw corn starch was achieved by using the yeast Saccharomyces cerevisiae codisplaying Rhizopus oryzae glucoamylase and Streptococcus bovis alpha-amylase by using the C-terminal-half region of alpha-agglutinin and the flocculation functional domain of Flo1p as the respective anchor proteins. In 72-h fermentation, this strain produced 61.8 g of ethanol/liter, with 86.5% of theoretical yield from raw corn starch. PMID:15294847

  5. Lectin typing of Campylobacter jejuni using a novel quartz crystal microbalance technique

    Energy Technology Data Exchange (ETDEWEB)

    Yakovleva, Maria E., E-mail: maria.yakovleva@gmail.com [Department of Infectious Diseases and Medical Microbiology, Lund University, 223 62 Lund (Sweden); Moran, Anthony P. [Department of Microbiology, School of Natural Sciences, National University of Ireland, Galway (Ireland); Safina, Gulnara R. [Department of Analytical and Marine Chemistry, University of Gothenburg, 412 96 Gothenburg (Sweden); Wadstroem, Torkel [Department of Infectious Diseases and Medical Microbiology, Lund University, 223 62 Lund (Sweden); Danielsson, Bengt [Acromed Invest AB, Magistratsvaegen 10, 226 43 Lund (Sweden)

    2011-05-23

    Seven Campylobacter jejuni strains were characterised by a lectin typing assay. The typing system was based on a quartz crystal microbalance technique (QCM) with four commercially available lectins (wheat germ agglutinin, Maackia amurensis lectin, Lens culinaris agglutinin, and Concanavalin A), which were chosen for their differing carbohydrate specificities. Initially, the gold surfaces of the quartz crystals were modified with 11-mercaptoundecanoic acid followed by lectin immobilisation using a conventional amine-coupling technique. Bacterial cells were applied for lectin typing without preliminary treatment, and resonant frequency and dissipation responses were recorded. The adhesion of microorganisms on lectin surfaces was confirmed by atomic force microscopy. Scanning was performed in the tapping mode and the presence of bacteria on lectin-coated surfaces was successfully demonstrated. A significant difference in the dissipation response was observed for different C. jejuni strains which made it possible to use this parameter for discriminating between bacterial strains. In summary, the QCM technique proved a powerful tool for the recognition and discrimination of C. jejuni strains. The approach may also prove applicable to strain discrimination of other bacterial species, particularly pathogens.

  6. Assessment of lectin inactivation by heat and digestion.

    Science.gov (United States)

    Pusztai, A; Grant, G

    1998-01-01

    Proteins/glycoproteins from plants, particularly lectins, are more resistant to heat denaturation than animal proteins (1, 2). With legume seeds, whose lectin content is appreciable, this presents potentially serious problems in nutritional practice. Therefore, before they can be used safely, legume-based food/ feeds usually require thorough and expensive heat processing to inactivate antinutritive components. Indeed, dry or moist heating of seeds at 70°C for several h has little or no effect on their lectin activity (Fig. 1) and treatment at much higher temperatures is needed to inactivate the biological and antinutritional effects of legume lectins (1, 2). The safety aspect is even more serious with some monocot lectins, such as wheatgerm agglutinin or a number of oilseed lectins, such as peanut agglutinin and many others because they are extremely heat stable and normal cooking or other conventional heat treatments may fail to inactivate them (3) Thus, the best way to avoid potential harmful effects of these heat-resistant lectins is to limit their dietary intake to a minimum. Fig. 1. Loss of lectin activity during aqueous heat treatment of soybean at various temperatures. PMID:21374488

  7. Density variant glycan microarray for evaluating cross-linking of mucin-like glycoconjugates by lectins.

    Science.gov (United States)

    Godula, Kamil; Bertozzi, Carolyn R

    2012-09-26

    Interactions of mucin glycoproteins with cognate receptors are dictated by the structures and spatial organization of glycans that decorate the mucin polypeptide backbone. The glycan-binding proteins, or lectins, that interact with mucins are often oligomeric receptors with multiple ligand binding domains. In this work, we employed a microarray platform comprising synthetic glycopolymers that emulate natural mucins arrayed at different surface densities to evaluate how glycan valency and spatial separation affect the preferential binding mode of a particular lectin. We evaluated a panel of four lectins (Soybean agglutinin (SBA), Wisteria floribunda lectin (WFL), Vicia villosa-B-4 agglutinin (VVA), and Helix pomatia agglutin (HPA)) with specificity for α-N-acetylgalactosamine (α-GalNAc), an epitope displayed on mucins overexpressed in many adenocarcinomas. While these lectins possess the ability to agglutinate A(1)-blood cells carrying the α-GalNAc epitope and cross-link low valency glycoconjugates, only SBA showed a tendency to form intermolecular cross-links among the arrayed polyvalent mucin mimetics. These results suggest that glycopolymer microarrays can reveal discrete higher-order binding preferences beyond the recognition of individual glycan epitopes. Our findings indicate that glycan valency can set thresholds for cross-linking by lectins. More broadly, well-defined synthetic glycopolymers enable the integration of glycoconjugate structural and spatial diversity in a single microarray screening platform. PMID:22967056

  8. Binucleate trophoblast giant cells in the water buffalo (Bubalus bubalis) placenta.

    Science.gov (United States)

    Carvalho, A F; Klisch, K; Miglino, M A; Pereira, F T V; Bevilacqua, E

    2006-01-01

    The binucleate trophoblast giant cells (BNC) of the water buffalo, Bubalus bubalis, placenta were studied, with emphasis on the synthesis of BNC-specific proteins. Placentomal tissues of 27 water buffalos (2-10 months of pregnancy) were processed for light and electron microscopy. The frequency of BNCs was 20% of the trophoblastic cells in 2-3-month placentas and increased to 27% in the later stages. Ultrastructurally, binucleate cells displayed a prominent granular endoplasmic reticulum and Golgi apparatus, typical of cells involved with protein synthesis and exportation. The buffalo BNCs contained periodic acid-Schiff (PAS)-positive granules and reacted with antisera against bovine placental lactogen, prolactin-related protein-I, and pregnancy-associated glycoproteins. Lectin histochemistry with Dolichos biflorus agglutinin, Vicia villosa agglutinin, and Phaseolus vulgaris leucoagglutinin showed specific staining of BNCs. Different stages of BNC migration and fusion with uterine epithelial cells were observed. Trinucleate feto-maternal hybrid cells were the typical outcome of cell fusions. These cells underwent degeneration, with typical morphological features of apoptosis. The results revealed a strong homology between water buffalo and cattle BNCs concerning cell morphology, protein expression, glycosylation pattern, and characteristics of cell migration and fusion. PMID:16240388

  9. The binucleate cell of okapi and giraffe placenta shows distinctive glycosylation compared with other ruminants: a lectin histochemical study.

    Science.gov (United States)

    Jones, Carolyn J P; Wilsher, Sandra A; Wooding, F B P; Benirschke, K; Allen, W R

    2015-02-01

    The placenta of ruminants contains characteristic binucleate cells (BNC) with a highly conserved glycan structure which evolved early in Ruminant phylogenesis. Giraffe and Okapi placentae also contain these cells and it is not known whether they have a similar glycan array. We have used lectin histochemistry to examine the glycosylation of these cells in these species and compare them with bovine BNC which have a typical ruminant glycan composition. Two placentae, mid and near term, from Giraffe (Giraffa camelopardalis) and two term placenta of Okapi (Okapia johnstoni) were embedded in resin and stained with a panel of 23 lectins and compared with near-term bovine (Bos taurus) placenta. Significant differences were found in the glycans of Giraffe and Okapi BNC compared with those from the bovine, with little or no expression of terminal αN-acetylgalactosamine bound by Dolichos biflorus and Vicia villosa agglutinins which instead bound to placental blood vessels. Higher levels of N-acetylglucosamine bound by Lycopersicon esculentum and Phytolacca americana agglutinins were also apparent. Some differences between Okapi and Giraffe were evident. Most N-linked glycans were similarly expressed in all three species as were fucosyl residues. Interplacentomal areas in Giraffe and Bovine showed differences from the placentomal cells though no intercotyledonary BNC were apparent in Okapi. In conclusion, Giraffidae BNC developed different glycan biosynthetic pathways following their split from the Bovidae with further differences evolving as Okapi and Giraffe diverged from each other, affecting both inter and placentomal BNC which may have different functions during development. PMID:25527317

  10. Microarrays with varying carbohydrate density reveal distinct subpopulations of serum antibodies.

    Science.gov (United States)

    Oyelaran, Oyindasola; Li, Qian; Farnsworth, David; Gildersleeve, Jeffrey C

    2009-07-01

    Antigen arrays have become important tools for profiling complex mixtures of proteins such as serum antibodies. These arrays can be used to better understand immune responses, discover new biomarkers, and guide the development of vaccines. Nevertheless, they are not perfect and improved array designs would enhance the information derived from this technology. In this study, we describe and evaluate a strategy for varying antigen density on an array and then use the array to study binding of lectins, monoclonal antibodies, and serum antibodies. To vary density, neoglycoproteins containing differing amounts of carbohydrate were synthesized and used to make a carbohydrate microarray with variations in both structure and density. We demonstrate that this method provides variations in density on the array surface within a range that is relevant for biological recognition events. The array was used to evaluate density dependent binding properties of three lectins (Vicia villosa lectin B4, Helix pomatia agglutinin, and soybean agglutinin) and three monoclonal antibodies (HBTn-1, B1.1, and Bric111) that bind the tumor-associated Tn antigen. In addition, serum antibodies were profiled from 30 healthy donors. The results show that variations in antigen density are required to detect the full spectrum of antibodies that bind a particular antigen and can be used to reveal differences in antibody populations between individuals that are not detectable using a single antigen density. PMID:19366269

  11. Distribution patterns of influenza virus receptors and viral attachment patterns in the respiratory and intestinal tracts of seven avian species

    Directory of Open Access Journals (Sweden)

    Costa Taiana

    2012-04-01

    Full Text Available Abstract This study assessed the presence of sialic acid α-2,3 and α-2,6 linked glycan receptors in seven avian species. The respiratory and intestinal tracts of the chicken, common quail, red-legged partridge, turkey, golden pheasant, ostrich, and mallard were tested by means of lectin histochemistry, using the lectins Maackia amurensis agglutinin II and Sambucus nigra agglutinin, which show affinity for α-2,3 and α-2,6 receptors, respectively. Additionally, the pattern of virus attachment (PVA was evaluated with virus histochemistry, using an avian-origin H4N5 virus and a human-origin seasonal H1N1 virus. There was a great variation of receptor distribution among the tissues and avian species studied. Both α-2,3 and α-2,6 receptors were present in the respiratory and intestinal tracts of the chicken, common quail, red-legged partridge, turkey, and golden pheasant. In ostriches, the expression of the receptor was basically restricted to α-2,3 in both the respiratory and intestinal tracts and in mallards the α-2,6 receptors were absent from the intestinal tract. The results obtained with the lectin histochemistry were, in general, in agreement with the PVA. The differential expression and distribution of α-2,3 and α-2,6 receptors among various avian species might reflect a potentially decisive factor in the emergence of new viral strains.

  12. Glycan-mediated uptake in urothelial primary cells: Perspectives for improved intravesical drug delivery in urinary tract infections.

    Science.gov (United States)

    Pichl, Clara Maria; Feilhauer, Sophie; Schwaigerlehner, Rose-Marie; Gabor, Franz; Wirth, Michael; Neutsch, Lukas

    2015-11-30

    Urinary tract infections (UTIs) are among the most common bacterial infections. Despite a wide range of therapeutic options, treatment success is compromised by multiresistance and the efficient mechanism of tissue colonization of uropathogenic Escherichia coli (UPEC). In advanced drug delivery systems, a similar, glycan-mediated targeting mechanism may be realized by conjugating the drug to a plant lectin. This may lead to the drug being more efficiently accumulated at the desired site of action, the bacterial reservoirs. In this study, we aimed at elucidating the potential of this biorecognitive approach. Glycan-triggered interaction cascades and uptake processes of several plant lectins with distinct carbohydrate specificities were characterized using single cells and monolayer culture. Due to pronounced cytoadhesive and cytoinvasive properties, wheat germ agglutinin (WGA) emerged as a promising targeter in porcine urothelial primary cells. The lectin-cell interaction proved highly stabile in artificial urine, simulating the conditions in actual application. Colocalisation studies with internalized WGA and lens culinaris agglutinin (LCA) revealed that intracellular accumulation sites were largely identical for GlcNAc- and Mannose-specific lectins. This indicates that WGA-mediated delivery may indeed constitute a potent tool to reach bacteria taken up via a FimH-triggered invasion process. Existing pitfalls in intravesical treatment schedules may soon be overcome. PMID:26383837

  13. Local cholinergic and non-cholinergic neural pathways to the rat supraoptic nucleus

    International Nuclear Information System (INIS)

    An estimated two thirds of the input to the supraoptic nucleus of the rat hypothalamus (SON) including a functionally significant cholinergic innervation, arise from local sources of unknown origin. The sources of these inputs were identified utilizing Golgi-Cox, retrograde tracing, choline acetyltransferase immunocytochemistry and anterograde tracing methodologies. Multipolar Golgi impregnated neurons located dorsal and lateral to the SON extend spiney processes into the nucleus. Injections of the retrograde tracers, wheat germ agglutinin or wheat germ agglutinin-horseradish peroxidase, into the SON labeled cells bilaterally in the arcuate nucleus, and ipsilaterally in the lateral hypothalamus, anterior hypothalamus, nucleus of the diagonal band, subfornical organ, medial preoptic area, lateral preoptic area and in the region dorsolateral to the nucleus. Immunocytochemistry for choline acetyltransferase revealed cells within the ventro-caudal portion of cholinergic cell group, Ch4, which cluster dorsolateral to the SON, and extend axon- and dendrite-like processes into the SON. Cells double-labeled by choline acetyltransferase immunocytochemistry and retrograde tracer injections into the SON are localized within the same cholinergic cell group dorsolateral to the SON. Injections of the anterograde tracer, Phaseolus vulgaris-leucoagglutinin, deposited dorsolateral to the SON results in labeled pre-and post-synaptic processes within the SON. The identification and characterization of endogenous immunoglobulin within the SON and other neurons innervating areas lacking a blood-brain barrier established a novel and potentially important system for direct communication of the supraoptic cells with blood-borne constitutents

  14. Differences in cellular glycoconjugates of quiescent, inflamed, and neoplastic colonic epithelium in colitis and cancer-prone tamarins.

    Science.gov (United States)

    Moore, R; King, N; Alroy, J

    1988-06-01

    In the preceding paper the authors demonstrated that the lectin staining patterns of normal colonic epithelium obtained from colitis and carcinoma-prone cotton top tamarins (CTTs), Saguinus oedipus, a New World primate, differs from colitis- and carcinoma-resistant primate species. In this study they determined the usefulness of cytochemical features in inflamed epithelium as indicators for malignant change. They compared the lectin staining pattern in inflamed mucosa and adjacent mucosa with colonic carcinoma from 8 CTTs with that of 9 clinically healthy CTTs with no histologic evidence of colitis. Deparaffinized sections were labeled with ten biotinylated lectins and stained by the avidin-biotin peroxidase complex method. Numerous significant differences were demonstrated in the lectin staining pattern between normal epithelium and colonic carcinoma; fewer between normal and chronic inflamed epithelium. However, between chronic inflamed epithelium and colonic carcinoma significant staining differences were observed with only two lectins, peanut agglutinin (PNA) and Ulex europaeus agglutinin-I (UEA-I). These findings suggest that there is a progression in alteration of lectin staining pattern from normal epithelium, via chronic colitis, to colonic carcinoma. Furthermore, the differences between chronic colitis and colonic carcinoma are expressed only with those lectins that are associated with malignant transformation of human colonic epithelium. PMID:3132858

  15. Identification and preliminary characterization of a Ca2+-dependent hemagglutinin in the celomic fluid of Sipunculus nudus

    Directory of Open Access Journals (Sweden)

    L Ballarin

    2010-10-01

    Full Text Available A soluble agglutinin was purified by affinity chromatography of the celomic fluid of the marine worm Sipunculus nudus. This agglutinin requires metal cations for its activity and is specific for derivatives of D-galactose. It resulted lightly thermostable, with a pH optimum around 7.5. On SDS-PAGE, it was resolved in two bands, of 33 and 31 kDa in reducing conditions and 29 and 26 kDa in non-reducing conditions. This behavior is probably due to the presence of disulfide bridges between cysteine residues, which are required for the correct functioning of the hemagglutinin, as β-mercaptoethanol completely abolish the agglutinating activity of cell-free celomic fluid. The purified lectin can influence in vitro phagocytosis of yeast by celomic leukocytes: in the presence of the molecule, ingestion of foreign particles results significantly decreased and yeast cells agglutinate and forms rosettes around the celomocytes. This suggests a role of the molecule in immunosurveillance.

  16. Anatomy, histochemistry and immunohistochemistry of the olfactory subsystems in mice

    Directory of Open Access Journals (Sweden)

    Arthur William Barrios

    2014-07-01

    Full Text Available The four regions of the murine nasal cavity featuring olfactory neurons were studied anatomically and by labelling with lectins and relevant antibodies with a view to establishing criteria for the identification of olfactory subsystems that are readily applicable to other mammals. In the main olfactory epithelium and the septal organ the olfactory sensory neurons (OSNs are embedded in quasi-stratified columnar epithelium; vomeronasal OSNs are embedded in epithelium lining the medial interior wall of the vomeronasal duct and do not make contact with the mucosa of the main nasal cavity; and in Grüneberg’s ganglion a small isolated population of OSNs lies adjacent to, but not within, the epithelium. With the exception of Grüneberg’s ganglion, all the tissues expressing olfactory marker protein (OMP (the above four nasal territories, the vomeronasal and main olfactory nerves, and the main and accessory olfactory bulbs are also labelled by Lycopersicum esculentum agglutinin, while Ulex europaeus agglutinin I labels all and only tissues expressing Gi2 (the apical sensory neurons of the vomeronasal organ, their axons, and their glomerular destinations in the anterior accessory olfactory bulb. These staining patterns of UEA-I and LEA may facilitate the characterization of olfactory anatomy in other species. A 710-section atlas of the anatomy of the murine nasal cavity has been made available on line.

  17. Anatomy, histochemistry, and immunohistochemistry of the olfactory subsystems in mice.

    Science.gov (United States)

    Barrios, Arthur W; Núñez, Gonzalo; Sánchez Quinteiro, Pablo; Salazar, Ignacio

    2014-01-01

    The four regions of the murine nasal cavity featuring olfactory neurons were studied anatomically and by labeling with lectins and relevant antibodies with a view to establishing criteria for the identification of olfactory subsystems that are readily applicable to other mammals. In the main olfactory epithelium and the septal organ the olfactory sensory neurons (OSNs) are embedded in quasi-stratified columnar epithelium; vomeronasal OSNs are embedded in epithelium lining the medial interior wall of the vomeronasal duct and do not make contact with the mucosa of the main nasal cavity; and in Grüneberg's ganglion a small isolated population of OSNs lies adjacent to, but not within, the epithelium. With the exception of Grüneberg's ganglion, all the tissues expressing olfactory marker protein (OMP) (the above four nasal territories, the vomeronasal and main olfactory nerves, and the main and accessory olfactory bulbs) are also labeled by Lycopersicum esculentum agglutinin, while Ulex europaeus agglutinin I labels all and only tissues expressing Gαi2 (the apical sensory neurons of the vomeronasal organ, their axons, and their glomerular destinations in the anterior accessory olfactory bulb). These staining patterns of UEA-I and LEA may facilitate the characterization of olfactory anatomy in other species. A 710-section atlas of the anatomy of the murine nasal cavity has been made available on line. PMID:25071468

  18. Evaluation of the antischistosomal activity of sulfated α-D-glucan from the lichen Ramalina celastri free and encapsulated into liposomes

    Directory of Open Access Journals (Sweden)

    R.V.S. Araújo

    2011-04-01

    Full Text Available The antischistosomal activity of the sulfated polysaccharide α-D-glucan (Glu.SO4 extracted from Ramalina celastri was evaluated after encapsulation into liposomes (Glu.SO4-LIPO in Schistosoma mansoni-infected mice. The effect of treatment with Glu.SO4 and Glu.SO4-LIPO (10 mg/kg on egg elimination, worm burden and hepatic granuloma formation was assessed using female albino Swiss mice, 35-40 days of age, weighing 25 ± 2 g, infected with 150 cercariae/animal (Biomphalaria glabrata, BH strain. Four groups (N = 10 were studied, two controls (empty liposomes and NaCl and two treated groups (Glu.SO4-LIPO and Glu.SO4 using a single dose. Parasitological analysis revealed that Glu.SO4-LIPO was as efficient as Glu.SO4 in reducing egg elimination and worm burden. Treatment with free Glu.SO4 and Glu.SO4-LIPO induced a statistically significant reduction in the number of granulomas (62 and 63%, respectively. Lectin histochemistry showed that wheat germ agglutinin intensely stained the egg-granuloma system in all treated groups. On the other hand, peanut agglutinin stained cells in the control groups, but not in the treated groups. The present results suggest a correlation between the decreasing number of hepatic egg-granulomas and the glycosylation profile of the egg-granuloma system in animals treated with free Glu.SO4 or Glu.SO4-LIPO.

  19. Affinity Separation of Lectins Using Porous Membranes Immobilized with Glycopolymer Brushes Containing Mannose or N-Acetyl-D-Glucosamine

    Directory of Open Access Journals (Sweden)

    Yoshiko Miura

    2013-07-01

    Full Text Available Porous membranes with glycopolymer brushes were prepared as biomaterials for affinity separation. Glycopolymer brushes contained acrylic acid and D-mannose or N-acetyl-D-glucosamine, and were formed on substrates by surface-initiated atom transfer radical polymerization. The presence of glycopolymer brush was confirmed by X-ray photoelectron spectroscopy, contact angle, and ellipsometry measurements. The interaction between lectin and the glycopolymer immobilized on glass slides was confirmed using fluorescent-labeled proteins. Glycopolymer-immobilized surfaces exhibited specific adsorption of the corresponding lectin, compared with bovine serum albumin. Lectins were continuously rejected by the glycopolymer-immobilized membranes. When the protein solution was permeated through the glycopolymer-immobilized membrane, bovine serum albumin was not adsorbed on the membrane surface. In contrast, concanavalin A and wheat germ agglutinin were rejected by membranes incorporating D-mannose or N-acetyl-D-glucosamine, respectively. The amounts of adsorbed concanavalin A and wheat germ agglutinin was increased five- and two-fold that of adsorbed bovine serum albumin, respectively.

  20. The effect of the state of differentiation on labeling of epidermal cell surface glycoproteins

    International Nuclear Information System (INIS)

    Epidermal cells were grown in a medium in which the Ca++ concentration controlled the stage of differentiation. Cell surface molecules of differentiated and undifferentiated cells were compared by lactoperoxidase-catalyzed iodination, by the interaction with 125I-lectins, and by the metabolic incorporation of L-(3H)-fucose. Molecular weights of the labeled components were determined by SDS-PAGE and autoradiography. After lactoperoxidase iodination, most of the radioactivity was found in polypeptide bands of 79,000, 65,000 and 56,000 daltons. The 79,000 band is the most intense for undifferentiated cells but disappears as differentiation proceeds. The 56,000 band is present in normal cells at all stages of differentiation but is absent from neoplastic cells. Glycoproteins reacted with 125I-lectins were found at 180,000, 130,000 and 85,000 daltons. The 130,000 band was the most prominent for differentiated cells labeled with wheat germ agglutinin but was essentially absent from the undifferentiated cells. With Ricinus communis agglutinin, this band was weaker for undifferentiated than for differentiated cells but was the most intense for both. After metabolic incorporation of tritiated fucose, radioactive glycoproteins were found at 130,000 and 85,000 daltons, with comparable intensities for differentiated and undifferentiated cells

  1. Direct targeting of cancer cells: a multiparameter approach.

    Science.gov (United States)

    Heinrich, Eileen L; Welty, Lily Anne Y; Banner, Lisa R; Oppenheimer, Steven B

    2005-01-01

    Lectins have been widely used in cell surface studies and in the development of potential anticancer drugs. Many past studies that have examined lectin toxicity have only evaluated the effects on cancer cells, not their non-cancer counterparts. In addition, few past studies have evaluated the relationship between lectin-cell binding and lectin toxicity on both cell types. Here we examine these parameters in one study: lectin-cell binding and lectin toxicity with both cancer cells and their normal counterparts. We found that the human colon cancer cell line CCL-220/Colo320DM bound to agarose beads derivatized with Phaseolus vulgaris agglutinin (PHA-L) and wheat germ agglutinin (WGA), while the non-cancer human colon cell line CRL-1459/CCD-18Co did not. When these lectins were tested for their effects on cell viability in culture, both cell lines were affected by the lectins but at 6, 48 and 72 h incubation times, PHA-L was most toxic to the cancer cell line in a concentration dependent manner. At 48 h incubation, WGA was more toxic to the cancer cell line. The results suggest that it may be possible to develop lectin protocols that selectively target cancer cells for death. In any case, examination of both malignant cells and their non-malignant counterparts, analysis of their binding characteristics to immobilized lectins, and examination of the toxicity of free lectins in culture, provides a multiparameter model for obtaining more comprehensive information than from more limited approaches. PMID:16181664

  2. Partial isolation and characterisation of a hemagglutinating factor from avocado seed.

    Science.gov (United States)

    Yaakobovich, Y; Neeman, I

    1983-01-01

    Extracts of ground avocado seeds (Fuerte and Hass varieties), prepared in different buffer solutions (pH 2.0-12.0), show hemagglutinating activity towards A, B, AB, and H (0) human erythrocytes. The extract showing the highest titer of aggulination was extracted at pH 10.5. The crude extract also causes hemolysis of fresh washed erythrocytes. The hemagglutinating factor is not inhibited by most of the simple sugars tested, e.g., D-glucose, D-mannose, D-galactose, and glucose-amine. The only sugars which show some inhibitory effect are N-Acetyl-neuraminic acid, melibiose, and stachiose. Basic amino acids, e.g., lysine and arginine also inhibit its activity. However the most potent inhibitors of the agglutinin are proteins and glycoproteins such as bovine serum albumin, collagen, thyroglobulin, ovalbumin, mucin, and fetuin. The agglutinin is adsorbed on polymer beads such as Sepharose 4B, Sephadex G100, Agarose, and Chitin, and it reacts with hog erythrocyte membranes. It can be partially eluted from those materials with alkaline buffers (pH 9.0-10.5). PMID:6578749

  3. A kinetic analysis of the tumor-associated galactopyranosyl-(1→3)-2-acetamido-2-deoxy--D-galactopyranoside antigen-lectin interaction

    Indian Academy of Sciences (India)

    Bandaru Narasimha Murthy; Narayanaswamy Jayaraman

    2008-01-01

    A kinetic study of the tumor-associated galactopyranosyl-(1→3)-2-acetamido-2-deoxy--Dgalactopyranoside (T-antigen) with lectin peanut agglutinin is described. The disaccharide antigen was synthesized by chemical methods and was functionalized suitably for immobilization onto a carboxymethylated sensor chip. The ligand immobilized surface was allowed interaction with the lectin peanut agglutinin, which acted as the analyte and the interaction was studied by the surface plasmon resonance method. The ligand-lectin interaction was characterized by the kinetic on-off rates and a bivalent analyte binding model was found to describe the observed kinetic constants. It was identified that the antigen-lectin interaction had a faster association rate constant (a1) and a slower dissociation rate constant (d1) in the initial binding step. The subsequent binding step showed much reduced kinetic rates. The antigen-lectin interaction was compared with the kinetic rates of the interaction of a galactopyranosyl-(1→ 4)--D-galactopyranoside derivative and a mannopyranoside derivative with the lectin.

  4. Design of near-infrared fluorescent bioactive conjugated functional iron oxide nanoparticles for optical detection of colon cancer

    Directory of Open Access Journals (Sweden)

    Corem-Salkmon E

    2012-10-01

    Full Text Available Enav Corem-Salkmon, Benny Perlstein, Shlomo MargelThe Institute of Nanotechnology and Advanced Materials, Department of Chemistry, Bar-Ilan University, Ramat-Gan, IsraelBackground: Colon cancer is one of the major causes of death in the Western world. Early detection significantly improves long-term survival for patients with the disease. Near-infrared (NIR fluorescent nanoparticles hold great promise as contrast agents for tumor detection. NIR offers several advantages for bioimaging compared with fluorescence in the visible spectrum, ie, lower autofluorescence of biological tissues, lower absorbance, and consequently deeper penetration into biomatrices.Methods and results: NIR fluorescent iron oxide nanoparticles with a narrow size distribution were prepared by nucleation, followed by controlled growth of thin iron oxide films onto cyanine NIR dye conjugated gelatin-iron oxide nuclei. For functionalization, and in order to increase the NIR fluorescence intensity, the NIR fluorescent iron oxide nanoparticles obtained were coated with human serum albumin containing cyanine NIR dye. Leakage of the NIR dye from these nanoparticles into phosphate-buffered saline solution containing 4% albumin was not detected. The work presented here is a feasibility study to test the suitability of iron oxide-human serum albumin NIR fluorescent nanoparticles for optical detection of colon cancer. It demonstrates that encapsulation of NIR fluorescent dye within these nanoparticles significantly reduces photobleaching of the dye. Tumor-targeting ligands, peanut agglutinin and anticarcinoembryonic antigen antibodies (αCEA, were covalently conjugated with the NIR fluorescent iron oxide-human serum albumin nanoparticles via a poly(ethylene glycol spacer. Specific colon tumor detection was demonstrated in chicken embryo and mouse models for both nonconjugated and the peanut agglutinin-conjugated or αCEA-conjugated NIR fluorescent iron oxide-human serum albumin

  5. Childhood brucellosis--a microbiological, epidemiological and clinical study.

    Science.gov (United States)

    Mantur, B G; Akki, A S; Mangalgi, Smita S; Patil, S V; Gobbur, R H; Peerapur, B V

    2004-06-01

    A total of 5726 blood specimens (from children aged 14 years and younger) were studied for the serological evidence of brucellosis. Ninety-three (1.6 per cent) showed diagnostic agglutinin titres with a geometric mean titre of 403 (SD +/- 547). Forty-three (59.7 per cent) blood specimens yielded the growth of Brucella melitensis. Thirty-nine patients (41.93 per cent) were shepherds, who constituted the major occupational group affected in the present series. More than 60 per cent of the patients had a history of both consumption of fresh goat's milk and close animal contact. The habit of consuming fresh goat's milk to obtain relief from chronic ailments was noted in nine patients. Seventy-three (78.49 per cent) were males and 20 (21.51 per cent) were females, with a male to female ratio of 3:1. The disease occurred mainly in the school age group (mean age 10.3 years). All the patients had an acute history of less than 2 months. Forty-nine (52.68 per cent) patients presented with persistent fever, 19 (20.43 per cent) with joint pain, and the rest with a combination of fever and joint pain with and without low backache, fever being the commonest complaint. One case presented with involuntary movements of limbs alone and the other with burning feet only. Pityriasis alba was the consistent physical finding, with fever in the majority of the patients. The major joint found to be involved was the knee (52.77 per cent). The synovial fluid obtained from the knee joint of five patients demonstrated Brucella agglutinins and also three grew B. melitensis. Eight patients presented with complications that included skin lesions (3), carditis (2), neurobrucellosis such as chorea (1), peripheral neuritis (1), and meningitis (1). Brucella melitensis biotype 1 was successfully isolated from the papular eruption of one out of three cases who presented with skin lesions. To our knowledge this is the fourth confirmed isolation of B. melitensis from skin lesions with brucellosis

  6. Syntheses of Sulfo-Glycodendrimers Using Click Chemistry and Their Biological Evaluation

    Directory of Open Access Journals (Sweden)

    Tomohiro Fukuda

    2012-10-01

    Full Text Available A series of novel glycol-clusters containing sulfonated N-acetyl-D-glucosamine (GlcNAc have been synthesized using click chemistry. Three dendrimers with aromatic dendrons were synthesized using chlorination, azidation and click chemistries. The resulting dendrimers were modified with azide-terminated sulfonated GlcNAc using click chemistry. The sulfonated dendrimers showed affinity for proteins, including the lectin wheat germ agglutinin and amyloid beta peptide (1-42. The dendrimers of G1 and G2 in particular showed the largest affinity for the proteins. The addition of the sulfonated GlcNAc dendrimers of G1 and G2 exhibited an inhibition effect on the aggregation of the amyloid beta peptide, reduced the b-sheet conformation, and led to a reduction in the level of nanofiber formation.

  7. PNA: a marker of neoplastic progression and differentiation in the gastro-intestinal tract.

    Science.gov (United States)

    Grigolato, P; Benetti, A; Berenzi, A; Villanacci, V; Tardanico, R

    1990-01-01

    We examined 35 cases of stomach carcinoma and 40 cases of colonic carcinoma with PNA associated with peroxidase (peanut agglutinin, lectin which binds to the terminal disaccharide galactose beta (1,3)-N-acetil-galacto-samine). In this way evaluation of the functional aspects of the normal-neoplastic sequence was undertaken. This method was carried out for histological and ultrastructural investigations. The results obtained in both cases showed a different reactivity in the evolution of neoplastic disease: in fact, positivity in dysplasia is finely granular intracytoplasmic, whereas in well-differentiated neoplastic transformation such a reactivity is preferentially localized along the cellular membranes, with restoration of gross positivity in the cytoplasm for the poorly-differentiated neoplasm. We therefore believe PNA to be a marker not only of neoplastic progression but of differentiation as well: we also hypothesize it to reveal glycoprotein groups with possible antigenic power, involved in immunologic interactions between tumor and host. PMID:2283482

  8. Títulos de anticorpos aglutinantes induzidos por vacinas comerciais contra leptospirose bovina Agglutinating antibody titers induced by commercial vaccines against bovine leptospirosis

    Directory of Open Access Journals (Sweden)

    Gabriela de Godoy Cravo Arduino

    2009-07-01

    serovars. All vaccines used were capable to product agglutinins for the Hardjo and Wolffi serovars observed at 3 days after vaccination, remaining until the 150th day; those serovars induced the highest titres of agglutinins. Vaccine D, in spite of not containing the Wolffi serovar, induced the production of agglutinins to this serovar. Agglutinins to the Canicola serovar were only observed in the animals vaccinated with the D bacterine. Vaccine D induced the highest average titers of antibodies to all tested serovars.

  9. Visual detection of serum asialohaptoglobin by plasmonic sandwich ELLSA--a new platform for cirrhosis diagnosis.

    Science.gov (United States)

    Bose, Partha Pratim; Mandal, Gautam; Kumar, Dharmendra; Duseja, Ajay; Chatterjee, Bishnu Pada

    2016-01-01

    The cirrhotic condition of the liver has long been acknowledged as the preface to liver cancer. The desialylation status of the serum acute phase protein, haptoglobin, has been introduced as a new diagnostic analyte for liver cirrhosis. The reliability of this new diagnostic molecule has been evaluated in 30 liver cirrhosis patients having a history of earlier viral hepatitis C (HCV-LC). A novel enzyme linked lectinosorbent assay has been developed coupled with the plasmon mechanism of gold nanoparticle aggregation as the colorimetric read out which can visually distinguish the cirrhotic liver patients from the normal healthy and hepatitis C controls. The assay can be useful for rapid point-of-care detection, and even an untrained person can execute it without a specialized instrument. This method employs Sambucus nigra agglutinin (SNA) to detect the extent of α-2,6 sialylation of serum haptoglobin, the new diagnostic molecule for liver cirrhosis. PMID:26568048

  10. Detection of Sugar-Lectin Interactions by Multivalent Dendritic Sugar Functionalized Single-Walled Carbon Nanotubes

    CERN Document Server

    Vasu, K S; Bagul, R S; Jayaraman, N; Sood, A K; 10.1063/1.4739793

    2012-01-01

    We show that single walled carbon nanotubes (SWNT) decorated with sugar functionalized poly (propyl ether imine) (PETIM) dendrimer is a very sensitive platform to quantitatively detect carbohydrate recognizing proteins, namely, lectins. The changes in electrical conductivity of SWNT in field effect transistor device due to carbohydrate - protein interactions form the basis of present study. The mannose sugar attached PETIM dendrimers undergo charge - transfer interactions with the SWNT. The changes in the conductance of the dendritic sugar functionalized SWNT after addition of lectins in varying concentrations were found to follow the Langmuir type isotherm, giving the concanavalin A (Con A) - mannose affinity constant to be 8.5 x 106 M-1. The increase in the device conductance observed after adding 10 nM of Con A is same as after adding 20 \\muM of a non - specific lectin peanut agglutinin, showing the high specificity of the Con A - mannose interactions. The specificity of sugar-lectin interactions was chara...

  11. Effects of processing on antinutritional factors in legumes: the soybean case.

    Science.gov (United States)

    Liener, I E

    1996-12-01

    The author recounts his personal trail of research which has ultimately led to better understanding of the factors which contribute to the poor nutritive value of unheated soybeans. Among the techniques that were employed were the isolation of a lectin from raw soybeans, the use of affinity chromatography to remove the trypsin inhibitors, and the nutritional evaluation of soybean varieties which lacked the lectin or the Kunitz trypsin inhibitor. Based on a consideration of the results obtained by these experiments, it was estimated that the trypsin inhibitors accounted for approximately 40% of the growth inhibition on raw soy, of which two-thirds could be attributed to the Kunitz inhibitor and one-third to the Bowman-Birk inhibitor. The soybean agglutinin was deemed responsible for 50% of the inhibition of growth, and the remaining 10% is most likely due to the poor digestibility of the undenatured protein. PMID:9137638

  12. A double diastereoselective Michael-type addition as an entry to conformationally restricted tn antigen mimics.

    Science.gov (United States)

    Aydillo, Carlos; Navo, Claudio D; Busto, Jesús H; Corzana, Francisco; Zurbano, María M; Avenoza, Alberto; Peregrina, Jesús M

    2013-11-01

    A totally stereocontrolled C-Michael addition of serine-equivalent C-nucleophiles to tri-O-benzyl-2-nitro-d-galactal was used as the key step to synthesize several pyrano[3,2-b]pyrrole structures. These scaffolds could be regarded as conformationally restricted Tn antigen mimics, as we have demonstrated by biological assays. The pyranose rings retain their (4)C1 chair conformation, as shown by molecular modeling and NMR spectroscopy. The expected bioactivity was established by a competition-tailored enzyme-linked lectin assay using both soybean and Vicia villosa agglutinins as model lectins. The facile described synthetic route and the strategic combination of computational and experimental techniques to reveal conformational features and bioactivity demonstrate the prepared glycomimics to be promising candidates for further exploitation of this scaffold to give glycans for lectin blocking and vaccination. PMID:24083620

  13. Covalent Attachment of Carbohydrate Derivatives to an Evanescent Wave Fiber Bragg Grating Biosensor

    Directory of Open Access Journals (Sweden)

    Christopher J. Stanford

    2009-01-01

    Full Text Available A carbohydrate-based biosensor was prepared by functionalization of the surface of an etched fiber Bragg grating with a glucopyranosyl-siloxane conjugate. Functionalization of the surface with the conjugate resulted in a Bragg grating shift of 24 pm. This shift in the refractive index is consistent with a theoretical shift calculated assuming monolayer coverage of the glucose conjugate on the sensor. The resulting functionalized fiber was shown to interact selectively with concanavalin A (Con A, a glucose binding protein (lectin. Exposure of the glucose-functionalized fiber to peanut agglutinin, a galactosebinding lectin, did not result in a change of the refractive index corresponding to a binding event.

  14. Detection of sialoglycomolecules in five plant trypanosomatids and in an insect phytophagous isolate.

    Science.gov (United States)

    Souza dos Santos, André Luis; Sales Alviano, Celuta; de Araújo Soares, Rosangela Maria

    2002-08-27

    The sialoglycoprotein profiles of five plant trypanosomatids (Phytomonas spp.) and of one flagellate (Herpetomonas sp.) isolated from the salivary gland of a phytophagous insect (Phthia picta) were analyzed by Western blotting using three distinct lectins (LFA, SNA and MAA), which recognize specifically sialic acid residues in glycoconjugates. All six flagellates presented at least one polypeptide recognized by the lectins, with the exception of Phytomonas françai, which did not show any reactivity with SNA agglutinin. Phytomonas serpens and P. françai showed the most distinct pattern of sialoglycoproteins. Phytomonas mcgheei, Herpetomonas sp. and the two other Phytomonas spp., isolated from latex, displayed an identical sialomolecule profile. We discuss the possible role of the sialoglycoproteins in the physiology of these trypanosomatids. PMID:12204367

  15. Blood Group Discrepancy-First Sign of Autoimmune Hemolytic Anemia after Hematopoietic Stem Cell Transplantation in a Child.

    Science.gov (United States)

    Datta, Suvro Sankha; Reddy, Mahua; Basu, Sabita; Krishnan, Shekhar

    2016-06-01

    A 12-year-old male child was presented in the emergency with features of anemia and mild icterus on day+67 of HSCT. The child was suffering from Fanconi anemia and undergone HSCT from ABO-matched, fully HLA matched sibling donor. The diagnosis of mixed type AIHA due to cytomegalovirus reactivation was made in the immunohematology laboratory and blood group discrepancy was the first sign of AIHA in this patient. Though the cold agglutinin titer was not significant but the clinical symptoms and laboratory evidences were suggestive of significant hemolysis due to underlying IgG autoantibody. In addition the high complement avidity of IgM autoantibody might also be a contributing factor for clinically significant hemolysis in this case. The patient was successfully treated with phenotype matched blood transfusion, rituximab and oral steroid therapy. PMID:27408394

  16. Bioengineering single crystal growth.

    Science.gov (United States)

    Wu, Ching-Hsuan; Park, Alexander; Joester, Derk

    2011-02-16

    Biomineralization is a "bottom-up" synthesis process that results in the formation of inorganic/organic nanocomposites with unrivaled control over structure, superior mechanical properties, adaptive response, and the capability of self-repair. While de novo design of such highly optimized materials may still be out of reach, engineering of the biosynthetic machinery may offer an alternative route to design advanced materials. Herein, we present an approach using micro-contact-printed lectins for patterning sea urchin embryo primary mesenchyme cells (PMCs) in vitro. We demonstrate not only that PMCs cultured on these substrates show attachment to wheat germ agglutinin and concanavalin A patterns but, more importantly, that the deposition and elongation of calcite spicules occurs cooperatively by multiple cells and in alignment with the printed pattern. This allows us to control the placement and orientation of smooth, cylindrical calcite single crystals where the crystallographic c-direction is parallel to the cylinder axis and the underlying line pattern. PMID:21265521

  17. Gastrointestinal parasites in Danish goats - prevalence and risk factors

    DEFF Research Database (Denmark)

    Sörensen, C.; Holm, S. A.; Thamsborg, S. M.;

    2012-01-01

    with EPG>300, and herds with a mean EPG>150 were offered a faecal egg count reduction test (FECRT). All herds were asked to complete a questionnaire about management and risk factors concerning parasites, particularly nematodes. Faecal egg counts were generally low; 2 out of 25 herds had a mean EPG>150......The aims were to examine prevalence of gastrointestinal parasites in Danish goats, based on faecal examination, in relation to geographical distribution and risk factors, and to investigate the occurrence of anthelmintic resistance in nematodes in selected farms. In April 2012 all Danish goat farms...... using a modified McMaster technique (sensitivity 5 eggs per gr. of faeces (EPG)). From each herd, samples with EPG>500 were pooled and stained with peanut agglutinin (PNA) for detection of Haemonchus contortus. An egg hatch assay (EHA) for detection of anthelmintic resistance was performed on samples...

  18. Screening for N-glycosylated proteins by liquid chromatography mass spectrometry

    DEFF Research Database (Denmark)

    Bunkenborg, Jakob; Pilch, Bartosz J; Podtelejnikov, Alexandre V; Wiśniewski, Jacek R

    2004-01-01

    In the last few years mass spectrometry has become the method of choice for characterization of post-translationally modified proteins. Whereas most protein chemical modifications are binary in the sense that only one change can be associated with a given residue, many different oligosaccharides...... complex mixtures by reducing sample complexity and enriching glycoprotein content. Glycosylated proteins are selected by an initial lectin chromatography step and digested with endoproteinase Lys-C. Glycosylated peptides are then selected from the digest mixture by a second lectin chromatography step. The...... glycan components are removed with N-glycosidase F and the peptides digested with trypsin before analysis by on-line reversed-phase liquid chromatography mass spectrometry. Using two different lectins, concanavalin A and wheat germ agglutinin, this procedure was applied to human serum and a total of 86 N...

  19. Definitive proof of endothelialization of a Dacron arterial prosthesis in a human being.

    Science.gov (United States)

    Wu, M H; Shi, Q; Wechezak, A R; Clowes, A W; Gordon, I L; Sauvage, L R

    1995-05-01

    A 10 mm woven Dacron axillofemoral bypass graft was removed from a 65-year-old patient during redo surgery after an implant period of 26 months, because of a large seroma that surrounded the entire length of the graft. Tissue blocks were taken from representative areas along the entire length of the graft surface and evaluated by light microscopy with hematoxylin and eosin and Masson trichrome staining, scanning electron microscopy, transmission electron microscopy, and immunocytochemical staining. Paraffin-embedded sections were stained with smooth muscle cell alpha-actin, which demonstrated smooth muscle cells in the pseudointima, and Ham 56 stain to identify macrophages. Endothelial factor VIII/von Willebrand factor and Ulex europaeus agglutinin identified human endothelial cells on the flow surface, in areas far removed from the anastomoses to the native vessels. This is the first definitive proof in a human of endothelialization of a synthetic arterial graft beyond the pannus ingrowth zone. PMID:7769746

  20. Partial characterization of the N-linked oligosaccharide structures on Pselectin glycoprotein ligand-1 (PSGL-1)

    Institute of Scientific and Technical Information of China (English)

    2001-01-01

    PSGL-1,a specific ligand for P-,E- and L-selectin,was isolated from in vivo [3H]-glucosamine labeled HL-60 cells by a combination of wheat germ agglutinin-agarose and P- or E-selectin-agarose chromatography.N-linked oligosaccharides were released from the purified,denatured ligand molecule by peptide: N-glycosidase F treatment and,following separation by Sephacryl S-200 chromatography,partially characterized using lectin,ion-exchange and size-exclusion chromatography in combination with glycosidase digestions.The data obtained suggest that the N-glycans on PSGL-1 are predominantly core-fucosylated,multiantennary complex type structures with extended,poly-N-acetyllactosamine containing outer chains.A portion of the outer chains appears to be substituted with fucose indicating that the N-glycans,in addition to the O-glycans on PSGL-1,may be involved in selectin binding.

  1. Diffuse pulmonary haemorrhage accompanied by haemothorax as a rare presentation of primary lung angiosarcoma

    Science.gov (United States)

    Radzikowska, Elżbieta; Szołkowska, Małgorzata; Oniszh, Karina; Szczęsna, Magdalena; Roszkowski-Śliż, Kazimierz

    2015-01-01

    Primary pulmonary angiosarcoma is an extremely rare disease. Chest computed tomography demonstrates solitary or multifocal lesions, sometimes associated with ground-glass opacities or pleural effusion. Diagnosis is based on histological examination that reveals spindle-shaped epithelioid cells with positive staining for endothelial markers (factor VIII, CD 31, CD34, Fli-1, Ulex europaeus agglutinin 1, vimentin). The prognosis is poor and effective treatment is still being researched. This is a report of a 65-year-old patient with a four-month history of haemoptysis, cough, and dyspnoea. The primary radiological findings suggested interstitial lung disease. After one month the clinical presentation evolved into diffuse pulmonary haemorrhage with concomitant haemothorax. The diagnosis of primary lung angiosarcoma was based on histological and immunohistochemical examination of the lung and pleural biopsy obtained by videothoracoscopy. PMID:26855658

  2. Molecular bases of the influence of calcium influx on the heart by means of a calcium duct-blocker

    International Nuclear Information System (INIS)

    The goal of this work was to identify and purify the physiological binding places for calcium duct-blocker in the heart muscle of cattle. The following was thereby attempted: 1. to purify the plasma membranes from heart cells; 2. to develop a measuring method for the tension-dependent calcium duct with the help of tritiated calcium duct-blocker or respectively, to improve existing methods; 3. to characterize by the use of different calcium duct-blockers this ion duct; 4. to develop methods to solubilize the calcium duct out of the plasma membrane using detergents; as well as 5. to purify the dissolved calcium duct by use of various procedures such as affinity chromatography, chromatography on wheatgerm-agglutinin-substituted sepharose and density gradient centrifugation. (orig./MG)

  3. How a Plant Lectin Recognizes High Mannose Oligosaccharides1[C][OA

    Science.gov (United States)

    Garcia-Pino, Abel; Buts, Lieven; Wyns, Lode; Imberty, Anne; Loris, Remy

    2007-01-01

    The crystal structure of Pterocarpus angolensis seed lectin is presented in complex with a series of high mannose (Man) oligosaccharides ranging from Man-5 to Man-9. Despite that several of the nine Man residues of Man-9 have the potential to bind in the monosaccharide-binding site, all oligomannoses are bound in the same unique way, employing the tetrasaccharide sequence Manα(1–2)Manα(1–6)[Manα(1–3)]Manα(1–. Isothermal titration calorimetry titration experiments using Man-5, Man-9, and the Man-9-containing glycoprotein soybean (Glycine max) agglutinin as ligands confirm the monovalence of Man-9 and show a 4-times higher affinity for Man-9 when it is presented to P. angolensis seed lectin in a glycoprotein context. PMID:17556509

  4. How a plant lectin recognizes high mannose oligosaccharides.

    Science.gov (United States)

    Garcia-Pino, Abel; Buts, Lieven; Wyns, Lode; Imberty, Anne; Loris, Remy

    2007-08-01

    The crystal structure of Pterocarpus angolensis seed lectin is presented in complex with a series of high mannose (Man) oligosaccharides ranging from Man-5 to Man-9. Despite that several of the nine Man residues of Man-9 have the potential to bind in the monosaccharide-binding site, all oligomannoses are bound in the same unique way, employing the tetrasaccharide sequence Manalpha(1-2)Manalpha(1-6)[Manalpha(1-3)]Manalpha(1-. Isothermal titration calorimetry titration experiments using Man-5, Man-9, and the Man-9-containing glycoprotein soybean (Glycine max) agglutinin as ligands confirm the monovalence of Man-9 and show a 4-times higher affinity for Man-9 when it is presented to P. angolensis seed lectin in a glycoprotein context. PMID:17556509

  5. Versatile Route to Colloidal Stability and Surface Functionalization of Hydrophobic Nanomaterials.

    Science.gov (United States)

    Culver, Heidi R; Steichen, Stephanie D; Herrera-Alonso, Margarita; Peppas, Nicholas A

    2016-06-01

    We introduce a general method for the stabilization and surface functionalization of hydrophobic nanoparticles using an amphiphilic copolymer, poly(maleic anhydride-alt-1-octadecene)-poly(ethylene glycol) methacrylate (PMAO-PEGMA). Coating nanoparticles with PMAO-PEGMA results in colloidally stable nanoparticles decorated with reactive carboxylic acid and methacrylate functionalities, providing a versatile platform for chemical reactions. The versatility and ease of surface functionalization is demonstrated by varying both the core material and the chemistry used. Specifically, the carboxylic acid functionalities are used to conjugate wheat germ agglutinin to conducting polymer nanoparticles via carbodiimide-mediated coupling, and the methacrylate groups are used to link cysteamine to the surface of poly(ε-caprolactone) nanoparticles via thiol-ene click chemistry and to link temperature-responsive polymer shells to the surface of gold nanoparticles via free radical polymerization. PMID:27203863

  6. AcEST: DK958409 [AcEST

    Lifescience Database Archive (English)

    Full Text Available 94_ASPNC Similarity to hypothetical endoglucanase IV - Trichoderma reesei OS=Aspergillus niger (strain CBS 5...r|A2QR94|A2QR94_ASPNC Similarity to hypothetical endoglucanase IV - Trichoderma reesei OS=Aspergillus niger ...tX Result : Swiss-Prot sp_hit_id Q99KG7 Definition sp|Q99KG7|HPS4_MOUSE Hermansky-Pudlak syn...............done Score E Sequences producing significant alignments: (bits) Value sp|Q99KG7|HPS4_MOUSE Herma..... 30 7.7 sp|P32323|AGA1_YEAST A-agglutinin anchorage subunit OS=Saccharom... 30 7.7 >sp|Q99KG7|HPS4_MOUSE Herma

  7. Study on the Obtaining of Transgenic Wheat with GNA Alien Gene by Biolistic Particle

    Institute of Scientific and Technical Information of China (English)

    XU Qiong-fang; LI Lian-cheng; CHEN Xiao; TIAN Fang; MA You-zhi; YE Xing-guo; ZHANG Zeng-yan; XU Hui-jun; XIN Zhi-yong

    2002-01-01

    The immature embryos of wheat plants, cv. Jing 411, 12 - 14 days after pollination, were cultured on SD2 medium for callus induction. After 10 days culture, 800 wheat calli were bombarded by biolistic particle coated with theDNA of plasmid pBI121-2 harboring both Galanthus nivalis agglutinin gene and bar gene. 67 green plants were finally regenerated from the bombardment calli on selection medium containing 4mg/L Basta. The results of bioassay by both inoculating wheat aphids onto the plants and applying Basta solution of 50 mg/L and 75 mg/L onto the wheat leaves in the field, and the molecular analysis, such as PCR and Southern blotting, indicated that 8 T2 plants contaning the target genes were obtained.

