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Sample records for affinity ige receptor

  1. Structural Changes in the Lectin Domain of CD23, the Low-Affinity IgE Receptor, upon Calcium Binding

    Energy Technology Data Exchange (ETDEWEB)

    Wurzburg, Beth A.; Tarchevskaya, Svetlana S.; Jardetzky, Theodore S. (NWU)

    2010-03-08

    CD23, the low-affinity receptor for IgE (Fc{var_epsilon}RII), regulates IgE synthesis and also mediates IgE-dependent antigen transport and processing. CD23 is a unique Fc receptor belonging to the C-type lectin-like domain superfamily and binds IgE in an unusual, non-lectin-like manner, requiring calcium but not carbohydrate. We have solved the high-resolution crystal structures of the human CD23 lectin domain in the presence and absence of Ca{sup 2+}. The crystal structures differ significantly from a previously determined NMR structure and show that calcium binding occurs at the principal binding site, but not at an auxiliary site that appears to be absent in human CD23. Conformational differences between the apo and Ca{sup 2+} bound structures suggest how IgE-Fc binding can be both calcium-dependent and carbohydrate-independent.

  2. Transmembrane adaptor proteins in the high-affinity IgE receptor signaling

    Czech Academy of Sciences Publication Activity Database

    Dráber, Petr; Hálová, Ivana; Levi-Schaffer, F.; Dráberová, Lubica

    2012-01-01

    Roč. 2, 11.1. (2012), s. 95. ISSN 1664-3224 R&D Projects: GA MŠk 1M0506; GA ČR GA301/09/1826; GA ČR GAP302/10/1759; GA AV ČR KAN200520701 Grant ostatní: AV ČR(CZ) M200520901 Institutional research plan: CEZ:AV0Z50520514 Institutional support: RVO:68378050 Keywords : IgE receptor * LAT/LAT1 * LAX * NTAL/Lab/LAT2 * PAG/Cbp * mast cells * plasma membrane * transmembrane adaptor proteins Subject RIV: EB - Genetics ; Molecular Biology

  3. Are basophil histamine release and high affinity IgE receptor expression involved in asymptomatic skin sensitization?

    DEFF Research Database (Denmark)

    Jensen, Bettina Margrethe; Assing, K; Jensen, Lone Hummelshøj; Glue, C; Skov, P S; Poulsen, L K

    2006-01-01

    . However, a relationship between the AS status and FcepsilonRI has not been investigated. We aimed to characterize basophils from AS by looking at histamine release (HR) (sensitivity and reactivity) and the FcepsilonRI molecule, and compare it with nonatopic (NA) or allergic (A) persons.......Immunoglobulin (Ig)E-sensitized persons with positive skin prick test, but no allergy symptoms, are classified as being asymptomatic skin sensitized (AS). The allergic type 1 disease is dependant on IgE binding to the high affinity IgE-receptor (FcepsilonRI) expressed on basophils and mast cells...

  4. Development of an in vitro model system for studying the interaction of Equus caballus IgE with its high-affinity receptor FcɛRI.

    Science.gov (United States)

    Sabban, Sari; Ye, Hongtu; Helm, Birgit

    2013-05-15

    The binding of immunoglobulin E (IgE) to its high-affinity receptor (FcɛRI) is the central protein interaction in IgE-mediated allergic reactions. The cross-linking of the IgE/FcɛRI complex, through cognate allergens, on the surface of mast cells and basophil cells results in mediator release, and thus leads to the symptoms of type I hypersensitivity responses in mammals. To develop a baseline value for subsequent equine anti-allergy drug and vaccine research, the interaction of equine IgE with its high-affinity FcɛRI receptor was investigated following the cloning and expression of equine IgE with specificity for NIP-HSA (4-hydroxy-5-iodo-3-nitrophenylacetic acid conjugated to human serum albumin). Receptor recognition and effector functions were assessed in Rat Basophil Leukemia (RBL-2H3.1) cells transfected with the α chain of equine and canine FcɛRI. Results obtained showed that the equine FcɛRI receptor recognizes both equine and canine IgE and supports similar β-hexosaminidase release levels from RBL cells transfected with equine FcɛRI, peaking at 36.68% at 100ngml(-1) antigen and 32.00% at 100ngml(-1) antigen respectively. Furthermore, the binding kinetics of the equine IgE to the equine FcɛRI receptor and the canine IgE to the same receptor was measured to be KA=6.33×10(9)M(-1) and KA=1.84×10(9)M(-1) respectively. This research established basic reagents and vitro assay systems to underpin the development of rational therapeutic intervention strategies to combat equine allergic manifestations. PMID:23485176

  5. Development of an in vitro model system for studying the interaction of Equus caballus IgE with its high-affinity receptor FcεRI.

    Science.gov (United States)

    Sabban, Sari; Ye, Hongtu; Helm, Birgit

    2014-01-01

    The interaction of IgE with its high-affinity Fc receptor (FcεRI) followed by an antigenic challenge is the principal pathway in IgE mediated allergic reactions. As a consequence of the high affinity binding between IgE and FcεRI, along with the continuous production of IgE by B cells, allergies usually persist throughout life, with currently no permanent cure available. Horses, especially race horses, which are commonly inbred, are a species of mammals that are very prone to the development of hypersensitivity responses, which can seriously affect their performance. Physiological responses to allergic sensitization in horses mirror that observed in humans and dogs. In this paper we describe the development of an in situ assay system for the quantitative assessment of the release of mediators of the allergic response pertaining to the equine system. To this end, the gene encoding equine FcεRIα was transfected into and expressed onto the surface of parental Rat Basophil Leukemia (RBL-2H3.1) cells. The gene product of the transfected equine α-chain formed a functional receptor complex with the endogenous rat β- and γ-chains. The resultant assay system facilitated an assessment of the quantity of mediator secreted from equine FcεRIα transfected RBL-2H3.1 cells following sensitization with equine IgE and antigenic challenge using β-hexosaminidase release as a readout. Mediator release peaked at 36.68% ± 4.88% at 100 ng ml(-1) of antigen. This assay was modified from previous assays used to study human and canine allergic responses. We have also shown that this type of assay system has multiple applications for the development of diagnostic tools and the safety assessment of potential therapeutic intervention strategies in allergic disease. PMID:25406512

  6. Proinflammatory and Th2 cytokines regulate the high affinity IgE receptor (FcepsilonRI and IgE-dependant activation of human airway smooth muscle cells.

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    Naresh Singh Redhu

    Full Text Available BACKGROUND: The high affinity IgE receptor (FcepsilonRI is a crucial structure for IgE-mediated allergic reactions. We have previously demonstrated that human airway smooth muscle (ASM cells express the tetrameric (alphabetagamma2 FcepsilonRI, and its activation leads to marked transient increases in intracellular Ca(2+ concentration, release of Th-2 cytokines and eotaxin-1/CCL11. Therefore, it was of utmost importance to delineate the factors regulating the expression of FcepsilonRI in human (ASM cells. METHODOLOGY/PRINCIPAL FINDINGS: Incubation of human bronchial and tracheal smooth muscle (B/TSM cells with TNF-alpha, IL-1beta or IL-4 resulted in a significant increase in FcepsilonRI-alpha chain mRNA expression (p<0.05; and TNF-alpha, IL-4 enhanced the FcepsilonRI-alpha protein expression compared to the unstimulated control at 24, 72 hrs after stimulation. Interestingly, among all other cytokines, only TNF-alpha upregulated the FcepsilonRI-gamma mRNA expression. FcepsilonRI-gamma protein expression remained unchanged despite the nature of stimulation. Of note, as a functional consequence of FcepsilonRI upregulation, TNF-alpha pre-sensitization of B/TSM potentially augmented the CC (eotaxin-1/CCL11 and RANTES/CCL5, but not TARC/CCL17 and CXC (IL-8/CXCL8, IP-10/CXCL10 chemokines release following IgE stimulation (p<0.05, n = 3. Furthermore, IgE sensitization of B/TSM cells significantly enhanced the transcription of selective CC and CXC chemokines at promoter level compared to control, which was abolished by Lentivirus-mediated silencing of Syk expression. CONCLUSIONS/SIGNIFICANCE: Our data depict a critical role of B/TSM in allergic airway inflammation via potentially novel mechanisms involving proinflammatory, Th2 cytokines and IgE/FcepsilonRI complex.

  7. Fc receptor-γ subunits with both polar or non-polar amino acids at position of T22 are capable of restoring surface expression of the high-affinity IgE receptor and degranulation in γ subunit-deficient rat basophilic leukemia cells

    Czech Academy of Sciences Publication Activity Database

    Rashid, A.; Housden, J.E.; Helm, B.A.; Dráber, Petr

    2013-01-01

    Roč. 53, č. 3 (2013), s. 270-273. ISSN 0161-5890 R&D Projects: GA MŠk LD12073; GA ČR GA301/09/1826; GA ČR GAP302/10/1759; GA ČR(CZ) GBP302/12/G101; GA MŠk 1M0506 Grant ostatní: European Cooperation in Science and Technology(XE) BM1007; AV ČR(CZ) M200520901 Institutional research plan: CEZ:AV0Z50520514 Keywords : allergy * high-affinity IgE receptor * plasma membrane * transmembrane signaling * 3-helix assembly model Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 3.003, year: 2013

  8. IgE receptor signaling in food allergy pathogenesis.

    Science.gov (United States)

    Oettgen, Hans C; Burton, Oliver T

    2015-10-01

    The pathogenesis of food allergy remains poorly understood. Recent advances in the use of murine models have led to discoveries that mast cells and IgE receptor signaling not only drive immediate hypersensitivity reactions but also exert an immunoregulatory function, promoting the development of allergic sensitivity to foods. We review the evidence that IgE, IgE receptors, key signaling kinases and mast cells impair oral tolerance to ingested foods, preventing the induction of regulatory T cells (Treg) and promoting the acquisition of pro-allergic T helper (Th) 2 responses. We discuss innovative strategies that that could be implemented to counteract these immunoregulatory effects of IgE-mediated mast cell activation, and potentially reverse established sensitization, curing food allergy. PMID:26296054

  9. Studies investigating peripherial blood derived cells that express the high affinity receptor for immunoglobulin E (FceRI) in allergic disorders

    OpenAIRE

    Sihra, Bhupinder Singh

    2008-01-01

    It is just forty years since the identification of immunoglobulin E (IgE) as the reagin responsible for allergen induced immediate hypersensitivity reactions. IgE exerts its biological actions through the binding of its Fc fragment to specific Fc receptors on effector cells. There are two predominant Fc receptors for IgE – Fc?RI, which has a very high affinity for IgE and Fc?RII, which shows less avid binding. For much of the first two decades after the identification of IgE, it w...

  10. Studies investigating peripheral blood derived cells that express the high affinity receptor for immunoglobulin E (Fc?RI) In Allergic Disorders

    OpenAIRE

    Sihra, Bhupinder Singh

    2008-01-01

    It is just forty years since the identification of immunoglobulin E (IgE) as the reagin responsible for allergen induced immediate hypersensitivity reactions. IgE exerts its biological actions through the binding of its Fc fragment to specific Fc receptors on effector cells. There are two predominant Fc receptors for IgE – Fc?RI, which has a very high affinity for IgE and Fc?RII, which shows less avid binding. For much of the first two decades after the identification of IgE, it was thought t...

  11. Structural determinants of sigma receptor affinity

    International Nuclear Information System (INIS)

    The structural determinants of sigma receptor affinity have been evaluated by examining a wide range of compounds related to opioids, neuroleptics, and phenylpiperidine dopaminergic structures for affinity at sigma receptor-binding sites labeled with (+)-[3H]3-PPP. Among opioid compounds, requirements for sigma receptor affinity differ strikingly from the determinants of affinity for conventional opiate receptors. Sigma sites display reverse stereoselectivity to classical opiate receptors. Multi-ringed opiate-related compounds such as morphine and naloxone have negligible affinity for sigma sites, with the highest sigma receptor affinity apparent for benzomorphans which lack the C ring of opioids. Highest affinity among opioids and other compounds occurs with more lipophilic N-substituents. This feature is particularly striking among the 3-PPP derivatives as well as the opioids. The butyrophenone haloperidol is the most potent drug at sigma receptors we have detected. Among the series of butyrophenones, receptor affinity is primarily associated with the 4-phenylpiperidine moiety. Conformational calculations for various compounds indicate a fairly wide range of tolerance for distances between the aromatic ring and the amine nitrogen, which may account for the potency at sigma receptors of structures of considerable diversity. Among the wide range of structures that bind to sigma receptor-binding sites, the common pharmacophore associated with high receptor affinity is a phenylpiperidine with a lipophilic N-substituent

  12. Structural determinants of sigma receptor affinity

    Energy Technology Data Exchange (ETDEWEB)

    Largent, B.L.; Wikstroem, H.G.; Gundlach, A.L.; Snyder, S.H.

    1987-12-01

    The structural determinants of sigma receptor affinity have been evaluated by examining a wide range of compounds related to opioids, neuroleptics, and phenylpiperidine dopaminergic structures for affinity at sigma receptor-binding sites labeled with (+)-(/sup 3/H)3-PPP. Among opioid compounds, requirements for sigma receptor affinity differ strikingly from the determinants of affinity for conventional opiate receptors. Sigma sites display reverse stereoselectivity to classical opiate receptors. Multi-ringed opiate-related compounds such as morphine and naloxone have negligible affinity for sigma sites, with the highest sigma receptor affinity apparent for benzomorphans which lack the C ring of opioids. Highest affinity among opioids and other compounds occurs with more lipophilic N-substituents. This feature is particularly striking among the 3-PPP derivatives as well as the opioids. The butyrophenone haloperidol is the most potent drug at sigma receptors we have detected. Among the series of butyrophenones, receptor affinity is primarily associated with the 4-phenylpiperidine moiety. Conformational calculations for various compounds indicate a fairly wide range of tolerance for distances between the aromatic ring and the amine nitrogen, which may account for the potency at sigma receptors of structures of considerable diversity. Among the wide range of structures that bind to sigma receptor-binding sites, the common pharmacophore associated with high receptor affinity is a phenylpiperidine with a lipophilic N-substituent.

  13. High-affinity neuropeptide Y receptor antagonists.

    OpenAIRE

    Daniels, A J; Matthews, J. E.; Slepetis, R J; Jansen, M; Viveros, O. H.; Tadepalli, A.; Harrington, W; Heyer, D; Landavazo, A; Leban, J J

    1995-01-01

    Neuropeptide Y (NPY) is one of the most abundant peptide transmitters in the mammalian brain. In the periphery it is costored and coreleased with norepinephrine from sympathetic nerve terminals. However, the physiological functions of this peptide remain unclear because of the absence of specific high-affinity receptor antagonists. Three potent NPY receptor antagonists were synthesized and tested for their biological activity in in vitro, ex vivo, and in vivo functional assays. We describe he...

  14. IgE receptor-activated calcium permeability pathway in rat basophilic leukemia cells

    International Nuclear Information System (INIS)

    When antigen-stimulated 45Ca uptake is measured in RBL cells loaded with > 3 mM quin2, re-extrusion of 45Ca is minimized and the initial rate of 45Ca uptake reflects the true unidirectional influx of Ca. This influx correlates more closely with secretion than with the number of IgE receptors aggregated by antigen. The antigen-induced permeability pathway is saturable, having a Km of about 0.7 mM and a Vmax of 0.9 nmol Ca/106 cells/min and it persists for at least an hour provided that receptor aggregation is maintained. The negatively charged fluorescent probe bis-oxonol is insensitive to changes in the mitochondrial membrane potential and is, therefore, a useful plasma membrane potential indicator. Antigen-stimulation of RBL cells equilibrated with bis-oxonol causes a rapid depolarization that peaks within 2-3 minutes and persists until receptor aggregates are disrupted. Antigen-induced depolarization is seen in the absence of extracellular calcium, but is almost completely abolished when both sodium and calcium are replaced by glucose. Addition of calcium restores this response even when sodium is absent. This suggests that the IgE receptor-activated permeability pathway has a similar conductance for sodium and calcium ions

  15. Convulsant bicuculline modifies CNS muscarinic receptor affinity

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    Rodríguez de Lores Arnaiz Georgina

    2006-04-01

    Full Text Available Abstract Background Previous work from this laboratory has shown that the administration of the convulsant drug 3-mercaptopropionic acid (MP, a GAD inhibitor, modifies not only GABA synthesis but also binding of the antagonist [3H]-quinuclidinyl benzilate ([3H]-QNB to central muscarinic receptors, an effect due to an increase in affinity without modifications in binding site number. The cholinergic system has been implicated in several experimental epilepsy models and the ability of acetylcholine to regulate neuronal excitability in the neocortex is well known. To study the potential relationship between GABAergic and cholinergic systems with seizure activity, we analyzed the muscarinic receptor after inducing seizure by bicuculline (BIC, known to antagonize the GABA-A postsynaptic receptor subtype. Results We analyzed binding of muscarinic antagonist [3H]-QNB to rat CNS membranes after i.p. administration of BIC at subconvulsant (1.0 mg/kg and convulsant (7.5 mg/kg doses. Subconvulsant BIC dose failed to develop seizures but produced binding alteration in the cerebellum and hippocampus with roughly 40% increase and 10% decrease, respectively. After convulsant BIC dose, which invariably led to generalized tonic-clonic seizures, binding increased 36% and 15% to cerebellar and striatal membranes respectively, but decreased 12% to hippocampal membranes. Kd value was accordingly modified: with the subconvulsant dose it decreased 27% in cerebellum whereas it increased 61% in hippocampus; with the convulsant dose, Kd value decreased 33% in cerebellum but increased 85% in hippocampus. No change in receptor number site was found, and Hill number was invariably close to unity. Conclusion Results indicate dissimilar central nervous system area susceptibility of muscarinic receptor to BIC. Ligand binding was modified not only by a convulsant BIC dose but also by a subconvulsant dose, indicating that changes are not attributable to the seizure process

  16. Tetrahydroprotoberberine alkaloids with dopamine and σ receptor affinity.

    Science.gov (United States)

    Gadhiya, Satishkumar; Madapa, Sudharshan; Kurtzman, Thomas; Alberts, Ian L; Ramsey, Steven; Pillarsetty, Nagavara-Kishore; Kalidindi, Teja; Harding, Wayne W

    2016-05-01

    Two series of analogues of the tetrahydroprotoberberine (THPB) alkaloid (±)-stepholidine that (a) contain various alkoxy substituents at the C10 position and, (b) were de-rigidified with respect to (±)-stepholidine, were synthesized and evaluated for affinity at dopamine and σ receptors in order to evaluate effects on D3 and σ2 receptor affinity and selectivity. Small n-alkoxy groups are best tolerated by D3 and σ2 receptors. Among all compounds tested, C10 methoxy and ethoxy analogues (10 and 11 respectively) displayed the highest affinity for σ2 receptors as well as σ2 versus σ1 selectivity and also showed the highest D3 receptor affinity. De-rigidification of stepholidine resulted in decreased affinity at all receptors evaluated; thus the tetracyclic THPB framework is advantageous for affinity at dopamine and σ receptors. Docking of the C10 analogues at the D3 receptor, suggest that an ionic interaction between the protonated nitrogen atom and Asp110, a H-bond interaction between the C2 phenol and Ser192, a H-bond interaction between the C10 phenol and Cys181 as well as hydrophobic interactions of the aryl rings to Phe106 and Phe345, are critical for high affinity of the compounds. PMID:27032890

  17. Damnacanthal inhibits IgE receptor-mediated activation of mast cells.

    Science.gov (United States)

    Garcia-Vilas, Javier A; Medina, Miguel A; Melo, Fabio R; Pejler, Gunnar; Garcia-Faroldi, Gianni

    2015-05-01

    Damnacanthal, an anthraquinone obtained from the noni fruit (Morinda citrifolia L.), has been described to possess anti-cancer and anti-inflammatory properties. Since mast cells are key players in various inflammatory conditions as well as in cancer, we considered the possibility that the biological actions of damnacanthal, at least partly, could be due to effects on mast cells. Many of the biological activities of mast cells are mediated by IgE receptor cross-linking, which results in degranulation with release of preformed granule mediators, as well as de novo synthesis and release of additional compounds. Here we show that damnacanthal has profound inhibitory activity on mast cell activation through this pathway. The release of the granule compounds beta-hexosaminidase and tryptase release was completely abrogated by damnacanthal at doses that were non-toxic to mast cells. In addition, damnacanthal inhibited activation-dependent pro-inflammatory gene induction, as well as cytokine/chemokine release in response to mast cell stimulation. The mechanism underlying damnacanthal inhibition was linked to impaired phosphorylation of Syk and Akt. Furthermore, damnacanthal inhibited mast cell activation in response to calcium ionophore A23187. Altogether, the data presented here demonstrate that damnacanthal inhibits mast cell activation induced by different stimuli and open a new window for the use of this compound as a mast cell stabilizer. PMID:25656801

  18. Comparison between sensitivity of autologous skin serum test and autologous plasma skin test in patients with Chronic Idiopathic Urticaria for detection of antibody against IgE or IgE receptor (FcεRIα).

    Science.gov (United States)

    Sajedi, Vahid; Movahedi, Masoud; Aghamohammadi, Asghar; Aghamohamadi, Asghar; Gharagozlou, Mohammad; Ghareguzlou, Mohammad; Shafiei, Alireza; Soheili, Habib; Sanajian, Nahal

    2011-06-01

    Intradermal injection of autologous serum and plasma elicit a cutaneous reactivity in almost 45-60% of patients with Chronic Idiopathic Urticaria (CIU). This reactivity is associated with the presence of auto antibodies against IgE or IgE receptors. This study was carried out to compare the cutaneous reactivity of autologous serum and plasma skin tests in a series of patients with CIU for diagnosis of auto antibodies against IgE or IgE receptor. Fifty eight patients with CIU were injected intradermally with autologous serum and plasma (anticoagulated by citrate). Histamine was used as positive control and normal saline as negative control. The study group was checked by routine laboratory tests (CBC, U/A etc), allergens with skin prick tests, and serum IgE level, and auto antibodies against thyroid as well. Duration of urticaria was another factor which was assessed.There was no significant difference between positive ASST and positive APST patients for the above mentioned tests. 77.6% of the patients were Positive for APST and 65.5% were ASST positive. Duration of urticaria was longer in patients with positive ASST and APST than ASST and APST negative patients, although the difference was not statistically significant.Autologus serum skin test (ASST) and autologous plasma skin test (APST) could be used for estimation of duration and severity of urticaria and planning for the treatment. PMID:21625019

  19. SplitCore Technology Allows Efficient Production of Virus-Like Particles Presenting a Receptor-Contacting Epitope of Human IgE.

    Science.gov (United States)

    Baltabekova, A Zh; Shagyrova, Zh S; Kamzina, A S; Voykov, M; Zhiyenbay, Ye; Ramanculov, E M; Shustov, A V

    2015-08-01

    Immunoglobulin E (IgE) plays a central role in type I hypersensitivity including allergy and asthma. Novel treatment strategy envisages development of a therapeutic vaccine designed to elicit autologous blocking antibodies against the IgE. We sought to develop an IgE-epitope antigen that induces antibodies against a receptor-contacting epitope on human IgE molecule. We designed the VLP immunogens which utilize hepatitis B virus core protein (HBcAg) as a carrier, and present arrays of the receptor-contacting epitopes of the human IgE on their surfaces. FG loop from the IgE domain Cε3 was engineered into the HBcAg. Two constructs explore a well-established approach of insertion into a main immunodominant region of the HBcAg. Third construct is different in that the carrier is produced in a form of an assembly of two polypeptide chains which upon expression remain associated in a stable VLP-forming subunit (SplitCore technology). No VLPs were isolated from E.coli expressing the IgE-epitope antigens with contiguous sequences. On the contrary, the SplitCore antigen carrying the FG loop efficiently formed the VLPs. Immunization of mice with the VLPs presenting receptor-contacting epitope of the IgE elicited antibodies recognizing the human IgE in ELISA. PMID:25837568

  20. High-Affinity Fc Receptor Expression Indicates Relative Immaturity in Human Monocytes.

    Science.gov (United States)

    Clanchy, Felix I L

    2016-05-01

    Within monocyte heterogeneity, subsets represent discrete, well-characterized phenotypes. Although many studies have highlighted differences between subsets, there is evidence that subpopulations represent contiguous stages in a maturational series. As CD14(hi)CD64(hi) monocytes have higher proliferative potential than CD14(hi)CD64(lo) monocytes, the surface marker profile on 4 subsets defined by CD14 and CD64 was measured. The profiles were compared to that of subsets defined by the high-affinity IgE receptor (FcɛRIα), CD16, and CD14; further differences in size, granularity, and buoyancy were measured in subsets delineated by these markers. There was a positive correlation between proliferative monocyte (PM) prevalence and CD64 expression on the classical monocyte subset, and also between PM prevalence and circulating FcɛRIα(+) monocytes. The expression of CD64, the high-affinity IgG receptor, on canonical human monocyte subsets was determined before and after short-term culture, and in response to interleukin (IL)-6, IL-10, macrophage colony-stimulating factor, granulocyte/macrophage colony-stimulating factor and interferon-γ; the influence of these cytokines on monocyte subset transition was also measured. The loss of FcɛRIα expression preceded an increase in CD16 expression in whole blood cultures. These data indicate that high-affinity Fc receptors are expressed on less mature monocytes and that FcɛRIα(+) monocytes are developmentally antecedent to the canonical classical and intermediate monocyte subsets. PMID:26714112

  1. MAST CELLS, MAST/STEM CELL GROWTH FACTOR RECEPTOR (C-KIT/CD117 AND IGE MAY BE INTEGRAL TO THE PATHOGENESIS OF ENDEMIC PEMPHIGUS FOLIACEUS

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    Ana Maria Roselino

    2013-11-01

    Full Text Available Introduction: Pemphigus foliaceus (PF is endemic in some South American countries, especially in Colombia and Brazil; in Brazil, it is also known as fogo selvagem (FS. We aimed to study the presence of mast cells and the expression of the mast/stem cell growth factor receptor (c-kit/CD117 in PF skin biopsies, as well as the role of IgE in the disease pathogenesis. Methods: Forty-four skin biopsies from patients affected by endemic PF (EPF (30 patients from El Bagre, Colombia, and 14 from the northeastern region of São Paulo State, Brazil, 48 control biopsies from Colombian and Brazilian endemic areas, and additional control biopsies from none endemic areas in Colombia and the USA non were studied. Immunohistochemistry (IHC was performed to evaluate skin biopsies with anti-mast cell tryptase (MCT, anti-c-kit and anti-IgE antibodies. We also searched for serum IgE in 30 EPF and 30 non-atopic controls from the El Bagre region via ELISA. In our El Bagre patients and controls, we also searched for IgE in skin samples by direct immunofluorescence. Results: In 100% of the EPF biopsies, MCT, c-kit and IgE were identified with stronger expression relative to control biopsies, especially in the inflammatory infiltrates around upper dermal blood vessels and dermal eccrine glands. IgE staining was positive along the BMZ in some EPF skin samples. The DIF results confirmed a linear deposition of IgE at the BMZ. Increased IgE serum levels were also noted in PF patients relative to controls.. Conclusions: In patients with EPF, the observed increased expression of MCT, c-kit and IgE in lesional skin, associated with higher serum IgE levels may indicate possible IgE participation in the antigenic response.

  2. Comparison between Sensitivity of Autologous Skin Serum Test and Autologous Plasma Skin Test in Patients with Chronic Idiopathic Urticaria for Detection of Antibody against IgE or IgE Receptor (FcεRIα

    Directory of Open Access Journals (Sweden)

    Vahid Sajedi

    2011-06-01

    Full Text Available Intradermal injection of autologous serum and plasma elicit a cutaneous reactivity in almost 45-60% of patients with Chronic Idiopathic Urticaria (CIU.   This reactivity is associated with the presence of auto antibodies against IgE or IgE receptors. This study was carried out to compare the cutaneous reactivity of autologous serum and plasma skin tests in a series of patients with CIU for diagnosis of auto ntibodies against IgE or IgE receptor.Fifty eight patients with CIU were injected intradermally with autologous serum and plasma (anticoagulated by  citrate.  Histamine  was  used  as  positive  control  and  normal  saline  as negative control. The study group was checked by routine laboratory tests (CBC, U/A etc, allergens with skin prick tests, and serum IgE level, and auto antibodies against thyroid as well. Duration of urticaria was another factor which was assessed.There was no significant difference between positive ASST and positive APST patients for the above mentioned tests. 77.6% of the patients were Positive for APST and 65.5% were ASST positive. Duration of urticaria was longer in patients with positive ASST and APST than ASST and APST negative patients, although the difference was not statistically significant.Autologous serum skin test (ASST and autologous plasma skin test (APST could be used for estimation of duration and severity of urticaria and planning for the treatment.

  3. Serum IgE, tumor epidermal growth factor receptor expression, and inherited polymorphisms associated with glioma survival.

    Science.gov (United States)

    Wrensch, Margaret; Wiencke, John K; Wiemels, Joe; Miike, Rei; Patoka, Joe; Moghadassi, Michelle; McMillan, Alex; Kelsey, Karl T; Aldape, Kenneth; Lamborn, Kathleen R; Parsa, Andrew T; Sison, Jennette D; Prados, Michael D

    2006-04-15

    In population-based glioma patients, we examined survival in relation to potentially pertinent constitutive polymorphisms, serologic factors, and tumor genetic and protein alterations in epidermal growth factor receptor (EGFR), MDM2, and TP53. Subjects were newly diagnosed adults residing in the San Francisco Bay Surveillance Epidemiology and End Results Area during 1991 to 1994 and 1997 to 1999 with central neuropathology review (n = 873). Subjects provided blood for serologic studies of IgE and IgG to four herpes viruses and constitutive specimens for genotyping 22 polymorphisms in 13 genes (n = 471). We obtained 595 of 697 astrocytic tumors for marker studies. We determined treatments, vital status, and other factors using registry, interview, medical record, and active follow-up data. Cox regressions for survival were adjusted for age, gender, ethnicity, study series, resection versus biopsy only, radiation, and chemotherapy. Using a stringent P < 0.001, glioma survival was associated with ERCC1 C8092A [hazard ratio (HR), 0.72; 95% confidence limits (95% CL), 0.60-0.86; P = 0.0004] and GSTT1 deletion (HR, 1.64; 95% CL, 1.25-2.16; P = 0.0004); glioblastoma patients with elevated IgE had 9 months longer survival than those with normal or borderline IgE levels (HR, 0.62; 95% CL, 0.47-0.82; P = 0.0007), and EGFR expression in anaplastic astrocytoma was associated with nearly 3-fold poorer survival (HR, 2.97; 95% CL, 1.70-5.19; P = 0.0001). Based on our and others' findings, we recommend further studies to (a) understand relationships of elevated IgE levels and other immunologic factors with improved glioblastoma survival potentially relevant to immunologic therapies and (b) determine which inherited ERCC1 variants or other variants in the 19q13.3 region influence survival. We also suggest that tumor EGFR expression be incorporated into clinical evaluation of anaplastic astrocytoma patients. PMID:16618782

  4. Isolation of high-affinity human IgE and IgG antibodies recognising Bet v 1 and Humicola lanuginosa lipase from combinatorial phage libraries

    DEFF Research Database (Denmark)

    Jakobsen, Charlotte G; Bodtger, Uffe; Kristensen, Peter; Poulsen, Lars K.; Roggen, Erwin L

    2004-01-01

    . Human IgE and IgG libraries have been created from patients allergic to birch pollen or lipase. These libraries have been used to select binders recognising the major birch pollen allergen Bet v 1 and Humicola lanuginosa lipase. A panel of allergen-specific IgE and IgG antibodies were identified; these...

  5. Altered catecholamine receptor affinity in rabbit aortic intimal hyperplasia

    International Nuclear Information System (INIS)

    Intimal thickening is a universal response to endothelial denudation and is also thought to be a precursor of atherosclerosis. The authors have demonstrated selective supersensitivity in arterial intimal hyperplasia to norepinephrine and they now report a possible mechanism for this. Binding studies in rabbit aorta with the selective alpha 1-adrenergic radioligand 125I-HEAT demonstrated that there was no change in receptor density (20 ± 4 fmole/10(6) cells) in intact vascular smooth muscle cells at either 5 or 14 days after denudation. However, competition studies showed a 2.6-fold increase in alpha 1-adrenergic receptor affinity for norepinephrine in intimal hyperplastic tissue (P less than 0.05). This increased affinity for norepinephrine was associated with a greater increase in 32P-labeled phosphatidylinositol (148% intimal thickening versus 76% control) and phosphatidic acid (151% intimal thickening versus 56% control) following norepinephrine stimulation of free floating rings of intimal hyperplastic aorta. These data suggest that the catecholamine supersensitivity in rabbit aortic intimal hyperplasia is receptor mediated and may be linked to the phosphatidylinositol cycle

  6. Altered catecholamine receptor affinity in rabbit aortic intimal hyperplasia

    Energy Technology Data Exchange (ETDEWEB)

    O' Malley, M.K.; Cotecchia, S.; Hagen, P.O. (Duke University Medical Center, Durham, NC (USA))

    1991-08-01

    Intimal thickening is a universal response to endothelial denudation and is also thought to be a precursor of atherosclerosis. The authors have demonstrated selective supersensitivity in arterial intimal hyperplasia to norepinephrine and they now report a possible mechanism for this. Binding studies in rabbit aorta with the selective alpha 1-adrenergic radioligand 125I-HEAT demonstrated that there was no change in receptor density (20 {plus minus} 4 fmole/10(6) cells) in intact vascular smooth muscle cells at either 5 or 14 days after denudation. However, competition studies showed a 2.6-fold increase in alpha 1-adrenergic receptor affinity for norepinephrine in intimal hyperplastic tissue (P less than 0.05). This increased affinity for norepinephrine was associated with a greater increase in 32P-labeled phosphatidylinositol (148% intimal thickening versus 76% control) and phosphatidic acid (151% intimal thickening versus 56% control) following norepinephrine stimulation of free floating rings of intimal hyperplastic aorta. These data suggest that the catecholamine supersensitivity in rabbit aortic intimal hyperplasia is receptor mediated and may be linked to the phosphatidylinositol cycle.

  7. THE RECEPTOR BINDING AFFINITIES, ANTIPROGESTERONE AND ANTIGLUCOCORTICOID ACTIVITIES OF MIFEPRISTONE AND LILOPRISTONE

    Institute of Scientific and Technical Information of China (English)

    LIUYong-Qiang; WUXi-Rui

    1989-01-01

    With radioligand binding assays, the receptor binding affmities of mifepristone and lilopristone to the rabbit uterus cytosol progesterone receptor and the rat fiver cytosol glucocorticoid receptor have been measured. The relative binding affinities ( RBA ) of

  8. Functional significance of IL-4 receptor on B cells in IL-4-induced human IgE production.

    Science.gov (United States)

    Yanagihara, Y; Ikizawa, K; Kajiwara, K; Koshio, T; Basaki, Y; Akiyama, K

    1995-12-01

    IL-4 with the IgE-inducing activity is shown to upregulate the expression of IL-4 receptor (IL-4R) on lymphocytes. Antisense strategy was used that aimed at investigating the significance of IL-4-induced upregulation of IL-4R on B cells in human IgE production. When an antisense phosphorothioate oligodeoxynucleotide to IL-4R (S-oligo 1) was added to B cells together with IL-4, the agent selectively abrogated the upregulation of IL-4R without affecting its constitutive level expression. Moreover, S-oligo 1 had a suppressive effect on the T-cell-independent synthesis of IgE by B cells costimulated with IL-4 and anti-CD40 antibody. This suppression was accompanied by inhibition of mature but not germline C epsilon transcription. These findings indicate that constitutively expressed IL-4R provides a signal or signals responsible for the induction of germline C epsilon transcription and suggest that IL-4R upregulation may be required for the subsequent class switch recombination that leads to mature C epsilon transcription and IgE synthesis. The IL-4R signal transduction mechanism underlying germline C epsilon transcription was also analyzed in a human Burkitt lymphoma B-cell line, DND39. Induction of germline C epsilon transcripts in DND39 cells by IL-4 required at least two distinct signaling cascades. One was mediated by enhancement of tyrosine phosphorylation of a 57 kd protein associated with phospholipase C-gamma 1 (PLC-gamma 1) that resulted in PLC-gamma 1 activation, inositol lipid hydrolysis, and protein kinase C delta translocation. The other was dependent on phosphatidylinositol 3-kinase, whose activation induced protein kinase C zeta translocation. In fact, kinase inhibitors such as herbimycin A, K-252a, and wortmannin were effective in inhibiting IL-4-induced germline C epsilon transcription. Therefore, in addition to activation of protein tyrosine kinases, coordinated actions of PLC-gamma 1 and phosphatidylinositol 3-kinase may be involved in IL-4-driven

  9. Influences of hydrocarbon linkers on the receptor binding affinities of gonadotropin-releasing hormone peptides

    OpenAIRE

    Guo, Haixun; Hathaway, Helen; Royce, Melanie E.; Prossnitz, Eric R.; Miao, Yubin

    2013-01-01

    Three new DOTA-conjugated GnRH peptides with various hydrocarbon linkers were synthesized to evaluate the influences of the linkers on their receptor binding affinities. The hydrocarbon linker displayed a profound impact on the receptor binding affinities of DOTA-conjugated GnRH peptides. The Aun linker was better than Gaba, Ahx and Aoc linkers in retaining strong receptor binding affinity of the GnRH peptide. DOTA-Aun-(D-Lys6-GnRH) displayed 22.8 nM GnRH receptor binding affinity. 111In-DOTA...

  10. Binding affinity prediction of novel estrogen receptor ligands using receptor-based 3-D QSAR methods.

    Science.gov (United States)

    Sippl, Wolfgang

    2002-12-01

    We have recently reported the development of a 3-D QSAR model for estrogen receptor ligands showing a significant correlation between calculated molecular interaction fields and experimentally measured binding affinity. The ligand alignment obtained from docking simulations was taken as basis for a comparative field analysis applying the GRID/GOLPE program. Using the interaction field derived with a water probe and applying the smart region definition (SRD) variable selection procedure, a significant and robust model was obtained (q(2)(LOO)=0.921, SDEP=0.345). To further analyze the robustness and the predictivity of the established model several recently developed estrogen receptor ligands were selected as external test set. An excellent agreement between predicted and experimental binding data was obtained indicated by an external SDEP of 0.531. Two other traditionally used prediction techniques were applied in order to check the performance of the receptor-based 3-D QSAR procedure. The interaction energies calculated on the basis of receptor-ligand complexes were correlated with experimentally observed affinities. Also ligand-based 3-D QSAR models were generated using program FlexS. The interaction energy-based model, as well as the ligand-based 3-D QSAR models yielded models with lower predictivity. The comparison with the interaction energy-based model and with the ligand-based 3-D QSAR models, respectively, indicates that the combination of receptor-based and 3-D QSAR methods is able to improve the quality of prediction. PMID:12413831

  11. The Quantum Nature of Drug-Receptor Interactions: Deuteration Changes Binding Affinities for Histamine Receptor Ligands

    Science.gov (United States)

    Repič, Matej; Zakšek, Maja; Kotnik, Kristina; Fijan, Estera; Mavri, Janez

    2016-01-01

    In this article we report a combined experimental and computational study concerning the effects of deuteration on the binding of histamine and two other histaminergic agonists to 3H-tiotidine-labeled histamine H2 receptor in neonatal rat astrocytes. Binding affinities were measured by displacing radiolabeled tiotidine from H2 receptor binding sites present on cultured neonatal rat astrocytes. Quantum-chemical calculations were performed by employing the empirical quantization of nuclear motion within a cluster model of the receptor binding site extracted from the homology model of the entire H2 receptor. Structure of H2 receptor built by homology modelling is attached in the supporting information (S1 Table) Experiments clearly demonstrate that deuteration affects the binding by increasing the affinity for histamine and reducing it for 2-methylhistamine, while basically leaving it unchanged for 4-methylhistamine. Ab initio quantum-chemical calculations on the cluster system extracted from the homology H2 model along with the implicit quantization of the acidic N–H and O–H bonds demonstrate that these changes in the binding can be rationalized by the altered strength of the hydrogen bonding upon deuteration known as the Ubbelohde effect. Our computational analysis also reveals a new mechanism of histamine binding, which underlines an important role of Tyr250 residue. The present work is, to our best knowledge, the first study of nuclear quantum effects on ligand receptor binding. The ligand H/D substitution is relevant for therapy in the context of perdeuterated and thus more stable drugs that are expected to enter therapeutic practice in the near future. Moreover, presented approach may contribute towards understanding receptor activation, while a distant goal remains in silico discrimination between agonists and antagonists based on the receptor structure. PMID:27159606

  12. Attenuation of IgE receptor signalling in mast cells as molecular basis for the antiallergic action of glucocorticoids

    Energy Technology Data Exchange (ETDEWEB)

    Sancono, A.

    2003-04-01

    Glucocorticoids suppress the expression of Fc{epsilon}RI alpha chain gene by inhibiting its promoter activity. This effect requires de novo protein synthesis and regulatory elements at the Fc{epsilon}RI {alpha}-chain promoter that bind the transcription factors GATA-1, Elf-1, Pu.1 and YY1. The downregulation of the Fc{epsilon}RI {alpha}-chain gene expression correlates with a reduced surface expression of the Fc{epsilon}RI. Furthermore, glucocorticoid treatment promotes inactivation of the lck/yes-related novel (Lyn) Src-like tyrosine kinase, responsible of phosphorylation and activation of the Fc{epsilon}RI, via enhanced phosphorylation of the regulatory tyrosine residue of Lyn. The reduction of surface expressed IgE receptor and the inactivation of Lyn would suppress the signal transduction events originating from the Fc{epsilon}RI and ending up with the activation of the downstream targets ERK1/2. In addition, it has been shown here that glucocorticoids inhibit IgE receptor signalling by enhancing the expression of the MAPK phosphatase MKP-1, which in turn inhibits the activation of ERK1/2. The glucocorticoid-mediated enhancement in MKP-1 expression occurs at the MKP-1 promoter level. This regulation requires the glucocorticoid receptor (GR) dimerisation function and the presence of discrete elements on the promoter proximal sequence, suggesting that it is a direct action of the GR. A crucial role of MKP-1 in glucocorticoid-mediated repression of ERK1/2 phosphorylation was demonstrated in bone marrow derived mast cells from MKP-1-deficient mice. ERK1/2 in these cells are activated by IgE receptor cross-linking or by Stem Cell Factor through the c-kit receptor, and they were no longer inhibited by glucocorticoids, while this was the case in cells from wild-type mice. However, ERK1/2 activity in other cell types such as thymocytes and splenocytes of MKP-1 deficient mice, could still be inhibited by glucocorticoids. These results dmeonstrate that repression of ERK1

  13. Synthesis and receptor binding affinity of new selective GluR5 ligands

    DEFF Research Database (Denmark)

    Bunch, L; Johansen, T H; Bräuner-Osborne, Hans;

    2001-01-01

    Two hybrid analogues of the kainic acid receptor agonists, 2-amino-3-(5-tert-butyl-3-hydroxy-4-isoxazolyl)propionic acid (ATPA) and (2S,4R)-4-methylglutamic acid ((2S,4R)-4-Me-Glu), were designed, synthesized, and characterized in radioligand binding assays using cloned ionotropic and metabotropi.......0 and 2.0 microM. respectively. Their affinities in the [3H]AMPA binding assay on native cortical receptors were shown to correlate with their GluR2 affinity rather than their GluR5 affinity. No affinity for GluR6 was detected (IC50 > 100 microM)....

  14. Only high-affinity receptors for interleukin 2 mediate internalization of ligand

    International Nuclear Information System (INIS)

    Interleukin 2 (IL-2) receptors are expressed on activated T cells and in select T-cell leukemias. Recently, it has been demonstrated that at least two classes of receptor for IL-2 exist with markedly different affinities for ligand. All known biological actions of IL-2 have been correlated with occupancy of high-affinity sites; the function of the low-affinity sites remains unknown. Receptor-mediated endocytosis is the primary means of internalization of cell-surface receptors and their ligands. The internalization of IL-2 bound to high- and low-affinity receptor sites was studied in a human T-cell lymphotrophic virus type 1 (HTLV-1)-infected human T-cell leukemia cell line and in a cloned murine cytotoxic T-cell line (CTLL). Internalization of IL-2 occurred only when bound to high-affinity sites. In addition, an anti-receptor antibody (anti-Tac), which binds equally well to high- and low-affinity sites, demonstrated no detectable internalization. The implications of these findings as they relate to IL-2 receptor structure and function are discussed

  15. Human IgE is efficiently produced in glycosylated and biologically active form in lepidopteran cells.

    Science.gov (United States)

    Bantleon, Frank; Wolf, Sara; Seismann, Henning; Dam, Svend; Lorentzen, Andrea; Miehe, Michaela; Jabs, Frederic; Jakob, Thilo; Plum, Melanie; Spillner, Edzard

    2016-04-01

    TH2-biased immunity to parasites and allergens is often associated with increased levels of antigen-specific and high affinity IgE. The role in reacting against minute amounts of target structures and to provoke severe anaphylactic reactions renders IgE a mechanistically outstanding isotype. IgE represents the least abundant serum antibody isotype and exhibits a variety of peculiarities including structure, extensive glycosylation and effector functions. Despite large progress in antibody technologies, however, the recombinant access to isotypes beyond IgG such as IgE still is scarce. The capacity of expression systems has to meet the complex structural conformations and the extensive posttranslational modifications that are indispensable for biological activity. In order to provide alternatives to mammalian expression systems with often low yield and a more complex glycosylation pattern we established the recombinant production of the highly complex IgE isotype in insect cells. Recombinant IgE (rIgE) was efficiently assembled and secreted into the supernatant in yields of >30 mg/L. Purification from serum free medium using different downstream processing methods provided large amounts of rIgE. This exhibited a highly specific interaction with its antigen, therapeutic anti-IgE and its high affinity receptor, the FcεRI. Lectins and glyco-proteomic analyses proved the presence of prototypic insect type N-glycans on the epsilon heavy chain. Mediator release assays demonstrated a biological activity of the rIgE comparable to IgE derived from mammalian cells. In summary the expression in insect cells provides rIgE with variant glycosylation pattern, but retained characteristics and biological activity. Therefore our data contribute to the understanding of functional and structural aspects and potential use of the IgE isotype. PMID:26943931

  16. Affinity profiles of hexahydro-sila-difenidol analogues at muscarinic receptor subtypes

    OpenAIRE

    Lambrecht, G.; Feifel, R.; Wagner-Röder, M.; Strohmann, C.; Zilch, H.; Tacke, Reinhold; Waelbroeck, M.; Christophe, J; Boddeke, H.; Mutschler, E.