  8. High resolution melting analysis as a new approach to discriminate gluten-containing cereals.

    Science.gov (United States)

    Martín-Fernández, Begoña; Costa, Joana; de-Los-Santos-Álvarez, Noemí; López-Ruiz, Beatriz; Oliveira, M Beatriz P P; Mafra, Isabel

    2016-11-15

    With this work, it is intended to propose a novel approach based on high resolution melting (HRM) analysis to detect wheat and discriminate it from other gluten-containing cereals. The method consisted of a real-time PCR assay targeting the gene encoding for the germ agglutinin isolectin A protein (Tri a 18 allergen), using the fluorescent Evagreen dye combined with HRM analysis. The results enabled wheat differentiation from other phylogenetically related cereals, namely barley, rye and oat with high level of confidence. Additionally, a quantitative real-time PCR approach was proposed, allowing detecting and quantifying wheat down to 20mg/kg in rice flour and 20pg of wheat DNA (∼1.1 DNA copies). Its application was successfully achieved in the analysis of processed foods to verify labelling compliance, being considered as a cost-effective tool for the specific detection of cereals in gluten-free foods. PMID:27283646

  9. Effects of proteolytic enzymes and neuraminidase on the I and i erythrocyte antigen sites

    International Nuclear Information System (INIS)

    Homogeneous cold agglutinins, purified and labelled with 125I, have been used in a study of the effects of neuraminidase and proteolytic enzymes on the I and i reactivities of human adult erythrocytes. Measurements were made of antigen site numbers, equilibrium constants and thermodynamic parameters. There was enhanced reactivity after enzyme treatment as well as after the release of N-acetylneuraminic acid. Steric factors were shown to be of primary importance in the accessibility of the I and i antigenic determinant. After enzyme treatment, the antigenic structures became more homogeneous in their reaction with antibodies. The heterogeneity of binding constants observed with antigenic determinants of non-treated erythrocytes is probably due to the wide range of spatial distribution of these receptors within the membrane. (author)

  10. Warm Autoimmune Haemolytic Anaemia and autoimmune hepatitis in an asymptomatic carrier of hepatitis B virus

    International Nuclear Information System (INIS)

    Warm antibody autoimmune haemolytic anaemia, a rare disease (0.2-1 per 100,000 populations), is due to the presence of warm agglutinins that react with protein antigens on the surface of red blood cells causing their premature destruction. Here, we present a case report of a 10 year old girl who came with features of haemolytic anaemia and history of blood transfusion since 3 years. On admission, laboratory test revealed that she had autoimmune hepatitis type 1 and was also an asymptomatic carrier of hepatitis B virus with positive HBs Ag. Steroid therapy resulted in clinical and laboratory remission. Direct antiglobulin test was negative after anaemia resolution, hepatitis B virus antigenemia persisted. To our knowledge, warm antibody autoimmune hemolytic anaemia has not previously been described in association with autoimmune hepatitis and asymptomatic carrier state of hepatitis B virus. (author)

  11. Maize—A potential source of human nutrition and health: A review

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    Tajamul Rouf Shah

    2016-12-01

    Full Text Available Maize or corn (Zea mays L. is an important cereal crop of the world. It is a source of nutrition as well as phytochemical compounds. Phytochemicals play an important role in preventing chronic diseases. It contains various major phytochemicals such as carotenoids, phenolic compounds, and phytosterols. It is believed to have potential anti-HIV activity due to the presence of Galanthus nivalis agglutinin (GNA lectin or GNA-maize. A tablespoon of maize oil satisfies the requirements for essential fatty acids for a healthy child or adult. Decoction of maize silk, roots, leaves, and cob are used for bladder problems, nausea, vomiting, and stomach complaints. Zein an alcohol-soluble prolamine found in maize endosperm has unique novel applications in pharmaceutical and nutraceutical areas. Resistant starch (RS from maize reduces the risk of cecal cancer, atherosclerosis, and obesity-related complications. This review presents a detailed view on the nutritional and potential health benefits of maize.

  12. Methodological factors affecting the results of staining frozen-thawed fertile and subfertile Japanese Black bull spermatozoa for acrosomal status.

    Science.gov (United States)

    Almadaly, Essam; El-Kon, Ismail; Heleil, Bassiouni; Fattouh, El-Sayed; Mukoujima, Koushi; Ueda, Takuya; Hoshino, Youichirou; Takasu, Masaki; Murase, Tetsuma

    2012-12-01

    In the present study, some methodological factors affecting the acrosomal staining of frozen-thawed Japanese Black bull spermatozoa were investigated by examining; the effect of fixation/permeabilization procedure on intact acrosome percentage after fluorescein isothiocyanate peanut agglutinin (FITC-PNA) staining, the acrosomal staining patterns by using two types of fluorescent probes FITC-PSA (Pisum Sativum Agglutinin) and FITC-PNA and the effect of staining methods, either smear or vial, on intact acrosome percentage. Then intact acrosome percentage was compared between the samples stained by thus established method and those simply fixed with glutaraldehyde (glutaraldehyde fixation method). A possibility that FITC-PNA staining or the glutaraldehyde fixation methods could detect any difference in intact acrosome percentage or acrosomal staining patterns between fertile and subfertile bulls was also examined. The results showed that (1) 4% paraformaldehyde fixation plus 1% Triton X-100 permeabilization was better than absolute ethanol alone, (2) FITC-PNA acrosomal labeling was more specific than FITC-PSA, (3) sperm suspensions should be smeared and gently processed before acrosomal staining rather than spotted onto glass slides after staining in vial in order to avoid excessive mechanical damage of the sperm acrosome, and (4) staining spermatozoa with FITC-PNA had no major advantages over examination of simply glutaraldehyde fixed sperm samples and both failed to detect any significant difference in intact acrosome percentage between the fertile and the subfertile bulls used here. The present study demonstrates important methodological considerations which need to be taken into account in order to design a reliable and reproducible protocol for the study of the acrosome. PMID:23182469

  13. Negative biomarker based male fertility evaluation: Sperm phenotypes associated with molecular-level anomalies

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    Peter Sutovsky

    2015-01-01

    Full Text Available Biomarker-based sperm analysis elevates the treatment of human infertility and ameliorates reproductive performance in livestock. The negative biomarker-based approach focuses on proteins and ligands unique to defective spermatozoa, regardless of their morphological phenotype, lending itself to analysis by flow cytometry (FC. A prime example is the spermatid specific thioredoxin SPTRX3/TXNDC8, retained in the nuclear vacuoles and superfluous cytoplasm of defective human spermatozoa. Infertile couples with high semen SPTRX3 are less likely to conceive by assisted reproductive therapies (ART and more prone to recurrent miscarriage while low SPTRX3 has been associated with multiple ART births. Ubiquitin, a small, proteolysis-promoting covalent posttranslational protein modifier is found on the surface of defective posttesticular spermatozoa and in the damaged protein aggregates, the aggresomes of spermiogenic origin. Semen ubiquitin content correlates negatively with fertility and conventional semen parameters, and with sperm binding of lectins LCA (Lens culinaris agglutinin; reveals altered sperm surface and PNA (Arachis hypogaea/peanut agglutinin; reveals acrosomal malformation or damage. The Postacrosomal Sheath WWI Domain Binding Protein (PAWP, implicated in oocyte activation during fertilization, is ectopic or absent from defective human and animal spermatozoa. Consequently, FC-parameters of PAWP correlate with ART outcomes in infertile couples and with fertility in bulls. Assays based on the above biomarkers have been combined into multiplex FC semen screening protocols, and the surface expression of lectins and ubiquitin has been utilized to develop nanoparticle-based bull semen purification method validated by field artificial insemination trials. These advances go hand-in-hand with the innovation of FC-technology and genomics/proteomics-based biomarker discovery.

  14. Sialic acid-specific affinity chromatography for the separation of erythropoietin glycoforms using serotonin as a ligand.

    Science.gov (United States)

    Meininger, M; Stepath, M; Hennig, R; Cajic, S; Rapp, E; Rotering, H; Wolff, M W; Reichl, U

    2016-02-15

    Recombinant human erythropoietin (rhEPO) is an important CHO cell-derived glycoprotein and the degree of sialylation of this hormone is crucial for its in vivo bioactivity. In order to improve the purification process serotonin as a potential affinity ligand was tested for preparative chromatographic separation of rhEPO glycoforms into fractions of different degrees of sialylation. Therefore, two chromatographic matrices were prepared by immobilizing serotonin on CNBr- and NHS-Sepharose™. First it was shown both matrices bind rhEPO only in its sialylated form. Results indicate that binding is pH independent between pH 3.5 to 8 suggesting it is not only based on electrostatic interactions. Second, after optimal binding conditions were identified, semi-purified rhEPO was loaded onto both matrices and eluted using a stepwise elution gradient of sodium chloride. For comparison same affinity purification experiments were performed using wheat germ agglutinin-coupled agarose, a lectin known for its affinity towards sialylated glycoproteins. To monitor changes in N-glycan fingerprint, eluate fractions were analyzed by multiplexed capillary gel electrophoresis coupled to laser-induced fluorescence (xCGE-LIF). For the serotonin matrices an increasing degree of sialylation was observed from the first to the third elution fraction while purity of rhEPO could be increased at the same time. The late elution fractions of serotonin-coupled CNBr- and NHS-Sepharose™ also showed an overall sialylation degree exceeding that of the starting material. In contrast, for rhEPO bound to wheat germ agglutinin-coupled agarose, no distinct change in the degree of sialylation could be observed after elution. Overall, these encouraging results highlight the potential of serotonin as a chromatographic ligand for the improvement of pharmaceutical purification processes of rhEPO. PMID:26851523

  15. C1GALT1 enhances proliferation of hepatocellular carcinoma cells via modulating MET glycosylation and dimerization.

    Science.gov (United States)

    Wu, Yao-Ming; Liu, Chiung-Hui; Huang, Miao-Juei; Lai, Hong-Shiee; Lee, Po-Huang; Hu, Rey-Heng; Huang, Min-Chuan

    2013-09-01

    Altered glycosylation is a hallmark of cancer. The core 1 β1,3-galactosyltransferase (C1GALT1) controls the formation of mucin-type O-glycans, far overlooked and underestimated in cancer. Here, we report that C1GALT1 mRNA and protein are frequently overexpressed in hepatocellular carcinoma tumors compared with nontumor liver tissues, where it correlates with advanced tumor stage, metastasis, and poor survival. Enforced expression of C1GALT1 was sufficient to enhance cell proliferation, whereas RNA interference-mediated silencing of C1GALT1 was sufficient to suppress cell proliferation in vitro and in vivo. Notably, C1GALT1 attenuation also suppressed hepatocyte growth factor (HGF)-mediated phosphorylation of the MET kinase in hepatocellular carcinoma cells, whereas enforced expression of C1GALT1 enhanced MET phosphorylation. MET blockade with PHA665752 inhibited C1GALT1-enhanced cell viability. In support of these results, we found that the expression level of phospho-MET and C1GALT1 were associated in primary hepatocellular carcinoma tissues. Mechanistic investigations showed that MET was decorated with O-glycans, as revealed by binding to Vicia villosa agglutinin and peanut agglutinin. Moreover, C1GALT1 modified the O-glycosylation of MET, enhancing its HGF-induced dimerization and activation. Together, our results indicate that C1GALT1 overexpression in hepatocellular carcinoma activates HGF signaling via modulation of MET O-glycosylation and dimerization, providing new insights into how O-glycosylation drives hepatocellular carcinoma pathogenesis. PMID:23832667

  16. Effects of L-Carnitine and Pentoxifylline on Carbohydrate Distribution of Mouse Testicular Sperm Membrane

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    Elham Aliabadi

    2013-06-01

    Full Text Available Background: The glycoconjugate content of sperms indicates their physiological and fertility properties. Lectin reactivity is indicative of intact, capacitated, and acrosome-reacted sperms. In the epididymis, sperms experience maturation, glycoconjugate modification, and simultaneously, higher L-carnitine (LC concentrations. The aim of this project was to evaluate the effects of LC and Pentoxifylline (PF on the integrity, capacitation, and acrosomal reaction of sperms by studying their lectin reactivity.Methods: Mouse testicular sperm samples were divided into three parts. Each sample was added Ham’s F10 (control or media containing 1.76 mM LC or PF. At 30 and 90 minutes after incubation, sperm motility was assessed. Peanut agglutinin (PNA, wheat germ agglutinin (WGA, and Concanavalin A (Con A were used to detect non-acrosome-reacted, non-capacitated, and acrosome-reacted sperms, respectively and the frequency was evaluated by flow cytometry. Statistical analysis was performed using the ANOVA. Results: Sperm motility increased after 30 and 90 minutes of incubation in the LC- and PF-treated cultures (P=0.001. LC administration created a significant increase in the percentage of the non-acrosome-reacted sperms compared to the control sperms after 30 and 90 minutes (P=0.02 and P=0.03, respectively. The frequency of the non-capacitated sperms in the LC-treated group increased compared to the control sperms after 30 minutes significantly (P=0.01. Conclusion: Although the administration of LC and PF enhanced sperm motility, LC also impacted glycoconjugates on the sperm surface. Glycoconjugates are involved in the interaction between the sperm and the zona pellucida and subsequently fertilization, thereby probably influencing the male fertility state.

  17. 衣藻有性生殖的分子机制%Molecular Mechanism of Chlamydomonas Mating

    Institute of Scientific and Technical Information of China (English)

    李修岭; 李夜光

    2009-01-01

    衣藻作为分子生物学研究的模式材料,被广泛用于植物光合作用、鞭毛组装与功能、细胞周期与节律、细胞信号传导与光感受、细胞识别等重要生物学过程的研究,而且衣藻有性生殖的分子机制与人类某些疾病的发生机制存在联系.该文对国内外近年来有关莱茵衣藻在有性生殖过程中凝集素的动态分布,包括鞭毛粘连、补充、传递、脱粘连、凝集素合成的正调节,以及与性凝集素行为有关的基因研究进展进行综述,以阐明衣藻有性生殖的分子机制,为人类的疾病研究提供参考.%As a model organism for studying photosynthesis of plant,flagella assembly and function,cell cycle and circadian rhythms,signal transduction,light perception and cell recognition,Chlamydomonas has been investigated widely.This review reported the advances in molecular mechanism of Chlamydomonas reinhardtii mating.We focused on adhension,tipping,disadhension and regulation of synthesis of sex agglutinins as well as some genes related with adhension of agglutinins.The molecular mechanism of Chlamydomonas mating might provide reference to the study of some human diseases.

  18. Application of fluorescence resonance energy transfer techniques to the study of lectin-binding site distribution on Paramecium primaurelia (Protista, Ciliophora) cell surface.

    Science.gov (United States)

    Locatelli, D; Delmonte Corrado, M U; Politi, H; Bottiroli, G

    1998-01-01

    Fluorescence resonance energy transfer (FRET) is a photophysical phenomenon occurring between the molecules of two fluorochromes with suitable spectral characteristics (donor-acceptor dye pair), and consisting in an excitation energy migration through a non-radiative process. Since the efficiency of the process is strictly dependent on the distance and reciprocal orientation of the donor and acceptor molecules, FRET-based techniques can be successfully applied to the study of biomolecules and cell component organisation and distribution. These techniques have been employed in studying Paramecium primaurelia surface membrane for the reciprocal distribution of N-acetylneuraminic acid (NeuAc) and N-acetylglucosamine (GlcNAc) glycosidic residues, which were found to be involved in mating cell pairing. NeuAc and GlcNAc were detected by their specific binding lectins, Limulus polyphemus agglutinin (LPA) and wheat germ agglutinin (WGA), respectively. Microspectrofluorometric analysis afforded the choice of fluorescein isothiocyanate and Texas red conjugated with LPA and WGA, respectively, as a suitable donor-acceptor couple efficiently activating FRET processes. Studies performed both in solution and in cells allowed to define the experimental conditions favourable for a FRET analysis. The comparative study carried out both on the conjugating-region and the non conjugating region of the surface membrane, indicates that FRET distribution appears quite homogeneous in mating-competent mating type (mt) I, whereas, in mating-competent mt II cells, FRET distribution seems to be preferentially localised on the conjugating-region functionally involved in mating cell pairing. This difference in the distribution of lectin-binding sites is suggested to be related to mating-competence acquisition. PMID:9857246

  19. Characterization of influenza virus sialic acid receptors in minor poultry species

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    Nieto Gloria

    2010-12-01

    Full Text Available Abstract It is commonly accepted that avian influenza viruses (AIVs bind to terminal α2,3 sialic acid (SA residues whereas human influenza viruses bind to α2,6 SA residues. By a series of amino acid changes on the HA surface protein, AIVs can switch receptor specificity and recognize α2,6 SA positive cells, including human respiratory epithelial cells. Animal species, like pigs and Japanese quail, that contain both α2,3 and α2,6 SA become ideal environments for receptor switching. Here, we describe the SA patterns and distributions in 6 common minor domestic poultry species: Peking duck, Toulouse geese, Chinese ring-neck pheasant, white midget turkey, bobwhite quail, and pearl guinea fowl. Lectins specific to α2,3 and α2,6 SA (Maakia amurensis agglutinin and Sambuca nigra agglutinin, respectively were used to detect SA by an alkaline phosphotase-based method and a fluorescent-based method. Differences in SA moieties and their ability to bind influenza viruses were visualized by fluorescent labeling of 4 different H3N2 influenza viruses known to be specific for one receptor or the other. The geese and ducks showed α2,3 SA throughout the respiratory tract and marginal α2,6 SA only in the colon. The four other avian species showed both α2,3 and α2,6 SA in the respiratory tract and the intestines. Furthermore, the turkey respiratory tract showed a positive correlation between age and α2,6 SA levels. The fact that these birds have both avian and human flu receptors, combined with their common presence in backyard farms and live bird markets worldwide, mark them as potential mixing bowl species and necessitates improved surveillance and additional research about the role of these birds in influenza host switching.

  20. Effect of tunicamycin on sialomucin and natural killer susceptibility of rat mammary tumor ascites cells.

    Science.gov (United States)

    Bharathan, S; Moriarty, J; Moody, C E; Sherblom, A P

    1990-09-01

    The MAT-B1 and MAT-C1 ascites sublines of the 13762 rat mammary adenocarcinoma contain a dominant cell surface "complex" consisting of two glycoproteins: ascites sialoglycoprotein (ASGP)-1, a Mr 600,000-700,000 peanut agglutinin-binding sialomucin, and ASGP-2, a Mr 120,000 concancavalin A-binding glycoprotein (Sherblom et al., J. Biol. Chem., 255: 783-790, 1980; Sherblom and Carraway, J. Biol. Chem., 255: 12051-12059, 1980). Although both cell lines are resistant to lysis by natural killer cells, treatments which result in loss of cell surface ASGP-1 render the cells susceptible to natural killer cell lysis (Sherblom and Moody, Cancer Res., 46:4543-4546, 1986). Treatment of the ascites cells with 5 micrograms/ml tunicamycin for 24 h effectively inhibits glycosylation of ASGP-2 without affecting cell viability or total protein synthesis. Under these conditions, expression of ASGP-1 is depressed by at least 50% in both cell lines, as monitored by [3H]glucosamine incorporation and by binding of peanut agglutinin to intact cells. The size distribution of O-linked oligosaccharides in ASGP-1 from tunicamycin-treated versus control MAT-B1 cells is indistinguishable, as determined by Bio-Gel P-4 chromatography following alkaline-borohydride treatment. Complex isolated from either treated or control cells bands at the same density in a CsCl gradient containing Triton X-100 and contains a diffuse band corresponding to ASGP-2 by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Tunicamycin-treated cells, consistent with the reduced expression of ASGP-1, are significantly more susceptible to natural killer cell-mediated lysis, when compared to untreated controls. The results suggest that N-linked glycosylation is a prerequisite for sialomucin synthesis and/or complex formation. PMID:2386935

  1. Effects of noise on the distribution of the cell surface glycoconjugates in the developing mouse spiral ganglion

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    Talaei T

    2007-09-01

    Full Text Available Background: Some pregnant women are exposed to occupational noise, a risk factor for the development of the auditory system. The auditory system is one of the areas in embryonic development in which noise might induce aberrant development. Noise can change the gene expression pattern of an embryo and thereby modify the physiology of the auditory system. Therefore, noise can change the molecular structure of the developing ear. One of the critical molecules involved in development of auditory system is glycoconjugate. The aim of this study was to investigate the molecular changes of the developing spiral ganglion after exposure to industrial levels of noise.Methods: A total of 42 pregnant mice were divided into control and experimental groups. Each group was further divided into three subgroups. The three experimental subgroups were exposed to daily noise with an intensity of 100 db for 2.5 hours until sacrifice (for the first group to be sacrificed or day seven of postnatal life (for the other two groups. The mice offspring were sacrificed at the first, seventh and 14th days of postnatal life. The inner ears were prepared histologically. The specimens were stained with the lectins wheat germ antigen (WGA, peanut agglutinin (PNA, Dolichos biflorus agglutinin (DBA and BSAI-B4. Results: The results indicated that, although there were no histological changes at the light-microscopic level in the ear development, statistical analysis showed that there was a significant decrease in the uptake of the BSA1-B4 lectin by neurons of spiral ganglion in 14th day of postnatal life in the experimental group compared to  that of the control group (p<0.05. Conclusions: After noise exposure, in spite of normal neuronal structure, these cells were modified at the molecular level, especially in glycoconjugate expression, influencing the normal physiology of neurons and causing auditory disorders.

  2. Cytoskeletal inhibitors, anti-adhesion molecule antibodies, and lectins inhibit hepatocyte spheroid formation.

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    Nakamura M

    2002-02-01

    Full Text Available We investigated the role of cytoskeletons, adhesion molecules, membrane-glycosylations, and proteoglycans in forming the shape of adult rat hepatocyte spheroids. Isolated hepatocytes were cultured on dishes coated with chondroitin sulfate phosphatidyl ethanolamine (CS-PE. Spheroid-forming ability was observed after adding cytoskeletal inhibitors (cytochalasin D, colchicine, okadaic acid, mycalolide B, anti-adhesion molecule antibodies (anti-E-cadherin, anti-connexin 32, anti-zo-1, a glycosphingolipid synthetic inhibitor (N-butyldeoxynojirimycin, a proteoglycan synthetic inhibitor (p-nitrophenyl-beta-D-xylopyranoside, and several lectins. Localization of actin was studied using confocal microscopy after rhodamine-phalloidin staining. Adding cytoskeletal inhibitors on the initial day resulted in weakly clustered cell aggregates rather than smoothly formed spheroids. These effects disappeared at lower reagent concentrations. When reagents were added on day 3, after the formation of spheroids, only mycalolide B was associated with an irregular spheroid surface; the others had no effect. Adding the anti-E-cadherin, anti-connexin 32 on the initial day showed inhibition of spheroid formation, but anti-zo-1 and proteoglycan synthetic inhibitor had no effects. Among the several lectins, only Wheat Germ Agglutinin (WGA, Ricinus communis Agglutinin I (RCA-I, and Concanavalin A (ConA showed inhibition. These results suggest that cytoskeletal conformation and some adhesion molecules are necessary to form spheroids. Based on the interactions between lectins and hepatocytes in the present study, hepatocytes appear to contain an N-linked complex or N-linked hybrid glycosylated chains.

  3. Our first experiences in applying an original method for removal of ABO-isoagglutinins in ABO-incompatible kidney recipients

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    Ignjatović Ljiljana

    2009-01-01

    Full Text Available Background/Aim. Due to improved methods for removal of ABO isoagglutinins and novel immunosuppressive protocols, short and long term outcome in blood group incompatible is similar to blood group compatible kidney transplantation. The aim of this study was to determine the efficacy of our original method for removal of ABO isoagglutinins from the blood in ABO-incompatible kidney allograft recipients. Method. Between 2006 and 2008 twelve patients were transplanted from ABO incompatible living donors. Titers of ABO isoagglutinins were 4-128 (IgG. Immunosuppressive therapy started 14 days before kidney transplantation with rituximab, followed by a triple therapy (prednisone + tacrolimus + mycophenolate mofetil and the first plasma exchange (PE procedure, in which one plasma volume was substituted with albumin and saline on day 7 before transplantation. For selective extracorporeal immunoadsorption, the removed plasma was mixed with donor blood type filtered red blood cells, centrifuged and the supernatant separated and preserved. In the next PE procedure, the removed plasma was replaced with immunoadsorbed plasma, and so on. Titers of ABO agglutinins, renal allograft function and survival were followed-up. Results. The pre-transplant treatment consisting of 1-5 PE procedures and immunosuppressive therapy resulted in target ABO agglutinins titers below 4. During a 10-24 month follow-up three patients had an early acute rejection, one patient acute rejection and hemolytic anemia, two patients surgical complications and one of them lost his graft. In the post-transplant period, the titers of ABO antibodies remained below 4. All the patients had stable kidney allograft function with mean serum creatinine ±SD of 129 ± 45 μmol/l at the end of the study. Conclusion. Our method for removal of ABO antibodies was effective in a limited series of patients and short-term follow-up.

  4. Monosaccharide profiling of silkworm (Bombyx mori L.) nervous system during development and aging.

    Science.gov (United States)

    Soya, Seçkin; Şahar, Umut; Karaçalı, Sabire

    2016-09-01

    Glycoconjugates have various functions in differentiation, development, aging and in all aspects of normal functioning of organisms. The reason for increased research on this topic is that glycoconjugates locate mostly on the cell surface and play crucial biological roles in the nervous system including brain development, synaptic plasticity, learning, and memory. Considering their roles in the nervous system, information about their existence in the insect nervous system is rather sparse. Therefore, in order to detect monosaccharide content of N- and O-glycans, we carried out capLC-ESI-MS/MS analysis to determine the concentration changes of glucose, mannose, galactose, N-acetylglucosamine (GlcNAc), N-acetylgalactosamine (GalNAc), fucose, xylose, arabinose, and ribose monosaccharides in the nervous system of Bombyx mori during development and aging processes. In addition to LC-MS, lectin blotting was done to detect quantitative changes in N- and O-glycans. Developmental stages were selected as 3rd (the youngest sample), 5th (young) larval instar, motionless prepupa (the oldest sample), and pupa (adult development). Derivatization of monosaccharides was performed with a solution of PMP agent and analyzed with capLC-ESI-MS/MS. For lectin blotting, determination of glycan types was carried out with Galanthus nivalis agglutinin and Peanut agglutinin lectins. In all stages, the most abundant monosaccharide was glucose. Although all monosaccharides were present most abundantly in the youngest stage (3rd instar), they are generally reduced gradually during the aging process. It was observed that amounts of monosaccharides increased again in the pupa stage. According to lectin blotting, N- and O-linked glycoproteins expressions were different and there were some specific glycoprotein expression differences between stages. These findings suggest that the glycosylation state of proteins in the nervous system changes during development and aging in insects in a similar

  5. Difference of clinical features in childhood Mycoplasma pneumoniae pneumonia

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    Kang Jin-Han

    2010-07-01

    Full Text Available Abstract Background M. pneumoniae pneumonia (MP has been reported in 10-40% of community-acquired pneumonia cases. We aimed to evaluate the difference of clinical features in children with MP, according to their age and chest radiographic patterns. Methods The diagnosis of MP was made by examinations at both admission and discharge and by two serologic tests: the indirect microparticle agglutinin assay (≥1:40 and the cold agglutinins titer (≥1:32. A total of 191 children with MP were grouped by age: ≤2 years of age (29 patients, 3-5 years of age (81 patients, and ≥6 years of age (81 patients. They were also grouped by pneumonia pattern: bronchopneumonia group (96 patients and segmental/lobar pneumonia group (95 patients. Results Eighty-six patients (45% were seroconverters, and the others showed increased antibody titers during hospitalization. Among the three age groups, the oldest children showed the longest duration of fever, highest C-reactive protein (CRP values, and the most severe pneumonia pattern. The patients with segmental/lobar pneumonia were older and had longer fever duration and lower white blood cell (WBC and lymphocyte counts, compared with those with bronchopneumonia. The patient group with the most severe pulmonary lesions had the most prolonged fever, highest CRP, highest rate of seroconverters, and lowest lymphocyte counts. Thrombocytosis was observed in 8% of patients at admission, but in 33% of patients at discharge. Conclusions In MP, older children had more prolonged fever and more severe pulmonary lesions. The severity of pulmonary lesions was associated with the absence of diagnostic IgM antibodies at presentation and lymphocyte count. Short-term paired IgM serologic test may be mandatory for early and definitive diagnosis of MP.

  6. Clinical and epidemiological features of typhoid fever in Pemba, Zanzibar: assessment of the performance of the WHO case definitions.

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    Kamala Thriemer

    Full Text Available BACKGROUND: The gold standard for diagnosis of typhoid fever is blood culture (BC. Because blood culture is often not available in impoverished settings it would be helpful to have alternative diagnostic approaches. We therefore investigated the usefulness of clinical signs, WHO case definition and Widal test for the diagnosis of typhoid fever. METHODOLOGY/PRINCIPAL FINDINGS: Participants with a body temperature ≥37.5°C or a history of fever were enrolled over 17 to 22 months in three hospitals on Pemba Island, Tanzania. Clinical signs and symptoms of participants upon presentation as well as blood and serum for BC and Widal testing were collected. Clinical signs and symptoms of typhoid fever cases were compared to other cases of invasive bacterial diseases and BC negative participants. The relationship of typhoid fever cases with rainfall, temperature, and religious festivals was explored. The performance of the WHO case definitions for suspected and probable typhoid fever and a local cut off titre for the Widal test was assessed. 79 of 2209 participants had invasive bacterial disease. 46 isolates were identified as typhoid fever. Apart from a longer duration of fever prior to admission clinical signs and symptoms were not significantly different among patients with typhoid fever than from other febrile patients. We did not detect any significant seasonal patterns nor correlation with rainfall or festivals. The sensitivity and specificity of the WHO case definition for suspected and probable typhoid fever were 82.6% and 41.3% and 36.3 and 99.7% respectively. Sensitivity and specificity of the Widal test was 47.8% and 99.4 both forfor O-agglutinin and H- agglutinin at a cut-off titre of 1:80. CONCLUSIONS/SIGNIFICANCE: Typhoid fever prevalence rates on Pemba are high and its clinical signs and symptoms are non-specific. The sensitivity of the Widal test is low and the WHO case definition performed better than the Widal test.

  7. Nucleocytoplasmic transport blockage by SV40 peptide-modified gold nanoparticles induces cellular autophagy

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    Tsai TL

    2012-10-01

    Full Text Available Tsung-Lin Tsai,1,5 Chia-Cheng Hou,1,5 Hao-Chen Wang,1,5 Zih-Syuan Yang,2 Chen-Sheng Yeh,4 Dar-Bin Shieh,2,3 Wu-Chou Su1,51Institute of Basic Medical Sciences, 2Institute of Oral Medicine and Department of Stomatology, 3Center for Micro/Nano Science and Technology, 4Department of Chemistry, National Cheng Kung University, Tainan, Taiwan; 5Department of Internal Medicine, National Cheng Kung University Hospital, College of Medicine, National Cheng Kung University, Tainan, TaiwanAbstract: Gold nanoparticles modified with the nuclear localization signal from simian virus 40 large T antigen (GNP-PEG/SV40 accumulate on the cytoplasmic side of the nuclear membrane in HeLa cells. Accumulation of GNP-PEG/SV40 around the nucleus blocks nucleocytoplasmic transport and prevents RNA export and nuclear shuttling of signaling proteins. This long-term blockage of nucleocytoplasmic transport results in cell death. This cell death is not caused by apoptosis or necrosis because caspases 3 and 9 are not activated, and the expression of annexin V/propidium iodide is not enhanced in HeLa cells after treatment. Using transmission electron microscopy, autophagosomes and autolysosomes were seen to appear after 72 hours of treatment with GNP-PEG/SV40. Increasing levels of enhanced green fluorescent protein-microtubule-associated protein 1 light chain 3 (EGFP-LC3-positive punctate and LC3-II confirmed GNP-PEG/SV40-induced autophagy. In SiHa cells, treatment did not induce accumulation of GNP-PEG/SV40 around the nucleus and autophagy. Treating cells with wheat germ agglutinin, a nuclear pore complex inhibitor, induced autophagy in both HeLa and SiHa cells. GNP-PEG/SV40-induced autophagy plays a role in cell death, not survival, and virus-mediated small hairpin RNA silencing of Beclin-1 attenuates cell death. Taken together, the results indicate that long-term blockade of nucleocytoplasmic transport results in autophagic cell death.Keywords: gold nanoparticles

  8. Differential expression of glycans in the hippocampus of rats trained on an inhibitory learning paradigm.

    Science.gov (United States)

    Hidalgo, Alejandra; Burgos, Valeria; Viola, Haydée; Medina, Jorge; Argibay, Pablo

    2006-12-01

    The glycan chains of glycoconjugates play important roles in cell-cell and cell-matrix interactions. In the CNS, previous studies on learning and memory suggest the importance of oligosaccharides attached to glycoconjugates in the modulation of synaptic connections. We studied the hippocampal glycan distribution of rats subject to an inhibitory avoidance task. The expression of glycans was examined by lectin-histochemistry using Vicia villosa lectin (VVL) for terminal alpha/beta N-acetylgalactosamine (alpha/beta GalNAc); Galanthus nivalus lectin (GNL) for terminal mannose alpha-1,3 (Man alpha-1,3); Peanut agglutinin (PNA) for galactose beta-1,3N-acetylgalactosamine (Gal beta-1,3 GalNAc); Erythrina cristagalli lectin (ECL) for galactose beta-1,4 N-acetylglucosamine (Gal beta-1,4 GlcNAc); Sambucus nigra lectin (SNA) for sialic acid alpha-2.6 galactose (SA alpha-2,6 Gal); Maackia amurensis lectin II (MAL II) for sialic acid alpha-2,3 (SA alpha-2,3); Wheat germ agglutinin (WGA) for terminal N-acetylglucosamine with/ without sialic acid (GlcNAc wo SA); succynilated WGA (sWGA) for terminal N-acetylglucosamine without sialic acid (terminal GlcNAc without SA); Griffonia simplicifolia lectin II (GSL II) for terminal alpha/beta N-acetylglucosamine (alpha/beta GlcNAc terminal); and Lotus tetragonolobus lectin (LTL) alpha-fucose. Two groups of 10 animals were examined: non-trained (Control) and Trained rats. ECL, sWGA and GSL II were negative for both groups in all the hippocampal subfields studied. For both groups, VVL was negative in CA4 and granular cells of the Dentate Gyrus (DG) and LTL was negative in the CA4 subfield. Expression of alpha/beta GalNAc, alpha-fucose and GlcNAc in other hippocampal subflields was positive, with no differences between groups. However, expression of Man alpha-1,3 was significantly higher in the CA1, CA2, CA3, and CA4 subfields in the Trained group. On the other hand, expression of Gal beta-1,3 GalNAc was significantly low in CA4 and DG in the

  9. Case report: diffuse splenic metastasis of occult breast cancer with incompatible blood group antigenic determinants.

    Science.gov (United States)

    Baranyay, Ferenc

    2009-01-01

    Cancer cells with immunogenic properties having altered protein glycosilation, modified blood group substances have been widely studied [Kannagi R, Miyake M, Zenita KM, Itai S, Hiraiwa N, Shigeta K, et al. Cancer-associated carbohydrate antigens: modified blood group substances and oncodevelopmental antigens on tumor cells. Gann Monogr Cancer Res 1988; 34: p. 15-28; Hakomori S. Antigen structure and genetic basis of histo-blood groups A, B and O their changes associated with human cancer. Biochem Biophys Acta 1999; 1473: p. 247-266; Brooks SA, Carter TM, Royle L, Harvey DJ, Fry SA, Kinch C, et al. Altered glycosilation of proteins in cancer: what is the potential for new anti-tumour strategies. Anticancer Agents Med Chem 2008; 8: p. 2-21]. In the study reported here, a 78-year-old female patient was admitted to the hospital with circulatory failure. At autopsy, the spleen (weight: 420 g) was extremely firm with a diffusely blackberry-colored cut surface. There were no signs of carcinomatous process at autopsy. By histology, the spleen showed diffuse metastatic carcinomatous infiltration. Using immunohistochemistry, an antibody to breast carcinoma antigen (BioGenex) labelled metastatic cells of the spleen and bone marrow. The patient was blood group O. Labelling for binding of lectins with and without blood group antigen specificity and monoclonal antibodies was carried out. The B blood group specific Banderiaea simplicifolia agglutinin I and an anti-B blood group monoclonal antibody labelled all the metastatic cells of spleen and bone marrow intensely. There was no detection of blood group A antigen by either binding of Dolichos biflorus agglutinin or anti-blood group A monoclonal antibodies. These observations raise the possibility that the detected incompatible B blood group antigen determinants on the metastatic cells were immunogenic. The surviving carcinoma cells may have found a place of refuge from immune surveillance in the spleen and in the bone marrow

  10. AUTOIMMUNE CYTOPENIAS IN CHRONIC LYMPHOCYTIC LEUKEMIA, FACTS AND MYTHS

    Directory of Open Access Journals (Sweden)

    Pavankumar Tandra

    2013-11-01

    Full Text Available CLL has been defined as presence of more than 5000 small mature appearing monoclonal B lymphocytes with a specific immunophenotype in peripheral blood. It is a well-known fact that CLL is associated with autoimmune cytopenias. CLL cells are CD5+ B lymphocytes, and usually are not the “guilty” cells which produce autoantibodies. T cell defect is another characteristic of CLL and the total number of T cells is increased, and there is inversion of the CD4/CD8 ratio. Autoimmune hemolytic anemia (AIHA is the most common autoimmune complication of CLL and has been reported in 10-25% of CLL patients. However, the stage-adjusted estimated rate of AIHA in CLL is about 5%. Conversely, CLL is three times more common in patients who present with AIHA. Direct agglutinin test (DAT is positive in 7-14% of CLL patients but AIHA may also occur in DAT negative patients. Autoimmune thrombocytopenia (AIT is the second most common complication of CLL and has been reported in 2-3% of patients. DAT is positive in AIT but presence of antiplatelet antibodies is neither diagnostic nor reliable. Autoimmune neutropenia (AIN and pure red cell aplasia (PRCA are very rare complications of CLL and like other autoimmune complications of CLL may occur at any clinical stage. It is believed that most case reports of AIN and PRCA in CLL actually belong to large granular lymphocytic leukemia (LGL. Non-hematologic autoimmune complications of CLL including cold agglutinin disease (CAD, paraneoplastic pemphigus (PNP, acquired angioedema, and anti-myelin associated globulin are rare. Before starting any treatment, clinicians should distinguish between autoimmune cytopenias and massive bone marrow infiltration since autoimmune complications of CLL are not necessarily equal to advanced disease with poor prognosis. According to IWCLL guideline, steroids are the mainstay of treatment of simple autoimmunity. Intravenous immunoglobulin (IVIg, cyclosporine, and rituximab are used in

  11. N-Glycans in Xenopus laevis testis characterised by lectin histochemistry.

    Science.gov (United States)

    Valbuena, Galder; Madrid, Juan Francisco; Martínez de Ubago, María; Gómez-Santos, Laura; Alonso, Edurne; Díaz-Flores, Lucio; Sáez, Francisco J

    2016-03-01

    Analysis of glycan chains of glycoconjugates is difficult because of their considerable variety. Despite this, several functional roles for these glycans have been reported. N-Glycans are oligosaccharides linked to asparagine residues of proteins. They are synthesised in the endoplasmic reticulum (ER) in a unique way, and later modified in both the ER and Golgi apparatus, developing different oligosaccharide chains. An essential role for complex N-glycans in mammalian spermatogenesis has been reported. The aim of the present study was to analyse the N-glycans of the Xenopus laevis testis by means of lectin histochemistry. Five lectins were used that specifically recognise mannose-containing and complex glycans, namely Galanthus nivalis agglutinin (GNA) from snowdrops, concanavalin A (Con A) from the Jack bean, Lens culinaris agglutinin (LCA) from lentils and Phaseolus vulgaris erythroagglutinin (PHA-E) and P. vulgaris leukoagglutinin (PHA-L) from the common bean. GNA and Con A labelled the interstitium and most of the germ cell types, whereas LCA and PHA-E showed affinity only for the interstitium. A granular cytoplasmic region was labelled in spermatogonia and spermatocytes by GNA and PHA-L, whereas GNA and LCA labelled a spermatid region that is probably associated with the centriolar basal body of the nascent flagellum. There was no specific labelling in the acrosome. Some unexpected results were found when deglycosylative pretreatments were used: pre-incubation of tissue sections with peptide N glycosidase F, which removes N-linked glycans, reduced or removed labelling with most lectins, as expected. However, after this pretreatment, the intensity of labelling remained or increased for Con A in the follicle (Sertoli) and post-meiotic germ cells. The β-elimination procedure, which removes O-linked glycans, revealed new labelling patterns with GNA, LCA and PHA-L, suggesting that some N-glycans were masked by O-glycans, and thus they became accessible to these

  12. Study on cell surface display of β-amylase on Saccharomces cerevisiae and its practical properties%酿酒酵母表面展示β-淀粉酶及其酶学性质研究

    Institute of Scientific and Technical Information of China (English)

    杨华; 樊游; 沈微; 石贵阳; SurenSingh; 王正祥

    2012-01-01

    通过PCR技术扩增来源于大麦的β-淀粉酶基因,将其与酿酒酵母细胞壁蛋白d凝集素基因在读框内融合,构建得到表面展示载体pBA-AG,进一步将该重组质粒通过遗传转化,整合到酿酒酵母W303-1A的染色体中,获得了β-淀粉酶经过仅凝集素锚定信号结合到细胞壁上的重组酵母。重组酵母表面展示的β-淀粉酶活力为131U/g干细胞。对展示的β-淀粉酶酶学性质研究表明,其最适反应温度为50℃,最适作用pH为5.0,与游离酶相比,其温度稳定性和pH稳定性均得到提高。本研究利用α凝集素系统首次将β-淀粉酶成功展示在酿酒酵母表面,为以酿酒酵母为基础的全细胞催化剂研究与应用打下了一定基础.%The gene encoding mature β-amylase from barley was cloned via PCR and then fused with the α-agglutinin of Saccharomyces cerevisiae in frame and a recombinant plasmid named pBA-AG was constructed. Recombinant plasmid pBA-AG was successfully transformed into S. cerevisiae W303-1A and was integrated into the chromosome. The expressed β-amylase was successfully anchored on the cell wall and displayed on the surface of recombinant S. cerevisiae. The measured activity of displayed β-amylase was 131U/g dry cells. The optimal temperature and pH for displayed β-amylase was 50℃ and 5.0,respectively. The recombinant β-amylase displayed on cell surface exhibited enhanced thermostability compared to free enzyme. In this study,the firstly constructed recombinant S. cerevisiae strain displaying β-amylase on the cell surface with a-agglutinin as carrier protein showed a great potential for industrial application as a whole-cell biocatalyst.

  13. Sialic acid receptor detection in the human respiratory tract: evidence for widespread distribution of potential binding sites for human and avian influenza viruses

    Directory of Open Access Journals (Sweden)

    Guan Yi

    2007-10-01

    Full Text Available Abstract Background Influenza virus binds to cell receptors via sialic acid (SA linked glycoproteins. They recognize SA on host cells through their haemagglutinins (H. The distribution of SA on cell surfaces is one determinant of host tropism and understanding its expression on human cells and tissues is important for understanding influenza pathogenesis. The objective of this study therefore was to optimize the detection of α2,3-linked and α2,6-linked SA by lectin histochemistry by investigating the binding of Sambucus nigra agglutinin (SNA for SAα2,6Gal and Maackia amurensis agglutinin (MAA for SAα2,3Gal in the respiratory tract of normal adults and children. Methods We used fluorescent and biotinylated SNA and MAA from different suppliers on archived and prospectively collected biopsy and autopsy specimens from the nasopharynx, trachea, bronchus and lungs of fetuses, infants and adults. We compared different methods of unmasking for tissue sections to determine if these would affect lectin binding. Using serial sections we then compared the lectin binding of MAA from different suppliers. Results We found that unmasking using microwave treatment in citrate buffer produced increased lectin binding to the ciliated and glandular epithelium of the respiratory tract. In addition we found that there were differences in tissue distribution of the α2,3 linked SA when 2 different isoforms of MAA (MAA1 and MAA2 lectin were used. MAA1 had widespread binding throughout the upper and lower respiratory tract and showed more binding to the respiratory epithelium of children than in adults. By comparison, MAA2 binding was mainly restricted to the alveolar epithelial cells of the lung with weak binding to goblet cells. SNA binding was detected in bronchial and alveolar epithelial cells and binding of this lectin was stronger to the paediatric epithelium compared to adult epithelium. Furthermore, the MAA lectins from 2 suppliers (Roche and EY Labs tended

  14. Evidence for lectin activity of a plant receptor-like protein kinase by application of neoglycoproteins and bioinformatic algorithms.

    Science.gov (United States)

    André, Sabine; Siebert, Hans-Christian; Nishiguchi, Mitsuru; Tazaki, Kiyoshi; Gabius, Hans-Joachim

    2005-09-15

    Detection of genes for putative receptor-like protein kinases, which contain an extracellular domain related to leguminous lectins, in plant genomes inspired the hypothesis that this part acts as sensor. Initial support for this concept came from proof for protein kinase activity. The next step, focusing on the protein of lombardy poplar (Populus nigra var. italica), is scrutiny for lectin activity. Consequently, we first pinpointed sets of high-scoring sequence pairs by extensive databank search. The calculations resulted in P-values in the range from 10(-14) to 10(-18) exclusively for leguminous lectins, the Pterocarpus angolensis agglutinin being front runner with P=3 x 10(-18) and thus most suitable template for modeling. The superimposition of the two folds gave notable similarity in the region responsible for binding carbohydrate and Ca(2+)/Mn(2+)-ions. Binding activity toward carbohydrates was detected by assaying a panel of (neo)glycoproteins as polyvalent probes, especially for alpha-l-rhamnose and glycans of asialofetuin. It was strictly dependent on Ca(2+)-ions, enhanced by Mn(2+)-ions and reached a K(D)-value of 34.3 nM for the neoglycoprotein with rhamnose as ligand. These results give further research direction to define physiological ligands, plant/bacterial rhamnose-containing saccharides and rhamnose-mimetic glycans or peptides being potential candidates. PMID:15878637

  15. Intracellular iron concentration of neurons with and without perineuronal nets

    Energy Technology Data Exchange (ETDEWEB)

    Fiedler, Anja [Paul Flechsig Institute for Brain Research, University of Leipzig, Jahnallee 59, D-04109 Leipzig (Germany) and Institute for Experimental Physics II, University of Leipzig, Linnestrasse 5, D-04103 Leipzig (Germany)]. E-mail: afiedler@uni-leipzig.de; Reinert, Tilo [Institute for Experimental Physics II, University of Leipzig, Linnestrasse 5, D-04103 Leipzig (Germany); Morawski, Markus [Paul Flechsig Institute for Brain Research, University of Leipzig, Jahnallee 59, D-04109 Leipzig (Germany); Brueckner, Gert [Paul Flechsig Institute for Brain Research, University of Leipzig, Jahnallee 59, D-04109 Leipzig (Germany); Arendt, Thomas [Paul Flechsig Institute for Brain Research, University of Leipzig, Jahnallee 59, D-04109 Leipzig (Germany); Butz, Tilman [Institute for Experimental Physics II, University of Leipzig, Linnestrasse 5, D-04103 Leipzig (Germany)

    2007-07-15

    Neurodegenerative diseases like Parkinson's disease, Alzheimer's disease and Huntington's disease are characterized by abnormally high concentrations of iron in the affected brain areas. Iron is believed to contribute to oxidative stress by catalysing radical generation and subsequently causing neuronal death. Interestingly, subpopulations of neurons are less vulnerable against degeneration. One of these subpopulations possesses a specialized extracellular matrix arranged as a perineuronal net (PN), a structure with poorly understood functions. In order to differentiate between neurons with and without PN according to their iron concentrations we have performed a {mu}PIXE study at the Leipzig LIPSION laboratory. PN-ensheathed neurons in selected brain areas were detected by lectin-histochemical staining with Wisteria floribunda agglutinin (WFA). The staining was intensified by DAB-nickel by an established method enabling the visualisation of the PNs by nuclear microscopy. The cellular concentration of iron in the rat brain was about 1 mmol/l (ca. 30 {mu}g/g dw). First results of subcellular analysis showed that the intracellular iron concentration of PN-ensheathed neurons tends to be slightly increased in comparison to neurons without PNs. The difference in intracellular iron concentrations could be an effect of the PNs.

  16. Intracellular iron concentration of neurons with and without perineuronal nets

    International Nuclear Information System (INIS)

    Neurodegenerative diseases like Parkinson's disease, Alzheimer's disease and Huntington's disease are characterized by abnormally high concentrations of iron in the affected brain areas. Iron is believed to contribute to oxidative stress by catalysing radical generation and subsequently causing neuronal death. Interestingly, subpopulations of neurons are less vulnerable against degeneration. One of these subpopulations possesses a specialized extracellular matrix arranged as a perineuronal net (PN), a structure with poorly understood functions. In order to differentiate between neurons with and without PN according to their iron concentrations we have performed a μPIXE study at the Leipzig LIPSION laboratory. PN-ensheathed neurons in selected brain areas were detected by lectin-histochemical staining with Wisteria floribunda agglutinin (WFA). The staining was intensified by DAB-nickel by an established method enabling the visualisation of the PNs by nuclear microscopy. The cellular concentration of iron in the rat brain was about 1 mmol/l (ca. 30 μg/g dw). First results of subcellular analysis showed that the intracellular iron concentration of PN-ensheathed neurons tends to be slightly increased in comparison to neurons without PNs. The difference in intracellular iron concentrations could be an effect of the PNs

  17. Cell Surface Display of Four Types of Solanum nigrum Metallothionein on Saccharomyces cerevisiae for Biosorption of Cadmium.

    Science.gov (United States)

    Wei, Qinguo; Zhang, Honghai; Guo, Dongge; Ma, Shisheng

    2016-05-28

    We displayed four types of Solanum nigrum metallothionein (SMT) for the first time on the surface of Saccharomyces cerevisiae using an α-agglutinin-based display system. The SMT genes were amplified by RT-PCR. The plasmid pYES2 was used to construct the expression vector. Transformed yeast strains were confirmed by PCR amplification and custom sequencing. Surface-expressed metallothioneins were indirectly indicated by the enhanced cadmium sorption capacity. Flame atomic absorption spectrophotometry was used to examine the concentration of Cd(2+) in this study. The transformed yeast strains showed much higher resistance ability to Cd(2+) compared with the control. Strikingly, their Cd(2+) accumulation was almost twice as much as that of the wild-type yeast cells. Furthermore, surface-engineered yeast strains could effectively adsorb ultra-trace cadmium and accumulate Cd(2+) under a wide range of pH levels, from 3 to 7, without disturbing the Cu(2+) and Hg(2+). Four types of surfaceengineered Saccharomyces cerevisiae strains were constructed and they could be used to purify Cd(2+)-contaminated water and adsorb ultra-trace cadmium effectively. The surface-engineered Saccharomyces cerevisiae strains would be useful tools for the bioremediation and biosorption of environmental cadmium contaminants. PMID:26838339

  18. A common sugar-nucleotide-mediated mechanism of inhibition of (glycosamino)glycan biosynthesis, as evidenced by 6F-GalNAc (Ac3).

    Science.gov (United States)

    van Wijk, Xander M; Lawrence, Roger; Thijssen, Victor L; van den Broek, Sebastiaan A; Troost, Ran; van Scherpenzeel, Monique; Naidu, Natasha; Oosterhof, Arie; Griffioen, Arjan W; Lefeber, Dirk J; van Delft, Floris L; van Kuppevelt, Toin H

    2015-07-01

    Glycosaminoglycan (GAG) polysaccharides have been implicated in a variety of cellular processes, and alterations in their amount and structure have been associated with diseases such as cancer. In this study, we probed 11 sugar analogs for their capacity to interfere with GAG biosynthesis. One analog, with a modification not directly involved in the glycosidic bond formation, 6F-N-acetyl-d-galactosamine (GalNAc) (Ac3), was selected for further study on its metabolic and biologic effect. Treatment of human ovarian carcinoma cells with 50 μM 6F-GalNAc (Ac3) inhibited biosynthesis of GAGs (chondroitin/dermatan sulfate by ∼50-60%, heparan sulfate by ∼35%), N-acetyl-d-glucosamine (GlcNAc)/GalNAc containing glycans recognized by the lectins Datura stramonium and peanut agglutinin (by ∼74 and ∼43%, respectively), and O-GlcNAc protein modification. With respect to function, 6F-GalNAc (Ac3) treatment inhibited growth factor signaling and reduced in vivo angiogenesis by ∼33%. Although the analog was readily transformed in cells into the uridine 5'-diphosphate (UDP)-activated form, it was not incorporated into GAGs. Rather, it strongly reduced cellular UDP-GalNAc and UDP-GlcNAc pools. Together with data from the literature, these findings indicate that nucleotide sugar depletion without incorporation is a common mechanism of sugar analogs for inhibiting GAG/glycan biosynthesis. PMID:25868729

  19. Phenotypic and cytogenetic characteristics of a new Epstein-Barr virus negative cell line (SKW 4) derived from a B-cell lymphoma.

    Science.gov (United States)

    Nilsson, K; Klareskog, L; Ralph, P; Sundström, C; Zech, L

    1983-01-01

    A new Epstein-Barr virus nuclear antigen (EBNA) negative cell line SKW 4 has been established in vitro from a patient with diffuse histiocytic lymphoma. The SKW 4 seems to be an authentic human tumour cell line as evidenced by its EBV negativity, monoclonality and aneuploidy tested during early in vitro passage. The cell line expresses surface mu and kappa-chains, HLA-DR antigen, C3 and Fc receptors and B-cell lineage antigens. The karyotypic analyses demonstrated many numerical and structural aberrations. No Burkitt lymphoma associated translocations (t8;14, t2;8, t;22) were detected, but most of the markers found are those commonly associated with various types of human cancer. The SKW 4 thus represents the most common type of 'histiocytic lymphoma', that with a B-lymphoid cell phenotype, but is unique among HL derived lymphoma lines in its strong expression of a Helix pomatia A agglutinin binding surface glycoprotein of an apparent molecular weight of 75 000 daltons. PMID:6329938

  20. Long-term exposure to arsenic affects head kidney and impairs humoral immune responses of Clarias batrachus

    International Nuclear Information System (INIS)

    The present study was aimed at determining the effects of long-term arsenic exposure on the head kidney (HK) and ensuing humoral immune responses in Clarias batrachus L. Long-term exposure (150 days) to non-lethal concentrations of arsenic (42.42 μM) resulted in significant time-dependent alterations in HK cell number eventually affecting the HK somatic index. Prolonged exposure to arsenic also suppressed HK-B cell proliferation and led to significant reduction in serum immunoglobulin levels and antigen-specific serum bacterial agglutinin titers. A decline in the number of antigen-specific plaque-forming cells with duration of arsenic exposure was noted in the HK. Enzyme linked immunosorbent assays further revealed that arsenic exposure inhibited the release of 'IL-4 like factors' from HK-T cells. Histological studies documented time-dependent changes in the structure and cellular composition of HK characterized by extensive lymphocytopenia, decrease in melano-macrophage population and hemosiderin accumulation. From exposure-challenge studies with Aeromonas hydrophila it was evident that pathogens could efficiently disseminate and colonize distant host tissues in the exposed fish. Moreover, the ability to decrease the pathogen load was also significantly reduced in the arsenic-exposed fish. Thus long-term exposure to non-lethal concentrations of arsenic affects HK and interferes with the humoral immune system of C. batrachus rendering them immunocompromised and susceptible to pathogenic challenge

  1. Agonist-promoted desensitization and phosphorylation of α1-adrenergic receptors coupled to stimulation of phosphatidylinositol metabolism

    International Nuclear Information System (INIS)

    In the DDT1 MF-2 hamster vas deferens smooth muscle cell line the α1-adrenergic receptor (α1-AR) agonist norepinephrine (NE) promotes rapid attenuation of α1-AR-mediated phosphatidylinositol (PI) metabolism which is paralleled by rapid phosphorylation of the α1-AR. Cells were labeled by incubation with 32P/sub i/. Coincubation with NE (100 μM) significantly increases the rate of 32P-labeling of both PI and phosphatidic acid. Pretreatment of cells with 100 μM NE (in the presence of 1 μM propranolol to prevent β-AR interactions) results in a drastic attenuation of the NE response on PI metabolism. α1-AR from labeled cells can be solubilized and purified by affinity chromatography on Affigel-A55414 and wheat germ agglutinin agarose chromatography. SDS-PAGE of purified α1-AR shows a NE-promoted increase in phosphorylation of the M/sub r/ 80K ligand binding peptide. Stoichiometry of phosphorylation increases from ∼ 1 mol phosphate/mol α1-AR in the basal condition to ∼ 2.5 after NE treatment. Both desensitization and phosphorylation are rapid being maximal within 10-20 min of agonist exposure. These results together with previous findings that phorbol esters promote rapid α1-AR uncoupling and phosphorylation suggest that receptor phosphorylation is an important mechanism of regulation of α1-AR receptor responsiveness

  2. Direct evidence that ganglioside is an integral component of the thyrotropin receptor

    International Nuclear Information System (INIS)

    Gangliosides were extracted from purified human and porcine thyrotropin (TSH) receptors (TSH-R) and were detected by probing with an 125I-labeled sialic acid-specific lectin, Limax flavus agglutinin. Gangliosides copurified with human and porcine TSH-R migrated between monosialoganglioside GM1 and disialoganglioside GD1a. Ceramide glycanase digestion of the purified human TSH-R-associated glycolipid confirmed its ganglioside nature. It was resistant to Vibrio cholerae sialidase, which digest all gangliosides except GM1, but was sensitive to Arthrobacter ureafaciens sialidase, which digests all gangliosides including GM1. These findings indicate that the human TSH-R contains ganglioside that belongs to the galactosyl(β1→ 3)-N-acetylgalactosaminyl(β1→ 4)-[N-acetylneuraminyl(α2→ 3)]galactosyl(β1 → 4)glucosyl(β1 → 1)ceramide (GM1) family. Its intimate association with receptor protein implies a key role for ganglioside in the structure and function of the TSH-R

  3. cDNA cloning and characterization of a mannose-binding lectin from Zingiber officinale Roscoe (ginger) rhizomes

    Indian Academy of Sciences (India)

    Zhonghai Chen; Guoyin Kai; Xiaojun Liu; Juan Lin; Xiaofen Sun; Kexuan Tang

    2005-03-01

    Using RNA extracted from Zingiber officinale rhizomes and primers designed according to the conservative regions of monocot mannose-binding lectins, the full-length cDNA of Z. officinale agglutinin (ZOA) was cloned by rapid amplification of cDNA ends (RACE). The full-length cDNA of zoa was 746 bp and contained a 510 bp open reading frame (ORF) encoding a lectin precursor of 169 amino acids with a signal peptide. ZOA was a mannose-binding lectin with three typical mannose-binding sites (QDNY). Semi-quantitative RT-PCR analysis revealed that zoa expressed in all the tested tissues of Z. officinale including leaf, root and rhizome, suggesting it to be a constitutively expressing form. ZOA protein was successfully expressed in Escherichia coli with the molecular weight expected. To our knowledge, this is the first mannose-binding lectin cDNA cloned from the family Zingiberaceae. Our results demonstrate that monocot mannose-binding lectins also occur within the family Zingiberaceae.

  4. Transgenic plants expressing the AaIT/GNA fusion protein show increased resistance and toxicity to both chewing and sucking pests.

    Science.gov (United States)

    Liu, Shu-Min; Li, Jie; Zhu, Jin-Qi; Wang, Xiao-Wei; Wang, Cheng-Shu; Liu, Shu-Sheng; Chen, Xue-Xin; Li, Sheng

    2016-04-01

    The adoption of pest-resistant transgenic plants to reduce yield losses and decrease pesticide use has been successful. To achieve the goal of controlling both chewing and sucking pests in a given transgenic plant, we generated transgenic tobacco, Arabidopsis, and rice plants expressing the fusion protein, AaIT/GNA, in which an insecticidal scorpion venom neurotoxin (Androctonus australis toxin, AaIT) is fused to snowdrop lectin (Galanthus nivalis agglutinin, GNA). Compared with transgenic tobacco and Arabidopsis plants expressing AaIT or GNA, transgenic plants expressing AaIT/GNA exhibited increased resistance and toxicity to one chewing pest, the cotton bollworm, Helicoverpa armigera. Transgenic tobacco and rice plants expressing AaIT/GNA showed increased resistance and toxicity to two sucking pests, the whitefly, Bemisia tabaci, and the rice brown planthopper, Nilaparvata lugens, respectively. Moreover, in the field, transgenic rice plants expressing AaIT/GNA exhibited a significant improvement in grain yield when infested with N. lugens. This study shows that expressing the AaIT/GNA fusion protein in transgenic plants can be a useful approach for controlling pests, particularly sucking pests which are not susceptible to the toxin in Bt crops. PMID:25641865

  5. Intracellular iron concentration of neurons with and without perineuronal nets

    Science.gov (United States)

    Fiedler, Anja; Reinert, Tilo; Morawski, Markus; Brückner, Gert; Arendt, Thomas; Butz, Tilman

    2007-07-01

    Neurodegenerative diseases like Parkinson's disease, Alzheimer's disease and Huntington's disease are characterized by abnormally high concentrations of iron in the affected brain areas. Iron is believed to contribute to oxidative stress by catalysing radical generation and subsequently causing neuronal death. Interestingly, subpopulations of neurons are less vulnerable against degeneration. One of these subpopulations possesses a specialized extracellular matrix arranged as a perineuronal net (PN), a structure with poorly understood functions. In order to differentiate between neurons with and without PN according to their iron concentrations we have performed a μPIXE study at the Leipzig LIPSION laboratory. PN-ensheathed neurons in selected brain areas were detected by lectin-histochemical staining with Wisteria floribunda agglutinin (WFA). The staining was intensified by DAB- nickel by an established method enabling the visualisation of the PNs by nuclear microscopy. The cellular concentration of iron in the rat brain was about 1 mmol/l (ca. 30 μg/g dw). First results of subcellular analysis showed that the intracellular iron concentration of PN-ensheathed neurons tends to be slightly increased in comparison to neurons without PNs. The difference in intracellular iron concentrations could be an effect of the PNs.

  6. Echinoderm immunity

    Directory of Open Access Journals (Sweden)

    JE García-Arrarás

    2010-09-01

    Full Text Available Echinoderms are exclusively marine animals that, after the chordates, represent the second largest group of deuterostomes. Their diverse species composition and singular ecological niches provide at the same time challenges and rewards when studying the broad range of responses that make up their immune mechanisms. Two types of responses comprise the immune system of echinoderms: a cellular response and a humoral one. Cell-based immunity is carried by the celomocytes, a morphologically heterogeneous population of free roaming cells that are capable of recognizing and neutralizing pathogens. Celomocytes present diverse morphologies and functions, which include phagocytosis, encapsulation, clotting, cytotoxicity, wound healing among others. Humoral immunity is mediated by a wide variety of secreted compounds that can be found in the celomic fluid and play important roles in defense against infection. Compounds such as lectins, agglutinins, perforins, complement and some cytokines make up some of the humoral responses of echinoderms. Recent advances in the field of molecular biology, genomics and transcriptomics have allowed for the discovery of new immune genes and their products. These discoveries have expanded our knowledge of echinoderm immunity and are setting up the stage for future experiments to better understand the evolution of the immune mechanisms of deuterostomes

  7. Isolation, proliferation, and induction of Bama mini-pig spermatogonial stem cells in vitro.

    Science.gov (United States)

    Zhao, H M; Yang, H; Luo, F H; Li, M X; Zhang, S; Yang, X G; Lu, Y Q; Lu, S S; Wu, Y J; Lu, K H

    2016-01-01

    Spermatogonial stem cells (SSCs), the unique seed cells of testes, can undergo meiosis and form spermatozoa, thus transmitting genetic information to offspring. Research concerning these cells explores the mechanism underlying spermatogenesis, making possible the induction of their differentiation into spermatozoa in vitro. SSCs have therefore attracted much interest among scientists. Although the proliferation of such cells in vitro has been demonstrated, we are unaware of any long-term laboratory culture of porcine SSCs. The objective of this study was to isolate, characterize, culture, and induce the differentiation of Bama mini-pig SSCs. SSCs were isolated using differential plating and cultured for over 100 days on an STO feeder cell layer without serum. Cell clusters appeared after three passages and continuously formed during subsequent cultivation. Staining showed that these clusters were positive for UCHL1 and CDH1, could be bound by Dolichos biflorus agglutinin, and that some cells expressed OCT4. Ultrastructure observations revealed SSCs in testis tissue to be round in shape, while those cultured in vitro were flat and bound together. Our attempts at inducing differentiation showed that SSCs cultured in vitro could undergo meiosis. In this study, we describe an effective culture system for Bama mini-pig SSCs capable of producing enough cells to establish a platform for further SSC research, such as genetic manipulation or exploration of the mechanism underlying spermatogenesis. PMID:27525927

  8. Assessment of released acrosin activity as a measurement of the sperm acrosome reaction

    Institute of Scientific and Technical Information of China (English)

    Rui-Zhi Liu; Wan-Li Na; Hong-Guo Zhang; Zhi-Yong Lin; Bai-Oong Xue; Zong-Oe Xu

    2008-01-01

    Aim: To develop a method for assessing sperm function by measuring released acrosin activity during the acrosome reaction (AR). Methods: Human semen samples were obtained from 24 healthy donors with proven fertility after 3-7 days of sexual abstinence. After collection, samples were liquefied for 30 min at room temperature. Standard semen parameters were evaluated according to World Health Organization (WHO) criteria. Calcium ionophore A23187 and progesterone (P4) were used to stimulate the sperm to undergo AR. After treatment, sperm were incubated with the supravital dye Hoechst33258, fixed in a glutaraldehyde-phosphate-buffered saline solution, and the acrosomal status was determined by fluorescence microscopy with fluorescein isothiocyanate-labeled Pisum sativum agglutinin (FITC-PSA). The percentage of sperm undergoing AR (AR%) was compared to sperm acrosin activities as assessed by spectrocolorimetry. The correlation between AR% and acrosin activity was determined by statistical analysis. Results: The AR% and released acrosin activity were both markedly increased with A23187 and P4 stimulation. Sperm motility and viability were significantly higher after stimulation with P4 versus stimula-tion with A23187 (P < 0.001). There was a significant positive correlation between released acrosin activity and AR% determined by FITC-PSA staining (r = 0.916, P < 0.001). Conclusion: Spectrocolorimetric measurement of released acrosin activity might serve as a reasonable alternative method to evaluate AR.

  9. Long-term fungal inhibitory activity of water-soluble extract from Amaranthus spp. seeds during storage of gluten-free and wheat flour breads.

    Science.gov (United States)

    Giuseppe Rizzello, Carlo; Coda, Rossana; De Angelis, Maria; Di Cagno, Raffaella; Carnevali, Paola; Gobbetti, Marco

    2009-05-31

    This study aimed at investigating the use of the water-soluble extract of amaranth seeds for extending the shelf-life of gluten-free and wheat flour breads. The antifungal activity of the amaranth water-soluble extract was shown by agar diffusion, conidia germination and dry biomass assays, using Penicillium roqueforti DPPMAF1 as the indicator fungus. The crude water-soluble extract had minimal inhibitory concentration (MIC) of 5 mg of peptides/ml and showed inhibition towards a large number of fungal species isolated from bakeries. Four novel antifungal peptides, encrypted in amaranth agglutinin sequences, were identified from the water-soluble extract by nano-Liquid Chromatography-Electrospray Ionisation-Mass Spectra/Mass Spectra (nano-LC-ESI-MS/MS). The water-soluble extract of amaranth was used as an ingredient for the manufacture of gluten-free and wheat flour breads and the inhibitory activity was confirmed during long-term shelf-life under pilot plant conditions. The effect of the water-soluble extract on gluten-free bread rheology and sensory properties was also shown. PMID:19328576

  10. Microstructured Block Copolymer Surfaces for Control of Microbe Adhesion and Aggregation

    Directory of Open Access Journals (Sweden)

    Ryan R. Hansen

    2014-03-01

    Full Text Available The attachment and arrangement of microbes onto a substrate is influenced by both the biochemical and physical surface properties. In this report, we develop lectin-functionalized substrates containing patterned, three-dimensional polymeric structures of varied shapes and densities and use these to investigate the effects of topology and spatial confinement on lectin-mediated microbe immobilization. Films of poly(glycidyl methacrylate-block-4,4-dimethyl-2-vinylazlactone (PGMA-b-PVDMA were patterned on silicon surfaces into line arrays or square grid patterns with 5 μm wide features and varied pitch. The patterned films had three-dimensional geometries with 900 nm film thickness. After surface functionalization with wheat germ agglutinin, the size of Pseudomonas fluorescens aggregates immobilized was dependent on the pattern dimensions. Films patterned as parallel lines or square grids with a pitch of 10 μm or less led to the immobilization of individual microbes with minimal formation of aggregates. Both geometries allowed for incremental increases in aggregate size distribution with each increase in pitch. These engineered surfaces combine spatial confinement with affinity-based capture to control the extent of microbe adhesion and aggregation, and can also be used as a platform to investigate intercellular interactions and biofilm formation in microbial populations of controlled sizes.

  11. Freeze-dried egg yolk based extenders containing various antioxidants improve post-thawing quality and incubation resilience of goat spermatozoa.

    Science.gov (United States)

    Alcay, Selim; Gokce, Elif; Toker, M Berk; Onder, N Tekin; Ustuner, Burcu; Uzabacı, Ender; Gul, Zulfiye; Cavus, Seda

    2016-06-01

    The aim of this study was to evaluate different antioxidants-supplemented freeze-dried egg yolk based extenders for the post-thawing quality and incubation resilience of goat spermatozoa. Pooled semen were diluted in a two-step dilution method to a final concentration of 1/5 (semen/extender) in control and antoxidant supplemented freeze-dried extenders (methionine, cysteamine and butylated hydroxytoluene). Semen samples were assessed for sperm motility, plasma membrane functional integrity using hypoosmotic swelling test (HOST), damaged acrosome using FITC-Pisum sativum agglutinin (PSA-FITC) and DNA integrity using terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling (TUNEL). Membrane lipid peroxidation status was also analyzed using the malondialdehyde (MDA) concentration. In the study, antioxidant supplemented freeze-dried egg yolk based extenders have beneficial effect on goat sperm parameters. In addition, we achieved a higher quality in post thawed goat semen even after 6 h incubation when the extender was supplemented by 5 mM BHT or cysteamine. PMID:27018219

  12. Molecular Dynamics and Monte Carlo simulations resolve apparent diffusion rate differences for proteins confined in nanochannels

    Energy Technology Data Exchange (ETDEWEB)

    Tringe, J.W., E-mail: tringe2@llnl.gov [Lawrence Livermore National Laboratory, 7000 East Avenue, Livermore, CA (United States); Ileri, N. [Lawrence Livermore National Laboratory, 7000 East Avenue, Livermore, CA (United States); Department of Chemical Engineering & Materials Science, University of California, Davis, CA (United States); Levie, H.W. [Lawrence Livermore National Laboratory, 7000 East Avenue, Livermore, CA (United States); Stroeve, P.; Ustach, V.; Faller, R. [Department of Chemical Engineering & Materials Science, University of California, Davis, CA (United States); Renaud, P. [Swiss Federal Institute of Technology, Lausanne, (EPFL) (Switzerland)

    2015-08-18

    Highlights: • WGA proteins in nanochannels modeled by Molecular Dynamics and Monte Carlo. • Protein surface coverage characterized by atomic force microscopy. • Models indicate transport characteristics depend strongly on surface coverage. • Results resolve of a four orders of magnitude difference in diffusion coefficient values. - Abstract: We use Molecular Dynamics and Monte Carlo simulations to examine molecular transport phenomena in nanochannels, explaining four orders of magnitude difference in wheat germ agglutinin (WGA) protein diffusion rates observed by fluorescence correlation spectroscopy (FCS) and by direct imaging of fluorescently-labeled proteins. We first use the ESPResSo Molecular Dynamics code to estimate the surface transport distance for neutral and charged proteins. We then employ a Monte Carlo model to calculate the paths of protein molecules on surfaces and in the bulk liquid transport medium. Our results show that the transport characteristics depend strongly on the degree of molecular surface coverage. Atomic force microscope characterization of surfaces exposed to WGA proteins for 1000 s show large protein aggregates consistent with the predicted coverage. These calculations and experiments provide useful insight into the details of molecular motion in confined geometries.

  13. An acoustically-driven biochip - Impact of flow on the cell-association of targeted drug carriers

    CERN Document Server

    Fillafer, Christian; Neumann, Jürgen; Guttenberg, Zeno; Dissauer, Silke; Lichtscheidl, Irene; Wirth, Michael; Gabor, Franz; Schneider, Matthias; 10.1039/B906006E

    2011-01-01

    The interaction of targeted drug carriers with epithelial and endothelial barriers in vivo is largely determined by the dynamics of the body fluids. To simulate these conditions in binding assays, a fully biocompatible in vitro model was developed which can accurately mimic a wide range of physiological flow conditions on a thumbnail-format cell-chip. This acoustically-driven microfluidic system was used to study the interaction characteristics of protein-coated particles with cells. Poly(D,L-lactide-co-glycolide) (PLGA) microparticles (2.86 {\\pm} 0.95 {\\mu}m) were conjugated with wheat germ agglutinin (WGA-MP, cytoadhesive protein) or bovine serum albumin (BSA-MP, nonspecific protein) and their binding to epithelial cell monolayers was investigated under stationary and flow conditions. While mean numbers of 1500 {\\pm} 307 mm-2 WGA-MP and 94 {\\pm} 64 mm-2 BSA-MP respectively were detected to be cell-bound in the stationary setup, incubation at increasing flow velocities increasingly antagonized the attachment...

  14. Transmission of the allergy reaction of a delayed type against Salmonella abortus ovis through blood plasma of gamma-irradiated guinea pigs

    International Nuclear Information System (INIS)

    Use is made of blood plasma taken from guinea pigs (sensibilized with a live culture of Salmonella abortus ovis and then irradiated with 800 rad gamma-rays) to transmit the skin allergy reaction to normal, nonsensibilized guinea pigs. The allergy reaction has been demonstrated in the recipients of plasma as early as the 3-4th hour following the injection of the allergen into the skin. It reaches its peak at the 12-24th hour and later on strongly diminishes, remaining in few of the animals only up to the 48th hour. The infiltrate at the site of injection in the skin of positively reacting animals contains at the 24th hour cells of the polymorphonuclear type, which predominate, while the cells of the mononuclear type are few in number. There are no precipitins in the plasma of the donors, and the titer of the agglutinins and the cytophile antibodies is very low. Regardless of these findings it is concluded that the transmitted allergy reaction is of the fast type (after Arthuss), and not of the delayed one. (author)

  15. Characteristics of oligosaccharides from rat parotid (RP) N-linked glycoproteins (GP) after β-adrenoreceptor (β-AR) stimulation

    International Nuclear Information System (INIS)

    The authors have shown that β-AR stimulation of RP cells leads to marked enhancement of N-linked glycosylation in 4 secretory GP (Mr∼220Kd, HMW;∼32-38Kd, MMW;∼17Kd, LMW). To characterize oligosaccharides in GP, cells were incubated 60 min +/- isoproterenol (ISO) and analyzed 2 ways. First, cell extracts were subjected to SDS-PAGE and Western blotting with peroxidase-conjugated Con A or wheat germ agglutinin (WGA). Second, double-labeled (3H) man/14C leu) extracts were chromatographed on G200 followed by analysis of GP on Con A-Sepharose. HMW from control (CON) and ISO cells were Endo H insensitive, Endo F sensitive, altered by incubation with deoxynojirimycin (dNM), and bound both Con A and WGA conjugates. Similar findings were observed with LMW while MMW were sensitive to Endo H and Endo F, unaffected by dNM, bound Con A (strongly) and WGA (weakly). MMW and LMW were primarily eluted from Con A-Sepharose with 0.5M α-methyl mannoside (α-MM) while HMW were eluted sequentially with 10 mM α-methyl glucoside and α-MM. HMW, MMW, and LMW had ∼4 fold higher 3H/14C ratios after ISO. These results suggest HMW and LMW likely contain biantennary complex and hybrid oligosaccharides while MMW contain only high mannose oligosaccharide types

  16. Effects of Transgenic Tobacco Plants Expressing ACA Gene from Amaranthus caudatus on the Population Development of Myzus persicae

    Institute of Scientific and Technical Information of China (English)

    GUOHong-Nian; JIAYan-Tao; ZHOUYong-Gang; ZHANGZhen-Shan; OUYANGQing; JIANGYing; TIANYing-Chuan

    2004-01-01

    To investigate the possible function of the agglutinin from Amaranthus caudatus L. (ACA) in plant defending against insect pests, ACA cDNA was cloned by RT-PCR and the 5' and 3' sequences were confirmed by rapid amplification of cDNA ends (RACE). The phloem-specific expression vector of ACA gene, pBCACAc, was constructed based on the plant binary vector pBC438 and transfered into tobacco plants via Agrobacterium-mediated transformation method. Results from PCR and Southern blotting analysis showed that AOA gene was integrated into the genomes of transformed plants and the transgene integration varied from one to four estimated copies per genome. Western blotting analysis indicated that ACA gene was transcribed and translated in the transgenic plants. The bioassay of Myzus persicae Sulzer on detached leaves demonstrated that the 78% transgenic tobacco plants displayed an average aphid-resistant rate of more than 75%. Some apterous progeny of M. persicae were found dead on the resistant plants. These results indicate that ACA gene should be an effective aphid-resistant gene and could be valuable for application in crop breeding for aphid resistance.

  17. Effects of hypothermic liquid storage and cryopreservation on basal and induced plasma membrane phospholipid disorder and acrosome exocytosis in boar spermatozoa.

    Science.gov (United States)

    Guthrie, H D; Welch, G R

    2005-01-01

    Flow cytometry was utilised to determine whether short-term (Day 1) or long-term hypothermic liquid storage (Day 5), or cryopreservation of boar spermatozoa (1) caused changes in plasma membrane phospholipid disorder (MPLD) and acrosome exocytosis (AE), indicative of an advanced stage of capacitation or acrosome status, and (2) facilitated or inhibited the induction of capacitation and the acrosome reaction. Merocyanine with Yo-Pro-1 and peanut agglutinin-fluorescein isothiocyanate with propidium iodide were used to identify MPLD and AE, respectively, in viable spermatozoa. The incidence of basal sperm MPLD and AE in fresh semen was very low (1.1 and 2.2%, respectively) and was increased (P semen (3-8%). Compared to no bicarbonate, incubation with bicarbonate increased MPLD, but the response was greatest (P semen than for Day 1 (45%) and Day 5 (57%) semen. In summary, hypothermic liquid storage and cryopreservation of boar spermatozoa did not advance capacitation or acrosome status in viable spermatozoa, but did alter their responses to induction of capacitation and the acrosome reaction. PMID:15899159

  18. Anti IH: An antibody worth mention

    Directory of Open Access Journals (Sweden)

    Nithya Mohanan

    2016-01-01

    Full Text Available A 72-year-old female with co-morbidities posted for surgical correction of fracture neck of femur without any history of transfusions was noted to have a hemoglobin level of 7 g/dl and packed red blood cells transfusion was ordered. Pretransfusion tests demonstrated A1B group with D positive on forward grouping. Reverse grouping showed a varying grade of agglutination with A, B, and O cells. Agglutination being stronger at 4°C. Antibody screening showed pan-agglutination, direct Coomb's test and auto control were negative. The serum reacted with adult O cells (OIadult but not with adult Bombay cells (Oh Iadult or O cord (Oicord cells. A possibility of a compound cold antibody anti IH was made and A1B compatible cells were transfused to the patient. This case report illustrates anti-IH cold agglutinin with broad thermal amplitude. Uniqueness of this case report was O group incompatibility with A1B group, which was detected earlier and a catastrophic transfusion reaction being subverted.

  19. Assessment of sperm damages during different stages of cryopreservation in water buffalo by fluorescent probes.

    Science.gov (United States)

    Kumar, Dharmendra; Kumar, Pradeep; Singh, Pawan; Yadav, S P; Yadav, P S

    2016-05-01

    The present study was designed to investigate the sperm damages occurring in acrosome, plasma membrane, mitochondrial activity, and DNA of fresh, equilibrated and frozen-thawed buffalo semen by fluorescent probes. The stability of sperm acrosome and plasma membrane stability, mitochondrial activity and DNA status were assessed by fluorescein conjugated lectin Pisum sativum agglutinin, Annexin-V/propidium iodide, JC-1 and TUNEL assay, respectively, under the fluorescent microscope. The damages percentage of acrosome integrity was significantly increased during equilibration and freezing-thawing process. The stability of sperm plasma membrane is dependent on stability of phosphatidylserine (PS) on the inner leaflet of plasma membrane. The frozen-thawed sperm showed externalization of PS leading to significant increase in apoptotic, early necrotic and necrotic changes and lowered high mitochondrial membrane potential as compared with the fresh sperm but all these parameters were not affected during equilibration. However, the DNA integrity was not affected during equilibration and freezing-thawing procedure. In conclusion, the present study revealed that plasma membrane and mitochondria of buffalo sperm are more susceptible to damage during cryopreservation. Furthermore, the use of fluorescent probes to evaluate integrity of plasma and acrosome membranes, as well as mitochondrial membrane potential and DNA status increased the accuracy of semen analyses. PMID:25373338

  20. Fine-needle aspiration cytology of postirradiation sarcomas, including angiosarcoma, with immunocytochemical confirmation

    Energy Technology Data Exchange (ETDEWEB)

    Silverman, J.F.; Lannin, D.L.; Larkin, E.W.; Feldman, P.; Frable, W.J. (East Carolina Univ. School of Medicine, Greenville, NC (USA))

    1989-01-01

    Postirradiation sarcomas are an unusual but well-recognized late effect of cancer therapy. In this article, a fine-needle aspiration (FNA) series of four cases is presented. There were three female patients and one male patient, with an age range of 28-55 yr (mean, 41). Two of the patients were irradiated for uterine cervical carcinoma while the other two received irradiation for malignant lymphoma. The time interval to the development of the postirradiation sarcoma ranged from 10 to greater than 20 yr. There were a postirradiation synovial sarcoma of the buttock region, malignant fibrous histiocytoma of the bone (femur), and rhabdomyosarcoma and angiosarcoma of the retroperitoneum. A spectrum of cytologic findings was encountered, reflecting the specific types of sarcomas. Immunocytochemical studies performed on the aspirated material from the angiosarcoma demonstrated the utility of immunoperoxidase stains for ULEX europaeus agglutinin-1 (UEA-1) and, to a lesser degree, factor VIII-related antigen antibody, confirming the vascular nature of this malignancy. The FNA findings from all four cases demonstrated cytologic features that allowed recognition of this unusual complication of irradiation treatment. This article confirms the utility of FNA cytology in following patients with previous malignancies and differentiating a postirradiation sarcoma from recurrent carcinoma.

  1. Fine-needle aspiration cytology of postirradiation sarcomas, including angiosarcoma, with immunocytochemical confirmation

    International Nuclear Information System (INIS)

    Postirradiation sarcomas are an unusual but well-recognized late effect of cancer therapy. In this article, a fine-needle aspiration (FNA) series of four cases is presented. There were three female patients and one male patient, with an age range of 28-55 yr (mean, 41). Two of the patients were irradiated for uterine cervical carcinoma while the other two received irradiation for malignant lymphoma. The time interval to the development of the postirradiation sarcoma ranged from 10 to greater than 20 yr. There were a postirradiation synovial sarcoma of the buttock region, malignant fibrous histiocytoma of the bone (femur), and rhabdomyosarcoma and angiosarcoma of the retroperitoneum. A spectrum of cytologic findings was encountered, reflecting the specific types of sarcomas. Immunocytochemical studies performed on the aspirated material from the angiosarcoma demonstrated the utility of immunoperoxidase stains for ULEX europaeus agglutinin-1 (UEA-1) and, to a lesser degree, factor VIII-related antigen antibody, confirming the vascular nature of this malignancy. The FNA findings from all four cases demonstrated cytologic features that allowed recognition of this unusual complication of irradiation treatment. This article confirms the utility of FNA cytology in following patients with previous malignancies and differentiating a postirradiation sarcoma from recurrent carcinoma

  2. The Toxoplasma gondii cyst wall protein CST1 is critical for cyst wall integrity and promotes bradyzoite persistence.

    Science.gov (United States)

    Tomita, Tadakimi; Bzik, David J; Ma, Yan Fen; Fox, Barbara A; Markillie, Lye Meng; Taylor, Ronald C; Kim, Kami; Weiss, Louis M

    2013-01-01

    Toxoplasma gondii infects up to one third of the world's population. A key to the success of T. gondii as a parasite is its ability to persist for the life of its host as bradyzoites within tissue cysts. The glycosylated cyst wall is the key structural feature that facilitates persistence and oral transmission of this parasite. Because most of the antibodies and reagents that recognize the cyst wall recognize carbohydrates, identification of the components of the cyst wall has been technically challenging. We have identified CST1 (TGME49_064660) as a 250 kDa SRS (SAG1 related sequence) domain protein with a large mucin-like domain. CST1 is responsible for the Dolichos biflorus Agglutinin (DBA) lectin binding characteristic of T. gondii cysts. Deletion of CST1 results in reduced cyst number and a fragile brain cyst phenotype characterized by a thinning and disruption of the underlying region of the cyst wall. These defects are reversed by complementation of CST1. Additional complementation experiments demonstrate that the CST1-mucin domain is necessary for the formation of a normal cyst wall structure, the ability of the cyst to resist mechanical stress, and binding of DBA to the cyst wall. RNA-seq transcriptome analysis demonstrated dysregulation of bradyzoite genes within the various cst1 mutants. These results indicate that CST1 functions as a key structural component that confers essential sturdiness to the T. gondii tissue cyst critical for persistence of bradyzoite forms. PMID:24385904

  3. The Toxoplasma gondii cyst wall protein CST1 is critical for cyst wall integrity and promotes bradyzoite persistence.

    Directory of Open Access Journals (Sweden)

    Tadakimi Tomita

    Full Text Available Toxoplasma gondii infects up to one third of the world's population. A key to the success of T. gondii as a parasite is its ability to persist for the life of its host as bradyzoites within tissue cysts. The glycosylated cyst wall is the key structural feature that facilitates persistence and oral transmission of this parasite. Because most of the antibodies and reagents that recognize the cyst wall recognize carbohydrates, identification of the components of the cyst wall has been technically challenging. We have identified CST1 (TGME49_064660 as a 250 kDa SRS (SAG1 related sequence domain protein with a large mucin-like domain. CST1 is responsible for the Dolichos biflorus Agglutinin (DBA lectin binding characteristic of T. gondii cysts. Deletion of CST1 results in reduced cyst number and a fragile brain cyst phenotype characterized by a thinning and disruption of the underlying region of the cyst wall. These defects are reversed by complementation of CST1. Additional complementation experiments demonstrate that the CST1-mucin domain is necessary for the formation of a normal cyst wall structure, the ability of the cyst to resist mechanical stress, and binding of DBA to the cyst wall. RNA-seq transcriptome analysis demonstrated dysregulation of bradyzoite genes within the various cst1 mutants. These results indicate that CST1 functions as a key structural component that confers essential sturdiness to the T. gondii tissue cyst critical for persistence of bradyzoite forms.

  4. Fluorescent Staining of Tea Pathogenic Fungi in Tea Leaves Using Fluorescein-labeled Lectin

    Science.gov (United States)

    Yamada, Kengo; Yoshida, Katsuyuki; Sonoda, Ryoichi

    Fluorochrome-labeled lectin, fluorescein conjugated wheat germ agglutinin (F-WGA) was applied to stain tea pathogenic fungi in tea leaf tissue. Infected leaves were fixed and decolorized with a mixture of ethanol and acetic acid, and cleared with 10% KOH for whole mount before staining with F-WGA. Hyphae of Pestalotiopsis longiseta, Pseudocercospora ocellata, Botrytis cinerea and Colletotrichum theae-sinensis fluoresced brightly in whole mount and sectioned samples of infected leaf tissue. In browned tissue, hyphae did not fluoresce frequently in whole mount sample. Autofluorescence of leaf tissue was strong in browned tissue of sections, it was removed by 10% KOH treatment before staining. Penetration hyphae of C. theae-sinensis in cell wall of trichome and hyphae in basal part of trichome did not fluoresced frequently. In whole mount samples of tea leaf infected with Exobasidium vexans and E. reticulatum, hymenia appeared on leaf surface fluoresced, but hyphae in leaf tissue did not fluoresce. In sectioned samples, hyphae fluoresced brightly when sections were treated with 10% KOH before staining.

  5. Comparison of the nature of interactions of two sialic acid specific lectins Saraca indica and Sambucus nigra with N-acetylneuraminic acid by spectroscopic techniques

    Energy Technology Data Exchange (ETDEWEB)

    Singha, Shuvendu [Department of Natural Science, West Bengal University of Technology, Kolkata 700064 (India); Department of Chemistry, Jadavpur University, Jadavpur, Kolkata 700032 (India); Bose, Partha P. [Department of Biotechnology, National Institute of Pharmaceutical Education and Research (NIPER), Hajipur 844101 (India); Ganguly, Tapan [School of Laser Science and Engineering, Jadavpur University, Jadavpur, Kolkata 700032 (India); Campana, Patricia T. [Escola de Artes, Ciências e Humanidades, Universidade de São Paulo, 03828-000 São Paulo (Brazil); Ghosh, Rina [Department of Chemistry, Jadavpur University, Jadavpur, Kolkata 700032 (India); Chatterjee, Bishnu P., E-mail: cbishnup@gmail.com [Department of Natural Science, West Bengal University of Technology, Kolkata 700064 (India)

    2015-04-15

    The present paper deals with the isolation and purification of a new sialic acid binding lectin from the seed integument of Saraca indica (Ashok) and the purified lectin was designated Saracin II. Comparative studies on the interactions of saracin II and another sialic acid specific lectin Sambucus nigra agglutinin (SNA) with N-acetylneuraminic acid (NANA) were made using UV–vis absorption, steady state and time resolved fluorescence along with circular dichroism (CD) spectroscopy to reveal the nature and mechanisms of binding of these two lectins with NANA. The experimental observations obtained from UV–vis, steady state and time resolved fluorescence measurements demonstrated that SNA–NANA system formed relatively stronger ground state complex than saracin II–NANA pair. CD measurements further substantiated the propositions made from steady state and time resolved spectroscopic investigations. It was inferred that during interaction of SNA with NANA, the lectin adopted a relatively looser conformation with the extended polypeptide structures leading to the exposure of the hydrophobic cavities which favoured stronger binding with NANA. - Highlights: • Of the two lectins, stronger binding of SNA with NANA is observed. • Full exposure of the hydrophobic cavities of SNA favors the stronger interactions. • Saracin II can be used for the new generation of lectin based-therapeutics.

  6. Radioiodine labeled CdSe/CdS quantum dots. Lectin targeted dual probes

    Energy Technology Data Exchange (ETDEWEB)

    Akca, Ozlet; Unak, Perihan; Medine, E. Ilker; Kilcar, Ayfer Yurt; Ichedef, Cigdem [Ege Univ., Izmir (Turkey). Dept. of Nuclear Applications; Sakarya, Serhan [Adnan Menderes Univ., Aydin (Turkey). Dept. of Nuclear Medicine; Bekis, Recep [Dokuz Eyluel Univ., Izmir (Turkey). Dept. of Nuclear Medicine; Timur, Suna [Ege Univ., Izmir (Turkey). Biochemistry Dept.

    2014-11-01

    CdSe/CdS quantum dots (QD) were synthesized and bioconjugated with Sambucus nigra agglutinin (SNA) lectin (Lec). Mannose triflate and cysteamine molecules (MTC) were also utilized to prepare MTC-QDs and MTC-QDs-Lec probes as well as Lec bound QDs. Afterwards; potential use of these nanoparticles as radiolabeled fluorescence nano-probes for the cell imaging studies has been investigated. Biological activities of {sup 125}I{sup -}, {sup 125}I-MTC-QDs, MTC-QDs- Lec-{sup 125}I, QDs-Lec-{sup 125}I and Lec-{sup 125}I were examined on various cancer cell lines such as Caco-2, MCF-7 and A-549 in terms of cell incorporation. QDs-Lec-{sup 125}I exhibited the highest cell incorporation on whole cell lines. In addition, the QDs-Lec-{sup 131}I, was used for in vivo imaging. The whole body distribution of the radiolabeled QDs on New Zealand rabbits and Balb C mice were examined by taking dynamic and static images. Radioactivity cleared from the kidneys and the bladder, while significant amount radioactivity was retained in the heart and liver within 24 h.

  7. Cloning and molecular characterization of a novel lectin gene from Pinellia ternata

    Institute of Scientific and Technical Information of China (English)

    JIAN HONG YAO; XIU YUN ZHAO; ZHI HUA LIAO; JUAN LIN; ZHONG HAI CHEN; FEI CHEN; JUN SONG; XIAO FEN SUN; KE XUAN TANG

    2003-01-01

    The full-length cDNA of Pinellia ternata agglutinin (PTA) was cloned from inflorescences using RACE-PCR. Through comparative analysis of PTA gene (pta) and its deduced amino acid sequence with those of other Araceae species, pta was found to encode a precursor lectin with signal peptide and to have extensive homology with those of other Araceae species. PTA was a heterotetrameric mannose-binding lectin with three mannose-binding boxes like lectins from other Araceae and Amaryllidaceae species. Southern blot analysis of the genomic DNA revealed that pta belonged to a low-copy gene family. Northern blot analysis demonstrated that pta constitutively expressed in various plant tissues including root, leaf, stem and inflorescence. The pta cDNA sequence encoding for mature PTA protein was cloned into pET-32a plasmid and the resulting plasmid, pET-32a-PTA containing Trx-PTA fusion protein, was investigated for the expression in E. coli BL21. SDS-PAGE gel analysis showed that the Trx-PTA fusion protein was successfully expressed in E. coli BL21 when induced by IPTG. Artificial diet assay revealed that PTA fusion protein had significant levels of resistance against peach potato aphids when incorporated into artificial diet at 0.1% (w/v). The cloning of the pta gene will enable us to further test its effect in depth on aphids by transferring the gene into crop plants.

  8. Detection of sugar-lectin interactions by multivalent dendritic sugar functionalized single-walled carbon nanotubes

    Science.gov (United States)

    Vasu, K. S.; Naresh, K.; Bagul, R. S.; Jayaraman, N.; Sood, A. K.

    2012-07-01

    We show that single walled carbon nanotubes (SWNTs) decorated with sugar functionalized poly (propyl ether imine) (PETIM) dendrimer is a very sensitive platform to quantitatively detect carbohydrate recognizing proteins, namely, lectins. The changes in electrical conductivity of SWNT in field effect transistor device due to carbohydrate-protein interactions form the basis of present study. The mannose sugar attached PETIM dendrimers undergo charge-transfer interactions with the SWNTs. The changes in the conductance of the dendritic sugar functionalized SWNT after addition of lectins in varying concentrations were found to follow the Langmuir type isotherm, giving the concanavalin A (Con A)-mannose affinity constant to be 8.5 × 106 M-1. The increase in the device conductance observed after adding 10 nM of Con A is same as after adding 20 μM of a non-specific lectin peanut agglutinin, showing the high specificity of the Con A-mannose interactions. The specificity of sugar-lectin interactions was characterized further by observing significant shifts in Raman modes of the SWNTs.

  9. Microstructured block copolymer surfaces for control of microbe capture and aggregation

    Energy Technology Data Exchange (ETDEWEB)

    Hansen, Ryan R [ORNL; Shubert, Katherine R [ORNL; Morrell, Jennifer L. [University of Tennessee, Knoxville (UTK); Lokitz, Bradley S [ORNL; Doktycz, Mitchel John [ORNL; Retterer, Scott T [ORNL

    2014-01-01

    The capture and arrangement of surface-associated microbes is influenced by biochemical and physical properties of the substrate. In this report, we develop lectin-functionalized substrates containing patterned, three-dimensional polymeric structures of varied shapes and densities and use these to investigate the effects of topology and spatial confinement on lectin-mediated microbe capture. Films of poly(glycidyl methacrylate)-block-4,4-dimethyl-2-vinylazlactone (PGMA-b-PVDMA) were patterned on silicon surfaces into line or square grid patterns with 5 m wide features and varied edge spacing. The patterned films had three-dimensional geometries with 900 nm film thickness. After surface functionalization with wheat germ agglutinin, the size of Pseudomonas fluorescens aggregates captured was dependent on the pattern dimensions. Line patterns with edge spacing of 5 m or less led to the capture of individual microbes with minimal formation of aggregates, while grid patterns with the same spacing also captured individual microbes with further reduction in aggregation. Both geometries allowed for increases in aggregate size distribution with increased in edge spacing. These engineered surfaces combine spatial confinement with affinity-based microbe capture based on exopolysaccharide content to control the degree of microbe aggregation, and can also be used as a platform to investigate intercellular interactions and biofilm formation in microbial populations of controlled sizes.

  10. RELATION BETWEEN GLUCOLIPID PROFILE AND SMALL INTESTINE HISTOLOGICAL PATTERNS IN DIABETIC RATS EXPOSED TO AN INTERMITTENT DIETARY RESTRICTION

    Directory of Open Access Journals (Sweden)

    Noriyuki Hisano

    2009-01-01

    Full Text Available The effects of an intermittent and prolonged dietary restriction on biochemical variables and histological small intestinal patterns in 12-month-old male eSMT rats are examined. These spontaneously diabetic animals were separated in two groups after weaning: 10 rats fed ad libitum with standard rat chow and 10 rats fed a restricted diet by deprivation of the same food for 24 hours every 72. At 12 months of age, animals were weighed and euthanized after tail vein bleeding for plasma analysis (glycemia- both fasting and 120 minutes after an oral glucose challenge-, triglyceridemia and total cholesterolemia. Small intestines were removed, weighed and measured in length.Intestinal specimens were fixed, embedded in paraffin, semi serially cut at 6 µm and stained with PAS-Hematoxilyn and Hematoxilyn-Eosin. Histometry was performed through a linear devise attached to ocular lens and lectin histochemistry was accomplished employing Canavalis ensiformis, Dolichos biflorus, Arachis hypogea, Ulex europaeus-I, Triticum vulgaris, Ricinus communis and Soy Bean (Glicine Max Agglutinin. Essentially, eSMT rats, a suitable animal model for studying diabetes and/or its complications, revealed at 12 months of age after undergoing the dietary restriction: 1.- An expected improvement in body weight and determined biochemical variables (fasting and after glucose overload glycemias, triglyceridemia and total cholesterolemia without reaching euglycemic values. 2.- Changes in most of the analyzed histometric patterns with no relevant reflection on morphometric ones, and 3.- No modifications in lectinhistochemical patterns.

  11. Invasion of insect cells by Spiroplasma citri involves spiralin relocalization and lectin/glycoconjugate-type interactions.