    2012-01-01

    In an attempt to assess the structural requirements of hexahydro-sila-difenidol for potency and selectivity, a series of analogues modified in the amino group and the phenyl ring were investigated for their affinity to muscarinic M1- (rabbit vas deferens), Mr (guinea-pig atria) and Mr (guinea-pig ileum) receptors. All compounds were competitive antagonists in the three tissues. Their affinities to the three muscarinic receptor subtypes differed by more than two orders of magnitude and the obs...

  17. Gastrin receptor characterization: affinity cross-linking of the gastrin receptor on canine gastric parietal cells

    International Nuclear Information System (INIS)

    The authors applied affinity cross-linking methods to label the gastrin receptor on isolated canine gastric parietal cells in order to elucidate the nature of its chemical structure. 125I-labeled Leu15-gastrin and 125I-labeled gastrin/sub 2-17/ bound to intact parietal cells and their membranes with equal affinity, and half-maximal inhibition of binding was obtained at an incubation concentration of 3.2 x 10-10 M unlabeled gastrin. 125I-gastrin/sub 2-17/ was cross-linked to plasma membranes or intact parietal cells by incubation in disuccinimidyl suberate. The membrane pellets were solubilized with or without dithiothreitol and applied to electrophoresis on 7.5% sodium dodecyl sulfate polyacrylamide gels. Autoradiograms revealed a band of labeling at M/sub r/ 76,000 and labeling of this band was inhibited in a dose-dependent fashion by addition of unlabeled gastrin to the incubation mixture. Dithiothreitol in concentrations as high as 100 mM did not later the electrophoretic mobility of the labeled band. After taking into account the molecular weight of 125I-gastrin/sub 2-17/, the results suggest that the gastrin receptor on parietal cells is a single protein of M/sub r/ 74,000 without disulfide-linked subunits

  18. Short-term desensitization of muscarinic cholinergic receptors in mouse neuroblastoma cells: selective loss of agonist low-affinity and pirenzepine high-affinity binding sites

    International Nuclear Information System (INIS)

    The effects of brief incubation with carbamylcholine on subsequent binding of [3H]N-methylscopolamine were investigated in mouse neuroblastoma cells (clone N1E-115). This treatment demonstrated that the muscarinic receptors in this neuronal clone can be divided into two types; one which is readily susceptible to regulation by receptor agonists, whereas the other is resistant in this regard. In control cells, both pirenzepine and carbamylcholine interacted with high- and low-affinity subsets of muscarinic receptors. Computer-assisted analysis of the competition between pirenzepine and carbamylcholine with [3H]N-methylscopolamine showed that the receptor sites remaining upon desensitization are composed mainly of pirenzepine low-affinity and agonist high-affinity binding sites. Furthermore, there was an excellent correlation between the ability of various muscarinic receptor agonists to induce a decrease in consequent [3H]N-methylscopolamine binding and their efficacy in stimulating cyclic GMP synthesis in these cells. Thus, only the agonists that are known to recognize the receptor's low-affinity conformation in order to elicit increases in cyclic GMP levels were capable of diminishing ligand binding. Taken together, our present results suggest that the receptor population that is sensitive to regulation by agonists includes both the pirenzepine high-affinity and the agonist low-affinity receptor binding states. In addition, the sensitivity of these receptor subsets to rapid regulation by agonists further implicates their involvement in desensitization of muscarinic receptor-mediated cyclic GMP formation

  19. Modulation of gephyrin-glycine receptor affinity by multivalency

    DEFF Research Database (Denmark)

    Maric, Hans-Michael; Kasaragod, Vikram Babu; Schindelin, Hermann

    2014-01-01

    Gephyrin is a major determinant for the accumulation and anchoring of glycine receptors (GlyRs) and the majority of γ-aminobutyric acid type A receptors (GABAARs) at postsynaptic sites. Here we explored the interaction of gephyrin with a dimeric form of a GlyR β-subunit receptor-derived peptide. A...

  20. Serum IgE clearance is facilitated by human FcεRI internalization

    Science.gov (United States)

    Greer, Alexandra M.; Wu, Nan; Putnam, Amy L.; Woodruff, Prescott G.; Wolters, Paul; Kinet, Jean-Pierre; Shin, Jeoung-Sook

    2014-01-01

    The high-affinity IgE receptor FcεRI is constitutively expressed in mast cells and basophils and is required for transmitting stimulatory signals upon engagement of IgE-bound allergens. FcεRI is also constitutively expressed in dendritic cells (DCs) and monocytes in humans; however, the specific functions of the FcεRI expressed by these cells are not completely understood. Here, we found that FcεRI expressed by human blood DC antigen 1–positive (BDCA1+) DCs and monocytes, but not basophils, traffics to endolysosomal compartments under steady-state conditions. Furthermore, IgE bound to FcεRI on BDCA1+ DCs was rapidly endocytosed, transported to the lysosomes, and degraded in vitro. IgE injected into mice expressing human FcεRIα (FCER1A-Tg mice) was endocytosed by conventional DCs and monocytes, and endocytosis was associated with rapid clearance of circulating IgE from these mice. Importantly, this rapid IgE clearance was dependent on monocytes or DCs but not basophils. These findings strongly suggest that constitutive internalization of human FcεRI by DCs and monocytes distinctively contributes to serum IgE clearance. PMID:24569373

  1. Relative binding affinity of diethylstilbestrol to uterine nuclear estrogen receptor: effect of serum and serum albumin

    International Nuclear Information System (INIS)

    The relative binding affinity (RBA) of tritium labelled diethylstilbestrol (DES) was determined in nuclear fractions of rat uterus. DES displayed a two- to threefold greater affinity than estradiol for nuclear E receptor. The RBA of DES to nuclear E receptor was lowered significantly in the presence of rat serum (43 +/- 1) or human serum (52 +/- 7). Dilution of human serum resulted in a progressive increase in the RBA of DES which approached that observed in the absence of serum. Addition of purified human serum albumin mimicked the decrease in RBA of DES that was observed with serum. The IC50 of estradiol was not changed in the presence of either rat serum or albumin. These data show that DES possesses a greater affinity for nuclear E receptor than estradiol and that serum albumin can modulate DES binding to uterine E receptor

  2. Stability of the Neurotensin Receptor NTS1 Free in Detergent Solution and Immobilized to Affinity Resin

    OpenAIRE

    White, Jim F.; Reinhard Grisshammer

    2010-01-01

    BACKGROUND: Purification of recombinant membrane receptors is commonly achieved by use of an affinity tag followed by an additional chromatography step if required. This second step may exploit specific receptor properties such as ligand binding. However, the effects of multiple purification steps on protein yield and integrity are often poorly documented. We have previously reported a robust two-step purification procedure for the recombinant rat neurotensin receptor NTS1 to give milligram q...

  3. Stability of the Neurotensin Receptor NTS1 Free in Detergent Solution and Immobilized to Affinity Resin

    OpenAIRE

    White, Jim F.; Grisshammer, Reinhard

    2010-01-01

    Background Purification of recombinant membrane receptors is commonly achieved by use of an affinity tag followed by an additional chromatography step if required. This second step may exploit specific receptor properties such as ligand binding. However, the effects of multiple purification steps on protein yield and integrity are often poorly documented. We have previously reported a robust two-step purification procedure for the recombinant rat neurotensin receptor NTS1 to give milligram qu...

  4. Role of the T cell receptor ligand affinity in T cell activation by bacterial superantigens

    DEFF Research Database (Denmark)

    Andersen, P S; Geisler, C; Buus, S; Mariuzza, R A; Karjalainen, K

    2001-01-01

    the SEC3 variants correlated with enhanced binding without any optimum in the binding range covered by native TCR ligands. Comparable studies using anti-TCR antibodies of known affinity confirmed these observations. By comparing the biological potency of the two sets of ligands, we found a significant...... correlation between ligand affinity and ligand potency indicating that it is the density of receptor-ligand complexes in the T cell contact area that determines TCR signaling strength....

  5. Effect of sodium ion on the affinity of naloxone for the kappa opioid receptor

    International Nuclear Information System (INIS)

    Several investigators have observed that sodium ion enhances the binding of naloxone to opioid receptors. This effect has generally been attributed to allosteric modulation of the state of the mu receptor. However, a recent claim has been made that the enhancement does not involve a change in the mu receptor, but instead occurs because naloxone becomes a more kappa-specific drug when sodium ion is present in high concentration. Since the claim was not based on experimental evidence from binding studies involving known high-affinity kappa ligands, the authors have investigated the competition of naloxone for the kappa site using [3H]U-69593 as the marker for receptor binding. Assays were carried out in the presence and absence of 100 mM NaCl. The results of the study indicate that sodium ion does not increase the affinity of naloxone or U-69593 for the kappa receptor. 9 references, 1 figure

  6. Effect of sodium ion on the affinity of naloxone for the kappa opioid receptor

    Energy Technology Data Exchange (ETDEWEB)

    Cheney, B.V.; Lahti, R.A.

    1987-03-16

    Several investigators have observed that sodium ion enhances the binding of naloxone to opioid receptors. This effect has generally been attributed to allosteric modulation of the state of the mu receptor. However, a recent claim has been made that the enhancement does not involve a change in the mu receptor, but instead occurs because naloxone becomes a more kappa-specific drug when sodium ion is present in high concentration. Since the claim was not based on experimental evidence from binding studies involving known high-affinity kappa ligands, the authors have investigated the competition of naloxone for the kappa site using (/sup 3/H)U-69593 as the marker for receptor binding. Assays were carried out in the presence and absence of 100 mM NaCl. The results of the study indicate that sodium ion does not increase the affinity of naloxone or U-69593 for the kappa receptor. 9 references, 1 figure.

  7. Binding affinities of anti-acetylcholine receptor autoantibodies in myasthenia gravis

    International Nuclear Information System (INIS)

    Antibodies directed against acetylcholine (ACh) receptors are present in the sera of nearly 90% of patients with myasthenia gravis (MG), and are involved in the pathogenesis of this autoimmune disease. However, the antibody titers measured by the standard radioimmunoassay correspond poorly with the clinical severity of the disease. To determine whether this disparity could be accounted for by differences in the binding affinities of anti-ACh receptor antibodies in different patients, we have measured the binding affinities of these autoantibodies in 15 sera from MG patients. The affinity constants (K/sub o/), as determined by Scatchard analysis, were all in the range of 1010 M-1, comparable to the highest values reported in immunized animals. The affinity constants were truly representative of the population of autoantibodies detected by the radioimmunoassay, as shown by the remarkable linearity of the Scatchard plots (r2>0.90) and the close correlation between the antibody titers determined by extrapolation of the Scatchard plots and by saturation analysis (r = 0.99; p < 0.001). There was only a 6-fold variation in affinity constants measured in this series of patients despite widely differing antibody titers and severity of the disease. Factors other than the titer and affinity of anti-ACh receptor antibodies may correlate better with the clinical manifestations of MG

  8. Binding affinities of anti-acetylcholine receptor autoantibodies in myasthenia gravis

    Energy Technology Data Exchange (ETDEWEB)

    Bray, J.J.; Drachman, D.B.

    1982-01-01

    Antibodies directed against acetylcholine (ACh) receptors are present in the sera of nearly 90% of patients with myasthenia gravis (MG), and are involved in the pathogenesis of this autoimmune disease. However, the antibody titers measured by the standard radioimmunoassay correspond poorly with the clinical severity of the disease. To determine whether this disparity could be accounted for by differences in the binding affinities of anti-ACh receptor antibodies in different patients, we have measured the binding affinities of these autoantibodies in 15 sera from MG patients. The affinity constants (K/sub o/), as determined by Scatchard analysis, were all in the range of 10/sup 10/ M/sup -1/, comparable to the highest values reported in immunized animals. The affinity constants were truly representative of the population of autoantibodies detected by the radioimmunoassay, as shown by the remarkable linearity of the Scatchard plots (r/sup 2/>0.90) and the close correlation between the antibody titers determined by extrapolation of the Scatchard plots and by saturation analysis (r = 0.99; p < 0.001). There was only a 6-fold variation in affinity constants measured in this series of patients despite widely differing antibody titers and severity of the disease. Factors other than the titer and affinity of anti-ACh receptor antibodies may correlate better with the clinical manifestations of MG.

  9. Antigen transfer from exosomes to dendritic cells as an explanation for the immune enhancement seen by IgE immune complexes.

    Directory of Open Access Journals (Sweden)

    Rebecca K Martin

    Full Text Available IgE antigen complexes induce increased specific T cell proliferation and increased specific IgG production. Immediately after immunization, CD23(+ B cells capture IgE antigen complexes, transport them to the spleen where, via unknown mechanisms, dendritic cells capture the antigen and present it to T cells. CD23, the low affinity IgE receptor, binds IgE antigen complexes and internalizes them. In this study, we show that these complexes are processed onto B-cell derived exosomes (bexosomes in a CD23 dependent manner. The bexosomes carry CD23, IgE and MHC II and stimulate antigen specific T-cell proliferation in vitro. When IgE antigen complex stimulated bexosomes are incubated with dendritic cells, dendritic cells induce specific T-cell proliferation in vivo, similar to IgE antigen complexes. This suggests that bexosomes can provide the essential transfer mechanism for IgE antigen complexes from B cells to dendritic cells.

  10. Affinity labeling of rat liver thyroid hormone nuclear receptor.

    OpenAIRE

    Nikodem, V M; Cheng, S Y; Rall, J. E.

    1980-01-01

    The thyroid hormone receptor from rat liver nuclei has been covalently labeled with the N-bromoacetyl derivatives of L-thyroxine (T4) and 3,3',5-triiodo-L-thyronine (T3). Displacement binding studies showed that, in the presence of 100-fold molar excess of unlabeled N-bromoacetyl-T3 or T4, binding of [125I]T3 or [125I]T4 was nearly totally inhibited. Heat inactivation of the receptor (55 degrees C for 15 min) resulted in parallel losses in the binding of T3 (95%) and N-bromoacetyl-T3 (93%). T...

  11. Induction and upregulation by interleukin 2 of high-affinity interleukin 2 receptors on thymocytes and T cells.

    OpenAIRE

    Reem, G H; Yeh, N H; Urdal, D L; Kilian, P L; J.J. Farrar

    1985-01-01

    We show that purified recombinant interleukin 2 (rIL-2) alone induces the expression of high- and low-affinity interleukin 2 (IL-2) receptors in vitro on human T cells and thymocytes that have not been activated previously by lectins or other inducing agents. IL-2 receptors are expressed after 24 hr, as determined by the binding of 125I-labeled monoclonal anti-IL-2 receptor antibody 2A3, which binds equally to high- and low-affinity receptors. High-affinity receptors were distinguished from l...

  12. Role of T cell receptor affinity in the efficacy and specificity of adoptive T cell therapies

    Directory of Open Access Journals (Sweden)

    Jennifer D. Stone

    2013-08-01

    Full Text Available Over the last several years, there has been considerable progress in the treatment of cancer using gene modified adoptive T cell therapies. Two approaches have been used, one involving the introduction of a conventional alpha-beta T cell receptor (TCR against a pepMHC cancer antigen, and the second involving introduction of a chimeric antigen receptor (CAR consisting of a single-chain antibody as an Fv fragment (scFv linked to transmembrane and signaling domains. In this review, we focus on one aspect of TCR-mediated adoptive T cell therapies, the impact of the affinity of the alpha-beta TCR for the pepMHC cancer antigen on both efficacy and specificity. We discuss the advantages of higher affinity TCRs in mediating potent activity of CD4 T cells. This is balanced with the potential disadvantage of higher affinity TCRs in mediating greater self-reactivity against a wider range of structurally similar antigenic peptides, especially in synergy with the CD8 co-receptor. Both TCR affinity and target selection will influence potential safety issues. We suggest pre-clinical strategies that might be used to examine each TCR for possible on-target and off-target side effects due to self-reactivities, and to adjust TCR affinities accordingly.

  13. Several distinct properties of the IgE repertoire determine effector cell degranulation in response to allergen challenge

    DEFF Research Database (Denmark)

    Christensen, Lars Harder; Holm, Jens-Christian; Lund, Gitte; Riise, Erik; Lund, Kaare

    2008-01-01

    E affinity for allergen, and IgE clonality are all distinct properties of allergic patients' IgE repertoires. However, the inability to isolate individual IgE antibodies from allergic patients' sera presents a major barrier to understanding the importance of patient-specific IgE repertoires for the...... manifestation and severity of allergic symptoms. OBJECTIVE: We sought to investigate how individual properties of an IgE repertoire affect effector cell degranulation. METHODS: A panel of recombinant IgE (rIgE) antibodies specific for the major house dust mite allergen Der p 2 was developed and characterized in...... non-allergen-specific IgE, more even concentration of individual allergen-specific IgE clones, increased IgE affinity for allergen, and increased number of allergen epitopes recognized by the IgE repertoire (increased IgE clonality). CONCLUSION: This study demonstrates how distinct properties of the...

  14. Contribution of a p75 interleukin 2 binding peptide to a high-affinity interleukin 2 receptor complex

    International Nuclear Information System (INIS)

    There are at least two forms of cellular receptors for interleukin 2(IL-2); one with a very high affinity and the other with a lower affinity. The authors identified a non-Tac IL-2 binding peptide with a relative molecular weight of 75,000 (p75). Cell lines bearing either the p55 Tac or the p75 peptide alone manifested low-affinity IL-2 binding, whereas a cell line bearing both peptides manifested both high- and low-affinity receptors. Cross-linking studies were performed with 125I-labeled IL-2. After the internalization of labeled IL-2 through high-affinity receptors, the p75 peptide could not be detected by cross-linking studies. Furthermore, fusion of cell membranes from low-affinity IL-2 binding cell lines bearing the Tac peptide alone with membranes from a cell line bearing the p75 peptide alone generated hybrid membranes bearing high-affinity receptors. These results suggest a multichain model for the high-affinity Il-2 receptor in which high-affinity receptors would be expressed when both Tac and p75 Il-2 binding peptides are present and associated in a receptor complex

  15. Tension-compression asymmetry in the binding affinity of membrane-anchored receptors and ligands

    Science.gov (United States)

    Xu, Guang-Kui; Liu, Zishun; Feng, Xi-Qiao; Gao, Huajian

    2016-03-01

    Cell adhesion plays a crucial role in many biological processes of cells, e.g., immune responses, tissue morphogenesis, and stem cell differentiation. An essential problem in the molecular mechanism of cell adhesion is to characterize the binding affinity of membrane-anchored receptors and ligands under different physiological conditions. In this paper, a theoretical model is presented to study the binding affinity between a large number of anchored receptors and ligands under both tensile and compressive stresses, and corroborated by demonstrating excellent agreement with Monte Carlo simulations. It is shown that the binding affinity becomes lower as the magnitude of the applied stress increases, and drops to zero at a critical tensile or compressive stress. Interestingly, the critical compressive stress is found to be substantially smaller than the critical tensile stress for relatively long and flexible receptor-ligand complexes. This counterintuitive finding is explained by using the Euler instability theory of slender columns under compression. The tension-compression asymmetry in the binding affinity of anchored receptors and ligands depends subtly on the competition between the breaking and instability of their complexes. This study helps in understanding the role of mechanical forces in cell adhesion mediated by specific binding molecules.

  16. GHB receptor targets in the CNS: Focus on high-affinity binding sites

    DEFF Research Database (Denmark)

    Bay, Tina; Eghorn, Laura Friis; Klein, Anders Bue;

    2014-01-01

    γ-Hydroxybutyric acid (GHB) is an endogenous compound in the mammalian brain with both low- and high-affinity receptor targets. GHB is used clinically in the treatment of symptoms of narcolepsy and alcoholism, but also illicitly abused as the recreational drug Fantasy. Major pharmacological effects...

  17. Evidence for a precursor of the high-affinity metastasis-associated murine laminin receptor

    DEFF Research Database (Denmark)

    Rao, C N; Castronovo, V; Schmitt, M C; Wewer, U M; Claysmith, A P; Liotta, L A; Sobel, M E

    1989-01-01

    The high-affinity cellular receptor for the basement membrane component laminin is differentially expressed during tumor invasion and metastasis. A cDNA clone encoding the murine laminin receptor was isolated and identified on the basis of sequence homology to the human laminin receptor [Wewer et......, and rat. The nascent laminin receptor predicted from the cDNA sequence is 295 amino acids long, with a molecular weight of 33,000, and contains one intradisulfide bridge, a short putative transmembrane domain, and an extracellular carboxy-terminal region which has abundant glutamic acid residues and...... multiple repeat sequences. The precursor of the laminin receptor is apparently smaller than the 67-kilodalton protein isolated from tissue. The apparent molecular weight on SDS-polyacrylamide gels of the rabbit reticulocyte cell-free translation product of selectively hybridized laminin receptor mRNA is 37...

  18. Scintigraphic studies in rats. Kinetics of insulin analogues covering wide range of receptor affinities

    International Nuclear Information System (INIS)

    Whole-body kinetics of 123I-labeled human insulin and five insulin analogues were investigated by scintigraphic studies in rats. The amino acid substitutions and the relative receptor affinities (RAs), determined by binding to HepG2 cells, were: GluB12, des-B30 insulin, RA 0.15%; AspB9, GluB27 insulin, RA 18%; AspB26 insulin, RA 80%; AspB18 insulin, RA 327%; and HisA8, HisB4, GluB10, HisB27 insulin, RA 687%. All analogues were compared with human insulin (RA 100%). The analogue with RA 0.15% showed a significantly slower disappearance in the heart window, no liver uptake, and the greatest kidney radioactivity, the latter probably caused by high plasma concentrations. The low-affinity analogue (RA 18%) reached a surprisingly high hepatic peak value, although significantly lower than insulin. Kidney radioactivity was higher than for insulin. The analogue with RA 80% showed liver and kidney radioactivities that were not significantly different from those of insulin. The two high-affinity analogues (RAs 327 and 687%) showed peak liver radioactivities not significantly different from insulin. However, liver radioactivity after the peaks declined significantly more slowly. Compared with insulin, the kidney radioactivity curves were not significantly different. We conclude that high-affinity insulin analogues will bind to any available receptor that, because of the large number of receptors in the periphery and the distribution of cardiac output, favors extrahepatic elimination. In contrast, low-affinity analogues bind to receptors several times before they are eliminated. This leads to recirculation and, thus, hepatic elimination due to the high receptor density there

  19. Point mutations of human interleukin-1 with decreased receptor binding affinity

    International Nuclear Information System (INIS)

    Interleukin-1 (IL-1) is a monocyte-derived polypeptide hormone that interacts with a plasma membrane receptor. We have used oligonucleotide-directed mutagenesis to construct mutant human IL-1 proteins. Three different point mutants in a unique histidine residue (position 30) exhibited varying degrees of reduced IL-1 receptor binding affinity, whereas point mutants at five other residues behaved normally. Structural analysis of these mutant proteins by nuclear magnetic resonance spectroscopy detected no (or only minor) conformational changes relative to wild-type IL-1. These data suggest that the unique histidine residue influences the architecture of the receptor binding site on human IL-1. (Auth.)

  20. Guanine nucleotide regulation of dopamine receptor agonist affinity states in rat estradiol-induced pituitary tumors

    Energy Technology Data Exchange (ETDEWEB)

    Di Paolo, T.; Falardeau, P.

    1987-08-31

    The authors have investigated dopamine (DA) receptor agonist high- and low-affinity states in female rate estradiol-induced prolactin (PRL)-secreting pituitary tumors and intact pituitary tissue. Estradiol treatment increased the anterior pituitary weight 9-fold and plasma prolactin levels 74-fold and these measures are correlated (R = 0.745, n = 73, p < 0.001). Competition for (/sup 3/H)-spiperone binding to the DA receptor by apomorphine was compared in normal and adenomatous pituitary tissue. The inhibition constants (Ki) and the proportions of the two apomorphine sites are unchanged in tumors compared to intact pituitary tissue. Guanosine 5'-(..beta..-..gamma..-imino)triphosphate (Gpp(NH)p) causes complete conversion of the high into low affinity dopaminergic agonist site in normal pituitary and in tumors. These results suggest that rats with primary estradiol-induced pituitary tumors have normal and functional DA receptors. 9 references, 2 tables.

  1. Guanine nucleotide regulation of dopamine receptor agonist affinity states in rat estradiol-induced pituitary tumors

    International Nuclear Information System (INIS)

    The authors have investigated dopamine (DA) receptor agonist high- and low-affinity states in female rate estradiol-induced prolactin (PRL)-secreting pituitary tumors and intact pituitary tissue. Estradiol treatment increased the anterior pituitary weight 9-fold and plasma prolactin levels 74-fold and these measures are correlated (R = 0.745, n = 73, p 3H]-spiperone binding to the DA receptor by apomorphine was compared in normal and adenomatous pituitary tissue. The inhibition constants (Ki) and the proportions of the two apomorphine sites are unchanged in tumors compared to intact pituitary tissue. Guanosine 5'-[β-γ-imino]triphosphate (Gpp(NH)p) causes complete conversion of the high into low affinity dopaminergic agonist site in normal pituitary and in tumors. These results suggest that rats with primary estradiol-induced pituitary tumors have normal and functional DA receptors. 9 references, 2 tables

  2. Molecular basis for high affinity and selectivity of peptide antagonist, Bantag-1, for the orphan BB3 receptor.

    Science.gov (United States)

    Nakamura, Taichi; Ramos-Álvarez, Irene; Iordanskaia, Tatiana; Moreno, Paola; Mantey, Samuel A; Jensen, R T

    2016-09-01

    Bombesin-receptor-subtype-3 (BB3 receptor) is a G-protein-coupled-orphan-receptor classified in the mammalian Bombesin-family because of high homology to gastrin-releasing peptide (BB2 receptor)/neuromedin-B receptors (BB1 receptor). There is increased interest in BB3 receptor because studies primarily from knockout-mice suggest it plays roles in energy/glucose metabolism, insulin-secretion, as well as motility and tumor-growth. Investigations into its roles in physiological/pathophysiological processes are limited because of lack of selective ligands. Recently, a selective, peptide-antagonist, Bantag-1, was described. However, because BB3 receptor has low-affinity for all natural, Bn-related peptides, little is known of the molecular basis of its high-affinity/selectivity. This was systematically investigated in this study for Bantag-1 using a chimeric-approach making both Bantag-1 loss-/gain-of-affinity-chimeras, by exchanging extracellular (EC) domains of BB3/BB2 receptor, and using site-directed-mutagenesis. Receptors were transiently expressed and affinities determined by binding studies. Bantag-1 had >5000-fold selectivity for BB3 receptor over BB2/BB1 receptors and substitution of the first EC-domain (EC1) in loss-/gain-of affinity-chimeras greatly affected affinity. Mutagenesis of each amino acid difference in EC1 between BB3 receptor/BB2 receptor showed replacement of His(107) in BB3 receptor by Lys(107) (H107K-BB3 receptor-mutant) from BB2 receptor, decreased affinity 60-fold, and three replacements [H107K, E11D, G112R] decreased affinity 500-fold. Mutagenesis in EC1's surrounding transmembrane-regions (TMs) demonstrated TM2 differences were not important, but R127Q in TM3 alone decreased affinity 400-fold. Additional mutants in EC1/TM3 explored the molecular basis for these changes demonstrated in EC1, particularly important is the presence of aromatic-interactions by His(107), rather than hydrogen-bonding or charge-charge interactions, for determining

  3. GABA agonist promoted formation of low affinity GABA receptors on cerebellar granule cells is restricted to early development

    DEFF Research Database (Denmark)

    Belhage, B; Hansen, Gert Helge; Schousboe, A;

    1988-01-01

    The ability of the GABA receptor agonist 4,5,6,7-tetrahydroisoxazolo[5,4-c]pyridin-3-ol (THIP) to promote formation of low affinity GABA receptors on cerebellar granule cells was tested using primary cultures of these neurons. Granule cells were exposed to THIP (150 microM) for 6 hr after......, respectively, 4, 7, 10 and 14 days in culture. It was found that THIP treatment of 4- and 7-day-old cultures led to formation of low affinity GABA receptors, whereas such receptors could not be detected after THIP treatment in the older cultures (10 and 14 days) in spite of the fact that these cultured granule...... cells expressed a high density of high affinity GABA receptors. It is concluded that the ability of THIP to promote formation of low affinity GABA receptors on cerebellar granule cells is restricted to an early developmental period....

  4. Syntheses of 7-Substituted α-Cyperone Derivatives for Selective Sigma-1 Receptor over Cannabinoid-1 Receptor Binding Affinities

    International Nuclear Information System (INIS)

    We have successfully synthesized seven α-cyperone derivatives and found that the presence of a hydrogen bond donor/acceptor groups at the C7 position of α-cyperone significantly affects specificity and potency of CB1 receptor binding affinity over sigma-1 receptor binding affinity. In particular, the presence of the amino moiety at the C7 position of α-cyperone is beneficial for binding to sigmia-1 receptor. The molecular mechanism of compound 8 involved in the high binding affinity to sigma-1 receptor is under investigation. We first synthesized α-cyperone 1 by following the previously reported synthetic routes.15-19 In brief, azeotropic imination of (+)-dihydrocarvone and (R)-(+)-1-phenylethylamine followed by alkylation with a slight excess of ethyl vinyl ketone (EVK) in THF at 40 .deg. C produced the Micheal adduct. The resulting adduct was hydrolyzed and then treated with sodium methoxide at room temperature to give an easily separable mixture of α-cyperone 1 and its side product. Flash chromatography resulted in pure α-cyperone 1 in a 30% yield from (+)-dihydrocarvone

  5. Prostaglandins E2 signal mediated by receptor subtype EP2 promotes IgE production in vivo and contributes to asthma development

    Science.gov (United States)

    Gao, Yuhan; Zhao, Chunyan; Wang, Wei; Jin, Rong; Li, Qian; Ge, Qing; Guan, Youfei; Zhang, Yu

    2016-01-01

    Prostaglandins E2 (PGE2) has been shown to enhance IgE production by B cells in vitro. The physiological and pathological relevance of this phenomenon and the underlying molecular mechanism, however, remain to be elucidated. B cells from wild type and EP2-deficient mice were compared in culture for their responses to PGE2 in terms of IgE class switching and production. Ovalbumin (OVA)-induced asthma models were used to evaluate the impact of EP2-deficiency on IgE responses and the development of asthma. PGE2 promoted IgE class switching, generation of IgE+ cells and secretion of IgE by B cells stimulated with LPS+IL4. These effects were much attenuated as a consequence of EP2 deficiency. Consistent with the in vitro data, EP2-deficient mice showed a markedly suppressed IgE antibody response and developed less pronounced airway inflammation in the OVA-induced asthma model. Mechanistic studies demonstrated that PGE2, in an EP2-depedent manner, enhanced STAT6 activation induced by IL-4, thereby promoting the expression of IgE germline and post switch transcripts and the transcription of activation-induced cytidine deaminase (AID). Collectively, these data support an important regulatory role of the PGE2-EP2-STAT6 signaling pathway in IgE response and allergic diseases. PMID:26852804

  6. Ring size in cyclic endomorphin-2 analogs modulates receptor binding affinity and selectivity.

    Science.gov (United States)

    Piekielna, Justyna; Kluczyk, Alicja; Gentilucci, Luca; Cerlesi, Maria Camilla; Calo', Girolamo; Tomböly, Csaba; Łapiński, Krzysztof; Janecki, Tomasz; Janecka, Anna

    2015-06-01

    The study reports the solid-phase synthesis and biological evaluation of a series of new side chain-to-side chain cyclized opioid peptide analogs of the general structure Tyr-[D-Xaa-Phe-Phe-Asp]NH2, where Xaa = Lys (1), Orn (2), Dab (3), or Dap (4) (Dab = 2,4-diaminobutyric acid, Dap = 2,3-diaminopropionic acid), containing 17- to 14-membered rings. The influence of the ring size on binding to the MOP, DOP and KOP opioid receptors was studied. In general, the reduction of the size of the macrocyclic ring increased the selectivity for the MOP receptor. The cyclopeptide incorporating Xaa = Lys displayed subnanomolar MOP affinity but modest selectivity over the KOP receptor, while the analog with the Orn residue showed increased affinity and selectivity for MOP. The analog with Dab was a weak MOP agonist and did not bind to the other two opioid receptors. Finally, the peptide with Xaa = Dap was completely MOP receptor-selective with subnanomolar affinity. Interestingly, the deletion of one Phe residue from 1 led to the 14-membered Tyr-c[D-Lys-Phe-Asp]NH2 (5), a potent and selective MOP receptor ligand. The in vitro potencies of the new analogs were determined in a calcium mobilization assay performed in Chinese Hamster Ovary (CHO) cells expressing human recombinant opioid receptors and chimeric G proteins. A good correlation between binding and the functional test results was observed. The influence of the ring size, solid support and the N-terminal protecting group on the formation of cyclodimers was studied. PMID:25948019

  7. Purification of high affinity benzodiazepine receptor binding site fragments from rat brain

    International Nuclear Information System (INIS)

    In central nervous system benzodiazepine recognition sites occur on neuronal cell surfaces as one member of a multireceptor complex, including recognition sites for benzodiazepines, gamma aminobutyric acid (GABA), barbiturates and a chloride ionophore. During photoaffinity labelling, the benzodiazepine agonist, 3H-flunitrazepam, is irreversibly bound to central benzodiazepine high affinity recognition sites in the presence of ultraviolet light. In these studies a 3H-flunitrazepam radiolabel was used to track the isolation and purification of high affinity agonist binding site fragments from membrane-bound benzodiazepine receptor in rat brain. The authors present a method for limited proteolysis of 3H-flunitrazepam photoaffinity labeled rat brain membranes, generating photolabeled benzodiazepine receptor fragments containing the agonist binding site. Using trypsin chymotrypsin A4, or a combination of these two proteases, they have demonstrated the extent and time course for partial digestion of benzodiazepine receptor, yielding photolabeled receptor binding site fragments. These photolabeled receptor fragments have been further purified on the basis of size, using ultrafiltration, gel permeation chromatography, and sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) as well as on the basis of hydrophobicity, using a high performance liquid chromatography (HPLC) precolumn, several HPLC elution schemes, and two different HPLC column types. Using these procedures, they have purified three photolabeled benzodiazepine receptor fragments containing the agonist binding site which appear to have a molecular weight of less than 2000 daltons each

  8. Identification of high- and low-affinity NGF receptors during development of the chicken central nervous system

    International Nuclear Information System (INIS)

    In order to study regulation of the nerve growth factor (NGF) receptor during embryogenesis in chick brain, we have used affinity crosslinking of tissues with 125I-NGF. NGF interacts with high- and low-affinity receptors; high-affinity receptors are required for the majority of NGF's actions. Most measurements of receptor levels do not distinguish between high- and low-affinity forms of the receptor. We have used the lipophilic crosslinking agent HSAB to identify the high-affinity, functional receptor during development of the chicken central nervous system. A peak of expression during Embryonic Days 5-10 was detected in all regions of the chicken central nervous system, but, shortly after birth, only the cerebellar region displays significant levels of NGF receptor protein. The time course of expression confirms the dramatic regulation of the NGF receptor gene during defined embryonic periods. The detection of high-affinity NGF receptors in brain and neural retina provides strong evidence that NGF is involved in essential ontogenetic events in the development of the chicken central nervous system

  9. Differentiation of extrastriatal dopamine D2 receptor density and affinity in the human brain using PET.

    Science.gov (United States)

    Olsson, Hans; Halldin, Christer; Farde, Lars

    2004-06-01

    Dopaminergic neurotransmission in extrastriatal regions may play a crucial role in the pathophysiology and treatment of neuropsychiatric disorders. The high-affinity radioligands [(11)C]FLB 457, [(123)I]epidepride, and [(18)F]fallypride are now used in clinical studies to measure these low-density receptor populations in vivo. However, a single determination of the regional binding potential (BP) does not differentiate receptor density (B(max)) from the apparent affinity (K(D)). In this positron emission tomography (PET) study, we measured extrastriatal dopamine D2 receptor density (B(max)) and apparent affinity (K(D)) in 10 healthy subjects using an in vivo saturation approach. Each subject participated in two to three PET measurements with different specific radioactivity of [(11)C]FLB 457. The commonly used simplified reference tissue model (SRTM) was used in a comparison of BP values with the B(max) values obtained from the saturation analysis. The calculated regional receptor density values were of the same magnitude (0.33-1.68 nM) and showed the same rank order as reported from postmortem studies, that is, in descending order thalamus, lateral temporal cortex, anterior cinguli, and frontal cortex. The affinity ranged from 0.27 to 0.43 nM, that is, approximately 10-20 times the value found in vitro (20 pM). The area under the cerebellar time activity curve (TAC) was slightly lower (11 +/- 8%, mean +/- SD, P = 0.004, n = 10) after injection of low as compared with high specific radioactivity, indicating sensitivity to the minute density of dopamine D2 receptors in the this region. The results of the present study support that dopamine D2 receptor density and affinity can be differentiated in low-density regions using a saturation approach. There was a significant (P < 0.001) correlation between the binding potential calculated with SRTM and the receptor density (B(max)), which supports the use of BP in clinical studies where differentiation of B(max) and K

  10. Affinity and kinetics study of anthranilic acids as HCA2 receptor agonists.

    Science.gov (United States)

    van Veldhoven, Jacobus P D; Liu, Rongfang; Thee, Stephanie A; Wouters, Yessica; Verhoork, Sanne J M; Mooiman, Christiaan; Louvel, Julien; IJzerman, Adriaan P

    2015-07-15

    Structure-affinity relationship (SAR) and structure-kinetics relationship (SKR) studies were combined to investigate a series of biphenyl anthranilic acid agonists for the HCA2 receptor. In total, 27 compounds were synthesized and twelve of them showed higher affinity than nicotinic acid. Two compounds, 6g (IC50=75nM) and 6z (IC50=108nM) showed a longer residence time profile compared to nicotinic acid, exemplified by their kinetic rate index (KRI) values of 1.31 and 1.23, respectively. The SAR study resulted in the novel 2-F, 4-OH derivative (6x) with an IC50 value of 23nM as the highest affinity HCA2 agonist of the biphenyl series, although it showed a similar residence time as nicotinic acid. The SAR and SKR data suggest that an early compound selection based on binding kinetics is a promising addition to the lead optimization process. PMID:25737085

  11. Further characterization of the subunits of the receptor with high affinity for immunoglobulin E

    International Nuclear Information System (INIS)

    The α, β, γ subunits of the receptor with high affinity for immunoglobulin E were isolated and their compositions assessed by direct amino acid analysis and by incorporation of radioactive precursors. The compositions show no unusual features other than a rather high content of tryptophan in the α chain as assessed from the incorporation studies. The results combined with future sequence data will permit unambiguous determination of the multiplicity of the chains in the receptor. Chymotryptic peptide maps of the extrinsically iodinated subunits show several similar peptides, particularly for α and β. However, these putative homologies were not apparent when tryptic maps of the biosynthetically ([3H]leucine) labeled subunits were analyzed

  12. Rational development of high-affinity T-cell receptor-like antibodies.

    Science.gov (United States)

    Stewart-Jones, Guillaume; Wadle, Andreas; Hombach, Anja; Shenderov, Eugene; Held, Gerhard; Fischer, Eliane; Kleber, Sascha; Nuber, Natko; Stenner-Liewen, Frank; Bauer, Stefan; McMichael, Andrew; Knuth, Alexander; Abken, Hinrich; Hombach, Andreas A; Cerundolo, Vincenzo; Jones, E Yvonne; Renner, Christoph

    2009-04-01

    T-cell interaction with a target cell is a key event in the adaptive immune response and primarily driven by T-cell receptor (TCR) recognition of peptide-MHC (pMHC) complexes. TCR avidity for a given pMHC is determined by number of MHC molecules, availability of coreceptors, and TCR affinity for MHC or peptide, respectively, with peptide recognition being the most important factor to confer target specificity. Here we present high-resolution crystal structures of 2 Fab antibodies in complex with the immunodominant NY-ESO-1(157-165) peptide analogue (SLLMWITQV) presented by HLA-A*0201 and compare them with a TCR recognizing the same pMHC. Binding to the central methionine-tryptophan peptide motif and orientation of binding were almost identical for Fabs and TCR. As the MW "peg" dominates the contacts between Fab and peptide, we estimated the contributions of individual amino acids between the Fab and peptide to provide the rational basis for a peptide-focused second-generation, high-affinity antibody library. The final Fab candidate achieved better peptide binding by 2 light-chain mutations, giving a 20-fold affinity improvement to 2-4 nM, exceeding the affinity of the TCR by 1,000-fold. The high-affinity Fab when grafted as recombinant TCR on T cells conferred specific killing of HLA-A*0201/NY-ESO-1(157-165) target cells. In summary, we prove that affinity maturation of antibodies mimicking a TCR is possible and provide a strategy for engineering high-affinity antibodies that can be used in targeting specific pMHC complexes for diagnostic and therapeutic purposes. PMID:19307587

  13. Decrease in IgE Fc receptor expression on mouse bone marrow-derived mast cells and inhibition of PAF-acether formation and of β-hexosaminidase release by dexamethasone

    International Nuclear Information System (INIS)

    The effect of dexamethasone (DM) on the immunologic and nonimmunologic release of paf-acether and of the granule marker β-hexosaminidase (BHEX) from mouse bone marrow-derived mast cells (BMMC) was studied. Preincubation of BMMC with DM inhibited in a dose-dependent fashion the immunologic release of paf-acether and of BHEX as compared with control cells. The antigen-induced increase in acetyltransferase activity, used as an index of cellular activation, was inhibited by 37 +/- 16% in 1 μM DM-treated BMMC as compared with untreated cells. Preincubation of BMMC with DM for 24 hr caused a dose-dependent inhibition of 125I-IgE binding to the cells, with a half-maximal effect at 14 nM. The number of IgE Fc receptors was decreased by 55% in 1 μM DM-treated BMMC as compared with untreated cells. Cytofluorometer analysis of BMMC sensitized with a saturating amount of purified monoclonal IgE, followed by addition of a fluoresceinated anti-mouse IgG (heavy and light chains), revealed a single cellular population for both DM-treated and untreated BMMC. The possible link between the decreased sensitization of the cells consequent to the reduction in IgG Fc receptor expression and the alteration of the secretory response and acetyltransferase activity was investigated. BMMC were incubated with IgE under experimental conditions giving half-sensitization of the cells. Upon antigen challenge, a 10.5 +/- 3.7% decrease in acetyltransferase activity and a 29.2 +/- 3.5% decrease in paf-acether release were observed with half-sensitized cells as compared with cells sensitized with a saturating amount of IgE. These results indicate that DM inhibits the immunologic release of paf-acether and of BHEX from passively sensitized BMMC and decreases the IgE Fc receptor number available for sensitization. Thus, the modulation of IgE Fc receptor number could explain part of the anti-allergic properties of glucocorticosteroids

  14. Computational estimation of rainbow trout estrogen receptor binding affinities for environmental estrogens

    International Nuclear Information System (INIS)

    Environmental estrogens have been the subject of intense research due to their documented detrimental effects on the health of fish and wildlife and their potential to negatively impact humans. A complete understanding of how these compounds affect health is complicated because environmental estrogens are a structurally heterogeneous group of compounds. In this work, computational molecular dynamics simulations were utilized to predict the binding affinity of different compounds using rainbow trout (Oncorhynchus mykiss) estrogen receptors (ERs) as a model. Specifically, this study presents a comparison of the binding affinity of the natural ligand estradiol-17β to the four rainbow trout ER isoforms with that of three known environmental estrogens 17α-ethinylestradiol, bisphenol A, and raloxifene. Two additional compounds, atrazine and testosterone, that are known to be very weak or non-binders to ERs were tested. The binding affinity of these compounds to the human ERα subtype is also included for comparison. The results of this study suggest that, when compared to estradiol-17β, bisphenol A binds less strongly to all four receptors, 17α-ethinylestradiol binds more strongly, and raloxifene has a high affinity for the α subtype only. The results also show that atrazine and testosterone are weak or non-binders to the ERs. All of the results are in excellent qualitative agreement with the known in vivo estrogenicity of these compounds in the rainbow trout and other fishes. Computational estimation of binding affinities could be a valuable tool for predicting the impact of environmental estrogens in fish and other animals.