    Science.gov (United States)

    Duret, Sybille; Batailler, Brigitte; Dubrana, Marie-Pierre; Saillard, Colette; Renaudin, Joël; Béven, Laure; Arricau-Bouvery, Nathalie

    2014-07-01

    Spiroplamas are helical, cell wall-less bacteria belonging to the Class Mollicutes, a group of microorganisms phylogenetically related to low G+C, Gram-positive bacteria. Spiroplasma species are all found associated with arthropods and a few, including Spiroplasma citri are pathogenic to plant. Thus S. citri has the ability to colonize cells of two very distinct hosts, the plant and the insect vector. While spiroplasmal factors involved in transmission by the leafhopper Circulifer haematoceps have been identified, their specific contribution to invasion of insect cells is poorly understood. In this study we provide evidence that the lipoprotein spiralin plays a major role in the very early step of cell invasion. Confocal laser scanning immunomicroscopy revealed a relocalization of spiralin at the contact zone of adhering spiroplasmas. The implication of a role for spiralin in adhesion to insect cells was further supported by adhesion assays showing that a spiralin-less mutant was impaired in adhesion and that recombinant spiralin triggered adhesion of latex beads. We also showed that cytochalasin D induced changes in the surface-exposed glycoconjugates, as inferred from the lectin binding patterns, and specifically improved adhesion of S. citri wild-type but not of the spiralin-less mutant. These results indicate that cytochalasin D exposes insect cell receptors of spiralin that are masked in untreated cells. In addition, competitive adhesion assays with lectins strongly suggest spiralin to exhibit glycoconjugate binding properties similar to that of the Vicia villosa agglutinin (VVA) lectin. PMID:24438161

  12. Expression of GALNT2 in human extravillous trophoblasts and its suppressive role in trophoblast invasion.

    Science.gov (United States)

    Liao, W-C; Chen, C-H; Liu, C-H; Huang, M-J; Chen, C-W; Hung, J-S; Chou, C-H; Chen, C-H; Che, M-I; Chang, H-M; Lan, C-T; Huang, H-C; Tseng, G-F; Shyu, M-K; Huang, M-C

    2012-12-01

    Extravillus trophoblast (EVT) invasion plays a critical role in placental development. Integrins bind to extracellular matrix (ECM) proteins to mediate EVT cell adhesion, migration, and invasion. Changes in O-glycans on β1-integrin have been found to regulate cancer cell behavior. We hypothesize that O-glycosyltransferases can regulate EVT invasion through modulating the glycosylation and function of β1-integrin. Here, we found that the GALNT1 and GALNT2 mRNA were highly expressed in HTR8/SVneo and first trimester EVT cells. Immunohistochemstry and immunofluorescence staining showed that GALNT2 was expressed in subpopulations of EVT cells in deciduas, but not in syncytiotrophoblasts and cytotrophoblasts of placental villi. The percentage of GALNT2-positive EVT cells increased with gestational ages. Overexpression of GALNT2 in HTR8/SVneo cells significantly enhanced cell-collagen IV adhesion, but suppressed cell migration and invasion. Notably, we found that GALNT2 increased the expression of Tn antigen (GalNAc-Ser/Thr) on β1-integrin as revealed by Vicia Villosa agglutinin (VVA) binding. Furthermore, GALNT2 suppressed the phosphorylation of focal adhesion kinase (FAK), a crucial downstream signaling molecule of β1-integrin. Our findings suggest that GALNT2 is a critical initiating enzyme of O-glycosylation for regulating EVT invasion. PMID:23117232

  13. A lectin recognizes differential arrangements of O-glycans on mucin repeats.

    Science.gov (United States)

    Kato, Kentaro; Takeuchi, Hideyuki; Ohki, Takao; Waki, Michihiko; Usami, Katsuaki; Hassan, Helle; Clausen, Henrik; Irimura, Tatsuro

    2008-07-11

    Interaction of Vicia villosa agglutinin-B4 (VVA-B4) to glycopeptides with O-linked GalNAc residues was investigated by surface plasmon resonance. The affinity was shown to be influenced by the arrangement of O-glycosylation sites on a peptide, PTTTPITTTTK, representing the tandem repeat of MUC2. The association rate constant was relatively high with a particular category of GalNAc-peptides in which more than three amino acid residues were placed between GalNAc-Thr residues. PTT( *)T( *)PITT( *)T( *)TK (T( *) indicates GalNAc-Thr) had the highest association rate constant among the glycopeptides tested. The dissociation rate constant was low in the peptides containing consecutive GalNAc residues and PT( *)TTPIT( *)T( *)T( *)TK was the lowest of the glycopeptides tested. Dissociation constant (K(D)), calculated as k(d)/k(a) was the lowest with PTT( *)T( *)PITT( *)T( *)TK. Therefore, the arrangement but not the quantity of GalNAc residues apparently determines the affinity between VVA-B4 and peptides with attached GalNAc residues. PMID:18455506

  14. GABAergic complex basket formations in the human neocortex.

    Science.gov (United States)

    Blazquez-Llorca, Lidia; García-Marín, Virginia; DeFelipe, Javier

    2010-12-15

    Certain GABAergic interneurons in the cerebral cortex, basket cells, establish multiple connections with cell bodies that typically outline the somata and proximal dendrites of pyramidal cells. During studies into the distribution of the vesicular GABA transporter (VGAT) in the human cerebral cortex, we were struck by the presence of a very dense, pericellular arrangement of multiple VGAT-immunoreactive (-ir) terminals in certain cortical areas. We called these terminals "Complex basket formations" (Cbk-formations) to distinguish them from the simpler and more typical pericellular GABAergic innervations of most cortical neurons. Here we examined the distribution of these VGAT-ir Cbk-formations in various cortical areas, including the somatosensory (area 3b), visual (areas 17 and 18), motor (area 4), associative frontal (dorsolateral areas 9, 10, 45, 46, and orbital areas 11, 12, 13, 14, 47), associative temporal (areas 20, 21, 22, and 38), and limbic cingulate areas (areas 24, 32). Furthermore, we used dual or triple staining techniques to study the chemical nature of the innervated cells. We found that VGAT-ir Cbk-formations were most frequently found in area 4 followed by areas 3b, 13, and 18. In addition, they were mostly observed in layer III, except in area 17, where they were most dense in layer IV. We also found that 70% of the innervated neurons were pyramidal cells, while the remaining 30% were multipolar cells. Most of these multipolar cells expressed the calcium-binding protein parvalbumin and the lectin Vicia villosa agglutinin. PMID:21031559

  15. The Thomsen-Friedenreich antigen-binding lectin jacalin interacts with desmoglein-1 and abrogates the pathogenicity of pemphigus foliaceus autoantibodies in vivo.

    Science.gov (United States)

    Li, Ning; Park, Moonhee; Zhao, Minglang; Hilario-Vargas, Julio; McInnes, David M; Prisayanh, Phillip S; Liu, Zhi; Diaz, Luis A

    2010-12-01

    Pemphigus foliaceus (PF) is an autoimmune skin blistering disease mediated by pathogenic autoantibodies against the desmosomal core glycoprotein desmoglein-1 (Dsg1). This study demonstrated that the O-glycan-specific plant lectin jacalin binds Dsg1 and inhibits the interaction of Dsg1/PF IgG. N-glycosylation is not involved in the interaction of Dsg1/jacalin or Dsg1/PF IgG. Subcutaneous injection of jacalin into neonatal mice drastically reduced PF IgG deposition at the epidermal cell surface and blocked PF IgG-induced skin blisters, both clinically and histologically. Interestingly, another plant lectin, peanut agglutinin, which shares the same carbohydrate specificity toward the O-linked carbohydrate structure known as Thomsen-Friedenreich antigen (TF antigen, Galβ1-3GalNAcα-O-Ser/Thr), also bound Dsg1 and blocked the skin blistering. In contrast, the plant lectin vicia villosa-B4 (VVL-B4), which shares the carbohydrate specificity toward the O-linked monosaccharide known as Thomsen-nouveau antigen (GalNAc-α1-O-Ser/Thr), did not bind Dsg1 and did not show a protective effect against the disease induced by the autoantibodies. Collectively, these results suggest that the binding of jacalin to O-linked TF carbohydrate motifs on Dsg1 impairs the Dsg1/PF autoantibody interactions and abrogates its pathogenicity in vivo. TF-specific binding ligands may have a potential therapeutic value for PF. PMID:20631728

  16. Intrahepatic cholangiocarcinoma detected by elevated levels of alpha-fetoprotein-L3 after hepatectomy for hepatocellular carcinoma in a patient with Budd-Chiari syndrome.

    Science.gov (United States)

    Yamamoto, Masakazu; Otsubo, Takehito; Ariizumi, Shunichi; Nakano, Masayuki; Takasaki, Ken

    2005-01-01

    We report the case of a 57-year-old woman with Budd-Chiari syndrome, hepatocellular carcinoma (HCC), and intrahepatic cholangiocarcinoma (ICC). She underwent partial hepatectomy for HCC in April 2000. After surgery, alpha-fetoprotein (AFP) and protein induced by vitamin K absence II (PIVKA-II) returned to normal levels, but lens culinaris agglutinin-reactive alpha-fetoprotein (AFP-L3) increased, and ultrasonography showed a nodule 2 cm in greatest dimension in the left lateral segment of the liver. We diagnosed this nodule as recurrence from HCC and performed a partial hepatectomy in October 2001. Microscopic examination showed that tubular adenocarcinoma and immunohistochemical staining was focally positive for AFP. AFP-L3 was 0% and AFP was 5 ng/ml 3 months after re-operation. This case was interesting in that ICC was detected by elevated levels of AFP-L3, and ICC produced AFP from the time it was minute in size. PMID:16119710

  17. The endophytic symbiont Epichloë festucae establishes an epiphyllous net on the surface of Lolium perenne leaves by development of an expressorium, an appressorium-like leaf exit structure.

    Science.gov (United States)

    Becker, Matthias; Becker, Yvonne; Green, Kimberly; Scott, Barry

    2016-07-01

    Epichloë festucae forms a mutualistic symbiotic association with Lolium perenne. This biotrophic fungus systemically colonizes the intercellular spaces of aerial tissues to form an endophytic hyphal network. E. festucae also grows as an epiphyte, but the mechanism for leaf surface colonization is not known. Here we identify an appressorium-like structure, which we call an expressorium that allows endophytic hyphae to penetrate the cuticle from the inside of the leaf to establish an epiphytic hyphal net on the surface of the leaf. We used a combination of scanning electron, transmission electron and confocal laser scanning microscopy to characterize this novel fungal structure and determine the composition of the hyphal cell wall using aniline blue and wheat germ agglutinin labelled with Alexafluor-488. Expressoria differentiate immediately below the cuticle in the leaf blade and leaf sheath intercalary cell division zones where the hyphae grow by tip growth. Differentiation of this structure requires components of both the NoxA and NoxB NADPH oxidase complexes. Major remodelling of the hyphal cell wall occurs following exit from the leaf. These results establish that the symbiotic association of E. festucae with L. perenne involves an interconnected hyphal network of both endophytic and epiphytic hyphae. PMID:26991322

  18. Mesenteric lymph node transcriptome profiles in BALB/c mice sensitized to three common food allergens

    Directory of Open Access Journals (Sweden)

    Boermans Herman J

    2011-01-01

    Full Text Available Abstract Background Food allergy is a serious health concern among infants and young children. Although immunological mechanism of food allergy is well documented, the molecular mechanism(s involved in food allergen sensitization have not been well characterized. Therefore, the present study analyzed the mesenteric lymph node (MLN transcriptome profiles of BALB/c mice in response to three common food allergens. Results Microarray analysis identified a total of 1361, 533 and 488 differentially expressed genes in response to β-lactoglobulin (BLG from cow's milk, ovalbumin (OVA from hen's egg white and peanut agglutinin (PNA sensitizations, respectively (p Conclusions Immunological profiles indicate that the allergen dosages used are sufficient to sensitize the BALB/c mice and to conduct transcriptome profiling. Microarray studies identified several differentially expressed genes in the sensitization phase of the food allergy. These findings will help to better understand the underlying molecular mechanism(s of food allergen sensitizations and may be useful in identifying the potential biomarkers of food allergy.

  19. A review of spontaneous and experimental porcine eperythrozoonosis

    International Nuclear Information System (INIS)

    Porcine eperythrozoonosis (PE) is a hemotropic disease caused by Eperythrozoon suis (E. suis). It is widespread and latent. Parasitemia is activated by environmental stress. Clinical signs include anemia and icterus. Reproductive disorders, neonatal death, postweaning diarrhea and increased susceptibility to other pathogens have been attributed to infection with E. suis. Factors which activate and regulate parasitemia are unknown. The disease cannot be reproduced consistently in intact pigs; however, experimental PE can be reproduced consistently in splenectomized pigs. Clinical signs and laboratory findings resemble those reported in spontaneous PE. We are investigating immunoregulation and misdirected immune responses in pigs infected with E. suis. Infected pigs have suppressed T-lymphocyte blastogenesis and synthesize a cold agglutinin immunoglobulin M (cIgM) directed against parasitized erythrocytes. There are reduced antifluorescein antibody concentration and affinity in primary and secondary sera from pigs immunized with keyhole limpet hemocyanin fluorescein when measured by microelisa and fluorescence quenching techniques. Purified cIgM agglutinates erythrocytes most actively at 4 C. Attachment and replication of E. suis on erythrocytes may result in the expression of a novel antigen responsible for synthesis of cIgM. E. suis might be a variable in studies of immunologic mechanisms in pigs

  20. Specific binding of a naturally occurring amyloidogenic fragment of Streptococcus mutans adhesin P1 to intact P1 on the cell surface characterized by solid state NMR spectroscopy

    Energy Technology Data Exchange (ETDEWEB)

    Tang, Wenxing; Bhatt, Avni [University of Florida, Department of Biochemistry and Molecular Biology, College of Medicine (United States); Smith, Adam N. [University of Florida, Department of Chemistry, College of Liberal Arts and Sciences (United States); Crowley, Paula J.; Brady, L. Jeannine, E-mail: jbrady@dental.ufl.edu [University of Florida, Department of Oral Biology, College of Dentistry (United States); Long, Joanna R., E-mail: jrlong@ufl.edu [University of Florida, Department of Biochemistry and Molecular Biology, College of Medicine (United States)

    2016-02-15

    The P1 adhesin (aka Antigen I/II or PAc) of the cariogenic bacterium Streptococcus mutans is a cell surface-localized protein involved in sucrose-independent adhesion and colonization of the tooth surface. The immunoreactive and adhesive properties of S. mutans suggest an unusual functional quaternary ultrastructure comprised of intact P1 covalently attached to the cell wall and interacting with non-covalently associated proteolytic fragments thereof, particularly the ∼57-kDa C-terminal fragment C123 previously identified as Antigen II. S. mutans is capable of amyloid formation when grown in a biofilm and P1 is among its amyloidogenic proteins. The C123 fragment of P1 readily forms amyloid fibers in vitro suggesting it may play a role in the formation of functional amyloid during biofilm development. Using wild-type and P1-deficient strains of S. mutans, we demonstrate that solid state NMR (ssNMR) spectroscopy can be used to (1) globally characterize cell walls isolated from a Gram-positive bacterium and (2) characterize the specific binding of heterologously expressed, isotopically-enriched C123 to cell wall-anchored P1. Our results lay the groundwork for future high-resolution characterization of the C123/P1 ultrastructure and subsequent steps in biofilm formation via ssNMR spectroscopy, and they support an emerging model of S. mutans colonization whereby quaternary P1-C123 interactions confer adhesive properties important to binding to immobilized human salivary agglutinin.

  1. Simple quantification of in planta fungal biomass.

    Science.gov (United States)

    Ayliffe, Michael; Periyannan, Sambasivam K; Feechan, Angela; Dry, Ian; Schumann, Ulrike; Lagudah, Evans; Pryor, Anthony

    2014-01-01

    An accurate assessment of the disease resistance status of plants to fungal pathogens is an essential requirement for the development of resistant crop plants. Many disease resistance phenotypes are partial rather than obvious immunity and are frequently scored using subjective qualitative estimates of pathogen development or plant disease symptoms. Here we report a method for the accurate comparison of total fungal biomass in plant tissues. This method, called the WAC assay, is based upon the specific binding of the plant lectin wheat germ agglutinin to fungal chitin. The assay is simple, high-throughput, and sensitive enough to discriminate between single Puccinia graminis f.sp tritici infection sites on a wheat leaf segment. It greatly lends itself to replication as large volumes of tissue can be pooled from independent experiments and assayed to provide truly representative quantification, or, alternatively, fungal growth on a single, small leaf segment can be quantified. In addition, as the assay is based upon a microscopic technique, pathogen infection sites can also be examined at high magnification prior to quantification if desired and average infection site areas are determined. Previously, we have demonstrated the application of the WAC assay for quantifying the growth of several different pathogen species in both glasshouse grown material and large-scale field plots. Details of this method are provided within. PMID:24643560

  2. A simple method for comparing fungal biomass in infected plant tissues.

    Science.gov (United States)

    Ayliffe, Michael; Periyannan, Sambasivam K; Feechan, Angela; Dry, Ian; Schumann, Ulrike; Wang, Ming-Bo; Pryor, Anthony; Lagudah, Evans

    2013-06-01

    Plant phenotypes resistant and susceptible to fungal pathogens are usually scored using qualitative, subjective methods that are based upon disease symptoms or by an estimation of the amount of visible fungal growth. Given that plant resistance genes often confer partial resistance to fungal pathogens, a simple, sensitive, nonsubjective quantitative method for measuring pathogen growth would be highly advantageous. This report describes an in planta quantitative assay for fungal biomass based upon detection of chitin using wheat germ agglutinin conjugated to a fluorophore. Using this assay, the growth of wheat rust pathogens on wheat was assayed and the additivity of several adult plant and seedling resistance genes to Puccinia striiformis, P. graminis, and P. triticina was assayed on both glasshouse- and field-grown material. The assay can discriminate between individual rust pustules on a leaf segment or, alternatively, compare fungal growth on field plots. The quantification of Erysiphe necator (powdery mildew) growth on Vitis vinifera (grapevine) is also demonstrated, with resistant and susceptible cultivars readily distinguished. Given that chitin is a major cell wall component of many plant fungal pathogens, this robust assay will enable simple and accurate measurement of biomass accumulation in many plant-fungus interactions. PMID:23405866

  3. A Novel Lectin with Antiproliferative and HIV-1 Reverse Transcriptase Inhibitory Activities from Dried Fruiting Bodies of the Monkey Head Mushroom Hericium erinaceum

    Directory of Open Access Journals (Sweden)

    Yanrui Li

    2010-01-01

    Full Text Available A lectin designated as Hericium erinaceum agglutinin (HEA was isolated from dried fruiting bodies of the mushroom Hericium erinaceum with a chromatographic procedure which entailed DEAE-cellulose, CM-cellulose, Q-Sepharose, and FPLC Superdex 75. Its molecular mass was estimated to be 51 kDa and its N-terminal amino acid sequences was distinctly different from those of other isolated mushroom lectins. The hemagglutinating activity of HEA was inhibited at the minimum concentration of 12.5 mM by inulin. The lectin was stable at pH 1.9–12.1 and at temperatures up to 70∘C, but was inhibited by Hg2+, Cu2+, and Fe3+ ions. The lectin exhibited potent mitogenic activity toward mouse splenocytes, and demonstrated antiproliferative activity toward hepatoma (HepG2 and breast cancer (MCF7 cells with an IC50 of 56.1 μM and 76.5 μM, respectively. It manifested HIV-1 reverse transcriptase inhibitory activity with an IC50 of 31.7 μM. The lectin exhibited potent mitogenic activity toward murine splenocytes but was devoid of antifungal activity.

  4. Insecticidal spider venom toxin fused to snowdrop lectin is toxic to the peach-potato aphid, Myzus persicae (Hemiptera: Aphididae) and the rice brown planthopper, Nilaparvata lugens (Hemiptera: Delphacidae).

    Science.gov (United States)

    Down, Rachel E; Fitches, Elaine C; Wiles, Duncan P; Corti, Paola; Bell, Howard A; Gatehouse, John A; Edwards, John P

    2006-01-01

    The SFI1/GNA fusion protein, comprising of snowdrop lectin (Galanthus nivalis agglutinin, GNA) fused to an insecticidal spider venom neurotoxin (Segestria florentina toxin 1, SFI1) was tested for toxicity against the rice brown planthopper Nilaparvata lugens (Stål) and the peach-potato aphid Myzus persicae (Sulzer) by incorporation into artificial diets. Significant effects on the mortality of N. lugens were observed, with 100% of the insects fed on the SFI1/GNA fusion protein diet dead by day 7. The survival of the aphid M. persicae was also reduced when fed on the SFI1/GNA fusion protein. After 14 days, only 49% of the aphids that were fed on the fusion protein were still alive compared with approximately 90% of the aphids fed on the control diet or on diet containing GNA only. The SFI1/GNA fusion protein also slowed the development of M. persicae, and the reproductive capacity of the aphids fed on the SFI1/GNA fusion protein was severely reduced. The ability of GNA to act as a carrier protein, and deliver the SFI1 neurotoxin to the haemolymph of N. lugens, following oral ingestion, was investigated. The successful delivery of intact SFI1/GNA fusion protein to the haemolymph of these insects was shown by western blotting. Haemolymph taken from the insects that were fed on the fusion protein contained two GNA-immunoreactive proteins of molecular weights corresponding to GNA and to the SFI1/GNA fusion protein. PMID:16206236

  5. Fusion proteins containing insect-specific toxins as pest control agents: snowdrop lectin delivers fused insecticidal spider venom toxin to insect haemolymph following oral ingestion.

    Science.gov (United States)

    Fitches, Elaine; Edwards, Martin G; Mee, Christopher; Grishin, Eugene; Gatehouse, Angharad M R; Edwards, John P; Gatehouse, John A

    2004-01-01

    The mannose-specific snowdrop lectin (Galanthus nivalis agglutinin: GNA), when fed to insects, binds to the gut epithelium and passes into the haemolymph. The ability of GNA to act as a carrier protein to deliver an insecticidal spider venom neurotoxin (Segestria florentina toxin 1: SFI1) to the haemolymph of lepidopteran larvae was investigated. Constructs encoding SFI1 and an SFI1/GNA fusion protein were expressed in Pichia pastoris. The insecticidal activity of purified recombinant proteins on injection was found to be comparable to published values for SfI1 purified from spider venom [Toxicon 40 (2002) 125]. Whereas neither GNA nor SFI1 alone showed acute toxicity when fed to larvae of tomato moth (Lacanobia oleracea), feeding SFI1/GNA fusion at 2.5% of dietary proteins was insecticidal to first stadium larvae, causing 100% mortality after 6 days. The protein also showed a significant, dose dependent, toxicity towards fourth and fifth stadium larvae, with growth reduced by up to approximately 90% over a 4-day assay period compared to controls. Delivery of intact SFI1/GNA to the haemolymph in these insects was shown by western blotting; haemolymph samples from fusion-fed larvae contained a GNA-immunoreactive protein of the same molecular weight as the SFI1/GNA fusion. SFI1/GNA and similar fusion proteins offer a novel and effective approach for delivering haemolymph active toxins by oral administration, which could be used in crop protection by expression in transgenic plants. PMID:15037094

  6. Actin polymerisation at the cytoplasmic face of eukaryotic nuclei

    Directory of Open Access Journals (Sweden)

    David-Watine Brigitte

    2006-05-01

    Full Text Available Abstract Background There exists abundant molecular and ultra-structural evidence to suggest that cytoplasmic actin can physically interact with the nuclear envelope (NE membrane system. However, this interaction has yet to be characterised in living interphase cells. Results Using a fluorescent conjugate of the actin binding drug cytochalasin D (CD-BODIPY we provide evidence that polymerising actin accumulates in vicinity to the NE. In addition, both transiently expressed fluorescent actin and cytoplasmic micro-injection of fluorescent actin resulted in accumulation of actin at the NE-membrane. Consistent with the idea that the cytoplasmic phase of NE-membranes can support this novel pool of perinuclear actin polymerisation we show that isolated, intact, differentiated primary hepatocyte nuclei support actin polymerisation in vitro. Further this phenomenon was inhibited by treatments hindering steric access to outer-nuclear-membrane proteins (e.g. wheat germ agglutinin, anti-nesprin and anti-nucleoporin antibodies. Conclusion We conclude that actin polymerisation occurs around interphase nuclei of living cells at the cytoplasmic phase of NE-membranes.

  7. Electrochemical Impedance Immunosensor Based on Self-Assembled Monolayers for Rapid Detection of Escherichia coli O157:H7 with Signal Amplification Using Lectin

    Directory of Open Access Journals (Sweden)

    Zhanming Li

    2015-08-01

    Full Text Available Escherichia coli O157:H7 is a predominant foodborne pathogen with severe pathogenicity, leading to increasing attention given to rapid and sensitive detection. Herein, we propose an impedance biosensor using new kinds of screen-printed interdigitated microelectrodes (SPIMs and wheat germ agglutinin (WGA for signal amplification to detect E. coli O157:H7 with high sensitivity and time-efficiency. The SPIMs integrate the high sensitivity and short response time of the interdigitated electrodes and the low cost of the screen-printed electrodes. Self-assembling of bi-functional 3-dithiobis-(sulfosuccinimidyl-propionate (DTSP on the SPIMs was investigated and was proved to be able to improve adsorption quantity and stability of biomaterials. WGA was further adopted to enhance the signal taking advantage of the abundant lectin-binding sites on the bacteria surface. The immunosensor exhibited a detection limit of 102 cfu·mL−1, with a linear detection range from 102 to 107 cfu·mL−1 (r2 = 0.98. The total detection time was less than 1 h, showing its comparable sensitivity and rapid response. Furthermore, the low cost of one SPIM significantly reduced the detection cost of the biosensor. The biosensor may have great promise in food safety analysis and lead to a portable biosensing system for routine monitoring of foodborne pathogens.

  8. Human Brucellosis: Still an Unfamiliar and Misdiagnosed Disease in India

    Directory of Open Access Journals (Sweden)

    Smita Mangalgi

    2015-01-01

    Full Text Available Background: Human brucellosis is a disease with protean clinical manifestations. Despite many awareness programmes, it is still missed or wrongly diagnosed. This leads to chronic morbidity leading to misery and loss of working days. Aim and Objectives: To assess the microbiological, clinical and epidemiological aspects of human brucellosis. Materials and Methods: Patients with positive brucella screening test constituted the study material. A detailed laboratory, clinical, epidemiological study along with response to the treatment was analyzed. Results: Seroprevalence of brucellosis was found to be 1.75%. Brucellosis was clinically diagnosed in only 12.73% of cases. Fever, joint pain and low backache were the commonest symptoms. Close contact with animals and raw milk ingestion were the major sources of infection. Knowledge regarding brucellosis and its prevention was lacking in patients. Brucellosis was not considered as one of the differential diagnosis by the treating physicians. Conclusion: Brucellosis should be considered as one of the differential diagnosis in cases presenting with fever, low backache, arthritis and arthralgia. Laboratories should screen all the serum samples for brucella agglutinins by Rose Bengal Plate Test. Awareness regarding the prevention of brucellosis in the general population and regarding the existence of the disease among the doctors practicing in rural areas is needed

  9. cDNA cloning and expression analysis of a mannose-binding lectin from Pinellia pedatisecta

    Indian Academy of Sciences (India)

    Juan Lin; Xuanwei Zhou; Shi Gao; Xiaojun Liu; Weisheng Wu; Xiaofen Sun; Kexuan Tang

    2007-03-01

    Pinellia pedatisecta agglutinin (PPA) is a very basic protein that accumulates in the tuber of P. pedatisecta. PPA is a hetero-tetramer protein of 40 kDa, composed of two polypeptide chains A (about 12 kDa) and two polypeptides chains B (about 12 kDa). The full-length cDNA of PPA was cloned from P. pedatisecta using SMART RACE-PCR technology; it was 1146 bp and contained a 771 bp open reading frame (ORF) encoding a lectin precursor of 256 amino acid residues with a 24 amino acid signal peptide. The PPA precursor contained 3 mannose-binding sites (QXDXNXVXY) and two conserved domains of 43% identity, PPA-DOM1 (polypeptides A) and PPA-DOM2 (polypeptides B). PPA shared varying identities, ranging from 40% to 85%, with mannose-binding lectins from other species of plant families such as Araceae, Alliaceae, Iridaceae, Liliaceae, Amaryllidaceae and Bromeliaceae. Southern blot analysis indicated that ppa belonged to a multi-copy gene family. Expression pattern analysis revealed that ppa expressed in most tested tissues, with high expression being found in spadix, spathe and tuber. Cloning of the ppa gene not only provides a basis for further investigation of its structure, expression and regulatory mechanism, but also enables us to test its potential role in controlling pests and fungal diseases by transferring the gene into plants in the future.

  10. Role of Golgi derived clathrin coated vesicles in the transport of calsequestrin to the sarcoplasmic reticulum of developing myotubes

    International Nuclear Information System (INIS)

    The sarcoplasmic reticulum (SR) is the major intracellular calcium sequestering organelle in skeletal and heart muscle. Calsequestrin (CSQ) is a glycoprotein with a molecular weight of 42,000. This protein is located in the lumen of the SR and it binds Ca2+ to maintain the concentration of this cation in the lumen at 10/sup /minus/3/M. Highly purified coated vesicles (CVs) were isolated from chick muscle and a Western blot using polyclonal anti-CSQ revealed the presence of CSQ within the CVs. Another major protein in the SR, the Ca2+-ATPase, was not contained in CVs suggesting different routes of insertion into the SR. Cultured chick myotubes were labelled with Trans35S-label contain [35S]-methionine and [35S]-cysteine to follow the transport of CSQ. Labelled CSQ remained high in the CVs until after 45 minutes of chase, then declined. The amount of labelled CSQ in the SR continued to rise over the chase period. No CSQ was secreted. All the CSQ in CVs and SR was sensitive to the activity of endoglycosidase H, and a significant fraction also bound wheat germ agglutinin. A small amount of CSQ was also co-transported with the muscle protein acetylcholinesterase within CVs. Non-secreted forms of acetylcholinesterase had the same carbohydrate structure as CSQ and were shown to be degraded in the SR

  11. X irradiation of human epidermis in vitro. 2

    International Nuclear Information System (INIS)

    On the example of the reduction of epidermal adhesion of FITC wheat germ agglutinine (WGA) the direct membrane effect of a single X irradiation (44 kV and 220 kV) was analyzed in vitro. Human normal skin and psoriasis centres were compared. Normal skin showed no alteration of microscopically visible FITC-WGA adhesion on epidermal cells over the whole dose range. Foci of psoriasis responded to doses of ≥ 5 Gy (44 and 220 kV) with a drastic reduction of epidermal lectin binding to lower and medium cell layers. Maximum efficacy was with 5 Gy (44 kV) or 10 Gy (220 kV). A dose elevation up to 20 Gy did not result in an increase of efficacy. Topographically the radiosensitive FITC-WGA adhesion could chiefly be seen in the dermal ridges. The findings support the impression of an increased radiosensitivity of the lesional psoriatic epidermis compared with normal skin. This is connected with an abnormal differentiation of keratinocytes in psoriasis. (author)

  12. Release of Glycoprotein (GP1 from the Tegumental Surface of Taenia solium by Phospholipase C from Clostridium perfringens Suggests a Novel Protein-Anchor to Membranes

    Directory of Open Access Journals (Sweden)

    Abraham Landa

    2010-01-01

    Full Text Available In order to explore how molecules are linked to the membrane surface in larval Taenia solium, whole cysticerci were incubated in the presence of phospholipase C from Clostridium perfringens (PLC. Released material was collected and analyzed in polyacrylamide gels with sodium dodecyl sulfate. Two major bands with apparent molecular weights of 180 and 43 kDa were observed. Western blot of released material and localization assays in cysticerci tissue sections using antibodies against five known surface glycoproteins of T. solium cysticerci indicated that only one, previously called GP1, was released. Similar localization studies using the lectins wheat-germ-agglutinin and Concanavalin A showed that N-acetyl-D-glucosamine, N-acetylneuraminic, sialic acid, αmethyl-D-mannoside, D-manose/glucose, and N-acetyl-D-glucosamine residues are abundantly present on the surface. On the other hand, we find that treatment with PLC releases molecules from the surface; they do not reveal Cross Reacting Determinant (CRD, suggesting a novel anchor to the membrane for the glycoprotein GP1.

  13. Enhancement of bone marrow allografts from nude mice into mismatched recipients by T cells void of graft-versus-host activity

    International Nuclear Information System (INIS)

    Transplantation of 8 x 10(6) C57BL/6-Nu+/Nu+ (nude) bone marrow cells into C3H/HeJ recipients after conditioning with 8 Gy of total body irradiation has resulted in a markedly higher rate of graft rejection or graft failure compared to that found in recipients of normal C57BL/6 or C57BL/6-Bg+/Bg+ (beige) T-cell-depleted bone marrow. Mixing experiments using different numbers of nude bone marrow cells with or without mature thymocytes (unagglutinated by peanut agglutinin) revealed that engraftment of allogeneic T-cell-depleted bone marrow is T-cell dependent. To ensure engraftment, a large inoculum of nude bone marrow must be supplemented with a trace number of donor T cells, whereas a small bone marrow dose from nude donors requires a much larger number of T cells for engraftment. Marked enhancement of donor type chimerism was also found when F1 thymocytes were added to nude bone marrow cells, indicating that the enhancement of bone marrow engraftment by T cells is not only mediated by alloreactivity against residual host cells but may rather be generated by growth factors, the release of which may require specific interactions between T cells and stem cells or between T cells and bone marrow stroma cells

  14. Purification and Biochemical Characterisation of Ricin from Castor Seeds

    Directory of Open Access Journals (Sweden)

    Om Kumar

    2004-07-01

    Full Text Available Ricin is a highly toxic plant toxin of Ricinus comtnunis seeds, commonly known as castor seeds. The toxin was extracted and purified using affinity and size exclusion  chromatography. The purity of ricin was evaluated by the sodium dodecylsulphate-polyacrylamide gel electrophoresis. Purified ricin gives a single band under non-reduced condition and two bands under reduced condition. The molecular weight of ricin was 65,0000 approx. The subunit structure of ricin on treatment with p-mercaptoethanol (1 % at molecular level revealed that the reducing agent converts ricin into two peptides. The molecular weight of these two peptides was estimated to be 34000 and 32000. The western-blot analysis revealed two dots for its two peptides in 29 kDa to 36 kDa regions. The heamagglutination litres for ricin and Ricinus communis agglutinins were 1:8 and 1:256. The purity of purified ricin was further confirmed by the electrophoresis and the western-blot analysis. The Indian variety of castor seeds, known as Ricinus communis used in this study, contains approx. 0.12 per cent ricin.

  15. Korean mistletoe lectin promotes proliferation and invasion of trophoblast cells through regulation of Akt signaling.

    Science.gov (United States)

    Lyu, Su-Yun; Choi, Jong Ho; Lee, Hyun-Jung; Park, Won-Bong; Kim, Gi Jin

    2013-08-01

    Recently, Viscum album var. coloratum agglutinin (VCA) was shown to have various effects on cancer cells. However, most researchers are focused on high concentrations (1-1000 ng/ml) of VCA and its anti-cancer effects. Therefore, we wanted to know whether low concentrations of VCA have an effect on proliferation and invasion of human trophoblast cells (HTR-8/SVneo cell line). Cell proliferations at low concentration of VCA (1-10 pg/ml) were increased over 2-fold relative to the control. Also, gelatinolytic activities of matrix metalloproteinase-2 were increased after VCA treatment, while TIMP-1 expression was reduced. Furthermore, the expression of integrin subunits α5 and β1 were increased 1.5-fold when cells were treated with low dose of VCA (10 pg/ml). Lastly, VCA was able to promote trophoblast invasion through activation of the Akt signaling pathway. In conclusion, low concentrations of VCA can stimulate the ability of trophoblast cells to invade through the extracellular matrix in vitro. PMID:23571125

  16. Anti-cancer effects of enteric-coated polymers containing mistletoe lectin in murine melanoma cells in vitro and in vivo.

    Science.gov (United States)

    Han, Seung-Yeon; Hong, Chang-Eui; Kim, Hwan-Gyu; Lyu, Su-Yun

    2015-10-01

    In this study, we evaluated the effects of Korean mistletoe (Viscum album L. var. coloratum) coated with a biodegradable polymer (Eudragit(®)) wall on the growth of mouse melanoma in vivo. Oral administration of 4% (430 mg/kg/day) enteric-coated mistletoe resulted in a significant reduction in tumor volume on day 14 compared to the negative control group in B16F10 melanoma-inoculated BDF1 mice. When we measured the survival rate, enteric-coated mistletoe-received mice had a higher survival rate after day 12. Also, we investigated the mechanism involving the cancer cell growth inhibition when melanoma cells were treated with Korean mistletoe lectin (Viscum album L. var. coloratum agglutinin, VCA) and its extract in vitro. As a result, a significant G0/G1 arrest was observed in both B16BL6 and B16F10 melanoma cells with VCA or mistletoe extract. In addition, VCA or mistletoe extract induced an increase in both early and late apoptosis in cells. When we studied the molecular mechanism, our results showed that VCA and mistletoe extract can increase activated multiple caspases (caspase-1, 3, 4, 5, 6, 7, 8, and 9), dose-dependently. We also found out that VCA and mistletoe treatment causes a significant decrease in the expression of procaspase-3 and 8. PMID:26152904

  17. Profiling of Leptospira interrogans, L. santarosai, L. meyeri and L. borgpetersenii by SE-AFLP, PFGE and susceptibility testing—a continuous attempt at species and serovar differentiation

    Science.gov (United States)

    Moreno, Luisa Z; Miraglia, Fabiana; Lilenbaum, Walter; Neto, José SF; Freitas, Julio C; Morais, Zenaide M; Hartskeerl, Rudy A; da Costa, Barbara LP; Vasconcellos, Silvio A; Moreno, Andrea M

    2016-01-01

    Leptospirosis is a widespread systemic zoonosis, considered as reemerging in certain developing countries. Although the cross agglutinin absorption test is still considered the standard method for Leptospira identification, it presents several disadvantages. The aim of this study was to characterize Leptospira spp. isolated from various hosts by genotyping and broth microdilution susceptibility testing in an attempt to differentiate Leptospira species, serogroups and serovars. Forty-seven isolates were studied. They were previously serotyped, and species confirmation was performed by 16S rRNA sequencing. Single-enzyme amplified fragment length polymorphism (SE-AFLP) and pulsed-field gel electrophoresis (PFGE) analysis enabled the distinction of L. interrogans from L. santarosai, L. meyeri and L. borgpetersenii in two main clusters. Among L. interrogans, it was possible to differentiate into two new clusters the serogroup Icterohaemorrhagiae from the serogroups Canicola and Pomona. L. santarosai isolates presented higher genetic variation than the other species in both techniques. Interestingly, the minimum inhibitory concentration (MIC) cluster analysis also provided Leptospira serogroup differentiation. Further studies are necessary regarding serovar Bananal isolates, as they presented the highest MIC values for most of the antimicrobials tested. All studied techniques successfully distinguished Leptospira species and serogroups. Despite being library-dependent methods, these approaches are less labor intensive and more economically viable, particularly SE-AFLP, and can be implemented in most reference laboratories worldwide to enable faster Leptospira typing. PMID:26956446

  18. Delta-Like Ligand (DLL)1 Expression in Early Mouse Decidua and Its Localization to Uterine Natural Killer Cells

    Science.gov (United States)

    Yamada, Aureo T.; Croy, B. Anne

    2012-01-01

    Uterine vascular changes, critical for pregnancy success, occur at each implant site during endometrial decidualization. Mesometrial decidualization recruits high numbers of angiogenic, uterine Natural Killer (uNK) cells that trigger midpregnancy spiral arterial remodeling. We postulated that uNK cells contribute to early decidual angiogenesis as endothelial-cell extrinsic sources of Delta-like ligand 1 (DLL1), a molecule that induces endothelial tip cell differentiation and orthogonal vascular growth in other tissues. Virgin uteri expressed Dll1 mesometrially and anti-mesometrially and relative expression increased in both anatomic regions as pregnancy progressed. Analyses of transcripts from gd10.5 uNK cells flow sorted on the basis of expression of Dolichos biflorus agglutinin (DBA) lectin revealed that DBA+ but not DBA- uNK cells expressed Dll1. Immunostaining at gd4.5 found DLL1-expressing cells rare. At gd6.5, DBA+ uNK cells at all stages of maturation expressed DLL1. By gd10.5, DLL1 immunoreactivity was strongly expressed by some but not all DBA+ uNK cells and more weakly by DBA- cells. DLL1+ cells were mesometrially stratified and concentrated within central decidua basalis. Our data suggest that uNK cells have the potential to induce endothelial tip cell differentiation and to promote non-planar vascular growth within early decidua basalis. PMID:23284862

  19. Specific binding of a naturally occurring amyloidogenic fragment of Streptococcus mutans adhesin P1 to intact P1 on the cell surface characterized by solid state NMR spectroscopy

    International Nuclear Information System (INIS)

    The P1 adhesin (aka Antigen I/II or PAc) of the cariogenic bacterium Streptococcus mutans is a cell surface-localized protein involved in sucrose-independent adhesion and colonization of the tooth surface. The immunoreactive and adhesive properties of S. mutans suggest an unusual functional quaternary ultrastructure comprised of intact P1 covalently attached to the cell wall and interacting with non-covalently associated proteolytic fragments thereof, particularly the ∼57-kDa C-terminal fragment C123 previously identified as Antigen II. S. mutans is capable of amyloid formation when grown in a biofilm and P1 is among its amyloidogenic proteins. The C123 fragment of P1 readily forms amyloid fibers in vitro suggesting it may play a role in the formation of functional amyloid during biofilm development. Using wild-type and P1-deficient strains of S. mutans, we demonstrate that solid state NMR (ssNMR) spectroscopy can be used to (1) globally characterize cell walls isolated from a Gram-positive bacterium and (2) characterize the specific binding of heterologously expressed, isotopically-enriched C123 to cell wall-anchored P1. Our results lay the groundwork for future high-resolution characterization of the C123/P1 ultrastructure and subsequent steps in biofilm formation via ssNMR spectroscopy, and they support an emerging model of S. mutans colonization whereby quaternary P1-C123 interactions confer adhesive properties important to binding to immobilized human salivary agglutinin

  20. An experimental platform using human intestinal epithelial cell lines to differentiate between hazardous and non-hazardous proteins.

    Science.gov (United States)

    Hurley, Bryan P; Pirzai, Waheed; Eaton, Alex D; Harper, Marc; Roper, Jason; Zimmermann, Cindi; Ladics, Gregory S; Layton, Raymond J; Delaney, Bryan

    2016-06-01

    Human intestinal epithelial cell lines (T84, Caco-2, and HCT-8) grown on permeable Transwell™ filters serve as models of the gastrointestinal barrier. In this study, this in vitro model system was evaluated for effectiveness at distinguishing between hazardous and non-hazardous proteins. Indicators of cytotoxicity (LDH release, MTT conversion), monolayer barrier integrity ([(3)H]-inulin flux, horseradish peroxidase flux, trans-epithelial electrical resistance [TEER]), and inflammation (IL-8, IL-6 release) were monitored following exposure to hazardous or non-hazardous proteins. The hazardous proteins examined include streptolysin O (from Streptococcus pyogenes), Clostridium difficile Toxins A and B, heat-labile toxin from enterotoxigenic Escherichia coli, listeriolysin O (from Listeria monocytogenes), melittin (from bee venom), and mastoparan (from wasp venom). Non-hazardous proteins included bovine and porcine serum albumin, bovine fibronectin, and ribulose bisphosphate carboxylase/oxygenase (RuBisco) from spinach. Food allergenic proteins bovine milk β-lactoglobulin and peanut Ara h 2 were also tested as was the anti-nutritive food protein wheat germ agglutinin. Results demonstrated that this model system effectively distinguished between hazardous and non-hazardous proteins through combined analysis of multiple cells lines and assays. This experimental strategy may represent a useful adjunct to multi-component analysis of proteins with unknown hazard profiles. PMID:27060235

  1. Studies on a novel macrophage-specific calmodulin binding glycoprotein

    Energy Technology Data Exchange (ETDEWEB)

    Orlow, S.J.

    1986-01-01

    The murine macrophage-like cell line J774 and peritoneal exudate cells elicited with thioglycollate or starch contain a major calmodulin-binding protein which is absent in trifluoperazine-resistant variants of J774, resident peritoneal macrophages and these elicited with concanavalin A, lipopolysaccharide, proteose peptone or Bacillus Clamette Guerin. Resident murine peritoneal cells maintained in tissue culture for 3 days begin to accumulate this protein as do human peripheral blood monocytes after 7 days of culture. A specific competitive displacement radioimmunoassay was developed using a rabbit antiserum raised to the partially purified calmodulin binding protein and (/sup 125/I) calmodulin covalently crosslinked to the principal calmodulin binding protein in the preparation. The radioimmunoassay confirmed the unique cellular distribution of this protein suggesting that it may be a marker for certain stages of macrophage differentiation. Monoclonal antibodies were prepared and one of these was used to further purify the protein by immunoaffinity chromatography. A protein of molecular weight 50,000 to 60,000 was isolated. It could be selectively adsorbed to wheat germ agglutinin agarose and subsequently eluted with N-acetyl glucosamine. This property plus its sensitivity to endoglycosidase F led to the conclusion that it is a glycoprotein. The cellular distribution, subcellular localization and evidence of glycosylation suggest that this protein may be a macrophage-specific receptor with a high affinity for calcium-calmodulin.

  2. Studies on a novel macrophage-specific calmodulin binding glycoprotein

    International Nuclear Information System (INIS)

    The murine macrophage-like cell line J774 and peritoneal exudate cells elicited with thioglycollate or starch contain a major calmodulin-binding protein which is absent in trifluoperazine-resistant variants of J774, resident peritoneal macrophages and these elicited with concanavalin A, lipopolysaccharide, proteose peptone or Bacillus Clamette Guerin. Resident murine peritoneal cells maintained in tissue culture for 3 days begin to accumulate this protein as do human peripheral blood monocytes after 7 days of culture. A specific competitive displacement radioimmunoassay was developed using a rabbit antiserum raised to the partially purified calmodulin binding protein and (125I) calmodulin covalently crosslinked to the principal calmodulin binding protein in the preparation. The radioimmunoassay confirmed the unique cellular distribution of this protein suggesting that it may be a marker for certain stages of macrophage differentiation. Monoclonal antibodies were prepared and one of these was used to further purify the protein by immunoaffinity chromatography. A protein of molecular weight 50,000 to 60,000 was isolated. It could be selectively adsorbed to wheat germ agglutinin agarose and subsequently eluted with N-acetyl glucosamine. This property plus its sensitivity to endoglycosidase F led to the conclusion that it is a glycoprotein. The cellular distribution, subcellular localization and evidence of glycosylation suggest that this protein may be a macrophage-specific receptor with a high affinity for calcium-calmodulin

  3. Identification of a modular pathogenicity island that is widespread among urease-producing uropathogens and shares features with a diverse group of mobile elements.

    Science.gov (United States)

    Flannery, Erika L; Mody, Lona; Mobley, Harry L T

    2009-11-01

    Pathogenicity islands (PAIs) are a specific group of genomic islands that contribute to genomic variability and virulence of bacterial pathogens. Using a strain-specific comparative genomic hybridization array, we report the identification of a 94-kb PAI, designated ICEPm1, that is common to Proteus mirabilis, Providencia stuartii, and Morganella morganii. These organisms are highly prevalent etiologic agents of catheter-associated urinary tract infections (caUTI), the most common hospital acquired infection. ICEPm1 carries virulence factors that are important for colonization of the urinary tract, including a known toxin (Proteus toxic agglutinin) and the high pathogenicity island of Yersinia spp. In addition, this PAI shares homology and gene organization similar to the PAIs of other bacterial pathogens, several of which have been classified as mobile integrative and conjugative elements (ICEs). Isolates from this study were cultured from patients with caUTI and show identical sequence similarity at three loci within ICEPm1, suggesting its transfer between bacterial genera. Screening for the presence of ICEPm1 among P. mirabilis colonizing isolates showed that ICEPm1 is more prevalent in urine isolates compared to P. mirabilis strains isolated from other body sites (P<0.0001), further suggesting that it contributes to niche specificity and is positively selected for in the urinary tract. PMID:19687197

  4. Rubella serology by solid-phase radioimmunoassay: its potential for screening programmes

    International Nuclear Information System (INIS)

    Sera from 269 adult females who had experienced naturally acquired or vaccine-induced infection by rubella virus, including immune persons challenged intranasally with rubella vaccine (RA27/3) as well as sera from 100 patients attending antenatal clinics, were tested for rubella antibodies by the conventional haemagglutination inhibition tests (HAI), as well as a newly developed solid-phase radioimmunoassay (RIA) for rubella immunoglobulin G(IgG) antibodies. Following both naturally acquired and vaccine-induced infection, titres by RIA were approximately ten-fold higher than by HAI. The RIA test was particularly useful in assessing the true immune status of those with apparently low levels of HAI antibody and has the added advantage that pretreatment of sera to remove inhibitors of haemagglutination and red cell agglutinins is unnecessary. The RIA test has potential for the large-scale screening programmes which need to be carried out if the Department of Health and Social Security recommendation, that women attending antenatal and family planning clinics be screened for rubella antibodies, is to be effectively met. (author)

  5. Delineation of downstream signalling components during acrosome reaction mediated by heat solubilized human zona pellucida

    Directory of Open Access Journals (Sweden)

    Talwar Pankaj

    2010-01-01

    Full Text Available Abstract Background Human egg is enveloped by a glycoproteinaceous matrix, zona pellucida (ZP, responsible for binding of the human spermatozoa to the egg and induction of acrosomal exocytosis in the spermatozoon bound to ZP. In the present manuscript, attempts have been made to delineate the downstream signalling components employed by human ZP to induce acrosome reaction. Methods Heat-solubilized human ZP (SIZP was used to study the induction of acrosome reaction in capacitated human spermatozoa using tetramethylrhodamine isothiocyanate conjugated Pisum sativum agglutinin (TRITC-PSA in absence or presence of various pharmacological inhibitors. In addition, intracellular calcium ([Ca2+]i levels in sperm using Fluo-3 acetoxymethyl ester as fluorescent probe were also estimated in response to SIZP. Results SIZP induces acrosomal exocytosis in capacitated human sperm in a dose dependent manner accompanied by an increase in [Ca2+]i. Human SIZP mediated induction of acrosome reaction depends on extracellular Ca2+ and involves activation of Gi protein-coupled receptor, tyrosine kinase, protein kinases A & C and phosphoinositide 3 (PI3- kinase. In addition, T-type voltage operated calcium channels and GABA-A receptor associated chloride (Cl- channels play an important role in SIZP mediated induction of acrosome reaction. Conclusions Results described in the present study provide a comprehensive account of the various downstream signalling components associated with human ZP mediated acrosome reaction.

  6. Nuclear pore complex contains a family of glycoproteins that includes p62: glycosylation through a previously unidentified cellular pathway

    International Nuclear Information System (INIS)

    Using a monoclonal antibody (mAb 414), the authors previously identified a protein of 62 kDa (p62) that was localized to the nuclear pore complex by immunoelectron microscopy. They also showed that p62 binds specifically to wheat germ agglutinin. Therefore, they proposed that this nuclear pore complex protein might be a member of a recently characterized family of glycoproteins that are labeled by in vitro galactosylation of rat liver nuclei and contain O-linked monosaccharidic GlcNAc residues. In support of this, they now show that incubation with N-acetylglucosaminidase reduces the molecular mass of p62 by ≅ 3 kDa because of the removal of terminal GlcNAc residues. Moreover, p62 can be galactosylated in vitro by using UDP-[3H]galactose and galactosyltransferase. They also show that most of the GlcNAc residues are added within 5 min of synthesis, when p62 is soluble and cytosolic. Thus, the addition of GlcNAc is carried out in the cytoplasm and is clearly distinct from the N- and O-linked glycosylation pathways of the endoplasmic reticulum and Golgi complex. Using another mAb with a broad specificity for nuclear GlcNAc-containing proteins, they show by immunofluorescence and protein blotting of subnuclear fractions that some of these proteins are in the interior of the nucleus, and others are most likely located in the pore complex

  7. Immunohistochemical Patterns in the Interfollicular Caucasian Scalps: Influences of Age, Gender, and Alopecia

    Directory of Open Access Journals (Sweden)

    Claudine Piérard-Franchimont

    2013-01-01

    Full Text Available Skin ageing and gender influences on the scalp have been seldom studied. We revisited the changes in the interfollicular scalp. The study was performed on a population of 650 volunteers (300 women and 350 men for over 7 years. Three age groups were selected in both genders, namely, subjects aged 20–35, 50–60, and 60–70 years. The hair status was further considered according to nonalopecic and alopecic patterns and severity (discrete, moderate, and severe. Biopsies from the parietal area were processed for immunohistochemistry. Stromal cells were distinguished according to the presence of vimentin, Factor XIIIa, CD117, and versican. Blood and lymphatic vessels were highlighted by Ulex europaeus agglutinin-1 and human podoplanin immunoreactivities, respectively. Actinic elastosis was identified by the lysozyme coating of elastic fibres. The epidermis was explored using the CD44 variant 3 and Ki67 immunolabellings. Biplot analyses were performed. Immunohistochemistry revealed a prominent gender effect in young adults. Both Factor XIIIa+ dermal dendrocytes and the microvasculature size decreased with scalp ageing. Alopecia changes mimicked stress-induced premature senescence.

  8. Nuclear and nucleolar localization signals and their targeting function in phosphatidylinositol 4-kinase PI4K230

    International Nuclear Information System (INIS)

    PI4K230, an isoform of phosphatidylinositol 4-kinase, known primarily as a cytoplasmic membrane-bound enzyme, was detected recently also in the nucleolus of several cells. Here we provide mechanistic insight on the targeting function of its putative nuclear localization signal (NLS) sequences using molecular modeling, digitonin-permeabilized HeLa cells and binding to various importins. The synthetic sequence 916NFNHIHKRIRRVADKYLSG934 comprising a putative monopartite NLS (NLS1), targeted covalently bound fluorescent BSA to the nucleoplasm via classical importin α/β mechanism employing importins α1 and α3 but not α5. This transport was inhibited by wheat germ agglutinin and GTPγS. The sequence 1414SKKTNRGSQLHKYYMKRRTL1433, a putative bipartite NLS (NLS2) proved ineffective in nuclear targeting if conjugated to fluorescently labeled BSA. Nonetheless, NLS2 or either of its basic clusters directed to the nucleolus soybean trypsin inhibitor that can pass the nuclear pore complex passively; moreover, an expressed 58 kDa fragment of PI4K230 (AA1166-1667) comprising NLS2 was also imported into the nucleus by import factors of reticulocyte lysate or by importin α1/β or α3/β complexes and localized to the nucleolus. We conclude that the putative bipartite NLS itself is a nucleolar targeting signal, and for nuclear import PI4K230 requires a larger sequence around it or, alternatively, the monopartite NLS

  9. Insulin and leptin enhance human sperm motility, acrosome reaction and nitric oxide production

    Institute of Scientific and Technical Information of China (English)

    Fanuel Lampiao; Stefan S. du Plessis

    2008-01-01

    Aim: To investigate the in vitro effects of insulin and leptin on human sperm motility, viability, acrosome reaction and nitric oxide (NO) production. Methods: Washed human spermatozoa from normozoospermic donors were treated with insulin (10 μIU) and leptin (10 nmol). Insulin and leptin effects were blocked by inhibition of their intracellular effector, phosphotidylinositol 3-kinase (PI3K), by wortmannin (10 μmol) 30 min prior to insulin and leptin being given. Computer-assisted semen analysis was used to assess motility after 1, 2 and 3 h of incubation. Viability was assessed by fluorescence-activated cell sorting using propidium iodide as a fluorescent probe. Acrosome-reacted cells were observed under a fluorescent microscope using fluorescein-isothiocyanate-Pisum sativum agglutinin as a probe. NO was measured after treating the sperm with 4,5-diaminofluorescein-2/diacetate (DAF-2/DA) and analyzed by fluorescence-activated cell sorting. Results: Insulin and leptin significantly increased total motility, progressive motility and acrosome reaction, as well as NO production. Conclusion: This study showed the in vitro beneficial effects of insulin and leptin on human sperm function. These hormones could play a role in enhancing the fertilization capacity of human spermatozoa.