  15. Development of novel cellular model for affinity studies of histamine H(4) receptor ligands.

    Science.gov (United States)

    Karcz, Tadeusz; Kieć-Kononowicz, Katarzyna

    2013-01-01

    The G protein-coupled histamine H4 receptor (H4R) is the last member of histamine receptors family discovered so far. Its expression pattern, together with postulated involvement in a wide variety of immunological and inflammatory processes make histamine H4 receptor an interesting target for drug development. Potential H4R ligands may provide an innovative therapies for different immuno-based diseases, including allergy, asthma, pruritus associated with allergy or autoimmune skin conditions, rheumatoid arthritis and pain. However, none of successfully developed selective and potent histamine H4 receptor ligands have been introduced to the market up to date. For that reason there is still a strong demand for pharmacological models to be used in studies on potent H4R ligands. In current work we present the development of novel mammalian cell line, stably expressing human histamine H4 receptor, with use of retroviral transduction approach. Obtained cell line was pharmacologically characterized in radioligand binding studies and its utility for affinity testing of potent receptor ligands was confirmed in comparative studies with the use of relevant insect cells expression model. Obtained results allow for statement that developed cellular model may be successfully employed in search for new compounds active at histamine H4 receptor. PMID:24432340

  16. Sensitivity of binding of high-affinity dopamine receptor radioligands to increased synaptic dopamine.

    Science.gov (United States)

    Gatley, S J; Gifford, A N; Carroll, F I; Volkow, N D

    2000-12-15

    PET and SPECT studies have documented that D2 radioligands of moderate affinity, but not radioligands of high affinity, are sensitive to pharmacological challenges that alter synaptic dopamine levels. The objective of this work was to determine whether the brain kinetics of high-affinity radioligands for dopamine D1 ([(3)H]SCH 23390) and D2 ([(123)I]epidepride) receptors were altered by a prolonged elevation of synaptic dopamine induced by the potent cocaine analog RTI-55. Mice were injected intravenously with radioligands either 30 min after or 4 h before intraperitoneal administration of RTI-55 (2 mg/kg). In separate experiments, the pharmacological effects of RTI-55 were assessed biochemically by measuring uptake of dopamine in synaptosomes prepared from RTI-treated mice and behaviorally by monitoring locomotor activity. Consistent with the expected elevation of synaptic dopamine, RTI-55 induced a long-lasting decrement in dopamine uptake measured ex vivo, and a prolonged increase in locomotor activity. RTI-55 injected prior to the radioligands induced a significant (P epidepride at 15 min, relative to saline-treated controls, but there were no differences between the two groups at later time-points. For [(3)H]SCH 23390, both initial striatal uptake and subsequent clearance were slightly increased by preadministration of RTI-55. Administration of RTI-55 4 h after the radioligands (i.e., when it was presumed that a state of near equilibrium binding of the radioligands had been reached), was associated with a significant reduction of striatal radioactivity for both radiotracers. Our results are consistent with increased competition between dopamine and radioligand for binding to both D1 and D2 receptors after treatment with RTI-55. We suggest that the magnitude of the competition is reduced by failure of the receptor binding of high-affinity radioligands to rapidly attain equilibrium. PMID:11044896

  17. Significant receptor affinities of metabolites and a degradation product of mometasone furoate

    Directory of Open Access Journals (Sweden)

    Högger Petra

    2004-07-01

    Full Text Available Abstract Mometasone furoate (MF is a highly potent glucocorticoid used topically to treat inflammation in the lung, nose and on the skin. However, so far no information has been published on the human glucocorticoid receptor activity of the metabolites or degradation products of MF. We have now determined the relative receptor binding affinities of the known metabolite 6β-OH MF and the degradation product 9,11-epoxy MF to understand their possible contribution to undesirable systemic side effects. In competition experiments with human lung glucocorticoid receptors we have determined the relative receptor affinities (RRA of these substances with reference to dexamethasone (RRA = 100. We have discovered that 6β-OH MF and 9,11-epoxy MF display RRAs of 206 ± 15 and 220 ± 22, respectively. This level of activity is similar to that of the clinically used inhaled corticosteroid flunisolide (RRA 180 ± 11. Furthermore we observed that 9,11-epoxy MF is a chemically reactive metabolite. In recovery experiments with human plasma and lung tissue we found a time dependent decrease in extractability of the compound. Hence, we provide data that might contribute to the understanding of the pharmacokinetics as well as the clinical effects of MF.

  18. Human receptor kinetics and lung tissue retention of the enhanced-affinity glucocorticoid fluticasone furoate

    Directory of Open Access Journals (Sweden)

    Högger Petra

    2007-07-01

    Full Text Available Abstract Fluticasone furoate (FF – USAN approved name, a new topically active glucocorticoid has been recently identified. The aim of this study was to characterise the binding affinity of this compound to the human lung glucocorticoid receptor in relation to other glucocorticoids. Additionally, we sought to determine the binding behaviour of fluticasone furoate to human lung tissue. The glucocorticoid receptor binding kinetics of fluticasone furoate revealed a remarkably fast association and a slow dissociation resulting in a relative receptor affinity (RRA of 2989 ± 135 with reference to dexamethasone (RRA: 100 ± 5. Thus, the RRA of FF exceeds the RRAs of all currently clinically used corticosteroids such as mometasone furoate (MF; RRA 2244, fluticasone propionate (FP; RRA 1775, ciclesonide's active metabolite (RRA 1212 – rat receptor data or budesonide (RRA 855. FP and FF displayed pronounced retention in human lung tissue in vitro. Lowest tissue binding was found for MF. There was no indication of instability or chemical modification of FF in human lung tissue. These advantageous binding attributes may contribute to a highly efficacious profile for FF as a topical treatment for inflammatory disorders of the respiratory tract.

  19. Evidence that the angiotensin at 2-receptor agonist compound 21 is also a low affinity thromboxane TXA2-receptor antagonist

    DEFF Research Database (Denmark)

    Fredgart, M.; Leurgans, T.; Stenelo, M.;

    2015-01-01

    Objective: The objective of this study was to test whether Compound 21 (C21), a high-affinity, non-peptide angiotensinAT2-receptor agonist, is also an antagonist of thromboxane A2 (TXA2) receptors thus reducing both vasoconstriction and platelet aggregation. Design and method: Binding of C21 to the...... TXA2 receptor was determined by TBXA2R Arrestin Biosensor Assay. Mouse mesenteric arteries were mounted in wire myographs, and responses to increasing concentrations of C21 (1nM- 10muM) were recorded during submaximal contractions with 0.1muM U46619 (TXA2 analogue) or 1muMphenylephrine. To control for...

  20. Vaccination for birch pollen allergy. Induction of affinity-matured or blocking IgG antibodies does not account for the reduced binding of IgE to Bet v 1

    DEFF Research Database (Denmark)

    Svenson, Morten; Jacobi, Henrik H; Bødtger, Uffe; Poulsen, Lars K; Rieneck, Klaus; Bendtzen, Klaus

    2003-01-01

    (Betula verrucosa) (Bet v 1), the major allergen in birch pollen, to serum IgG and IgE, separately and in competition. Sera from six birch pollen-allergic patients were obtained before and after 5 years of SAV, and binding was assessed with 125I-Bet v 1. Before SAV, IgG bound more than eight times the...... amount of Bet v 1 compared with IgE, and together they accounted for more than 85% of the serum binding capacity. While SAV induced minimal changes in IgE binding, the IgG binding capacities increased 6-32 times. In contrast, the binding avidities (K(d) 28-40pM) changed less than 20%, pre- and post......-SAV IgG provided similar inhibition of Bet v 1 binding to IgE at equimolar levels, and cross inhibition studies between IgG and IgE showed low inter-individual differences. Following SAV, all sera reduced Bet v 1 binding to CD23(+) cells, correlating with reduced binding of Bet v 1 to IgE (P...

  1. Isolation, Affinity Purification, and Identification of Piglet Small Intestine Mucosa Receptor for Enterotoxigenic Escherichia coli K88ac+ Fimbriae

    OpenAIRE

    Fang, Lin; Gan, Zhibo; Marquardt, Ronald R.

    2000-01-01

    An affinity chromatography technique was utilized to isolate and purify the receptors of Escherichia coli K88ac+ fimbriae from the mucus of the small intestines of newborn piglets. Purified K88ac+ fimbriae were covalently immobilized onto a beaded agarose matrix (Sepharose 4B). The immobilized fimbriae were used for the affinity purification of the K88ac+ receptors. Only two major proteins were tightly and specifically bound to the immobilized fimbriae after the column containing bound recept...

  2. Structure-based rational design of a Toll-like receptor 4 (TLR4 decoy receptor with high binding affinity for a target protein.

    Directory of Open Access Journals (Sweden)

    Jieun Han

    Full Text Available Repeat proteins are increasingly attracting much attention as alternative scaffolds to immunoglobulin antibodies due to their unique structural features. Nonetheless, engineering interaction interface and understanding molecular basis for affinity maturation of repeat proteins still remain a challenge. Here, we present a structure-based rational design of a repeat protein with high binding affinity for a target protein. As a model repeat protein, a Toll-like receptor4 (TLR4 decoy receptor composed of leucine-rich repeat (LRR modules was used, and its interaction interface was rationally engineered to increase the binding affinity for myeloid differentiation protein 2 (MD2. Based on the complex crystal structure of the decoy receptor with MD2, we first designed single amino acid substitutions in the decoy receptor, and obtained three variants showing a binding affinity (K(D one-order of magnitude higher than the wild-type decoy receptor. The interacting modes and contributions of individual residues were elucidated by analyzing the crystal structures of the single variants. To further increase the binding affinity, single positive mutations were combined, and two double mutants were shown to have about 3000- and 565-fold higher binding affinities than the wild-type decoy receptor. Molecular dynamics simulations and energetic analysis indicate that an additive effect by two mutations occurring at nearby modules was the major contributor to the remarkable increase in the binding affinities.

  3. Modulation of Opioid Receptor Ligand Affinity and Efficacy Using Active and Inactive State Receptor Models

    OpenAIRE

    Anand, Jessica P.; Purington, Lauren C.; Pogozheva, Irina D.; Traynor, John R.; Mosberg, Henry I.

    2012-01-01

    Mu opioid receptor (MOR) agonists are widely used for the treatment of pain; however chronic use results in the development of tolerance and dependence. It has been demonstrated that co-administration of a MOR agonist with a delta opioid receptor (DOR) antagonist maintains the analgesia associated with MOR agonists, but with reduced negative side effects. Using our newly refined opioid receptor models for structure-based ligand design, we have synthesized several pentapeptides with tailored a...

  4. Affinity capture of (Arg sup 8 )vasopressin-receptor complex using immobilized antisense peptide

    Energy Technology Data Exchange (ETDEWEB)

    Feng Xian Lu; Aiyar, N.; Chaiken, I. (SmithKline Beecham, King of Prussia, PA (United States))

    1991-05-01

    Solubilized noncovalent complexes of (Arg{sup 8})-vasopressin (AVP) with receptor proteins from rat liver membranes were isolated by selective binding to silica-immobilized antisense (AS) peptide. The affinity chromatographic support was prepared with a chemically synthesized AS peptide whose sequence is encoded by the AS DNA corresponding to the 20 amino-terminal residues of the AVP bovine neurophysin II biosynthetic precursor (pro-AVP/BNPII-(20-1)), region that includes the AVP sequence at residues 1-9. The AS peptide-AVP interaction mechanism hypothesized, contact by hydropathic complementarity at multiple sites along the peptide chains, led to the prediction that AVP bound to its receptor would still have enough free surface to interact with immobilized AS peptide. To test this prediction of a three-way interaction, ({sup 3}H)AVP-receptor was obtained as a solubilized, partially purified fraction from rat liver membrane. Covalently crosslinked ({sup 3}H)AVP complex also was bound to the AS peptide column; binding was blocked by competition with unlabeled AVP in the elution buffer. Since the AVP-linked 31- and 38-kDa proteins have the same apparent molecular mass on SDS/PAGE as found previously by photoaffinity labeling, the authors conclude that the AS peptide column has affinity-captured AVP-receptor complexes. The 15-kDa protein appears to be an active AVP-receptor fragment of one or both of the larger proteins. It is generally concluded that immobilized AS peptides may be useful to isolate peptide and protein receptor complexes in other systems as well.

  5. Expression of high affinity receptors for murine interleukin 4 (BSF-1) on hemopoietic and nonhemopoietic cells

    International Nuclear Information System (INIS)

    In this report a method for the affinity purification and radiolabeling of recombinant mouse interleukin (IL)-4 is described. It is shown on the basis of several criteria that IL-4 retains full biologic activity after radioiodination and can therefore be used as a valid model for measuring the binding characteristics of native IL-4. By using Scatchard plot analysis of equilibrium binding data, it is demonstrated that 125I-IL-4 binds to a high affinity cell surface receptor which is expressed by both hemopoietic and nonhemopoietic cells. The dissociation constant for 125I-IL-4 (Kd = 20 to 60 pM) corresponds to the concentration of IL-4 which gives 50% biologic activity (i.e., 10 to 30 pM). Binding of 125I-IL-4 is rapid (t1/2 of 2 min), whereas dissociation occurs at a slow rate (t1/2 approximately 4 hr). The IL-4 receptor shows a high degree of specificity. Whereas unlabeled mouse IL-4 competed with mouse 125I-IL-4 in an equimolar fashion for binding to IL-4 receptors, several other lymphokines, including mouse IL-2, IL-3, interferon-gamma, granulocyte-macrophage colony-stimulating factor, and human IL-1, IL-2, and IL-4 were unable to inhibit, even at molar excesses of 400 to 800-fold. At 37 degrees C, 125I-IL-4 is rapidly internalized (approximately 200 molecules/cell/min) by HT-2 cells, with at least 85% of cell surface receptors being functional in this respect. Receptors for IL-4 were found to be expressed by subclasses of T and B cells, mast cells, macrophages, and by cells of the myeloid and erythroid lineages. This wide distribution of receptor expression closely matches the known spectrum of biologic activities of IL-4, including proliferation and/or differentiation of T and B cells, mast cells and granulocytes, and induction of macrophage antigen-presenting capacity

  6. Network-of-queues approach to B-cell-receptor affinity discrimination

    Science.gov (United States)

    Felizzi, Federico; Comoglio, Federico

    2012-06-01

    The immune system is one of the most complex signal processing machineries in biology. The adaptive immune system, consisting of B and T lymphocytes, is activated in response to a large spectrum of pathogen antigens. B cells recognize and bind the antigen through B-cell receptors (BCRs) and this is fundamental for B-cell activation. However, the system response is dependent on BCR-antigen affinity values that span several orders of magnitude. Moreover, the ability of the BCR to discriminate between affinities at the high end (e.g., 109M-1-1010M-1) challenges the formulation of a mathematical model able to robustly separate these affinity-dependent responses. Queuing theory enables the analysis of many related processes, such as those resulting from the stochasticity of protein binding and unbinding events. Here we define a network of queues, consisting of BCR early signaling states and transition rates related to the propensity of molecular aggregates to form or disassemble. By considering the family of marginal distributions of BCRs in a given signaling state, we report a significant separation (measured as Jensen-Shannon divergence) that arises from a broad spectrum of antigen affinities.

  7. Structural correlates of affinity in fetal versus adult endplate nicotinic receptors

    Science.gov (United States)

    Nayak, Tapan Kumar; Chakraborty, Srirupa; Zheng, Wenjun; Auerbach, Anthony

    2016-04-01

    Adult-type nicotinic acetylcholine receptors (AChRs) mediate signalling at mature neuromuscular junctions and fetal-type AChRs are necessary for proper synapse development. Each AChR has two neurotransmitter binding sites located at the interface of a principal and a complementary subunit. Although all agonist binding sites have the same core of five aromatic amino acids, the fetal site has ~30-fold higher affinity for the neurotransmitter ACh. Here we use molecular dynamics simulations of adult versus fetal homology models to identify complementary-subunit residues near the core that influence affinity, and use single-channel electrophysiology to corroborate the results. Four residues in combination determine adult versus fetal affinity. Simulations suggest that at lower-affinity sites, one of these unsettles the core directly and the others (in loop E) increase backbone flexibility to unlock a key, complementary tryptophan from the core. Swapping only four amino acids is necessary and sufficient to exchange function between adult and fetal AChRs.

  8. Ligand binding affinities of arctigenin and its demethylated metabolites to estrogen receptor alpha.

    Science.gov (United States)

    Jin, Jong-Sik; Lee, Jong-Hyun; Hattori, Masao

    2013-01-01

    Phytoestrogens are defined as plant-derived compounds with estrogen-like activities according to their chemical structures and activities. Plant lignans are generally categorized as phytoestrogens. It was reported that (-)-arctigenin, the aglycone of arctiin, was demethylated to (-)-dihydroxyenterolactone (DHENL) by Eubacterium (E.) sp. ARC-2. Through stepwise demethylation, E. sp. ARC-2 produced six intermediates, three mono-desmethylarctigenins and three di-desmethylarctigenins. In the present study, ligand binding affinities of (-)-arctigenin and its seven metabolites, including DHENL, were investigated for an estrogen receptor alpha, and found that demethylated metabolites had stronger binding affinities than (-)-arctigenin using a ligand binding screen assay method. The IC(50) value of (2R,3R)-2-(4-hydroxy-3-methoxybenzyl)-3-(3,4-dihydroxybenzyl)-butyrolactone was 7.9 × 10⁻⁴ M. PMID:23325100

  9. Ligand Binding Affinities of Arctigenin and Its Demethylated Metabolites to Estrogen Receptor Alpha

    Directory of Open Access Journals (Sweden)

    Masao Hattori

    2013-01-01

    Full Text Available Phytoestrogens are defined as plant-derived compounds with estrogen-like activities according to their chemical structures and activities. Plant lignans are generally categorized as phytoestrogens. It was reported that (−-arctigenin, the aglycone of arctiin, was demethylated to (−-dihydroxyenterolactone (DHENL by Eubacterium (E. sp. ARC-2. Through stepwise demethylation, E. sp. ARC-2 produced six intermediates, three mono-desmethylarctigenins and three di-desmethylarctigenins. In the present study, ligand binding affinities of (−-arctigenin and its seven metabolites, including DHENL, were investigated for an estrogen receptor alpha, and found that demethylated metabolites had stronger binding affinities than (−-arctigenin using a ligand binding screen assay method. The IC50 value of (2R,3R-2-(4-hydroxy-3-methoxybenzyl-3-(3,4-dihydroxybenzyl-butyrolactone was 7.9 × 10−4 M.

  10. Receptor binding profiles and quantitative structure-affinity relationships of some 5-substituted-N,N-diallyltryptamines.

    Science.gov (United States)

    Cozzi, Nicholas V; Daley, Paul F

    2016-02-01

    N,N-Diallyltryptamine (DALT) and 5-methoxy-N,N-diallyltryptamine (5-MeO-DALT) are two tryptamines synthesized and tested by Alexander Shulgin. In self-experiments, 5-MeO-DALT was reported to be psychoactive in the 12-20mg range, while the unsubstituted compound DALT had few discernible effects in the 42-80 mg range. Recently, 5-MeO-DALT has been used in nonmedical settings for its psychoactive effects, but these effects have been poorly characterized and little is known of its pharmacological properties. We extended the work of Shulgin by synthesizing additional 5-substituted-DALTs. We then compared them to DALT and 5-MeO-DALT for their binding affinities at 45 cloned receptors and transporter proteins. Based on in vitro binding affinity, we identified 27 potential receptor targets for the 5-substituted-DALT compounds. Five of the DALT compounds had affinity in the 10-80 nM range for serotonin 5-HT1A and 5-HT2B receptors, while the affinity of DALT itself at 5-HT1A receptors was slightly lower at 100 nM. Among the 5-HT2 subtypes, the weakest affinity was at 5-HT2A receptors, spanning 250-730 nM. Five of the DALT compounds had affinity in the 50-400 nM range for serotonin 5-HT1D, 5-HT6, and 5-HT7 receptors; again, it was the unsubstituted DALT that had the weakest affinity at all three subtypes. The test drugs had even weaker affinity for 5-HT1B, 5-HT1E, and 5-HT5A subtypes and little or no affinity for the 5-HT3 subtype. These compounds also had generally nanomolar affinities for adrenergic α2A, α2B, and α2C receptors, sigma receptors σ1 and σ2, histamine H1 receptors, and norepinephrine and serotonin uptake transporters. They also bound to other targets in the nanomolar-to-low micromolar range. Based on these binding results, it is likely that multiple serotonin receptors, as well as several nonserotonergic sites are important for the psychoactive effects of DALT drugs. To learn whether any quantitative structure-affinity relationships existed, we evaluated

  11. Proliferative responses and binding properties of hematopoietic cells transfected with low-affinity receptors for leukemia inhibitory factor, oncostatin M, and ciliary neurotrophic factor.

    OpenAIRE

    Gearing, D P; Ziegler, S F; Comeau, M R; Friend, D; Thoma, B; Cosman, D; Park, L.; Mosley, B

    1994-01-01

    Specific low-affinity receptors for leukemia inhibitory factor (LIF), oncostatin M (OSM; gp130), and ciliary neurotrophic factor (CNTF; receptor alpha, CNTFR alpha) may be utilized in various combinations to generate high-affinity binding sites and signal transduction. We have tested the ability of combinations of these receptors to transduce a proliferative signal in BAF-B03 cells. Coexpression of the LIF receptor and gp130 in these cells conferred high-affinity LIF and OSM binding and respo...

  12. Mast cells and IgE in defense against venoms: Possible “good side” of allergy?

    Directory of Open Access Journals (Sweden)

    Stephen J. Galli

    2016-01-01

    Full Text Available Physicians think of mast cells and IgE primarily in the context of allergic disorders, including fatal anaphylaxis. This ‘bad side’ of mast cells and IgE is so well accepted that it can be difficult to think of them in other contexts, particularly those in which they may have beneficial functions. However, there is evidence that mast cells and IgE, as well as basophils (circulating granulocytes whose functions partially overlap with those of mast cells, can contribute to host defense as components of adaptive type 2 immune responses to helminths, ticks and certain other parasites. Accordingly, allergies often are conceptualized as “misdirected” type 2 immune responses, in which IgE antibodies are produced against any of a diverse group of apparently harmless antigens, as well as against components of animal venoms. Indeed, certain unfortunate patients who have become sensitized to venoms develop severe IgE-associated allergic reactions, including fatal anaphylaxis, upon subsequent venom exposure. In this review, we will describe evidence that mast cells can enhance innate resistance to reptile or arthropod venoms during a first exposure to such venoms. We also will discuss findings indicating that, in mice which survive an initial encounter with venom, acquired type 2 immune responses, IgE antibodies, the high affinity IgE receptor (FcɛRI, and mast cells can contribute to acquired resistance to the lethal effects of both honeybee venom and Russell's viper venom. These findings support the hypothesis that mast cells and IgE can help protect the host against venoms and perhaps other noxious substances.

  13. Lack of dopamine D4 receptor affinity contributes to the procognitive effect of lurasidone.

    Science.gov (United States)

    Murai, Takeshi; Nakako, Tomokazu; Ikeda, Kazuhito; Ikejiri, Masaru; Ishiyama, Takeo; Taiji, Mutsuo

    2014-03-15

    We previously demonstrated among several antipsychotics exhibiting potent dopamine D2 receptor antagonism that only lurasidone, (1R,2S,3R,4S)-N-[(1R,2R)-2-[4-(1,2-benzisothiazol-3-yl)-1-piperazinylmethyl]-1-cyclohexylmethyl]-2,3-bicyclo[2.2.1] heptanedicarboximide hydrochloride, improved performance in the object retrieval detour (ORD) task by marmosets. The mechanisms by which only lurasidone causes enhancements in cognitive function have not yet been established; however, most antipsychotics, except for lurasidone, have been shown to exhibit potent antagonistic activity against the dopamine D4 receptor. The objectives of this study were to evaluate the role of the dopamine D4 receptor on executive function with the selective agonist, Ro10-5824 and antagonist, L-745,870, and elucidate a possible mechanism for the procognitive effect of lurasidone. The effects of these drugs were evaluated in naïve marmosets using the ORD task. Changes in the success rate during the difficult trial in the task were used to assess the cognitive effect of the drugs. Ro10-5824 (0.3-3 mg/kg) increased the success rate in the difficult trial, potentiated the effect of lurasidone, and reversed the cognitive impairment induced by clozapine. Interestingly, the co-administration of L-745,870 with lurasidone decreased the success rate in the difficult trial, whereas the single administration of L-745,870 had no effect. These results suggest that activation of the dopamine D4 receptor may improve executive function, whereas concomitant blockade of dopamine D4 and D2 receptors may have the opposite effect. In addition to the other unique binding profiles of other monoamine receptors, the lack of affinity for the dopamine D4 receptor by lurasidone could also contribute, at least partly, to its cognitive-enhancing effect. PMID:24304719

  14. Localization of the binding site for the human high-affinity Fc receptor on IgG.

    Science.gov (United States)

    Duncan, A R; Woof, J M; Partridge, L J; Burton, D R; Winter, G

    1988-04-01

    A major pathway in the clearance of pathogens involves the coating of the pathogen with specific antibodies, and the binding of the antibody Fc region to cell receptors. This can trigger engulfment of the pathogen by phagocytes or lysis by killer cells. By oligonucleotide site-directed mutagenesis we have engineered a single amino acid change in a mouse IgG2b antibody (Glu 235----Leu) which now enables the antibody to bind to the FcRI (high affinity) receptor on human monocytes with a 100-fold improvement in affinity. This indicates that Leu 235 is a major determinant in the binding of antibody to FcRI and that the receptor may interact directly with the region linking the CH2 domain to the hinge. Tailoring the affinity of antibodies for cell receptors could help dissect their role in clearing pathogen. PMID:2965792

  15. 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

  16. Differences in affinity of cardiac beta-adrenergic receptors for [3H]dihydroalprenolol

    International Nuclear Information System (INIS)

    We performed quantitative light microscopic autoradiography of [3H]dihydroalprenolol (DHA) binding to frozen sections of canine myocardium to test the hypothesis that there are differences in the density or affinity of beta-adrenergic receptors on various tissue compartments. In one study, with concentrations of [3H]DHA from 0.34 to 5.1 nM, specific binding to cardiac myocytes was saturable, whereas nonspecific binding was linear with ligand concentration. Arterioles had more specific grain counts than muscle cells (P less than 0.0001), and Scatchard analysis showed that the arterioles had a much higher affinity for [3H]DHA than myocytes. In a second study with lower concentrations of [3H]DHA (0.19-1.98 nM), binding to the arterioles saturated, whereas binding to the cardiac myocytes did not. Specific binding to arterioles was significantly higher (P less than 0.0001) than binding to myocytes at all concentrations of [3H]DHA. The dissociation constants for the subendocardial and subepicardial myocytes were 1.57 and 1.71 nM, respectively, while the dissociation constant for the arterioles was 0.26 nM. The maximum number of binding sites was 911 grains/0.9 X 10(-2) mm2 for subepicardial myocytes, 936 for subendocardial myocytes, and 986 for arterioles. The large nerves accompanying an epicardial artery also demonstrated specific [3H]DHA binding. Thus this study has demonstrated major differences in the distribution and affinity of beta-adrenergic receptors, which may help to explain various physiological responses to beta-adrenergic stimulation

  17. High affinity dopamine D2 receptor radioligands. 1. Regional rat brain distribution of iodinated benzamides

    Energy Technology Data Exchange (ETDEWEB)

    Kessler, R.M.; Ansari, M.S.; de Paulis, T.; Schmidt, D.E.; Clanton, J.A.; Smith, H.E.; Manning, R.G.; Gillespie, D.; Ebert, M.H. (Vanderbilt University School of Medicine, Nashville, TN (USA))

    1991-08-01

    Five 125I-labeled substituted benzamides, which are close structural analogues of (S)-sulpiride, eticlopride, and isoremoxipride, were evaluated for their selective in vivo uptake into dopamine D2 receptor rich tissue of the rat brain. Iodopride (KD 0.88 nM), an iodine substituted benzamide structurally related to sulpiride, displayed a maximal striatum: cerebellar uptake ratio of 7.6. Demonstration of saturation of the receptor with (125I)iodopride in striatum required uptake in frontal cortex to be used, rather than cerebellar uptake, to define nonspecific binding. Two other ligands structurally related to eticlopride, iclopride (KD 0.23 nM) and itopride (KD 0.16 nM), displayed maximal striatal: cerebellar uptake ratios of 9.8 and 3.3, respectively. The most potent ligands, epidepride (KD 0.057 nM) and ioxipride (KD 0.070 nM) showed striatal:cerebellar uptake ratios of 234 and 65, respectively. The observed uptake ratios correlated poorly with the affinity constants for the dopamine D2 receptor alone, but were highly correlated (r = 0.92) with the product of the receptor dissociation constant (KD) and the apparent lipophilicity (kw), as determined by reverse-phase HPLC at pH 7.5. Total striatal uptake also appeared dependent on lipophilicity, with maximal uptake occurring for ligands having log kw 2.4-2.8.

  18. High affinity dopamine D2 receptor radioligands. 1. Regional rat brain distribution of iodinated benzamides

    International Nuclear Information System (INIS)

    Five 125I-labeled substituted benzamides, which are close structural analogues of (S)-sulpiride, eticlopride, and isoremoxipride, were evaluated for their selective in vivo uptake into dopamine D2 receptor rich tissue of the rat brain. Iodopride (KD 0.88 nM), an iodine substituted benzamide structurally related to sulpiride, displayed a maximal striatum: cerebellar uptake ratio of 7.6. Demonstration of saturation of the receptor with [125I]iodopride in striatum required uptake in frontal cortex to be used, rather than cerebellar uptake, to define nonspecific binding. Two other ligands structurally related to eticlopride, iclopride (KD 0.23 nM) and itopride (KD 0.16 nM), displayed maximal striatal: cerebellar uptake ratios of 9.8 and 3.3, respectively. The most potent ligands, epidepride (KD 0.057 nM) and ioxipride (KD 0.070 nM) showed striatal:cerebellar uptake ratios of 234 and 65, respectively. The observed uptake ratios correlated poorly with the affinity constants for the dopamine D2 receptor alone, but were highly correlated (r = 0.92) with the product of the receptor dissociation constant (KD) and the apparent lipophilicity (kw), as determined by reverse-phase HPLC at pH 7.5. Total striatal uptake also appeared dependent on lipophilicity, with maximal uptake occurring for ligands having log kw 2.4-2.8

  19. Genome-wide scan on total serum IgE levels identifies FCER1A as novel susceptibility locus.

    Directory of Open Access Journals (Sweden)

    Stephan Weidinger

    2008-08-01

    Full Text Available High levels of serum IgE are considered markers of parasite and helminth exposure. In addition, they are associated with allergic disorders, play a key role in anti-tumoral defence, and are crucial mediators of autoimmune diseases. Total IgE is a strongly heritable trait. In a genome-wide association study (GWAS, we tested 353,569 SNPs for association with serum IgE levels in 1,530 individuals from the population-based KORA S3/F3 study. Replication was performed in four independent population-based study samples (total n = 9,769 individuals. Functional variants in the gene encoding the alpha chain of the high affinity receptor for IgE (FCER1A on chromosome 1q23 (rs2251746 and rs2427837 were strongly associated with total IgE levels in all cohorts with P values of 1.85 x 10(-20 and 7.08 x 10(-19 in a combined analysis, and in a post-hoc analysis showed additional associations with allergic sensitization (P = 7.78 x 10(-4 and P = 1.95 x 10(-3. The "top" SNP significantly influenced the cell surface expression of FCER1A on basophils, and genome-wide expression profiles indicated an interesting novel regulatory mechanism of FCER1A expression via GATA-2. Polymorphisms within the RAD50 gene on chromosome 5q31 were consistently associated with IgE levels (P values 6.28 x 10(-7-4.46 x 10(-8 and increased the risk for atopic eczema and asthma. Furthermore, STAT6 was confirmed as susceptibility locus modulating IgE levels. In this first GWAS on total IgE FCER1A was identified and replicated as new susceptibility locus at which common genetic variation influences serum IgE levels. In addition, variants within the RAD50 gene might represent additional factors within cytokine gene cluster on chromosome 5q31, emphasizing the need for further investigations in this intriguing region. Our data furthermore confirm association of STAT6 variation with serum IgE levels.

  20. Selection of DNA aptamers against epidermal growth factor receptor with high affinity and specificity

    Energy Technology Data Exchange (ETDEWEB)

    Wang, Deng-Liang [The First Clinical Medical College of Fujian Medical University, Fuzhou (China); Department of Neurosurgery, The First Affiliated Hospital of Fujian Medical University, Fuzhou (China); Song, Yan-Ling; Zhu, Zhi; Li, Xi-Lan; Zou, Yuan [State Key Laboratory for Physical Chemistry of Solid Surfaces, Key Laboratory for Chemical Biology of Fujian Province, Key Laboratory of Analytical Chemistry, and Department of Chemical Biology, College of Chemistry and Chemical Engineering, Xiamen University, Xiamen 361005 (China); Yang, Hai-Tao; Wang, Jiang-Jie [The First Clinical Medical College of Fujian Medical University, Fuzhou (China); Yao, Pei-Sen [Department of Neurosurgery, The First Affiliated Hospital of Fujian Medical University, Fuzhou (China); Pan, Ru-Jun [The First Clinical Medical College of Fujian Medical University, Fuzhou (China); Yang, Chaoyong James, E-mail: cyyang@xmu.edu.cn [State Key Laboratory for Physical Chemistry of Solid Surfaces, Key Laboratory for Chemical Biology of Fujian Province, Key Laboratory of Analytical Chemistry, and Department of Chemical Biology, College of Chemistry and Chemical Engineering, Xiamen University, Xiamen 361005 (China); Kang, De-Zhi, E-mail: kdzy99988@163.com [The First Clinical Medical College of Fujian Medical University, Fuzhou (China); Department of Neurosurgery, The First Affiliated Hospital of Fujian Medical University, Fuzhou (China)

    2014-10-31

    Highlights: • This is the first report of DNA aptamer against EGFR in vitro. • Aptamer can bind targets with high affinity and selectivity. • DNA aptamers are more stable, cheap and efficient than RNA aptamers. • Our selected DNA aptamer against EGFR has high affinity with K{sub d} 56 ± 7.3 nM. • Our selected DNA aptamer against EGFR has high selectivity. - Abstract: Epidermal growth factor receptor (EGFR/HER1/c-ErbB1), is overexpressed in many solid cancers, such as epidermoid carcinomas, malignant gliomas, etc. EGFR plays roles in proliferation, invasion, angiogenesis and metastasis of malignant cancer cells and is the ideal antigen for clinical applications in cancer detection, imaging and therapy. Aptamers, the output of the systematic evolution of ligands by exponential enrichment (SELEX), are DNA/RNA oligonucleotides which can bind protein and other substances with specificity. RNA aptamers are undesirable due to their instability and high cost of production. Conversely, DNA aptamers have aroused researcher’s attention because they are easily synthesized, stable, selective, have high binding affinity and are cost-effective to produce. In this study, we have successfully identified DNA aptamers with high binding affinity and selectivity to EGFR. The aptamer named TuTu22 with K{sub d} 56 ± 7.3 nM was chosen from the identified DNA aptamers for further study. Flow cytometry analysis results indicated that the TuTu22 aptamer was able to specifically recognize a variety of cancer cells expressing EGFR but did not bind to the EGFR-negative cells. With all of the aforementioned advantages, the DNA aptamers reported here against cancer biomarker EGFR will facilitate the development of novel targeted cancer detection, imaging and therapy.

  1. Selection of DNA aptamers against epidermal growth factor receptor with high affinity and specificity

    International Nuclear Information System (INIS)

    Highlights: • This is the first report of DNA aptamer against EGFR in vitro. • Aptamer can bind targets with high affinity and selectivity. • DNA aptamers are more stable, cheap and efficient than RNA aptamers. • Our selected DNA aptamer against EGFR has high affinity with Kd 56 ± 7.3 nM. • Our selected DNA aptamer against EGFR has high selectivity. - Abstract: Epidermal growth factor receptor (EGFR/HER1/c-ErbB1), is overexpressed in many solid cancers, such as epidermoid carcinomas, malignant gliomas, etc. EGFR plays roles in proliferation, invasion, angiogenesis and metastasis of malignant cancer cells and is the ideal antigen for clinical applications in cancer detection, imaging and therapy. Aptamers, the output of the systematic evolution of ligands by exponential enrichment (SELEX), are DNA/RNA oligonucleotides which can bind protein and other substances with specificity. RNA aptamers are undesirable due to their instability and high cost of production. Conversely, DNA aptamers have aroused researcher’s attention because they are easily synthesized, stable, selective, have high binding affinity and are cost-effective to produce. In this study, we have successfully identified DNA aptamers with high binding affinity and selectivity to EGFR. The aptamer named TuTu22 with Kd 56 ± 7.3 nM was chosen from the identified DNA aptamers for further study. Flow cytometry analysis results indicated that the TuTu22 aptamer was able to specifically recognize a variety of cancer cells expressing EGFR but did not bind to the EGFR-negative cells. With all of the aforementioned advantages, the DNA aptamers reported here against cancer biomarker EGFR will facilitate the development of novel targeted cancer detection, imaging and therapy

  2. Characterization of the somatogenic receptor in rat liver. Hydrodynamic properties and affinity cross-linking

    International Nuclear Information System (INIS)

    Rat liver somatogenic receptors have been characterized by gel permeation chromatography, sucrose density gradients in H2O and D2O, and affinity cross-linking using 125I-bovine growth hormone (bGH) as a specific somatogenic receptor ligand. Cross-linking of 125I-bovine growth hormone to a Triton X-100-treated low density fraction isolated from livers of late pregnant rats followed by sodium dodecylsulfate-polyacrylamide gel electrophoresis under reducing conditions showed three major binders with Mr 95,000, 86,000, and 43,000 and a minor binder of Mr 55,000, after correction for bound ligand assuming a 1:1 binding ratio of ligand-receptor. The Mr 86,000, 55,000, and 43,000 species were recovered in the detergent-soluble supernatant after high-speed centrifugation, whereas the Mr 95,000 species remained Triton X-100 insoluble. Detergent-soluble 125I-bGH-receptor complexes were further analyzed by sedimentation into sucrose density gradients. The sedimentation coefficient was S20,w = 5.2 S and the partial specific volume v = 0.72 ml/g. Gel permeation chromatography on a Sepharose S-400 column indicated a Stokes radius of 61 A for the 125I-bGH-receptor-Triton X-100 complex. Based on these figures, the molecular weight of the complex was calculated as 131,100. The molecular weight of the ligand-free receptor-Triton X-100 complex was calculated as Mr 109,100. Affinity cross-linking and sodium dodecyl sulfate-polyacrylamide gel electrophoresis of the 61 A peak from Sephacryl S-400 chromatography (cf. above) showed two binding entities, one major and one minor with Mr values 86,000 and 43,000, respectively, in the absence of reductant. When electrophoresis was run in the presence of reductant the Mr 43,000 species was the major binding entity

  3. Conformational destabilization of Immunoglobulin G increases the low pH-binding affinity with the Neonatal Fc Receptor

    DEFF Research Database (Denmark)

    Walters, Benjamin T; Jensen, Pernille Foged; Larraillet, Vincent;

    2016-01-01

    Crystallographic evidence suggests that the pH-dependent affinity of IgG molecules for the neonatal Fc receptor (FcRn) receptor primarily arises from salt bridges involving IgG histidine residues, resulting in moderate affinity at mildly acidic conditions. However, this view does not explain the...... diversity in affinity found in IgG variants, such as the YTE mutant (M252Y,S254T,T256E), which increases affinity to FcRn by up to 10×. Here we compare hydrogen exchange measurements at pH 7.0 and pH 5.5 with and without FcRn bound with surface plasmon resonance estimates of dissociation constants and Fc...

  4. Dual Labeled Peptides as Tools to Study Receptors: Nanomolar Affinity Derivatives of TIPP (Tyr-Tic-Phe-Phe) Containing an Affinity Label and Biotin as Probes of δ Opioid Receptors

    OpenAIRE

    Aldrich, Jane V.; Kumar, Vivek; Murray, Thomas F.; Guang, Wei; Wang, Jia Bei

    2009-01-01

    A general strategy for the design of dual labeled peptides was developed and derivatives of the δ opioid receptor (DOR) selective antagonist TIPP (Tyr-Tic-Phe-PheOH) containing both an affinity label and biotin were prepared by solid phase synthesis. Tyr-Tic-Phe-Phe(p-X)-Asp-NH(CH2CH2O)2-CH2CH2NH-biotin (where X = N=C=S or NHCOCH2Br) exhibit nanomolar DOR affinity. The ability to detect receptors labeled with these peptides following solubilization and SDS-PAGE demonstrate the applicability o...

  5. IGES transformer and NURBS in grid generation

    Science.gov (United States)

    Yu, Tzu-Yi; Soni, Bharat K.

    1993-01-01

    In the field of Grid Generation and the CAD/CAM, there are numerous geometry output formats which require the designer to spend a great deal of time manipulating geometrical entities in order to achieve a useful sculptured geometrical description for grid generation. Also in this process, there is a danger of losing fidelity of the geometry under consideration. This stresses the importance of a standard geometry definition for the communication link between varying CAD/CAM and grid system. The IGES (Initial Graphics Exchange Specification) file is a widely used communication between CAD/CAM and the analysis tools. The scientists at NASA Research Centers - including NASA Ames, NASA Langley, NASA Lewis, NASA Marshall - have recognized this importance and, therefore, in 1992 they formed the committee of the 'NASA-IGES' which is the subset of the standard IGES. This committee stresses the importance and encourages the CFD community to use the standard IGES file for the interface between the CAD/CAM and CFD analysis. Also, two of the IGES entities -- the NURBS Curve (Entity 126) and NURBS Surface (Entity 128) -- which have many useful geometric properties -- like the convex hull property, local control property and affine invariance, also widely utilized analytical geometries can be accurately represented using NURBS. This is important in today grid generation tools because of the emphasis of the interactive design. To satisfy the geometry transformation between the CAD/CAM system and Grid Generation field, the CAGI (Computer Aided Geometry Design) developed, which include the Geometry Transformation, Geometry Manipulation and Geometry Generation as well as the user interface. This paper will present the successful development IGES file transformer and application of NURBS definition in the grid generation.

  6. Synthesis and evaluation of [18F] labeled benzamides: High affinity sigma receptor ligands for PET imaging

    International Nuclear Information System (INIS)

    We have synthesized and characterized four new fluorinated halobenzamides as sigma receptor ligands for use with positron emission tomography (PET). All the compounds were found to have high sigma-1 affinities (Ki = 0.38-0.98 nM), and the 4-fluoro-substituted benzamides were found to be more potent sigma-2 ligands (Ki = 3.77-4.02 nM) than their corresponding 2-fluoro analogs (Ki = 20.3-22.8 nM). The [18F] radiochemical syntheses of two of the analogs gave overall yields between 3-10% (EOS), radiochemical purities >99%, and specific activities between 800-1200 Ci/mmol (29.6-44.4 TBq/mmol). Rat biodistribution and blocking experiments were performed with 2-[18F](N-fluorobenzylpiperidin-4yl)-4-iodobenzamide, the analog with the best Ki value for sigma-1 sites (0.38 nM). Results of these experiments demonstrate specific uptake of the compound in tissues believed to contain sigma receptors, such as lungs, kidneys, heart, brain, and spleen and indicate its potential as a candidate for use in PET imaging of tissues containing these receptors

  7. Energetics of ligand-receptor binding affinity on endothelial cells: An in vitro model.

    Science.gov (United States)

    Fotticchia, Iolanda; Guarnieri, Daniela; Fotticchia, Teresa; Falanga, Andrea Patrizia; Vecchione, Raffaele; Giancola, Concetta; Netti, Paolo Antonio

    2016-08-01

    Targeted therapies represent a challenge in modern medicine. In this contest, we propose a rapid and reliable methodology based on Isothermal Titration Calorimetry (ITC) coupled with confluent cell layers cultured around biocompatible templating microparticles to quantify the number of overexpressing receptors on cell membrane and study the energetics of receptor-ligand binding in near-physiological conditions. In the in vitro model here proposed we used the bEnd3 cell line as brain endothelial cells to mimic the blood brain barrier (BBB) cultured on dextran microbeads ranging from 67μm to 80μm in size (Cytodex) and the primary human umbilical vein cells (HUVEC) for comparison. The revealed affinity between transferrin (Tf) and transferrin receptor (TfR) in both systems is very high, Kd values are in the order of nM. Conversely, the value of TfRs/cell reveals a 100-fold increase in the number of TfRs per bEnd3 cells compared to HUVEC cells. The presented methodology can represent a novel and helpful strategy to identify targets, to address drug design and selectively deliver therapeutics that can cross biological barriers such as the blood brain barrier. PMID:27100851

  8. Introduction of D-Phenylalanine enhanced the receptor binding affinities of gonadotropin-releasing hormone peptides

    OpenAIRE

    Lu, Jie; Hathaway, Helen J.; Royce, Melanie E.; Prossnitz, Eric R.; Miao, Yubin

    2014-01-01

    The purpose of this study was to examine whether the introduction of D-Phe could improve the GnRH receptor binding affinities of DOTA-conjugated D-Lys6-GnRH peptides. Building upon the construct of DOTA-Ahx-(D-Lys6-GnRH1) we previously reported, an aromatic amino acid of D-Phe was inserted either between the DOTA and Ahx or between the Ahx and D-Lys6 to generate new DOTA-D-Phe-Ahx-(D-Lys6-GnRH) or DOTA-Ahx-D-Phe-(D-Lys6-GnRH) peptides. Compared to DOTA-Ahx-(D-Lys6-GnRH1) (36.1 nM), the introd...

  9. High-affinity receptors for bombesin-like peptides in normal guinea pig lung membranes

    Energy Technology Data Exchange (ETDEWEB)

    Lach, E.; Trifilieff, A.; Landry, Y.; Gies, J.P. (Univ. Louis Pasteur, Illkirch (France))

    1991-01-01

    The binding of the radiolabeled bombesin analogue ({sup 125}I-Tyr{sup 4})bombesin to guinea-pig lung membranes was investigated. Binding of ({sup 125}I-Tyr{sup 4})bombesin was specific, saturable, reversible and linearly related to the protein concentration. Scatchard analysis of equilibrium binding data at 25C indicated the presence of a single class of non-interacting binding sites for bombesin (B{sub max} = 7.7 fmol/mg protein). The value of the equilibrium dissociation constant (K{sub D} = 90 pM) agrees with a high-affinity binding site. Bombesin and structurally related peptides such as ({sup 125}I-Tyr{sup 4})bombesin, neuromedin B and neuromedin C inhibited the binding of ({sup 125}I-Tyr{sup 4})bombesin in an order of potencies as follows: ({sup 125}I-Tyr{sup 4})bombesin {gt} bombesin {ge} neuromedin C {much gt} neuromedin B. These results indicate that guinea-pig lung membranes possess a single class of bombesin receptors with a high affinity for bombesin and a lower one for neuromedin B.