  10. Cryopreservation of aoudad (Ammotragus lervia sahariensis) sperm obtained by transrectal ultrasound-guided massage of the accessory sex glands and electroejaculation.

    Science.gov (United States)

    Santiago-Moreno, J; Castaño, C; Toledano-Díaz, A; Esteso, M C; López-Sebastián, A; Guerra, R; Ruiz, M J; Mendoza, N; Luna, C; Cebrián-Pérez, J A; Hildebrandt, T B

    2013-01-15

    This study examines (1) the effectiveness of transrectal, ultrasound-guided massage of the accessory sex glands (TUMASG) combined with electroejaculation for obtaining aoudad (Ammotragus lervia sahariensis) sperm samples for cryopreservation, and (2) the effectiveness of a Tris-citric acid-glucose-based medium (TCG; usually used for freezing ibex sperm) and a TES-Tris-glucose-based medium (TTG; typically used in the cryopreservation of mouflon sperm) as sperm extenders. After TUMASG, just one to three electrical pulses were required for ejaculation to occur in five of the six animals studied; one ejaculated after TUMASG alone. Transrectal, ultrasound-guided massage of the accessory sex glands would therefore appear to be useful in obtaining sperm samples from this species, requiring few subsequent electrical electroejaculation stimuli and sometimes none at all. After thawing, the membrane integrity (assessed by nigrosin-eosin staining) of sperm extended with TTG was greater than that of sperm extended with TCG (P < 0.05). The total percentage of sperm showing an intact acrosome, as assessed by fluorescein isothiocyanate-conjugated peanut (Arachis hypogea) agglutinin, was also higher in the TTG-extended sperm (P < 0.05), and the percentage of dead sperm with a damaged acrosome was lower (P < 0.05). No differences were seen between TCG and TTG in terms of apoptotic manifestations (DNA damage, caspase activity, mitochondrial membrane potential, and plasmalemma stability). Therefore, TTG appears to be a better extender than TCG for cryopreserving aoudad sperm. PMID:23158213

  11. Expression of fucose in human cervical squamous carcinoma tissues and its clinical significance%岩藻糖基抗原在宫颈鳞癌的表达及意义

    Institute of Scientific and Technical Information of China (English)

    黄凤英; 袁建寰; 陈惠祯; 熊艳; 李玉春; 纪燕琴; 刘惠芬

    2003-01-01

    目的探讨人宫颈鳞癌组织的岩藻糖基(fucose,Fuc)化的水平及其意义.方法 1992年1月至1996年12月采用凝集素组织化学染色技术,以能与糖链α-Fuc特异性结合的生物素标记的荆豆凝集素(biotinylated ulex europeaus agglutinin,BUEA)检测了100例宫颈鳞癌、150例宫颈不典型增生及50例正常宫颈组织的Fuc表达,应用图像分析系统定量分析其表达水平.结果 Fuc在非癌组织中无表达,在宫颈癌组织中的表达率为76%,Fuc在细胞分化程度低、临床分期晚、有转移、复发及预后差的患者中的显著高表达(P<0.01或P<0.05).结论 Fuc的表达可作为反映宫颈癌恶性潜能和患者预后的一项新的指标.

  12. Ultrasensitive impedimetric lectin based biosensor for glycoproteins containing sialic acid

    Science.gov (United States)

    Bertok, Tomas; Gemeiner, Pavol; Mikula, Milan; Gemeiner, Peter; Tkac, Jan

    2016-01-01

    We report on an ultrasensitive label-free lectin-based impedimetric biosensor for the determination of the sialylated glycoproteins fetuin and asialofetuin. A sialic acid binding agglutinin from Sambucus nigra I was covalently immobilised on a mixed self-assembled monolayer (SAM) consisting of 11-mercaptoundecanoic acid and 6-mercaptohexanol. Poly(vinyl alcohol) was used as a blocking agent. The sensor layer was characterised by atomic force microscopy, electrochemical impedance spectroscopy and X-ray photoelectron spectroscopy. The biosensor exhibits a linear range that spans 7 orders of magnitude for both glycoproteins, with a detection limit as low as 0.33 fM for fetuin and 0.54 fM for asialofetuin. We also show, by making control experiments with oxidised asialofetuin, that the biosensor is capable of quantitatively detecting changes in the fraction of sialic acid on glycoproteins. We conclude that this work lays a solid foundation for future applications of such a biosensor in terms of the diagnosis of diseases such as chronic inflammatory rheumatoid arthritis, genetic disorders and cancer, all of which are associated with aberrant glycosylation of protein biomarkers.

  13. Sialic acid specific lectins from Episesarma tetragonum (Decapoda, Grapsidae: isolation, purification and characterization

    Directory of Open Access Journals (Sweden)

    R. Viswambari Devi

    2013-07-01

    Full Text Available Two sialic acid specific lectins Episesarma tetragonum agglutinin–1 and 2 were purified from the hemolymph of the Mangrove crab, Episesarma tetragonum. The major lectin was purified using CNBr-activated sepharose 4B conjugated to fetuin. N-acetyl glucosamine containing buffer was used for elution. The hemagglutination activity of purified lectin was inhibited by glycoproteins containing Siaα, 2-3Galβ, 1-4 GlcNAc linkages. On SDS-PAGE, the molecular weight of calcium dependent lectin was observed to be 70 kDa. Lectin had the maximum activity at a wide range of pH (6.5 – 9.5 and temperature (0 - 40 °C.  The physicochemical characteristics of the minor agglutinin showed that its hemagglutinating activity was calcium dependent, optimum at pH 8 – 9.5 and temperature 0 – 37 °C. The only potent inhibitor of minor lectin was bovine submaxillary mucin. An attempt was also made to purify minor lectin by affinity chromatography using bovine submaxillary mucin coupled to CNBr-activated sepharose 4B column. The lectin was eluted with elution buffer containing ethylene diamine tetra acetate. Strong inhibition of purified minor lectin by bovine submaxillary mucin and non-inhibitory action of de-O-acetylated bovine submaxillary mucin suggested that the lectin was O-acetyl sialic acid specific.

  14. The cryoprotective effect of Ficoll on the rabbit spermatozoa quality.

    Science.gov (United States)

    Kuliková, Barbora; Di Iorio, Michele; Kubovicova, Elena; Kuzelova, Lenka; Iaffaldano, Nicolaia; Chrenek, Peter

    2015-10-01

    The aim of this study was to evaluate the effect of the addition of Ficoll 70 into the cryopreservation medium containing sucrose and dimethyl sulfoxide (DMSO) on rabbit spermatozoa characteristics following freezing/thawing. This large molecular weight polymer elevates the viscosity of medium and, therefore, could better protect spermatozoa during the freezing process. Only ejaculates of good initial motility (>80%) were used in the experiments. Heterospermic pools were diluted in a freezing medium composed of commercial diluent, 16% dimethyl sulphoxide (DMSO) and 2% sucrose (control) or in the same medium enriched with 4% Ficoll 70 (Ficoll) and frozen in liquid nitrogen vapours for 10 min before being plunged in liquid nitrogen. The quality of fresh and frozen/thawed spermatozoa samples was evaluated in vitro using the Computer Assisted Semen Analysis (CASA) system, fluorescent probes (peanut agglutinin (PNA)-Alexa Fluor®; annexin V-FLOUS) and by electron microscopy. Better cryoprotective effect was observed when Ficoll 70 was added, compared with the semen cryopreserved with sucrose and DMSO only. The higher values (P semen group were found. In conclusion, our study showed that the addition of Ficoll 70 might improve several characteristics of rabbit spermatozoa measured in vitro following freezing/thawing. PMID:25255836

  15. An International Proficiency Test to Detect, Identify and Quantify Ricin in Complex Matrices

    Science.gov (United States)

    Worbs, Sylvia; Skiba, Martin; Bender, Jennifer; Zeleny, Reinhard; Schimmel, Heinz; Luginbühl, Werner; Dorner, Brigitte G.

    2015-01-01

    While natural intoxications with seeds of Ricinus communis (R. communis) have long been known, the toxic protein ricin contained in the seeds is of major concern since it attracts attention of those intending criminal, terroristic and military misuse. In order to harmonize detection capabilities in expert laboratories, an international proficiency test was organized that aimed at identifying good analytical practices (qualitative measurements) and determining a consensus concentration on a highly pure ricin reference material (quantitative measurements). Sample materials included highly pure ricin as well as the related R. communis agglutinin (RCA120) spiked into buffer, milk and meat extract; additionally, an organic fertilizer naturally contaminated with R. communis shred was investigated in the proficiency test. The qualitative results showed that either a suitable combination of immunological, mass spectrometry (MS)-based and functional approaches or sophisticated MS-based approaches alone successfully allowed the detection and identification of ricin in all samples. In terms of quantification, it was possible to determine a consensus concentration of the highly pure ricin reference material. The results provide a basis for further steps in quality assurance and improve biopreparedness in expert laboratories worldwide. PMID:26703726

  16. Partial purification of the 5-hydroxytryptophan-reuptake system from human blood platelets using a citalopram-derived affinity resin

    Energy Technology Data Exchange (ETDEWEB)

    Biessen, E.A.L; Horn, A.S.; Robillard, G.T. (Univ. of Groningen (Netherlands))

    1990-04-03

    This paper describes a procedure for the synthesis and application of a citalopram-derived affinity resin in purifying the 5HT-reuptake system from human blood platelets. A two-step scheme has been developed for partial purification, based on wheat germ agglutinin-lectin (WGA) affinity and citalopram affinity chromatographies. Upon solubilization of the carrier with 1% digitonin, a 50-70-fold increase in specific ({sup 3}H) imipramine binding activity with a 70% recovery could be accomplished through WGA-lectin chromatography. The WGA pool was then subjected to affinity chromatography on citalopram-agarose. At least 90% of the binding capacity adsorbed to the column. Specific elution using 10 {mu}M citalopram resulted in a 22% recovery of binding activity. A 10,000-fold overall purification was obtained by using this two-step procedure. Analysis of the fractions on SDS-PAGE after {sup 125}I labeling revealed specific elution of 78- and 55-kDa proteins concomitant with the appearance of ({sup 3}H) imipramine binding activity. The pharmacological profile of the partially purified reuptake system correlated well with that derived from the crude membrane-bound reuptake system, suggesting a copurification of the 5HT binding activity and ({sup 3}H)imipramine binding activity.

  17. Radioiodine labeled CdSe/CdS quantum dots. Lectin targeted dual probes

    International Nuclear Information System (INIS)

    CdSe/CdS quantum dots (QD) were synthesized and bioconjugated with Sambucus nigra agglutinin (SNA) lectin (Lec). Mannose triflate and cysteamine molecules (MTC) were also utilized to prepare MTC-QDs and MTC-QDs-Lec probes as well as Lec bound QDs. Afterwards; potential use of these nanoparticles as radiolabeled fluorescence nano-probes for the cell imaging studies has been investigated. Biological activities of 125I-, 125I-MTC-QDs, MTC-QDs- Lec-125I, QDs-Lec-125I and Lec-125I were examined on various cancer cell lines such as Caco-2, MCF-7 and A-549 in terms of cell incorporation. QDs-Lec-125I exhibited the highest cell incorporation on whole cell lines. In addition, the QDs-Lec-131I, was used for in vivo imaging. The whole body distribution of the radiolabeled QDs on New Zealand rabbits and Balb C mice were examined by taking dynamic and static images. Radioactivity cleared from the kidneys and the bladder, while significant amount radioactivity was retained in the heart and liver within 24 h.

  18. Spur cell anaemia and acute haemolysis in patients with hyperreactive malarious splenomegaly. Experience in an isolated Yanomamo population of Venezuela.

    Science.gov (United States)

    Torres R, J R; Magris, M; Villegas, L; Torres V, M A; Dominguez, G

    2000-12-01

    A prospective study, aimed to investigate the aetiology of an unusual clustering of cases of severe acute haemolytic anaemia affecting a high percentage of the adult population, was carried out in two isolated Yanomamo communities of the Upper Orinoco basin in Venezuela. Twenty-six patients with active or recent episodes of severe haemolysis were evaluated. All of them exhibited massive liver and spleen enlargement and fulfilled the diagnostic criteria of the hyperreactive malarious splenomegaly (HMS) syndrome. In four cases with advanced non-alcohol-related chronic liver disease, hypersplenism, severe haemolytic anaemia and acanthocytosis, the characteristic clinical and laboratory findings of spur cell anaemia were documented. Chronic infection by the HBV and HCV was present in three of them. However, in most of the 22 additional HMS cases, the acute haemolytic condition appeared associated with the occurrence of a cold agglutinin-mediated autoimmune response. The clustering of a significant number of cases of severe acute haemolysis in HMS patients from this small isolated aboriginal community is most unusual, and represents a serious complicating factor for a population already beleaguered by a high prevalence of malaria due to multiresistant strains of Plasmodium falciparum. Moreover, the coexistence of HMS and severe chronic HBV or HCV infection may further aggravate the course of the haemolytic disorder, because of the occurrence of spur cell anaemia. PMID:11114387

  19. Bis(β-lactosyl-[60]fullerene as novel class of glycolipids useful for the detection and the decontamination of biological toxins of the Ricinus communis family

    Directory of Open Access Journals (Sweden)

    Hirofumi Dohi

    2014-07-01

    Full Text Available Glycosyl-[60]fullerenes were first used as decontaminants against ricin, a lactose recognition proteotoxin in the Ricinus communis family. A fullerene glycoconjugate carrying two lactose units was synthesized by a [3 + 2] cycloaddition reaction between C60 and the azide group in 6-azidohexyl β-lactoside per-O-acetate. A colloidal aqueous solution with brown color was prepared from deprotected bis(lactosyl-C60 and was found stable for more than 6 months keeping its red color. Upon mixing with an aqueous solution of Ricinus communis agglutinin (RCA120, the colloidal solution soon caused precipitations, while becoming colorless and transparent. In contrast, a solution of concanavalin A (Con A caused no apparent change, indicating that the precipitation was caused specifically by carbohydrate–protein interactions. This notable phenomenon was quantified by means of sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE, and the results were discussed in terms of detection and decontamination of the deadly biological toxin in the Ricinus communis family.

  20. In vitro induction of Entamoeba histolytica cyst-like structures from trophozoites.

    Science.gov (United States)

    Aguilar-Díaz, Hugo; Díaz-Gallardo, Martha; Laclette, Juan P; Carrero, Julio C

    2010-01-01

    Inhibition of encystment can be conceived as a potentially useful mechanism to block the transmission of Entamoeba histolytica under natural conditions. Unfortunately, amoeba encystment has not been achieved in vitro and drugs inhibiting the formation of cysts are not available. Luminal conditions inducing encystment in vivo are also unknown, but cellular stress such as exposure to reactive oxygen species from immune cells or intestinal microbiota could be involved. A role for certain divalent cations as cofactors of enzymes involved in excystment has also been described. In this study, we show that trophozoite cultures, treated with hydrogen peroxide in the presence of trace amounts of several cations, transform into small-sized spherical and refringent structures that exhibit resistance to different detergents. Ultrastructural analysis under scanning and transmission electron microscopy revealed multinucleated structures (some with four nuclei) with smooth, thick membranes and multiple vacuoles. Staining with calcofluor white, as well as an ELISA binding assay using wheat germ agglutinin, demonstrated the presence of polymers of N-acetylglucosamine (chitin), which is the primary component of the natural cyst walls. Over-expression of glucosamine 6-phosphate isomerase, likely to be the rate-limiting enzyme in the chitin synthesis pathway, was also confirmed by RT-PCR. These results suggest that E. histolytica trophozoites activated encystment pathways when exposed to our treatment. PMID:20169067

  1. Alarm pheromone is detected by the vomeronasal organ in male rats.

    Science.gov (United States)

    Kiyokawa, Yasushi; Kodama, Yuka; Kubota, Takahiro; Takeuchi, Yukari; Mori, Yuji

    2013-10-01

    It is widely known that a stressed animal releases specific pheromones, possibly for alarming nearby conspecifics. We previously investigated an alarm pheromone in male rats and found that this alarm pheromone evokes several responses, including increases in the defensive and risk assessment behaviors in a modified open-field test, and enhancement of the acoustic startle reflex. However, the role of the vomeronasal organ in these pheromone effects remains unclear. To clarify this point, vomeronasal organ-excising or sham surgeries were performed in male rats for use in 2 experimental models, after which they were exposed to alarm pheromone. We found that the vomeronasal organ-excising surgery blocked the effects of this alarm pheromone in both the modified open-field test and acoustic startle reflex test. In addition, the results of habituation/dishabituation test and soybean agglutinin binding to the accessory olfactory bulb suggested that the vomeronasal organ-excising surgery completely ablated the vomeronasal organ while preserving the functioning of the main olfactory system. From the above results, we showed that the vomeronasal organ plays an important role in alarm pheromone effects in the modified open-field test and acoustic startle reflex test. PMID:23821727

  2. The Toxoplasma gondii cyst wall protein CST1 is critical for cyst wall integrity and promotes bradyzoite persistence

    Energy Technology Data Exchange (ETDEWEB)

    Tomita, Tadakimi; Bzik, David J.; Ma, Yan Fen; Fox, Barbara A.; Markillie, Lye Meng; Taylor, Ronald C.; Kim, Kami; Weiss, Louis M.

    2013-12-26

    Toxoplasma gondii infects up to one third of the world’s population. A key to the success of T.gondii is its ability to persist for the life of its host as bradyzoites within tissue cysts. The glycosylated cyst wall is the key structural feature that facilitates persistence and oral transmission of this parasite. We have identified CST1 (TGME49_064660) as a 250 kDa SRS (SAG1 related sequence) domain protein with a large mucin-like domain. CST1 is responsible for the Dolichos biflorus Agglutinin (DBA) lectin binding characteristic of T. gondii cysts. Deletion of CST1 results in a fragile brain cyst phenotype revealed by a thinning and disruption of the underlying region of the cyst wall. These defects are reversed by complementation of CST1. Additional complementation experiments demonstrate that the CST1-mucin domain is necessary for the formation of a normal cyst wall structure, the ability of the cyst to resist mechanical stress and binding of DBA to the cyst wall. RNA-seq transcriptome analysis demonstrated dysregulation of bradyzoite genes within the various cst1 mutants. These results indicate that CST1 functions as a key structural component that reinforces the cyst wall structure and confers essential sturdiness to the T. gondii tissue cyst.

  3. Combining microtomy and confocal laser scanning microscopy for structural analyses of plant-fungus associations.

    Science.gov (United States)

    Rath, Magnus; Grolig, Franz; Haueisen, Janine; Imhof, Stephan

    2014-05-01

    The serious problem of extended tissue thickness in the analysis of plant-fungus associations was overcome using a new method that combines physical and optical sectioning of the resin-embedded sample by microtomy and confocal microscopy. Improved tissue infiltration of the fungal-specific, high molecular weight fluorescent probe wheat germ agglutinin conjugated to Alexa Fluor® 633 resulted in high fungus-specific fluorescence even in deeper tissue sections. If autofluorescence was insufficient, additional counterstaining with Calcofluor White M2R or propidium iodide was applied in order to visualise the host plant tissues. Alternatively, the non-specific fluorochrome acid fuchsine was used for rapid staining of both, the plant and the fungal cells. The intricate spatial arrangements of the plant and fungal cells were preserved by immobilization in the hydrophilic resin Unicryl™. Microtomy was used to section the resin-embedded roots or leaves until the desired plane was reached. The data sets generated by confocal laser scanning microscopy of the remaining resin stubs allowed the precise spatial reconstruction of complex structures in the plant-fungus associations of interest. This approach was successfully tested on tissues from ectomycorrhiza (Betula pendula), arbuscular mycorrhiza (Galium aparine; Polygala paniculata, Polygala rupestris), ericoid mycorrhiza (Calluna vulgaris), orchid mycorrhiza (Limodorum abortivum, Serapias parviflora) and on one leaf-fungus association (Zymoseptoria tritici on Triticum aestivum). The method provides an efficient visualisation protocol applicable with a wide range of plant-fungus symbioses. PMID:24249491

  4. Liver Full Reference Set Application: Hiro Yamada - Wako (2011) — EDRN Public Portal

    Science.gov (United States)

    Wako has received new 510(k) clearance for Lens culinaris agglutinin-reactive fraction of alpha-fetoprotein (AFP-L3) and Des-gamma-carboxy prothrombin (DCP) tests on an innovative μTASWako i30 analyzer from FDA. The AFP-L3 and DCP assayed on an older platform LiBASys have been cleared with indication of use for risk assessment of hepatocellular carcinoma (HCC) in patient at risk for the liver malignancy. Wako believes that early detection of HCC is critical for improving HCC patient outcome. Therefore, Wako is currently seeking collaborative opportunities to retrospectively measure clinical samples using the AFP-L3 and DCP for further determining of effectiveness of the HCC biomarkers in early detection which are collected prospectively during HCC surveillance. The Reference Sample Set in the EDRN biorepository are well characterized and studied. Access to these samples would allow Wako to quickly determine the clinical effectiveness of AFP-L3 and DCP in detecting early HCC

  5. Labeling of lectin receptors during the cell cycle.

    Science.gov (United States)

    Garrido, J

    1976-12-01

    Labeling of lectin receptors during the cell cycle. (Localizabión de receptores para lectinas durante el ciclo celular). Arch. Biol. Med. Exper. 10: 100-104, 1976. The topographic distribution of specific cell surface receptors for concanavalin A and wheat germ agglutinin was studied by ultrastructural labeling in the course of the cell cycle. C12TSV5 cells were synchronized by double thymidine block or mechanical selection (shakeoff). They were labeled by means of lectin-peroxidase techniques while in G1 S, G2 and M phases of the cycle. The results obtained were similar for both lectins employed. Interphase cells (G1 S, G2) present a stlihtly discontinous labeling pattern that is similar to the one observed on unsynchronized cells of the same line. Cells in mitosis, on the contrary, present a highly discontinous distribution of reaction product. This pattern disappears after the cells enters G1 and is not present on mitotic cells fixed in aldehyde prior to labeling. PMID:1030938

  6. Sialylated immunoglobulin G can neutralize influenza virus infection through receptor mimicry

    Science.gov (United States)

    Zhao, Conghui; Liu, Xingmu; Zhang, Zaiping; Li, Tongfei; Sun, Ruiman; Gu, Huan; Gu, Jiang

    2016-01-01

    Influenza viruses possess a great threat to human health, but there is still no effective drug to deal with the outbreak of possible new influenza subtypes. In this study, we first fractionated sialylated immunoglobulin G (IgG), mainly Fab sialylated fraction, with sambucus nigra agglutinin affinity chromatography. We then demonstrated that sialylated IgG possessed more effective neutralizing activity against 2009 A (H1N1) subtype than that of IgG mixture, and sialosides on the Fab is crucial in this neutralization reaction as when such residues were removed with neuraminidase A digestion the blocking effect was significantly reduced. It appears that sialic acid residues attached to Fab could serve as binding moieties to receptor binding site of influenza virus. These findings indicate that sialylated IgG probably is an effective anti-influenza broad-spectrum drug utilizing its receptor mimicry to competitively inhibit the attachment of influenza viruses with sialic acid receptors on target cells. This property would be particularly useful if it can be applied to prevent newly emerged influenza virus strain infections in future epidemics. PMID:26870994

  7. Nanostructured materials with biomimetic recognition abilities for chemical sensing

    Science.gov (United States)

    Bajwa, Sadia Zafar; Mustafa, Ghulam; Samardzic, Renata; Wangchareansak, Thipvaree; Lieberzeit, Peter A.

    2012-06-01

    Binding features found in biological systems can be implemented into man-made materials to design nanostructured artificial receptor matrices which are suitable, e.g., for chemical sensing applications. A range of different non-covalent interactions can be utilized based on the chemical properties of the respective analyte. One example is the formation of coordinative bonds between a polymerizable ligand (e.g., N-vinyl-2-pyrrolidone) and a metal ion (e.g., Cu(II)). Optimized molecularly imprinted sensor layers lead to selectivity factors of at least 2 compared to other bivalent ions. In the same way, H-bonds can be utilized for such sensing purposes, as shown in the case of Escherichia coli. The respective molecularly imprinted polymer leads to the selectivity factor of more than 5 between the W and B strains, respectively. Furthermore, nanoparticles with optimized Pearson hardness allow for designing sensors to detect organic thiols in air. The `harder' MoS2 yields only about 40% of the signals towards octane thiol as compared to the `softer' Cu2S. However, both materials strongly prefer molecules with -SH functionality over others, such as hydrocarbon chains. Finally, selectivity studies with wheat germ agglutinin (WGA) reveal that artificial receptors yield selectivities between WGA and bovine serum albumin that are only about a factor of 2 which is smaller than natural ligands.

  8. Mechanisms underlying the impaired contractility of diabetic cardiomyopathy

    Institute of Scientific and Technical Information of China (English)

    Marie-Louise; Ward; David; J; Crossman

    2014-01-01

    Cardiac dysfunction is a well-known consequence of diabetes,with sustained hyperglycaemia leading to the development of a cardiomyopathy that is independent of cardiovascular disease or hypertension.Animal models of diabetes are commonly used to study the pathophysiology of diabetic cardiomyopathy,with the hope that increased knowledge will lead ultimately to better therapeutic strategies being developed.At physiological temperature,left ventricular trabeculae isolated from the streptozotocin rat model of type 1 diabetes showed decreased stress and prolonged relaxation,but with no evidence that decreased contractility was a result of altered myocardial Ca2+handling.Although sarcoplasmic reticulum(SR)Ca2+reuptake appeared slower in diabetic trabeculae,it was offset by an increase in actionpotential duration,thereby maintaining SR Ca2+content and favouring increased contraction force.Frequency analysis of t-tubule distribution by confocal imaging of ventricular tissue labeled with wheat germ agglutinin or ryanodine receptor antibodies showed a reduced T-power for diabetic tissue,but the differences were minor in comparison to other models of heart failure.The contractile dysfunction appeared to be the result of disrupted F-actin in conjunction with the increased typeⅠcollagen,with decreased myofilament Ca2+sensitivity contributing to the slowed relaxation.

  9. Long-term exposure to arsenic affects head kidney and impairs humoral immune responses of Clarias batrachus

    Energy Technology Data Exchange (ETDEWEB)

    Ghosh, Debabrata [Immunobiology Laboratory, School of Life Sciences, Visva-Bharati University, Santiniketan 731235 (India); Datta, Soma [Immunobiology Laboratory, School of Life Sciences, Visva-Bharati University, Santiniketan 731235 (India); Bhattacharya, Shelley [Environmental Toxicology Laboratory, School of Life Sciences, Visva-Bharati University, Santiniketan 731235 (India); Mazumder, Shibnath [Immunobiology Laboratory, School of Life Sciences, Visva-Bharati University, Santiniketan 731235 (India)]. E-mail: shibnath1@yahoo.co.in

    2007-02-15

    The present study was aimed at determining the effects of long-term arsenic exposure on the head kidney (HK) and ensuing humoral immune responses in Clarias batrachus L. Long-term exposure (150 days) to non-lethal concentrations of arsenic (42.42 {mu}M) resulted in significant time-dependent alterations in HK cell number eventually affecting the HK somatic index. Prolonged exposure to arsenic also suppressed HK-B cell proliferation and led to significant reduction in serum immunoglobulin levels and antigen-specific serum bacterial agglutinin titers. A decline in the number of antigen-specific plaque-forming cells with duration of arsenic exposure was noted in the HK. Enzyme linked immunosorbent assays further revealed that arsenic exposure inhibited the release of 'IL-4 like factors' from HK-T cells. Histological studies documented time-dependent changes in the structure and cellular composition of HK characterized by extensive lymphocytopenia, decrease in melano-macrophage population and hemosiderin accumulation. From exposure-challenge studies with Aeromonas hydrophila it was evident that pathogens could efficiently disseminate and colonize distant host tissues in the exposed fish. Moreover, the ability to decrease the pathogen load was also significantly reduced in the arsenic-exposed fish. Thus long-term exposure to non-lethal concentrations of arsenic affects HK and interferes with the humoral immune system of C. batrachus rendering them immunocompromised and susceptible to pathogenic challenge.

  10. Role of Chitin-Binding Proteins in the Specific Attachment of the Marine Bacterium Vibrio harveyi to Chitin

    Science.gov (United States)

    Montgomery, Michael T.; Kirchman, David L.

    1993-01-01

    We examined the mechanism of attachment of the marine bacterium Vibrio harveyi to chitin. Wheat germ agglutinin and chitinase bind to chitin and competitively inhibited the attachment of V. harveyi to chitin, but not to cellulose. Bovine serum albumin and cellulase do not bind to chitin and had no effect on bacterial attachment to chitin. These data suggest that this bacterium recognizes specific attachment sites on the chitin particle. The level of attachment of a chitinase-overproducing mutant of V. harveyi to chitin was about twice as much as that of the uninduced wild type. Detergent-extracted cell membranes inhibited attachment and contained a 53-kDa peptide that was overproduced by the chitinase-overproducing mutant. Three peptides (40, 53, and 150 kDa) were recovered from chitin which had been exposed to membrane extracts. Polyclonal antibodies raised against extracellular chitinase cross-reacted with the 53- and 150-kDa chitin-binding peptides and inhibited attachment, probably by sterically hindering interactions between the chitin-binding peptides and chitin. The 53- and 150-kDa chitin-binding peptides did not have chitinase activity. These results suggest that chitin-binding peptides, especially the 53-kDa chitin-binding peptide and chitinase and perhaps the 150-kDa peptide, mediate the specific attachment of V. harveyi to chitin. Images PMID:16348865

  11. In vitro induction of Entamoeba histolytica cyst-like structures from trophozoites.

    Directory of Open Access Journals (Sweden)

    Hugo Aguilar-Díaz

    Full Text Available Inhibition of encystment can be conceived as a potentially useful mechanism to block the transmission of Entamoeba histolytica under natural conditions. Unfortunately, amoeba encystment has not been achieved in vitro and drugs inhibiting the formation of cysts are not available. Luminal conditions inducing encystment in vivo are also unknown, but cellular stress such as exposure to reactive oxygen species from immune cells or intestinal microbiota could be involved. A role for certain divalent cations as cofactors of enzymes involved in excystment has also been described. In this study, we show that trophozoite cultures, treated with hydrogen peroxide in the presence of trace amounts of several cations, transform into small-sized spherical and refringent structures that exhibit resistance to different detergents. Ultrastructural analysis under scanning and transmission electron microscopy revealed multinucleated structures (some with four nuclei with smooth, thick membranes and multiple vacuoles. Staining with calcofluor white, as well as an ELISA binding assay using wheat germ agglutinin, demonstrated the presence of polymers of N-acetylglucosamine (chitin, which is the primary component of the natural cyst walls. Over-expression of glucosamine 6-phosphate isomerase, likely to be the rate-limiting enzyme in the chitin synthesis pathway, was also confirmed by RT-PCR. These results suggest that E. histolytica trophozoites activated encystment pathways when exposed to our treatment.

  12. Exploitation of phosphorescent labelling reagent of fullerol-fluorescein isothiocyanate and new method for the determination of trace alkaline phosphatase as well as forecast of human diseases

    International Nuclear Information System (INIS)

    A new phosphorescent labelling reagent consisting of fullerol, fluorescein isothiocyanate and N,N-dimethylaniline (F-ol-(FITC)n-DMA) was developed. The mode of action is based on the reactivity of the active -OH group in F-ol with the -COOH group of FITC to form an F-ol-(FITC)n-DMA complex containing several FITC molecules. F-ol-(FITC)n-DMA increased the number of luminescent molecules in the biological target of WGA-AP-WGA-F-ol-(FITC)n-DMA (WGA and AP are wheat germ agglutinin and alkaline phosphatase, respectively) which improved the sensitivity using solid substrate room temperature phosphorimetry (SSRTP) detection. The proposed method provided high sensitivity and strong specificity for WGA-AP. The limit of detection (LD) was 0.15 ag AP spot-1 for F-ol and 0.097 ag AP spot-1 for FITC in F-ol-(FITC)n-DMA, which was lower than the method using single luminescent molecules of F-ol-DMA and FITC-DMA to label WGA (0.20 ag AP spot-1 for F-ol-DMA and 0.22 ag AP spot-1 for FITC-DMA). Results for the determination of AP in human serum were in good agreement with those obtained by enzyme-linked immunosorbent assay. The mechanism of F-ol-(FITC)n-DMA labelling of WGA was discussed.

  13. Transporter protein and drug-conjugated gold nanoparticles capable of bypassing the blood-brain barrier.

    Science.gov (United States)

    Zhang, Yanhua; Walker, Janelle Buttry; Minic, Zeljka; Liu, Fangchao; Goshgarian, Harry; Mao, Guangzhao

    2016-01-01

    Drug delivery to the central nervous system (CNS) is challenging due to the inability of many drugs to cross the blood-brain barrier (BBB). Here, we show that wheat germ agglutinin horse radish peroxidase (WGA-HRP) chemically conjugated to gold nanoparticles (AuNPs) can be transported to the spinal cord and brainstem following intramuscular injection into the diaphragm of rats. We synthesized and determined the size and chemical composition of a three-part nanoconjugate consisting of WGA-HRP, AuNPs, and drugs for the treatment of diaphragm paralysis associated with high cervical spinal cord injury (SCI). Upon injection into the diaphragm muscle of rats, we show that the nanoconjugate is capable of delivering the drug at a much lower dose than the unconjugated drug injected systemically to effectively induce respiratory recovery in rats following SCI. This study not only demonstrates a promising strategy to deliver drugs to the CNS bypassing the BBB but also contributes a potential nanotherapy for the treatment of respiratory muscle paralysis resulted from cervical SCI. PMID:27180729

  14. NMR-guided molecular docking of a protein-peptide complex based on ant colony optimization.

    Science.gov (United States)

    Korb, Oliver; Möller, Heiko M; Exner, Thomas E

    2010-07-01

    Standard docking approaches used for the prediction of protein-ligand complexes in the drug development process have problems identifying the correct binding mode of large flexible ligands. Herein we show how additional experimental data from NMR experiments can be used to predict the binding mode of a mucin 1 (MUC-1) pentapeptide recognized by the breast-cancer-selective monoclonal antibody SM3. Distance constraints derived from trNOE and saturation transfer difference NMR experiments are combined with the docking approach PLANTS. The resulting complex structures show excellent agreement with the NMR data and with a published X-ray crystal structure. The method was then further tested on two complexes in order to demonstrate its more general applicability: T-antigen disaccharide bound to Maclura pomifera agglutinin, and the inhibitor SBi279 bound to S100B protein. Our new approach has the advantages of being fully automatic, rapid, and unbiased; moreover, it is based on relatively easily obtainable experimental data and can greatly increase the reliability of the generated structures. PMID:20486157

  15. The evaluation of a PCR-based method for identification of Salmonella enterica serotypes from environmental samples and various food matrices.

    Science.gov (United States)

    Jean-Gilles Beaubrun, Junia; Cheng, Chorng-Ming; Chen, Kai-Shun; Ewing, Laura; Wang, Hua; Agpaoa, Maria C; Huang, Mei-Chiung J; Dickey, Erin; Du, Jamie M; Williams-Hill, Donna M; Hamilton, Brittany; Micallef, Shirley A; Rosenberg Goldstein, Rachel E; George, Ashish; Joseph, Sam W; Sapkota, Amy R; Jacobson, Andrew P; Tall, Ben D; Kothary, Mahendra H; Dudley, Kim; Hanes, Darcy E

    2012-09-01

    The most commonly used method for serotyping Salmonella spp. is based on the Kaufmann-White scheme, and is composed of serological reactions using antibodies to LPS agglutinins. The multiplex PCR used in this investigation was established by Kim et al. to serotype the 30 most common clinical Salmonella serotypes, as determined by CDC. The PCR assay consists of two five-plex reactions and a single two-plex PCR reaction, based on six genetic loci from Salmonella enterica serotype Typhimurium and four loci from S. enterica serotype Typhi. In this investigation, we further evaluated the method for serotyping Salmonella spp. using a reference collection, environmental samples collected from a Mid-Atlantic region tomato farm study, four food matrices spiked with different Salmonella serotypes and a proficiency test. The PCR assay was first evaluated using DNA isolated from pure cultures of isolates obtained from various clinical and environmental samples, and then DNA isolated from broth cultures of food matrices of "Red round" and Roma tomatoes, Romaine lettuce, green onions and Serrano peppers spiked with serotypes Newport, Typhimurium, Javiana and Saintpaul, respectively. The results showed that the PCR assay correctly serotyped Salmonella spp. from the clinical, environmental, spiked food matrices, and proficiency test samples. These findings are significant because the PCR assay was successful in the identification of Salmonella in the spiked samples in a broth culture containing other non-salmonella organism. This method may be a useful resource for the food safety community. PMID:22608224

  16. Enhancement of resistance to aphids by introducing the snowdrop lectin gene gna into maize plants

    Indian Academy of Sciences (India)

    Zhaoyu Wang; Kewei Zhang; Xiaofen Sun; Kexuan Tang; Juren Zhang

    2005-12-01

    In order to enhance the resistance to pests, transgenic maize (Zea mays L.) plants from elite inbred lines containing the gene encoding snowdrop lectin (Galanthus nivalis L. agglutinin; GNA) under control of a phloemspecific promoter were generated through the Agrobacterium tumefaciens-mediated method. The toxicity of GNA-expressing plants to aphids has also been studied. The independently derived plants were subjected to molecular analyses. Polymerase chain reaction (PCR) and Southern blot analyses confirmed that the gna gene was integrated into maize genome and inherited to the following generations. The typical Mendelian patterns of inheritance occurred in most cases. The level of GNA expression at 0.13%–0.28% of total soluble protein was observed in different transgenic plants. The progeny of nine GNA-expressing independent transformants that were derived separately from the elite inbred lines DH4866, DH9942, and 8902, were selected for examination of resistance to aphids. These plants synthesized GNA at levels above 0.22% total soluble protein, and enhanced resistance to aphids was demonstrated by exposing the plants to corn leaf aphid (Rhopalosiphum maidis Fitch) under greenhouse conditions. The nymph production was significantly reduced by 46.9% on GNA-expressing plants. Field evaluation of the transgenic plants supported the results from the inoculation trial. After a series of artificial self-crosses, some homozygous transgenic maize lines expressing GNA were obtained. In the present study, we have obtained new insect-resistant maize material for further breeding work.

  17. Evidence for the involvement of surface carbohydrates in the recognition of Haematococcus pluvialis by the parasitic blastoclad Paraphysoderma sedebokerensis.

    Science.gov (United States)

    Gutman, Jenia; Zarka, Aliza; Boussiba, Sammy

    2011-08-01

    The unicellular green alga Haematococcus pluvialis (Chlorophyta, Volvocales) is currently the best commercial source of the natural red ketocarotenoid astaxanthin. Paraphysoderma sedebokerensis (Blastocladiomycota), a parasitic blastoclad that is specific for this microalga, was recently isolated and identified in our laboratory. In this study, we investigated the recognition process between the parasite and H. pluvialis. Obligatory requirements for recognition were identified as an ion concentration in the medium of 20 mM, the presence of calcium ions, and neutral to basic conditions; these requirements imply that a protein is involved in the process. In a search for potential lectin-sugar interactions as a major event in the recognition process, we screened for exposed glycosidic moieties on the cell wall of the alga and on the parasite zoospore surface. Competition experiments with the appropriate lectins and monosugars identified Ricinus communis agglutinin (RCA(120)) as the lectin that recognizes Gal-N-acetyl-d-glucosamine, an oligosaccharide located on the host. We propose that an RCA(120)-like lectin-sugar interaction mediates the highly specific interaction between the blastocladian parasite and its algal host. PMID:21802061

  18. Endothelial progenitor cells regenerate infracted myocardium with neovascularisation development.

    Science.gov (United States)

    Abd El Aziz, M T; Abd El Nabi, E A; Abd El Hamid, M; Sabry, D; Atta, H M; Rahed, L A; Shamaa, A; Mahfouz, S; Taha, F M; Elrefaay, S; Gharib, D M; Elsetohy, Khaled A

    2015-03-01

    We achieved possibility of isolation, characterization human umbilical cord blood endothelial progenitor cells (EPCs), examination potency of EPCs to form new blood vessels and differentiation into cardiomyoctes in canines with acute myocardial infarction (AMI). EPCs were separated and cultured from umbilical cord blood. Their phenotypes were confirmed by uptake of double stains dioctadecyl tetramethylindocarbocyanine-labeled acetylated LDL and FITC-labeled Ulex europaeus agglutinin 1 (DILDL-UEA-1). EPCs of cord blood were counted. Human VEGFR-2 and eNOS from the cultured EPCs were assessed by qPCR. Human EPCs was transplanted intramyocardially in canines with AMI. ECG and cardiac enzymes (CK-MB and Troponin I) were measured to assess severity of cellular damage. Histopathology was done to assess neovascularisation. Immunostaining was done to detect EPCs transdifferentiation into cardiomyocytes in peri-infarct cardiac tissue. qPCR for human genes (hVEGFR-2, and eNOS) was done to assess homing and angiogenic function of transplanted EPCs. Cultured human cord blood exhibited an increased number of EPCs and significant high expression of hVEGFR-2 and eNOS genes in the culture cells. Histopathology showed increased neovascularization and immunostaining showed presence of EPCs newly differentiated into cardiomyocyte-like cells. Our findings suggested that hEPCs can mediate angiogenesis and differentiate into cardiomyoctes in canines with AMI. PMID:25750747

  19. Expression of serotonin receptor genes in cranial ganglia.

    Science.gov (United States)

    Maeda, Naohiro; Ohmoto, Makoto; Yamamoto, Kurumi; Kurokawa, Azusa; Narukawa, Masataka; Ishimaru, Yoshiro; Misaka, Takumi; Matsumoto, Ichiro; Abe, Keiko

    2016-03-23

    Taste cells release neurotransmitters to gustatory neurons to transmit chemical information they received. Sweet, umami, and bitter taste cells use ATP as a neurotransmitter. However, ATP release from sour taste cells has not been observed so far. Instead, they release serotonin when they are activated by sour/acid stimuli. Thus it is still controversial whether sour taste cells use ATP, serotonin, or both. By reverse transcription-polymerase chain reaction and subsequent in situ hybridization (ISH) analyses, we revealed that of 14 serotonin receptor genes only 5-HT3A and 5-HT3B showed significant/clear signals in a subset of neurons of cranial sensory ganglia in which gustatory neurons reside. Double-fluorescent labeling analyses of ISH for serotonin receptor genes with wheat germ agglutinin (WGA) in cranial sensory ganglia of pkd1l3-WGA mice whose sour neural pathway is visualized by the distribution of WGA originating from sour taste cells in the posterior region of the tongue revealed that WGA-positive cranial sensory neurons rarely express either of serotonin receptor gene. These results suggest that serotonin receptors expressed in cranial sensory neurons do not play any role as neurotransmitter receptor from sour taste cells. PMID:26854841

  20. Interactions of egg yolk phosphatidylcholine with cholesteryl polyethoxy neoglycolipids containing N-acetyl- D-glucosamine

    Science.gov (United States)

    Kemoun, Rachida; Gelhausen, Micaèle; Besson, Françoise; Lafont, Dominique; Buchet, René; Boullanger, Paul; Roux, Bernard

    1999-03-01

    Series of neoglycolipids containing cholesteryl and N-acetyl- D-glucosaminyl groups were synthesized with various ethoxy linkers. Their self aggregations and intermolecular interactions, without and with egg yolk phosphatidylcholine (EYPC), were characterized in dry and hydrated states, by using infrared spectroscopy. The neoglycolipids in the dry state formed intermolecular hydrogen bonds between the CO and N-H or O-H groups of N-acetyl- D-glucosamine (GlcNAc). In the presence of EYPC, these intermolecular interactions were broken and new hydrogen bonds, involving the phosphate group of EYPC and N-H or O-H groups of GlcNAc of neoglycolipid, were formed. The presence of water molecules altered these intermolecular hydrogen bonds. The CO groups of EYPC were not affected by the presence of neoglycolipids, either in hydrated or in dry states, indicating that the GlcNAc polar groups interacted mostly with EYPC phosphate residues. The phase transition-temperature of mixtures of EYPC containing either cholesterol or neoglycolipid were similar, indicating that the cholesteryl group of the neoglycolipid interacted in the same manner as cholesterol with hydrocarbon chains of EYPC. Some structural models of molecular interactions of neoglycolipids were discussed in relation with the molecular recognition of wheat germ agglutinin.

  1. Clonal redemption of autoantibodies by somatic hypermutation away from self-reactivity during human immunization.

    Science.gov (United States)

    Reed, Joanne H; Jackson, Jennifer; Christ, Daniel; Goodnow, Christopher C

    2016-06-27

    Clonal anergy is an enigmatic self-tolerance mechanism because no apparent purpose is served by retaining functionally silenced B cells bearing autoantibodies. Human autoantibodies with IGHV4-34*01 heavy chains bind to poly-N-acetyllactosamine carbohydrates (I/i antigen) on erythrocytes and B lymphocytes, cause cold agglutinin disease, and are carried by 5% of naive B cells that are anergic. We analyzed the specificity of three IGHV4-34*01 IgG antibodies isolated from healthy donors immunized against foreign rhesus D alloantigen or vaccinia virus. Each IgG was expressed and analyzed either in a hypermutated immune state or after reverting each antibody to its unmutated preimmune ancestor. In each case, the preimmune ancestor IgG bound intensely to normal human B cells bearing I/i antigen. Self-reactivity was removed by a single somatic mutation that paradoxically decreased binding to the foreign immunogen, whereas other mutations conferred increased foreign binding. These data demonstrate the existence of a mechanism for mutation away from self-reactivity in humans. Because 2.5% of switched memory B cells use IGHV4-34*01 and >43% of these have mutations that remove I/i binding, clonal redemption of anergic cells appears efficient during physiological human antibody responses. PMID:27298445

  2. Regulator of insulin receptor affinity in rat skeletal muscle sarcolemmal vesicles

    International Nuclear Information System (INIS)

    Wheat germ agglutinin (WGA) affinity purification of detergent solubilized insulin receptors (IR) from rat skeletal muscle sarcolemmal vesicles resulted in an apparent increase in total insulin binding activity of greater than 5-fold, suggesting that an inhibitory component had been removed. This was verified when the flow-through fraction from the WGA column was dialyzed and added back to the partially purified receptor. The addition of a 100-fold dilution of the inhibitor preparation caused a 50% reduction in binding to trace amounts of 125I-insulin. Scatchard analysis revealed that the effect of the inhibitor was to decrease the affinity of the muscle IR. The inhibitor appeared to be tissue specific, inasmuch as the I50's for WGA-purified IR from rat fat and liver were 10 times the I50 for muscle IR. The I50 for insulin binding to intact IM-9 cells was 30 times the value for muscle IR. The inhibitor eluted in the void volume of Sephadex G-50 columns. Its activity was not destroyed by heating at 900C for 10 minutes, or by prolonged incubation with trypsin or dithiothreitol. The inhibitor described here may have a role in modulating insulin sensitivity in skeletal muscle

  3. Study on proliferative responses to host Ia antigens in allogeneic bone marrow chimera in mice: sequential analysis of the reactivity and characterization of the cells involved in the responses

    International Nuclear Information System (INIS)

    Irradiation bone marrow chimeras were established by reconstitution of lethally irradiated AKR mice with C57BL/10 marrow cells to permit serial analysis of the developing reactivities of lymphocytes from such chimeras, [B10----AKR], against donor, host, or third party antigens. We found that substantial proliferative responses to Ia antigens of the recipient strain and also to third party antigens were generated by the thymocytes obtained from the irradiation chimeras at an early stage after bone marrow reconstitution. The majority of the responding thymocytes had surfaces lacking demonstrable peanut agglutinin receptors and were donor type Thy-1+, Ly-2-, and L3T4+ in both anti-recipient and anti-third party MLR. In anti-host responses, however, Ly-2+ thymocytes seemed to be at least partially involved. This capacity of thymus cells to mount a response to antigens of the recipient strain declined shortly thereafter, whereas the capacity to mount MLR against third party antigens persisted. The spleen cells of [B10----AKR] chimeras at the same time developed a more durable capability to exhibit anti-host reactivities and a permanent capability of reacting to third party allo-antigens. The stimulator antigens were Ia molecules on the stimulator cells in both anti-recipient and anti-third party MLR. The responding splenocytes were of donor origin and most of them had Thy-1+, Ly-1+2-, and L3T4+ phenotype

  4. 亲和层析微柱法测定肝癌特异性AFP及其在肝癌诊断中的临床价值%Quantitative analysis of hepatoma-specific α-fetoprotein (HS-AFP) by a new mini-column affinity chromatography and its clinical value in diagnosis of hepatocellular carcinoma

    Institute of Scientific and Technical Information of China (English)

    Wei Wu; Dengfu Yao; Liwei Qiu; Xiaoxiao Gu; Xinhua Wu

    2008-01-01

    Objective: To establish a convenient and economic method to determine hepatoma-specific α-fetoprotein (HS-AFP) for diagnosis of hepatocellular carcinoma (HCC). Methods: HS-AFP from serum of HCC patients was separated by a mini-column Lens culinaris agglutinin (LCA)-affinity chromatography. The levels of serum total AFP and separated HS-AFP were detected by radioimmunoassay (RIA). Results: Circulating AFP was separated into three peaks (AFP-1, AFP-2, and AFP-3) by LCA-affinity chromatography. During the elution course, the AFP-1 and AFP-2 could be eluted with TE buffer. HS-AFP (AFP-3) from sera of HCC patients was eluted dearly on the LCA-sepharose gel mini-column with a solution containing a-methyl-D-mannoside. It was a part of total AFP and only found in sera of HCC patients. A ratio of more than 15% for HS-AFP to total AFP in serum was considered as a specific marker for HCC diagnosis with higher sensitivity (92.7%) and specificity(88.2%). Conclusion: The new assay for circulating HS-AFP analysis is more sensitive, repeatable, and convenient. Its clinical application would be useful to early diagnosis of HCC.

  5. Lectin-induced activation of platelets may require only limited phosphorylation of the 47K protein

    International Nuclear Information System (INIS)

    Wheat germ agglutinin (WGA) is an N-acetylglucosamine (Glc-NAc) specific lectin which can activate platelets. Like thrombin, stimulation of platelets by WGA is accompanied by enhanced phosphorylation of two polypeptides of M/sub r/ 47K and 20K. Addition of GlcNAc at different time intervals arrested that aggregation of platelets by WGA and paralleled the modification of phosphorylation of the 47K polypeptide. So, the phosphorylation of the 47K polypeptide may regulate the WGA-receptor mediated stimulation of platelets. However, the ratio of phosphoserine to phosphothreonine in the 47K protein was markedly different in WGA-activated than thrombin-stimulated platelets. Thus, the molecular mechanism of action of thrombin and WGA could be different. To explore this idea, 32P/sub i/-labeled platelets were stimulated with WGA and the activation arrested with N-acetyl-glucosamine at different times. Two-dimensional gel electrophoresis of total protein at 5s showed only two phosphorylated species of 47K protein. At 60s, maximally four phosphorylated species were noted. In contrast, with thrombin using the same technique, seven to nine phosphorylated components have been reported. These results suggest that the different activators of platelets may act by different mechanisms. In addition, activation of platelets may require only limited levels of phosphorylation of the 47K polypeptide

  6. Differences in the sites of phosphorylation of the insulin receptor in vivo and in vitro

    International Nuclear Information System (INIS)

    Phosphorylation of the insulin receptor was studied in intact well differentiated hepatoma cells (Fao) and in a solubilized and partially purified receptor preparation obtained from these cells by affinity chromatography on wheat germ agglutinin agarose. Tryptic peptides containing the phosphorylation sites of the beta-subunit of the insulin receptor were analyzed by reverse-phase high performance liquid chromatography. Phosphoamino acid content of these peptides was determined by acid hydrolysis and high voltage electrophoresis. Separation of the phosphopeptides from unstimulated Fao cells revealed one major and two minor phosphoserine-containing peptides and a single minor phosphothreonine-containing peptide. Insulin (10(-7) M) increased the phosphorylation of the beta-subunit of the insulin receptor 3- to 4-fold in the intact Fao cell. After insulin stimulation, two phosphotyrosine-containing peptides were identified. Tyrosine phosphorylation reached a steady state within 20 s after the addition of insulin and remained nearly constant for 1 h. Under our experimental conditions, no significant change in the amount of [32P]phosphoserine or [32P]phosphothreonine associated with the beta-subunit was found during the initial response of cells to insulin. When the insulin receptor was extracted from the Fao cells and incubated in vitro with [gamma-32P]ATP and Mn2+, very little phosphorylation occurred in the absence of insulin

  7. Differences in the sites of phosphorylation of the insulin receptor in vivo and in vitro

    Energy Technology Data Exchange (ETDEWEB)

    White, M.F.; Takayama, S.; Kahn, C.R.

    1985-08-05

    Phosphorylation of the insulin receptor was studied in intact well differentiated hepatoma cells (Fao) and in a solubilized and partially purified receptor preparation obtained from these cells by affinity chromatography on wheat germ agglutinin agarose. Tryptic peptides containing the phosphorylation sites of the beta-subunit of the insulin receptor were analyzed by reverse-phase high performance liquid chromatography. Phosphoamino acid content of these peptides was determined by acid hydrolysis and high voltage electrophoresis. Separation of the phosphopeptides from unstimulated Fao cells revealed one major and two minor phosphoserine-containing peptides and a single minor phosphothreonine-containing peptide. Insulin (10(-7) M) increased the phosphorylation of the beta-subunit of the insulin receptor 3- to 4-fold in the intact Fao cell. After insulin stimulation, two phosphotyrosine-containing peptides were identified. Tyrosine phosphorylation reached a steady state within 20 s after the addition of insulin and remained nearly constant for 1 h. Under our experimental conditions, no significant change in the amount of (TSP)phosphoserine or (TSP)phosphothreonine associated with the beta-subunit was found during the initial response of cells to insulin. When the insulin receptor was extracted from the Fao cells and incubated in vitro with (gamma-TSP)ATP and MnS , very little phosphorylation occurred in the absence of insulin.

  8. Synthesis and characterization of near IR fluorescent albumin nanoparticles for optical detection of colon cancer

    Energy Technology Data Exchange (ETDEWEB)

    Cohen, Sarit; Pellach, Michal [Department of Chemistry, Bar-Ilan Institute of Nanotechnology and Advanced Materials, Bar-Ilan University, Ramat-Gan 52900 (Israel); Kam, Yossi [Institute for Drug Research, School of Pharmacy, Faculty of Medicine, The Hebrew University of Jerusalem, P.O. Box 12065, Jerusalem 91120 (Israel); Grinberg, Igor; Corem-Salkmon, Enav [Department of Chemistry, Bar-Ilan Institute of Nanotechnology and Advanced Materials, Bar-Ilan University, Ramat-Gan 52900 (Israel); Rubinstein, Abraham [Institute for Drug Research, School of Pharmacy, Faculty of Medicine, The Hebrew University of Jerusalem, P.O. Box 12065, Jerusalem 91120 (Israel); Margel, Shlomo, E-mail: shlomo.margel@mail.biu.ac.il [Department of Chemistry, Bar-Ilan Institute of Nanotechnology and Advanced Materials, Bar-Ilan University, Ramat-Gan 52900 (Israel)

    2013-03-01

    Near IR (NIR) fluorescent human serum albumin (HSA) nanoparticles hold great promise as contrast agents for tumor diagnosis. HSA nanoparticles are considered to be biocompatible, non-toxic and non-immunogenic. In addition, NIR fluorescence properties of these nanoparticles are important for in vivo tumor diagnostics, with low autofluorescence and relatively deep penetration of NIR irradiation due to low absorption of biomatrices. The present study describes the synthesis of new NIR fluorescent HSA nanoparticles, by entrapment of a NIR fluorescent dye within the HSA nanoparticles, which also significantly increases the photostability of the dye. Tumor-targeting ligands such as peanut agglutinin (PNA) and anti-carcinoembryonic antigen antibodies (anti-CEA) were covalently conjugated to the NIR fluorescent albumin nanoparticles, increasing the potential fluorescent signal in tumors with upregulated corresponding receptors. Specific colon tumor detection by the NIR fluorescent HSA nanoparticles was demonstrated in a chicken embryo model and a rat model. In future work we also plan to encapsulate cancer drugs such as doxorubicin within the NIR fluorescent HSA nanoparticles for both colon cancer imaging and therapy. - Highlights: Black-Right-Pointing-Pointer Near IR human serum albumin nanoparticles were synthesized and characterized. Black-Right-Pointing-Pointer Nanoparticles were shown to be physically and chemically stable and photostable. Black-Right-Pointing-Pointer Tumor-targeting ligands were covalently conjugated to the nanoparticles. Black-Right-Pointing-Pointer Specific colon cancer tumor detection was demonstrated in chicken-embryo and rat models.

  9. Sialomucin and lytic susceptibility of rat mammary tumor ascites cells.

    Science.gov (United States)

    Moriarty, J; Skelly, C M; Bharathan, S; Moody, C E; Sherblom, A P

    1990-11-01

    The potential role of cell surface sialomucin in preventing natural killer (NK)-mediated lysis of tumor cell targets has been addressed by comparing the properties of 2 NK-resistant [ascites (ASC) and short-term cultured (STC)] and 2 NK-susceptible [tunicamycin-treated (TUN) and long-term cultured (LTC)] preparations of 13762 MAT-B1 rat mammary tumor cells. Both the ASC and STC cell preparations contain elevated levels of the sialomucin ASGP-1 relative to TUN and LTC preparations as determined by [3H]glucosamine labeling and by binding of peanut agglutinin. The major difference in the susceptibility to NK-mediated lysis appeared to be due to the differences in the susceptibility to lysis by lytic granules, rather than to differences in the ability to bind or trigger effector cells, since TUN and LTC cells were approximately 10-fold more sensitive to lysis by lytic granules than were ASC and STC cells. All preparations inhibited the lysis of the susceptible target YAC-1 by normal rat splenocytes, indicating an ability to bind these effector cells. Triggering of effectors, as monitored either by incorporation of 32P into phosphatidylinositol or by transmethylation of phosphatidylcholine, was similar for the positive control YAC-1, STC, TUN, and LTC, whereas ASC appeared to be defective in triggering effectors. These results suggest that tumor sialomucin blocks the final phase of lysis, but not the initial recognition of tumor cells by NK effectors. PMID:2208144

  10. Recommended Immunological Assays to Screen for Ricin-Containing Samples

    Directory of Open Access Journals (Sweden)

    Stéphanie Simon

    2015-11-01

    Full Text Available Ricin, a toxin from the plant Ricinus communis, is one of the most toxic biological agents known. Due to its availability, toxicity, ease of production and absence of curative treatments, ricin has been classified by the Centers for Disease Control and Prevention (CDC as category B biological weapon and it is scheduled as a List 1 compound in the Chemical Weapons Convention. An international proficiency test (PT was conducted to evaluate detection and quantification capabilities of 17 expert laboratories. In this exercise one goal was to analyse the laboratories’ capacity to detect and differentiate ricin and the less toxic, but highly homologuous protein R. communis agglutinin (RCA120. Six analytical strategies are presented in this paper based on immunological assays (four immunoenzymatic assays and two immunochromatographic tests. Using these immunological methods “dangerous” samples containing ricin and/or RCA120 were successfully identified. Based on different antibodies used the detection and quantification of ricin and RCA120 was successful. The ricin PT highlighted the performance of different immunological approaches that are exemplarily recommended for highly sensitive and precise quantification of ricin.

  11. Ricinus communis Intoxications in Human and Veterinary Medicine—A Summary of Real Cases

    Directory of Open Access Journals (Sweden)

    Martin Schaer

    2011-10-01

    Full Text Available Accidental and intended Ricinus communis intoxications in humans and animals have been known for centuries but the causative agent remained elusive until 1888 when Stillmark attributed the toxicity to the lectin ricin. Ricinus communis is grown worldwide on an industrial scale for the production of castor oil. As by-product in castor oil production ricin is mass produced above 1 million tons per year. On the basis of its availability, toxicity, ease of preparation and the current lack of medical countermeasures, ricin has gained attention as potential biological warfare agent. The seeds also contain the less toxic, but highly homologous Ricinus communis agglutinin and the alkaloid ricinine, and especially the latter can be used to track intoxications. After oil extraction and detoxification, the defatted press cake is used as organic fertilizer and as low-value feed. In this context there have been sporadic reports from different countries describing animal intoxications after uptake of obviously insufficiently detoxified fertilizer. Observations in Germany over several years, however, have led us to speculate that the detoxification process is not always performed thoroughly and controlled, calling for international regulations which clearly state a ricin threshold in fertilizer. In this review we summarize knowledge on intended and unintended poisoning with ricin or castor seeds both in humans and animals, with a particular emphasis on intoxications due to improperly detoxified castor bean meal and forensic analysis.

  12. Surface binding properties of aged and fresh (recently excreted) Toxoplasma gondii oocysts.

    Science.gov (United States)

    Harito, Jemere Bekele; Campbell, Andrew T; Prestrud, Kristin W; Dubey, J P; Robertson, Lucy J

    2016-06-01

    The surfaces of aged (10 years) and fresh (recently excreted) oocysts of Toxoplasma gondii were investigated using monoclonal antibody (mAb) and lectin-binding assays. Fresh oocysts bound a wall-specific mAb labelled with fluorescein isothiocyanate while aged oocysts did not. In contrast, the walls of aged oocysts bound a lectin (wheat germ agglutinin, WGA), but not the walls of fresh oocysts. Exposure of oocysts to detergent solutions or trypsin did not affect the binding properties of the walls of the oocysts. However, exposure of fresh oocysts to acidified pepsin enabled labelling of the walls with WGA, presumably due to the relevant moieties on the oocyst walls becoming exposed. WGA binding, but not mAb binding, was partially abrogated with periodate exposure. These findings reveal a significant difference in the binding properties of oocyst walls from "aged" and "fresh" oocysts. The results are of relevance when considering technologies for isolating or detecting T. gondii oocysts in environmental samples based on oocyst surface properties, as used for other protozoan parasites. Our results suggest the possibility of developing a WGA-based separation procedure for isolating Toxoplasma oocysts from environmental matrices, in which pepsin pre-treatment would be included to ensure that both fresh and aged oocysts were isolated. PMID:27003461

  13. Cross neutralizing antibodies in hamsters vaccinated with leptospiral bacterins produced with three serovars of serogroup Sejroe

    Directory of Open Access Journals (Sweden)

    Rosana Tabata

    2002-09-01

    Full Text Available Three leptospiral bacterins, produced with different serovars of Serogroup Sejroe, namely the hardjo (bacterin A, wolffi (bacterin B and guaricura (bacterin C, were evaluated in male hamsters (Mesocricetus auratus by comparing the agglutinating and neutralizing antibodies titers using microscopic agglutination (MAT and in vitro growth inhibition (GIT tests. The immunization schedule was based on two 1.0 mL doses of non-diluted formalininactivated whole culture bacterin given through subcutaneous route with 10-day interval. The challenge was performed ten days after the second vaccine dose, when the animals were inoculated with 0.2 mL of non-inactivated cultures of each serovar through intraperitoneal route. On the 21st post-challenge day (PCD, all animals were bled and their sera were joined in pools (n=8 and tested by MAT and GIT. All vaccinated and control animals presented no clinical signs of leptospirosis after the challenge, but the serovar guaricura was isolated from the kidneys of control animals on the 21st PCD. The MAT results showed cross agglutinins between serovars hardjo and wolffi, and between wolffi and guaricura. The GIT results revealed the presence of cross neutralizing antibodies between serovars wolffi or guaricura against hardjo, wolffi and guaricura. It was found that the tested strain of serovar hardjo did not produce detectable levels of neutralizing antibodies, indicating its poor immunogenicity.

  14. True blue: S-opsin is widely expressed in different animal species.

    Science.gov (United States)

    Amann, B; Hirmer, S; Hauck, S M; Kremmer, E; Ueffing, M; Deeg, C A

    2014-02-01

    Colour vision in animals is an interesting, fascinating subject. In this study, we examined a wide variety of species for expression of S-opsin (blue sensitive) and M-/L-opsin (green-red sensitive) in retinal cones using two novel monoclonal antibodies specific for peptides from human opsins. Mouse, rat and hare did not express one of the investigated epitopes, but we could clearly prove existence of cones through peanut agglutinin labelling. Retinas of guinea pig, dog, wolf, marten, cat, roe deer, pig and horse were positive for S-opsin, but not for M-/L-opsin. Nevertheless all these species are clearly at least dichromats, because we could detect further S-opsin negative cones by labelling with cone arrestin specific antibody. In contrast, pheasant and char had M-/L-opsin positive cones, but no S-opsin expressing cones. Sheep, cattle, monkey, men, pigeon, duck and chicken were positive for both opsins. Visual acuity analyzed through density of retinal ganglion cells revealed least visual discrimination by horses and highest resolution in pheasant and pigeon. Most mammals studied are dichromats with visual perception similar to red-green blind people. PMID:23173557

  15. Pediatric Typhoid Fever: Evaluation of 30 Cases

    Directory of Open Access Journals (Sweden)

    Mahmut Abuhandan

    2012-04-01

    Full Text Available Objective: This study aimed to evaluate pediatric patients with typhoid fever with respect to their epidemiological, clinical and laboratory findings and response to treatment in our region, where typhoid fever is endemic.Methods: A retrospective evaluation was performed in 30 pediatric patients diagnosed as typhoid fever between January 2011 and December 2011 in terms of age, gender, complaints on presentation, physical examination and laboratory findings, and the therapeutics selected. Diagnosis was confirmed by Gruber-Widal agglutination test and/or growth of the causative agent in culture.Results: The patients comprised of 15 males and 15 females with a mean age of 10.03±5.08 years. The most common presenting symptoms were fever (n=28, fatigue (n=22, headache (n=19, sweating (n=17, abdominal pain (n=16, diarrhea (n=15, vomiting (n=11 and arthralgia (n=8. Physical examination revealed fever (n=27, hepatomegaly (n=12, splenomegaly (n=10, and rose spots (n=1. Titers of Salmonella typhi O agglutinins were ≥1/160 in all patients. Blood cultures were positive for S. typhi in three patients.Conclusions: There may be many different clinical signs of typhoid fever. In areas where the disease is endemic, it should be considered primarily, especially in patients presenting with fever, abdominal pain, and diarrhea.

  16. Serum ferritin and #betta#2-microglobulin in patients with multiple myeloma

    International Nuclear Information System (INIS)

    In 76 patients with clinically well defined multiple myeloma (median age at diagnosis: 68.5 years) serum-ferritin (SF) and #betta#2-microglobulin (#betta#2M) were measured by RIA-methods. 70 sex and age-matched healthy individuals served as controls. Both serum-ferritin (median: 343 μg/l vs. 193 μg/l; p -7) and #betta#2M (median: 4.25 mg/l vs. 3.5 mg/l; p 2M and between #betta#2M and tumour mass, percentage of plasma cell infiltration in bone marrow, agglutinine titer, serum-creatinine, hemoglobin and age of the patients. Both tumour proteins might gain clinical importance particularly in those patients in which precise monitoring of disease is impossible either due to lack of paraprotein production or due to the particular paraprotein type. This seems to account for patients with light cahin paraproteins, and furthermore for those patients with biclonal gammopathies or with IgE- and/or IgD-paraproteins. (Author)

  17. Location and identification of colloidal gold particles on the cell surface with a scanning electron microscope and energy dispersive analyzer

    International Nuclear Information System (INIS)

    The use of colloidal gold particles for locating cell surface components by scanning electron microscopy (SEM) has been restricted due to difficulties in the identification of these gold particles under SEM. It is shown here how the gold particles bound to cell surfaces can be located and identified under SEM using the secondary electron imaging (SEI) mode with an energy dispersive X-ray microanalyzer (EDS). This enables reliable identification of gold particles and good quality micrographs of the cells to be achieved at the same time. The distribution of receptors for two lectins, concanavalin A (ConA) and wheat germ agglutinin (WGA), on the surface of cultured Raji cells and human erythrocytes is presented as an example. Raji cells and erythrocytes were fixed with glutaraldehyde, post-fixed with a glutaraldehyde-tannic acid mixture and then incubated with ConA- or WGA-coated gold particles. After dehydration and critical point drying, the specimen filters were mounted on copper stubs and coated with carbon. The cells were examined on a JEOL TEMSCAN 100CX II electron microscope. The gold particles could be identified with the EDS analyzer, which was able to detect the Au spectrum when the electron beam was focused on single gold particles using a magnification of 100,000 or more. High-resolution photographs of the same cells were obtained up to the same magnification of 100,000

  18. Investigation of the response to the enterobacterial common antigen after typhoid vaccination

    Directory of Open Access Journals (Sweden)

    Arlete M. Milhomem

    1987-03-01

    Full Text Available Antibodies against the Salmonella typhi enterobacterial common antigen (ECA and the O and H antigens were investigated in sera from healthy male subjects who had been previously vaccinated with the typhoid vaccine. No serological response to ECA was observed. Sera from subjects not previously vaccinated presented titers of ECA hemagglutinins which quantitatively were related to the presence ofH titers, but not to O agglutinins but with no statistical significance. The results are discussed in relation to the possible protective immunological mechanisms in typhoid fever.Anticorpos contra o antígeno comum de enterobactérias (ECA bem como contra os antigenos somáticos (O e flagelar (H de Salmonella typhi foram investigados no soro de recrutas do sexo masculino, após a vacinação. Não fo i detectada resposta humoral para ECA. Os soros obtidos antes da vacinação mostraram hemaglutininas para ECA acompanhando a presença de aglutininas para o antígeno H, ao contrário do que se observou em relação ao antígeno O. Discutem-se os resultados quanto ao possível mecanismo da imunoproteção da febre tifóide.

  19. Importin-11, a nuclear import receptor for the ubiquitin-conjugating enzyme, UbcM2.

    Science.gov (United States)

    Plafker, S M; Macara, I G

    2000-10-16

    Importins are members of a family of transport receptors (karyopherins) that mediate the nucleocytoplasmic transport of protein and RNA cargoes. We identified importin-11 as a potential new human member of this family, on the basis of limited similarity to the Saccharomyces cerevisiae protein, Lph2p, and cloned the complete open reading frame. Importin-11 interacts with the Ran GTPase, and constitutively shuttles between the nuclear and cytoplasmic compartments. A yeast dihybrid screen identified UbcM2, an E2-type ubiquitin-conjugating enzyme, as a binding partner and potential transport cargo for importin-11. Importin-11 and UbcM2 interact directly, and the complex is disassembled by Ran:GTP but not by Ran:GDP. UbcM2 is constitutively nuclear and shuttles between the nuclear and cytoplasmic compartments. Nuclear import of UbcM2 requires Ran and importin-11, and is inhibited by wheatgerm agglutinin, energy depletion or dominant interfering mutants of Ran and importin-beta. These data establish importin-11 as a new member of the karyopherin family of transport receptors, and identify UbcM2 as a nuclear member of the E2 ubiquitin-conjugating enzyme family. PMID:11032817

  20. Avaliação sorológica de vacinações preventivas da difteria, do tétano e da coqueluche, efetuadas em crianças prematuras

    Directory of Open Access Journals (Sweden)

    Vicente Amato Neto

    1977-08-01

    Full Text Available Vinte crianças prematuras receberam, no primeiro ano de vida. vacinas que habitualmente fazem parte do esquema básico de imunizações ativas. Em amostra de soro obtida quando elas atingiram a idade de 12 meses, foram dosados os teores de antitoxina diftérica, de antitoxina tetânica e de aglutininas anti Bordetella pertussis. Valores plenamente satisfatórios de anticorpos relativos à difteria e ao tétano puderam ser encontrados e, quanto à coqueluche, nunca notaram os Autores ausência de aglutininas, mas conclusão mais decisiva não ocorreu, em virtude da falta de melhor conhecimento da cifra indicativa de proteção. O estudd em questão representa subsídio no sentido de arrefecer o temor e o cepticismo, bastante divulgados, acerca da vacinação de prematuros.Twenty premature-born children received, during their first year of life, vaccines routinely apptied as part of a basic immunization schedule. Sera obtained at the age of 12 months were titered for antibodies against diphteria, tetanus and pertussis. Values considered protective were observed for diphteria and tetanus. Anti - Bordetella pertussis agglutinins were always present, however, in the absence of a consensus as to what are protective levels, no conclusion could be drawn. The present study contributes towards erasing the prejudice and scepticism concerning the immunization of the premature-born.

  1. Biomolecular interaction analysis for carbon nanotubes and for biocompatibility prediction.

    Science.gov (United States)

    Chen, Xiaoping; Fang, Jinzhang; Cheng, Yun; Zheng, Jianhui; Zhang, Jingjing; Chen, Tao; Ruan, Benfang Helen

    2016-07-15

    The interactions between carbon nanotubes (CNTs) and biologics have been commonly studied by various microscopy and spectroscopy methods. We tried biomolecular interaction analysis to measure the kinetic interactions between proteins and CNTs. The analysis demonstrated that wheat germ agglutinin (WGA) and other proteins have high affinity toward carboxylated CNT (f-MWCNT) but essentially no binding to normal CNT (p-MWCNT). The binding of f-MWCNT-protein showed dose dependence, and the observed kinetic constants were in the range of 10(-9) to 10(-11) M with very small off-rates (10(-3) to 10(-7) s(-1)), indicating a relatively tight and stable f-MWCNT-protein complex formation. Interestingly in hemolysis assay, p-MWCNT showed good biocompatibility, f-MWCNT caused 30% hemolysis, but WGA-coated f-MWCNT did not show hemolysis. Furthermore, the f-MWCNT-WGA complex demonstrated enhanced cytotoxicity toward cancer cells, perhaps through the glycoproteins expressed on the cells' surface. Taken together, biomolecular interaction analysis is a precise method that might be useful in evaluating the binding affinity of biologics to CNTs and in predicting biological actions. PMID:27108187

  2. Identification and Characterization of TEX101 in Bovine Epididymal Spermatozoa

    Directory of Open Access Journals (Sweden)

    Subir K. Nagdas

    2014-01-01

    Full Text Available Several studies exhibit the presence of Ricinus Communis Agglutinin I (RCA binding glycocalyx in mammalian spermatozoa. However, the molecular characterization of RCA binding glycocalyx in sperm membranes and its mechanism of action are poorly understood. The objective of the study was to identify and to characterize RCA binding glycoprotein of the bovine sperm plasma membranes (PM. Lectin blots of caput and cauda sperm PM revealed a 38 kDa polypeptide exhibiting the highest affinity to RCA among the several major RCA binding polypeptides. The 38 kDa RCA binding polypeptide of cauda sperm PM was purified and exhibited a charge train of three distinct spots with isoelectric points (pH 5.3 and 5.8. Proteomic identification yielded ten peptides that matched the sequence of Testis Expressed 101 protein (TEX101. Western blots data revealed that bovine sperm TEX101 is present in both testicular and epididymal sperm PM fractions. The native TEX101 polypeptide contains ~17 kDa N-linked oligosaccharides and the polypeptide is anchored to sperm membrane via a glycosylphosphatidylinositol lipid linkage. Immunofluorescence staining of sperm with anti-TEX101 demonstrated that the polypeptide is localized at the head of cauda sperm. Our biochemical results provide evidence on the presence of TEX101 in bovine epididymal sperm plasma membranes and may have a potential role in sperm-egg interaction.

  3. Molecular Dynamics and Monte Carlo simulations resolve apparent diffusion rate differences for proteins confined in nanochannels

    International Nuclear Information System (INIS)

    Highlights: • WGA proteins in nanochannels modeled by Molecular Dynamics and Monte Carlo. • Protein surface coverage characterized by atomic force microscopy. • Models indicate transport characteristics depend strongly on surface coverage. • Results resolve of a four orders of magnitude difference in diffusion coefficient values. - Abstract: We use Molecular Dynamics and Monte Carlo simulations to examine molecular transport phenomena in nanochannels, explaining four orders of magnitude difference in wheat germ agglutinin (WGA) protein diffusion rates observed by fluorescence correlation spectroscopy (FCS) and by direct imaging of fluorescently-labeled proteins. We first use the ESPResSo Molecular Dynamics code to estimate the surface transport distance for neutral and charged proteins. We then employ a Monte Carlo model to calculate the paths of protein molecules on surfaces and in the bulk liquid transport medium. Our results show that the transport characteristics depend strongly on the degree of molecular surface coverage. Atomic force microscope characterization of surfaces exposed to WGA proteins for 1000 s show large protein aggregates consistent with the predicted coverage. These calculations and experiments provide useful insight into the details of molecular motion in confined geometries

  4. Gram-typing of mastitis bacteria in milk samples using flow cytometry

    DEFF Research Database (Denmark)

    Langerhuus, Sine Nygaard; Ingvartsen, Klaus Lønne; Bennedsgaard, Torben Werner;

    2013-01-01

    Fast identification of pathogenic bacteria in milk samples from cows with clinical mastitis is central to proper treatment. In Denmark, time to bacterial diagnosis is typically 24 to 48 h when using traditional culturing methods. The PCR technique provides a faster and highly sensitive identifica......Fast identification of pathogenic bacteria in milk samples from cows with clinical mastitis is central to proper treatment. In Denmark, time to bacterial diagnosis is typically 24 to 48 h when using traditional culturing methods. The PCR technique provides a faster and highly sensitive...... cytometry-based method, which can detect and distinguish gram-negative and gram-positive bacteria in mastitis milk samples. The differentiation was based on bacterial fluorescence intensities upon labeling with biotin-conjugated wheat germ agglutinin and acridine orange. Initially 19 in-house bacterial...... characteristic curves for the 19 bacterial cultures. The method was then tested on 53 selected mastitis cases obtained from the department biobank (milk samples from 6 gram-negative and 47 gram-positive mastitis cases). Gram-negative bacteria in milk samples were detected with a sensitivity of 1 and a...

  5. Isolation and characterization of a new mannose-binding lectin gene from Taxus media

    Indian Academy of Sciences (India)

    Guoyin Kai; Lingxia Zhao; Jingui Zheng; Lei Zhang; Zhiqi Miao; Xiaofen Sun; Kexuan Tanga

    2004-12-01

    In this paper, we report the cloning and characterization of the first mannose-binding lectin gene from a gymnosperm plant species, Taxus media. The full-length cDNA of T. media agglutinin (TMA) consisted of 676 bp and contained a 432 bp open reading frame (ORF) encoding a 144 amino acid protein. Comparative analysis showed that TMA had high homology with many previously reported plant mannose-binding lectins and that tma encoded a precursor lectin with a 26-aa signal peptide. Molecular modelling revealed that TMA was a new mannose-binding lectin with three typical mannose-binding boxes like lectins from species of angiosperms. Tissue expression pattern analyses revealed that tma is expressed in a tissue-specific manner in leaves and stems, but not in fruits and roots. Phylogenetic tree analyses showed that TMA belonged to the structurally and evolutionarily closely related monocot mannose-binding lectin superfamily. This study provides useful information to understand the molecular evolution of plant lectins.

  6. Construction of the yeast whole-cell Rhizopus oryzae lipase biocatalyst with high activity

    Institute of Scientific and Technical Information of China (English)

    Mei-ling CHEN; Qin GUO; Rui-zhi WANG; Juan XU; Chen-wei ZHOU; Hui RUAN; Guo-qing HE

    2011-01-01

    Surface display is effectively utilized to construct a whole-cell biocatalyst.Codon optimization has been proven to be effective in maximizing production of heterologous proteins in yeast.Here,the cDNA sequence of Rhizopus oryzae lipase (ROL) was optimized and synthesized according to the codon bias of Saccharomyces cerevisiae,and based on the Saccharomyces cerevisiae cell surface display system with α-agglutinin as an anchor,recombinant yeast displaying fully codon-optimized ROL with high activity was successfully constructed.Compared with the wild-type ROL-displaying yeast,the activity of the codon-optimized ROL yeast whole-cell biocatalyst (25 U/g dried cells) was 12.8-fold higher in a hydrolysis reaction using p-nitrophenyl palmitate (pNPP) as the substrate.To our knowledge,this was the first attempt to combine the techniques of yeast surface display and codon optimization for whole-cell biocatalyst construction.Consequently,the yeast whole-cell ROL biocatalyst was constructed with high activity.The optimum pH and temperature for the yeast whole-cell ROL biocatalyst were pH 7.0 and 40 ℃.Furthermore,this whole-cell biocatalyst was applied to the hydrolysis of tributyrin and the resulted conversion of butyric acid reached 96.91% after 144 h.

  7. 检测胆汁糖蛋白糖链结构变化鉴别良恶性胆道疾病分析%Detection and analysis of biliary glycoprotein glycan structure changes in differential diagnosis of benign and malignant biliary disease

    Institute of Scientific and Technical Information of China (English)

    候鹏; 高峰; 马树民

    2014-01-01

    目的:探讨胆汁糖蛋白糖链结构变化对于鉴别良恶性胆道疾病的作用。方法选取青岛市海慈医疗集团普外科收治的胆道疾病患者100例,按照良、恶性将其分为对照组(良性胆道疾病)和观察组(胆管癌),每组各50例。取两组患者胆汁滴于硝酸纤维膜上,通过比较麦胚凝集素( WGA )、欧曼陀罗凝集素( DSA)、小扁豆凝集素( LCA)、刀豆凝集素( CONA)试验阳性率,探讨胆汁糖蛋白糖链结构变化与良恶性胆道疾病的关系。结果对照组 WGA、DSA、LCA、CONA 凝集素试验阳性率分别为22.0%、14.0%、2.0%、76.0%,观察组分别为76.0%、66.0%、76.0%、82.0%。两组CONA凝集素试验阳性率差异无统计学意义(P>0.05),观察组WGA、LCA、DSA凝集试验阳性率均明显高于对照组,差异均有统计学意义(χ2=29.17、28.17、57.55,均P<0.05)。结论胆汁糖蛋白糖链结构变化与胆道疾病良、恶性密切相关,可以通过WGA、LCA、DSA凝集试验阳性率判断胆管疾病良、恶性,值得在临床上广泛推广。%Objective To investigate the effects of changes in protein bile sugar chain structure for differen-tiating benign and malignant biliary tract disease .Methods 100 patients with biliary tract diseases who were treated in Department of General Surgery ,Qingdao Haici Medical Group were selected in this study .According to benign or malignant biliary disease ,the patients were divided into control group ( benign biliary tract disease ) and observation group ( cholangiocarcinoma ) ,50 cases in each group .Two groups of patients with bile drops on the nitrocellulose mem-brane,through the comparison of wheat germ agglutinin (WGA),Datura stramonium agglutinin (DSA),lentil lectin (LCA),concanavalin A(CONA) positive rate of test,to explore the relationship between sugar chain of glycoprotein in bile and bile duct benign and

  8. A preliminary neuropathological study of Japanese encephalitis in humans and a mouse model.

    Science.gov (United States)

    German, Allison C; Myint, Khin Saw Aye; Mai, Nguyen Thi Hoang; Pomeroy, Ian; Phu, Nguyen Hoan; Tzartos, John; Winter, Peter; Collett, Jennifer; Farrar, Jeremy; Barrett, Alan; Kipar, Anja; Esiri, Margaret M; Solomon, Tom

    2006-12-01

    Japanese encephalitis virus is a mosquito-borne flavivirus that causes approximately 10000 deaths annually in Asia. After a brief viraemia, the virus enters the central nervous system, but the means of crossing the blood-brain barrier is uncertain. We used routine histological staining, immunohistology and electron microscopy to examine brain material from four fatal human cases, and made comparisons with material from a mouse model. In human material there was oedema, perivascular inflammation, haemorrhage, microglial nodules and acellular necrotic foci, as has been described previously. In addition, there was new evidence suggestive of viral replication in the vascular endothelium, with endothelial cell damage; this included occasional viral antigen staining, uneven binding of the vascular endothelial cells to Ulex europaeus agglutinin I and ultrastructural changes. Viral antigen was also found in neurons. There was an active astrocytic response, as shown by glial fibrillary acidic protein staining, and activation of microglial cells was demonstrated by an increase in major histocompatibility complex class II expression. Similar inflammatory infiltrates and a microglial reaction were observed in mouse brain tissue. In addition, beta-amyloid precursor protein staining indicated impaired axonal transport. Whether these findings are caused by viral replication in the vascular endothelium or the immune response merits further investigation. PMID:16814333

  9. Chemical Characterization of N-Linked Oligosaccharide As the Antigen Epitope Recognized by an Anti-Sperm Auto-Monoclonal Antibody, Ts4.

    Directory of Open Access Journals (Sweden)

    Hiroshi Yoshitake

    Full Text Available Ts4, an anti-sperm auto-monoclonal antibody, possesses immunoreactivity to the acrosomal region of mouse epididymal spermatozoa. In addition, the mAb shows specific immunoreactivity to reproduction-related regions such as testicular germ cells and early embryo. Our qualitative study previously showed that the antigen epitope for Ts4 contained a N-linked common oligosaccharide (OS chain on testicular glycoproteins as determined by Western blotting for testicular glycoproteins after treatment with several glycohydrolases. Since the distribution of the Ts4-epitope is unique, the OS chain in Ts4-epitope may have role(s in the reproductive process. The aim of this study was to clarify the molecular structure of the Ts4-epitope, particularly its OS moiety. Using Ts4 immunoprecipitation combined with liquid chromatography and multiple-stage mass spectrometry, the candidate carbohydrate structure in the Ts4-epitope is proposed to be N-linked fucosylated agalacto-biantennary with bisecting N-acetylglucosamine (GlcNAc or with N-acetylgalactosamine-GlcNAc motif. Further binding analyses using various lectins against the mouse testicular Ts4-immunoprecipitants revealed that Phaseolus vulgaris erythroagglutinin and Pisum sativum agglutinin showed positive staining of the bands corresponding to Ts4 reactive proteins. Moreover, the immunoreactivity of Ts4 against the testicular extract was completely abrogated after digestion with β-N-acetylglucosaminidase. These results show that the Ts4-epitope contains agalacto-biantennary N-glycan with bisecting GlcNAc carrying fucose residues.

  10. Glycoprotein expression by adenomatous polyps of the colon

    Science.gov (United States)

    Roney, Celeste A.; Xie, Jianwu; Xu, Biying; Jabour, Paul; Griffiths, Gary; Summers, Ronald M.

    2008-03-01

    Colon cancer is the second leading cause of cancer related deaths in the United States. Specificity in diagnostic imaging for detecting colorectal adenomas, which have a propensity towards malignancy, is desired. Adenomatous polyp specimens of the colon were obtained from the mouse model of colorectal cancer called adenomatous polyposis coli-multiple intestinal neoplasia (APC Min). Histological evaluation, by the legume protein Ulex europaeus agglutinin I (UEA-1), determined expression of the glycoprotein α-L-fucose. FITC-labelled UEA-1 confirmed overexpression of the glycoprotein by the polyps on fluorescence microscopy in 17/17 cases, of which 13/17 included paraffin-fixed mouse polyp specimens. In addition, FITC-UEA-1 ex vivo multispectral optical imaging of 4/17 colonic specimens displayed over-expression of the glycoprotein by the polyps, as compared to non-neoplastic mucosa. Here, we report the surface expression of α-L-fucosyl terminal residues by neoplastic mucosal cells of APC specimens of the mouse. Glycoprotein expression was validated by the carbohydrate binding protein UEA-1. Future applications of this method are the development of agents used to diagnose cancers by biomedical imaging modalities, including computed tomographic colonography (CTC). UEA-1 targeting to colonic adenomas may provide a new avenue for the diagnosis of colorectal carcinoma by CT imaging.

  11. Common antigens of streptococcal and non-streptococcal oral bacteria: immunochemical studies of extracellular and cell-wall-associated antigens from Streptococcus sanguis, Streptococcus mutans, Lactobacillus salivarius, and Actinomyces viscosus.

    Science.gov (United States)

    Schöller, M; Klein, J P; Frank, R M

    1981-01-01

    Soluble extracellular antigens (ESA) were prepared from the culture supernatant of exponential growing cells of Streptococcus sanguis OMZ 9 by a combination of ammonium sulfate precipitation and chromatography on a Bio-Gel P6 column. Soluble cell wall antigens (WEA) were obtained from the bacterial pellet by extraction with 1 M phosphate buffer (pH 6). Antisera against whole cells of S. sanguis and S. mutans of different serotypes, 10% trichloroacetic extracts of bacterial cell walls, dextran, ESA, and WEA were prepared by injecting the different antigens several times in rabbits. ESA and WEA were prepared from a representative strain of Bratthall's seven serological groups, Lactobacillus salivarius, and Actinomyces viscosus. All sera showed various agglutinin titers against heat-killed cells, and titers were generally higher with homologous cells. The comparison of the different antigens using agar gel diffusion and immunoelectrophoresis showed the presence of extracellular common antigens in both ESA and WEA between the different strains. Absorption of anti-ESA sera with WEA, and anti-WEA sera with ESA, showed the existence of a specific antigen common to all bacteria in each fraction. Enzymatic treatment of the antigen before immunodiffusion demonstrated the protein nature of the two antigens present in ESA and WEA. Images PMID:6783541

  12. Exclusion of mRNPs and ribosomal particles from a thin zone beneath the nuclear envelope revealed upon inhibition of transport

    International Nuclear Information System (INIS)

    We have studied the nucleocytoplasmic transport of a specific messenger RNP (mRNP) particle, named Balbiani ring (BR) granule, and ribosomal RNP (rRNP) particles in the salivary glands of the dipteran Chironomus tentans. The passage of the RNPs through the nuclear pore complex (NPC) was inhibited with the nucleoporin-binding wheat germ agglutinin, and the effects were examined by electron microscopy. BR mRNPs bound to the nuclear basket increased in number, while BR mRNPs translocating through the central channel decreased, suggesting that the initiation of translocation proper had been inhibited. The rRNPs accumulated heavily in nucleoplasm, while no or very few rRNPs were recorded within nuclear baskets. Thus, the transport of rRNPs had been blocked prior to the entry into the baskets. Remarkably, the rRNPs had been excluded both from baskets and the space in between the baskets. We propose that normally basket fibrils move freely and repel RNPs from the exclusion zone unless the particles have affinity for and bind to nucleoporins within the baskets.

  13. Novel strategy for yeast construction using delta-integration and cell fusion to efficiently produce ethanol from raw starch.

    Science.gov (United States)

    Yamada, Ryosuke; Tanaka, Tsutomu; Ogino, Chiaki; Fukuda, Hideki; Kondo, Akihiko

    2010-02-01

    We developed a novel strategy for constructing yeast to improve levels of amylase gene expression and the practical potential of yeast by combining delta-integration and polyploidization through cell fusion. Streptococcus bovis alpha-amylase and Rhizopus oryzae glucoamylase/alpha-agglutinin fusion protein genes were integrated into haploid yeast strains. Diploid strains were constructed from these haploid strains by mating, and then a tetraploid strain was constructed by cell fusion. The alpha-amylase and glucoamylase activities of the tetraploid strain were increased up to 1.5- and tenfold, respectively, compared with the parental strain. The diploid and tetraploid strains proliferated faster, yielded more cells, and fermented glucose more effectively than the haploid strain. Ethanol productivity from raw starch was improved with increased ploidy; the tetraploid strain consumed 150 g/l of raw starch and produced 70 g/l of ethanol after 72 h of fermentation. Our strategy for constructing yeasts resulted in the simultaneous overexpression of genes integrated into the genome and improvements in the practical potential of yeasts. PMID:19707752

  14. Effect of ConA—receptor interaction on the structure of cell membrane

    Institute of Scientific and Technical Information of China (English)

    DAIJIANWU; KECHUNLIN; 等

    1992-01-01

    Recently,the effect of ligand receptor interaction on the membrane structure of liposomes has been studied extensively,However,little is known about how it exists on biological membranes,In this paper,the effect of Concanavalin A(ConA) receptorinteratcion on the structure of cell membranes was studied by Circular DIchrosim(CD) and 31P Nuclear Magnetic Resonance(NMR).CD results of both the purified macrophage membranes and human erythrocyte hgosts(EG) showed that the conformation of membrane proteins changed after ConA binding.For further research,31P-NMR was used to detect the orgainzation of phosp[holipid molecules on macrophage membranes.After ConA binding,the tendercy to form non bilayer structure increased with the amount of ConA.The changes of 31P-NMR spectra of living macrophages might be partly due to the above stated reason too.In addition,ConA-receptor interaction also induced similar results of 31P-NMR spectra in EG.In contrast,wheat germ agglutinin (WGA),another kind of lectin,rarely showed the same influence.

  15. A survey of domestic species of Basidiomycetes fungi for the presence of lectins inn their carpophores

    Directory of Open Access Journals (Sweden)

    Grażyna Końska

    2014-02-01

    Full Text Available Preliminary investigations were conducted to determine the presence of active lectins in carpophores of fungi from the class Basidiomycetes, collected from natural localities in southern and south-eastern Poland. The degree of agglutination activity (expressed as the titre of agglutination of aqueous extracts was determined at room temperature (18-20°C and at +4°C in respect to human and animal erythrocytes suspended in physiological saline, part of which were additionally treated with proteolytic enzymes. From among the 104 tested species, extracts from 41 of them showed agglutination activity, among which 18 were high. In six cases, specific activity against human ABH group antigens was found. Extracts from 5 species agglutinated only animal erythrocytes, with pigeon erythrocytes being exceptionally sensitive to the lectins. Extracts from two species had distinctly higher agglutination activity at 4°C, which suggests that lectins of the "cold" agglutinin type are present in these species. Analysis of extracts from caps and stems showed that caps had a higher lectin content.

  16. Synthesis and characterization of near IR fluorescent albumin nanoparticles for optical detection of colon cancer

    International Nuclear Information System (INIS)

    Near IR (NIR) fluorescent human serum albumin (HSA) nanoparticles hold great promise as contrast agents for tumor diagnosis. HSA nanoparticles are considered to be biocompatible, non-toxic and non-immunogenic. In addition, NIR fluorescence properties of these nanoparticles are important for in vivo tumor diagnostics, with low autofluorescence and relatively deep penetration of NIR irradiation due to low absorption of biomatrices. The present study describes the synthesis of new NIR fluorescent HSA nanoparticles, by entrapment of a NIR fluorescent dye within the HSA nanoparticles, which also significantly increases the photostability of the dye. Tumor-targeting ligands such as peanut agglutinin (PNA) and anti-carcinoembryonic antigen antibodies (anti-CEA) were covalently conjugated to the NIR fluorescent albumin nanoparticles, increasing the potential fluorescent signal in tumors with upregulated corresponding receptors. Specific colon tumor detection by the NIR fluorescent HSA nanoparticles was demonstrated in a chicken embryo model and a rat model. In future work we also plan to encapsulate cancer drugs such as doxorubicin within the NIR fluorescent HSA nanoparticles for both colon cancer imaging and therapy. - Highlights: ► Near IR human serum albumin nanoparticles were synthesized and characterized. ► Nanoparticles were shown to be physically and chemically stable and photostable. ► Tumor-targeting ligands were covalently conjugated to the nanoparticles. ► Specific colon cancer tumor detection was demonstrated in chicken-embryo and rat models.

  17. Surface display of recombinant Drosophila melanogaster acetylcholinesterase for detection of organic phosphorus and carbamate pesticides.

    Directory of Open Access Journals (Sweden)

    Jingquan Li

    Full Text Available Acetylcholinesterase (AChE is commonly used for the detection of organophosphate (OP and carbamate (CB insecticides. However, the cost of this commercially available enzyme is high, making high-throughput insecticide detection improbable. In this study we constructed a new AChE yeast expression system in Saccharomyces cerevisiae for the expression of a highly reactive recombinant AChE originating from Drosophila melanogaster (DmAChE. Specifically, the coding sequence of DmAChE was fused with the 3'-terminal half of an α-agglutinin anchor region, along with an antigen tag for the detection of the recombinant protein. The target sequence was cloned into the yeast expression vector pYes-DEST52, and the signal peptide sequence was replaced with a glucoamylase secretion region for induced expression. The resultant engineered vector was transformed into S. cerevisiae. DmAChE was expressed and displayed on the cell surface after galactose induction. Our results showed that the recombinant protein displayed activity comparable to the commercial enzyme. We also detected different types of OP and CB insecticides through enzyme inhibition assays, with the expressed DmAChE showing high sensitivity. These results show the construction of a new yeast expression system for DmAChE, which can subsequently be used for detecting OP and CB insecticides with reduced economic costs.

  18. Partial purification of the mu opioid receptor irreversibly labeled with [3H]b-funaltrexamine

    International Nuclear Information System (INIS)

    The mu opioid receptor in bovine striatal membranes was specifically and irreversibly labeled by incubation with 5 nM [3H]β-funaltrexamine (approx.-FNA) at 370C for 90 min in the presence of 100 mM NaCl. The specific and irreversible binding of [3H]β-FNA as defined by that blocked by 1 +M naloxone was about 60% of total irreversible binding. The specific irreversible binding was saturable, stereospecific, time-, temperature, and tissue-dependent. Mu opioid ligands were much more potent than delta or kappa ligands in inhibiting the specific irreversible labeling. SDS polyacrylamide gel electrophoresis of solubilized membranes in the presence of 2-mercaptoethanol yielded a major radiolabeled broad band of MW 68-97K daltons, characteristic of a glycoprotein band. This band was not observed in membranes labeled in the presence of excess unlabeled naloxone. The glycoprotein nature of the [3H]β-FNA-labeled opioid receptor was confirmed by its binding to a wheat germ agglutinin-Sepharose column and its elution with N-acetylglucosamine

  19. Segmental pairs of giant insect cells discharge presumptive immune proteins at each larval molt.

    Science.gov (United States)

    Nardi, James B; Bee, Charles M; Miller, Lou Ann; Imai, Brian S; Yau, Peter M

    2016-05-15

    A pair of massive secretory cells exists within each thoracic and the nine abdominal segments of Manduca larvae. Each of these cells is nestled between the dorsal integument and underlying muscles. Contents of large vacuoles in these cells are abruptly discharged at each molt and have always been considered to contribute to shedding and/or formation of cuticle. Peanut agglutinin is a specific lectin label for these secretory vacuoles; vacuoles label intensely immediately before each molt as vacuoles attain their maximal size. Contents of vacuoles are restored after each molt and throughout most of each intermolt. During the molt cycle these cells secrete contents of their vacuoles into the interior hemocoel rather than onto the exterior cuticle. Vacuoles discharge via a distinctive mechanism involving partitioning of contents into numerous vesicles that move to the cell surface. Dermal secretory cells were dissected from larvae before and after the 4th-5th instar molt. Proteins from pre-molt and post-molt secretory cells were separated by two-dimensional electrophoresis to establish which proteins are discharged at the molt. While secreted proteins are novel, all have presumptive roles in immune responses. Dermal secretory cells may represent a new, unsuspected component of the innate immune system that release their proteins during the vulnerable molting period of an insect's life. PMID:27039264

  20. Three-dimensional direct measurement of cardiomyocyte volume, nuclearity, and ploidy in thick histological sections

    Science.gov (United States)

    Bensley, Jonathan Guy; de Matteo, Robert; Harding, Richard; Black, Mary Jane

    2016-04-01

    Quantitative assessment of myocardial development and disease requires accurate measurement of cardiomyocyte volume, nuclearity (nuclei per cell), and ploidy (genome copies per cell). Current methods require enzymatically isolating cells, which excludes the use of archived tissue, or serial sectioning. We describe a method of analysis that permits the direct simultaneous measurement of cardiomyocyte volume, nuclearity, and ploidy in thick histological sections. To demonstrate the utility of our technique, heart tissue was obtained from four species (rat, mouse, rabbit, sheep) at up to three life stages: prenatal, weaning and adulthood. Thick (40 μm) paraffin sections were stained with Wheat Germ Agglutinin-Alexa Fluor 488 to visualise cell membranes, and DAPI (4‧,6-diamidino-2-phenylindole) to visualise nuclei and measure ploidy. Previous methods have been restricted to thin sections (2–10 μm) and offer an incomplete picture of cardiomyocytes. Using confocal microscopy and three-dimensional image analysis software (Imaris Version 8.2, Bitplane AG, Switzerland), cardiomyocyte volume, nuclearity, and ploidy were measured. This method of staining and analysis of cardiomyocytes enables accurate morphometric measurements in thick histological sections, thus unlocking the potential of archived tissue. Our novel time-efficient method permits the entire cardiomyocyte to be visualised directly in 3D, eliminating the need for precise alignment of serial sections.

  1. Radiologic and clinical study of mycoplasam pneumonia in children

    International Nuclear Information System (INIS)

    We studied 143 cases of serologically proven (cold agglutinin or 1HA test) M. Pneumonia in children below 15 year of age who admitted to Wallace Memorial Baptist Hospital between March 1986 and August 1988. The following results were noted : 1. Radiologic findings · The patterns of lung infiltration were bronchopneumonic in 52.6%, interstitial in 24.5%, lobar in 3.5%, mixed in 1.3%, and normal or slightly increased bronchovascular markings in 18.1%. · The degree of pulmonary involvement were minimal in 49.2%, moderate in 24.5% and extensive in 8.3%. · The distribution of infiltration in the minimal and moderate pulmonary involvement were upper lobe in 28.6%, middle lobe in 17.2% and lower lobe in 54.3%. · Other findings were bilateral involvement in 31.5%, pleural involvement in 15.8%, new area of consolidation in 3.5% and hilar lymph node involvement in 34.2%. 2. The most common duration of the period between the onset of symptom and confirmation of the diagnosis was 7-14 days. 3. The sex distribution was 1.3 : 1 in male to female ratio. 4. The most peak age incidence was seen in 10-13, and the second one in 4 - 7 year. 5. As for the concomitant illness and complication, asthmatic bronchitis was most common (23 cases, 16.1%) followed by pharyngitis, sinusitis, skin rash and gastroenteritis, etc

  2. Surface display of malolactic enzyme from Oenococcus oeni on Saccharomyces cerevisiae.

    Science.gov (United States)

    Zhang, Xiuyan; Hou, Xiaoyan; Liang, Fang; Chen, Fusheng; Wang, Xiaohong

    2013-04-01

    In order to display malolactic enzyme (MLE) on the cell surface of Saccharomyces cerevisiae, a yeast cell surface display plasmid pADH1-AGG was constructed by fusing the α-factor signal encoding sequence (267 bp) and the C-terminal half of α-agglutinin encoding sequence (1,645 bp) into the plasmid pADH1. The pADH1-AGG could successfully express and anchor the enhanced green fluorescent protein (EGFP) onto the yeast cell surface when the EGFP was used to verify its function. Then the pADH1-MLE was constructed by inserting the MLE encoding sequence (1,600 bp) into the pADH1-AGG and introduced into S. cerevisiae cells. The positive strain carrying pADH1-MLE was confirmed by use of the 6× His monoclonal antibody and fluorescein isothiocyanate-conjugated goat anti-mouse IgG. All results indicated that the MLE was displayed successfully on the cell surface of positive transformant. The MLE activity of genetically engineered yeast strain could turn 21.11 % L-malate into lactic acid after 12 h reaction with L-malate. The constructed yeast strain might be used to conduct malolactic fermentation (MLF) in wine to solve the important issues of sluggish MLF, microbial spoilage, and adverse metabolic substances produced by the lactic acid bacteria. PMID:23446978

  3. Profiling of Leptospira interrogans, L. santarosai, L. meyeri and L. borgpetersenii by SE-AFLP, PFGE and susceptibility testing-a continuous attempt at species and serovar differentiation.