  10. Development of 99mTc labelled somatostatin analogues with high affinity for somatostatin receptors

    International Nuclear Information System (INIS)

    A recent development in oncology involves the use of metabolically stabilized peptide hormone analogues labelled with metallic radionuclides for the diagnosis or therapy of malignant disease. This approach was successfully applied for the first time in the visualization of somatostatin positive tumours and their metastases with 111In DTPA-octreotide. In an effort to obtain a 99mTc somatostatin receptor affine radioligand we describe herein the synthesis, radiochemistry and preliminary biological evaluation of two novel 99mTc labelled somatostatin analogues, N4-TOC and N4-RC-160. In these compounds a tetraamine bifunctional unit was covalently attached to the N-terminal (D)Phe1 of the peptide chain using Boc-protection strategies. The peptide conjugates were purified by high performance liquid chromatography (HPLC) and characterized by UV/Vis and ES-MS spectroscopies. As revealed by HPLC, 99mTc labelling was quantitative under mild conditions, leading to a single 99mTc species in high specific activities. Affinity of 99mTc N4-TOC for the somatostatin receptor, as determined by in vitro binding assays in rat brain cortex membranes, was found unaffected by the presence of the bulky metal chelate. The binding properties of 99mTc N4-RC-160 could not be determined by this assay due to an extremely high non-specific binding of this radioligand, and will be shortly investigated by other methods. Tissue distribution in healthy mice revealed that 99mTc N4-TOC is clearing mainly through the kidneys and the urinary tract whereas 99mTc N4-RC-160 shows a high accumulation in the liver as a result of its lipophilicity. Analysis of urine samples by HPLC showed that 99mTc N4-TOC is excreted integer from the body of mice, while 99mTc N4-RC-160 is totally transformed to an unidentified hydrophilic metabolite in vivo. The location of this metabolism is currently investigated. In vivo blocking experiments using animals pre-treated with 50 μg octreotide prior to the injection of the

  11. Close-up of the alpha-1,3-Gal epitope as defined by a monoclonal chimeric IgE and human serum using saturation transfer difference (STD) NMR

    DEFF Research Database (Denmark)

    Plum, Melanie; Michel, Yvonne; Wallach, Katharina;

    2011-01-01

    epitopes in allergy. The aim of this study was to dissect immunoreactivity, interaction and fine epitope of alpha-Gal-specific antibodies to obtain insights into the recognition of carbohydrate epitopes by IgE antibodies and their consequences on a molecular and cellular level. The antigen binding moiety...... by mediator release assays, surface plasmon resonance (SPR) and STD NMR analyses. The alpha-Gal-specific chimeric IgE and IgG antibodies were proven functional regarding interaction with antigen and Fc receptors. SPR measurements demonstrated affinities in the micromolar range. In contrast to a reference...

  12. Vaccination for birch pollen allergy. Induction of affinity-matured or blocking IgG antibodies does not account for the reduced binding of IgE to Bet v 1

    DEFF Research Database (Denmark)

    Svenson, Morten; Jacobi, Henrik H; Bødtger, Uffe;

    2003-01-01

    Specific allergy vaccination (SAV) is associated with increased levels of allergen specific IgG in serum. It is not clear, however, to what extent qualitative changes in allergen binding to IgG may be induced as well. We therefore analyzed the binding of the major allergen in pollen of birch...... the amount of Bet v 1 compared with IgE, and together they accounted for more than 85% of the serum binding capacity. While SAV induced minimal changes in IgE binding, the IgG binding capacities increased 6-32 times. In contrast, the binding avidities (K(d) 28-40pM) changed less than 20%, pre- and post...

  13. Solubilization of high affinity corticotropin-releasing factor receptors from rat brain: Characterization of an active digitonin-solubilized receptor complex

    International Nuclear Information System (INIS)

    The binding characteristics of CRF receptors in rat frontal cerebral cortex membranes solubilized in 1% digitonin were determined. The binding of [125I]Tyro-ovine CRF ([125I]oCRF) to solubilized membrane proteins was dependent on incubation time, temperature, and protein concentration, was saturable and of high affinity, and was absent in boiled tissue. The solubilized receptors retained their high affinity for [125I] oCRF in the solubilized state, exhibiting a dissociation constant (KD) of approximately 200 pM, as determined by direct binding saturation isotherms. Solubilized CRF receptors maintained the rank order of potencies for various related and unrelated CRF peptides characteristic of the membrane CRF receptor: rat/human CRF congruent to ovine CRF congruent to Nle21,38-rat CRF greater than alpha-helical oCRF-(9-41) greater than oCRF-(7-41) much greater than vasoactive intestinal peptide, arginine vasopressin, or the substance-P antagonist. Furthermore, the absolute potencies (Ki values) for the various CRF-related peptides in solubilized receptors were almost identical to those observed in the membrane preparations, indicating that the CRF receptor retained its high affinity binding capacity in the digitonin-solubilized state. Chemical affinity cross-linking of digitonin-solubilized rat cortical membrane proteins revealed a specifically labeled protein with an apparent mol wt of 58,000 which was similar to the labeled protein in native membrane homogenates. Although solubilized CRF receptors retained their high affinity for agonists, their sensitivity for guanine nucleotide was lost. Size exclusion chromatography substantiated these results, demonstrating that in the presence or absence of guanine nucleotides, [125I]oCRF labeled the same size receptor complex

  14. Affinity profiles for human somatostatin receptor subtypes SST1-SST5 of somatostatin radiotracers selected for scintigraphic and radiotherapeutic use

    International Nuclear Information System (INIS)

    Using cell lines transfected with somatostatin receptor subtypes sst1, sst2, sst3, sst4 and sst5, we have evaluated the in vitro binding characteristics of labelled (indium, yttrium, gallium) and unlabelled DOTA-[Tyr3]-octreotide, DOTA-octreotide, DOTA-lanreotide, DOTA-vapreotide, DTPA-[Tyr3]-octreotate and DOTA-[Tyr3]-octreotate. Small structural modifications, chelator substitution or metal replacement were shown to considerably affect the binding affinity. A marked improvement of sst2 affinity was found for Ga-DOTA-[Tyr3]-octreotide (IC50 2.5 nM) compared with the Y-labelled compound and Octreoscan. An excellent binding affinity for sst2 in the same range was also found for In-DTPA-[Tyr3]-octreotate (IC50 1.3 nM) and for Y-DOTA-[Tyr3]-octreotate (IC50 1.6 nM). Remarkably, Ga-DOTA-[Tyr3]-octreotate bound at sst2 with a considerably higher affinity (IC50 0.2 nM). An up to 30-fold improvement in sst3 affinity was observed for unlabelled or Y-labelled DOTA-octreotide compared with their Tyr3-containing analogue, suggesting that replacement of Tyr3 by Phe is crucial for high sst3 affinity. Substitution in the octreotide molecule of the DTPA by DOTA improved the sst3 binding affinity 14-fold. Whereas Y-DOTA-lanreotide had only low affinity for sst3 and sst4, it had the highest affinity for sst5 among the tested compounds (IC50 16 nM). Increased binding affinity for sst3 and sst5 was observed for DOTA-[Tyr3]-octreotide, DOTA-lanreotide and DOTA-vapreotide when they were labelled with yttrium. These marked changes in subtype affinity profiles are due not only to the different chemical structures but also to the different charges and hydrophilicity of these compounds. Interestingly, even the coordination geometry of the radiometal complex remote from the pharmacophoric amino acids has a significant influence on affinity profiles as shown with Y-DOTA versus Ga-DOTA in either [Tyr3]-octreotide or [Tyr3]-octreotate. Such changes in sst affinity profiles must be identified

  15. NK1 receptor fused to beta-arrestin displays a single-component, high-affinity molecular phenotype

    DEFF Research Database (Denmark)

    Martini, Lene; Hastrup, Hanne; Holst, Birgitte;

    2002-01-01

    against substance P and especially against antagonists with up to 1000-fold lower apparent affinity than determined in functional assays and in homologous binding assays. When the NK1 receptor was closely fused to G proteins, this phenomenon was eliminated among agonists, but the agonists still competed...

  16. T-cell receptor affinity and avidity defines antitumor response and autoimmunity in T-cell immunotherapy

    Science.gov (United States)

    Zhong, Shi; Malecek, Karolina; Johnson, Laura A.; Yu, Zhiya; Vega-Saenz de Miera, Eleazar; Darvishian, Farbod; McGary, Katelyn; Huang, Kevin; Boyer, Josh; Corse, Emily; Shao, Yongzhao; Rosenberg, Steven A.; Restifo, Nicholas P.; Osman, Iman; Krogsgaard, Michelle

    2013-01-01

    T cells expressing antigen-specific T-cell receptors (TCRs) can mediate effective tumor regression, but they often also are accompanied by autoimmune responses. To determine the TCR affinity threshold defining the optimal balance between effective antitumor activity and autoimmunity in vivo, we used a unique self-antigen system comprising seven human melanoma gp100(209–217)-specific TCRs spanning physiological affinities (1–100 μM). We found that in vitro and in vivo T-cell responses are determined by TCR affinity, except in one case that was compensated by substantial CD8 involvement. Strikingly, we found that T-cell antitumor activity and autoimmunity are closely coupled but plateau at a defined TCR affinity of 10 µM, likely due to diminished contribution of TCR affinity to avidity above the threshold. Together, these results suggest that a relatively low-affinity threshold is necessary for the immune system to avoid self-damage, given the close relationship between antitumor activity and autoimmunity. The low threshold, in turn, indicates that adoptive T-cell therapy treatment strategies using in vitro-generated high-affinity TCRs do not necessarily improve efficacy. PMID:23576742

  17. Alveolar type II cells express a high-affinity receptor for pulmonary surfactant protein A

    International Nuclear Information System (INIS)

    Primary cultures of rat alveolar type II cells bind radiolabeled pulmonary surfactant protein A (SP-A) with high affinity. The binding of 125I-labeled SP-A is time- and temperature-dependent and is not accompanied by significant degradation. The binding process is saturable at low concentrations of SP-A, and unlabeled SP-A readily competes with labeled SP-A for cellular binding sites. Subsequent to binding, two pools of cell-associated 125I-labeled SP-A can be identified based upon sensitivity to trypsin at 0 degree C. It is likely that the trypsin-sensitive pool comprises 125I-labeled SP-A bound to the cell surface and the trypsin-insensitive pool comprises the internalized protein. Scatchard analysis of cell surface binding of SP-A at 0.1-10 μg/ml shows positive cooperativity at concentrations between 0.1 and 1 μg/ml. Hill plots give nH = 1.34 ± 0.08 with an apparent dissociation constant K'd = 1.02 ± 0.32 μg/ml. The binding of SP-A to type II cells shows an absolute requirement for Ca2+. The putative receptor for SP-A is unaffected by treatment of type II cells with a variety of proteases and N-Glycanase. Alveolar macrophages also exhibit high-affinity binding of SP-A, but rat lung fibroblasts and the alveolar epithelial cell line L2 exhibit only nonspecific binding

  18. Expression cloning of cDNA encoding a seven-helix receptor from human placenta with affinity for opioid ligands

    OpenAIRE

    1992-01-01

    Here we report the expression cloning of cDNA encoding a putative opioid receptor from a human placenta cDNA library. Placental opioid receptors are of the kappa type. As the dynorphin opioid peptides are kappa-selective, a dynorphin ligand was used in an affinity-enrichment (panning) procedure to select transiently transfected COS-7 cells expressing kappa receptor binding sites. The cloned cDNA encodes a 440-residue protein of the seven-helix guanine nucleotide-binding protein (G-protein)-co...

  19. Gestational treatment with cocaine and fluoxetine alters oxytocin receptor number and binding affinity in lactating rat dams.

    Science.gov (United States)

    Johns, Josephine M; Lubin, Deborah A; Walker, Cheryl H; Joyner, Paul; Middleton, Christopher; Hofler, Vivian; McMurray, Matthew

    2004-01-01

    Cocaine administered chronically throughout gestation has been correlated with deficits in maternal behavior, increased maternal aggressive behavior and decreased oxytocin levels in rats. In addition to its effects on oxytocin levels, cocaine is a potent serotonergic, dopaminergic and noradrenergic reuptake inhibitor. Alterations in the dopaminergic and serotonergic systems have been suggested as possibly having a role in cocaine-induced maternal aggression. This study was in part, an attempt to understand some of the mechanisms by which cocaine increases postpartum aggression, particularly as they relate to changes in the oxytocin system. Oxytocin receptor number and binding affinity in the medial preoptic area of the hypothalamus, ventral tegmental area, hippocampus and amygdala were determined for lactating rat dams on postpartum day 6 (PPD 6) that were gestationally treated with cocaine, fluoxetine, saline or an amfonelic acid/fluoxetine drug combination. Cocaine and fluoxetine treatment both resulted in a significant up-regulation of oxytocin receptor number and lower receptor affinity in the amygdala of lactating rat dams compared to saline controls and the amfonelic acid/fluoxetine combination treatment group. Cocaine treatment also resulted in a significant down-regulation of oxytocin receptors in the medial preoptic area and both cocaine and fluoxetine treated dams had the highest affinity for oxytocin receptors in this brain region. Results of the present study support previous data indicating that alterations in oxytocinergic and perhaps serotonergic system dynamics in the amygdala may play a role in cocaine-induced postpartum aggression. PMID:15380831

  20. Affinity purification of human granulocyte macrophage colony-stimulating factor receptor alpha-chain. Demonstration of binding by photoaffinity labeling

    International Nuclear Information System (INIS)

    The human granulocyte macrophage colony-stimulating factor (GM-CSF) receptor alpha-chain, a low affinity component of the receptor, was solubilized and affinity-purified from human placenta using biotinylated GM-CSF. Scatchard analysis of 125I-GM-CSF binding to the placental membrane extract disclosed that the GM-CSF receptor had a dissociation constant (Kd) of 0.5-0.8 nM, corresponding to the Kd value of the GM-CSF receptor alpha-chain on the intact placental membrane. Affinity labeling of the solubilized protein using a photoreactive cross-linking agent, N-hydroxysuccinimidyl-4-azidobenzoate (HSAB), demonstrated a single specific band of 70-95 kDa representing a ligand-receptor complex. Approximately 2 g of the placental membrane extract was subjected to a biotinylated GM-CSF-fixed streptavidin-agarose column, resulting in a single major band at 70 kDa on a silver-stained sodium dodecyl sulfate gel. The radioiodination for the purified material disclosed that the purified protein had an approximate molecular mass of 70 kDa and a pI of 6.6. Binding activity of the purified material was demonstrated by photoaffinity labeling using HSAB-125I-GM-CSF, producing a similar specific band at 70-95 kDa as was demonstrated for the crude protein

  1. Affinity purification of human granulocyte macrophage colony-stimulating factor receptor alpha-chain. Demonstration of binding by photoaffinity labeling

    Energy Technology Data Exchange (ETDEWEB)

    Chiba, S.; Shibuya, K.; Miyazono, K.; Tojo, A.; Oka, Y.; Miyagawa, K.; Takaku, F. (Univ. of Tokyo (Japan))

    1990-11-15

    The human granulocyte macrophage colony-stimulating factor (GM-CSF) receptor alpha-chain, a low affinity component of the receptor, was solubilized and affinity-purified from human placenta using biotinylated GM-CSF. Scatchard analysis of {sup 125}I-GM-CSF binding to the placental membrane extract disclosed that the GM-CSF receptor had a dissociation constant (Kd) of 0.5-0.8 nM, corresponding to the Kd value of the GM-CSF receptor alpha-chain on the intact placental membrane. Affinity labeling of the solubilized protein using a photoreactive cross-linking agent, N-hydroxysuccinimidyl-4-azidobenzoate (HSAB), demonstrated a single specific band of 70-95 kDa representing a ligand-receptor complex. Approximately 2 g of the placental membrane extract was subjected to a biotinylated GM-CSF-fixed streptavidin-agarose column, resulting in a single major band at 70 kDa on a silver-stained sodium dodecyl sulfate gel. The radioiodination for the purified material disclosed that the purified protein had an approximate molecular mass of 70 kDa and a pI of 6.6. Binding activity of the purified material was demonstrated by photoaffinity labeling using HSAB-{sup 125}I-GM-CSF, producing a similar specific band at 70-95 kDa as was demonstrated for the crude protein.

  2. Evidence for monomeric and oligomeric hormone-binding domains in affinity-purified gonadotropin receptor from rat ovary

    International Nuclear Information System (INIS)

    Rat ovarian lutropin/choriogonadotropin receptor was purified from a Triton X-100-solubilized membrane preparation by affinity chromatography with Affi-Gel 10 coupled to purified human choriogonadotropin. The affinity-purified receptor preparations contained a single class of high-affinity binding sites for 125I-labeled human choriogonadotropin, with an equilibrium dissociation constant (Kd) of 2.5 x 10-9 M, which is comparable to the Kd values for membrane-bound and solubilized receptors. The purified receptor appeared as two dominant bands with molecular weights of 135,000 and 92,000 after sodium dodecyl sulfate/polyacrylamide gel electrophoresis (SDS/PAGE) under nonreducing conditions. When the individual affinity-purified receptor bands were electroeluted from the gel and analyzed again by SDS/PAGE under nonreducing conditions, both the Mr 92,000 and the 135,000 proteins retained their original molecular form even when 8 M urea was included in the gel. However, when the electrophoretically purified Mr 92,000 and 135,000 bands were subjected to SDS/PAGE under reducing conditions, the Mr 135,000 species was almost completely converted to a Mr 92,000 band, but the Mr 92,000 species did not undergo any alteration in molecular weight. The results suggest that the lutropin/choriogonadotropin receptor from rat ovary exists in two molecular forms, and the higher molecular weight form appears to be composed of disulfide-linked Mr 92,000 subunit, which comprises the hormone-binding domain

  3. Domain interplay in the urokinase receptor. Requirement for the third domain in high affinity ligand binding and demonstration of ligand contact sites in distinct receptor domains

    DEFF Research Database (Denmark)

    Behrendt, N; Ronne, E; Dano, K

    1996-01-01

    . This result shows that in addition to D1, which has an established function in ligand binding (Behrendt, N., Ploug, M., Patthy, L., Houen, G., Blasi, F., and Dano, K. (1991) J. Biol. Chem. 266, 7842-7847), D3 has an important role in governing a high affinity in the intact receptor. Real-time biomolecular...

  4. Galpha-subunits differentially alter the conformation and agonist affinity of kappa-opioid receptors.

    Science.gov (United States)

    Yan, Feng; Mosier, Philip D; Westkaemper, Richard B; Roth, Bryan L

    2008-02-12

    Although ligand-induced conformational changes in G protein-coupled receptors (GPCRs) are well-documented, there is little direct evidence for G protein-induced changes in GPCR conformation. To investigate this possibility, the effects of overexpressing Galpha-subunits (Galpha16 or Galphai2) with the kappa-opioid receptor (KOR) were examined. The changes in KOR conformation were subequently examined via the substituted cysteine accessibility method (SCAM) in transmembrane domains 6 (TM6) and 7 (TM7) and extracellular loop 2 (EL2). Significant conformational changes were observed on TM7, the extracellular portion of TM6, and EL2. Seven SCAM-sensitive residues (S3107.33, F3147.37, and I3167.39 to Y3207.43) on TM7 presented a cluster pattern when the KOR was exposed to baseline amounts of G protein, and additional residues became sensitive upon overexpression of various G proteins. In TM7, S3117.34 and N3267.49 were found to be sensitive in Galpha16-overexpressed cells and Y3137.36, N3227.45, S3237.46, and L3297.52 in Galphai2-overexpressed cells. In addition, the degree of sensitivity for various TM7 residues was augmented, especially in Galphai2-overexpressed cells. A similar phenomenon was also observed for residues in TM6 and EL2. In addition to an enhanced sensitivity of certain residues, our findings also indicated that a slight rotation was predicted to occur in the upper part of TM7 upon G protein overexpression. These relatively modest conformational changes engendered by G protein overexpression had both profound and differential effects on the abilities of agonists to bind to KOR. These data are significant because they demonstrate that Galpha-subunits differentially modulate the conformation and agonist affinity of a prototypical GPCR. PMID:18205395

  5. Improved binding affinity and interesting selectivities of aminopyrimidine-bearing carbohydrate receptors in comparison with their aminopyridine analogues.

    Science.gov (United States)

    Lippe, Jan; Seichter, Wilhelm; Mazik, Monika

    2015-12-28

    Due to the problems with the exact prediction of the binding properties of an artificial carbohydrate receptor, the identification of characteristic structural features, having the ability to influence the binding properties in a predictable way, is of high importance. The purpose of our investigation was to examine whether the previously observed higher affinity of 2-aminopyrimidine-bearing carbohydrate receptors in comparison with aminopyridine substituted analogues represents a general tendency of aminopyrimidine-bearing compounds. Systematic binding studies on new compounds consisting of 2-aminopyrimidine groups confirmed such a tendency and allowed the identification of interesting structure-activity relationships. Receptors having different symmetries showed systematic preferences for specific glycosides, which are remarkable for such simple receptor systems. Particularly suitable receptor architectures for the recognition of selected glycosides were identified and represent a valuable base for further developments in this field. PMID:26467387

  6. Computational Estimates of Binding Affinities for Estrogen Receptor Isoforms in Rainbow Trout

    CERN Document Server

    Shyu, Conrad

    2009-01-01

    Molecular dynamics simulations are performed to determine the binding affinities between the hormone 17 beta-estradiol (E2) and different estrogen receptor (ER) isoforms in the rainbow trout (Oncorhynchus mykiss). Previous studies have demonstrated that a recent, unique gene duplication of the ER alpha subtype created two isoforms ER alpha 1 and ER alpha 2, and an early secondary split of ER beta produced two distinct isoforms of ER beta 1 and ER beta 2 based on the phylogenetic analysis. The objective of our computational studies is to provide insight into the underlying evolutionary selection pressure on the ER isoforms. Our results show that E2 binds preferentially to ER alpha 1. This finding corresponds to the experimental results as the ERs evolved from gene duplication events are frequently free from selective pressure and should exhibit no deleterious effects. The E2, however, only binds slightly better to ER beta 2. Both isoforms remain competitive. This finding reflects the fact that since ER beta 2 ...

  7. High-affinity receptors for peptides of the bombesin family in Swiss 3T3 cells

    International Nuclear Information System (INIS)

    Gastrin-releasing peptide (GRP) labeled with 125I at tyrosine-15 (125I-GRP) binds to intact quiescent Swiss 3T3 cells in a specific and saturable manner. Scatchard analysis indicates the presence of a single class of high-affinity binding sites of Kd = 0.5 X 10(-9) M and a value for the number of sites per cell of about 100,000. 125I-GRP binding was not inhibited by other mitogens for these cells, and cell lines that are mitogenically unresponsive to GRP do not exhibit specific GRP binding. Structure-activity relationships show a close parallel between the ability of a range of GRP-related peptides to both inhibit GRP binding and to stimulate mitogenesis. Further, GRP binding is selectively blocked in a competitive fashion by a novel bombesin antagonist, [D-Arg1, D-Pro2, D-Trp7,9, Leu11] substance P. In addition, this compound selectively inhibits GRP and bombesin-induced mitogenesis. These results demonstrate that the mitogenic response of Swiss 3T3 cells to peptides of the bombesin family is mediated by a class of receptors distinct from those of other mitogens for these cells

  8. Early signs of pathological cognitive aging in mice lacking high-affinity nicotinic receptors.

    Directory of Open Access Journals (Sweden)

    Eleni eKonsolaki

    2016-04-01

    Full Text Available In order to address pathological cognitive decline effectively, it is critical to adopt early preventive measures in individuals considered at risk. It is therefore essential to develop approaches that identify such individuals before the onset of irreversible dementia. Α deficient cholinergic system has been consistently implicated as one of the main factors associated with a heightened vulnerability to the aging process. In the present study we used mice lacking high affinity nicotinic receptors (β2-/-, which have been proposed as an animal model of accelerated/premature cognitive aging. Our aim was to identify behavioural signs that could serve as indicators or predictors of impending cognitive decline. We used test batteries in order to assess cognitive functions and additional tasks to investigate spontaneous behaviours, such as species-specific activities and exploration/locomotion in a novel environment. Our data confirm and extend the hypothesis that β2-/- animals exhibit age-related cognitive impairments, manifested in both spatial learning and recognition memory tasks. In addition, we reveal deficits in spontaneous behaviour and habituation processes earlier in life. To our knowledge, this is the first study to perform an extensive behavioural examination of an animal model of premature cognitive aging, and our results suggest that β2-nAChR dependent cognitive deterioration progressively evolves from initial subtle behavioural changes to global dementia due to the combined effect of the neuropathology and aging.

  9. High-affinity cannabinoid binding site in brain: A possible marijuana receptor

    Energy Technology Data Exchange (ETDEWEB)

    Nye, J.S.

    1988-01-01

    The mechanism by which delta{sup 9} tetrahydrocannabinol (delta{sup 9}THC), the major psychoactive component of marijuana or hashish, produces its potent psychological and physiological effects is unknown. To find receptor binding sites for THC, we designed a water-soluble analog for use as a radioligand. 5{prime}-Trimethylammonium-delta{sup 8}THC (TMA) is a positively charged analog of delta-{sup 8}THC modified on the 5{prime} carbon, a portion of the molecule not important for its psychoactivity. We have studied the binding of ({sup 3}H)-5{prime}-trimethylammonium-delta-{sup 8}THC (({sup 3}H)TMA) to rat neuronal membranes. ({sup 3}H)TMA binds saturably and reversibly to brain membranes with high affinity to apparently one class of sites. Highest binding site density occurs in brain, but several peripheral organs also display specific binding. Detergent solubilizes the sites without affecting their pharmacologial properties. Molecular sieve chromatography reveals a bimodal peak of ({sup 3}H)TMA binding activity of approximately 60,000 daltons apparent molecular weight.

  10. High-affinity cannabinoid binding site in brain: A possible marijuana receptor

    International Nuclear Information System (INIS)

    The mechanism by which delta9 tetrahydrocannabinol (delta9THC), the major psychoactive component of marijuana or hashish, produces its potent psychological and physiological effects is unknown. To find receptor binding sites for THC, we designed a water-soluble analog for use as a radioligand. 5'-Trimethylammonium-delta8THC (TMA) is a positively charged analog of delta-8THC modified on the 5' carbon, a portion of the molecule not important for its psychoactivity. We have studied the binding of [3H]-5'-trimethylammonium-delta-8THC ([3H]TMA) to rat neuronal membranes. [3H]TMA binds saturably and reversibly to brain membranes with high affinity to apparently one class of sites. Highest binding site density occurs in brain, but several peripheral organs also display specific binding. Detergent solubilizes the sites without affecting their pharmacologial properties. Molecular sieve chromatography reveals a bimodal peak of [3H]TMA binding activity of approximately 60,000 daltons apparent molecular weight

  11. Roles of the NMDA Receptor and EAAC1 Transporter in the Modulation of Extracellular Glutamate by Low and High Affinity AMPA Receptors in the Cerebellum in Vivo: Differential Alteration in Chronic Hyperammonemia.

    Science.gov (United States)

    Cabrera-Pastor, Andrea; Taoro, Lucas; Llansola, Marta; Felipo, Vicente

    2015-12-16

    The roles of high- and low-affinity AMPA receptors in modulating extracellular glutamate in the cerebellum remain unclear. Altered glutamatergic neurotransmission is involved in neurological alterations in hyperammonemia, which differently affects high- and low-affinity AMPA receptors. The aims were to assess by in vivo microdialysis (a) the effects of high- and low-affinity AMPA receptor activation on extracellular glutamate in the cerebellum; (b) whether chronic hyperammonemia alters extracellular glutamate modulation by high- and/or low-affinity AMPA receptors; and (c) the contribution of NMDA receptors and EAAC1 transporter to AMPA-induced changes in extracellular glutamate. In control rats, high affinity receptor activation does not affect extracellular glutamate but increases glutamate if NMDA receptors are blocked. Low affinity AMPA receptor activation increases transiently extracellular glutamate followed by reduction below basal levels and return to basal values. The reduction is associated with transient increased membrane expression of EAAC1 and is prevented by blocking NMDA receptors. Blocking NMDA receptors with MK-801 induces a transient increase in extracellular glutamate which is associated with reduced membrane expression of EAAC1 followed by increased membrane expression of the glutamate transporter GLT-1. Chronic hyperammonemia does not affect responses to activation of low affinity AMPA receptors. Activation of high affinity AMPA receptors increases extracellular glutamate in hyperammonemic rats by an NMDA receptor-dependent mechanism. In conclusion, these results show that there is a tightly controlled interplay between AMPA and NMDA receptors and an EAAC1 transporter in controlling extracellular glutamate. Hyperammonemia alters high- but not low-affinity AMPA receptors. PMID:26428532

  12. Structure-activity relationships for hallucinogenic N,N-dialkyltryptamines: photoelectron spectra and serotonin receptor affinities of methylthio and methylenedioxy derivatives

    Energy Technology Data Exchange (ETDEWEB)

    Kline, T.B.; Benington, F.; Morin, R.D.; Beaton, J.M.; Glennon, R.A.; Domelsmith, L.N.; Houk, K.N.; Rozeboom, M.D.

    1982-11-01

    Serotonin receptor affinity and photelectron spectral data were obtained on a number of substituted N,N-dimethyltryptamines. Evidence is presented that electron-donating substituents in the 5-position lead to enhanced behavioral disruption activity and serotonin receptor affinity as compared to unsubstituted N,N-dimethyltryptamine and analogues substituted in the 4- or 6-position. Some correlation was found between ionization potentials and behavioral activity, which may have implications concerning the mechanism of receptor binding.

  13. Cutting edge: dominant effect of Ile50Val variant of the human IL-4 receptor alpha-chain in IgE synthesis.

    Science.gov (United States)

    Mitsuyasu, H; Yanagihara, Y; Mao, X Q; Gao, P S; Arinobu, Y; Ihara, K; Takabayashi, A; Hara, T; Enomoto, T; Sasaki, S; Kawai, M; Hamasaki, N; Shirakawa, T; Hopkin, J M; Izuhara, K

    1999-02-01

    Two variants of the IL-4R alpha-chain (IL-4Ralpha) gene have been recently identified in association with different atopic disorders. To clarify the etiological relationship between the two variants, we analyzed responsiveness to IL-4 of transfectants with four kinds of IL-4Ralpha carrying either Val or Ile at 50 and either Gln or Arg at 551. The substitution of Ile for Val augmented STAT6 activation, proliferation, and transcription activity of the Iepsilon promoter by IL-4, whereas that of Arg for Gln did not change these IL-4 signals. Arg551 was not associated with atopic asthma in the Japanese population. CD23 expression and IgE synthesis by IL-4 were augmented in Ile50-bearing PBMC, compared with those bearing Val50. Taken together, substitution of Arg551 does not enhance the IL-4 signal for generation of germline epsilon transcript, whereas the substitution of Ile50 contributes to enhancement of IgE synthesis. PMID:9973373

  14. Synthesis, characterization and binding affinities of rhenium(I) thiosemicarbazone complexes for the estrogen receptor (α/β).

    Science.gov (United States)

    Núñez-Montenegro, Ara; Carballo, Rosa; Vázquez-López, Ezequiel M

    2014-11-01

    The binding affinities towards estrogen receptors (ERs) α and β of a set of thiosemicarbazone ligands (HL(n)) and their rhenium(I) carbonyl complexes [ReX(HL(n))(CO)3] (X=Cl, Br) were determined by a competitive standard radiometric assay with [(3)H]-estradiol. The ability of the coordinated thiosemicarbazone ligands to undergo deprotonation and the lability of the ReX bond were used as a synthetic strategy to obtain [Re(hpy)(L(n))(CO)3] (hpy=3- or 4-hydroxypyridine). The inclusion of the additional hpy ligand endows the new thiosemicarbazonate complexes with an improved affinity towards the estrogen receptors and, consequently, the values of the inhibition constant (Ki) could be determined for some of them. In general, the values of Ki for both ER subtypes suggest an appreciable selectivity towards ERα. PMID:25061691

  15. Deficient T Cell Receptor Excision Circles (TRECs) in autosomal recessive hyper IgE syndrome caused by DOCK8 mutation: implications for pathogenesis and potential detection by newborn screening.

    Science.gov (United States)

    Dasouki, Majed; Okonkwo, Kingsley C; Ray, Abhishek; Folmsbeel, Caspian K; Gozales, Diana; Keles, Sevgi; Puck, Jennifer M; Chatila, Talal

    2011-11-01

    Loss of function of DOCK8 is the major cause of autosomal recessive hyper IgE syndrome, a primary immunodeficiency with adaptive and innate immune dysfunction. Patients affected with ARHIES have atopic dermatitis and recurrent, potentially life-threatening viral and bacterial infections. Three consanguineous Pakistani siblings presented with severe atopic dermatitis and superinfection. Direct sequencing of DOCK8 in all three affected siblings demonstrated homozygosity for a deleterious, novel exon 14 frame shift mutation. Current newborn screening for severe combined immunodeficiency syndrome (SCID) and related T cell disorders relies on the quantitation of T Cell Receptor Excision Cells (TRECs) in dried blood spots (DBS). Significantly, both older affected siblings had undetectable TRECs, and TREC copy number was reduced in the youngest sibling. These findings suggest that AR-HIES may be detected by TREC newborn screening, and this diagnosis should be considered in the evaluation of newborns with abnormal TRECs who do not have typical SCID. PMID:21763205

  16. ZK91587: a novel synthetic antimineralocorticoid displays high affinity for corticosterone (type I) receptors in the rat hippocampus

    International Nuclear Information System (INIS)

    In vitro cytosol binding assays have shown the properties of binding of a novel steroid, ZK91587 (15β, 16βb-methylene-mexrenone) in the brain of rats. Scatchard and Woolf analyses of the binding data reveal the binding of [3H] ZK91587 to the total hippocampal coritcosteroid receptor sites with high affinity, and low capacity. When 100-fold excess RU28362 was included simultaneously with [3H] ZK91587, the labelled steroid binds with the same affinity and capacity. Relative binding affinities (RBA) of various steroids for the Type I or Type II corticosteroid receptor in these animals are: Type I: ZK91587 = corticosterone (B) > cortisol (F); Type II: B > F >>> ZK91587. In the binding kinetic study, ZK91587 has a high association rate of binding in the rat. The steroid dissociates following a one slope pattern, indicating, the present data demonstrate that in the rat hippocampus, ZK91587 binds specifically to the Type I (corticosterone-preferring/mineralocorticoid-like receptor

  17. Reduced secondary cytokine induction by BAY 50-4798, a high-affinity receptor-specific interleukin-2 analog.

    Science.gov (United States)

    Steppan, Sonja; Eckart, Michael R; Bajsarowicz, Krystyna; Sternberg, Lawrence R; Greve, Jeffrey M; Cassell, Delanie J

    2006-03-01

    Recombinant interleukin-2 (IL-2) (aldesleukin, Proleukin, Chiron, Emeryville, CA) is approved for treatment of cancer patients and under investigation in HIV-infected individuals. However, treatment with aldesleukin is associated with toxicity, which may be due to its elicitation of inflammatory mediators from cells that express the intermediate-affinity IL-2 receptor. BAY 50-4798, a novel IL-2 analog, is a selective agonist for the high-affinity receptor. It induces the proliferation of activated T cells with a potency similar to that of aldesleukin but has reduced activity on cells expressing the intermediate-affinity receptor. In the current study, we compared cytokine responses elicited in peripheral blood mononuclear cell (PBMC) cultures stimulated with BAY 50-4798 or aldesleukin. BAY 50-4798 induced approximately 5-fold lower mean levels of endogenous IL-2 than aldesleukin, and at least 50% lower levels of proinflammatory cytokines, such as tumor necrosis fctor-alpha (TNF-alpha), IL-1beta, IL-6, and interferon-gamma (IFN-gamma). Furthermore, statistically significant reductions in the levels of IL-5, IL-8, IL-10, IL-13, and granulocyte-macrophage colony-stimulating factor (GM-CSF) were observed in response to BAY 50-4798. These findings increase our understanding of the biologic action of BAY 50-4798 and suggest a mechanism by which it may exhibit better safety than aldesleukin in humans. PMID:16542139

  18. Distinction between high-affinity [3H]phencyclidine binding sites and muscarinic receptors in guinea-pig ileum muscle

    International Nuclear Information System (INIS)

    [3H]Phencyclidine ([3H]PCP) binding was studied in guinea-pig ileum muscle membranes. Specific binding of [3H]PCP was time dependent, reversible and saturable, with an equilibrium dissociation constant of 154 nM and maximum binding of 12.9 pmol/mg of protein at pH 9. Its pH dependency suggests that the un-ionized PCP is the pharmacologically active form. The binding site was on a protein which was sensitive to heat, proteolytic enzymes and the carboxylic group reagent dicyclohexylcarbodiimide, but insensitive to phospholipase A and C, concanavalin A, dithiothreitol and N-ethylmaleimide. Specific [3H]PCP binding was displaced effectively by several PCP analogs and Ca++ channel antagonists including verapamil, to which these sites had a high affinity. These high-affinity PCP-binding sites were found at a much higher concentration in the same membrane preparation than muscarinic receptor sites identified by their specific binding of [3H]quinuclidinyl benzilate. PCP bound to both sites, but with a lower affinity to the muscarinic receptor sites. The PCP and muscarinic receptor sites differed in their sensitivities to pH and drug specifities

  19. Early Signs of Pathological Cognitive Aging in Mice Lacking High-Affinity Nicotinic Receptors.

    Science.gov (United States)

    Konsolaki, Eleni; Tsakanikas, Panagiotis; Polissidis, Alexia V; Stamatakis, Antonios; Skaliora, Irini

    2016-01-01

    In order to address pathological cognitive decline effectively, it is critical to adopt early preventive measures in individuals considered at risk. It is therefore essential to develop approaches that identify such individuals before the onset of irreversible dementia. A deficient cholinergic system has been consistently implicated as one of the main factors associated with a heightened vulnerability to the aging process. In the present study we used mice lacking high affinity nicotinic receptors (β2-/-), which have been proposed as an animal model of accelerated/premature cognitive aging. Our aim was to identify behavioral signs that could serve as indicators or predictors of impending cognitive decline. We used test batteries in order to assess cognitive functions and additional tasks to investigate spontaneous behaviors, such as species-specific activities and exploration/locomotion in a novel environment. Our data confirm the hypothesis that β2-/- animals exhibit age-related cognitive impairments in spatial learning. In addition, they document age-related deficits in other areas, such as recognition memory, burrowing and nesting building, thereby extending the validity of this animal model for the study of pathological aging. Finally, our data reveal deficits in spontaneous behavior and habituation processes that precede the onset of cognitive decline and could therefore be useful as a non-invasive behavioral screen for identifying animals at risk. To our knowledge, this is the first study to perform an extensive behavioral assessment of an animal model of premature cognitive aging, and our results suggest that β2-nAChR dependent cognitive deterioration progressively evolves from initial subtle behavioral changes to global dementia due to the combined effect of the neuropathology and aging. PMID:27199738

  20. Different sensitivity of activation via Thy-1 and the high-affinity IgE receptor to acute lowering of cholesterol in rat basophilic leukemia cells

    Czech Academy of Sciences Publication Activity Database

    Dráber, Petr

    Chevy Chase, 2000, s. 90. [Meeting of International Research Scholars from Argentina , Belarus, Brazil, Canada, Chile, Czech Republic, Estonia, Hungary, Latvia, Lithuania, Mexico, Poland, Russia, Slovakia, Ukraine, and Venezuela. Chevy Chase (US), 20.06.2000-23.06.2000] Institutional research plan: CEZ:AV0Z5052915 Subject RIV: EB - Genetics ; Molecular Biology

  1. Different sensitivity of activation via Thy-1 and the high-affinity IgE receptor to acute lowering of cholesterol in rat basophilic leukemia cells

    Czech Academy of Sciences Publication Activity Database

    Surviladze, Zurab; Dráberová, Lubica; Kovářová, Martina; Dráber, Petr

    Chevy Chase, 2000. s. 90. [Meeting of International Research Scholars from Argentina , Belarus, Brazil, Canada, Chile, Czech Republic, Estonia, Hungary, Latvia, Lithuania, Mexico, Poland, Russia, Slovakia, Ukraine, and Venezuela. 20.06.2000-23.06.2000, Chevy Chase] Institutional research plan: CEZ:AV0Z5052915 Subject RIV: EB - Genetics ; Molecular Biology

  2. Relative binding affinity of steroids for the corticosterone receptor system in rat hippocampus

    NARCIS (Netherlands)

    De Kloet, E R; Veldhuis, H D; Wagenaars, J L; Bergink, E W

    1984-01-01

    In cytosol of the hippocampus corticosterone displays highest affinity for the sites that remain available for binding in the presence of excess RU 26988, which is shown to be a "pure" glucocorticoid. A rather high affinity (greater than or equal to 25%) was found for 11 beta-hydroxyprogesterone, 21

  3. LGR6 is a high affinity receptor of R-spondins and potentially functions as a tumor suppressor.

    Directory of Open Access Journals (Sweden)

    Xing Gong

    Full Text Available BACKGROUND: LGR6 (leucine-rich repeat containing, G protein-coupled receptor 6 is a member of the rhodopsin-like seven transmembrane domain receptor superfamily with the highest homology to LGR4 and LGR5. LGR6 was found as one of the novel genes mutated in colon cancer through total exon sequencing and its promoter region is hypermethylated in 20-50% of colon cancer cases. In the skin, LGR6 marks a population of stem cells that can give rise to all cell lineages. Recently, we and others demonstrated that LGR4 and LGR5 function as receptors of R-spondins to potentiate Wnt/β-catenin signaling. However, the binding affinity and functional response of LGR6 to R-spondins, and the activity of colon cancer mutants of LGR6 have not been determined. PRINCIPAL FINDINGS: We found that LGR6 also binds and responds to R-spondins 1-3 with high affinity to enhance Wnt/β-catenin signaling through increased LRP6 phosphorylation. Similar to LGR4 and LGR5, LGR6 is not coupled to heterotrimeric G proteins or to β-arrestin following R-spondin stimulation. Functional and expression analysis of three somatic mutations identified in colon cancer samples indicates that one mutant fails to bind and respond to R-spondin (loss-of-function, but the other two have no significant effect on receptor function. Overexpression of wild-type LGR6 in HeLa cells leads to increased cell migration following co-treatment with R-spondin1 and Wnt3a when compared to vector control cells or cells overexpressing the loss-of-function mutant. CONCLUSIONS: LGR6 is a high affinity receptor for R-spondins 1-3 and potentially functions as a tumor suppressor despite its positive effect on Wnt/β-catenin signaling.

  4. Increased hsp70 of glucocorticoid receptor complex induced by scald and heat stress and its possible effect on the affinity of glucocorticoid receptor

    Institute of Scientific and Technical Information of China (English)

    WANG Xiao-hui; TANG Hong-tai; LU Jian; XIA Zhao-fan

    2010-01-01

    Background Glucocorticoid (GC) insensitivity/GC resistance is an important etiological and prognostic factor in multiple diseases and pathophysiological processes such as scald, shock and asthma. The function of GC was mediated by glucocorticoid receptor (GR). Scald not only decreased the expression of GR but also reduced the affinity of GR, which played an important role in GC resistance in scalded rats. Whereas the molecular mechanism responsible for the decrease of GR affinity resulted from scald remains unclear. Recent studies showed that the changes of heat shock proteins (hsp) especially hsp90 and hsp70 of GR heterocomplex were associated with GR low affinity in vitro. Methods The affinity of GR in hepatic cytosols and in the cytosols of SMMC-7721 cells were determined by radioligand binding assay and scatchard plot. GR heterocomplex in cytosols were captured by coimmunoprecipation and the levels of hsp90 and hsp70 of GR complex were detected by quantitative Western blotting.Results Similar with that of hepatic cytosol of scalded rats, a remarkable decrease of GR affinity was also found in the cytosol of heat stressed SMMC-7721 cells. The level of hsp70 of GR complex in hepatic cytosol of scalded rats (30% total body surface area immersion scald) and in cytosol of heat stressed human hepatocarcinoma cell line SMMC-7721 were both increased by 1.5 fold, whereas no change of hsp90 in GR heterocomplex was found. According to the correlation analysis, there may be a positive relationship between increased hsp70 of GR complex and decreased GR affinity in the cytosols.Conclusions The primary results indicated that the level of hsp70 of GR heterocomplex was increased in the hepatic cytosol of scalded rats and the cytosol of heat stressed SMMC-7721 cells. The increase of hsp70 of GR complex might be associated with the decrease of GR affinity.

  5. Affinity of 3-acyl substituted 4-quinolones at the benzodiazepine site of GABAA receptors

    DEFF Research Database (Denmark)

    Lager, Erik; Nilsson, Jakob; Nielsen, Elsebet Østergaard;

    2008-01-01

    The finding that alkyl 1,4-dihydro-4-oxoquinoline-3-carboxylate and N-alkyl-1,4-dihydro-4-oxoquinoline-3-carboxamide derivatives may be high-affinity ligands at the benzodiazepine binding site of the GABA(A) receptor, prompted a study of 3-acyl-1,4-dihydro-4-oxoquinoline (3-acyl-4-quinolones). In......- and a gamma-subunit in the GABA(A) receptor, selected compounds were tested on the alpha(1)beta(2)gamma(2s), alpha(2)beta(2)gamma(2s) and alpha(3)beta(2)gamma(2s) GABA(A) receptor subtypes. The 3-acyl-4-quinolones display various degrees of selectivity for alpha(1)- versus alpha(2)- and alpha(3...

  6. Covalent labeling of a high-affinity, guanyl nucleotide sensitive parathyroid hormone receptor in canine renal cortex

    International Nuclear Information System (INIS)

    Putative parathyroid hormone (PTH) receptors in canine renal membranes were affinity labeled with 125I-bPTH(1-34) using the heterobifunctional cross-linking reagent N-hydroxysuccinimidyl 4-azido-benzoate. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis revealed the presence of a major 85,000 molecular weight (M/sub r/) PTH binding component, the labeling of which was inhibited by nanomolar concentrations of unlabeled PTH and by micromolar concentrations of 5'-guanylyl imidodiphosphate [Gpp-(NH)p]. Labeling was not influenced by the unrelated peptides insulin and arginine vasopressin. Minor PTH binding components of M/sub r/ 55,000 and 130,000 were also seen, and labeling of these was likewise sensitive to unlabeled PTH and to Gpp(NH)p. Omission of protease inhibitors during the isolation of plasma membranes resulted in the loss of the M/sub r/ 85,000 PTH binding species and the appearance of an M/sub r/ 70,000 form. Several minor PTH binding components also were observed. Equilibrium binding studies showed that such membranes had an affinity for PTH indistinguishable from that in membranes isolated with protease inhibitors and displaying a major M/sub r/ 85,000 PTH binding species. The authors conclude that the major form of the adenylate cyclase coupled PTH receptor in canine renal membranes is an M/sub r/ 85,000 protein. An endogenous enzyme, probably a lysosomal cathepsin, can cleave this form to produce an M/sub r/ 70,000 receptor that retains full functional activity with respect to high-affinity, guanyl nucleotide sensitive PTH binding. The ability to covalently label the PTH receptor in high yield represents a major step toward the structural characterization of this important detector molecule

  7. Synthetic cannabinoids: In silico prediction of the cannabinoid receptor 1 affinity by a quantitative structure-activity relationship model.