    Science.gov (United States)

    Moreno, Luisa Z; Miraglia, Fabiana; Lilenbaum, Walter; Neto, José Sf; Freitas, Julio C; Morais, Zenaide M; Hartskeerl, Rudy A; da Costa, Barbara Lp; Vasconcellos, Silvio A; Moreno, Andrea M

    2016-01-01

    Leptospirosis is a widespread systemic zoonosis, considered as reemerging in certain developing countries. Although the cross agglutinin absorption test is still considered the standard method for Leptospira identification, it presents several disadvantages. The aim of this study was to characterize Leptospira spp. isolated from various hosts by genotyping and broth microdilution susceptibility testing in an attempt to differentiate Leptospira species, serogroups and serovars. Forty-seven isolates were studied. They were previously serotyped, and species confirmation was performed by 16S rRNA sequencing. Single-enzyme amplified fragment length polymorphism (SE-AFLP) and pulsed-field gel electrophoresis (PFGE) analysis enabled the distinction of L. interrogans from L. santarosai, L. meyeri and L. borgpetersenii in two main clusters. Among L. interrogans, it was possible to differentiate into two new clusters the serogroup Icterohaemorrhagiae from the serogroups Canicola and Pomona. L. santarosai isolates presented higher genetic variation than the other species in both techniques. Interestingly, the minimum inhibitory concentration (MIC) cluster analysis also provided Leptospira serogroup differentiation. Further studies are necessary regarding serovar Bananal isolates, as they presented the highest MIC values for most of the antimicrobials tested. All studied techniques successfully distinguished Leptospira species and serogroups. Despite being library-dependent methods, these approaches are less labor intensive and more economically viable, particularly SE-AFLP, and can be implemented in most reference laboratories worldwide to enable faster Leptospira typing. PMID:26956446

  4. Purification and molecular cloning of a new galactose-specific lectin from Bauhinia variegata seeds

    Indian Academy of Sciences (India)

    Luciano S Pinto; Celso S Nagano; Taianá M Oliveira; Tales R Moura; Alexandre H Sampaio; Henri Debray; Vicente P Pinto; Odir A Dellagostin; Benildo S Cavada

    2008-09-01

    A new galactose-specific lectin was purified from seeds of a Caesalpinoideae plant, Bauhinia variegata, by affinity chromatography on lactose–agarose. Protein extracts haemagglutinated rabbit and human erythrocytes (native and treated with proteolytic enzymes), showing preference for rabbit blood treated with papain and trypsin. Among various carbohydrates tested, the lectin was best inhibited by D-galactose and its derivatives, especially lactose. SDS-PAGE showed that the lectin, named BVL, has a pattern similar to other lectins isolated from the same genus, Bauhinia purpurea agglutinin (BPA). The molecular mass of BVL subunit is 32 871 Da, determined by MALDI-TOF spectrometry. DNA extracted from B. variegata young leaves and primers designed according to the B. purpurea lectin were used to generate specific fragments which were cloned and sequenced, revealing two distinct isoforms. The bvl gene sequence comprised an open reading frame of 876 base pairs which encodes a protein of 291 amino acids. The protein carried a putative signal peptide. The mature protein was predicted to have 263 amino acid residues and 28 963 Da in size.

  5. Crystallization and preliminary X-ray crystallographic analysis of a galactose-specific lectin from Dolichos lablab

    International Nuclear Information System (INIS)

    The galactose-specific lectin from the seeds of a leguminous plant, D. lablab, has been crystallized. Molecular-replacement solution using 3.0 Å X-ray diffraction data showed the lectin to be a tetramer. The galactose-specific lectin from the seeds of Dolichos lablab has been crystallized using the hanging-drop vapour-diffusion technique. The crystals belong to space group P1, with unit-cell parameters a = 73.99, b = 84.13, c = 93.15 Å, α = 89.92, β = 76.01, γ = 76.99°. X-ray diffraction data to a resolution of 3.0 Å have been collected under cryoconditions (100 K) using a MAR imaging-plate detector system mounted on a rotating-anode X-ray generator. Molecular-replacement calculations carried out using the available structures of legume lectins as search models revealed that the galactose-specific lectin from D. lablab forms a tetramer similar to soybean agglutinin; two such tetramers are present in the asymmetric unit

  6. The subcommissural organ of the rat secretes Reissner's fiber glycoproteins and CSF-soluble proteins reaching the internal and external CSF compartments

    Directory of Open Access Journals (Sweden)

    Rodríguez Esteban M

    2008-01-01

    Full Text Available Abstract Background The subcommissural organ (SCO is a highly conserved brain gland present throughout the vertebrate phylum; it secretes glycoproteins into the cerebrospinal fluid (CSF, where they aggregate to form Reissner's fiber (RF. SCO-spondin is the major constituent protein of RF. Evidence exists that the SCO also secretes proteins that remain soluble in the CSF. The aims of the present investigation were: (i to identify and partially characterize the SCO-secretory compounds present in the SCO gland itself and in the RF of the Sprague-Dawley rat and non-hydrocephalic hyh mouse, and in the CSF of rat; (ii to make a comparative analysis of the proteins present in these three compartments; (iii to identify the proteins secreted by the SCO into the CSF at different developmental periods. Methods The proteins of the SCO secreted into the CSF were studied (i by injecting specific antibodies into ventricular CSF in vivo; (ii by immunoblots of SCO, RF and CSF samples, using specific antibodies against the SCO secretory proteins (AFRU and anti-P15. In addition, the glycosylated nature of SCO-compounds was analysed by concanavalin A and wheat germ agglutinin binding. To analyse RF-glycoproteins, RF was extracted from the central canal of juvenile rats and mice; to investigate the CSF-soluble proteins secreted by the SCO, CSF samples were collected from the cisterna magna of rats at different stages of development (from E18 to PN30. Results Five glycoproteins were identified in the rat SCO with apparent molecular weights of 630, 450, 390, 320 and 200 kDa. With the exception of the 200-kDa compound, all other compounds present in the rat SCO were also present in the mouse SCO. The 630 and 390 kDa compounds of the rat SCO have affinity for concanavalin A but not for wheat germ agglutinin, suggesting that they correspond to precursor forms. Four of the AFRU-immunoreactive compounds present in the SCO (630, 450, 390, 320 kDa were absent from the RF and

  7. Targeted gene disruption in Candida parapsilosis demonstrates a role for CPAR2_404800 in adhesion to a biotic surface and in a murine model of ascending urinary tract infection.

    Science.gov (United States)

    Bertini, Alessia; Zoppo, Marina; Lombardi, Lisa; Rizzato, Cosmeri; De Carolis, Elena; Vella, Antonietta; Torelli, Riccardo; Sanguinetti, Maurizio; Tavanti, Arianna

    2016-02-17

    Candida parapsilosis is an emerging opportunistic pathogen, second in frequency only to C. albicans and commonly associated with both mucosal and systemic infections. Adhesion to biotic surfaces is a key step for the development of mycoses. The C. parapsilosis genome encodes 5 predicted agglutinin-like sequence proteins and their precise role in the adhesion process still remains to be elucidated. In this study, we focused on the putative adhesin Cpar2_404800, in view of its high homology to the most important adhesion molecule in C. albicans. Two independent lineages of C. parapsilosis CPAR2_404800 heterozygous and null mutants were obtained by site-specific deletion. CPAR2_404800 mutants did not differ from wild-type strain in terms of in vitro growth or in their ability to undergo morphogenesis. However, when compared for adhesion to a biotic surface, CPAR2_404800 null mutants exhibited a marked reduction in their adhesion to buccal epithelial cells (>60% reduction of adhesion index). Reintroduction of one copy of CPAR2_404800 gene in the null background restored wild type phenotype. A murine model of urinary tract infection was used to elucidate the in vivo contribution of CPAR2_404800. A 0.5 and 1 log10 reduction in colony forming unit numbers (per gram) was observed respectively in bladder and kidneys obtained from mice infected with null mutant compared to wild-type infected ones. Taken together, these findings provide the first evidence for a direct role of CPAR2_404800 in C. parapsilosis adhesion to host surfaces and demonstrate its contribution to the pathogenesis of murine urinary candidiasis. PMID:26632333

  8. Dissociation of insulin receptor phosphorylation and stimulation of glucose transport in BC3H-1 myocytes

    International Nuclear Information System (INIS)

    The authors have investigated insulin receptor phosphorylation in differentiated cultured BC3H-1 myocytes. As for other insulin-responsive cell systems in partially purified wheat germ agglutinin receptor preparations, insulin stimulates the phosphorylation of its own receptor (95K β-subunits) in a dose dependent manner (0-400 nM), as identified by immunoprecipitation with antiinsulin receptor antibodies and SDS-PAGE. In the same preparations they show that 12-0-tetradecanyl phorbol acetate (TPA), which in many respect β-subunits in the same dose dependent manner (0-5 μM). In addition, antiinsulin receptor antibodies (B-10) also induced phosphorylation of mimics insulin action, also induced phosphorylation of the insulin receptor and HPLC tryptic maps of the 32P-labeled β-subunit were identical to those for insulin-induced receptor phosphorylation. However, while insulin and TPA are potent stimulators of glucose transport in these muscle cells, the antireceptor antibodies alone failed to provoke glucose transport at any concentration. The specificity and activity of these antibodies were confirmed in their system by their ability to inhibit insulin binding and insulin-stimulated glucose transport in a concentration-dependent manner. Their results indicate that phosphorylation of insulin receptor is not a crucial event in mediating insulin action, at least with respect to glucose transport. While the effects of the B-10 antibody in the BC3H-1 myocyte differ from those in the adipocyte, their results provide independent confirmation of their essential conclusion that phosphorylation of the insulin receptor may not be necessary nor sufficient for its acute action in promoting glucose transport

  9. Characterization of the receptor for the 1,4-dihydropyridine Ca2+ channel modulators in skeletal muscle

    International Nuclear Information System (INIS)

    The 1,4-dihydropyridine receptor was purified in high yield from isolated rabbit skeletal muscle triads. The purified receptor contains four integral subunits: α1 (170 kDa), α2 (175/150 kDa), β (52 kDa) and γ (32 kDa) in a 1:1:1:1 stoichiometry. The α2 subunit is a glycoprotein that binds wheat germ agglutinin and its electrophoretic mobility on sodium dodecyl sulfate-polyacrylamide gels is increased upon reduction whereas the α1 subunit does not exhibit these properties but is recognized by specific monoclonal antibodies. These monoclonal antibodies to the α1 subunit, as well as a specific monoclonal antibody to the β subunit, are capable of immunoprecipitating dihydropyridine binding activity from digitonin-solubilized skeletal muscle triads as well as co-immunoprecipitating the α1 and β subunits. The α1 and β subunits are highly labile to mild trypsin digestion in isolated triads but the dihydropyridine binding activity of the receptor does not depend on an intact α1 (dihydropyridine binding) subunit. The α2 and γ subunits are resistant to trypsin digestion and the proteolytic fragments of the α subunit remain associated with the α2 subunit even after solubilization of the receptor with digitonin. A binding site for the dihydropyridines is proposed based on the identification of a 28 kDa peptide that is both photoaffinity labeled with [3H]azidopine and contains the epitope of a specific monoclonal antibody. Polyclonal antibodies raised against a synthetic peptide corresponding to the deduced carboxyl-terminus of the α1 subunit of the receptor did not react with the purified dihydropyridine receptor and skeletal muscle membranes from various sources

  10. Roles of salivary components in Streptococcus mutans colonization in a new animal model using NOD/SCID.e2f1-/- mice.

    Directory of Open Access Journals (Sweden)

    Tatsuro Ito

    Full Text Available Streptococcus mutans plays an important role in biofilm formation on the tooth surface and is the primary causative agent of dental caries. The binding of S. mutans to the salivary pellicle is of considerable etiologic significance and is important in biofilm development. Recently, we produced NOD/SCID.e2f1(-/- mice that show hyposalivation, lower salivary antibody, and an extended life span compared to the parent strain: NOD.e2f1(-/-. In this study we used NOD/SCID.e2f1(-/- 4 or 6 mice to determine the roles of several salivary components in S. mutans colonization in vivo. S. mutans colonization in NOD/SCID.e2f1(-/- mice was significantly increased when mice were pre-treated with human saliva or commercial salivary components. Interestingly, pre-treatment with secretory IgA (sIgA at physiological concentrations promoted significant colonization of S. mutans compared with sIgA at higher concentrations, or with human saliva or other components. Our data suggest the principal effects of specific sIgA on S. mutans occur during S. mutans colonization, where the appropriate concentration of specific sIgA may serve as an anti-microbial agent, agglutinin, or an adherence receptor to surface antigens. Further, specific sIgA supported biofilm formation when the mice were supplied 1% sucrose water and a non-sucrose diet. The data suggests that there are multiple effects exerted by sIgA in S. mutans colonization, with synergistic effects evident under the condition of sIgA and limited nutrients on colonization in NOD/SCID.e2f1(-/- mice. This is a new animal model that can be used to assess prevention methods for dental biofilm-dependent diseases such as dental caries.

  11. Growth-associated protein 43 immunoreactivity in the superficial dorsal horn of the rat spinal cord is localized in atrophic C-fiber, and not in sprouted A-fiber, central terminals after peripheral nerve injury.

    Science.gov (United States)

    Doubell, T P; Woolf, C J

    1997-09-15

    Peripheral nerve injury induces the up-regulation in dorsal root ganglion cells of growth-associated protein 43 (GAP-43) and its transport to the superficial laminae of the dorsal horn of the spinal cord, where it is located primarily in unmyelinated axons and growth-cone like structures. Peripheral nerve injury also induces the central terminals of axotomized myelinated axons to sprout and form novel synaptic contacts in lamina II of the dorsal horn. To investigate whether the sprouting of A-fiber central terminals into lamina II is the consequence of GAP-43 incorporation into their terminal membranes, we have used an ultrastructural analysis with double labelling to identify the localization of GAP-43 immunoreactivity. Transganglionic transport of wheat germ agglutinin conjugated to horseradish peroxidase (WGA-HRP) was used to identify C-fiber terminals. Transganglionic transport of the B fragment of cholera toxin conjugated to horseradish peroxidase (B-HRP) was used to label A-fiber sciatic nerve central terminals in combination with GAP-43 immunocytochemistry. GAP-43 was found to colocalize only with WGA-HRP- and not with B-HRP-labelled synapses or axons. In addition, many single-labelled GAP-43 synapses were observed. Many of the WGA-HRP-labelled terminals that were characterized by degenerative changes were GAP-43 immunoreactive. Our results indicate that peripheral nerve injury induces novel synapse formation of A fibers in lamina II but that up-regulated levels of GAP-43 are present mainly in other axon projections to the superficial dorsal horn. PMID:9303528

  12. Evaluation of glycophenotype in breast cancer by quantum dot-lectin histochemistry

    Directory of Open Access Journals (Sweden)

    Andrade CG

    2013-12-01

    Full Text Available Camila G Andrade,1 Paulo E Cabral Filho,2 Denise PL Tenório,3 Beate S Santos,4 Eduardo IC Beltrão,1 Adriana Fontes,2 Luiz B Carvalho Jr1 1Keizo Asami Immunopathology Laboratory, 2Biophysics and Radiobiology Department, 3Fundamental Chemistry Department, 4Pharmaceutical Science Department, Federal University of Pernambuco, Recife, Pernambuco, Brazil Abstract: Cell surface glycoconjugates play an important role in differentiation/dedifferentiation processes and lectins are employed to evaluate them by several methodologies. Fluorescent probes are considered a valuable tool because of their ability to provide a particular view, and are more detailed and sensitive in terms of cell structure and molecular content. The aim of this study was to evaluate and compare the expression and distribution of glycoconjugates in normal human breast tissue, and benign (fibroadenoma, and malignantly transformed (invasive ductal carcinoma breast tissues. For this, we used mercaptosuccinic acid-coated Cadmium Telluride (CdTe quantum dots (QDs conjugated with concanavalin A (Con A or Ulex europaeus agglutinin I (UEA I lectins to detect α-D-glucose/mannose and L-fucose residues, respectively. The QD-lectin conjugates were evaluated by hemagglutination activity tests and carbohydrate inhibition assays, and were found to remain functional, keeping their fluorescent properties and carbohydrate recognition ability. Fluorescence images showed that different regions of breast tissue expressed particular types of carbohydrates. While the stroma was preferentially and intensely stained by QD-Con A, ductal cells were preferentially labeled by QD-UEA I. These results indicate that QD-lectin conjugates can be used as molecular probes and can help to elucidate the glycoconjugate profile in biological processes. Keywords: nanoparticles, concanavalin A, Ulex europaeus, carbohydrates, mammary tissues

  13. Purification and partial characterization of a receptor protein for mouse interferon γ

    International Nuclear Information System (INIS)

    A receptor protein for mouse interferon γ has been purified from solubilized plasma membranes of the mouse monomyelocytic cell line WEHI-3. Sequential wheat germ agglutinin and ligand affinity chromatography of membranes extracted with octyl β-D-glucopyranoside resulted in at least a 680-fold purification of the receptor, as measured by precipitating it in association with liposomes composed of phosphatidylcholine. The purified receptor bound 125I-labeled recombinant mouse interferon γ (rMuIFN-γ) with a Kd of 10 nM, a value comparable to that obtained with isolated membranes, PAGE analysis of radiolabeled (with either 35S or 125I) receptor preparations consistently revealed a major band of 95 kDa. This species was degraged with time to smaller fragments, GR-20, a monoclonal antibody against the receptor, completely inhibited specific binding of 125I-labeled rMuIFN-γ to WEHI-3 cells, blocked the induction of priming by rMuIFN-γ of macrophage-mediated tumor cell killing, removed binding activity for 125I-labeled rMuIFN-γ from solubilized membranes, and immunoprecipitated a single 95-kDa protein from the extract of surface labeled (125I) WEHI-3 cells. Cross-linking of 125I-labeled rMuIFN-γ to its receptor yielded a complex of 125 ± 5 kDa, consistent with the binding of the dimeric form of mouse interferon γ (32 kDa) to a membrane protein of 95 kDa. These data suggest that the receptor for mouse interferon γ is a glycoprotein of 95 kDa

  14. A lectin from Platypodium elegans with unusual specificity and affinity for asymmetric complex N-glycans.

    Science.gov (United States)

    Benevides, Raquel Guimarães; Ganne, Géraldine; Simões, Rafael da Conceição; Schubert, Volker; Niemietz, Mathäus; Unverzagt, Carlo; Chazalet, Valérie; Breton, Christelle; Varrot, Annabelle; Cavada, Benildo Sousa; Imberty, Anne

    2012-07-27

    Lectin activity with specificity for mannose and glucose has been detected in the seed of Platypodium elegans, a legume plant from the Dalbergieae tribe. The gene of Platypodium elegans lectin A has been cloned, and the resulting 261-amino acid protein belongs to the legume lectin family with similarity with Pterocarpus angolensis agglutinin from the same tribe. The recombinant lectin has been expressed in Escherichia coli and refolded from inclusion bodies. Analysis of specificity by glycan array evidenced a very unusual preference for complex type N-glycans with asymmetrical branches. A short branch consisting of one mannose residue is preferred on the 6-arm of the N-glycan, whereas extensions by GlcNAc, Gal, and NeuAc are favorable on the 3-arm. Affinities have been obtained by microcalorimetry using symmetrical and asymmetrical Asn-linked heptasaccharides prepared by the semi-synthetic method. Strong affinity with K(d) of 4.5 μm was obtained for both ligands. Crystal structures of Platypodium elegans lectin A complexed with branched trimannose and symmetrical complex-type Asn-linked heptasaccharide have been solved at 2.1 and 1.65 Å resolution, respectively. The lectin adopts the canonical dimeric organization of legume lectins. The trimannose bridges the binding sites of two neighboring dimers, resulting in the formation of infinite chains in the crystal. The Asn-linked heptasaccharide binds with the 6-arm in the primary binding site with extensive additional contacts on both arms. The GlcNAc on the 6-arm is bound in a constrained conformation that may rationalize the higher affinity observed on the glycan array for N-glycans with only a mannose on the 6-arm. PMID:22692206

  15. A Lectin from Platypodium elegans with Unusual Specificity and Affinity for Asymmetric Complex N-Glycans*

    Science.gov (United States)

    Benevides, Raquel Guimarães; Ganne, Géraldine; Simões, Rafael da Conceição; Schubert, Volker; Niemietz, Mathäus; Unverzagt, Carlo; Chazalet, Valérie; Breton, Christelle; Varrot, Annabelle; Cavada, Benildo Sousa; Imberty, Anne

    2012-01-01

    Lectin activity with specificity for mannose and glucose has been detected in the seed of Platypodium elegans, a legume plant from the Dalbergieae tribe. The gene of Platypodium elegans lectin A has been cloned, and the resulting 261-amino acid protein belongs to the legume lectin family with similarity with Pterocarpus angolensis agglutinin from the same tribe. The recombinant lectin has been expressed in Escherichia coli and refolded from inclusion bodies. Analysis of specificity by glycan array evidenced a very unusual preference for complex type N-glycans with asymmetrical branches. A short branch consisting of one mannose residue is preferred on the 6-arm of the N-glycan, whereas extensions by GlcNAc, Gal, and NeuAc are favorable on the 3-arm. Affinities have been obtained by microcalorimetry using symmetrical and asymmetrical Asn-linked heptasaccharides prepared by the semi-synthetic method. Strong affinity with Kd of 4.5 μm was obtained for both ligands. Crystal structures of Platypodium elegans lectin A complexed with branched trimannose and symmetrical complex-type Asn-linked heptasaccharide have been solved at 2.1 and 1.65 Å resolution, respectively. The lectin adopts the canonical dimeric organization of legume lectins. The trimannose bridges the binding sites of two neighboring dimers, resulting in the formation of infinite chains in the crystal. The Asn-linked heptasaccharide binds with the 6-arm in the primary binding site with extensive additional contacts on both arms. The GlcNAc on the 6-arm is bound in a constrained conformation that may rationalize the higher affinity observed on the glycan array for N-glycans with only a mannose on the 6-arm. PMID:22692206

  16. Autoimmune Hemolytic Anemia in Children: Mayo Clinic Experience.

    Science.gov (United States)

    Sankaran, Janani; Rodriguez, Vilmarie; Jacob, Eapen K; Kreuter, Justin D; Go, Ronald S

    2016-04-01

    We studied 35 pediatric patients with autoimmune hemolytic anemia seen at Mayo Clinic from 1994 to 2014. The median age was 10.0 years and 65.7% were males. Most had warm antibodies (80.0%) and some secondary to viral (14.3%) or autoimmune disorders (31.4%). Seven (20.0%) patients presented with Evans syndrome, 3 of whom also had common variable immunodeficiency. The median hemoglobin at diagnosis was 6.1 g/dL and 62.8% patients required red cell transfusions. The severity of anemia was worse among children below 10 years (median 5.5 vs. 7.0 g/dL, P=0.01). Steroid was the initial treatment for 88.5% patients, with overall response rate of 82.7% (68.5% complete, 14.2% partial) and median response duration of 10.7 months (range, 0.2 to 129.7+ mo). After median follow-up of 26.6 months, 8 (22.8%) patients relapsed. Salvage treatments included splenectomy, intravenous immunoglobulin, rituximab, and mycophenolate mofetil. Infectious complications occurred in 9 (25.7%) patients and 1 patient died of cytomegalovirus infection. Four patients had cold agglutinin disease and 3 (75.0%) responded to steroids. Autoimmune hemolytic anemia is a rare disorder in pediatric population and most respond well to steroids regardless of the type of antibody. Infectious complications are common and screening for immunodeficiency is recommended among those with Evans syndrome. PMID:26925716

  17. Tri-partite complex for axonal transport drug delivery achieves pharmacological effect

    Directory of Open Access Journals (Sweden)

    Frederickson Martyn

    2010-01-01

    Full Text Available Abstract Background Targeted delivery of pharmaceutical agents into selected populations of CNS (Central Nervous System neurons is an extremely compelling goal. Currently, systemic methods are generally used for delivery of pain medications, anti-virals for treatment of dermatomal infections, anti-spasmodics, and neuroprotectants. Systemic side effects or undesirable effects on parts of the CNS that are not involved in the pathology limit efficacy and limit clinical utility for many classes of pharmaceuticals. Axonal transport from the periphery offers a possible selective route, but there has been little progress towards design of agents that can accomplish targeted delivery via this intraneural route. To achieve this goal, we developed a tripartite molecular construction concept involving an axonal transport facilitator molecule, a polymer linker, and a large number of drug molecules conjugated to the linker, then sought to evaluate its neurobiology and pharmacological behavior. Results We developed chemical synthesis methodologies for assembling these tripartite complexes using a variety of axonal transport facilitators including nerve growth factor, wheat germ agglutinin, and synthetic facilitators derived from phage display work. Loading of up to 100 drug molecules per complex was achieved. Conjugation methods were used that allowed the drugs to be released in active form inside the cell body after transport. Intramuscular and intradermal injection proved effective for introducing pharmacologically effective doses into selected populations of CNS neurons. Pharmacological efficacy with gabapentin in a paw withdrawal latency model revealed a ten fold increase in half life and a 300 fold decrease in necessary dose relative to systemic administration for gabapentin when the drug was delivered by axonal transport using the tripartite vehicle. Conclusion Specific targeting of selected subpopulations of CNS neurons for drug delivery by axonal

  18. Carbohydrate binding activity in human spermatozoa: localization, specificity, and involvement in sperm-egg fusion.

    Science.gov (United States)

    Gabriele, A; D'Andrea, G; Cordeschi, G; Properzi, G; Giammatteo, M; De Stefano, C; Romano, R; Francavilla, F; Francavilla, S

    1998-06-01

    Sperm carbohydrate binding activity is involved in gamete recognition. We identified a human sperm protein extracted under reducing conditions, and with a molecular mass of 65 kDa on sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE), and which binds D-mannose coupled to albumin (DMA) in presence of cations and a neutral pH. Epifluorescence microscopy showed that fluorescein-DMA binds to dead or permeabilized sperm heads. The DMA-binding activity of human sperm heads was highly specific for a polysaccharide structure containing charged sugar residues. After capacitation, or induction of the acrosome reaction using solubilized zonae pellucidae, fluorescein-DMA was bound respectively to 10.3% (+/- 3.5%) and to 37.6% (+/- 2.1%) of viable sperm heads. The sequential analysis of viable spermatozoa for fluorescein-DMA binding and for rhodamine-Pisum sativum agglutinin binding, showed that DMA-binding sites are present in viable acrosome-reacted spermatozoa. Three dimensional analysis of fluorescence and ultrastructural studies showed that DMA-binding sites are mostly restricted to the sub-acrosomal space of the equatorial segment. Incubation of spermatozoa and zona-free hamster eggs in the presence of DMA was associated with a dose-dependent significant reduction in the number of spermatozoa bound to the oolemma, compared with a control, and to a dose-dependent inhibition of oocyte penetration. This effect was highly specific for DMA, suggesting that DMA-binding sites in human spermatozoa are involved in sperm-egg fusion. PMID:9665337

  19. Loss of CD34 expression in aging human choriocapillaris endothelial cells.

    Directory of Open Access Journals (Sweden)

    Elliott H Sohn

    Full Text Available Structural and gene expression changes in the microvasculature of the human choroid occur during normal aging and age-related macular degeneration (AMD. In this study, we sought to determine the impact of aging and AMD on expression of the endothelial cell glycoprotein CD34. Sections from 58 human donor eyes were categorized as either young (under age 40, age-matched controls (> age 60 without AMD, or AMD affected (>age 60 with early AMD, geographic atrophy, or choroidal neovascularization. Dual labeling of sections with Ulex europaeus agglutinin-I lectin (UEA-I and CD34 antibodies was performed, and the percentage of capillaries labeled with UEA-I but negative for anti-CD34 was determined. In addition, published databases of mouse and human retinal pigment epithelium-choroid were evaluated and CD34 expression compared between young and old eyes. Immunohistochemical studies revealed that while CD34 and UEA-I were colocalized in young eyes, there was variable loss of CD34 immunoreactivity in older donor eyes. While differences between normal aging and AMD were not significant, the percentage of CD34 negative capillaries in old eyes, compared to young eyes, was highly significant (p = 3.8×10(-6. Endothelial cells in neovascular membranes were invariably CD34 positive. Published databases show either a significant decrease in Cd34 (mouse or a trend toward decreased CD34 (human in aging. These findings suggest that UEA-I and endogenous alkaline phosphatase activity are more consistent markers of aging endothelial cells in the choroid, and suggest a possible mechanism for the increased inflammatory milieu in the aging choroid.

  20. Characterization of glutamatergic neurons in the rat atrial intrinsic cardiac ganglia that project to the cardiac ventricular wall.

    Science.gov (United States)

    Wang, Ting; Miller, Kenneth E

    2016-08-01

    The intrinsic cardiac nervous system modulates cardiac function by acting as an integration site for regulating autonomic efferent cardiac output. This intrinsic system is proposed to be composed of a short cardio-cardiac feedback control loop within the cardiac innervation hierarchy. For example, electrophysiological studies have postulated the presence of sensory neurons in intrinsic cardiac ganglia (ICG) for regional cardiac control. There is still a knowledge gap, however, about the anatomical location and neurochemical phenotype of sensory neurons inside ICG. In the present study, rat ICG neurons were characterized neurochemically with immunohistochemistry using glutamatergic markers: vesicular glutamate transporters 1 and 2 (VGLUT1; VGLUT2), and glutaminase (GLS), the enzyme essential for glutamate production. Glutamatergic neurons (VGLUT1/VGLUT2/GLS) in the ICG that have axons to the ventricles were identified by retrograde tracing of wheat germ agglutinin-horseradish peroxidase (WGA-HRP) injected in the ventricular wall. Co-labeling of VGLUT1, VGLUT2, and GLS with the vesicular acetylcholine transporter (VAChT) was used to evaluate the relationship between post-ganglionic autonomic neurons and glutamatergic neurons. Sequential labeling of VGLUT1 and VGLUT2 in adjacent tissue sections was used to evaluate the co-localization of VGLUT1 and VGLUT2 in ICG neurons. Our studies yielded the following results: (1) ICG contain glutamatergic neurons with GLS for glutamate production and VGLUT1 and 2 for transport of glutamate into synaptic vesicles; (2) atrial ICG contain neurons that project to ventricle walls and these neurons are glutamatergic; (3) many glutamatergic ICG neurons also were cholinergic, expressing VAChT; (4) VGLUT1 and VGLUT2 co-localization occurred in ICG neurons with variation of their protein expression level. Investigation of both glutamatergic and cholinergic ICG neurons could help in better understanding the function of the intrinsic cardiac

  1. Display of wasp venom allergens on the cell surface of Saccharomyces cerevisiae

    Directory of Open Access Journals (Sweden)

    Poulsen Lars K

    2010-09-01

    Full Text Available Abstract Background Yeast surface display is a technique, where the proteins of interest are expressed as fusions with yeast surface proteins and thus remain attached to the yeast cell wall after expression. Our purpose was to study whether allergens expressed on the cell surface of baker's yeast Saccharomyces cerevisiae preserve their native allergenic properties and whether the yeast native surface glycoproteins interfere with IgE binding. We chose to use the major allergens from the common wasp Vespula vulgaris venom: phospholipase A1, hyaluronidase and antigen 5 as the model. Results The proteins were expressed on the surface as fusions with a-agglutinin complex protein AGA2. The expression was confirmed by fluorescent cytometry (FACS after staining the cells with antibody against a C-tag attached to the C-terminal end of the allergens. Phospholipase A1 and hyaluronidase retained their enzymatic activities. Phospholipase A1 severely inhibited the growth of the yeast cells. Antigen 5 - expressing yeast cells bound IgE antibodies from wasp venom allergic patient sera but not from control sera as demonstrated by FACS. Moreover, antigen 5 - expressing yeast cells were capable of mediating allergen-specific histamine release from human basophils. Conclusions All the three major wasp venom allergens were expressed on the yeast surface. A high-level expression, which was observed only for antigen 5, was needed for detection of IgE binding by FACS and for induction of histamine release. The non-modified S. cerevisiae cells did not cause any unspecific reaction in FACS or histamine release assay despite the expression of high-mannose oligosaccharides. In perspective the yeast surface display may be used for allergen discovery from cDNA libraries and possibly for sublingual immunotherapy as the cells can serve as good adjuvant and can be produced in large amounts at a low price.

  2. Potential effects of oilseed rape expressing oryzacystatin-1 (OC-1 and of purified insecticidal proteins on larvae of the solitary bee Osmia bicornis.

    Directory of Open Access Journals (Sweden)

    Roger Konrad

    Full Text Available Despite their importance as pollinators in crops and wild plants, solitary bees have not previously been included in non-target testing of insect-resistant transgenic crop plants. Larvae of many solitary bees feed almost exclusively on pollen and thus could be highly exposed to transgene products expressed in the pollen. The potential effects of pollen from oilseed rape expressing the cysteine protease inhibitor oryzacystatin-1 (OC-1 were investigated on larvae of the solitary bee Osmia bicornis (= O. rufa. Furthermore, recombinant OC-1 (rOC-1, the Bt toxin Cry1Ab and the snowdrop lectin Galanthus nivalis agglutinin (GNA were evaluated for effects on the life history parameters of this important pollinator. Pollen provisions from transgenic OC-1 oilseed rape did not affect overall development. Similarly, high doses of rOC-1 and Cry1Ab as well as a low dose of GNA failed to cause any significant effects. However, a high dose of GNA (0.1% in the larval diet resulted in significantly increased development time and reduced efficiency in conversion of pollen food into larval body weight. Our results suggest that OC-1 and Cry1Ab expressing transgenic crops would pose a negligible risk for O. bicornis larvae, whereas GNA expressing plants could cause detrimental effects, but only if bees were exposed to high levels of the protein. The described bioassay with bee brood is not only suitable for early tier non-target tests of transgenic plants, but also has broader applicability to other crop protection products.

  3. Potential effects of oilseed rape expressing oryzacystatin-1 (OC-1) and of purified insecticidal proteins on larvae of the solitary bee Osmia bicornis.

    Science.gov (United States)

    Konrad, Roger; Ferry, Natalie; Gatehouse, Angharad M R; Babendreier, Dirk

    2008-01-01

    Despite their importance as pollinators in crops and wild plants, solitary bees have not previously been included in non-target testing of insect-resistant transgenic crop plants. Larvae of many solitary bees feed almost exclusively on pollen and thus could be highly exposed to transgene products expressed in the pollen. The potential effects of pollen from oilseed rape expressing the cysteine protease inhibitor oryzacystatin-1 (OC-1) were investigated on larvae of the solitary bee Osmia bicornis (= O. rufa). Furthermore, recombinant OC-1 (rOC-1), the Bt toxin Cry1Ab and the snowdrop lectin Galanthus nivalis agglutinin (GNA) were evaluated for effects on the life history parameters of this important pollinator. Pollen provisions from transgenic OC-1 oilseed rape did not affect overall development. Similarly, high doses of rOC-1 and Cry1Ab as well as a low dose of GNA failed to cause any significant effects. However, a high dose of GNA (0.1%) in the larval diet resulted in significantly increased development time and reduced efficiency in conversion of pollen food into larval body weight. Our results suggest that OC-1 and Cry1Ab expressing transgenic crops would pose a negligible risk for O. bicornis larvae, whereas GNA expressing plants could cause detrimental effects, but only if bees were exposed to high levels of the protein. The described bioassay with bee brood is not only suitable for early tier non-target tests of transgenic plants, but also has broader applicability to other crop protection products. PMID:18628826

  4. Impact of transgenic oilseed rape expressing oryzacystatin-1 (OC-1) and of insecticidal proteins on longevity and digestive enzymes of the solitary bee Osmia bicornis.

    Science.gov (United States)

    Konrad, Roger; Connor, Melanie; Ferry, Natalie; Gatehouse, Angharad M R; Babendreier, Dirk

    2009-04-01

    The risk that insect-resistant transgenic plants may pose for solitary bees was assessed by determining longevity of adult Osmia bicornis (O. rufa) chronically exposed to transgenic oilseed rape expressing oryzacystatin-1 (OC-1) or to the purified insecticidal proteins recombinant rOC-1, Kunitz soybean trypsin inhibitor (SBTI), Galanthus nivalis agglutinin (GNA), or Bacillus thuringiensis toxin Cry1Ab dissolved in sugar solution (at 0.01 and 0.1%, w:v, Cry1Ab only at 0.01%). Compared to control bees, longevity was significantly reduced by SBTI and GNA at both concentrations and by rOC-1 at 0.1%, but not by Cry1Ab or rOC-1 at 0.01%. Longevity on the OC-1 oilseed rape was not significantly different from the control plants. The effects of SBTI and rOC-1 on longevity were investigated through characterization of the digestive proteinases of O. bicornis and analysis of the response in proteinase profiles to ingestion of these proteinase inhibitors. A relatively complex profile of at least four types of soluble proteolytic enzymes was identified. Serine proteinases were found to be predominant, with metallo and especially cysteine proteinases making a smaller albeit significant contribution. The compensatory response to in vivo enzyme inhibition was similar for SBTI and rOC-1 although less pronounced for rOC-1. It consisted of a non-specific overproduction of native proteinases, both sensitive and insensitive, and the induction of a novel aspartic proteinase. PMID:19135058

  5. Agonist-promoted desensitization and phosphorylation of. cap alpha. /sub 1/-adrenergic receptors coupled to stimulation of phosphatidylinositol metabolism

    Energy Technology Data Exchange (ETDEWEB)

    Leeb-Lundberg, L.M.F.; Cotecchia, S.; Caron, M.G.; Lefkowitz, R.J.

    1986-03-05

    In the DDT/sub 1/ MF-2 hamster vas deferens smooth muscle cell line the ..cap alpha../sub 1/-adrenergic receptor (..cap alpha../sub 1/-AR) agonist norepinephrine (NE) promotes rapid attenuation of ..cap alpha../sub 1/-AR-mediated phosphatidylinositol (PI) metabolism which is paralleled by rapid phosphorylation of the ..cap alpha../sub 1/-AR. Cells were labeled by incubation with /sup 32/P/sub i/. Coincubation with NE (100 ..mu..M) significantly increases the rate of /sup 32/P-labeling of both PI and phosphatidic acid. Pretreatment of cells with 100 ..mu..M NE (in the presence of 1 ..mu..M propranolol to prevent ..beta..-AR interactions) results in a drastic attenuation of the NE response on PI metabolism. ..cap alpha../sub 1/-AR from labeled cells can be solubilized and purified by affinity chromatography on Affigel-A55414 and wheat germ agglutinin agarose chromatography. SDS-PAGE of purified ..cap alpha../sub 1/-AR shows a NE-promoted increase in phosphorylation of the M/sub r/ 80K ligand binding peptide. Stoichiometry of phosphorylation increases from approx. 1 mol phosphate/mol ..cap alpha../sub 1/-AR in the basal condition to approx. 2.5 after NE treatment. Both desensitization and phosphorylation are rapid being maximal within 10-20 min of agonist exposure. These results together with previous findings that phorbol esters promote rapid ..cap alpha../sub 1/-AR uncoupling and phosphorylation suggest that receptor phosphorylation is an important mechanism of regulation of ..cap alpha../sub 1/-AR receptor responsiveness.

  6. Enhanced accuracy of kinetic information from CT-CPMG experiments by transverse rotating-frame spectroscopy

    International Nuclear Information System (INIS)

    Micro-to-millisecond motions of proteins transmit pivotal signals for protein function. A powerful technique for the measurement of these motions is nuclear magnetic resonance spectroscopy. One of the most widely used methodologies for this purpose is the constant-time Carr–Purcell–Meiboom–Gill (CT-CPMG) relaxation dispersion experiment where kinetic and structural information can be obtained at atomic resolution. Extraction of accurate kinetics determined from CT-CPMG data requires refocusing frequencies that are much larger than the nuclei’s exchange rate between states. We investigated the effect when fast processes are probed by CT-CPMG experiments via simulation and show that if the intrinsic relaxation rate (R(CT-CPMG)(2,0) is not known a priori the extraction of accurate kinetics is hindered. Errors on the order of 50 % in the exchange rate are attained when processes become fast, but are minimized to 5 % with a priori R2,0CT-CPMG information. To alleviate this shortcoming, we developed an experimental scheme probing R2,0CT-CPMG with large amplitude spin-lock fields, which specifically contains the intrinsic proton longitudinal Eigenrelaxation rate. Our approach was validated with ubiquitin and the Oscillatoria agardhii agglutinin (OAA). For OAA, an underestimation of 66 % in the kinetic rates was observed if R2,0CT-CPMG is not included during the analysis of CT-CPMG data and result in incorrect kinetics and imprecise amplitude information. This was overcome by combining CT-CPMG with R2,0CT-CPMG measured with a high power R1ρ experiment. In addition, the measurement of R2,0CT-CPMG removes the ambiguities in choosing between different models that describe CT-CPMG data

  7. Effect of 3-bromopyruvate and atovaquone on infection during in vitro interaction of Toxoplasma gondii and LLC-MK2 cells.

    Science.gov (United States)

    de Lima, Loyze Paola O; Seabra, Sergio H; Carneiro, Henrique; Barbosa, Helene S

    2015-09-01

    Toxoplasma gondii infection can be severe during pregnancy and in immunocompromised patients. Current therapies for toxoplasmosis are restricted to tachyzoites and have little or no effect on bradyzoites, which are maintained in tissue cysts. Consequently, new therapeutic alternatives have been proposed as the use of atovaquone has demonstrated partial efficacy against tachyzoites and bradyzoites. This work studies the effect of 3-bromopyruvate (3-BrPA), a compound that is being tested against cancer cells, on the infection of LLC-MK2 cells with T. gondii tachyzoites, RH strain. No effect of 3-BrPA on host cell proliferation or viability was observed, but it inhibited the proliferation of T. gondii. The incubation of cultures with lectin Dolichos biflorus agglutinin (DBA) showed the development of cystogenesis, and an ultrastructural analysis of parasite intracellular development confirmed morphological characteristics commonly found in tissue cysts. Moreover, the presence of degraded parasites and the influence of 3-BrPA on endodyogeny were observed. Infected cultures were alternatively treated with a combination of this compound plus atovaquone. This resulted in a 73% reduction in intracellular parasites after 24 h of treatment and a 71% reduction after 48 h; cyst wall formation did not occur in these cultures. Therefore, we conclude that the use of 3-BrPA may serve as an important tool for the study of (i) in vitro cystogenesis; (ii) parasite metabolism, requiring a deeper understanding of the target of action of this compound on T. gondii; (iii) the alternative parasite metabolic pathways; and (iv) the molecular/cellular mechanisms that trigger parasite death. PMID:26077255

  8. The benefits of liposomes for chilling canine sperm for 4 days at 4°C.

    Science.gov (United States)

    Belala, Redha; Delay, Juliette; Amirat, Lamia; Ropers, Marie-Hélène; Guillou, Jocya Le; Anton, Marc; Schmitt, Eric; Thorin, Chantal; Michaud, Sandrine; Kaidi, Rachid; Tainturier, Daniel; Bencharif, Djemil

    2016-05-01

    This study comprises 3 experiments exploring the possible benefits and mechanism of action of liposomes for chilling (4°C) canine sperm over a period of 4 days. In the first experiment, 20 ejaculates collected from 5 Beagle dogs were chilled in an extender containing 6% low density lipoproteins (LDL) (Control), or one of 7 extenders containing different concentrations (2, 4, 6, 8, 10, 15, 20%) of liposomes (LIPO). These ejaculates were chilled over 4 days and motility was assessed daily using a Hamilton Thorne analyzer (HTM-IVOS, 14.0). The 2% LIPO obtained the best results (p=0.038) after four days (72.55% motile spermatozoa and 31.4% progressive spermatozoa). In experiment 2, 10 ejaculates were collected from same 5 dogs and chilled in 6% LDL or 2% LIPO-based extenders. Sperm integrity characteristics were assessed prior to refrigeration and every 48h for four days (D0, D2, and D4). Acrosome integrity was assessed using the FITC-PSA test (Fluorescein IsoThiocyanate-Pisum Sativum Agglutinin), plasma membrane (PM) integrity using both the hypo-osmotic swelling test (HOSt) and SYBR14/Propidium Iodide test (SYBR14/PI), and DNA integrity using the Acridine-Orange test (AO). The 2% LIPO extender provided equivalent preservation of sperm integrity parameters to the reference extender (6% LDL). In experiment 3, a Langmuir-Blodgett trough was used to evaluate the mechanistic interactions between LDL, LIPO, prostatic fluid, and the canine spermatozoal membrane during chilling. Results indicate that LDL and LIPO interact differently with the biomimetic membrane. The most likely conclusion of these findings is that LDL and liposomes employ different protective mechanisms during the chilling (4°C) of canine spermatozoa. PMID:26952759

  9. Square-wave voltammetry assays for glycoproteins on nanoporous gold.

    Science.gov (United States)

    Pandey, Binod; Bhattarai, Jay K; Pornsuriyasak, Papapida; Fujikawa, Kohki; Catania, Rosa; Demchenko, Alexei V; Stine, Keith J

    2014-03-15

    Electrochemical enzyme-linked lectinsorbent assays (ELLA) were developed using nanoporous gold (NPG) as a solid support for protein immobilization and as an electrode for the electrochemical determination of the product of the reaction between alkaline phosphatase (ALP) and p-aminophenyl phosphate (p-APP), which is p-aminophenol (p-AP). Glycoproteins or concanavalin A (Con A) and ALP conjugates were covalently immobilized onto lipoic acid self-assembled monolayers on NPG. The binding of Con A - ALP (or soybean agglutinin - ALP) conjugate to glycoproteins covalently immobilized on NPG and subsequent incubation with p-APP substrate was found to result in square-wave voltammograms whose peak difference current varied with the identity of the glycoprotein. NPG presenting covalently bound glycoproteins was used as the basis for a competitive electrochemical assay for glycoproteins in solution (transferrin and IgG). A kinetic ELLA based on steric hindrance of the enzyme-substrate reaction and hence reduced enzymatic reaction rate after glycoprotein binding is demonstrated using immobilized Con A-ALP conjugates. Using the immobilized Con A-ALP conjugate, the binding affinity of immunoglobulin G (IgG) was found to be 105 nM, and that for transferrin was found to be 650 nM. Minimal interference was observed in the presence of 5 mg mL(-1) BSA as a model serum protein in both the kinetic and competitive ELLA. Inhibition studies were performed with methyl D-mannoside for the binding of TSF and IgG to Con A-ALP; IC50 values were found to be 90 μM and 286 μM, respectively. Surface coverages of proteins were estimated using solution depletion and the BCA protein concentration assay. PMID:24611035

  10. Hepatitis C Virus Resistance to Carbohydrate-Binding Agents.

    Directory of Open Access Journals (Sweden)

    Laure Izquierdo

    Full Text Available Carbohydrate binding agents (CBAs, including natural lectins, are more and more considered as broad-spectrum antivirals. These molecules are able to directly inhibit many viruses such as Human Immunodeficiency Virus (HIV, Hepatitis C Virus (HCV, Dengue Virus, Ebola Virus or Severe Acute Respiratory Syndrome Coronavirus through binding to envelope protein N-glycans. In the case of HIV, it has been shown that CBAs select for mutant viruses with N-glycosylation site deletions which are more sensitive to neutralizing antibodies. In this study we aimed at evaluating the HCV resistance to CBAs in vitro. HCV was cultivated in the presence of increasing Galanthus nivalis agglutinin (GNA, Cyanovirin-N, Concanavalin-A or Griffithsin concentrations, during more than eight weeks. At the end of lectin exposure, the genome of the isolated strains was sequenced and several potential resistance mutations in the E1E2 envelope glycoproteins were identified. The effect of these mutations on viral fitness as well as on sensitivity to inhibition by lectins, soluble CD81 or the 3/11 neutralizing antibody was assessed. Surprisingly, none of these mutations, alone or in combination, conferred resistance to CBAs. In contrast, we observed that some mutants were more sensitive to 3/11 or CD81-LEL inhibition. Additionally, several mutations were identified in the Core and the non-structural proteins. Thus, our results suggest that in contrast to HIV, HCV resistance to CBAs is not directly conferred by mutations in the envelope protein genes but could occur through an indirect mechanism involving mutations in other viral proteins. Further investigations are needed to completely elucidate the underlying mechanisms.

  11. Anti-HIV Ⅰ/Ⅱ Activity and Molecular Cloning of a Novel Mannose/Sialic Acid-binding Lectin from Rhizome of Polygonatum cyrtonema Hua

    Institute of Scientific and Technical Information of China (English)

    Jie AN; Jin-Ku BAO; Jin-Zhi LIU; Chuan-Fang WU; Jian LI; Lei DAI; Els Van DAMME; Jan BALZARINI; Erik De CLERCQ; Fang CHEN

    2006-01-01

    The anti-human immunodeficiency virus (HIV) Ⅰ/Ⅱ activity of a mannose and sialic acid binding lectin isolated from rhizomes of Polygonatum cyrtonema Hua was elucidated by comparing its HIV infection inhibitory activity in MT-4 and CEM cells with that of other mannose-binding lectins (MBLs). The anti-HIV activity of Polygonatum cyrtonema Hua lectin (PCL) was 10- to 100-fold more potent than other tested MBLs, but without significant cytotoxicity towards MT-4 or CEM cells. To amplify cDNA of PCL by 3'/5'-rapid amplification of cDNA ends (RACE), the 30 amino acids of N-terminal were determined by sequencing and the degenerate oligonucleotide primers were designed. The full-length cDNA of PCL contained 693 bp with an open reading frame encoding a precursor protein of 160 amino acid residues, consisting of a 28-residue signal peptide, a 22-residue C-terminal cleavage peptide and a 110-residue mature polypeptide which contained three tandemly arranged subdomains with an obvious sequence homology to the monocot MBL. However, only one active mannose-binding site (QDNVY) was found in subdomain Ⅰ of PCL, that of subdomain Ⅱ and Ⅲ changed to HNNVY and PDNVY, respectively. There was no intron in PCL, which was in good agreement with other monocot MBLs. Molecular modeling of PCL indicated that its three-dimen-sional structure resembles that of the snowdrop agglutinin. By docking, an active sialic acid-binding site was found in PCL. The instabilization of translation initiation region (TIR) in mRNA of PCL benefits its high expression in rhizomes.