    Science.gov (United States)

    Paulke, Alexander; Proschak, Ewgenij; Sommer, Kai; Achenbach, Janosch; Wunder, Cora; Toennes, Stefan W

    2016-03-14

    The number of new synthetic psychoactive compounds increase steadily. Among the group of these psychoactive compounds, the synthetic cannabinoids (SCBs) are most popular and serve as a substitute of herbal cannabis. More than 600 of these substances already exist. For some SCBs the in vitro cannabinoid receptor 1 (CB1) affinity is known, but for the majority it is unknown. A quantitative structure-activity relationship (QSAR) model was developed, which allows the determination of the SCBs affinity to CB1 (expressed as binding constant (Ki)) without reference substances. The chemically advance template search descriptor was used for vector representation of the compound structures. The similarity between two molecules was calculated using the Feature-Pair Distribution Similarity. The Ki values were calculated using the Inverse Distance Weighting method. The prediction model was validated using a cross validation procedure. The predicted Ki values of some new SCBs were in a range between 20 (considerably higher affinity to CB1 than THC) to 468 (considerably lower affinity to CB1 than THC). The present QSAR model can serve as a simple, fast and cheap tool to get a first hint of the biological activity of new synthetic cannabinoids or of other new psychoactive compounds. PMID:26795018

  8. Selective κ opioid antagonists nor-BNI, GNTI and JDTic have low affinities for non-opioid receptors and transporters.

    Directory of Open Access Journals (Sweden)

    Thomas A Munro

    Full Text Available BACKGROUND: Nor-BNI, GNTI and JDTic induce selective κ opioid antagonism that is delayed and extremely prolonged, but some other effects are of rapid onset and brief duration. The transient effects of these compounds differ, suggesting that some of them may be mediated by other targets. RESULTS: In binding assays, the three antagonists showed no detectable affinity (K(i≥10 µM for most non-opioid receptors and transporters (26 of 43 tested. There was no non-opioid target for which all three compounds shared detectable affinity, or for which any two shared sub-micromolar affinity. All three compounds showed low nanomolar affinity for κ opioid receptors, with moderate selectivity over μ and δ (3 to 44-fold. Nor-BNI bound weakly to the α(2C-adrenoceptor (K(i = 630 nM. GNTI enhanced calcium mobilization by noradrenaline at the α(1A-adrenoceptor (EC₅₀ = 41 nM, but did not activate the receptor, displace radioligands, or enhance PI hydrolysis. This suggests that it is a functionally-selective allosteric enhancer. GNTI was also a weak M₁ receptor antagonist (K(B = 3.7 µM. JDTic bound to the noradrenaline transporter (K(i = 54 nM, but only weakly inhibited transport (IC₅₀ = 1.1 µM. JDTic also bound to the opioid-like receptor NOP (K(i = 12 nM, but gave little antagonism even at 30 µM. All three compounds exhibited rapid permeation and active efflux across Caco-2 cell monolayers. CONCLUSIONS: Across 43 non-opioid CNS targets, only GNTI exhibited a potent functional effect (allosteric enhancement of α(1A-adrenoceptors. This may contribute to GNTI's severe transient effects. Plasma concentrations of nor-BNI and GNTI may be high enough to affect some peripheral non-opioid targets. Nonetheless, κ opioid antagonism persists for weeks or months after these transient effects dissipate. With an adequate pre-administration interval, our results therefore strengthen the evidence that nor-BNI, GNTI and JDTic are highly

  9. The development of a radioallergosorbent test (RAST) for murine IgE antibodies

    International Nuclear Information System (INIS)

    A purified monoclonal IgE preparation, isolated from the ascitic fluid of mice bearing a hybridoma secreting IgE with specificity to ovalbumin, was used for the production of goat anti-murine IgE (GAME) antiserum, which was then rendered monospecific for the epsilon chain. Another monoclonal hybridoma IgE preparation with specificity for the 2,4-dinitrophenyl group was isolated from ascitic fluid in a relatively pure state by affinity chromatography and used in the form of an immunosorbent to isolate antibodies from the monospecific goat serum. The GAME of antibodies were 125I-labeled and used to develop a radioallergosorbent test (RAST) for the quantitation of murine IgE antibodies specifically adsorbed onto antigen-coupled paper discs. The RAST was specific for antibodies of the IgE class only and was as sensitive as and more accurate than PCA assay. RAST results on sera of mice treated with tolerogenic conjugates indicated a reduction in the affinity and concentration of the IgE antibody populations on suppression of the IgE response. The effect of interference by non-IgE antibody populations on the RAST curves has been discussed. (Auth.)

  10. Developments of segregation process and chemical analysis for virus receptor recognizing environmental signal substance by affinity binding assaying

    International Nuclear Information System (INIS)

    Virus receptors recognizing environmental signal were segregated and analyzed chemically by affinity binding assaying using radio-isotope marked compound. Pathogenic protein in rice stem virus was indicated possibility of a protein which participates in transfer process between the virus cells. An enzyme, Chitobiose was separated from a bacteria, actinomycete. It was cleared by 3H chitobiose affinity binding assaying that two kinds of chitobiose binding protein existed in cell membrane of the actinomycete. A different kind of protein was founded in the cell membrane which was raised in existence of chitobiose. Observation showed that the protein was different from a protein in the cell membrane which was raised in non-existence of chitobiose. These two kinds of protein might be formed a complex compound on the surface of cell membrane. (M. Suetake)

  11. Progress in the discovery of selective, high affinity A2B adenosine receptor antagonists as clinical candidates

    OpenAIRE

    Kalla, Rao V.; Zablocki, Jeff

    2008-01-01

    The selective, high affinity A2B adenosine receptor (AdoR) antagonists that were synthesized by several research groups should aid in determining the role of the A2B AdoR in inflammatory diseases like asthma or rheumatoid arthritis (RA) and angiogenic diseases like diabetic retinopathy or cancer. CV Therapeutics scientists discovered the selective, high affinity A2B AdoR antagonist 10, a 8-(4-pyrazolyl)-xanthine derivative [CVT-6883, Ki(hA2B) = 22 nM; Ki(hA1) = 1,940 nM; Ki(hA2A) = 3,280; and...

  12. Helminth Allergens, Parasite-Specific IgE, and Its Protective Role in Human Immunity

    OpenAIRE

    Fitzsimmons, Colin Matthew; Falcone, Franco Harald; Dunne, David William

    2014-01-01

    The Th2 immune response, culminating in eosinophilia and IgE production, is not only characteristic of allergy but also of infection by parasitic worms (helminths). Anti-parasite IgE has been associated with immunity against a range of helminth infections and many believe that IgE and its receptors evolved to help counter metazoan parasites. Allergens (IgE-antigens) are present in only a small minority of protein families and known IgE targets in helminths belong to these same families (e.g.,...

  13. GABA-agonists induce the formation of low-affinity GABA-receptors on cultured cerebellar granule cells via preexisting high affinity GABA receptors

    DEFF Research Database (Denmark)

    Belhage, B; Meier, E; Schousboe, A

    1986-01-01

    The kinetics of specific GABA-binding to membranes isolated from cerebellar granule cells, cultured for 12 days from dissociated cerebella of 7-day-old rats was studied using [3H]GABA as the ligand. The granule cells were cultured in the presence of the specific GABA receptor agonist 4, 5, 6, 7......-tetrahydroisoxazolo [5,4-c]pyridin-3-ol (THIP, 150 microM) or THIP plus the antagonist bicuculline methobromide (150 microM of each) or in the absence of the agonist or antagonist. Membranes isolated from granule cells cultured in a medium without the GABA agonist revealed a single binding site for GABA with a...... binding constant (KD) of 7.9 +/- 0.4 nM and a Bmax of 3.42 +/- 0.08 pmol X mg-1 protein. Membranes from cells cultured in the presence of THIP had two binding sites for GABA with KD-values of 6.8 +/- 0.9 nM and 476 +/- 311 nM, respectively. The corresponding Bmax values were 4.41 +/- 0.42 pmol X mg-1 and...

  14. The cytoplasmic tail of FcgammaRIIIAalpha is involved in signaling by the low affinity receptor for immunoglobulin G

    DEFF Research Database (Denmark)

    Hou, X; Dietrich, J; Geisler, Carsten

    1996-01-01

    The low affinity receptor for IgG, FcgammaRIIIA, is a multimeric receptor composed of the ligand binding subunit FcgammaRIIIAalpha (CD16) in association with the signal-transducing subunits zeta or gamma. Previous studies suggested that the cytoplasmic tail of FcgammaRIIIAalpha was not required for...... FcgammaRIIIAalpha-zeta association or signaling by FcgammaRIIIA. However, in these studies, the truncated FcgammaRIIIAalpha chains still expressed the four most membrane-proximal amino acids of the cytoplasmic tail (amino acids 230-233). By successive truncations from the C terminus of Fcgamma......RIIIAalpha, we have studied the role played by the membrane-proximal amino acids of the cytoplasmic tail of FcgammaRIIIAalpha in (i) FcgammaRIIIA expression, (ii) FcgammaRIIIAalpha-zeta association, and (iii) signal transduction. We provide evidence that this region is not required for FcgammaRIIIA expression or...

  15. Molecular mechanisms of IgE mediated food allergy.

    Science.gov (United States)

    Kumar, Sandeep; Verma, Alok Kumar; Das, Mukul; Dwivedi, Premendra D

    2012-08-01

    The purpose of this review is to collate current knowledge and recent advances in molecular mechanism behind the immediate type hypersensitivity of foods. Food allergy is a growing concern of human health in developed as well as developing countries now days. Food allergic reactions are mostly IgE mediated and also known as immediate type hypersensitivity or type I reaction. This review encompasses a wide range of molecular events during IgE mediated reactions like primary exposure of allergens, processing of allergens by antigen presenting cells, role of transcription factors like GATA-3, STAT-6, NF-AT, c-maf, c-kit and NF-κB, Treg cells, toll like receptors, cytokines and chemokines, class switch to IgE, FcεR1 receptor, priming of IgE on mast cells or basophils, signaling events followed by secondary exposure of allergens, degranulation and release of mediators like leukotrienes, histamines, prostaglandins, β-hexosaminidase and ultimately anaphylaxis. This review may be helpful to beginners as well as experts working in the field of allergy and immunology because of the stepwise explanations of molecular mechanisms involved in IgE mediated reactions. PMID:22668720

  16. Structural analogs of human insulin-like growth factor I with reduced affinity for serum binding proteins and the type 2 insulin-like growth factor receptor

    International Nuclear Information System (INIS)

    Four structural analogs of human insulin-like growth factor I (hIGF-I) have been prepared by site-directed mutagenesis of a synthetic IGF-I gene and subsequent expression and purification of the mutant protein from the conditioned media of transformed yeast. [Phe-1, Val1, Asn2, Gln3, His4, Ser8, His9, Glu12, Tyr15, Leu16]IGF-I (B-chain mutant), in which the first 16 amino acids of hIGF-I were replaced with the first 17 amino acids of the B-chain of insulin, has >1000-, 100-, and 2-fold reduced potency for human serum binding proteins, the rat liver type 2 IGF receptor, and the human placental type 1 IGF receptor, respectively. The B-chain mutant also has 4-fold increased affinity for the human placental insulin receptor. [Gln3, Ala4] IGF-I has 4-fold reduced affinity for human serum binding proteins, but is equipotent to hIGF-I at the types 1 and 2 IGF and insulin receptors. [Tyr15, Leu16] IGH-I has 4-fold reduced affinity for human serum binding proteins and 10-fold increased affinity for the insulin receptor. The peptide in which these four-point mutations are combined, [Gln3, Ala4, Tyr15,Leu16]IGF-I, has 600-fold reduced affinity for the serum binding proteins. All four of these mutants stimulate DNA synthesis in the rat vascular smooth muscle cell line A10 with potencies reflecting their potency at the type 1 IGF receptor. These studies identify some of the domains of hIGF-I which are responsible for maintaining high affinity binding with the serum binding protein and the type 2 IGF receptor. In addition, These peptides will be useful in defining the role of the type 2 IGF receptor and serum binding proteins in the physiological actions of hIGF-I

  17. [3H]-DOB(4-bromo-2,5-dimethoxyphenylisopropylamine) and [3H] ketanserin label two affinity states of the cloned human 5-hydroxytryptamine2 receptor

    International Nuclear Information System (INIS)

    The binding properties of the 5-hydroxytryptamine2 (5-HT2) receptor have been the subject of much interest and debate in recent years. The hallucinogenic amphetamine derivative 4-bromo-2,5-dimethoxyphenylisopropylamine (DOB) has been shown to bind to a small number of binding sites with properties very similar to [3H]ketanserin-labeled 5-HT2 receptors, but with much higher agonist affinities. Some researchers have interpreted this as evidence for the existence of a new subtype of 5-HT2 receptor (termed 5-HT2A), whereas others have interpreted these data as indicative of agonist high affinity and agonist low affinity states for the 5-HT2 receptor. In this investigation, a cDNA clone encoding the serotonin 5-HT2 receptor was transiently transfected into monkey kidney Cos-7 cells and stably transfected into mouse fibroblast L-M(TK-) cells. In both systems, expression of this single serotonin receptor cDNA led to the appearance of both [3H]DOB and [3H]ketanserin binding sites with properties that matched their binding characteristics in mammalian brain homogenates. Addition of guanosine 5'-(beta, gamma-imido) triphosphate [Gpp(NH)p] to this system caused a rightward shift and steepening of agonist competition curves for [3H] ketanserin binding, converting a two-site binding curve to a single low affinity binding state. Gpp(NH)p addition also caused a 50% decrease in the number of high affinity [3H]DOB binding sites, with no change in the dissociation constant of the remaining high affinity states. These data on a single human 5-HT2 receptor cDNA expressed in two different transfection host cells indicate that [3H]DOB and [3H]ketanserin binding reside on the same gene product, apparently interacting with agonist and antagonist conformations of a single human 5-HT2 receptor protein

  18. Altered endothelin receptor expression and affinity in spontaneously hypertensive rat cerebral and coronary arteries

    DEFF Research Database (Denmark)

    Cao, Lei; Cao, Yong-Xiao; Xu, Cang-Bao;

    2013-01-01

    BACKGROUND: Hypertension is associated with arterial hyperreactivity, and endothelin (ET) receptors are involved in vascular pathogenesis. The present study was performed to examine the hypothesis that ET receptors were altered in cerebral and coronary arteries of spontaneously hypertensive rats...... (SHR). METHODOLOGY/PRINCIPAL FINDINGS: Cerebral and coronary arteries were removed from SHR. Vascular contraction was recorded using a sensitive myograph system. Real-time PCR and Western blotting were used to quantify mRNA and protein expression of receptors and essential MAPK pathway molecules. The...... results demonstrated that both ETA and ETB receptor-mediated contractile responses in SHR cerebral arteries were shifted to the left in a nonparallel manner with increased maximum contraction compared with Wistar-Kyoto (WKY) rats. In SHR coronary arteries, the ETA receptor-mediated contraction curve was...

  19. Micro-structured peptide surfaces for the detection of high-affinity peptide-receptor interactions in living cells.

    Science.gov (United States)

    Lipp, Anna-Maria; Ji, Bozhi; Hager, Roland; Haas, Sandra; Schweiggl, Simone; Sonnleitner, Alois; Haselgrübler, Thomas

    2015-12-15

    Peptide ligands have great potential as selective agents for diagnostic imaging and therapeutic targeting of human cancers. A number of high-throughput assays for screening potential candidate peptides have been developed. Although these screening assays are indispensable for the identification of peptide ligands at a large scale, it is crucial to validate peptide binding and selectivity for targeted receptors in a live-cell context. For testing high-affinity peptide-receptor interactions in the plasma membrane of living cells, we developed cell-resistant, micro-structured glass surfaces with high-density and high-contrast peptide features. Cell adhesion and recruitment of fluorescent receptors to micro-patterned peptides in the live-cell membrane were evaluated by reflection interference contrast (RIC) and total internal reflection (TIRF) microscopy, respectively. To demonstrate both the specificity and modularity of the assay, co-patterning of fluorescent receptors with three different immobilized micro-structured ligands was shown: first, interaction of green fluorescent protein (GFP)-tagged epidermal growth factor (EGF) receptor expressed in Jurkat cells with immobilized EGF was detected and quantified. Second, using Jurkat cells, we demonstrated specific interaction of yellow fluorescent protein (YFP)-tagged β3 integrin with c(RGDfK) peptide. Third, we identified indirect recruitment of GFP-tagged α5 integrin to an 11-mer peptide. In summary, our results show that the developed micro-structured surfaces are a useful tool for the validation and quantification of peptide-receptor interactions in their natural cellular environment. PMID:26210593

  20. Old Mice Lacking High-Affinity Nicotine Receptors Resist Acoustic Trauma

    OpenAIRE

    Shen, Haiyan; Lin, Zhaoyu; Lei, Debin; Han, Josiah; Ohlemiller, Kevin K.; Bao, Jianxin

    2011-01-01

    There is presently no clearly effective preventative medication against noise-induced hearing loss (NIHL). However, negative feedback systems that presumably evolved to modulate the sensitivity of the organ of Corti may incidentally confer protection. One feedback system implicated in protection from NIHL involves synaptic connections between the lateral olivocochlear efferent terminals and the afferent fibers of spiral ganglion neurons (SGNs). These connections operate via high affinity nico...

  1. Isothermal Chemical Denaturation to Determine Binding Affinity of Small Molecules to G-Protein Coupled Receptors

    OpenAIRE

    Ross, Patrick; Weihofen, Wilhelm; Siu, Fai; Xie, Amy; Katakia, Hetal; Wright, S. Kirk; Hunt, Ian; Brown, Richard K; Freire, Ernesto

    2014-01-01

    The determination of accurate binding affinities is critical in drug discovery and development. Several techniques are available for characterizing the binding of small molecules to soluble proteins. The situation is different for integral membrane proteins. Isothermal chemical denaturation (ICD) has been shown to be a valuable biophysical method to determine in a direct and label-free fashion the binding of ligands to soluble proteins. In this communication, the application of isothermal che...

  2. Early Signs of Pathological Cognitive Aging in Mice Lacking High-Affinity Nicotinic Receptors

    OpenAIRE

    Konsolaki, Eleni; Tsakanikas, Panagiotis; Polissidis, Alexia V.; Stamatakis, Antonios; Skaliora, Irini

    2016-01-01

    In order to address pathological cognitive decline effectively, it is critical to adopt early preventive measures in individuals considered at risk. It is therefore essential to develop approaches that identify such individuals before the onset of irreversible dementia. A deficient cholinergic system has been consistently implicated as one of the main factors associated with a heightened vulnerability to the aging process. In the present study we used mice lacking high affinity nicotinic rece...

  3. CYCLIC CHOLECYSTOKININ ANALOGUES EXHIBIT HIGH BLOOD STABILITY AND BINDING AFFINITY WITH CHOLECYSTOKININ RECEPTOR

    OpenAIRE

    Eun-Ha Joh; Jae Cheong Lim; Jin Joo Kim; Sang Mu Choi; Sun-Ju Choi

    2014-01-01

    Recently, incidence of Cholecystokinin (CCK) receptor is recognized as a factor that determines the aggressive phenotype of pancreatic cancer. In this study, a novel Cholecystokinin (CCK) analogues; DOTA-Nle-cyclo (Glu-Trp-Met-Asp-Phe-Lys-NH2) (DOTA-cCCK) and DOTA-Nle-cyclo (Glu-Trp-Nle-Asp-Phe-Lys-NH2) (DOTA-[Nle]-cCCK) were synthesized and radiolabeled and the targeting abilities on the CCK receptor were evaluated for new CCK receptor targeting agents searching. Peptides were prepared throu...

  4. Interactions of dopamine agonists with brain D1 receptors labeled by 3H-antagonists. Evidence for the presence of high and low affinity agonist-binding states

    International Nuclear Information System (INIS)

    The interactions of dopaminergic agonists and antagonists with 3H-antagonist labeled D1 dopamine receptors of rat striatum have been characterized. [3H]Flupentixol has been found to selectively label D1 dopamine receptors when its binding to D2 dopamine receptors is blocked by the inclusion of D2 selective concentrations of unlabeled spiroperidol or domperidone. Antagonist/3H-antagonist competition curves are of uniformly steep slope (nH . 1.0) suggesting the presence of a single D1 dopamine receptor. Agonist/3H-antagonist competition curves are extremely shallow (nH less than or equal to 0.5) for agonists of high relative efficacy, suggesting the presence of heterogeneous populations of agonist-binding states of the D1 dopamine receptor. Computer-modeling techniques were used to estimate affinities and relative site densities for these heterogeneous binding states. This analysis indicates that the ratio of agonist affinities for low and high affinity agonist-binding states is correlated with agonist relative efficacies in activating adenylate cyclase in membrane homogenates. Under the assay conditions employed, the addition of saturating concentrations of guanine nucleotides reduced, but did not abolish, the relative density of high affinity agonist-binding sites. These binding data can, at least in part, be explained by postulating two states of the D1 dopamine receptor, inducible by agonists but not by antagonists and modulated by guanine nucleotides

  5. Progress in the discovery of selective, high affinity A(2B) adenosine receptor antagonists as clinical candidates.

    Science.gov (United States)

    Kalla, Rao V; Zablocki, Jeff

    2009-03-01

    The selective, high affinity A(2B) adenosine receptor (AdoR) antagonists that were synthesized by several research groups should aid in determining the role of the A(2B) AdoR in inflammatory diseases like asthma or rheumatoid arthritis (RA) and angiogenic diseases like diabetic retinopathy or cancer. CV Therapeutics scientists discovered the selective, high affinity A(2B) AdoR antagonist 10, a 8-(4-pyrazolyl)-xanthine derivative [CVT-6883, K(i)(hA(2B)) = 22 nM; K(i)(hA(1)) = 1,940 nM; K(i)(hA(2A)) = 3,280; and K(i)(hA(3)) = 1,070 nM] that has favorable pharmacokinetic (PK) properties (t (1/2) = 4 h and F > 35% rat). Compound 10 demonstrated functional antagonism at the A(2B) AdoR (K(B) = 6 nM) and efficacy in a mouse model of asthma. In two phase 1 clinical trials, CVT-6883 was found to be safe, well tolerated, and suitable for once daily dosing. A second compound 20, 8-(5-pyrazolyl)-xanthine, has been nominated for development from Baraldi's group in conjunction with King Pharmaceuticals that has favorable A(2B) AdoR affinity and selectivity [K(i)(hA(2B)) = 5.5 nM; K(i)(hA(1)) > 1,000 nM; K(i)(hA(2A)) > 1,000; and K(i)(hA(3)) > 1,000 nM], and it has been demonstrated to be a functional antagonist. A third compound 32, a 2-aminopyrimidine, from the Almirall group has high A(2B) AdoR affinity and selectivity [K(i)(hA(2B)) = 17 nM; K(i)(hA(1)) > 1,000 nM; K(i)(hA(2A)) > 2,500; and K(i)(hA(3)) > 1,000 nM], and 32 has been moved into preclinical safety testing. Since three highly selective, high affinity A(2B) AdoR antagonists have been nominated for development with 10 (CVT-6883) being the furthest along in the development process, the role of the A(2B) AdoR in various disease states will soon be established. PMID:18568423

  6. Changes in parathyroid hormone receptor binding affinity during egg laying: implications for calcium homeostasis in chicken.

    Science.gov (United States)

    Yasuoka, T; Kawashima, M; Takahashi, T; Iwata, A; Oka, N; Tanaka, K

    1996-12-01

    Parathyroid hormone (PTH) receptor bindings were examined in the membrane fraction of the calvaria and the kidney of the hen by the use of [125I]PTH-related protein (PTHrP) binding assays. The binding specificity, reversibility, and saturation of the receptor were demonstrated. The equilibrium dissociation constant (Kd) and the maximum binding capacity (Bmax) were obtained by Scatchard analyses. In both calvaria and kidney, Kd and Bmax values decreased at 3 h before oviposition in egg-laying hens, but not in nonlaying hens. Administration of 17 beta-estradiol or progesterone in vivo caused a decrease in the Kd and Bmax values. Ionized calcium concentrations in the blood plasma showed a decrease at 13 h before oviposition. The results suggest that the PTH receptor binding in the calvaria and the kidney is affected by ovarian steroid hormones and may play a role in maintaining the calcium homeostasis in the egg-laying hen. PMID:8970893

  7. Preparation and characterization of chemical affinity probes for the hepatic galactosyl receptor

    International Nuclear Information System (INIS)

    The authors have developed a protocol to covalently attach asialo-orosomucoid (ASOR) to the Galactosyl receptor. 125I-ASOR was modified to an NHS ester by reaction with either disuccinimidyl suberate to give NHS-125I-ASOR or dithiobis(succinimidylpropionate) to give NHS-S-S125I-ASOR. The specificity of binding of the purified derivatives to isolated rat hepatocytes at 40C, assessed in the presence of ASOR, was >90%. Derivatized ASOR was shown to be covalently cross-linked by the inability of EGTA, to release radioactivity. Optimum cross-linking of the specifically bound derivative was ≥40%. The specificity of cross-linking to receptor with both cross-linkers was ∼80% as determined by immunoprecipitation of EGTA-treated cells with a goat anti-receptor antisera. SDS-PAGE and autoradiography of extracts from cells cross-linked with either derivative revealed bands of M/sub r/ ∼ 108-110 kDa, and 82 kDa under both reducing and nonreducing conditions. Thus, receptor and ASOR could be cross-linked with both high efficiency and high specificity. In order to assess cellular internalization of cross-linked receptor-ligand complexes, cells were treated with dithiothreitol or trypsin to remove, respectively, only the surface bound NHS-S-S-125I-ASOR or NHS-125I-ASOR. The kinetics of internalization for both derivatized or underivatized ASOR were identical. These results directly show for the first time that both surface receptor and ligand are internalized

  8. Searsia species with affinity to the N-methyl-d-aspartic acid (NMDA) receptor

    DEFF Research Database (Denmark)

    Jäger, Anna; Knap, D.M.; Nielsen, Birgitte;

    2012-01-01

    Species of Searsia are used in traditional medicine to treat epilepsy. Previous studies on S. dentata and S. pyroides have shown that this is likely mediated via the N-methyl-d-aspartic acid (NMDA) receptor. Ethanolic extracts of leaves of six Searsia species were tested in a binding assay for...... accessible Searsia species can be used in traditional medicine....

  9. CYCLIC CHOLECYSTOKININ ANALOGUES EXHIBIT HIGH BLOOD STABILITY AND BINDING AFFINITY WITH CHOLECYSTOKININ RECEPTOR

    Directory of Open Access Journals (Sweden)

    Eun-Ha Joh

    2014-01-01

    Full Text Available Recently, incidence of Cholecystokinin (CCK receptor is recognized as a factor that determines the aggressive phenotype of pancreatic cancer. In this study, a novel Cholecystokinin (CCK analogues; DOTA-Nle-cyclo (Glu-Trp-Met-Asp-Phe-Lys-NH2 (DOTA-cCCK and DOTA-Nle-cyclo (Glu-Trp-Nle-Asp-Phe-Lys-NH2 (DOTA-[Nle]-cCCK were synthesized and radiolabeled and the targeting abilities on the CCK receptor were evaluated for new CCK receptor targeting agents searching. Peptides were prepared through a solid phase synthesis method and their purity was over 98%. DOTA is the chelating agent for 68Ga-labelling, which the peptides were radiolabeled with 68Ga by a high radiolabeling yield (>98%. Peptides were stable over 98% by incubation in mouse blood at 37°C for 2 h. A competitive displacement of 125I-CCK8 on the AR42J human pancreatic carcinoma cells revealed that 50% inhibitory concentration value (IC50 were 12.31 nM of DOTA-cCCK and 1.69 nM of DOTA-[Nle]-cCCK. Stable in the blood of both DOTA-cCCK and DOTA-[Nle]-cCCK, but the binding rate with the CCK receptor on AR42J cells, DOTA-[Nle]-cCCK confirmed better than DOTA-cCCK. Therefore, it is concluded that 68Ga-DOTA-[Nle]-cCCK can be potential candidate as a targeting modality for the CCK receptor over-expressing tumors and further studies to evaluate their biological characteristics are needed.

  10. Hyper IgE syndrome

    Directory of Open Access Journals (Sweden)

    Yun Dang

    2014-01-01

    Full Text Available Hyper IgE syndrome (HIES is a rare disorder characterized by eczema, recurrent infections of the skin and lungs, typically with Staphylococcus aureus, Candida albicans and certain viruses, and elevated levels of serum IgE. Other clinical manifestations include characteristic facies (prominent forehead, broad nasal bridge and facial asymmetry, chronic eczematous dermatitis, retained primary dentition, recurrent pathological fractures, hyper-extensibility and scoliosis. The central nervous system (CNS involvement in HIES has been rarely reported. Here we presented a case of HIES with rare associations of epilepsy in a young patient to raise awareness for this disorder.

  11. Synthesis and in vitro affinities of various MDL 100907 derivatives as potential F-18-radioligands for 5-HT2A receptor imaging with PET

    DEFF Research Database (Denmark)

    Herth, Matthias Manfred; Kramer, Vasko; Piel, Markus; Palner, Mikael; Riss, Patrick J.; Knudsen, Gitte M.; Roesch, Frank

    2009-01-01

    improve molecular imaging properties of [(11)C]MDL 100907. Their in vitro affinities to a broad spectrum of neuroreceptors and their lipophilicities were determined and compared to the clinically used reference compounds MDL 100907 and altanserin. The novel compounds MA-1 (53) and (R)-MH.MZ (56) show K......(i)-values in the nanomolar range towards the 5-HT(2A) receptor and insignificant binding to other 5-HT receptor subtypes or receptors. Interestingly, compounds MA-1 (53), MH.MZ (55) and (R)-MH.MZ (56) provide a receptor selectivity profile similar to MDL 100907. These compounds could possibly be preferable...... radiotracers altanserin and MDL 100907. The novel compounds MA-1 (53) and (R)-MH.MZ (56) thus appear to be promising high affine and selective tracers of (18)F-labelled analogues for 5-HT(2A) imaging with PET....

  12. Prediction of binding affinity and efficacy of thyroid hormone receptor ligands using QSAR and structure-based modeling methods

    International Nuclear Information System (INIS)

    The thyroid hormone receptor (THR) is an important member of the nuclear receptor family that can be activated by endocrine disrupting chemicals (EDC). Quantitative Structure–Activity Relationship (QSAR) models have been developed to facilitate the prioritization of THR-mediated EDC for the experimental validation. The largest database of binding affinities available at the time of the study for ligand binding domain (LBD) of THRβ was assembled to generate both continuous and classification QSAR models with an external accuracy of R2 = 0.55 and CCR = 0.76, respectively. In addition, for the first time a QSAR model was developed to predict binding affinities of antagonists inhibiting the interaction of coactivators with the AF-2 domain of THRβ (R2 = 0.70). Furthermore, molecular docking studies were performed for a set of THRβ ligands (57 agonists and 15 antagonists of LBD, 210 antagonists of the AF-2 domain, supplemented by putative decoys/non-binders) using several THRβ structures retrieved from the Protein Data Bank. We found that two agonist-bound THRβ conformations could effectively discriminate their corresponding ligands from presumed non-binders. Moreover, one of the agonist conformations could discriminate agonists from antagonists. Finally, we have conducted virtual screening of a chemical library compiled by the EPA as part of the Tox21 program to identify potential THRβ-mediated EDCs using both QSAR models and docking. We concluded that the library is unlikely to have any EDC that would bind to the THRβ. Models developed in this study can be employed either to identify environmental chemicals interacting with the THR or, conversely, to eliminate the THR-mediated mechanism of action for chemicals of concern. - Highlights: • This is the largest curated dataset for ligand binding domain (LBD) of the THRβ. • We report the first QSAR model for antagonists of AF-2 domain of THRβ. • A combination of QSAR and docking enables prediction of both

  13. Prediction of binding affinity and efficacy of thyroid hormone receptor ligands using QSAR and structure-based modeling methods

    Energy Technology Data Exchange (ETDEWEB)

    Politi, Regina [Laboratory for Molecular Modeling, Division of Chemical Biology and Medicinal Chemistry, University of North Carolina, Chapel Hill, NC 27599 (United States); Department of Environmental Sciences and Engineering, University of North Carolina, Chapel Hill, NC 27599 (United States); Rusyn, Ivan, E-mail: iir@unc.edu [Department of Environmental Sciences and Engineering, University of North Carolina, Chapel Hill, NC 27599 (United States); Tropsha, Alexander, E-mail: alex_tropsha@unc.edu [Laboratory for Molecular Modeling, Division of Chemical Biology and Medicinal Chemistry, University of North Carolina, Chapel Hill, NC 27599 (United States)

    2014-10-01

    The thyroid hormone receptor (THR) is an important member of the nuclear receptor family that can be activated by endocrine disrupting chemicals (EDC). Quantitative Structure–Activity Relationship (QSAR) models have been developed to facilitate the prioritization of THR-mediated EDC for the experimental validation. The largest database of binding affinities available at the time of the study for ligand binding domain (LBD) of THRβ was assembled to generate both continuous and classification QSAR models with an external accuracy of R{sup 2} = 0.55 and CCR = 0.76, respectively. In addition, for the first time a QSAR model was developed to predict binding affinities of antagonists inhibiting the interaction of coactivators with the AF-2 domain of THRβ (R{sup 2} = 0.70). Furthermore, molecular docking studies were performed for a set of THRβ ligands (57 agonists and 15 antagonists of LBD, 210 antagonists of the AF-2 domain, supplemented by putative decoys/non-binders) using several THRβ structures retrieved from the Protein Data Bank. We found that two agonist-bound THRβ conformations could effectively discriminate their corresponding ligands from presumed non-binders. Moreover, one of the agonist conformations could discriminate agonists from antagonists. Finally, we have conducted virtual screening of a chemical library compiled by the EPA as part of the Tox21 program to identify potential THRβ-mediated EDCs using both QSAR models and docking. We concluded that the library is unlikely to have any EDC that would bind to the THRβ. Models developed in this study can be employed either to identify environmental chemicals interacting with the THR or, conversely, to eliminate the THR-mediated mechanism of action for chemicals of concern. - Highlights: • This is the largest curated dataset for ligand binding domain (LBD) of the THRβ. • We report the first QSAR model for antagonists of AF-2 domain of THRβ. • A combination of QSAR and docking enables

  14. Mass dose effects and in vivo affinity in brain PET receptor studies — a study of cerebral 5-HT4 receptor binding with [11C]SB207145

    International Nuclear Information System (INIS)

    Attention to tracer dose principles is crucial in positron emission tomography (PET), and deviations can induce serious errors. In this study, we devise a method for determining receptor occupancy of the mass dose of the radioligand itself and the in vivo affinity. Methods: The approach was used for [11C]SB207145, a new PET radioligand for imaging the cerebral 5-HT4 receptors in humans. Test–retest PET studies with varying specific activities of [11C]SB207145 were conducted in seven healthy subjects, and the output parameter regional BPND was modeled. Individual occupancy plots were first computed to estimate the mass dose that saturates 50% of receptors (ID50), and subsequently, the maximal mass dose that can be injected (arbitrarily set at an occupancy D. Results: Increasing the mass dose resulted in a decrease in BPND, whilst the relative cerebellar uptake was unchanged. The ID50 was 85.4±30.2 μg, and the upper mass dose limit was 4.5±1.6 μg, which does not require ultrahigh specific activity. The estimated in vivo KD was 2.8 nM (range 1.0–4.8), without any regional differences. Conclusion: The presented method for estimating the upper mass dose limit is suggested as part of validation of PET radioligands.

  15. Differential effects of chronic hyperammonemia on modulation of the glutamate-nitric oxide-cGMP pathway by metabotropic glutamate receptor 5 and low and high affinity AMPA receptors in cerebellum in vivo.

    Science.gov (United States)

    Cabrera-Pastor, Andrea; Llansola, Marta; Reznikov, Vitaliy; Boix, Jordi; Felipo, Vicente

    2012-07-01

    Previous studies show that chronic hyperammonemia impairs learning ability of rats by impairing the glutamate-nitric oxide (NO)-cyclic guanosine mono-phosphate (cGMP) pathway in cerebellum. Three types of glutamate receptors cooperate in modulating the NO-cGMP pathway: metabotropic glutamate receptor 5 (mGluR5), (RS)-α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) and N-methyl-d-aspartic acid (NMDA) receptors. The aim of this work was to assess whether hyperammonemia alters the modulation of this pathway by mGluR5 and AMPA receptors in cerebellum in vivo. The results support that in control rats: (1) low AMPA concentrations (0.1mM) activate nearly completely Ca(2+)-permeable (glutamate receptor subunit 2 (GluR2)-lacking) AMPA receptors and the NO-cGMP pathway; (2) higher AMPA concentrations (0.3 mM) also activate Ca(2+)-impermeable (GluR2-containing) AMPA receptors, leading to activation of NMDA receptors and of NO-cGMP pathway. Moreover, the data support that chronic hyperammonemia: (1) reduces glutamate release and activation of the glutamate-NO-cGMP pathway by activation of mGluR5; (2) strongly reduces the direct activation by AMPA receptors of the NO-cGMP pathway, likely due to reduced entry of Ca(2+) through GluR2-lacking, high affinity AMPA receptors; (3) strongly increases the indirect activation of the NO-cGMP pathway by high affinity AMPA receptors, likely due to increased entry of Na(+) through GluR2-lacking AMPA receptors and NMDA receptors activation; (4) reduces the indirect activation of the NO-cGMP pathway by low affinity AMPA receptors, likely due to reduced activation of NMDA receptors. PMID:22521775

  16. NMR and computational evidence that high-affinity bradykinin receptor antagonists adopt C-terminal beta-turns.

    Science.gov (United States)

    Kyle, D J; Blake, P R; Smithwick, D; Green, L M; Martin, J A; Sinsko, J A; Summers, M F

    1993-05-14

    Three tetrapeptides were prepared, each corresponding to the four C-terminal amino acid residues of highly potent, second-generation bradykinin receptor antagonists. The tetrapeptides are (IA) Ser-D-Phe-Oic-Arg, (IIA) Ser-D-Tic-Oic-Arg, and (IIIA) Ser-D-Hype(trans-propyl)-Oic-Arg. Solution conformations for each were determined by incorporating interproton distance restraints, determined by 2D NMR experiments performed in water at neutral pH, into a series of distance geometry/simulated annealing model building calculations. Similarly, systematic conformational analyses were performed for each using molecular mechanics calculations. Both the NMR-derived structures, as well as the calculated structures, are shown to adopt a beta-turn as the primary conformation. Excellent agreement between the predicted structures and the NMR-derived structures is demonstrated. Aside from being the first examples of linear tetrapeptides reported to be ordered in aqueous solvent, the results presented support the hypothesis that high-affinity bradykinin receptor antagonists must adopt C-terminal beta-turn conformations. PMID:8388469

  17. [3H]naloxone as an opioid receptor label: Analysis of binding site heterogeneity and use for determination of opioid affinities of casomorphin analogues

    International Nuclear Information System (INIS)

    The nonselective antagonist [3H]naloxone was used to identify opioid receptors in rat brain membranes. The multiple naloxone binding sites were related to different opioid receptors by means of selective opiod ligands as well as various β-casomorphin analogues. Analysis of binding site heterogeneity was performed using several computer curve fitting methods. The results indicate that structurally modified casomorphin peptides are able to discriminate between μ1 and μ2 binding sites. The affinities to the μ sites obtained with [3H]naloxone as label are in a good agreement with those from experiments with the μ selective radioligand [3H]DAGO. The μ1 site affinities of these casomorphin derivatives are well correlated with their antinociceptive potencies. This finding suggests the mediation of the analgesic activity via the high-affinity μ1 subtype. (author)

  18. The ketamine analogue methoxetamine and 3- and 4-methoxy analogues of phencyclidine are high affinity and selective ligands for the glutamate NMDA receptor.

    Directory of Open Access Journals (Sweden)

    Bryan L Roth

    Full Text Available In this paper we determined the pharmacological profiles of novel ketamine and phencyclidine analogues currently used as 'designer drugs' and compared them to the parent substances via the resources of the National Institute of Mental Health Psychoactive Drug Screening Program. The ketamine analogues methoxetamine ((RS-2-(ethylamino-2-(3-methoxyphenylcyclohexanone and 3-MeO-PCE (N-ethyl-1-(3-methoxyphenylcyclohexanamine and the 3- and 4-methoxy analogues of phencyclidine, (1-[1-(3-methoxyphenylcyclohexyl]piperidine and 1-[1-(4-methoxyphenylcyclohexyl]piperidine, were all high affinity ligands for the PCP-site on the glutamate NMDA receptor. In addition methoxetamine and PCP and its analogues displayed appreciable affinities for the serotonin transporter, whilst the PCP analogues exhibited high affinities for sigma receptors. Antagonism of the NMDA receptor is thought to be the key pharmacological feature underlying the actions of dissociative anaesthetics. The novel ketamine and PCP analogues had significant affinities for the NMDA receptor in radioligand binding assays, which may explain their psychotomimetic effects in human users. Additional actions on other targets could be important for delineating side-effects.

  19. In vivo measurement of haloperidol affinity to dopamine D2/D3 receptors by [123I]IBZM and single photon emission computed tomography

    DEFF Research Database (Denmark)

    Videbaek, C; Toska, K; Friberg, L;

    2001-01-01

    This study examines the feasibility of a steady-state bolus-integration method with the dopamine D2/D3 receptor single photon emission computer tomography (SPECT) tracer, [123I]IBZM, for determination of in vivo affinity of haloperidol. The nonspecific binding of [123I]IBZM was examined in the rat...

  20. Dissecting the Relation between a nuclear receptor and GATA: binding affinity studies of thyroid hormone receptor and GATA2 on TSHβ promoter.

    Directory of Open Access Journals (Sweden)

    Ana Carolina Migliorini Figueira

    Full Text Available BACKGROUND: Much is known about how genes regulated by nuclear receptors (NRs are switched on in the presence of a ligand. However, the molecular mechanism for gene down-regulation by liganded NRs remains a conundrum. The interaction between two zinc-finger transcription factors, Nuclear Receptor and GATA, was described almost a decade ago as a strategy adopted by the cell to up- or down-regulate gene expression. More recently, cell-based assays have shown that the Zn-finger region of GATA2 (GATA2-Zf has an important role in down-regulation of the thyrotropin gene (TSHβ by liganded thyroid hormone receptor (TR. METHODOLOGY/PRINCIPAL FINDINGS: In an effort to better understand the mechanism that drives TSHβ down-regulation by a liganded TR and GATA2, we have carried out equilibrium binding assays using fluorescence anisotropy to study the interaction of recombinant TR and GATA2-Zf with regulatory elements present in the TSHβ promoter. Surprisingly, we observed that ligand (T3 weakens TR binding to a negative regulatory element (NRE present in the TSHβ promoter. We also show that TR may interact with GATA2-Zf in the absence of ligand, but T3 is crucial for increasing the affinity of this complex for different GATA response elements (GATA-REs. Importantly, these results indicate that TR complex formation enhances DNA binding of the TR-GATA2 in a ligand-dependent manner. CONCLUSIONS: Our findings extend previous results obtained in vivo, further improving our understanding of how liganded nuclear receptors down-regulate gene transcription, with the cooperative binding of transcription factors to DNA forming the core of this process.

  1. Structural analogs of human insulin-like growth factor I with reduced affinity for serum binding proteins and the type 2 insulin-like growth factor receptor

    Energy Technology Data Exchange (ETDEWEB)

    Bayne, M.L.; Applebaum, J.; Chicchi, G.G.; Hayes, N.S.; Green, B.G.; Cascieri, M.A.

    1988-05-05

    Four structural analogs of human insulin-like growth factor I (hIGF-I) have been prepared by site-directed mutagenesis of a synthetic IGF-I gene and subsequent expression and purification of the mutant protein from the conditioned media of transformed yeast. (Phe/sup -1/, Val/sup 1/, Asn/sup 2/, Gln/sup 3/, His/sup 4/, Ser/sup 8/, His/sup 9/, Glu/sup 12/, Tyr/sup 15/, Leu/sup 16/)IGF-I (B-chain mutant), in which the first 16 amino acids of hIGF-I were replaced with the first 17 amino acids of the B-chain of insulin, has >1000-, 100-, and 2-fold reduced potency for human serum binding proteins, the rat liver type 2 IGF receptor, and the human placental type 1 IGF receptor, respectively. The B-chain mutant also has 4-fold increased affinity for the human placental insulin receptor. (Gln/sup 3/, Ala/sup 4/) IGF-I has 4-fold reduced affinity for human serum binding proteins, but is equipotent to hIGF-I at the types 1 and 2 IGF and insulin receptors. (Tyr/sup 15/, Leu/sup 16/) IGH-I has 4-fold reduced affinity for human serum binding proteins and 10-fold increased affinity for the insulin receptor. The peptide in which these four-point mutations are combined, (Gln/sup 3/, Ala/sup 4/, Tyr/sup 15/,Leu/sup 16/)IGF-I, has 600-fold reduced affinity for the serum binding proteins. All four of these mutants stimulate DNA synthesis in the rat vascular smooth muscle cell line A10 with potencies reflecting their potency at the type 1 IGF receptor. These studies identify some of the domains of hIGF-I which are responsible for maintaining high affinity binding with the serum binding protein and the type 2 IGF receptor. In addition, These peptides will be useful in defining the role of the type 2 IGF receptor and serum binding proteins in the physiological actions of hIGF-I.

  2. High-affinity insulin binding to an atypical insulin-like growth factor-I receptor in human breast cancer cells.

    OpenAIRE

    Milazzo, G.; Yip, C. C.; Maddux, B A; Vigneri, R; Goldfine, I D

    1992-01-01

    We studied the nature of insulin receptor binding in MCF-7 breast cancer cells. In both intact cells and solubilized receptor preparations, high-affinity insulin binding was seen. However, unlabeled insulin-like growth factor-I (IGF-I) was five-fold more potent in inhibiting 125I-insulin binding than insulin itself. With monoclonal antibodies to the insulin receptor, 30% of 125I-insulin binding was inhibited. In contrast when alpha-IR3, a monoclonal antibody that recognizes typical IGF-I rece...

  3. A transforming growth factor β (TGF-β) receptor from human placenta exhibits greater affinity for TGF-β2 than for TGF-β1

    International Nuclear Information System (INIS)

    Affinity-labeling techniques have been used to identify three types of high-affinity receptors for transforming growth factor β (TGF-β) on the surface of many cells in culture. Here the authors demonstrate that membrane preparations from tissue sources may also be used as an alternative system for studying the binding properties of TGF-β receptors. Using a chemical cross-linking technique with 125I-TGF-β1 and 125I-TGF-β2 and bis(sulfosuccinimidyl)suberate (BS3), they have identified and characterized two high-affinity binding components in membrane preparations derived from human term placenta. The larger species, which migrates as a diffuse band of molecular mass 250-350 kDa on sodium dodecyl sulfate-polyacrylamide electrophoresis gels, is characteristic of the TGF-β receptor type III, a proteoglycan containing glycosaminoglycan (GAG) chains of chondroitin and heparan sulfate. The smaller species of molecular mass 140 kDa was identified as the core glycoprotein of this type III receptor by using the techniques of enzymatic deglycosylation and peptide mapping. Competition experiments, using 125I-TGF-β1 or 125I-TGF-β2 and varying amounts of competing unlabeled TGF-β1 or TGF-β2, revealed that both the placental type III proteoglycan and its core glycoprotein belong to a novel class of type III receptors that exhibit a greater affinity for TGF-β2 than for TGF-β1. This preferential binding of TGF-β2 to placental type III receptors suggests differential roles for TGF-β2 and TGF-β 1 in placental function

  4. The Synthesis of 1,3,5-triazine Derivatives and JNJ7777120 Analogues with Histamine H4 Receptor Affinity and Their Interaction with PTEN Promoter.

    Science.gov (United States)

    Latacz, Gniewomir; Kechagioglou, Petros; Papi, Rigini; Łażewska, Dorota; Więcek, Małgorzata; Kamińska, Katarzyna; Wencel, Przemysław; Karcz, Tadeusz; Schwed, Johannes S; Stark, Holger; Kyriakidis, Dimitrios A; Kieć-Kononowicz, Katarzyna

    2016-08-01

    The involvement of histamine and H4 receptor (H4 R) in cancer has been investigated recently using the H4 R agonists and antagonists. The scope of the research project was synthesis and exploration of the consequences of a group of compounds with histamine H4 receptor (H4 R) affinity on the promoter of PTEN gene encoding the antitumor PTEN protein. The series of novel compounds based either on H4 R antagonists JNJ7777120 structure or 1,3,5-triazine scaffold were synthesized, evaluated for histamine H4 R affinity and used in this study. Compounds 5 and 7 belonging to the group of JNJ7777120 analogues showed the highest interaction with the promoter of PTEN gene and weak affinity against H4 R with Ki value >100 μm. These compounds showed no significant effect on neuroblastoma IMR-32 cells viability indicating no correlation between PTEN gene promoter affinity and antitumor activity. Compound 6, another JNJ7777120 analogue, showed the highest effect on IMR-32 viability with calculated IC50 = 23.27 μm. The 1,3,5-triazine derivatives exhibited generally low or medium interaction with PTEN gene promoter. However, the 1,3,5-triazine derivative 11 with the para-bromo substituent showed the highest affinity against H4 R with Ki value of 520 nm and may be considered as a new lead structure. PMID:26931395

  5. Chiral dimethylamine flutamide derivatives-modeling, synthesis, androgen receptor affinities and carbon-11 labeling

    International Nuclear Information System (INIS)

    Most prostate cancers are androgen dependent upon initial diagnosis. On the other hand, some very aggressive forms of prostate cancer were shown to have lost the expression of the androgen receptor (AR). Although the AR is routinely targeted in endocrine treatment, the clinical outcome remains suboptimal. Therefore, it is crucial to demonstrate the presence and activity of the AR in each case of prostate cancer, before and after treatment. While noninvasive positron emission tomography (PET) has the potential to determine AR expression of tumor cells in vivo, fully optimized PET imaging agents are not yet available. Based on molecular modeling, three novel derivatives of hydroxyflutamide (Compounds 1-3) were designed and synthesized. They contain an electron-rich group (dimethylamine) located on the methyl moiety, which may confer a better stability to the molecule in vivo. Compounds 1-3 have AR binding that is similar or higher than that of the currently used commercial drugs. An automated carbon-11 radiolabeling route was developed, and the compounds were successfully labeled with a 10-15% decay-corrected radiochemical yield, 99% radiochemical purity and a specific activity of 4Ci/μmol end of bombardment (n=15). These labeled biomarkers may facilitate the future quantitative molecular imaging of AR-positive prostate cancer using PET and may also allow for image-guided treatment of prostate cancer

  6. Glucocorticoid up-regulation of high-affinity interleukin 6 receptors on human epithelial cells

    International Nuclear Information System (INIS)

    Interleukin 6 (IL-6) is a potent pleiotropic cytokine, known, among others, to stimulate immunoglobulin production by B cells and to trigger acute-phase protein synthesis by hepatocytes. Similar to IL-1, it is produced by monocytes and macrophages following an inflammatory challenge. Analysis of IL-6 receptor (IL-6R) expression on different human cell lines indicates that dexamethasone could up-regulate the number of IL-6R on one epithelial cell line (UAC) and on two hepatoma cell lines (HepG2 and Hep3B). This effect was confirmed by Scatchard analysis of binding experiments, using [35S]methionine and [35S]cysteine metabolically labeled IL-6. It was confirmed at the level of mRNA expression by Northern blot analysis. These results provide evidence for a link between IL-6 and glucocorticoids. They could represent an example of a system in which one role of glucocorticoids is to define more accurately the target of cytokines, and they could explain, at least partly, the frequently observed synergy between IL-6 and glucocorticoids, notably in the case of hepatocytes

  7. Nicotine ameliorates NMDA receptor antagonist-induced deficits in contextual fear conditioning through high-affinity nicotinic acetylcholine receptors in the hippocampus.

    Science.gov (United States)

    André, Jessica M; Leach, Prescott T; Gould, Thomas J

    2011-03-01

    NMDA glutamate receptors (NMDARs) and nicotinic acetylcholine receptors (nAChRs) are both involved in learning and synaptic plasticity. Increasing evidence suggests processes mediated by these receptors may interact to modulate learning; however, little is known about the neural substrates involved in these interactive processes. The present studies investigated the effects of nicotine on MK-801 hydrogen maleate (MK-801) and DL-2-Amino-5-phosphonovaleric acid (APV)-induced disruption of contextual fear conditioning in male C57BL/6J mice, using direct drug infusion and selective nAChR antagonists to define the brain regions and the nAChR subtypes involved. Mice treated with MK-801 showed a deficit in contextual fear conditioning that was ameliorated by nicotine. Direct drug infusion demonstrated that the NMDAR antagonists disrupted hippocampal function and that nicotine acted in the dorsal hippocampus to ameliorate the deficit in learning. The high-affinity nAChR antagonist Dihydro-β-erythroidine hydrobromide (DhβE) blocked the effects of nicotine on MK-801-induced deficits while the α7 nAChR antagonist methyllycaconitine citrate salt hydrate (MLA) did not. These results suggest that NMDARs and nAChRs may mediate similar hippocampal processes involved in contextual fear conditioning. Furthermore, these results may have implications for developing effective therapeutics for the cognitive deficits associated with schizophrenia because a large subset of patients with schizophrenia exhibit cognitive deficits that may be related to NMDAR dysfunction and smoke at much higher rates than the healthy population, which may be an attempt to ameliorate cognitive deficits. PMID:21167848

  8. Human metabolites of synthetic cannabinoids JWH-018 and JWH-073 bind with high affinity and act as potent agonists at cannabinoid type-2 receptors

    Energy Technology Data Exchange (ETDEWEB)

    Rajasekaran, Maheswari; Brents, Lisa K.; Franks, Lirit N. [Department of Pharmacology and Toxicology, University of Arkansas for Medical Sciences, Little Rock, AR 72205 (United States); Moran, Jeffery H. [Department of Pharmacology and Toxicology, University of Arkansas for Medical Sciences, Little Rock, AR 72205 (United States); Arkansas Department of Public Health, Public Health Laboratory, Little Rock, AR 72205 (United States); Prather, Paul L., E-mail: pratherpaull@uams.edu [Department of Pharmacology and Toxicology, University of Arkansas for Medical Sciences, Little Rock, AR 72205 (United States)

    2013-06-01

    K2 or Spice is an emerging drug of abuse that contains synthetic cannabinoids, including JWH-018 and JWH-073. Recent reports indicate that monohydroxylated metabolites of JWH-018 and JWH-073 retain high affinity and activity at cannabinoid type-1 receptors (CB{sub 1}Rs), potentially contributing to the enhanced toxicity of K2 compared to marijuana. Since the parent compounds also bind to cannabinoid type-2 receptors (CB{sub 2}Rs), this study investigated the affinity and intrinsic activity of JWH-018, JWH-073 and several monohydroxylated metabolites at human CB{sub 2}Rs (hCB{sub 2}Rs). The affinity of cannabinoids for hCB{sub 2}Rs was determined by competition binding studies employing CHO-hCB{sub 2} membranes. Intrinsic activity of compounds was assessed by G-protein activation and adenylyl cyclase (AC)-inhibition in CHO-hCB{sub 2} cells. JWH-073, JWH-018 and several of their human metabolites exhibit nanomolar affinity and act as potent agonists at hCB{sub 2}Rs. Furthermore, a major omega hydroxyl metabolite of JWH-073 (JWH-073-M5) binds to CB{sub 2}Rs with 10-fold less affinity than the parent molecule, but unexpectedly, is equipotent in regulating AC-activity when compared to the parent molecule. Finally, when compared to CP-55,940 and Δ{sup 9}-tetrahydrocannabinol (Δ{sup 9}-THC), JWH-018, JWH-018-M5 and JWH-073-M5 require significantly less CB{sub 2}R occupancy to produce similar levels of AC-inhibition, indicating that these compounds may more efficiently couple CB{sub 2}Rs to AC than the well characterized cannabinoid agonists examined. These results indicate that JWH-018, JWH-073 and several major human metabolites of these compounds exhibit high affinity and demonstrate distinctive signaling properties at CB{sub 2}Rs. Therefore, future studies examining pharmacological and toxicological properties of synthetic cannabinoids present in K2 products should consider potential actions of these drugs at both CB{sub 1} and CB{sub 2}Rs. - Highlights: • JWH-018

  9. Human metabolites of synthetic cannabinoids JWH-018 and JWH-073 bind with high affinity and act as potent agonists at cannabinoid type-2 receptors

    International Nuclear Information System (INIS)

    K2 or Spice is an emerging drug of abuse that contains synthetic cannabinoids, including JWH-018 and JWH-073. Recent reports indicate that monohydroxylated metabolites of JWH-018 and JWH-073 retain high affinity and activity at cannabinoid type-1 receptors (CB1Rs), potentially contributing to the enhanced toxicity of K2 compared to marijuana. Since the parent compounds also bind to cannabinoid type-2 receptors (CB2Rs), this study investigated the affinity and intrinsic activity of JWH-018, JWH-073 and several monohydroxylated metabolites at human CB2Rs (hCB2Rs). The affinity of cannabinoids for hCB2Rs was determined by competition binding studies employing CHO-hCB2 membranes. Intrinsic activity of compounds was assessed by G-protein activation and adenylyl cyclase (AC)-inhibition in CHO-hCB2 cells. JWH-073, JWH-018 and several of their human metabolites exhibit nanomolar affinity and act as potent agonists at hCB2Rs. Furthermore, a major omega hydroxyl metabolite of JWH-073 (JWH-073-M5) binds to CB2Rs with 10-fold less affinity than the parent molecule, but unexpectedly, is equipotent in regulating AC-activity when compared to the parent molecule. Finally, when compared to CP-55,940 and Δ9-tetrahydrocannabinol (Δ9-THC), JWH-018, JWH-018-M5 and JWH-073-M5 require significantly less CB2R occupancy to produce similar levels of AC-inhibition, indicating that these compounds may more efficiently couple CB2Rs to AC than the well characterized cannabinoid agonists examined. These results indicate that JWH-018, JWH-073 and several major human metabolites of these compounds exhibit high affinity and demonstrate distinctive signaling properties at CB2Rs. Therefore, future studies examining pharmacological and toxicological properties of synthetic cannabinoids present in K2 products should consider potential actions of these drugs at both CB1 and CB2Rs. - Highlights: • JWH-018 and JWH-073 are synthetic cannabinoids present in abused K2 products. • JWH-018, JWH-073 and

  10. Nucleotide binding by the widespread high-affinity cyclic di-GMP receptor MshEN domain.

    Science.gov (United States)

    Wang, Yu-Chuan; Chin, Ko-Hsin; Tu, Zhi-Le; He, Jin; Jones, Christopher J; Sanchez, David Zamorano; Yildiz, Fitnat H; Galperin, Michael Y; Chou, Shan-Ho

    2016-01-01

    C-di-GMP is a bacterial second messenger regulating various cellular functions. Many bacteria contain c-di-GMP-metabolizing enzymes but lack known c-di-GMP receptors. Recently, two MshE-type ATPases associated with bacterial type II secretion system and type IV pilus formation were shown to specifically bind c-di-GMP. Here we report crystal structure of the MshE N-terminal domain (MshEN1-145) from Vibrio cholerae in complex with c-di-GMP at a 1.37 Å resolution. This structure reveals a unique c-di-GMP-binding mode, featuring a tandem array of two highly conserved binding motifs, each comprising a 24-residue sequence RLGxx(L/V/I)(L/V/I)xxG(L/V/I)(L/V/I)xxxxLxxxLxxQ that binds half of the c-di-GMP molecule, primarily through hydrophobic interactions. Mutating these highly conserved residues markedly reduces c-di-GMP binding and biofilm formation by V. cholerae. This c-di-GMP-binding motif is present in diverse bacterial proteins exhibiting binding affinities ranging from 0.5 μM to as low as 14 nM. The MshEN domain contains the longest nucleotide-binding motif reported to date. PMID:27578558

  11. EFFECTS OF INTEGRIN ALPHA ⅡbR995A MUTATION ON RECEPTOR AFFINITY AND pp 125 (FAK) PHOSPHORYLATION

    Institute of Scientific and Technical Information of China (English)

    Xue-yuan Tang; Zai-fu Jian; Guo-ping Wang; Hong-hui Yang; Wei Liu

    2004-01-01

    Objective To investigate the role of cytoplasmic domain of integrin alpha Ⅱb in platelet signal transduction.Methods Binding capacity of integrin alpha ⅡbR995Ato antibody platelet activation complex-1 (PAC-1) and pp125focal adhesion kinase (FAK) phosphorylation of cells were detected by flow cytometry, immune precipitation, and Western blotting.Results Without activation, wild-type alpha Ⅱ bbeta3 Chinese hamster ovary (CHO) cells failed to bind to PAC-1, but mutant chimera alpha ⅡbR995Aeta3 CHO cells were able to bind with PAC-1. Furthermore, phosphorylation of pp125 (FAK)in wild-type alpha Ⅱbbeta3 CHO cells occured only when cells were adhered to fibrinogen, but could not be detected in bovine serum albumin suspension. However in the mutant chimera group, it could be detected in both conditions.Conclusion The mutation in integrin alpha ⅡbR995Aalters its affinity state as a receptor, thus also mediating cytoplasmic signal transduction leading to the phosphorylation of pp125 (FAK) without ligand binding.

  12. The neonatal Fc receptor (FcRn) binds independently to both sites of the IgG homodimer with identical affinity

    Science.gov (United States)

    Abdiche, Yasmina Noubia; Yeung, Yik Andy; Chaparro-Riggers, Javier; Barman, Ishita; Strop, Pavel; Chin, Sherman Michael; Pham, Amber; Bolton, Gary; McDonough, Dan; Lindquist, Kevin; Pons, Jaume; Rajpal, Arvind

    2015-01-01

    The neonatal Fc receptor (FcRn) is expressed by cells of epithelial, endothelial and myeloid lineages and performs multiple roles in adaptive immunity. Characterizing the FcRn/IgG interaction is fundamental to designing therapeutic antibodies because IgGs with moderately increased binding affinities for FcRn exhibit superior serum half-lives and efficacy. It has been hypothesized that 2 FcRn molecules bind an IgG homodimer with disparate affinities, yet their affinity constants are inconsistent across the literature. Using surface plasmon resonance biosensor assays that eliminated confounding experimental artifacts, we present data supporting an alternate hypothesis: 2 FcRn molecules saturate an IgG homodimer with identical affinities at independent sites, consistent with the symmetrical arrangement of the FcRn/Fc complex observed in the crystal structure published by Burmeister et al. in 1994. We find that human FcRn binds human IgG1 with an equilibrium dissociation constant (KD) of 760 ± 60 nM (N = 14) at 25°C and pH 5.8, and shows less than 25% variation across the other human subtypes. Human IgG1 binds cynomolgus monkey FcRn with a 2-fold higher affinity than human FcRn, and binds both mouse and rat FcRn with a 10-fold higher affinity than human FcRn. FcRn/IgG interactions from multiple species show less than a 2-fold weaker affinity at 37°C than at 25°C and appear independent of an IgG's variable region. Our in vivo data in mouse and rat models demonstrate that both affinity and avidity influence an IgG's serum half-life, which should be considered when choosing animals, especially transgenic systems, as surrogates. PMID:25658443

  13. Radioiodinated ligands for the estrogen receptor: Effect of different 7-cyanoalkyl chains on the binding affinity of novel iodovinyl-6-dehydroestradiols

    International Nuclear Information System (INIS)

    Three novel 17α-ethynyl-Δ6,7-estra-3,17β-diols and their 17α-[125I]-iodovinyl derivatives, containing different C7-cyanoalkyl chains, were studied as potential radioligands for the estrogen receptor. The influence of the chain length on the biological behaviour of the compounds was assessed through in vitro ER binding assays of the ethynyl derivatives and breast cancer cell uptake studies of the 17α-[125I]-iodovinyl-Δ6,7-estra-3,17β-diols. A difference in alkyl chain induced a decrease in ER binding affinities of substances, however, the receptor-binding affinities (RBA) of all compounds were lower than that of estradiol itself. In addition, a non-specific cell binding was observed which is in accordance with the encountered ethynyl RBA values suggesting that the uptake is not ER mediated

  14. Radioiodinated ligands for the estrogen receptor: Effect of different 7-cyanoalkyl chains on the binding affinity of novel iodovinyl-6-dehydroestradiols

    Energy Technology Data Exchange (ETDEWEB)

    Neto, Carina [Instituto Tecnologico e Nuclear, Estrada Nacional 10, 2686-953 Sacavem (Portugal); Centro de Investigacao em Meio Ambiente Genetica e Oncobiologia (CIMAGO) (Portugal); Oliveira, Maria Cristina [Instituto Tecnologico e Nuclear, Estrada Nacional 10, 2686-953 Sacavem (Portugal); Centro de Investigacao em Meio Ambiente Genetica e Oncobiologia (CIMAGO) (Portugal)], E-mail: cmelo@itn.pt; Gano, Lurdes; Marques, Fernanda; Santos, Isabel [Instituto Tecnologico e Nuclear, Estrada Nacional 10, 2686-953 Sacavem (Portugal); Centro de Investigacao em Meio Ambiente Genetica e Oncobiologia (CIMAGO) (Portugal); Morais, Goreti Ribeiro [Faculty of Pharmacy, University of Lisbon, Lisbon (Portugal); Yasuda, Takumi [Interdisciplinary Graduate School of Engineering Sciences, Kyushu University, Fukuoka (Japan); Thiemann, Thies [Centro de Investigacao em Meio Ambiente Genetica e Oncobiologia (CIMAGO) (Portugal); Interdisciplinary Graduate School of Engineering Sciences, Kyushu University, Fukuoka (Japan); Botelho, Filomena [Centro de Investigacao em Meio Ambiente Genetica e Oncobiologia (CIMAGO) (Portugal); Instituto de Biofisica/Biomatematica, IBILI, FMUC, Coimbra (Portugal); Oliveira, Carlos F. [Centro de Investigacao em Meio Ambiente Genetica e Oncobiologia (CIMAGO) (Portugal); Clinica Ginecologica, FMUC, Coimbra (Portugal)

    2009-02-15

    Three novel 17{alpha}-ethynyl-{delta}{sup 6,7}-estra-3,17{beta}-diols and their 17{alpha}-[{sup 125}I]-iodovinyl derivatives, containing different C7-cyanoalkyl chains, were studied as potential radioligands for the estrogen receptor. The influence of the chain length on the biological behaviour of the compounds was assessed through in vitro ER binding assays of the ethynyl derivatives and breast cancer cell uptake studies of the 17{alpha}-[{sup 125}I]-iodovinyl-{delta}{sup 6,7}-estra-3,17{beta}-diols. A difference in alkyl chain induced a decrease in ER binding affinities of substances, however, the receptor-binding affinities (RBA) of all compounds were lower than that of estradiol itself. In addition, a non-specific cell binding was observed which is in accordance with the encountered ethynyl RBA values suggesting that the uptake is not ER mediated.

  15. The Length and Flexibility of the 2-Substituent of 9-Ethyladenine Derivatives Modulate Affinity and Selectivity for the Human A2A Adenosine Receptor.

    Science.gov (United States)

    Thomas, Ajiroghene; Buccioni, Michela; Dal Ben, Diego; Lambertucci, Catia; Marucci, Gabriella; Santinelli, Claudia; Spinaci, Andrea; Kachler, Sonja; Klotz, Karl-Norbert; Volpini, Rosaria

    2016-08-19

    The A2A adenosine receptor (A2A AR) is a key target for the development of pharmacological tools for the treatment of central nervous system disorders. Previous works have demonstrated that the insertion of substituents at various positions on adenine leads to A2A AR antagonists with affinity in the micromolar to nanomolar range. In this work, a series of 9-ethyladenine derivatives bearing phenylalkylamino, phenylakyloxy or phenylakylthio groups of different lengths at the 2-position were synthesised and tested against the human adenosine receptors. The derivatives showed sub-micromolar affinity for these membrane proteins. The further introduction of a bromine atom at the 8-position has the effect of improving the affinity and selectivity for all ARs and led to compounds that are able bind to the A2A AR subtype at low nanomolar levels. Functional studies confirmed that the new adenine derivatives behave as A2A AR antagonists with half-maximal inhibitory concentration values in the nanomolar range. Molecular modelling studies provide a description of the possible binding mode of these compounds at the A2A AR and an interpretation of the affinity data at this AR subtype. PMID:27037522

  16. Radioimmunoassay of total IgE and allergen-specific IgE antibodies with a uniform indicator system in allergies of childhood

    International Nuclear Information System (INIS)

    Solid-phase radioimmunoassays for the determination of allergen-specific and total IgE have been developed. In an indirect solid-phase radioimmunoassay for the measurement of allergen-specific antibodies PVC blisters coated with allergens and in a sandwich solid-phase radioimmunoassay blisters coated with antihuman IgE antibodies are incubated sequentially with patient serum, unlabelled antihuman IgE from rabbits purified by affinity chromatography, and finally with antirabbitglobulin from sheep. Antirabbitglobuline was purified by immunoadsorption. The 125I-labelled antibody with a specific activity of 30 kBq/μg antibody protein could be used universally for the determination of antibodies of each immunoglobulin class. In 160 patients mostly with seasonal asthma these assays supported RAST and PRIST kits and were helpful in the diagnosis of atopic diseases. (author)

  17. Measurement of density and affinity for dopamine D2 receptors by a single positron emission tomography scan with multiple injections of [11C]raclopride

    OpenAIRE

    Ikoma, Yoko; Watabe, Hiroshi; Hayashi, Takuya; Miyake, Yoshinori; Teramoto, Noboru; Minato, Kotaro; Iida, Hidehiro

    2009-01-01

    Positron emission tomography (PET) with [11C]raclopride has been used to investigate the density (Bmax) and affinity (Kd) of dopamine D2 receptors related to several neurological and psychiatric disorders. However, in assessing the Bmax and Kd, multiple PET scans are necessary under variable specific activities of administered [11C]raclopride, resulting in a long study period and unexpected physiological variations. In this paper, we have developed a method of multiple-injection graphical ana...

  18. Certain photooxidized derivatives of tryptophan bind with very high affinity to the Ah receptor and are likely to be endogenous signal substances

    International Nuclear Information System (INIS)

    The purpose of the present study was to determine whether ultraviolet light (UV) irradiation of amino acids produces compounds with affinity for the Ah receptor. Aqueous solutions of L-tryptophan were exposed to radiation from an unfiltered high-pressure mercury lamp. The photoproducts formed were solvent-extracted or concentrated on Sep-Pak C18 cartridges. The concentrated extracts or eluants were treated for their ability to compete with 3H-labeled 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD). Binding was assayed in liver cytosolic preparations from Sprague-Dawley rats using a technique based on hydroxylapatite separation. Photoproducts with receptor affinity were formed in a time-dependent manner. Histidine and tryptamine also gave products upon UV irradiation that competed with TCDD. Commercial tryptophan, at least aged, contained trace amounts of impurities with receptor affinity. Analysis by TLC and high-pressure liquid chromatography of the photo-products of tryptophan showed a minimum of three different binding compounds. Two of the products were studied in greater detail. One of them, showing UV absorbance and yellow fluorescence, gave a molecular ion (M+) of 284 and the other gave M+ 312 but showed little UV absorption and fluorescence. The concentration, based on mass spectrometry quantifications, of the two compounds that displaced more than 50% of TCDD was found to be extremely low, giving Kd values of 0.44 nM (M+ 312) and 0.07 nM (M+ 284). The existence of high affinity receptors for oxidized amino acids is postulated and their possible role in the proliferative cellular responses to TCDD and tryptophan is discussed briefly

  19. Analysis of D2 dopamine receptor occupancy with quantitative SPET using the high-affinity ligand [123I]epidepride: resolving conflicting findings.

    Science.gov (United States)

    Erlandsson, Kjell; Bressan, Rodrigo A; Mulligan, Rachel S; Ell, Peter J; Cunningham, Vincent J; Pilowsky, Lyn S

    2003-07-01

    Recent studies of limbic cortical dopamine D(2) receptor occupancy by clozapine using high-affinity PET and SPET radioligands have produced conflicting findings. It has been suggested that these divergent findings are due to between-study differences in the method used to estimate D(2) receptor-binding potential. We compared different methods for estimating striatal and temporal cortical D(2) receptor occupancy with high-affinity tracers. In vivo experimental SPET data, obtained with [(123)I]epidepride were analysed with reference tissue kinetic modeling and with the ratio method, applied to data corresponding to short (60 min) and long (240 min) acquisition times. Dopamine D(2) receptor occupancy by the atypical antipsychotic drug risperidone was evaluated. Simulation experiments were also performed, comparing occupancy values obtained for different receptor densities in relation to different data acquisition times. The simulation results revealed that previously published data regarding errors in occupancy estimation by analysis of time activity data acquired for 60 min cannot be extrapolated to studies performed over 240 min. The ratio method provided accurate temporal cortical D(2) receptor occupancy values when applied to data from a late time period, but underestimated the occupancy with earlier data. In striatum, both the late data ratio method and reference tissue kinetic modeling using all data underestimated D(2) receptor occupancy. However, more accurate analyses of striatal D(2) occupancy still showed selective limbic/cortical occupancy by risperidone. Our results substantiate the previous [(123)I]epidepride findings of high temporal cortical occupancy by other atypical antipsychotic drugs and suggest that a potential source of conflicting findings might be short scanning times imposed by [(11)C]FLB 457, leading to underestimation of temporal cortical D(2) receptor occupancy by this method. PMID:12880845

  20. Clobazam and its active metabolite N-desmethylclobazam display significantly greater affinities for α₂- versus α₁-GABA(A-receptor complexes.

    Directory of Open Access Journals (Sweden)

    Henrik Sindal Jensen

    Full Text Available Clobazam (CLB, a 1,5-benzodiazepine (BZD, was FDA-approved in October 2011 for the adjunctive treatment of seizures associated with Lennox-Gastaut syndrome (LGS in patients 2 years and older. BZDs exert various CNS effects through allosteric modulation of GABAA receptors. The structurally distinct, 1,4-BZD clonazepam (CLN is also approved to treat LGS. The precise mechanisms of action and clinical efficacy of both are unknown. Data show that the GABAA α₁-subunit-selective compound zolpidem [ZOL] exhibits hypnotic/sedative effects. Conversely, data from knock-in mice carrying BZD binding site mutations suggest that the α₂ subunit mediates anticonvulsant effects, without sedative actions. Hence, the specific pattern of interactions across the GABAA receptor complexes of BZDs might be reflected in their clinical efficacies and adverse effect profiles. In this study, GABAA-receptor binding affinities of CLB, N-desmethylclobazam (N-CLB, the major metabolite of CLB, CLN, and ZOL were characterized with native receptors from rat-brain homogenates and on cloned receptors from HEK293 cells transfected with combinations of α (α₁, α₂, α₃, or α₅, β₂, and γ₂ subtypes. Our results demonstrate that CLB and N-CLB have significantly greater binding affinities for α₂- vs. α₁-receptor complexes, a difference not observed for CLN, for which no distinction between α₂ and α₁ receptors was observed. Our experiments with ZOL confirmed the high preference for α₁ receptors. These results provide potential clues to a new understanding of the pharmacologic modes of action of CLB and N-CLB.

  1. Insecticidal 3-benzamido-N-phenylbenzamides specifically bind with high affinity to a novel allosteric site in housefly GABA receptors.

    Science.gov (United States)

    Ozoe, Yoshihisa; Kita, Tomo; Ozoe, Fumiyo; Nakao, Toshifumi; Sato, Kazuyuki; Hirase, Kangetsu

    2013-11-01

    γ-Aminobutyric acid (GABA) receptors (GABARs) are an important target for existing insecticides such as fiproles. These insecticides act as noncompetitive antagonists (channel blockers) for insect GABARs by binding to a site within the intrinsic channel of the GABAR. Recently, a novel class of insecticides, 3-benzamido-N-phenylbenzamides (BPBs), was shown to inhibit GABARs by binding to a site distinct from the site for fiproles. We examined the binding site of BPBs in the adult housefly by means of radioligand-binding and electrophysiological experiments. 3-Benzamido-N-(2,6-dimethyl-4-perfluoroisopropylphenyl)-2-fluorobenzamide (BPB 1) (the N-demethyl BPB) was a partial, but potent, inhibitor of [(3)H]4'-ethynyl-4-n-propylbicycloorthobenzoate (GABA channel blocker) binding to housefly head membranes, whereas the 3-(N-methyl)benzamido congener (the N-methyl BPB) had low or little activity. A total of 15 BPB analogs were tested for their abilities to inhibit [(3)H]BPB 1 binding to the head membranes. The N-demethyl analogs, known to be highly effective insecticides, potently inhibited the [(3)H]BPB 1 binding, but the N-methyl analogs did not even though they, too, are considered highly effective. [(3)H]BPB 1 equally bound to the head membranes from wild-type and dieldrin-resistant (rdl mutant) houseflies. GABA allosterically inhibited [(3)H]BPB 1 binding. By contrast, channel blocker-type antagonists enhanced [(3)H]BPB 1 binding to housefly head membranes by increasing the affinity of BPB 1. Antiparasitic macrolides, such as ivermectin B1a, were potent inhibitors of [(3)H]BPB 1 binding. BPB 1 inhibited GABA-induced currents in housefly GABARs expressed in Xenopus oocytes, whereas it failed to inhibit l-glutamate-induced currents in inhibitory l-glutamate receptors. Overall, these findings indicate that BPBs act at a novel allosteric site that is different from the site for channel blocker-type antagonists and that is probably overlapped with the site for macrolides

  2. Potentials and pitfalls using high affinity radioligands in PET and SPET determinations on regional drug induced D2 receptor occupancy--a simulation study based on experimental data.

    Science.gov (United States)

    Olsson, H; Farde, L

    2001-10-01

    The D2 dopamine receptor density ranges from 0.2 to 40 nM among human brain regions. For high density regions radioligands like [(11)C]raclopride provide accurate and reliable estimates of the receptor density. In research on neuropsychiatric disorders there is, however, a growing need for quantitative approaches that accurately measure D2 dopamine receptor occupancy induced by drugs or endogenous dopamine in regions with low receptor density. The new high affinity radioligands [(11)C]FLB 457 and [(123)I]epidepride have been shown to provide a signal for extrasriatal D2 dopamine receptor populations in the human brain in vivo. Initial observations indicate, however, that the time required to reach equilibrium is dependent on receptor density. Ratio analyses may thus not be readily used for comparisons among different brain regions. The aim of the present simulation study was to examine commonly used approaches for calculation of drug induced D2 dopamine receptor occupancy among regions with widely different receptor density. The input functions and the rate constants of [(11)C]FLB 457 and the reference ligand [(11)C]raclopride were first used in a simulation estimating the effect of receptor density on equilibrium time. In a second step we examined how errors produced by inaccurate determination of the binding potential parameter propagate to calculations of drug induced receptor occupancy. The simulations showed a marked effect of receptor density on equilibrium time for [(11)C]FLB 457, but not for [(11)C]raclopride. For [(11)C]FLB 457, a receptor density above about 7 nM caused the time of equilibrium to fall beyond time of data acquisition (1 h). The use of preequilibrium data caused the peak equilibrium and the end time ratio approaches but not the simplified reference tissue model (SRTM) approach to underestimate the binding potential and thus also the drug occupancy calculated for high-density regions. The study supports the use of ratio and SRTM analyses in

  3. A high-affinity soluble folate receptor in fluids of non-neoplastic ovarian cysts. Radioligand binding, molecular size, hydrophobic residue, and immunological properties

    International Nuclear Information System (INIS)

    The presence of a soluble folate receptor in fluids of non-neoplastic ovarian cysts was demonstrated. Radioligand binding exhibited characteristics typical of high-affinity folate-binding proteins. These included positive cooperativity, a tendency to increased binding affinity with decreasing receptor concentration, a slow ligand dissociation at pH 7.4 and inhibition by folate analogues. The folate receptor was probably synthesized in the lining epithelial cells of the cysts which showed positive immunostaining with antibodies against human milk folate-binding protein. The gel filtration profile of cystic fluid contained two radioligand-bound peaks, 25 and 100 kDa, whereas a single band of 70 kDa was seen on SDS-PAGE immunoblotting. Treatment with the enzyme phosphatidyl-inositol-specific phospholipase C resulted in a partial conversion of the 100 kDa peak to the 25 kDa peak. This suggests that insertion of a hydrophobic glycosyl-phosphatidyl-inositol tail into Triton X-100 micelles could give rise to large molecular size forms of the receptor on gel filtration. (au) 30 refs

  4. Synthesis of an artificial cell surface receptor that enables oligohistidine affinity tags to function as metal-dependent cell-penetrating peptides.

    Science.gov (United States)

    Boonyarattanakalin, Siwarutt; Athavankar, Sonalee; Sun, Qi; Peterson, Blake R

    2006-01-18

    Cell-penetrating peptides and proteins (CPPs) are important tools for the delivery of impermeable molecules into living mammalian cells. To enable these cells to internalize proteins fused to common oligohistidine affinity tags, we synthesized an artificial cell surface receptor comprising an N-alkyl derivative of 3beta-cholesterylamine linked to the metal chelator nitrilotriacetic acid (NTA). This synthetic receptor inserts into cellular plasma membranes, projects NTA headgroups from the cell surface, and rapidly cycles between the plasma membrane and intracellular endosomes. Jurkat lymphocytes treated with the synthetic receptor (10 microM) for 1 h displayed approximately 8,400,000 [corrected]NTA groups on the cell surface. Subsequent addition of the green fluorescent protein AcGFP fused to hexahistidine or decahistidine peptides (3 microM) and Ni(OAc)(2) (100 microM) enhanced the endocytosis of AcGFP by 150-fold (hexahistidine fusion protein) or 600-fold (decahistidine fusion protein) within 4 h at 37 degrees C. No adverse effects on cellular proliferation or morphology were observed under these conditions. By enabling common oligohistidine affinity tags to function as cell-penetrating peptides, this metal-chelating cell surface receptor provides a useful tool for studies of cellular biology [corrected] PMID:16402806

  5. Fluoxetine, a selective inhibitor of serotonin uptake, potentiates morphine analgesia without altering its discriminative stimulus properties or affinity for opioid receptors

    International Nuclear Information System (INIS)

    The analgesic effect of morphine in the rat tail jerk assay was enhanced by the serotonin uptake inhibitor, fluoxetine. Tail jerk latency was not affected by fluoxetine alone. Morphine's affinity for opioid receptors labeled in vitro with 3H-naloxone or 3H-D-Ala2-D-Leu5-enkephalin was not altered by fluoxetine, which has no affinity for these sites at concentrations as high as 1000 nM. In rats trained to discriminate morphine from saline, fluoxetine at doses of 5 or 10 mg/kg were recognized as saline. Increasing the fluoxetine dose to 20 mg/kg did not result in generalization to either saline or morphine. The dose response curve for morphine generalization was not significantly altered by fluoxetine doses of 5 or 10 mg/kg. Those rats treated with the combination of morphine and 20 mg/kg of fluoxetine did not exhibit saline or morphine appropriate responding. Fluoxetine potentiates the analgesic properties of morphine without enhancing its affinity for opioid receptors or its discriminative stimulus properties. 30 references, 2 figures, 2 tables

  6. Fluoxetine, a selective inhibitor of serotonin uptake, potentiates morphine analgesia without altering its discriminative stimulus properties or affinity for opioid receptors

    Energy Technology Data Exchange (ETDEWEB)

    Hynes, M.D.; Lochner, M.A.; Bemis, K.G.; Hymson, D.L.

    1985-06-17

    The analgesic effect of morphine in the rat tail jerk assay was enhanced by the serotonin uptake inhibitor, fluoxetine. Tail jerk latency was not affected by fluoxetine alone. Morphine's affinity for opioid receptors labeled in vitro with /sup 3/H-naloxone or /sup 3/H-D-Ala/sup 2/-D-Leu/sup 5/-enkephalin was not altered by fluoxetine, which has no affinity for these sites at concentrations as high as 1000 nM. In rats trained to discriminate morphine from saline, fluoxetine at doses of 5 or 10 mg/kg were recognized as saline. Increasing the fluoxetine dose to 20 mg/kg did not result in generalization to either saline or morphine. The dose response curve for morphine generalization was not significantly altered by fluoxetine doses of 5 or 10 mg/kg. Those rats treated with the combination of morphine and 20 mg/kg of fluoxetine did not exhibit saline or morphine appropriate responding. Fluoxetine potentiates the analgesic properties of morphine without enhancing its affinity for opioid receptors or its discriminative stimulus properties. 30 references, 2 figures, 2 tables.

  7. Synthesis, structure and affinity of novel 3-alkoxy-1,2-dihydro-3H-1,4-benzodiazepin-2-ones for CNS central and peripheral benzodiazepine receptors.

    Science.gov (United States)

    Andronati, Sergey; Semenishyna, Ekaterina; Pavlovsky, Victor; Simonov, Yuriy; Makan, Svetlana; Boyko, Irina; Burenkova, Natalya; Gdaniec, Maria; Cardinael, Pascal; Bouillon, Jean-Philippe; Mazepa, Alexander

    2010-04-01

    A series of novel 3-alkoxy-1,2-dihydro-3H-1,4-benzodiazepin-2-ones (7-15) was synthesized and their in vitro affinity for both the central benzodiazepine receptor (CBR) and the peripheral benzodiazepine receptor (PBR) of rat brain was studied. Racemic mixture of 7-bromo-3-(2-methoxy)ethoxy-5-phenyl-1,2-dihydro-3H-1,4-benzodiazepin-2-one (13) was separated into enantiomers 14, 15 by chiral HPLC. Absolute configuration of R-enantiomer 15 was determined by the method of X-ray diffraction analysis. The affinity of S-enantiomer 14 for CBR ( IC50)=245 nM) is 20-fold higher than the affinity of R-enantiomer 15 (IC50)=4,930 nM). A high selectivity for CBR versus PBR (IC50) (PBR)>10,000 nM) was shown by all reported compounds. Compound 12 was revealed as a potent (IC50)=9 nM) and selective CBR ligand among the synthesized 3-alkoxy-1,2-dihydro-3H-1,4-benzodiazepin-2-ones. PMID:20061068

  8. Development of rast assay for determination of anti-popolus canadensis IgE antibodies

    International Nuclear Information System (INIS)

    One of the frequent causes of pollen allergy in our region (Serbia, Yugoslavia) is the pollen of poplar (Populus canadensis). The aim of this study was to form RAST for the determination of specific anti-Populus canadensis IgE antibodies. Affinity purified and radiolabelled (I-125) MoAb E1 was used for forming assay for the determination of specific IgE. By titration of extract of poplar Populus canadensis we determined that the quantity of 0.65 mL extract is needed for coupling of Ig BrCN activated paper discs. Coupling was performed in Na2CO3/NaHCO3 buffer pH 10, on 4 deg C for 48h. Using this newly formed RAST, specific for Populus canadensis, we have determinated anti-Populus canadensis IgE antibodies as well as cross reactivity between pollens of Populus canadensis and Populus deltoides. (author). 12 refs., 3 figs., 1 tab

  9. IgE ELISA using antisera derived from epsilon chain antigenic peptides detects allergen-specific IgE in allergic horses.

    Science.gov (United States)

    Kalina, Warren V; Pettigrew, Howard D; Gershwin, Laurel J

    2003-05-12

    Equine disease with an allergic etiology is common. Environmental antigens most often implicated as allergens in horses include molds, dusty hay, grass pollen, hay dust mites, and insect saliva. Although intradermal testing with allergen is a useful diagnostic tool for some species, skin testing frequently produces false positive results in horses. Allergen deprivation as a diagnostic tool is often impossible and at best it is ineffective at diagnosing the specific allergic reactivity. Synthesis of IgE after exposure to allergen is the instigator of the allergic process. While IgE exerts its effect after binding strongly to mast cell Fc receptors, the presence of free IgE in the serum can be used to quantify and determine the allergen specificity of the allergic disease. A lack of widely available reagents for detection of equine IgE has limited this approach in horses. We have used the nucleotide sequence of equine IgE to prepare a peptide-based immunogen to elicit equine epsilon chain-specific antisera. Selection of peptides was based on antigenic attributes of the deduced amino acid sequence of the equine epsilon chain. Six peptides were selected for conjugation to carrier molecules and rabbit immunization. Of these, one peptide elicited antisera that was successfully used in enzyme linked immunosorbant assay (ELISA) to screen horse serum from 64 allergic horses for allergen-specific IgE. Twenty-four of the 64 horses showed positive reactivity to one or more of the following allergens: grass, grain mill dust, mosquito, and horsefly. This study demonstrates the usefulness of peptide-based immunogens for development of antisera to rare or difficult to purify antigens such as IgE. Resultant antisera has great usefulness in diagnostic assays for equine allergy and as a research tool. PMID:12730014

  10. Development of new peptide-based receptor of fluorescent probe with femtomolar affinity for Cu(+) and detection of Cu(+) in Golgi apparatus.

    Science.gov (United States)

    Jung, Kwan Ho; Oh, Eun-Taex; Park, Heon Joo; Lee, Keun-Hyeung

    2016-11-15

    Developing fluorescent probes for monitoring intracellular Cu(+) is important for human health and disease, whereas a few types of their receptors showing a limited range of binding affinities for Cu(+) have been reported. In the present study, we first report a novel peptide receptor of a fluorescent probe for the detection of Cu(+). Dansyl-labeled tripeptide probe (Dns-LLC) formed a 1:1 complex with Cu(+) and showed a turn-on fluorescent response to Cu(+) in aqueous buffered solutions. The dissociation constant of Dns-LLC for Cu(+) was determined to be 12 fM, showing that Dns-LLC had more potent binding affinity for Cu(+) than those of previously reported chemical probes for Cu(+). The binding mode study showed that the thiol group of the peptide receptor plays a critical role in potent binding with Cu(+) and the sulfonamide and amide groups of the probe might cooperate to form a complex with Cu(+). Dns-LLC detected Cu(+) selectively by a turn-on response among various biologically relevant metal ions, including Cu(2+) and Zn(2+). The selectivity of the peptide-based probe for Cu(+) was strongly dependent on the position of the cysteine residue in the peptide receptor part. The fluorescent peptide-based probe penetrated the living RKO cells and successfully detected Cu(+) in the Golgi apparatus in live cells by a turn-on response. Given the growing interest in imaging Cu(+) in live cells, a novel peptide receptor of Cu(+) will offer the potential for developing a variety of fluorescent probes for Cu(+) in the field of copper biochemistry. PMID:27208475

  11. Crystal structure of an affinity-matured prolactin complexed to its dimerized receptor reveals the topology of hormone binding site 2

    DEFF Research Database (Denmark)

    Broutin, Isabelle; Jomain, Jean-Baptiste; Tallet, Estelle; Van Agthoven, Jan; Raynal, Bertand; Hoos, Sylviane; Kragelund, Birthe Brandt; Kelly, Paul; Ducroix, Alexandre; England, Patrick; Goffin, Vincent

    2010-01-01

    We report the first crystal structure of a 1:2 hormone.receptor complex that involves prolactin (PRL) as the ligand, at 3.8-A resolution. Stable ternary complexes were obtained by generating affinity-matured PRL variants harboring an N-terminal tail from ovine placental lactogen, a closely related...... components of PRL site 2 ("three-pin plug"): the conserved glycine 129 of helix alpha3, the hydrogen bond network involving surrounding residues (glycine cavity), and the N terminus. The model provides a molecular basis for the properties of the different PRL analogs designed to date, including PRLR...

  12. Anti-ulcer drugs promote IgE formation toward dietary antigens in adult patients.

    Science.gov (United States)

    Untersmayr, Eva; Bakos, Noémi; Schöll, Isabella; Kundi, Michael; Roth-Walter, Franziska; Szalai, Krisztina; Riemer, Angelika B; Ankersmit, Hendrik J; Scheiner, Otto; Boltz-Nitulescu, George; Jensen-Jarolim, Erika

    2005-04-01

    Recently, we have demonstrated that anti-ulcer drugs, such as H2-receptor blockers and proton pump inhibitors, promote the development of immediate type food allergy toward digestion-labile proteins in mice. The aim of this study was to examine the allergological relevance of these findings in humans. In an observational cohort study, we screened 152 adult patients from a gastroenterological outpatient clinic with negative case histories for atopy or allergy, who were medicated with H2-receptor blockers or proton pump inhibitors for 3 months. IgE reactivities to food allergens before and after 3 months of anti-acid treatment were compared serologically. Ten percent of the patients showed a boost of preexisting IgE antibodies and 15% de novo IgE formation toward numerous digestion-labile dietary compounds, like milk, potato, celery, carrots, apple, orange, wheat, and rye flour. Thus, the relative risk to develop food-specific IgE after anti-acid therapy was 10.5 (95% confidence interval: 1.44-76.48). The long-term effect was evaluated 5 months after therapy. Food-specific IgE could still be measured in 6% of the patients, as well as significantly elevated serum concentrations of ST2, a Th2-specific marker. An unspecific boost during the pollen season could be excluded, as 50 untreated control patients revealed no changes in their IgE pattern. In line with our previous animal experiments, our data strongly suggest that anti-ulcer treatment primes the development of IgE toward dietary compounds in long-term acid-suppressed patients. PMID:15671152

  13. P2X7 receptor activation downmodulates Na(+)-dependent high-affinity GABA and glutamate transport into rat brain cortex synaptosomes.