  12. A Chemiluminescent Protein Microarray Method for Determining the Seroglycoid Fucosylation Index.

    Science.gov (United States)

    Zhang, Aiying; Skog, Sven; Wang, Shengqi; Ke, Yang; Zhang, Yonghong; Li, Kang; He, Ellen; Li, Ning

    2016-01-01

    The Lens culinaris agglutinin-reactive fraction of AFP (AFP-L3) is widely used to screen for hepatocellular carcinoma (HCC) in Japan and China. We developed a chemiluminescent protein microarray for determining the AFP-L3/AFP index (the ratio of AFP-L3 to total AFP, AFP-L3%) by fixing AFP-specific antibodies and Lens culinaris lectin on aldehyde-coated glass slides. Serum samples were tested for AFP using an enzyme-linked immunosorbent assay (ELISA) to validate the microarray. AFP-L3 was detected using Hotgen Biotech glycosyl capture spin column pretreatment technology and ELISA. When the AFP cut-off value was set to 20 ng/ml, the protein microarray displayed 89.74% sensitivity and 100% specificity for HCC diagnosis, and the ELISA displayed 87.17% sensitivity and 100% specificity. When the AFP-L3% cut-off value was set to 0.1, the protein microarray displayed 56.41% sensitivity and 100% specificity for HCC diagnosis, and the ELISA displayed 53.84% sensitivity and 100% specificity. The ROC curve for the HCC diagnosis showed that the AFP area under the ROC curve (AUC = 0.996; 95% CI: 0.986-1.005) was much higher than that of AFP-L3 (AUC = 0.857; 95% CI: 0.769-0.94) and AFP-L3% (AUC = 0.827; CI: 0.730-0.924). The microarray assay used in this study is a highly sensitive, accurate, and efficient assay for the determination of the AFP-L3%. PMID:27528397

  13. Thermostable mannose-binding lectin from Dendrobium findleyanum with activities dependent on sulfhydryl content

    Institute of Scientific and Technical Information of China (English)

    Runglawan Sudmoon; Nison Sattayasai; Wandee Bunyatratchata; Arunrat Chaveerach; Suporn Nuchadomrong

    2008-01-01

    A mannose-binding lectin was purified from Dendrobium (D.) findleyanum pseudobulb using mannan-agarose column chromatography. After heating in the presence of SDS with or without 2-mercaptoethanol on SDS-PAGE with a continuous gradient of 8%-20% acrylamide, the purified lectin showed only one protein band with a molecular mass of 14.5 kDa. Without heating, two bands were seen on the gel at the positions of 14.5 kDa and 53.7 kDa, but a higher amount of the 53.7 kDa protein was observed in the presence of 2- mercaptoethanol. Protein identification of both protein bands by liquid chromatography-tandem mass spectrometry showed three peptide fragments identical to parts of a lectin precursor from D. officinale; the lectin was named D. Findleyanum agglutinin (DFA). Using various concentrations of native- PAGE and Ferguson plot, only one protein band revealed a molecular mass of 56.2 kDa, indicating four 14.5 kDa polypeptide subunits in the DFA. Isoelectric focusing revealed that the DFA had three conformational forms with an isoelectric point of 5.18, 4.87 and 4.72, whereas 2-mercaptoethanoltreated DFA showed only one band with an isoelectric point of 5.18. DFA exhibited specificity towards mannose using the solid-phase method. The binding activity, anti-fungal activity and hemagglutination activity of DFA were not affected by heat, but were increased by free sulfhydryl groups.

  14. Sequential extraction results in improved proteome profiling of medicinal plant Pinellia ternata tubers, which contain large amounts of high-abundance proteins.

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    Xiaolin Wu

    Full Text Available Pinellia ternata tuber is one of the well-known Chinese traditional medicines. In order to understand the pharmacological properties of tuber proteins, it is necessary to perform proteome analysis of P. ternata tubers. However, a few high-abundance proteins (HAPs, mainly mannose-binding lectin (agglutinin, exist in aggregates of various sizes in the tubers and seriously interfere with proteome profiling by two-dimensional electrophoresis (2-DE. Therefore, selective depletion of these HAPs is a prerequisite for enhanced proteome analysis of P. ternata tubers. Based on differential protein solubility, we developed a novel protocol involving two sequential extractions for depletion of some HAPs and prefractionation of tuber proteins prior to 2-DE. The first extraction using 10% acetic acid selectively extracted acid-soluble HAPs and the second extraction using the SDS-containing buffer extracted remaining acid-insoluble proteins. After application of the protocol, 2-DE profiles of P. ternata tuber proteins were greatly improved and more protein spots were detected, especially low-abundance proteins. Moreover, the subunit composition of P. ternata lectin was analyzed by electrophoresis. Native lectin consists of two hydrogen-bonded subunits (11 kDa and 25 kDa and the 11 kDa subunit was a glycoprotein. Subsequently, major HAPs in the tubers were analyzed by mass spectrometry, with nine protein spots being identified as lectin isoforms. The methodology was easy to perform and required no specialized apparatus. It would be useful for proteome analysis of other tuber plants of Araceae.

  15. ALS1 and ALS3 gene expression and biofilm formation in Candida albicans isolated from vulvovaginal candidiasis

    Science.gov (United States)

    Roudbarmohammadi, Shahla; Roudbary, Maryam; Bakhshi, Bita; Katiraee, Farzad; Mohammadi, Rasoul; Falahati, Mehraban

    2016-01-01

    Background: A cluster of genes are involved in the pathogenesis and adhesion of Candida albicans to mucosa and epithelial cells in the vagina, the important of which is agglutinin-like sequence (ALS) genes. As well as vaginitis is a significant health problem among women, the antifungal resistance of Candida species is continually increasing. This cross-sectional study investigates the expression of ALS1 and ALS3 genes and biofilm formation in C. albicans isolate isolated from vaginitis. Materials and Methods: Fifty-three recognized isolates of C. albicans were collected from women with recurrent vulvovaginal candidiasis in Iran, cultured on sabouraud dextrose agar, and then examined for gene expression. Total messenger RNA (mRNA) extracted from C. albicans isolates and complementary DNA synthesized using reverse transcriptase enzyme. Reverse transcription-polymerase chain reaction (RT-PCR) using specific primer was used to evaluate the expression of ALS1 and ALS3 through housekeeping (ACT1) genes. 3-(4,5-dimethyl-2-thiazyl)-2,5-diphenyl-2H-tetrazolium bromide assay was performed to assess adherence capacity and biofilm formation in the isolated. Results: Forty isolates (75.8%) expressed ALS1 and 41 isolates (77.7%) expressed ALS3 gene. Moreover, 39 isolates (74%) were positive for both ALS1 and ALS3 mRNA by the RT-PCR. Adherence capability in isolates with ALS1 or ALS3 genes expression was greater than the control group (with any gene expression), besides, it was significantly for the most in the isolates that expressed both ALS1 and ALS3 genes simultaneously. Conclusion: The results attained indicated that there is an association between the expression of ALS1 and ALS3 genes and fluconazole resistance in C. albicans. A considerable percent of the isolates expressing the ALS1 and ALS3 genes may have contributed to their adherence to vagina and biofilm formation. PMID:27376044

  16. The glycosylation pattern of secretory granules in binucleate trophoblast cells is highly conserved in ruminants.

    Science.gov (United States)

    Klisch, K; Wooding, F B P; Jones, C J P

    2010-01-01

    The binucleate trophoblast cells (BNCs) in the ruminant placenta are a unique feature of this taxon. These cells produce several secretory proteins and transfer these across the fetomaternal barrier into the dam. We used lectin histochemistry with a panel of 24 lectins to characterise the glycosylation pattern of BNC secretory granules in a variety of ruminants. Seven species out of three ruminant families were thus investigated: greater malayan chevrotain (Tragulidae); fallow deer, red deer, chinese water deer (Cervidae); and domestic goat, springbok, impala (Bovidae). BNC granules in all species studied strongly expressed tri-/tetraantennary complex N-glycans and bisecting N-acetylglucosamine [GlcNAc] as shown by binding of leuco- and erythroagglutins of Phaseolus vulgaris respectively. The presence of terminal N-acetylgalactosamine [GalNAc]) in BNC granules is shown by intense staining with lectins from Dolichos biflorus, Vicia villosa and Wisteria floribunda. Terminal galactose or GalNAc was also present, bound by Glycine max agglutinin. Treatment of slides with neuraminidase strongly intensified staining of Erythrina cristagalli lectin (ECA) to terminal lactosamine in all species studied; this was otherwise absent except in goat. Sambucus nigra-1 lectin bound to BNC granules in all species except in Impala, indicating the presence of abundant alpha2,6 linked sialic acid. These results indicate that these unusual highly branched glycans, with bisecting GlcNAc and terminal GalNAc are a general feature of BNC granules in Ruminants, including the most basal Tragulid branch. It therefore appears that the specific glycosylation pattern of BNC granules evolved early in ruminant phylogenesis, together with the appearance of BNC. The conserved glycan structure in BNC secretory granules indicates that this pattern of glycosylation is likely to be of considerable functional importance for the secretory glycoproteins of ruminant BNC. PMID:19959226

  17. Overexpression of Galnt3 in Chondrocytes Resulted in Dwarfism Due to the Increase of Mucin-type O-Glycans and Reduction of Glycosaminoglycans*

    Science.gov (United States)

    Yoshida, Carolina Andrea; Kawane, Tetsuya; Moriishi, Takeshi; Purushothaman, Anurag; Miyazaki, Toshihiro; Komori, Hisato; Mori, Masako; Qin, Xin; Hashimoto, Ayako; Sugahara, Kazuyuki; Yamana, Kei; Takada, Kenji; Komori, Toshihisa

    2014-01-01

    Galnt3, UDP-N-acetyl-α-d-galactosamine:polypeptide N-acetylgalactosaminyltransferase 3, transfers N-acetyl-d-galactosamine to serine and threonine residues, initiating mucin type O-glycosylation of proteins. We searched the target genes of Runx2, which is an essential transcription factor for chondrocyte maturation, in chondrocytes and found that Galnt3 expression was up-regulated by Runx2 and severely reduced in Runx2−/− cartilaginous skeletons. To investigate the function of Galnt3 in chondrocytes, we generated Galnt3−/− mice and chondrocyte-specific Galnt3 transgenic mice under the control of the Col2a1 promoter-enhancer. Galnt3−/− mice showed a delay in endochondral ossification and shortened limbs at embryonic day 16.5, suggesting that Galnt3 is involved in chondrocyte maturation. Galnt3 transgenic mice presented dwarfism, the chondrocyte maturation was retarded, the cell cycle in chondrocytes was accelerated, premature chondrocyte apoptosis occurred, and the growth plates were disorganized. The binding of Vicia villosa agglutinin, which recognizes the Tn antigen (GalNAc-O-Ser/Thr), was drastically increased in chondrocytes, and aggrecan (Acan) was highly enriched with Tn antigen. However, safranin O staining, which recognizes glycosaminoglycans (GAGs), and Acan were severely reduced. Chondroitin sulfate was reduced in amount, but the elongation of chondroitin sulfate chains had not been severely disturbed in the isolated GAGs. These findings indicate that overexpression of Galnt3 in chondrocytes caused dwarfism due to the increase of mucin-type O-glycans and the reduction of GAGs, probably through competition with xylosyltransferases, which initiate GAG chains by attaching O-linked xylose to serine residues, suggesting a negative effect of Galnt family proteins on Acan deposition in addition to the positive effect of Galnt3 on chondrocyte maturation. PMID:25107907

  18. An improved lectin-based method for the detection of mucin-type O-glycans in biological samples.

    Science.gov (United States)

    Lee, Cheng-Siang; Muthusamy, Arivalagan; Abdul-Rahman, Puteri Shafinaz; Bhavanandan, Veer P; Hashim, Onn Haji

    2013-06-21

    Mucins and mucin-type glycoproteins, collectively referred to as mucin-type O-glycans, are implicated in many important biological functions and pathological conditions, including malignancy. Presently, there is no reliable method to measure the total mucin-type O-glycans of a sample, which may contain one or more of these macromolecules of unknown structures. We report the development of an improved microassay that is based on the binding of lectins to the unique and constant GalNAc-Ser/Thr structural feature of mucin-type O-glycans. Since the sugar-amino acid linkage in the mucin-type O-glycans is invariably cryptic, we first chemically removed the heterogeneous peripheral and core saccharides of model glycoconjugates before examining for their interactions using an enzyme-linked lectin assay (ELLA). Desialylation of the model glycoconjugates led to maximal binding of the lectins but additional treatments such as Smith degradation did not result in increased binding. Of the lectins tested for their ability to probe the desialylated O-glycans, jacalin showed the highest sensitivity followed by champedak galactose binding (CGB) lectin and Vicia villosa agglutinin. Further improvement in the sensitivity of ELLA was achieved by using microtiter plates that were pre-coated with the CGB lectin, which increased the specificity of the assay to mucin-type O-glycans. Finally, the applicability of the developed sandwich ELLA to crude samples was demonstrated by estimating trace quantities of the mucin-type O-glycans in the human serum. PMID:23665615

  19. O-glycosylation of protein subpopulations in alcohol-extracted rice proteins.

    Science.gov (United States)

    Kilcoyne, Michelle; Shah, Miti; Gerlach, Jared Q; Bhavanandan, Veer; Nagaraj, Vinay; Smith, Amy D; Fujiyama, Kazuhito; Sommer, Ulf; Costello, Catherine E; Olszewski, Neil; Joshi, Lokesh

    2009-02-15

    Mucin-type O-glycosylation has been well characterized in mammalian systems but not in plants. In this study, the purified alcohol-soluble, non-reduced protein (prolamin) fraction from rice seed was investigated for the occurrence of O-linked oligosaccharides. As storage prolamins are unlikely to be O-glycosylated, any O-glycosylation found was likely to belong to co-extracted proteins, whether because of association with the protein body or solubility. SDS-PAGE and MS analyses revealed 14 and 16kDa protein families in fractions that bound to the lectins peanut agglutinin (PNA), Vicia villosa lectin (VVL) and Jacalin, indicative of the presence of O-linked saccharides. Enzymatic cleavage, fluorescent labeling and high-performance liquid chromatography (HPLC) analysis demonstrated a peak consistent with Gal-beta-(1-->3)-GalNAc, with similar MS/MS fragmentation. Additionally, upon chemical analysis, a GlcNAc-containing O-linked carbohydrate moiety was discovered. Protein blotting with anti-O-GlcNAc antibody (clone CTD110.6) was positive in a subpopulation of the 14kDa alcohol-soluble protein fraction, but a hot capping experiment was negative. Therefore, the GlcNAc residue in this case is unlikely to be terminal. Additionally, a positive reaction with CTD110.6mAb cannot be taken as absolute proof of O-GlcNAc modification and further confirmatory experiments should be employed. We hypothesize that O-glycosylation may contribute to protein functionality or regulation. Further investigation is required to identify the specific proteins with these modifications. This 'reverse' approach could lead to the identification of proteins involved in mRNA targeting, signaling, translation, anchoring or maintenance of translational quiescence and may be applied to germinating rice seed extracts for further elucidation of protein function and regulation. PMID:18639953

  20. An array-based method to identify multivalent inhibitors.

    Science.gov (United States)

    Zhang, Yalong; Li, Qian; Rodriguez, Luis G; Gildersleeve, Jeffrey C

    2010-07-21

    Carbohydrate-protein interactions play a critical role in a variety of biological processes, and agonists/antagonists of these interactions are useful as biological probes and therapeutic agents. Most carbohydrate-binding proteins achieve tight binding through formation of a multivalent complex. Therefore, both ligand structure and presentation contribute to recognition. Since there are many potential combinations of structure, spacing, and orientation to consider and the optimal one cannot be predicted, high-throughput approaches for analyzing carbohydrate-protein interactions and designing inhibitors are appealing. In this report, we develop a strategy to vary neoglycoprotein density on a surface of a glycan array. This feature of presentation was combined with variations in glycan structure and glycan density to produce an array with approximately 600 combinations of glycan structure and presentation. The unique array platform allows one to distinguish between different types of multivalent complexes on the array surface. To illustrate the advantages of this format, it was used to rapidly identify multivalent probes for various lectins. The new array was first tested with several plant lectins, including concanavalin A (conA), Vicia villosa isolectin B4 (VVL-B(4)), and Ricinus communis agglutinin (RCA120). Next, it was used to rapidly identify potent multivalent inhibitors of Pseudomonas aeruginosa lectin I (PA-IL), a key protein involved in opportunistic infections of P. aeruginosa , and mouse macrophage galactose-type lectin (mMGL-2), a protein expressed on antigen presenting cells that may be useful as a vaccine targeting receptor. An advantage of the approach is that structural information about the lectin/receptor is not required to obtain a multivalent inhibitor/probe. PMID:20583754

  1. Overexpression of Galnt3 in chondrocytes resulted in dwarfism due to the increase of mucin-type O-glycans and reduction of glycosaminoglycans.

    Science.gov (United States)

    Yoshida, Carolina Andrea; Kawane, Tetsuya; Moriishi, Takeshi; Purushothaman, Anurag; Miyazaki, Toshihiro; Komori, Hisato; Mori, Masako; Qin, Xin; Hashimoto, Ayako; Sugahara, Kazuyuki; Yamana, Kei; Takada, Kenji; Komori, Toshihisa

    2014-09-19

    Galnt3, UDP-N-acetyl-α-D-galactosamine:polypeptide N-acetylgalactosaminyltransferase 3, transfers N-acetyl-D-galactosamine to serine and threonine residues, initiating mucin type O-glycosylation of proteins. We searched the target genes of Runx2, which is an essential transcription factor for chondrocyte maturation, in chondrocytes and found that Galnt3 expression was up-regulated by Runx2 and severely reduced in Runx2(-/-) cartilaginous skeletons. To investigate the function of Galnt3 in chondrocytes, we generated Galnt3(-/-) mice and chondrocyte-specific Galnt3 transgenic mice under the control of the Col2a1 promoter-enhancer. Galnt3(-/-) mice showed a delay in endochondral ossification and shortened limbs at embryonic day 16.5, suggesting that Galnt3 is involved in chondrocyte maturation. Galnt3 transgenic mice presented dwarfism, the chondrocyte maturation was retarded, the cell cycle in chondrocytes was accelerated, premature chondrocyte apoptosis occurred, and the growth plates were disorganized. The binding of Vicia villosa agglutinin, which recognizes the Tn antigen (GalNAc-O-Ser/Thr), was drastically increased in chondrocytes, and aggrecan (Acan) was highly enriched with Tn antigen. However, safranin O staining, which recognizes glycosaminoglycans (GAGs), and Acan were severely reduced. Chondroitin sulfate was reduced in amount, but the elongation of chondroitin sulfate chains had not been severely disturbed in the isolated GAGs. These findings indicate that overexpression of Galnt3 in chondrocytes caused dwarfism due to the increase of mucin-type O-glycans and the reduction of GAGs, probably through competition with xylosyltransferases, which initiate GAG chains by attaching O-linked xylose to serine residues, suggesting a negative effect of Galnt family proteins on Acan deposition in addition to the positive effect of Galnt3 on chondrocyte maturation. PMID:25107907

  2. Specificity analysis of lectins and antibodies using remodeled glycoproteins.

    Science.gov (United States)

    Iskratsch, Thomas; Braun, Andreas; Paschinger, Katharina; Wilson, Iain B H

    2009-03-15

    Due to their ability to bind specifically to certain carbohydrate sequences, lectins are a frequently used tool in cytology, histology, and glycan analysis but also offer new options for drug targeting and drug delivery systems. For these and other potential applications, it is necessary to be certain as to the carbohydrate structures interacting with the lectin. Therefore, we used glycoproteins remodeled with glycosyltransferases and glycosidases for testing specificities of lectins from Aleuria aurantia (AAL), Erythrina cristagalli (ECL), Griffonia simplicifolia (GSL I-B(4)), Helix pomatia agglutinin (HPA), Lens culinaris (LCA), Lotus tetragonolobus (LTA), peanut (Arachis hypogaeae) (PNA), Ricinus communis (RCA I), Sambucus nigra (SNA), Vicia villosa (VVA), and wheat germ (Triticum vulgaris) (WGA) as well as reactivities of anti-carbohydrate antibodies (anti-bee venom, anti-horseradish peroxidase [anti-HRP], and anti-Lewis(x)). After enzymatic remodeling, the resulting neoglycoforms display defined carbohydrate sequences and can be used, when spotted on nitrocellulose or in enzyme-linked lectinosorbent assays, to identify the sugar moieties bound by the lectins. Transferrin with its two biantennary complex N-glycans was used as scaffold for gaining diverse N-glycosidic structures, whereas fetuin was modified using glycosidases to test the specificities of lectins toward both N- and O-glycans. In addition, alpha(1)-acid glycoprotein and Schistosoma mansoni egg extract were chosen as controls for lectin interactions with fucosylated glycans (Lewis(x) and core alpha1,3-fucose). Our data complement and expand the existing knowledge about the binding specificity of a range of commercially available lectins. PMID:19123999

  3. Pregnancy associated glycoprotein-1, -6, -7, and -17 are major products of bovine binucleate trophoblast giant cells at midpregnancy.

    Science.gov (United States)

    Klisch, Karl; De Sousa, Noelita Melo; Beckers, Jean-François; Leiser, Rudolf; Pich, Andreas

    2005-08-01

    Pregnancy associated glycoproteins (PAGs) are extensively glycosylated secretory proteins of ruminant trophoblast cells. In cattle placenta several PAG cDNAs are expressed, but the variety of correspondent proteins and their degree of glycosylation are not well characterized. Thus, we purified PAGs by using a protocol which included a lectin (Vicia villosa agglutinin) affinity chromatography. Due to their specific glycosylation pattern, PAGs derived from binucleate trophoblast giant cells were highly enriched by this protocol. PAGs were purified from cotyledons of 2 day 100 placentas and from a single placenta at day 155 and 180. In all samples three major bands (75; 66; 56 kDa) were detected by one-dimensional SDS-PAGE. Mass-spectrometric analysis identified the 75 kDa band as a mixture of PAG-7 and PAG-6, the 66 kDa band as PAG-1 and the 56 kDa band as PAG-17. N-terminal sequencing of the day 100 sample confirmed the mass spectrometric identifications. Enzymatic release of N-glycans with peptide-N-glycanase-F from PAGs reduced the molecular weight to approximately 37 kDa which corresponds to the theoretical molecular mass of PAGs. Limited peptide-N-glycanase-F treatment revealed that all four N-glycosylation sites are quantitatively occupied in PAG-1. Compared to PAG-1 the number of potential N-glycosylation sites is lower in PAG-17 (three sites) and higher in PAG-6 and -7 (five and six sites, respectively). This suggests that the number of attached N-glycans is the main determinant of molecular mass of bovine PAGs. The degree of glycosylation may be a major factor regulating the plasma half life of PAGs. PMID:15822115

  4. Levantamento soroepidemiológico de leptospirose em trabalhadores do serviço de saneamento ambiental em localidade urbana da região sul do Brasil Serological survey of leptospirosis among environmental sanitation workers in an urban locality in southern Brazil

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    Laerte Pereira de Almeida

    1994-02-01

    Full Text Available A pesquisa de aglutininas anti-Leptospira, pela técnica de soroaglutinação microscópica, em soros de trabalhadores dos serviços de águas, bueiros e galerias, esgotos, coleta de lixo e limpeza pública, do Município de Pelotas, RS, Brasil, revelou 10,4% de reagentes a um ou mais sorovares; não houve diferenças significantes entre as proporções de reagentes de cada um dos setores de trabalho. Foram identificados 12 sorovares diferentes; castelonis e australis, apesar de mais freqüentes, não apresentaram diferenças estatisticamente significantes com os demais. Constatou-se que 86,9% das amostras apresentavam títulos aglutinantes compreendidos entre 100 e 400; as proporções de soros com títulos iguais a 100 e 400 foram superiores às dos títulos 800, 1.600 e 3.200 (p Sera from 386 environmental sanitation workers, concerned with water supply, drains and drainage galleries, sewers, garbage collection and road sweepers, were examined for leptospiral agglutinins by the microscopic agglutination test. Altogether 40 of the 386 workers (10.4% were positive to one or more serovars; however, the difference in seropositivity between the professional categories was not significant (p > 0.05. Twelve serovars were recorded among the seropositive workers with predominance of L. castelonis and L. australis; but the difference between the serovars was not statistically significant (p > 0.05. Of the seropositive workers, 86.9% had agglutination titres > 100 and < 400; the rates for titres 100 and 400 were higher than 800, 1,600 and 3,200 (p < 0.05.

  5. Short-term intermittent administration of CXCR4 antagonist AMD3100 facilitates myocardial repair in experimental myocardial infarction

    Institute of Scientific and Technical Information of China (English)

    Yuechen Luo; Xiaoning Zhao; Xin Zhou; Wenjie Ji; Ling Zhang; Tao Luo; Hongrnei Liu

    2013-01-01

    The binding of the stromal cell-derived factor-1α (SDF-1α)to the cysteine (C)-X-C motif chemokine receptor 4 (CXCR4) has emerged as a key signal for stem and progenitor cells trafficking to the circulation from the bone marrow.Our aim was to investigate the role of daily intermittent administration of AMD3100 (a specific reversible CXCR4 receptor antagonist) during the healing process after myocardial infarction (MI).Wistar rats were subjected to MI and AMD3100 was injected intraperitoneally after surgery.SDF-1α mRNA expression was measured by real-time polymerase chain reaction.Histology changes were analyzed with immunofluorescence,Masson's trichrome staining,and wheat germ agglutinin.The number of leukocytes in peripheral blood was measured by complete blood cell count analysis.The activities of matrix metalloproteinase-2/9 (MMP-2/9) were determined by gelatin zymography.The expression level of SDF-1αmRNA in the infarcted tissue was enhanced rapidly (6 h),peaked at 24 h,and then declined to the normal level at 7days post-MI.AMD3100 further enhanced the increase of SDF-1α in infarct area.Increased leukocytes were observed in AMD3100-treated groups.The mobilization of c-kit+ stem/progenitor cells and enhanced neovascularization were augmented by AMD3100.Additionally,AMD3100 improved ventricular remodeling,which was revealed by the decrease of infarct size,viable cardiomyocyte cross-sectional area and left ventricle (LV) expansion index,and the increase of LV free wall thickness.The activities of MMP-2/9 were up-regulated by AMD3100.In conclusion,short-term intermittent administration of AMD3100 could accelerate the wound healing process in experimental MI and be a potential therapy for the treatment of MI.

  6. Reconstruction of atonic bladder innervation after spinal cord injury: A bladder reflex arc with afferent and efferent pathways.

    Science.gov (United States)

    He, Jun; Li, Guitao; Luo, Dixin; Sun, Hongtao; Qi, Yong; Li, Yiyi; Jin, Xunjie

    2015-11-01

    Background Establishing bladder reflex arcs only with the efferent pathway to induce micturition after spinal cord injury (SCI) has been successful. However, the absence of sensory function and micturition desires can lead to serious complications. Objectives To reconstruct a bladder reflex arc with both afferent and efferent pathways to achieve atonic bladder innervation after SCI. Methods A reflex arc was established by microanastomosis of the S2 dorsal root to the peripheral process of the L5 dorsal ganglion and the L5 ventral root to the S2 ventral root. The functions of the reflex arc were evaluated using electrophysiology, wheat germ agglutinin-horseradish peroxidase (WGA-HRP) tracing, and calcitonin gene-related peptide (CGRP) immunocytochemistry analysis. Hind-paw motion was evaluated by CatWalk gait. Results Compound action potentials and compound muscle action potentials were recorded at the right L5 dorsal root following electrical stimulation of right S2 dorsal root. Similar to the control side, these were not significantly different before or after the spinal cord destruction between L6 and S4. WGA-HRP tracing and CGRP immunocytochemistry showed that construction of the afferent and efferent pathways of the bladder reflex arc encouraged axonal regeneration of motor and sensory nerves, which then made contact with the anterior and posterior horns of the spinal cord, ultimately reestablishing axoplasmic transportation. Gait analysis showed that at 3 months following the operation, only the regularity index was significantly different as compared with 1 day before the operation, other parameters showing no difference. Conclusion Bladder reflex arc with the afferent and efferent pathways reconstructs the micturition function without great influence on the motion of leg. PMID:25582052

  7. Organization of sensory cortex in the East African hedgehog (Atelerix albiventris).

    Science.gov (United States)

    Catania, K C; Collins, C E; Kaas, J H

    2000-05-29

    We investigated the organization of neocortex in the East African hedgehog (Atelerix albiventris) with microelectrode recordings from sensory areas that were later correlated with cytochrome oxidase patterns in sections of flattened cortex. The location of corticospinal projecting neurons was also examined and related to sensory areas by making small injections of wheat germ agglutinin-horseradish peroxidase into the spinal cord. Our goals were to determine how hedgehog cortex is organized, how much sensory areas overlap, and to compare results with recent findings in other insectivores. Evidence was found for three separate topographically organized somatosensory areas, two visual areas, and a caudolateral auditory area. A medial somatosensory area corresponded to S1, the primary somatosensory area, whereas two lateral areas partially encircled auditory cortex and corresponded to the parietal ventral area (PV) and the secondary somatosensory area (S2). Primary visual cortex (V1) was delineated by a caudomedial cytochrome oxidase dark oval, and a more lateral visual area between V1 and somatosensory cortex corresponded to V2, or area 18. Two patches of corticospinal projecting cells were found primarily overlapping S1 and S2. Some bimodal auditory and somatosensory responses were found in parts of PV and S2, but for the most part, areas had relatively sharp histochemically apparent and physiologically defined borders. The present results indicate that the caudal neocortex of hedgehogs has only a few sensory areas, corresponding to those commonly found in several other small-brained mammals. Hedgehog cortical organization differs significantly in somatotopy, number, and position of fields from that of closely related shrews and moles. Thus, clear specializations occur, even within the order Insectivora. PMID:10813786

  8. CNTF induces regeneration of cone outer segments in a rat model of retinal degeneration.

    Directory of Open Access Journals (Sweden)

    Yiwen Li

    Full Text Available BACKGROUND: Cone photoreceptors are responsible for color and central vision. In the late stage of retinitis pigmentosa and in geographic atrophy associated with age-related macular degeneration, cone degeneration eventually causes loss of central vision. In the present work, we investigated cone degeneration secondary to rod loss in the S334ter-3 transgenic rats carrying the rhodopsin mutation S334ter. METHODOLOGY/PRINCIPAL FINDINGS: Recombinant human ciliary neurotrophic factor (CNTF was delivered by intravitreal injection to the left eye of an animal, and vehicle to the right eye. Eyes were harvested 10 days after injection. Cone outer segments (COS, and cell bodies were identified by staining with peanut agglutinin and cone arrestin antibodies in whole-mount retinas. For long-term treatment with CNTF, CNTF secreting microdevices were implanted into the left eyes at postnatal day (PD 20 and control devices into the right eyes. Cone ERG was recorded at PD 160 from implanted animals. Our results demonstrate that an early sign of cone degeneration is the loss of COS, which concentrated in many small areas throughout the retina and is progressive with age. Treatment with CNTF induces regeneration of COS and thus reverses the degeneration process in early stages of cone degeneration. Sustained delivery of CNTF prevents cones from degeneration and helps them to maintain COS and light-sensing function. CONCLUSIONS/SIGNIFICANCE: Loss of COS is an early sign of secondary cone degeneration whereas cell death occurs much later. At early stages, degenerating cones are capable of regenerating outer segments, indicating the reversal of the degenerative process. Sustained delivery of CNTF preserves cone cells and their function. Long-term treatment with CNTF starting at early stages of degeneration could be a viable strategy for preservation of central vision for patients with retinal degenerations.

  9. Lectin-carbohydrate recognition mechanism of Plasmodium berghei in the midgut of malaria vector Anopheles stephensi using quantum dot as a new approach.

    Science.gov (United States)

    Basseri, Hamid R; Javazm, Mahdi Salari; Farivar, Leila; Abai, Mohammad R

    2016-04-01

    Potential targets of Plasmodium ookinetes at the mosquito midgut walls were investigated in relation to interfering malarial transmission. In this study, the essential application of Quantum Dots (QDs) was used to examine the interaction between Plasmodium berghei ookinetes and the Anopheles stephensi midgut, based on lectin-carbohydrate recognition. Two significant lectins were utilized to determine this interaction. Two QDs, cadmium telluride (CdTe)/CdS and cadmium selenide (CdSe)/CdS, were employed in staining Plasmodium ookinete to study its interaction in the midgut of the mosquito vector in vivo. Concurrently, two lectins, wheat germ agglutinin (WGA) and concanavalin A (Con A), were inadvertently exploited to mask lectin binding sites between ookinetes and mosquito midgut cells. The numbers of ookinetes in both lumen and epithelial cells were eventually counted, following adequate preparation of wax sections extracted from whole midgut, and subsequent examination using a differential interference contrast a fluorescence microscopic technique. Interestingly, we detected that neither of the QDs mutated ookinete invasion into the midgut cells of the investigated mosquitoes. QD staining of ookinetes remained permanent despite the effective embedding procedure. The massive binding potency of ookinetes to midgut cells of the cross-examined mosquitoes undoubtedly revealed that Con A did not interrupt ookinete penetration into the midgut wall. In contrast, WGA inhibited ookinete invasion into the midgut cells. The results proved that QD nanoparticles are biocompatible, non-toxic to P. berghei and stable to photobleaching. The QDs staining, which was successfully implemented for ookinete labelling, is a simple and effective tool which plays a crucial role in bioimaging including the study of parasite-vector interactions. PMID:26772447

  10. Detection of nearest neighbors to specific fluorescently tagged ligands in rod outer segment and lymphocyte plasma membranes by photosensitization of 5-iodonaphthyl 1-azide

    International Nuclear Information System (INIS)

    Lima bean agglutinin-fluorescein 5-isothiocyanate conjugate (FluNCS-lima bean lectin) interacts with specific receptor molecules on membranes both from the rod outer segment (ROS) of the frog retina and from S49 mouse lymphoma cells. When [125I]-5-iodonaphthyl 1-azide (125I-INA), which freely and randomly partitions into the lipid bilayer, is added to membranes and the suspension is irradiated at 480 nm, the FluNCS-conjugated lectin photosensitizes the [125I]INA but only at discrete sites. This results in the selective labeling of specific proteins: an 88-kDa protein on ROS membranes and a 56-kDa protein on S49 plasma membranes. Labeling is dependent upon the interaction of the FluNCS-lectin with glycosylated receptor sites, since N-acetylgalactosamine, but not methyl alpha-mannoside, blocked labeling of the 56-kDa protein on S49 membranes. In contrast, a random labeling pattern of membrane proteins was observed upon irradiation at 480 nm using other fluorescein conjugates, such as FluNCS-bovine serum albumin (FluNCS-BSA) or FluNCS-soybean trypsin inhibitor (FluNCS-STI), which interact with cell membranes in a nonselective manner, or with N-(fluorescein-5-thiocarbamoyl)-n-undecyclamine (FluNCS-NHC11), which is freely miscible in the membrane lipid. Random labeling was also obtained by direct photoexcitation of [125I]INA at 314 nm, with no distinct labeling of the 88- and 56-kDa proteins in the respective membranes. These results suggest that protein ligands can be used to guide sensitizers to discrete receptor sites and lead to their selective labeling by photosensitized activation of [125I]INA

  11. Origin of primary sensory neurons innervating the buccal stretch receptor.

    Science.gov (United States)

    Yamamoto, T; Onozuka, M; Nagasaki, S; Watanabe, K; Ozono, S

    1999-01-01

    The primary sensory neurons innervating mechanoreceptors in oro-facial regions have their cell bodies in either the trigeminal ganglion or the mesencephalic nucleus of the trigeminal nerve. The buccal stretch receptor (BSR), a type of mechanoreceptor in the jaw of rodents, has recently been recognized as signaling the position of the mandible. The location of the primary afferent neurons innervating this receptor is unknown. To investigate the cell bodies of the BSR afferent neurons in rats, we applied wheat germ agglutinin-horseradish peroxidase (WGA-HRP) to the proximal stump of the severed nerve branch of the buccal nerve that supplied the BSR. HRP-labeled cell bodies were observed in the posterolateral portion of the ipsilateral trigeminal ganglion. None was found in the contralateral trigeminal ganglion or in the brainstem. All labeled cell bodies were oval or round and closely resembled pseudo-unipolar neurons. The mean diameter of the labeled somata ranged between 25.5 and 52.5 microm, with small ( or = 41 microm) accounting for 8.8%, 54.9%, and 36.3%, respectively. Among the myelinated nerve fibers in the branch in which WGA-HRP was applied, 78.5% terminated in the BSR and had larger fiber diameters than the rest, indicating that most of the medium and large HRP-labeled cell bodies were BSR afferents. From these results and the ontogenetic origin of this receptor, it is suggested that the BSR differentiated from the mechanoreceptors in the oral mucosa or the fascia of masticatory muscles. PMID:10065945

  12. Radiologic and clinical study of mycoplasam pneumonia in children

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    Baik, Seung Kug; Yoon, Young Woon; Yang, Dae Dong; Lee, Jong Yul; Choi, Hang Yong; Kim, Bong Ki [College of Medicine, Wallace Memorial Baptist Hospital, Pusan (Korea, Republic of)

    1990-10-15

    We studied 143 cases of serologically proven (cold agglutinin or 1HA test) M. Pneumonia in children below 15 year of age who admitted to Wallace Memorial Baptist Hospital between March 1986 and August 1988. The following results were noted : 1. Radiologic findings {center_dot} The patterns of lung infiltration were bronchopneumonic in 52.6%, interstitial in 24.5%, lobar in 3.5%, mixed in 1.3%, and normal or slightly increased bronchovascular markings in 18.1%. {center_dot} The degree of pulmonary involvement were minimal in 49.2%, moderate in 24.5% and extensive in 8.3%. {center_dot} The distribution of infiltration in the minimal and moderate pulmonary involvement were upper lobe in 28.6%, middle lobe in 17.2% and lower lobe in 54.3%. {center_dot} Other findings were bilateral involvement in 31.5%, pleural involvement in 15.8%, new area of consolidation in 3.5% and hilar lymph node involvement in 34.2%. 2. The most common duration of the period between the onset of symptom and confirmation of the diagnosis was 7-14 days. 3. The sex distribution was 1.3 : 1 in male to female ratio. 4. The most peak age incidence was seen in 10-13, and the second one in 4 - 7 year. 5. As for the concomitant illness and complication, asthmatic bronchitis was most common (23 cases, 16.1%) followed by pharyngitis, sinusitis, skin rash and gastroenteritis, etc.

  13. Separate domains of the insulin receptor contain sites of autophosphorylation and tyrosine kinase activity

    International Nuclear Information System (INIS)

    The authors have studied the structure and function of the solubilized insulin receptor before and after partial proteolytic digestion to define domains in the β-subunit that undergo autophosphorylation and contain the tyrosine kinase activity. Wheat germ agglutinin purified insulin receptor from Fao cells was digested briefly at 220C with low concentrations of trypsin, staphylococcal V8 protease, or elastase. Autophosphorylation of the β-subunit was carried out before and after digestion, and the [32P]phosphoproteins were separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, detected by autoradiography, and analyzed by tryptic peptide mapping by use of reverse-phase high-performance liquid chromatography. The 85-kDa fragment was not immunoprecipitated by an antibody directed against the C-terminal domain of the β-subunit (αPep-1), indicating that this region of the receptor was lost. The 85-kDa fragment contained about half of the [32P]phosphate originally found in the β-subunit, and tryptic peptide mapping showed that two major tryptic phosphopeptides (previously called pY2 and pY3) were removed. Three other tryptic phosphopeptides (pY1, pY1a, and pY4) were found in the 85- and 70-kDa fragments. To determined the structural requirements for kinase activity, the insulin receptor was subjected to tryptic digestion for 30 s-30 min, such that the receptor was composed exclusively of 85- and 70-kDa fragments of the β-subunit. The 85-kDa fragment exhibited autophosphorylation at pY1, pY1a, and pY4. Both the 85- and 70-kDa fragments phosphorylated tyrosine residues in a synthetic decapeptide that has the sequence of the C-terminal domain of the β-subunit of human insulin rare in the receptor

  14. Photoaffinity labeling of insulin receptors in viable cultured human lymphocytes. Demonstration of receptor shedding and degradation

    International Nuclear Information System (INIS)

    A photosensitive derivative of radiolabeled insulin, SANAH-125I-insulin, was prepared by reacting N-succinimidyl-6-(4'-azido-2'-nitrophenylamino) hexanoate (SANAH) with 125I-insulin. Cultured IM-9 cells were incubated with SANAH-125I-insulin at 16 degrees C in the dark. They were then washed, photolyzed, solubilized, and analyzed by SDS-polyacrylamide gel electrophoresis and autoradiography. Under disulfide reducing conditions, a single specific band of Mr 125,000 was obtained. The characteristics of the labeling of this band with SANAH-125I-insulin (specificity, time course, concentration effect) were the same as that of 125I-insulin interaction with the IM-9 cells and the labeling process did not affect cell viability. The solubilized photolabeled insulin receptor fraction was enriched by first adsorbing to agarose-bound wheat germ agglutinin and the material eluted with N-acetyl-D-glucosamine was then analyzed by SDS-PAGE and autoradiography. Under nonreducing conditions, a major receptor band of Mr 320 K and a minor band of 280 K were obtained. Upon disulfide bond reduction with increasing concentrations of dithiothreitol, a major band of Mr 125 K and two minor bands of Mr 210 K and 94 K were seen. When cells photolabeled at 16 degrees C were further incubated at 37 degrees C, there was a time-dependent loss of intact receptors into the incubation buffer. In contrast, no similar shedding of labeled receptors was observed from isolated rat adipocytes. Following shedding, the labeled IM-9 insulin receptors rapidly disappeared from the incubation buffer (half-time approximately 1.5 h). These results demonstrate the feasibility of photoaffinity labeling, characterizing, and following the fate of insulin receptor in viable cells. Thus receptor photoaffinity labeling should provide a suitable approach for studies of the biologic fate of insulin receptors in cells that are targets for insulin action

  15. Display of phytase on the cell surface of Saccharomyces cerevisiae to degrade phytate phosphorus and improve bioethanol production.

    Science.gov (United States)

    Chen, Xianzhong; Xiao, Yan; Shen, Wei; Govender, Algasan; Zhang, Liang; Fan, You; Wang, Zhengxiang

    2016-03-01

    Currently, development of biofuels as an alternative fuel has gained much attention due to resource and environmental challenges. Bioethanol is one of most important and dominant biofuels, and production using corn or cassava as raw materials has become a prominent technology. However, phytate contained in the raw material not only decreases the efficiency of ethanol production, but also leads to an increase in the discharge of phosphorus, thus impacting on the environment. In this study, to decrease phytate and its phosphorus content in an ethanol fermentation process, Saccharomyces cerevisiae was engineered through a surface-displaying system utilizing the C-terminal half of the yeast α-agglutinin protein. The recombinant yeast strain, PHY, was constructed by successfully displaying phytase on the surface of cells, and enzyme activity reached 6.4 U/g wet biomass weight. Ethanol productions using various strains were compared, and the results demonstrated that the specific growth rate and average fermentation rate of the PHY strain were higher 20 and 18 %, respectively, compared to the control strain S. cerevisiae CICIMY0086, in a 5-L bioreactor process by simultaneous saccharification and fermentation. More importantly, the phytate phosphorus concentration decreased by 89.8 % and free phosphorus concentration increased by 142.9 % in dry vinasse compared to the control in a 5-L bioreactor. In summary, we constructed a recombinant S. cerevisiae strain displaying phytase on the cell surface, which could improve ethanol production performance and effectively reduce the discharge of phosphorus. The strain reported here represents a useful novel engineering platform for developing an environment-friendly system for bioethanol production from a corn substrate. PMID:26610799

  16. Baculovirus Coinfection Strategy for Improved Galactosylation of Recombinant Glycoprotein Produced by Insect Cell Culture

    Science.gov (United States)

    Ney, Yap Wei; Rahman, Badarulhisam Abdul; Aziz, Azila Abdul

    Baculovirus Expression Vector System (BEVS) is widely used for the production of recombinant glycoproteins, but it is not ideal for pharmaceutical glycoprotein production due to incomplete glycosylation. The factors that ensure successful glycosylation are the presence of sufficient amount of glycosyltransferases, sugar nucleotides as the substrate donor and the recombinant protein as the substrate acceptor. In this study, we analyzed the galactosylation process by the introduction of ß-1,4galactosyltransferase (ß-1,4GalT) as the glycosyltransferase of interest and uridine-5`-diphosphogalactose (UDP-Gal) as the substrate donor. Recombinant human transferrin (rhTf) as a model protein was used as the substrate acceptor. Insect cell lines have been reported to produce a small amount of ß-1,4GalT and thus insufficient for effective galactosylation. In this study, we developed a method to produce galactosylated rhTf and optimized the expression of rhTf with better N-glycan quality. Recombinant ß-1,4GalT was introduced during protein expression by the coinfection of the BEVS with baculovirus carrying bovine ß-1,4GalT. To evaluate the extent of galactosylation by the coinfection strategy, a binding assay was established. In this binding assay, glycoprotein acceptor was absorbed onto ELISA plate surface. A lectin known as Ricinus communis agglutinin-I (RCA-I) labeled with peroxidase, was added and allowed to recognize Gal ß1>4GlcNAc group on the N-glycan of the glycoprotein, followed by appropriate color reaction measurements. Coexpression between rhTf and ß-1,4GalT did not show encouraging results due to the reduction of UDP-Gal upon baculovirus infection. This interesting finding suggested that the introduction of ß-1,4GalT alone was not sufficient for successful galactosylation. Alternatively, post harvest glycosylation method strategy seems to be a promising technique in the improvement of glycoprotein quality.

  17. In vitro capacitation and acrosome reaction in sperm of the phyllostomid bat Artibeus jamaicensis.

    Science.gov (United States)

    Álvarez-Guerrero, Alma; González-Díaz, Francisco; Medrano, Alfredo; Moreno-Mendoza, Norma

    2016-04-01

    Sperm capacitation occurs during the passage of sperm through the female reproductive tract. Once the sperm binds to the pellucid zone, the acrosome reaction to enable penetration of the oocyte is completed. In this study, sperm of Artibeus jamaicensis bat was used to evaluate both capacitation status and the acrosome reaction under in vitro conditions, incubating sperm at 32 and 37°C with and without progesterone. Sperm was incubated at different times to assess sperm cells' functionality in terms of capacitation and acrosome reaction, using the chlortetracycline staining, lectin fluoresceinisocyanate conjugate-Pisum sativum agglutinin (FITC-PSA), and transmission electron microscopy. Sperm cells that presented uniform fluorescence throughout the head and mid-piece were classified as non-capacitated. Subsequently, sperm cells, which were observed with fluorescence only in the anterior portion of the head and mid-piece, were classified as capacitated. Sperm cells with no fluorescence in the head, but fluorescence in the mid-piece, were categorized as sperm cells that have carried out the acrosome reaction. During the acrosome reaction, sperm cells showed changes in their morphology, so it was not possible to distinguish the plasma and acrosomal membranes. Around the entire head, it was not possible to distinguish the fusion points between these membranes that made it possible for the acrosomal reaction to take place and thus to release the enzymes necessary to penetrate the pellucid zone. In conclusion, under appropriate in vitro conditions and by supplementing the culture medium with progesterone, A. jamaicensis bat sperm cells are able to be capacitated in a period from 6 to 8 h and to carry out the acrosome reaction. PMID:26744028

  18. DBA-Lectin Reactivity Defines Mouse Uterine Natural Killer Cell Subsets with Biased Gene Expression 1

    Science.gov (United States)

    Chen, Zhilin; Zhang, Jianhong; Hatta, Kota; Lima, Patricia D. A.; Yadi, Hakim; Colucci, Francesco; Yamada, Aureo T.; Croy, B. Anne

    2012-01-01

    Endometrial decidualization, a process essential for blastocyst implantation in species with hemochorial placentation, is accompanied by an enormous but transient influx of Natural Killer (NK) cells. Mouse uterine (u)NK cell subsets have been defined by diameter and cytoplasmic granule number, reflecting stage of maturity and by histochemical reactivity with Periodic Acid Schiff’s (PAS) reagent, with or without co-reactivity with Dolichos biflorus agglutinin (DBA) lectin. We asked whether DBA− and DBA+ mouse uNK cells were equivalent using quantitative (q)RT-PCR analyses of flow separated, midpregnancy (gestation day (gd)10) cells and using immunohistochemistry. CD3E (CD3)-IL2RB (CD122)+DBA− cells were identified as the dominant Ifng transcript source. Skewed IFNG production by uNK cell subsets was confirmed by analysis of uNK cells from eYFP-tagged IFNG-reporter mice. In contrast, CD3E-IL2RB+DBA+ uNK cells expressed genes compatible with significantly greater potential for IL22 synthesis, angiogenesis and participation in regulation mediated by the renin-angiotensin system (RAS). CD3E-IL2RB+DBA+ cells were further divided into VEGFA+ and VEGFA− subsets. CD3E-IL2RB+DBA+ uNK cells but not CD3E-IL2RB+DBA− uNK cells arose from circulating, bone marrow-derived progenitor cells by gd6. These findings indicate the heterogeneous nature of mouse uNK cells and suggest that studies using only DBA + uNK cells will give biased data that does not fully represent the uNK cell population. PMID:22875907

  19. Streptococcus salivarius Fimbriae Are Composed of a Glycoprotein Containing a Repeated Motif Assembled into a Filamentous Nondissociable Structure

    Science.gov (United States)

    Lévesque, Céline; Vadeboncoeur, Christian; Chandad, Fatiha; Frenette, Michel

    2001-01-01

    Streptococcus salivarius, a gram-positive bacterium found in the human oral cavity, expresses flexible peritrichous fimbriae. In this paper, we report purification and partial characterization of S. salivarius fimbriae. Fimbriae were extracted by shearing the cell surface of hyperfimbriated mutant A37 (a spontaneous mutant of S. salivarius ATCC 25975) with glass beads. Preliminary experiments showed that S. salivarius fimbriae did not dissociate when they were incubated at 100°C in the presence of sodium dodecyl sulfate. This characteristic was used to separate them from other cell surface components by successive gel filtration chromatography procedures. Fimbriae with molecular masses ranging from 20 × 106 to 40 × 106 Da were purified. Examination of purified fimbriae by electron microscopy revealed the presence of filamentous structures up to 1 μm long and 3 to 4 nm in diameter. Biochemical studies of purified fimbriae and an amino acid sequence analysis of a fimbrial internal peptide revealed that S. salivarius fimbriae were composed of a glycoprotein assembled into a filamentous structure resistant to dissociation. The internal amino acid sequence was composed of a repeated motif of two amino acids alternating with two modified residues: A/X/T-E-Q-M/φ, where X represents a modified amino acid residue and φ represents a blank cycle. Immunolocalization experiments also revealed that the fimbriae were associated with a wheat germ agglutinin-reactive carbohydrate. Immunolabeling experiments with antifimbria polyclonal antibodies showed that antigenically related fimbria-like structures were expressed in two other human oral streptococcal species, Streptococcus mitis and Streptococcus constellatus. PMID:11292790

  20. Expression analysis of carbohydrate antigens in ductal carcinoma in situ of the breast by lectin histochemistry

    Directory of Open Access Journals (Sweden)

    Kieber-Emmons Thomas

    2008-05-01

    Full Text Available Abstract Background The number of breast cancer patients diagnosed with ductal carcinoma in situ (DCIS continues to grow. Laboratory and clinical data indicate that DCIS can progress to invasive disease. Carbohydrate-mediated cell-cell adhesion and tumor-stroma interaction play crucial roles in tumorigenesis and tumor aggressive behavior. Breast carcinogenesis may reflect quantitative as well as qualitative changes in oligosaccharide expression, which may provide a useful tool for early detection of breast cancer. Because tumor-associated carbohydrate antigens (TACA are implicated in tumor invasion and metastasis, the purpose of this study was to assess the expression of selected TACA by lectin histochemistry on DCIS specimens from the archival breast cancer tissue array bank of the University of Arkansas for Medical Sciences. Methods For detection of TACA expression, specimens were stained with Griffonia simplicifolia lectin-I (GS-I and Vicia vilosa agglutinin (VVA. We studied associations of lectin reactivity with established prognostic factors, such as tumor size, tumor nuclear grade, and expression of Her-2/neu, p53 mutant and estrogen and progesterone receptors. Results We observed that both lectins showed significant associations with nuclear grade of DCIS. DCIS specimens with nuclear grades II and III showed significantly more intense reactivity than DCIS cases with nuclear grade I to GS-1 (Mean-score chi-square = 17.60, DF = 2; P = 0.0002 and VVA (Mean-score chi-square = 15.72, DF = 2; P = 0.0004. Conclusion The results suggest that the expression of VVA- and GS-I-reactive carbohydrate antigens may contribute to forming higher grade DCIS and increase the recurrence risk.