    Science.gov (United States)

    Barros-Barbosa, A R; Lobo, M G; Ferreirinha, F; Correia-de-Sá, P; Cordeiro, J M

    2015-10-15

    Sodium-dependent high-affinity amino-acid transporters play crucial roles in terminating synaptic transmission in the central nervous system (CNS). However, there is lack of information about the mechanisms underlying the regulation of amino-acid transport by fast-acting neuromodulators, like ATP. Here, we investigated whether activation of the ATP-sensitive P2X7 receptor modulates Na(+)-dependent high-affinity γ-aminobutyric acid (GABA) and glutamate uptake into nerve terminals (synaptosomes) of the rat cerebral cortex. Radiolabeled neurotransmitter accumulation was evaluated by liquid scintillation spectrometry. The cell-permeant sodium-selective fluorescent indicator, SBFI-AM, was used to estimate Na(+) influx across plasma membrane. 2'(3')-O-(4-benzoylbenzoyl)ATP (BzATP, 3-300 μM), a prototypic P2X7 receptor agonist, concentration-dependently decreased [(3)H]GABA (14%) and [(14)C]glutamate (24%) uptake; BzATP decreased transport maximum velocity (Vmax) without affecting the Michaelis constant (Km) values. The selective P2X7 receptor antagonist, A-438079 (3 μM), prevented inhibition of [(3)H]GABA and [(14)C]glutamate uptake by BzATP (100 μM). The inhibitory effect of BzATP coincided with its ability to increase intracellular Na(+) and was mimicked by Na(+) ionophores, like gramicidin and monensin. Increases in intracellular Na(+) (with veratridine or ouabain) or substitution of extracellular Na(+) by N-methyl-D-glucamine (NMDG)(+) all decreased [(3)H]GABA and [(14)C]glutamate uptake and attenuated BzATP effects. Uptake inhibition by BzATP (100 μM) was also attenuated by calmidazolium, which selectively inhibits Na(+) currents through the P2X7 receptor pore. In conclusion, disruption of the Na(+) gradient by P2X7 receptor activation downmodulates high-affinity GABA and glutamate uptake into rat cortical synaptosomes. Interference with amino-acid transport efficacy may constitute a novel target for therapeutic management of cortical excitability. PMID

  14. High Affinity Binding of the Receptor-associated Protein D1D2 Domains with the Low Density Lipoprotein Receptor-related Protein (LRP1) Involves Bivalent Complex Formation: CRITICAL ROLES OF LYSINES 60 AND 191.

    Science.gov (United States)

    Prasad, Joni M; Young, Patricia A; Strickland, Dudley K

    2016-08-26

    The LDL receptor-related protein 1 (LRP1) is a large endocytic receptor that binds and mediates the endocytosis of numerous structurally diverse ligands. Currently, the basis for ligand recognition by LRP1 is not well understood. LRP1 requires a molecular chaperone, termed the receptor-associated protein (RAP), to escort the newly synthesized receptor from the endoplasmic reticulum to the Golgi. RAP is a three-domain protein that contains the following two high affinity binding sites for LRP1: one is located within domains 1 and 2, and one is located in its third domain. Studies on the interaction of the RAP third domain with LRP1 reveal critical contributions by lysine 256 and lysine 270 for this interaction. From these studies, a model for ligand recognition by this class of receptors has been proposed. Here, we employed surface plasmon resonance to investigate the binding of RAP D1D2 to LRP1. Our results reveal that the high affinity of D1D2 for LRP1 results from avidity effects mediated by the simultaneous interactions of lysine 60 in D1 and lysine 191 in D2 with sites on LRP1 to form a bivalent D1D2-LRP1 complex. When lysine 60 and 191 are both mutated to alanine, the binding of D1D2 to LRP1 is ablated. Our data also reveal that D1D2 is able to bind to a second distinct site on LRP1 to form a monovalent complex. The studies confirm the canonical model for ligand recognition by this class of receptors, which is initiated by pairs of lysine residues that dock into acidic pockets on the receptor. PMID:27402839

  15. Premature aging phenotype in mice lacking high affinity nicotinic receptors: region specific changes in layer V pyramidal cell morphology

    Directory of Open Access Journals (Sweden)

    Eleni Konsolaki

    2014-02-01

    accelerated cognitive aging, based on structural alterations and spatial learning deficits only evident in old animals (Zoli et al., 1999; Picciotto and Zoli, 2002. However a systematic comparison of neuronal microanatomy in adult and aged animals has not been done to date. In the present study adult (4-6months and old (22-24months WT and β2-/- animals were used to examine the respective contributions of age and genotype on neuronal structure. We focus on layer V pyramidal cells because: (i they constitute the main cortical output (DeFelipe and Farinas, 1992; Romand et al., 2011 (ii they are often reported to exhibit increased sensitivity to aging (Nakamura et al., 1985; Baskys et al., 1990; De Brabander et al., 1998; Turner et al., 2005; (iii they possess a high density of cholinergic terminals (Houser et al., 1985 and, in contrast to layer III cells, they exhibit strong presynaptic modulation by β2 containing nAChRs and are activated by nAChR stimulation (Poorthuis et al., 2013; hence they would be a sensitive readout for the lack of high affinity nicotinic receptors. Furthermore, to examine the degree of age-related vulnerability across distinct cortical areas we used YFP-H mice that express yellow fluorescent protein (YFP in specific populations of thick-tufted layer V pyramidal neurons across the cortical mantle (Feng et al., 2000; Sugino et al., 2006. We used mutants crossed with YFP+ mice in order to have the same labeled populations in both genotypes, and we examined cells in primary visual cortex (V1 and anterior cingulate cortex (ACC, two cortical regions that receive similar cholinergic inputs (McKinney et al., 1983; Jacobowitz and Creed, 1983; Everitt and Robbins, 1997; Laplante et al., 2005 but have distinct cytoarchitecture and functional role (Elston et al., 2005. We ask whether neurons in old β2-/- mice exhibit greater structural deficits than aged-matched controls and whether deficits appear in old age or are already present earlier. Brains from 21 adult

  16. Iodination of [Tyr11]somatostatin yields a super high affinity ligand for somatostatin receptors in GH4C1 pituitary cells

    International Nuclear Information System (INIS)

    GH4C1 cells are a clonal strain of rat pituitary tumor cells which contain high affinity receptors for the inhibitory neuropeptide somatostatin (SRIF). In contrast to other peptides that bind to specific receptors on these cells, receptor-bound [125I-Tyr1]SRIF does not undergo rapid endocytosis. Rather, partial degradation to 125I-tyrosine occurs concomitantly with the dissociation of [125I-Tyr1]SRIF from cell surface receptors. In this study we characterize the binding, biological activity and receptor-mediated degradation of [125I-Tyr11]SRIF, a SRIF analog that is radiolabeled in the center of the molecule. The binding of trace concentrations of [125I-Tyr11]SRIF (less than 50 pM) required 6 hr to reach equilibrium at 37 degrees compared with the 60 min required for [125I-Tyr1]SRIF. Analysis of the kinetics of [125I- Tyr11]SRIF binding showed that the rate constant for association (kon = 1.7 x 10(8) M-8min-1) was similar to that for [125I-Tyr1]SRIF (0.8 x 10(8) M-1min-1). However, the two radioligands exhibited markedly different dissociation kinetics; the koff for [125I-Tyr11]SRIF was 0.002 min-1 compared with the value of 0.02 min-1 for [125I-Tyr1] SRIF. In agreement with its much slower rate of dissociation, [125I-Tyr11]SRIF bound to the SRIF receptor with higher affinity (Kd = 70 pM) than did [125I-Tyr1]SRIF (Kd = 350 pM). However, the apparent ED50 for [I-Tyr11]SRIF to inhibit cAMP accumulation (1.9 +/- 0.4 nM) was greater than the ED50 for SRIF (0.19 +/- 0.04 nM). The low potency of [I-Tyr11]SRIF probably resulted from the fact that subsaturating concentrations of this peptide did not achieve equilibrium binding during the 30-min incubation used to assay biological activity. As previously reported for [125I-Tyr1]SRIF, receptor-bound [125I-Tyr11]SRIF was not internalized and was released from the cells as a mixture of intact [125I-Tyr11]SRIF (30%) and the degradation product 125I-tyrosine

  17. Design and Synthesis of High-Affinity Dimeric Inhibitors Targeting the Interactions between Gephyrin and Inhibitory Neurotransmitter Receptors

    DEFF Research Database (Denmark)

    Maric, Hans-Michael; Kasaragod, Vikram Babu; Kedström, Linda Maria Haugaard;

    2015-01-01

    Gephyrin is the central scaffolding protein for inhibitory neurotransmitter receptors in the brain. Here we describe the development of dimeric peptides that inhibit the interaction between gephyrin and these receptors, a process which is fundamental to numerous synaptic functions and diseases of...

  18. Pharmacokinetics, toxicity, side effects, receptor affinities and in vitro radiosensitizing effects of the novel metoclopramide formulations, sensamide and neu-sensamide

    International Nuclear Information System (INIS)

    Metoclopramide is a drug which has experienced worldwide use in the clinic for over 30 years as an antiemetic. Recently, it has also been shown to possess radio- and chemosensititzing properties in both animal tumour models and humans at the higher dose of 2 mg/kg. Two new metoclopramide formulations are being clinically developed and they differ mainly in whether the pH of their formulations are acidic (pH 2.5-3.5) or neutral (pH 6.5-7.0). Here we report that intramuscular administration of neutral metoclopramide is about 100% bioavailable, safer and with reduced side effects compared to acidic metoclopramide delivered by intramuscular injection to rats within the dose ranger of 3.5 to 14 mg/kg. The intramuscular administration of metoclopramide was also about 100% bioavailable compared to the intravenous route of administration. Furthermore, neutral metoclopramide had significantly decreased affinity for dopamine D2 receptors and increased affinity for 5-hydroxytryptamine3 receptors, but the radiosensitizing potency was the same, when compared to equimolar concentrations of acidic metoclopramide. Taken together these data support the continued development of neutral metoclopramide for high dose intramuscular administration of metoclopramide for future clinical use as both an antiemetic and radiosensitizer. (au) 29 refs

  19. Synthesis of 16α-[125I]iodo-5α-dihydrotestosterone and evaluation of its affinity for the androgen receptor

    International Nuclear Information System (INIS)

    Analogs of 5α-dihydrotestosterone, halogenated at carbon 11, were synthesized as potential gamma-emitting ligands for the androgen receptor. These compounds, were chosen for synthesis because estradiol, similarly substituted, is strongly bound to the estrogen receptor, and both androgen and estrogen receptors have generally similar structural requirements for the D-ring. The 16α-halogenated steroids, including 16α-[125I]iodo-5α-dihydrotestosterone were synthesized from 16β-bromo-5α-dihydrotestosterone by halogen exchange. The cis-β-bromohydrin substrate was synthesized from 5α-androstane-3,17-dione by selective ketalization, dibromination at C-16 and stereoselective reduction. 16α-iodo dihydrotestosterone was devoid of androgen activity in vivo at concentrations at which 5α-dihydrotestosterone was fully stimulatory. The 16α-iodo, 16α-bromo, and 16β-bromo analogs were allowed to compete with [3H]-dihydrotestosterone for binding to the androgen receptor; the 16α-iodo compound had a relative binding affinity 1/100th and both bromo compounds 1/30th that of dihydrotestosterone. In addition, no specific binding was detected when the 16α-[125I]iodo analog was incubated with prostatic cytosol. (author)

  20. α4βδ GABA receptors are high-affinity targets for γ-hydroxybutyric acid (GHB)

    DEFF Research Database (Denmark)

    Absalom, N.; Karim, N.; Eghorn, L.F.;

    2012-01-01

    γ-Hydroxybutyric acid (GHB) binding to brain-specific high-affinity sites is well-established and proposed to explain both physiological and pharmacological actions. However, the mechanistic links between these lines of data are unknown. To identify molecular targets for specific GHB high-affinit...... physiology. This finding will aid in elucidating the molecular mechanisms behind the proposed function of GHB as a neurotransmitter and its unique therapeutic effects in narcolepsy and alcoholism....

  1. Nicotine Ameliorates NMDA Receptor Antagonist-Induced Deficits in Contextual Fear Conditioning through High Affinity Nicotinic Acetylcholine Receptors in the Hippocampus

    OpenAIRE

    André, Jessica M.; Leach, Prescott T.; Gould, Thomas J.

    2010-01-01

    NMDA glutamate receptors (NMDARs) and nicotinic acetylcholine receptors (nAChRs) are both involved in learning and synaptic plasticity. Increasing evidence suggests processes mediated by these receptors may interact to modulate learning; however, little is known about the neural substrates involved in these interactive processes. The present studies investigated the effects of nicotine on MK-801 hydrogen maleate (MK-801) and DL-2-Amino-5-phosphonovaleric acid (APV) induced disruption of conte...

  2. Construction of a high affinity zinc binding site in the metabotropic glutamate receptor mGluR1

    DEFF Research Database (Denmark)

    Jensen, Anders A.; Sheppard, P O; Jensen, L B;

    2001-01-01

    The metabotropic glutamate receptors (mGluRs) belong to family C of the G-protein-coupled receptor (GPCR) superfamily. The receptors are characterized by having unusually long amino-terminal domains (ATDs), to which agonist binding has been shown to take place. Previously, we have constructed a...... "closed" conformation, and thus stabilizing a more or less inactive "open" form of the ATD. This study presents the first metal ion site constructed in a family C GPCR. Furthermore, it is the first time a metal ion site has been created in a region outside of the seven transmembrane regions of a GPCR and...... the loops connecting these. The findings offer valuable insight into the mechanism of ATD closure and family C receptor activation. Furthermore, the findings demonstrate that ATD regions other than those participating in agonist binding could be potential targets for new generations of ligands for...

  3. Role of IgE in autoimmunity.

    Science.gov (United States)

    Sanjuan, Miguel A; Sagar, Divya; Kolbeck, Roland

    2016-06-01

    There is accumulating evidence to suggest that IgE plays a significant role in autoimmunity. The presence of circulating self-reactive IgE in patients with autoimmune disorders has been long known but, at the same time, largely understudied. However, studies have shown that the increased IgE concentration is not associated with higher prevalence for atopy and allergy in patients with autoimmune diseases, such as systemic lupus erythematosus. IgE-mediated mechanisms are conventionally known to facilitate degranulation of mast cells and basophils and promote TH2 immunity, mechanisms that are not only central to mounting an appropriate defense against parasitic worms, noxious substances, toxins, venoms, and environmental irritants but that also trigger exuberant allergic reactions in patients with allergies. More recently, IgE autoantibodies have been recognized to participate in the self-inflicted damaging immune responses that characterize autoimmunity. Such autoimmune responses include direct damage on tissue-containing autoantigens, activation and migration of basophils to lymph nodes, and, as observed most recently, induction of type 1 interferon responses from plasmacytoid dendritic cells. The importance of IgE as a central pathogenic mechanism in autoimmunity has now been clinically validated by the approval of omalizumab, an anti-IgE mAb, for patients with chronic spontaneous urticaria and for the clinical benefit of patients with bullous pemphigoid. In this review we summarize recent reports describing the prevalence of self-reactive IgE and discuss novel findings that incriminate IgE as central in the pathogenesis of inflammatory autoimmune disorders. PMID:27264000

  4. High-affinity prorenin binding to cardiac man-6-P/IGF-II receptors precedes proteolytic activation to renin

    OpenAIRE

    Saris, Jasper; Derkx, Frans; Bruin, René; Dekkers, Dick; Lamers, Jos; Saxena, Pramod Ranjan; Schalekamp, Maarten; Danser, Jan

    2001-01-01

    textabstractMannose-6-phosphate (man-6-P)/insulin-like growth factor-II (man-6-P/IgF-II) receptors are involved in the activation of recombinant human prorenin by cardiomyocytes. To investigate the kinetics of this process, the nature of activation, the existence of other prorenin receptors, and binding of native prorenin, neonatal rat cardiomyocytes were incubated with recombinant, renal, or amniotic fluid prorenin with or without man-6-P. Intact and activated prorenin were measured in cell ...

  5. The discovery of epidepride and its analogs as high-affinity radioligands for imaging extrastriatal dopamine D(2) receptors in human brain.

    Science.gov (United States)

    de Paulis, Tomas

    2003-01-01

    [(123)I]Epidepride, [(18)F]fallypride, and [(76)Br]isoremoxipride (FLB-457) and their corresponding [(11)C]labeled derivatives belong to a class of high-affinity radioligands for SPECT or PET imaging of dopamine D(2) receptors in the human brain. In contrast to previously used imaging agents, these ligands are capable of identifying extrastriatal dopamine D(2) receptors. The design of these substituted benzamides derive its origin from the atypical antipsychotic agent, remoxipride. Starting in the late 1970's, halogenated analogs of (S)-sulpiride were evaluated in binding assays and behavioral studies, leading to the discovery of remoxipride. Remoxipride was 10 times weaker than sulpiride in vitro but 50 times more potent in vivo. Search for a putative active metabolite of remoxipride led to the discovery of raclopride and eticlopride, the former becoming a useful radioligand as tritium or carbon-11 labeled form for receptor binding and PET studies, respectively. In the US, the mono-iodine analog of raclopride, [(123)I]iodobenzamide (IBZM), was found to have moderate putamen-to-cerebellum ratio in rat and human brain. Continued search for metabolites of remoxipride led to the discovery of its 3,6-dihydroxy derivative, NCQ-344, with an extremely potent in vivo activity in the rat. SAR studies of the metabolites of remoxipride led to the discovery of the 3-methoxy isomer, isoremoxipride (FLB-457) and its corresponding 6-hydroxy analog, FLB-463, both having affinities for the dopamine D(2) receptor in the 20-30 pM range. Later, the 5-[(123)I]iodo analog of FLB-463, [(123)I]ioxipride ([(123)I]NCQ-298), became a potential SPECT imaging agent. In the mean time, the deshydroxy analog of IBZM, [(125)I]iodopride, showed binding potential in the rat similar to [(125)I]IBZM. Epidepride was designed by combining the structure of isoremoxipride with that of iodopride. In 1988, epidepride was independently prepared and radiolabeled in three separate laboratories in Stockholm

  6. Monitoring β-arrestin recruitment via β-lactamase enzyme fragment complementation: purification of peptide E as a low-affinity ligand for mammalian bombesin receptors.

    Science.gov (United States)

    Ikeda, Yuichi; Kumagai, Hidetoshi; Okazaki, Hiroaki; Fujishiro, Mitsuhiro; Motozawa, Yoshihiro; Nomura, Seitaro; Takeda, Norifumi; Toko, Haruhiro; Takimoto, Eiki; Akazawa, Hiroshi; Morita, Hiroyuki; Suzuki, Jun-ichi; Yamazaki, Tsutomu; Komuro, Issei; Yanagisawa, Masashi

    2015-01-01

    Identification of cognate ligands for G protein-coupled receptors (GPCRs) provides a starting point for understanding novel regulatory mechanisms. Although GPCR ligands have typically been evaluated through the activation of heterotrimeric G proteins, recent studies have shown that GPCRs signal not only through G proteins but also through β-arrestins. As such, monitoring β-arrestin signaling instead of G protein signaling will increase the likelihood of identifying currently unknown ligands, including β-arrestin-biased agonists. Here, we developed a cell-based assay for monitoring ligand-dependent GPCR-β-arrestin interaction via β-lactamase enzyme fragment complementation. Inter alia, β-lactamase is a superior reporter enzyme because of its cell-permeable fluorescent substrate. This substrate makes the assay non-destructive and compatible with fluorescence-activated cell sorting (FACS). In a reporter cell, complementary fragments of β-lactamase (α and ω) were fused to β-arrestin 2 and GPCR, respectively. Ligand stimulation initiated the interaction of these chimeric proteins (β-arrestin-α and GPCR-ω), and this inducible interaction was measured through reconstituted β-lactamase activity. Utilizing this system, we screened various mammalian tissue extracts for agonistic activities on human bombesin receptor subtype 3 (hBRS3). We purified peptide E as a low-affinity ligand for hBRS3, which was also found to be an agonist for the other two mammalian bombesin receptors such as gastrin-releasing peptide receptor (GRPR) and neuromedin B receptor (NMBR). Successful purification of peptide E has validated the robustness of this assay. We conclude that our newly developed system will facilitate the discovery of GPCR ligands. PMID:26030739

  7. Monitoring β-arrestin recruitment via β-lactamase enzyme fragment complementation: purification of peptide E as a low-affinity ligand for mammalian bombesin receptors.

    Directory of Open Access Journals (Sweden)

    Yuichi Ikeda

    Full Text Available Identification of cognate ligands for G protein-coupled receptors (GPCRs provides a starting point for understanding novel regulatory mechanisms. Although GPCR ligands have typically been evaluated through the activation of heterotrimeric G proteins, recent studies have shown that GPCRs signal not only through G proteins but also through β-arrestins. As such, monitoring β-arrestin signaling instead of G protein signaling will increase the likelihood of identifying currently unknown ligands, including β-arrestin-biased agonists. Here, we developed a cell-based assay for monitoring ligand-dependent GPCR-β-arrestin interaction via β-lactamase enzyme fragment complementation. Inter alia, β-lactamase is a superior reporter enzyme because of its cell-permeable fluorescent substrate. This substrate makes the assay non-destructive and compatible with fluorescence-activated cell sorting (FACS. In a reporter cell, complementary fragments of β-lactamase (α and ω were fused to β-arrestin 2 and GPCR, respectively. Ligand stimulation initiated the interaction of these chimeric proteins (β-arrestin-α and GPCR-ω, and this inducible interaction was measured through reconstituted β-lactamase activity. Utilizing this system, we screened various mammalian tissue extracts for agonistic activities on human bombesin receptor subtype 3 (hBRS3. We purified peptide E as a low-affinity ligand for hBRS3, which was also found to be an agonist for the other two mammalian bombesin receptors such as gastrin-releasing peptide receptor (GRPR and neuromedin B receptor (NMBR. Successful purification of peptide E has validated the robustness of this assay. We conclude that our newly developed system will facilitate the discovery of GPCR ligands.

  8. Synthesis, binding affinity at glutamic acid receptors, neuroprotective effects, and molecular modeling investigation of novel dihydroisoxazole amino acids

    DEFF Research Database (Denmark)

    Conti, Paola; De Amici, Marco; Grazioso, Giovanni;

    2005-01-01

    The four stereoisomers of 5-(2-amino-2-carboxyethyl)-4,5-dihydroisoxazole-3-carboxylic acid(+)-4, (-)-4, (+)-5, and (-)-5 were prepared by stereoselective synthesis of two pairs of enantiomers, which were subsequently resolved by enzymatic procedures. These four stereoisomers and the four...... stereoisomers of the bicyclic analogue 5-amino-4,5,6,6a-tetrahydro-3aH-cyclopenta[d]isoxazole-3,5-dicarboxylic acid (+)-2, (-)-2, (+)-3, and (-)-3 were tested at ionotropic and metabotropic glutamate receptor subtypes. The most potent NMDA receptor antagonists [(+)-2, (-)-4, and (+)-5] showed a significant...... derivatives showed high antagonist potency with preference for the NR2A and NR2B subtypes, with derivative (-)-4 behaving as the most potent antagonist. The biological data are discussed on the basis of homology models reported in the literature for NMDA receptors and mGluRs....

  9. Detection of Interaction of Binding Affinity of Aromatic Hydrocarbon Receptor to the Specific DNA by Exonuclease Protection Mediated PCR Assay

    Institute of Scientific and Technical Information of China (English)

    SUN Xi; XU Shunqing

    2005-01-01

    A novel exonuclease protection mediated PCR assay (EPM-PCR) to detect the interaction of protein and DNA at a dioxin-responsive enhancer (DRE) upstream of the CYP1A1 gene in rat hepatic cytosol was established. A double-stranded DNA fragment containing two binding sites was designed and incubated with the aryl hydrocarbon receptor (AhR) transformed by 2,3,7,8-tet rachlorodibenzo p dioxin (TCDD) to generate TCDD: AhR: DNA complex which could protect receptor-binding DNA against exonuclease Ⅲ (Exo Ⅲ) digestion. With ExoⅢ treatment, free DNAs were digested and receptor-bound DNAs remained that could be amplified by PCR. By agarose gel electrophoreses a clear band (285bp) was detected using TCDD-treated sample, while nothing with control samples. To detect transformed AhR-DRE complex, 2 fmol DNAs and 3 ug cytosol proteins were found to be sufficient in the experiment. Compared with gel retardation assay, this new method is more sensitive for monitoring the Ah receptor-enhancer interaction without radioactive pollution.

  10. LYR3, a high-affinity LCO-binding protein of Medicago truncatula, interacts with LYK3, a key symbiotic receptor.

    Science.gov (United States)

    Fliegmann, Judith; Jauneau, Alain; Pichereaux, Carole; Rosenberg, Charles; Gasciolli, Virginie; Timmers, Antonius C J; Burlet-Schiltz, Odile; Cullimore, Julie; Bono, Jean-Jacques

    2016-05-01

    LYR3, LYK3, and NFP are lysin motif-containing receptor-like kinases (LysM-RLKs) from Medicago truncatula, involved in perception of symbiotic lipo-chitooligosaccharide (LCO) signals. Here, we show that LYR3, a high-affinity LCO-binding protein, physically interacts with LYK3, a key player regulating symbiotic interactions. In vitro, LYR3 is phosphorylated by the active kinase domain of LYK3. Fluorescence lifetime imaging/Förster resonance energy transfer (FLIM/FRET) experiments in tobacco protoplasts show that the interaction between LYR3 and LYK3 at the plasma membrane is disrupted or inhibited by addition of LCOs. Moreover, LYR3 attenuates the cell death response, provoked by coexpression of NFP and LYK3 in tobacco leaves. PMID:27129432

  11. Activation of high and low affinity dopamine receptors generates a closed loop that maintains a conductance ratio and its activity correlate

    Directory of Open Access Journals (Sweden)

    Ryan M Hooper

    2013-10-01

    Full Text Available Neuromodulators alter network output and have the potential to destabilize a circuit. The mechanisms maintaining stability in the face of neuromodulation are not well described. Using the pyloric network in the crustacean stomatogastric nervous system, we show that dopamine (DA does not simply alter circuit output, but activates a closed loop in which DA-induced alterations in circuit output consequently drive a change in an ionic conductance to preserve a conductance ratio and its activity correlate. DA acted at low affinity type 1 receptors (D1Rs to induce an immediate modulatory decrease in the transient potassium current (IA of a pyloric neuron. This, in turn, advanced the activity phase of that component neuron, which disrupted its network function and thereby destabilized the circuit. DA simultaneously acted at high affinity D1Rs on the same neuron to confer activity-dependence upon the hyperpolarization activated current (Ih such that the DA-induced changes in activity subsequently reduced Ih. This DA-enabled, activity-dependent, intrinsic plasticity exactly compensated for the modulatory decrease in IA to restore the IA:Ih ratio and neuronal activity phase, thereby closing an open loop created by the modulator. Activation of closed loops to preserve conductance ratios may represent a fundamental operating principle neuromodulatory systems use to ensure stability in their target networks.

  12. Identification of a point mutation in type IIB von Willebrand disease illustrating the regulation of von Willebrand factor affinity for the platelet membrane glycoprotein Ib-IX receptor

    International Nuclear Information System (INIS)

    von Willebrand factor (vWF) supports platelet adhesion on thrombogenic surfaces by binding to platelet membrane glycoprotein (GP) Ib in the GP Ib-IX receptor complex. This interaction is physiologically regulated so that it does not occur between circulating vWF and platelets but, rather, only at a site of vascular injury. The abnormal vWF found in type IIB von Willebrand disease, however, has a characteristically increased affinity for GP Ib and binds to circulating platelets. The authors have analyzed the molecular basis of this abnormality by sequence analysis of a type IIB vWF cDNA and have identified a single amino acid change, Trp550 to Cys550, located in the GP IB-binding domain of the molecule comprising residues 449-728. Bacterial expression of recombinant fragments corresponding to this vWF domain yielded molecules that, whether containing a normal Trp550 or a mutant Cys550 residue, bound directly to GP Ib in the absence of modulators and with similar affinity. These results identify a region of vWF that, although not thought to be directly involved in binding to GP Ib, may modulate the interaction through conformational changes

  13. High-affinity binding of [3H]estradiol-17 beta by an estrogen receptor in the liver of the turtle

    International Nuclear Information System (INIS)

    Specific [3H]estradiol-17 beta ([3H]E2) binding activity (EBA) with characteristics of an estrogen receptor (ER) was demonstrated in cytosols and nuclear extracts of the female turtle, Chrysemys picta. Three different receptor assays (dextran-coated charcoal assay, hydroxylapatite batch procedure, and DNA-cellulose chromatography) were evaluated in terms of their applicability in analyzing large numbers of samples. For the measurement of cytosolic EBA, the hydroxylapatite batch procedure was found to be the most reliable assay. On the other hand, the dextran-coated charcoal assay was found to be the most appropriate method for the measurement of nuclear EBA. Turtle hepatic EBA binds [3H]E2 with high affinity (cytosolic, 17.4 +/- 2.8 X 10(9) M-1; nuclear, 17.7 +/- 1.9 X 10(9) M-1), limited capacity (cytosolic, 133.7 +/- 4.6 fmol/g tissue; nuclear, 81.1 +/- 9.0 fmol/g tissue), and strict steroid specificity. The EBA bound natural estrogens (E2, estrone, estriol) as well as the nonsteroidal estrogen, diethylstilbestrol, but exhibited little affinity for androgens, progesterone, or corticosterone. The turtle hepatic EBA resembled mammalian and avian ERs in terms of binding characteristics; however, unlike mammalian and avian ERs it was shown to be heat-labile. Incubation at 30 degrees caused rapid loss of [3H]E2 binding activity in both cytosolic and nuclear fractions. The exchange between [3H]E2 and the endogenously bound estrogen was slow at 4 and 15 degrees, but the exchange process was facilitated in the presence of the chaotropic salt, NaSCN. Establishment of quantitation methods for both cytosolic and nuclear forms of EBA will enable future investigation of the mechanism and regulation of estrogen action in the liver of this turtle species

  14. Affinity of estrogens for human progesterone receptor A and B monomers and risk of breast cancer: a comparative molecular modeling study

    Directory of Open Access Journals (Sweden)

    Tarique N Hasan

    2011-03-01

    Full Text Available Tarique N Hasan1,4, Leena Grace B2, Tariq A Masoodi3,5, Gowhar Shafi4 , Ali A. Alshatwi4, P Sivashanmugham31Department of Biotechnology, Bharathiar University, Coimbator, TN, India; 2Department of Biotechnology, V. M. K. V. College of Engineering, Salem, TN, India; 3Department of Bioinformatics, Jamal Mohammed College, Bharathidasan University, Tiruchirappalli, India; 4Molecular Cancer Biology Laboratory, Department of Food Science and Nutrition, College of Food and Agricultural Sciences; 5Department of Community Health Sciences, College of Applied Medical Sciences, King Saud University, Saudi ArabiaBackground: The human progesterone receptor (hPR belongs to the steroid receptor family. It may be found as monomers (A and B and or as a dimer (AB. hPR is regarded as the prognostic biomarker for breast cancer. In a cellular dimer system, AB is the dominant species in most cases. However, when a cell coexpresses all three isoforms of hPR, the complexity of the action of this receptor increases. For example, hPR A suppresses the activity of hPR B, and the ratio of hPR A to hPR B may determine the physiology of a breast tumor. Also, persistent exposure of hPRs to nonendogenous ligands is a common risk factor for breast cancer. Hence we aimed to study progesterone and some nonendogenous ligand interactions with hPRs and their molecular docking.Methods and results: A pool of steroid derivatives, namely, progesterone, cholesterol, testosterone, testolectone, estradiol, estrone, norethindrone, exemestane, and norgestrel, was used for this in silico study. Dockings were performed on AutoDock 4.2. We found that estrogens, including estradiol and estrone, had a higher affinity for hPR A and B monomers in comparison with the dimer, hPR AB, and that of the endogenous progesterone ligand. hPR A had a higher affinity to all the docked ligands than hPR B.Conclusion: This study suggests that the exposure of estrogens to hPR A as well as hPR B, and more

  15. Response to platelet-activating factor in human platelets stored and aged in plasma. Decrease in aggregation, phosphoinositide turnover, and receptor affinity

    International Nuclear Information System (INIS)

    Human platelet concentrates were stored in polyolefin bags at 22 to 24 degrees C on a horizontal shaker for up to 8 days. At different intervals, aliquots of platelet-rich plasma (PRP) were removed aseptically and five variables, i.e., platelet counts, morphology, platelet-activating factor (PAF)-stimulated aggregation, phosphoinositide turnover, and [3H]PAF binding to platelet receptors, were studied. The number of platelets did not change during the 8 days of storage. Scanning electron microscopy of the platelets revealed a gradual morphologic change from biconcave flat discs to irregular, crenated forms. The PAF-induced aggregation of platelets declined with time of storage. A decrease to 50 percent of the Day 1 aggregatory response to PAF was evident on Day 2, and there was a further decline to about 20 percent by Day 6. Similarly, PAF receptor-coupled phosphoinositide turnover, as monitored by 32P incorporation into individual phosphoinositides, decreased dramatically with storage. After 2 to 3 days of storage, the phosphoinositide turnover was reduced to 50 percent of the original response, and it continued to decline to about 25 percent of original response by Day 5 or 6. The binding of [3H]PAF to washed human platelets indicated subtle changes between Days 2 and 4, which became more noticeable by Day 6. These results have raised the possibility of changes in the number of the receptors and/or their affinity for the ligand during storage. We conclude that although the number of platelets was maintained during storage for 8 days, a general deterioration of their responses to PAF occurred at the levels of cell surface receptor, transmembrane signaling (phosphoinositide turnover), and response (aggregation)

  16. Hyper IgE syndrome: Report of two cases with moderate elevation of IgE

    Directory of Open Access Journals (Sweden)

    Muhammed K

    2005-03-01

    Full Text Available Hyper IgE syndrome with recurrent infection (Job′s syndrome is a rare idiopathic primary immunodeficiency disease characterized by the triad of elevated serum IgE (>2000 IU/ml, recurrent cutaneous abscesses and recurrent sinopulmonary infections. The bacteria which commonly infect these patients are Staphylococcus aureus and Haemophilus influenzae. Therapy should include prolonged antibiotic therapy and early surgery. Non-specific agents like levamisole and ascorbic acid may reduce recurrent infections. We are reporting two girls, six and twelve years of age, presented with recurrent cutaneous and respiratory infections and moderately elevated levels of serum IgE.

  17. Phase I hydroxylated metabolites of the K2 synthetic cannabinoid JWH-018 retain in vitro and in vivo cannabinoid 1 receptor affinity and activity.

    Directory of Open Access Journals (Sweden)

    Lisa K Brents

    Full Text Available BACKGROUND: K2 products are synthetic cannabinoid-laced, marijuana-like drugs of abuse, use of which is often associated with clinical symptoms atypical of marijuana use, including hypertension, agitation, hallucinations, psychosis, seizures and panic attacks. JWH-018, a prevalent K2 synthetic cannabinoid, is structurally distinct from Δ(9-THC, the main psychoactive ingredient in marijuana. Since even subtle structural differences can lead to differential metabolism, formation of novel, biologically active metabolites may be responsible for the distinct effects associated with K2 use. The present study proposes that K2's high adverse effect occurrence is due, at least in part, to distinct JWH-018 metabolite activity at the cannabinoid 1 receptor (CB1R. METHODS/PRINCIPAL FINDINGS: JWH-018, five potential monohydroxylated metabolites (M1-M5, and one carboxy metabolite (M6 were examined in mouse brain homogenates containing CB1Rs, first for CB1R affinity using a competition binding assay employing the cannabinoid receptor radioligand [(3H]CP-55,940, and then for CB1R intrinsic efficacy using an [(35S]GTPγS binding assay. JWH-018 and M1-M5 bound CB1Rs with high affinity, exhibiting K(i values that were lower than or equivalent to Δ(9-THC. These molecules also stimulated G-proteins with equal or greater efficacy relative to Δ(9-THC, a CB1R partial agonist. Most importantly, JWH-018, M2, M3, and M5 produced full CB1R agonist levels of activation. CB1R-mediated activation was demonstrated by blockade with O-2050, a CB1R-selective neutral antagonist. Similar to Δ(9-THC, JWH-018 and M1 produced a marked depression of locomotor activity and core body temperature in mice that were both blocked by the CB1R-preferring antagonist/inverse agonist AM251. CONCLUSIONS/SIGNIFICANCE: Unlike metabolites of most drugs, the studied JWH-018 monohydroxylated compounds, but not the carboxy metabolite, retain in vitro and in vivo activity at CB1Rs. These observations

  18. Synthesis and preliminary evaluation of [123I]1-(4-cyanobenzyl)-4-[[(trans-iodopropen-2-yl)oxy]-methyl]piperidine: A novel high affinity sigma receptor radioligand for SPECT

    International Nuclear Information System (INIS)

    1-(4-Cyanobenzyl)-4-[[(trans-iodopropen-2-yl)oxy]methyl]piperidine (2) has been synthesized as a novel iodinated ligand for sigma receptors. This new compound exhibited high affinity (Ki = 0.38 nM) for the sigma-1 receptor and selectivity for sigma-1 vs. sigma-2 receptors (Ki σ-1/σ-2 0.02) using in vitro receptor binding assays. Moderate affinity for muscarinic M1 (K1 = 178 nM) receptors and low affinity (Ki = 1,460 nM) for dopamine D2 receptors was also noted. The lipophilicity of 2 (log P7.5 3.24) is moderate, indicating that the ligand should readily cross the blood/brain barrier. The corresponding radioiodinated ligand, 123I-2, was synthesized from the appropriate trans vinyl tributylstannane precursor under acidic conditions using oxidative iododestannylation methods. HPLC purification provided the radioligand in yields ranging between 63 and 75% EOS (n = 5) and with > 99% radiochemical purity and a specific activity > 77,000 MBq/μmol. Preliminary in vivo distribution and pharmacological blocking studies using 123I-2 were performed in male Australian Albino Wistar rats. 123I-2 exhibited good brain uptake (1.11 ± 0.14 %ID at 20 min PI) with no significant loss of radioactivity from the brain over the course of the study (4 h). The gross regional brain distribution of the radioligand showed highest uptake in the posterior cortex and frontal cortex, with slightly lower uptake in other brain regions examined. Most of the uptake of radioactivity in the brain, lung, heart, muscle, and kidney was prevented by pre-administration of compounds with affinity for sigma receptors, indicating the in vivo specificity of the ligand. In view of these results, 123I-2 shows promise for the in vivo tomographic evaluation of sigma receptor densities

  19. Dissecting the Relation between a Nuclear Receptor and GATA: Binding Affinity Studies of Thyroid Hormone Receptor and GATA2 on TSHβ Promoter

    OpenAIRE

    Ana Carolina Migliorini Figueira; Igor Polikarpov; Dmitry Veprintsev; Guilherme Martins Santos

    2010-01-01

    BACKGROUND: Much is known about how genes regulated by nuclear receptors (NRs) are switched on in the presence of a ligand. However, the molecular mechanism for gene down-regulation by liganded NRs remains a conundrum. The interaction between two zinc-finger transcription factors, Nuclear Receptor and GATA, was described almost a decade ago as a strategy adopted by the cell to up- or down-regulate gene expression. More recently, cell-based assays have shown that the Zn-finger region of GATA2 ...

  20. Synthesis and biological evaluation of technetium(III) mixed-ligand complexes with high affinity for the cerebral 5-HT1A receptor and the alpha1-adrenergic receptor

    International Nuclear Information System (INIS)

    Tc(III) and Re(III) complexes [M(NS3)(CNR)] (M=Re, 99mTc, NS3=2,2',2''-nitrilotris(ethanethiol), CNR = functionalized isocyanide bearing a derivative of WAY 100635) have been synthesized and characterized. Re was used as Tc surrogate for chemical characterization and in vitro receptor-binding studies. For two representatives subnanomolar affinities for the 5-HT1A as well as for the alpha1-adrenergic receptor were reached. Biodistribution studies in rats of the 99mTc complexes showed brain uptakes between 0.3 and 0.5% ID/organ (5 min p.i.). In vitro autoradiography of one 99mTc representative in sections of post mortem human brain indicate its accumulation in 5-HT1A receptor-rich brain regions. However, addition of the specific 5-HT1A receptor agonist 8-OH-DPAT as well as the alpha1-adrenoceptor antagonist prazosin could not substantially block this tracer accumulation. A preliminary SPET study in a monkey showed negligible brain uptake

  1. Primary Immunodeficiencies with Elevated IgE.

    Science.gov (United States)

    Mogensen, Trine H

    2016-01-01

    In recent years a number of primary immunodeficiencies (PIDs) characterized by elevated Immunoglobulin E (IgE) levels have been uncovered and termed as Hyper-IgE syndrome (HIES). In addition to the elevated levels of IgE, patients with these PIDs display a spectrum of infections by staphylococci and fungi, and in some cases viruses, particularly affecting skin and lungs. Most of these PIDs also have a non-infectious phenotype, comprising musculoskeletal, vascular, and neurological abnormalities. The genetic basis for the majority of conditions with elevated IgE has now been established and includes mutations in STAT3, DOCK8, TYK2, and most recently PGM3 molecules. However, in some patients with the relevant phenotype, mutations in these molecules are not identified, suggesting additional genetic etiologies of HIES not yet discovered. As the immunological and molecular basis of HIES is being unraveled, important insights are emerging that may have implications for our understanding of basic principles of immunology and protective immunity as well as for the pathogenesis and clinical management of patients with these complex and challenging PIDs. In this review, are presented the current knowledge on the clinical presentation, infectious phenotype, and the genetic and immunological pathogenesis of hyper-IgE syndromes as well as some other PIDs with elevated levels of IgE. PMID:25970001

  2. Carbon-11 labelling of eticlopride in two different positions - a selective high-affinity ligand for the study of dopamine D-2 receptors using PET

    International Nuclear Information System (INIS)

    A new highly selective high-affinity dopamine D-2 receptor antagonist, eticlopride ((-)-(S)-5-chloro-3-ethyl-N-(1-ethyl-2-pyrrolidinyl)methyl)-6-methoxysalicylamide), was labelled with 11C in two different positions ([N-ethyl-11C]eticlopride (I) and ([methyl-11C]eticlopride (II)). Product I was prepared by N-alkylation of the N-desethyl compound with [11C]ethyl iodide. II was prepared by O-alkylation of the diphenolic precursor with [11C]methyl iodide followed by separation of the two methylated products. The radiochemical yields were 15-20% (EOB) with an overall synthesis time of 45-60 min. Both compounds were isolated by semi-preparative HPLC and the radiochemical purity was in both cases > 99%. I was injected i.v. in a Cynomolgus monkey and brain radioactivity was measured by positron emission tomography (PET). The specific activity was 70 Ci/mmol at time of injection. There was a marked accumulation of radioactivity in the basal ganglia, regions known to have a high density of dopamine D-2 receptors. (author)

  3. Fc receptors for immunoglobulins and their appearance during vertebrate evolution.

    Science.gov (United States)

    Akula, Srinivas; Mohammadamin, Sayran; Hellman, Lars

    2014-01-01

    Receptors interacting with the constant domain of immunoglobulins (Igs) have a number of important functions in vertebrates. They facilitate phagocytosis by opsonization, are key components in antibody-dependent cellular cytotoxicity as well as activating cells to release granules. In mammals, four major types of classical Fc receptors (FcRs) for IgG have been identified, one high-affinity receptor for IgE, one for both IgM and IgA, one for IgM and one for IgA. All of these receptors are related in structure and all of them, except the IgA receptor, are found in primates on chromosome 1, indicating that they originate from a common ancestor by successive gene duplications. The number of Ig isotypes has increased gradually during vertebrate evolution and this increase has likely been accompanied by a similar increase in isotype-specific receptors. To test this hypothesis we have performed a detailed bioinformatics analysis of a panel of vertebrate genomes. The first components to appear are the poly-Ig receptors (PIGRs), receptors similar to the classic FcRs in mammals, so called FcRL receptors, and the FcR γ chain. These molecules are not found in cartilagous fish and may first appear within bony fishes, indicating a major step in Fc receptor evolution at the appearance of bony fish. In contrast, the receptor for IgA is only found in placental mammals, indicating a relatively late appearance. The IgM and IgA/M receptors are first observed in the monotremes, exemplified by the platypus, indicating an appearance during early mammalian evolution. Clearly identifiable classical receptors for IgG and IgE are found only in marsupials and placental mammals, but closely related receptors are found in the platypus, indicating a second major step in Fc receptor evolution during early mammalian evolution, involving the appearance of classical IgG and IgE receptors from FcRL molecules and IgM and IgA/M receptors from PIGR. PMID:24816777

  4. Fc receptors for immunoglobulins and their appearance during vertebrate evolution.

    Directory of Open Access Journals (Sweden)

    Srinivas Akula

    Full Text Available Receptors interacting with the constant domain of immunoglobulins (Igs have a number of important functions in vertebrates. They facilitate phagocytosis by opsonization, are key components in antibody-dependent cellular cytotoxicity as well as activating cells to release granules. In mammals, four major types of classical Fc receptors (FcRs for IgG have been identified, one high-affinity receptor for IgE, one for both IgM and IgA, one for IgM and one for IgA. All of these receptors are related in structure and all of them, except the IgA receptor, are found in primates on chromosome 1, indicating that they originate from a common ancestor by successive gene duplications. The number of Ig isotypes has increased gradually during vertebrate evolution and this increase has likely been accompanied by a similar increase in isotype-specific receptors. To test this hypothesis we have performed a detailed bioinformatics analysis of a panel of vertebrate genomes. The first components to appear are the poly-Ig receptors (PIGRs, receptors similar to the classic FcRs in mammals, so called FcRL receptors, and the FcR γ chain. These molecules are not found in cartilagous fish and may first appear within bony fishes, indicating a major step in Fc receptor evolution at the appearance of bony fish. In contrast, the receptor for IgA is only found in placental mammals, indicating a relatively late appearance. The IgM and IgA/M receptors are first observed in the monotremes, exemplified by the platypus, indicating an appearance during early mammalian evolution. Clearly identifiable classical receptors for IgG and IgE are found only in marsupials and placental mammals, but closely related receptors are found in the platypus, indicating a second major step in Fc receptor evolution during early mammalian evolution, involving the appearance of classical IgG and IgE receptors from FcRL molecules and IgM and IgA/M receptors from PIGR.

  5. Making IGES work for CAD/CAM data exchange

    Energy Technology Data Exchange (ETDEWEB)

    Ames, A.L.; Fletcher, S.K.

    1985-01-01

    This report outlines Sandia National Laboratories effort to integrate the CAE/CAD/CAM capabilities of three laboratories and six production plants in the DOE weapons complex with the Initial Graphics Exchange Specifications (IGES). The problems with IGES are addressed and solutions are explained. The implementation of the IGES concept is also outlined.

  6. Development of affinity binding assay for isolation and analysis of microbial recognizing receptors for environmental signal substances

    International Nuclear Information System (INIS)

    Aiming at isolation and analysis of recognizing receptors for environmental signaling substances, a development of effective labeling method was attempted for oligosaccharides having a role as inducer for exo-enzyme production, various signaling substances among plant and microbes as well as bacterial and viral proteins directly mediating the interaction between pathogenic microbes and plants. First, the optimum labeling conditions for those substances were investigated. To isolate oligosaccharide receptor, labeling of chitobiose was performed. Radio-labeled UDP-GlcNAc was used as the substrate for chitin synthesizing enzyme to produce labeled chitobiose using chitinase. Thin layer chromatography of silica gel was useful for isolation and purification of the labeled chitobiose. Thus, chitin of a highly specific activity able to use for isolation of its receptor was obtained. Then, an expression system for chlorocatechol dioxygenase gene was constructed aiming to produce chloromucoic acid, an inducer of catabolic genes for aromatic chlorides through conversion from chlorocatechol. In this study, clcA gene from P. putida AC866 strain was inserted in expression vector pGST/HisT1 after PCR amplification and the vector that could produce His-tagged chlorocatechol-dioxygenase was constructed. Next, for analysis of a protein able to interact with a protein related to pathogenicity of plant virus, a labeling method for the protein coded in rice stripe virus (RSV) was developed. By autoradiographic analysis of translation products for the virus RNA, two bands corresponding to 22.8 K and 20.5 K proteins were detected. Whereas the translation products of RNA from RSV-infected host presented 5 bands, neither of which was detectable in the products from un-infected hosts. The two bands of them were identified as the proteins of nucleoside capsid, 22.8 K and 20.5 K proteins. Isolation of these labeled proteins would be made in near future. (M.N.)

  7. Characterization of high affinity neurotensin receptor NTR1 in HL-60 cells and its down regulation during granulocytic differentiation

    OpenAIRE

    Choi, Se-Young; Chae, Hee-Don; Park, Tae-Ju; Ha, Hyunjung; Kim, Kyong-Tai

    1999-01-01

    We investigated responses to neurotensin in human promyelocytic leukaemia HL-60 cells.Neurotensin increased the cytosolic calcium concentration ([Ca2+]i) in a concentration-dependent manner and also produced inositol 1,4,5-trisphosphate (InsP3).Among the tested neurotensin analogues, neurotensin 8-13, neuromedin-N, and xenopsin also increased [Ca2+]i, whereas neurotensin 1–11 and neurotensin 1–8 did not elicit detectable responses.SR48692, an antagonist of NTR1 neurotensin receptors, blocked ...

  8. Affinity-purified tetanus neurotoxin interaction with synaptic membranes: properties of a protease-sensitive receptor component

    International Nuclear Information System (INIS)

    The pharmacokinetic interaction of an affinity-purified 125I-labeled tetanotoxin fraction with guinea pig brain synaptosomal preparations was investigated. Binding of tetanotoxin was time- and temperature-dependent, was proportional to protein concentration, and was saturable at about 8 x 10-9 M as estimated by a solid-surface binding assay. Binding was optimal at pH 6.5 under low ionic strength buffer and was almost entirely blocked by gangliosides or antitoxin. In analogy to intact nerve cells, binding of toxin to membranes resulted in a tight association operationally defined as sequestration. Binding and sequestration were abolished after membrane pretreatment with sialidase. The enzyme could not dissociate the membrane-bound toxin formed at 4 or 370C under low ionic strength conditions, which is in part compatible with internalization as defined in nerve cell cultures. In the latter system the toxin could be removed at 40C but not at 370C. Binding was significantly reduced upon pretreatment of guinea pig brain membranes by a variety of hydrolytic enzymes. It is proposed that, in addition to a ganglioside, interaction of tetanotoxin with synaptic membranes is facilitated by a protein and may also require an appropriate lipid environment. These latter membrane constituents may play a pivotal role in the sequestration of the toxin

  9. DOTA-NOC, a high-affinity ligand of somatostatin receptor subtypes 2, 3 and 5 for labelling with various radiometals

    International Nuclear Information System (INIS)

    Earlier studies have shown that modification of the octapeptide octreotide in positions 3 and 8 may result in compounds with increased somatostatin receptor affinity that, if radiolabelled, display improved uptake in somatostatin receptor-positive tumours. The aim of a recent research study in our laboratory was to employ the parallel peptide synthesis approach by further exchanging the amino acid in position 3 of octreotide and coupling the macrocyclic chelator DOTA(1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid) to these peptides for labelling with radiometals like gallium-67 or -68, indium-111, yttrium-90 and lutetium-177. The purpose was to find radiopeptides with an improved somatostatin receptor binding profile in order to extend the spectrum of targeted tumours. A first peptide, [111In,90Y-DOTA]-1-Nal3-octreotide (111In,90Y-DOTA-NOC), was isolated which showed an improved profile. InIII-DOTA-NOC exhibited the following IC50 values (nM) when studied in competition with [125I][Leu8, d-Trp22, Tyr25]somatostatin-28 (values for YIII-DOTA-NOC are shown in parentheses): sstr2, 2.9±0.1 (3.3±0.2); sstr3, 8±2 (26±1.9); sstr5, 11.2±3.5 (10.4±1.6). Affinity towards sstr1 and 4 was very low or absent. InIII-DOTA-NOC is superior to all somatostatin-based radiopeptides having this particular type of binding profile, including DOTA-lanreotide, and has three to four times higher binding affinity to sstr2 than InIII,YIII-DOTA-Tyr3-octreotide (InIII,YIII-DOTA-TOC). In addition, [111In]DOTA-NOC showed a specific and high rate of internalization into AR4-2J rat pancreatic tumour cells which, after 4 h, was about two times higher than that of [111In]DOTA-TOC and three times higher than that of [111In]DOTA-octreotide ([111In]DOTA-OC). The internalized radiopeptides were externalized intact upon 2 h of internalization followed by an acid wash. After 2-3 h of externalization a plateau is reached, indicating a steady-state situation explained by reactivation of the

  10. Palmitic acid analogs exhibit nanomolar binding affinity for the HIV-1 CD4 receptor and nanomolar inhibition of gp120-to-CD4 fusion.

    Directory of Open Access Journals (Sweden)

    Elena E Paskaleva

    Full Text Available BACKGROUND: We recently reported that palmitic acid (PA is a novel and efficient CD4 fusion inhibitor to HIV-1 entry and infection. In the present report, based on in silico modeling of the novel CD4 pocket that binds PA, we describe discovery of highly potent PA analogs with increased CD4 receptor binding affinities (K(d and gp120-to-CD4 inhibition constants (K(i. The PA analogs were selected to satisfy Lipinski's rule of drug-likeness, increased solubility, and to avoid potential cytotoxicity. PRINCIPAL FINDINGS: PA analog 2-bromopalmitate (2-BP was most efficacious with K(d approximately 74 nM and K(i approximately 122 nM, ascorbyl palmitate (6-AP exhibited slightly higher K(d approximately 140 nM and K(i approximately 354 nM, and sucrose palmitate (SP was least efficacious binding to CD4 with K(d approximately 364 nM and inhibiting gp120-to-CD4 binding with K(i approximately 1486 nM. Importantly, PA and its analogs specifically bound to the CD4 receptor with the one to one stoichiometry. SIGNIFICANCE: Considering observed differences between K(i and K(d values indicates clear and rational direction for improving inhibition efficacy to HIV-1 entry and infection. Taken together this report introduces a novel class of natural small molecules fusion inhibitors with nanomolar efficacy of CD4 receptor binding and inhibition of HIV-1 entry.

  11. Quantification of human opiate receptor concentration and affinity using high and low specific activity ( sup 11 C)diprenorphine and positron emission tomography

    Energy Technology Data Exchange (ETDEWEB)

    Sadzot, B.; Price, J.C.; Mayberg, H.S.; Douglass, K.H.; Dannals, R.F.; Lever, J.R.; Ravert, H.T.; Wilson, A.A.; Wagner, H.N. Jr.; Feldman, M.A. (Johns Hopkins Medical Institutions, Baltimore, MD (USA))

    1991-03-01

    (11C)Diprenorphine, a weak partial opiate agonist, and positron emission tomography were used to obtain noninvasive regional estimates of opiate receptor concentration (Bmax) and affinity (Kd) in human brain. Different compartmental models and fitting strategies were compared statistically to establish the most reliable method of parameter estimation. Paired studies were performed in six normal subjects using high (769-5,920 Ci/mmol) and low (27-80 Ci/mmol) specific activity (SA) (11C)diprenorphine. Two subjects were studied a third time using high SA (11C)diprenorphine after a pretreatment with 1-1.5 mg/kg of the opiate antagonist naloxone. After the plasma radioactivity was corrected for metabolites, the brain data were analyzed using a three-compartment model and nonlinear least-squares curve fitting. Linear differential equations were used to describe the high SA (low receptor occupancy) kinetics. The k3/k4 ratio varied from 1.0 +/- 0.2 (occipital cortex) to 8.6 +/- 1.6 (thalamus). Nonlinear differential equations were used to describe the low SA (high receptor occupancy) kinetics and the curve fits provided the konf2 product. The measured free fraction of (11C)diprenorphine in plasma (f1) was 0.30 +/- 0.03, the average K1/k2 ratio from the two naloxone studies was 1.1 +/- 0.2, and the calculated free fraction of (11C)diprenorphine in the brain (f2) was 0.3. Using the paired SA studies, the estimated kinetic parameters, and f2, separate estimates of Bmax and Kd were obtained. Bmax varied from 2.3 +/- 0.5 (occipital cortex) to 20.6 +/- 7.3 (cingulate cortex) nM. The average Kd (eight brain regions) was 0.85 +/- 0.17 nM.

  12. Quantification of human opiate receptor concentration and affinity using high and low specific activity [11C]diprenorphine and positron emission tomography

    International Nuclear Information System (INIS)

    [11C]Diprenorphine, a weak partial opiate agonist, and positron emission tomography were used to obtain noninvasive regional estimates of opiate receptor concentration (Bmax) and affinity (Kd) in human brain. Different compartmental models and fitting strategies were compared statistically to establish the most reliable method of parameter estimation. Paired studies were performed in six normal subjects using high (769-5,920 Ci/mmol) and low (27-80 Ci/mmol) specific activity (SA) [11C]diprenorphine. Two subjects were studied a third time using high SA [11C]diprenorphine after a pretreatment with 1-1.5 mg/kg of the opiate antagonist naloxone. After the plasma radioactivity was corrected for metabolites, the brain data were analyzed using a three-compartment model and nonlinear least-squares curve fitting. Linear differential equations were used to describe the high SA (low receptor occupancy) kinetics. The k3/k4 ratio varied from 1.0 +/- 0.2 (occipital cortex) to 8.6 +/- 1.6 (thalamus). Nonlinear differential equations were used to describe the low SA (high receptor occupancy) kinetics and the curve fits provided the konf2 product. The measured free fraction of [11C]diprenorphine in plasma (f1) was 0.30 +/- 0.03, the average K1/k2 ratio from the two naloxone studies was 1.1 +/- 0.2, and the calculated free fraction of [11C]diprenorphine in the brain (f2) was 0.3. Using the paired SA studies, the estimated kinetic parameters, and f2, separate estimates of Bmax and Kd were obtained. Bmax varied from 2.3 +/- 0.5 (occipital cortex) to 20.6 +/- 7.3 (cingulate cortex) nM. The average Kd (eight brain regions) was 0.85 +/- 0.17 nM

  13. Radiolabeled high affinity peptidomimetic antagonist selectively targets {alpha}{sub v}{beta}{sub 3} receptor-positive tumor in mice

    Energy Technology Data Exchange (ETDEWEB)

    Jang, Beom-Su [Department of Nuclear Medicine, Warren G. Magnuson Clinical Center, National Institutes of Health, Bethesda, MD 20892 (United States); Lim, Esther [Department of Radiology, Warren G. Magnuson Clinical Center, National Institutes of Health, Bethesda, MD 20892 (United States); Hee Park, Seung [Department of Nuclear Medicine, Warren G. Magnuson Clinical Center, National Institutes of Health, Bethesda, MD 20892 (United States); Shin, In Soo [Department of Nuclear Medicine, Warren G. Magnuson Clinical Center, National Institutes of Health, Bethesda, MD 20892 (United States); Danthi, S. Narasimhan [Department of Radiology, Warren G. Magnuson Clinical Center, National Institutes of Health, Bethesda, MD 20892 (United States); Hwang, In Sook [Department of Nuclear Medicine, Warren G. Magnuson Clinical Center, National Institutes of Health, Bethesda, MD 20892 (United States); Le, Nhat [Department of Nuclear Medicine, Warren G. Magnuson Clinical Center, National Institutes of Health, Bethesda, MD 20892 (United States); Yu, Sarah [Department of Nuclear Medicine, Warren G. Magnuson Clinical Center, National Institutes of Health, Bethesda, MD 20892 (United States); Xie Jianwu [Department of Radiology, Warren G. Magnuson Clinical Center, National Institutes of Health, Bethesda, MD 20892 (United States); Li, King C.P. [Department of Radiology, Warren G. Magnuson Clinical Center, National Institutes of Health, Bethesda, MD 20892 (United States); Carrasquillo, Jorge A. [Department of Nuclear Medicine, Warren G. Magnuson Clinical Center, National Institutes of Health, Bethesda, MD 20892 (United States); Paik, Chang H. [Department of Nuclear Medicine, Warren G. Magnuson Clinical Center, National Institutes of Health, Bethesda, MD 20892 (United States)]. E-mail: cpaik@mail.nih.gov

    2007-05-15

    Objectives: The aim of this research was to synthesize radiolabeled peptidomimetic integrin {alpha}{sub v}{beta}{sub 3} antagonists that selectively target integrin {alpha}{sub v}{beta}{sub 3} receptor and clear rapidly from the whole body. Methods: Integrin {alpha}{sub v}{beta}{sub 3} antagonists, 4-[2-(3,4,5,6-tetrahydropyrimidine-2-ylamino) ethyloxy]benzoyl-2-(S)-aminoethylsulfonyl-amino-{beta}-alanine (IA) and 4-[2-(3,4,5,6-tetrahydro-pyrimidin-2-ylamino)-ethyloxy]benzoyl-2-(S)-[N- (3-amino-neopenta-1-carbamyl)]-aminoethylsulfonylamino-{beta}-alanine hydrochloride (IAC), a hydrophobic carbamate derivative of IA, were conjugated with 2-p-isothiocyanatobenzyl-DOTA at the amino terminus and labeled with {sup 111}In. The {sup 111}In labeled IA and IAC were subjected to in vitro receptor binding, biodistribution and imaging studies using nude mice bearing the receptor-positive M21 human melanoma xenografts. Results: The {sup 111}In-labeled IA (40%) and -IAC (72%) specifically bound in vitro to {alpha}{sub v}{beta}{sub 3} (0.8 {mu}M) at a molar excess. This receptor binding was completely blocked by a molar excess of cold IA to {alpha}{sub v}{beta}{sub 3}. The higher receptor-binding affinity of the {sup 111}In-labeled IAC was reflected in higher tumor uptake and retention: 5.6{+-}1.4 and 4.5{+-}0.7 %ID/g vs. 3.8{+-}0.9 and 2.0{+-}0.3 %ID/g for the {sup 111}In-labeled IA at 0.33 and 2 h. The tumor uptakes were inhibited by the co-injection of 200 {mu}g of IA, indicating that the uptake was receptor mediated. These antagonists were excreted primarily via the renal system. The {sup 111}In activity retained in the whole body was quite comparable between the {sup 111}In-labeled IA (24% ID) and the {sup 111}In-labeled IAC (33% ID) at 2 h. The higher peak tumor uptake and longer retention resulted in higher tumor-to-background ratios for the {sup 111}In-labeled IAC at 2 h with 9.7, 2.3, 0.8, 1.9, 7.1, 2.2, 0.9, 3.7 and 9.9 for blood, liver, kidney, lung, heart, stomach

  14. Localized low-level re-expression of high-affinity mesolimbic nicotinic acetylcholine receptors restores nicotine-induced locomotion but not place conditioning.

    Science.gov (United States)

    Mineur, Y S; Brunzell, D H; Grady, S R; Lindstrom, J M; McIntosh, J M; Marks, M J; King, S L; Picciotto, M R

    2009-04-01

    High-affinity, beta2-subunit-containing (beta2*) nicotinic acetylcholine receptors (nAChRs) are essential for nicotine reinforcement; however, these nAChRs are found on both gamma-aminobutyric acid (GABA) and dopaminergic (DA) neurons in the ventral tegmental area (VTA) and also on terminals of glutamatergic and cholinergic neurons projecting from the pedunculopontine tegmental area and the laterodorsal tegmental nucleus. Thus, systemic nicotine administration stimulates many different neuronal subtypes in various brain nuclei. To identify neurons in which nAChRs must be expressed to mediate effects of systemic nicotine, we investigated responses in mice with low-level, localized expression of beta2* nAChRs in the midbrain/VTA. Nicotine-induced GABA and DA release were partially rescued in striatal synaptosomes from transgenic mice compared with tissue from beta2 knockout mice. Nicotine-induced locomotor activation, but not place preference, was rescued in mice with low-level VTA expression, suggesting that low-level expression of beta2* nAChRs in DA neurons is not sufficient to support nicotine reward. In contrast to control mice, transgenic mice with low-level beta2* nAChR expression in the VTA showed no increase in overall levels of cyclic AMP response element-binding protein (CREB) but did show an increase in CREB phosphorylation in response to exposure to a nicotine-paired chamber. Thus, CREB activation in the absence of regulation of total CREB levels during place preference testing was not sufficient to support nicotine place preference in beta2 trangenic mice. This suggests that partial activation of high-affinity nAChRs in VTA might block the rewarding effects of nicotine, providing a potential mechanism for the ability of nicotinic partial agonists to aid in smoking cessation. PMID:19077117

  15. Cord blood IgE. III. Prediction of IgE high-response and allergy. A follow-up at the age of 18 months

    DEFF Research Database (Denmark)

    Hansen, L G; Høst, A; Halken, S; Holmskov, A; Husby, S; Lassen, L B; Storm, K; Osterballe, O

    E was superior to the originally suggested 0.5 kU/l. Significantly more infants with elevated cord blood IgE had developed specific IgE antibodies at 18 months. The most frequent specific IgE antibody was towards cow's milk. Specific IgE antibodies were very rarely found when total IgE was not elevated...

  16. Synthesis and structure-affinity relationships of novel small molecule natural product derivatives capable of discriminating between serotonin 5-HT1A, 5-HT2A, 5-HT2C receptor subtypes

    OpenAIRE

    Cummings, David F.; Canseco, Diana C.; Sheth, Pratikkumar; Johnson, James E.; Schetz, John A.

    2010-01-01

    Efforts to develop ligands that distinguish between clinically relevant 5-HT2A and 5-HT2C serotonin receptor subtypes have been challenging, because their sequences have high homology. Previous studies reported that a novel aplysinopsin belonging to a chemical class of natural products isolated from a marine sponge was selective for the 5-HT2C over the 5-HT2A receptor subtype. Our goal was to explore the 5-HT2A/2C receptor structure-affinity relationships of derivatives based on the aplysinop...

  17. BODIPY(®) FL histamine as a new modality for quantitative detection of histamine receptor upregulation upon IgE sensitization in murine bone marrow-derived mast cells.

    Science.gov (United States)

    Mirzahosseini, Arash; Kovács, Marianna; Kánai, Károly; Csutora, Péter; Dalmadi, Balázs

    2015-01-01

    Flow cytometry is one of the most widely used methods for the qualitative and quantitative analysis of cell surface expressed proteins by making use of fluorescent specific antibodies. Lacking an antibody validated for flow cytometry, an alternative approach for labeling cell surface receptors is the use of fluorescently tagged ligands. In this study, histamine H4 receptor transfected Chinese hamster ovary cells and murine bone marrow-derived mast cells (mBMMCs) were selected for studying the possibility of staining individual histamine receptors using BODIPY(®) FL histamine and selective antagonists. Flow cytometric measurements and supporting calculations showed that BODIPY FL histamine is suitable tool for quantitating cell surface histamine receptors. The binding, and competitive inhibition of this fluorescent ligand were characterized, which were in good agreement with a semi-empirical model constructed from fundamental protein-binding relationships. Using this method it was shown for the first time that even though mature mBMMCs express H2R and H4R to the same extent, immunoglobulin E sensitization results in H4R upregulation only, while the surface expression of H2R remains unchanged. PMID:25212523

  18. Cellular receptor for 125I-labeled tumor necrosis factor: specific binding, affinity labeling, and relationship to sensitivity

    International Nuclear Information System (INIS)

    Tumor necrosis factor (TNF) is a proteinaceous toxin shed by stimulated myeloid cells. Murine TNF was radioiodinated to a specific activity of 1 mCi/nmol (1 Ci = 37 GBq) of monomer. 125I-labeled TNF (125-TNF) retained complete cytotoxic activity and it was immunochemically identical to the native toxin in a quantitative immunoprecipitation assay. It could be shown by competition binding that 125I-TNF bound to intact L929 cells with a specificity equal to that of native toxin. The conditions of time, temperature, and concentration involved in equilibrium specific binding to intact cells were studied in detail. J774.1 cells, the source of the toxin, demonstrated similar binding but were not sensitive to 125I-TNF cytotoxicity. Normal lymphoid organ cell suspensions and two human tumorigenic cell lines were not sensitive and failed to demonstrate specific binding. 125I-TNF, covalently cross-linked to its receptor on sensitive L-M cells with disuccinimidyl suberate, was isolated and analyzed by sodium dodecyl sulfate/polyacrylamide gel electrophoresis and autoradiography. Two specific bands were identified. The presence of the binding site appears to be necessary but not sufficient to explain the sensitivity of cells to the cytotoxic action of TNF

  19. Syntaxin-4 is essential for IgE secretion by plasma cells

    Energy Technology Data Exchange (ETDEWEB)

    Rahman, Arman; DeCourcey, Joseph; Larbi, Nadia Ben [Immunomodulation Group, School of Biotechnology, Dublin City University (Ireland); Loughran, Sinéad T.; Walls, Dermot [School of Biotechnology and National Centre for Sensor Research, Dublin City University (Ireland); Loscher, Christine E., E-mail: christine.loscher@dcu.ie [Immunomodulation Group, School of Biotechnology, Dublin City University (Ireland)

    2013-10-11

    Highlights: •Knock-down of syntaxin-4 in U266 plasma cells resulted in reduction of IgE secretion. •Knock-down of syntaxin-4 also leads to the accumulation of IgE in the cell. •Immuno-fluorescence staining shows co-localisation of IgE and syntaxin-4 in U266 cells. •Findings suggest a critical requirement for syntaxin-4 in IgE secretion from plasma cells. -- Abstract: The humoral immune system provides a crucial first defense against the invasion of microbial pathogens via the secretion of antigen specific immunoglobulins (Ig). The secretion of Ig is carried out by terminally differentiated B-lymphocytes called plasma cells. Despite the key role of plasma cells in the immune response, the mechanisms by which they constitutively traffic large volumes of Ig out of the cell is poorly understood. The involvement of Soluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE) proteins in the regulation of protein trafficking from cells has been well documented. Syntaxin-4, a member of the Qa SNARE syntaxin family has been implicated in fusion events at the plasma membrane in a number of cells in the immune system. In this work we show that knock-down of syntaxin-4 in the multiple myeloma U266 human plasma cell line results in a loss of IgE secretion and accumulation of IgE within the cells. Furthermore, we show that IgE co-localises with syntaxin-4 in U266 plasma cells suggesting direct involvement in secretion at the plasma membrane. This study demonstrates that syntaxin-4 plays a critical role in the secretion of IgE from plasma cells and sheds some light on the mechanisms by which these cells constitutively traffic vesicles to the surface for secretion. An understanding of this machinery may be beneficial in identifying potential therapeutic targets in multiple myeloma and autoimmune disease where over-production of Ig leads to severe pathology in patients.

  20. Syntaxin-4 is essential for IgE secretion by plasma cells

    International Nuclear Information System (INIS)

    Highlights: •Knock-down of syntaxin-4 in U266 plasma cells resulted in reduction of IgE secretion. •Knock-down of syntaxin-4 also leads to the accumulation of IgE in the cell. •Immuno-fluorescence staining shows co-localisation of IgE and syntaxin-4 in U266 cells. •Findings suggest a critical requirement for syntaxin-4 in IgE secretion from plasma cells. -- Abstract: The humoral immune system provides a crucial first defense against the invasion of microbial pathogens via the secretion of antigen specific immunoglobulins (Ig). The secretion of Ig is carried out by terminally differentiated B-lymphocytes called plasma cells. Despite the key role of plasma cells in the immune response, the mechanisms by which they constitutively traffic large volumes of Ig out of the cell is poorly understood. The involvement of Soluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE) proteins in the regulation of protein trafficking from cells has been well documented. Syntaxin-4, a member of the Qa SNARE syntaxin family has been implicated in fusion events at the plasma membrane in a number of cells in the immune system. In this work we show that knock-down of syntaxin-4 in the multiple myeloma U266 human plasma cell line results in a loss of IgE secretion and accumulation of IgE within the cells. Furthermore, we show that IgE co-localises with syntaxin-4 in U266 plasma cells suggesting direct involvement in secretion at the plasma membrane. This study demonstrates that syntaxin-4 plays a critical role in the secretion of IgE from plasma cells and sheds some light on the mechanisms by which these cells constitutively traffic vesicles to the surface for secretion. An understanding of this machinery may be beneficial in identifying potential therapeutic targets in multiple myeloma and autoimmune disease where over-production of Ig leads to severe pathology in patients

  1. High affinity dopamine D2 receptor radioligands. 2. [125I]epidepride, a potent and specific radioligand for the characterization of striatal and extrastriatal dopamine D2 receptors

    International Nuclear Information System (INIS)

    Epidepride, (S)-N-[(1-ethyl-2-pyrrolidinyl)methyl]-5-iodo-2,3-dimethoxybenzamide, the iodine analogue of isoremoxipride (FLB 457), was found to be a very potent dopamine D2 receptor antagonist. Optimal in vitro binding required incubation at 25C for 4 h at pH 7.4 in a buffer containing 120 mM NaCl, 5 mM KCl, 2 mM CaCl2 and 1 nM MgCl2. Scatchard analysis of in vitro binding to striatal, medical frontal cortical, hippocampal and cerebellar membranes revealed a KD of 24 pM in all regions, with Bmax's of 36.7, 1.04, 0.85, and 0.37 pmol/g tissue, respectively. The Hill coefficients ranged from 0.91-1.00 in all four regions. The IC50's for inhibition of [125I]epidepride binding to striatal, medial frontal cortical, and hippocampal membranes for SCH 23390, SKF 83566, serotonin, ketanserin, mianserin, naloxone, QNB, prasozin, clonidine, alprenolol, and norepinephrine ranged from 1 μM to >10 μM. Partial displacement of [125I]epidepride by nanomolar concentrations of clonidine was noted in the frontal cortex and hippocampus, but not in the striatum. Scatchard analysis of epidepride binding to α2 noradrenergic receptors in the frontal cortex and hippocampus revealed an apparent KD of 9 nM. At an epidepride concentration equal to the KD for the D2 receptor, i.e., 25 pM, no striatal α2 binding was seen and only 7% of the specific epidepride binding in the cortex or hippocampus was due to binding at the α2 site. Correlation of inhibition of [3H]spiperone and [125I]epidepride binding to striatal membranes by a variety of D2 ligands revealed a correlation coefficient of 0.99, indicating that epidepride labels a D2 site

  2. Intermediate- and high-affinity interleukin-2 receptors expressed in an IL-4-dependent T-cell line induce different signals.

    Science.gov (United States)

    Rebollo, A; Silva, A

    1993-01-01

    In order to understand the respective roles of IL-2R alpha and IL-2R beta subunits in transmission of the interleukin-2 (IL-2)-mediated growth signals, we have established two IL-4-dependent murine T-cell clones stably expressing the human IL-2R beta chain and three clones stably expressing the human IL-2R alpha chain. Whereas parental LD8 cells (which express only the murine IL-2R beta chain) do not proliferate in response to IL-2, cell lines stably expressing human IL-2R beta or the chimeric IL-2R alpha beta complex proliferate in response to IL-2. Stably transfected cells expressing the chimeric high-affinity receptor (human IL-2R alpha and murine IL-2R beta) expressed de novo endogenous murine IL-2R alpha when cultured in the presence of IL-2 but not IL-4. Both chimeric and endogenous receptors are functional in response to IL-2, since only addition of both anti-human and anti-murine IL-2R alpha monoclonal antibodies (mAb) inhibited IL-2-induced proliferation. Taken together, our results strongly suggest that human and murine IL-2R beta molecules are different since interaction of IL-2 with human p70 IL-2R is sufficient for transduction of proliferative signals in the absence of p55 IL-2R or, alternatively, that over-expression of the IL-2R beta chain renders cells responsive to IL-2. In addition, IL-2 stimulation of T cells through different forms of IL-2R results in the induction of distinct cellular responses. PMID:8262551

  3. Quantitative Analysis of Human Neonatal Fc Receptor (FcRn) Tissue Expression in Transgenic Mice by Online Peptide Immuno-Affinity LC-HRMS.

    Science.gov (United States)

    Fan, Yao-Yun; Neubert, Hendrik

    2016-04-19

    Neonatal Fc receptor (FcRn) is the homeostatic receptor responsible for the long half-life of endogenous IgG by protecting it from lysosomal degradation. Understanding systemic FcRn tissue expression is important to predict and design the half-life of therapeutic antibodies and Fc-coupled biotherapeutics. To this end, we measured human FcRn (hFcRn) tissue expression in Tg32, a human FcRn knock-in transgenic mouse model, for which a strong correlation of drug clearance to humans has been demonstrated. Building an hFcRn tissue expression profile in Tg32 was enabled by the development of a tissue preparation procedure composed of bead-based protein extraction and protein precipitation using acetone followed by pellet digestion with trypsin. Digests were then loaded onto an online peptide immuno-affinity flow configuration hyphenated with reversed phase nanoflow chromatography and coupled with high resolution mass spectrometry to quantify hFcRn derived peptides. The workflow allowed bypassing some of the challenges typically associated with membrane protein analysis. We demonstrated acceptable precision and bias for measuring hFcRn in tissue matrices, typically within 20% coefficient of variation and relative error. We also report hFcRn expression in several Tg32 tissues. We anticipate that establishing a quantitative approach for hFcRn in tissues will enable the systematic measurement of hFcRn concentrations to further increase the accuracy of physiologically based pharmacokinetic (PBPK) models for PK prediction of Fc-containing biotherapeutics. This is anticipated to improve the translation of pharmacokinetic data from preclinical model systems to humans. PMID:27012525

  4. OX48, a monoclonal antibody against a 70,000 MW rat activation antigen expressed by T cells bearing the high-affinity interleukin-2 receptor.

    Science.gov (United States)

    Somoza, C; Fernández-Ruiz, E; Rebollo, A; Sanz, E; Ramírez, F; Silva, A

    1990-01-01

    The monoclonal antibody (mAb) OX48 recognizes a 70,000 MW cell-surface protein present in a small percentage of activated rat T cells and in CD8+ rat x BW5147 interleukin-2 (IL-2)-dependent T-cell hybridomas, but not in resting spleen cells or in IL-2-independent T-cell hybrids. OX48 antibody added simultaneously with concanavalin A (Con A) to resting spleen cells inhibits the cell proliferation and reduces the IL-2 production. However, addition of IL-2 does not restore the mitogenic response. Growth of rat blast T cells or IL-2-dependent hybrids is not affected by the OX48 antibody. There is a close correlation between the expression of high-affinity IL-2 receptors (IL-2R) and the OX48 antigen in T-cell hybridomas. In spite of this striking correlation, OX48 mAb does not inhibit the binding of 125I-IL-2 to the IL-2-dependent hybrids, and is unable to immunoprecipitate any of the proteins chemically cross-linked to 125I-IL-2. Therefore, the OX48 molecule represents a new rat activation antigen, undefined in other species, and probably involved in the early steps of T-cell activation. Images Figure 5 Figure 7 PMID:2373518

  5. Expression of low-, intermediate-, and high-affinity IL-2 receptors on B cell lines derived from patients with undifferentiated lymphoma of Burkitt's and non-Burkitt's types

    Energy Technology Data Exchange (ETDEWEB)

    Benjamin, D.; Rosolen, A.; Wormsley, S.B.; DeBault, L.E.; Colamonici, O.R. (Saint Francis Research Institute, Oklahoma City, OK (USA))

    1990-08-01

    IL-2 receptors on T cells exist in at least three forms which differ in their ligand-binding affinity. The low-affinity IL-2 receptor (IL-2R) consists of the 55-kDa Tac protein (p55 alpha), the intermediate-affinity site corresponds to the 70-kDa molecule (p70 beta), and the high-affinity IL-2R consists of a noncovalent heterodimeric structure involving both p55 alpha and p70 beta. We studied 24 B cell lines (8 EBV-negative and 16 EBV-positive) for IL-2R expression in the presence or absence of the tumor promoter, teleocidin. 125I-IL-2 radioreceptor binding assays and crosslinking studies demonstrated the sole expression of p55 alpha in EBV-negative cell lines only, whereas p55 alpha present in EBV-positive cell lines was always associated with p70 beta to construct high-affinity IL-2R. p70 beta was not detected in any of the EBV-negative cell lines, but was expressed on most of the EBV-positive cell lines (13 of 16). Our data also indicate that the expression of p55 alpha and p70 beta by radiolabeling correlates with their expression in flow cytometry, and that a large excess of p55 alpha is required to construct high-affinity IL-2R. Coexpression of p55 alpha and p70 beta on human B cells contributed to constructing high-affinity IL-2R hybrid complex as shown by rapid association rate contributed by p55 alpha and slow dissociation rate by p70 beta; teleocidin's ability to induce p55 alpha on cell lines which express p70 beta only, resulting in appearance of high-affinity IL-2R; and blocking p55 alpha by anti-Tac mAb in cell lines which constitutively express high-affinity IL-2R eliminated both high- and low-affinity components. The existence of low, intermediate, and high IL-2R on human B cells bears important future implications for understanding the mechanism of IL-2 signaling and the role of IL-2 in B cell activation, proliferation, and differentiation.

  6. Site-specific conjugation of a lanthanide chelator and its effects on the chemical synthesis and receptor binding affinity of human relaxin-2 hormone

    International Nuclear Information System (INIS)

    Highlights: ► A mono-Eu-DTPA conjugated peptide ligand, Eu-DTPA-(A)-H2, has been developed. ► The choice of a site for incorporation of a chelator is critical. ► The labeled peptide retains full activity at the RXFP1 receptor. ► It is markedly cheaper to produce and easier to use than radioactive probes. -- Abstract: Diethylenetriamine pentaacetic acid (DTPA) is a popular chelator agent for enabling the labeling of peptides for their use in structure–activity relationship study and biodistribution analysis. Solid phase peptide synthesis was employed to couple this commercially available chelator at the N-terminus of either the A-chain or B-chain of H2 relaxin. The coupling of the DTPA chelator at the N-terminus of the B-chain and subsequent loading of a lanthanide (europium) ion into the chelator led to a labeled peptide (Eu-DTPA-(B)-H2) in low yield and having very poor water solubility. On the other hand, coupling of the DTPA and loading of Eu at the N-terminus of the A-chain led to a water-soluble peptide (Eu-DTPA-(A)-H2) with a significantly improved final yield. The conjugation of the DTPA chelator at the N-terminus of the A-chain did not have any impact on the secondary structure of the peptide determined by circular dichroism spectroscopy (CD). On the other hand, it was not possible to determine the secondary structure of Eu-DTPA-(B)-H2 because of its insolubility in phosphate buffer. The B-chain labeled peptide Eu-DTPA-(B)-H2 required solubilization in DMSO prior to carrying out binding assays, and showed lower affinity for binding to H2 relaxin receptor, RXFP1, compared to the water-soluble A-chain labeled peptide Eu-DTPA-(A)-H2. The mono-Eu-DTPA labeled A-chain peptide, Eu-DTPA-(A)-H2, thus can be used as a valuable probe to study ligand–receptor interactions of therapeutically important H2 relaxin analogs. Our results show that it is critical to choose an approriate site for incorporating chelators such as DTPA. Otherwise, the bulky size of

  7. Site-specific conjugation of a lanthanide chelator and its effects on the chemical synthesis and receptor binding affinity of human relaxin-2 hormone

    Energy Technology Data Exchange (ETDEWEB)

    Shabanpoor, Fazel [Florey Neuroscience Institutes, The University of Melbourne, Parkville, VIC 3010 (Australia); School of Chemistry, The University of Melbourne, Parkville, VIC 3010 (Australia); Bathgate, Ross A.D.; Belgi, Alessia [Florey Neuroscience Institutes, The University of Melbourne, Parkville, VIC 3010 (Australia); Department of Biochemistry and Molecular Biology, The University of Melbourne, Parkville, VIC 3010 (Australia); Chan, Linda J.; Nair, Vinojini B.; Wade, John D. [Florey Neuroscience Institutes, The University of Melbourne, Parkville, VIC 3010 (Australia); School of Chemistry, The University of Melbourne, Parkville, VIC 3010 (Australia); Hossain, Mohammed Akhter, E-mail: akhter.hossain@florey.edu.au [Florey Neuroscience Institutes, The University of Melbourne, Parkville, VIC 3010 (Australia); School of Chemistry, The University of Melbourne, Parkville, VIC 3010 (Australia)

    2012-04-06

    Highlights: Black-Right-Pointing-Pointer A mono-Eu-DTPA conjugated peptide ligand, Eu-DTPA-(A)-H2, has been developed. Black-Right-Pointing-Pointer The choice of a site for incorporation of a chelator is critical. Black-Right-Pointing-Pointer The labeled peptide retains full activity at the RXFP1 receptor. Black-Right-Pointing-Pointer It is markedly cheaper to produce and easier to use than radioactive probes. -- Abstract: Diethylenetriamine pentaacetic acid (DTPA) is a popular chelator agent for enabling the labeling of peptides for their use in structure-activity relationship study and biodistribution analysis. Solid phase peptide synthesis was employed to couple this commercially available chelator at the N-terminus of either the A-chain or B-chain of H2 relaxin. The coupling of the DTPA chelator at the N-terminus of the B-chain and subsequent loading of a lanthanide (europium) ion into the chelator led to a labeled peptide (Eu-DTPA-(B)-H2) in low yield and having very poor water solubility. On the other hand, coupling of the DTPA and loading of Eu at the N-terminus of the A-chain led to a water-soluble peptide (Eu-DTPA-(A)-H2) with a significantly improved final yield. The conjugation of the DTPA chelator at the N-terminus of the A-chain did not have any impact on the secondary structure of the peptide determined by circular dichroism spectroscopy (CD). On the other hand, it was not possible to determine the secondary structure of Eu-DTPA-(B)-H2 because of its insolubility in phosphate buffer. The B-chain labeled peptide Eu-DTPA-(B)-H2 required solubilization in DMSO prior to carrying out binding assays, and showed lower affinity for binding to H2 relaxin receptor, RXFP1, compared to the water-soluble A-chain labeled peptide Eu-DTPA-(A)-H2. The mono-Eu-DTPA labeled A-chain peptide, Eu-DTPA-(A)-H2, thus can be used as a valuable probe to study ligand-receptor interactions of therapeutically important H2 relaxin analogs. Our results show that it is critical to

  8. In Silico Prediction of Estrogen Receptor Subtype Binding Affinity and Selectivity Using Statistical Methods and Molecular Docking with 2-Arylnaphthalenes and 2-Arylquinolines

    Directory of Open Access Journals (Sweden)

    Yonghua Wang

    2010-09-01

    Full Text Available Over the years development of selective estrogen receptor (ER ligands has been of great concern to researchers involved in the chemistry and pharmacology of anticancer drugs, resulting in numerous synthesized selective ER subtype inhibitors. In this work, a data set of 82 ER ligands with ERα and ERβ inhibitory activities was built, and quantitative structure-activity relationship (QSAR methods based on the two linear (multiple linear regression, MLR, partial least squares regression, PLSR and a nonlinear statistical method (Bayesian regularized neural network, BRNN were applied to investigate the potential relationship of molecular structural features related to the activity and selectivity of these ligands. For ERα and ERβ, the performances of the MLR and PLSR models are superior to the BRNN model, giving more reasonable statistical properties (ERα: for MLR, Rtr2 = 0.72, Qte2 = 0.63; for PLSR, Rtr2 = 0.92, Qte2 = 0.84. ERβ: for MLR, Rtr2 = 0.75, Qte2 = 0.75; for PLSR, Rtr2 = 0.98, Qte2 = 0.80. The MLR method is also more powerful than other two methods for generating the subtype selectivity models, resulting in Rtr2 = 0.74 and Qte2 = 0.80. In addition, the molecular docking method was also used to explore the possible binding modes of the ligands and a relationship between the 3D-binding modes and the 2D-molecular structural features of ligands was further explored. The results show that the binding affinity strength for both ERα and ERβ is more correlated with the atom fragment type, polarity, electronegativites and hydrophobicity. The substitutent in position 8 of the naphthalene or the quinoline plane and the space orientation of these two planes contribute the most to the subtype selectivity on the basis of similar hydrogen bond interactions between binding ligands and both ER subtypes. The QSAR models built together with the docking procedure should be of great advantage for screening and designing ER ligands with improved affinity

  9. Interactions of dopaminergic agonists and antagonists with dopaminergic D3 binding sites in rat striatum. Evidence that (/sup 3/H)dopamine can label a high affinity agonist-binding state of the D1 dopamine receptor

    Energy Technology Data Exchange (ETDEWEB)

    Leff, S.E.; Creese, I.

    1985-02-01

    The interactions of dopaminergic agonists and antagonists with /sup 3/H-agonist labeled D3 dopaminergic binding sites of rat striatum have been characterized by radioligand-binding techniques. When the binding of (/sup 3/H)dopamine and (/sup 3/H)apomorphine to D2 dopamine receptors is blocked by the inclusion of D2 selective concentrations of unlabeled spiroperidol or domperidone, these ligands appear to label selectively the previously termed D3 binding site. Antagonist/(/sup 3/H)dopamine competition curves are of uniformly steep slope (nH . 1.0), suggesting the presence of a single D3 binding site. The relative potencies of antagonists to inhibit D3 specific (/sup 3/H)dopamine binding are significantly correlated with their potencies to block D1 dopamine receptors as measured by the inhibition of both dopamine-stimulated adenylate cyclase and (/sup 3/H)flupentixol-binding activities. The affinities of agonists to inhibit D3 specific (/sup 3/H)dopamine binding are also correlated with estimates of these agonists affinities for the high affinity binding component of agonist/(/sup 3/H)flupentixol competition curves. Both D3 specific (/sup 3/H) dopamine binding and the high affinity agonist-binding component of dopamine/(/sup 3/H)flupentixol competition curves show a similar sensitivity to guanine nucleotides. Taken together, these data strongly suggest that the D3 binding site is related to a high affinity agonist-binding state of the D1 dopamine receptor.

  10. Development of p-benzoylbenzoylated [N,C,rANP(1-28)]pBNP32 (pBNP1) derivatives and affinity photolabeling of the bovine NPR-A receptor.

    Science.gov (United States)

    Coupal, M; De Léan, A; McNicoll, N; Fournier, A

    1999-04-29

    Two Nalpha-benzophenone-substituted photoprobes, derived from the high affinity NPR-A chimeric agonist [N, C, rANP(1-28)]pBNP32 (pBNP1) were assembled by solid-phase peptide synthesis. [Nalpha-p-benzoylbenzoyl, Tyr2]pBNP1 (probe A), and [Nalpha-p-benzoylbenzoyl, Tyr18]pBNP1 (probe B) were synthesized and their affinity was tested on bovine zona glomerulosa membrane preparations. Both were found to exert ANP-type high affinities (Kd = 20 pM) with Kd of 10 pM and 30 pM for probe A, and probe B, respectively. Photolabeling of NPR-A with both analogs cross-linked specifically the 130 kDa monomeric NPR-A. The maximal irreversible ligand incorporations were estimated at 18% and 41% for probe A, and probe B, respectively. These results show that the N-terminus of the chimeric compound can be acylated with a large chemical function, such as the benzophenone moiety, without loosing its affinity for the NPR-A receptor. Furthermore, Leu2 or Leu18 can be substituted with tyrosine without disturbing the binding capacity of the ligand. Finally, it appears that the pBNP1 N-terminus is close to the receptor structure as irreversible incorporation is observed after photolabeling. PMID:10222239

  11. Helminth Allergens, Parasite-Specific IgE, and Its Protective Role in Human Immunity.

    Science.gov (United States)

    Fitzsimmons, Colin Matthew; Falcone, Franco Harald; Dunne, David William

    2014-01-01

    The Th2 immune response, culminating in eosinophilia and IgE production, is not only characteristic of allergy but also of infection by parasitic worms (helminths). Anti-parasite IgE has been associated with immunity against a range of helminth infections and many believe that IgE and its receptors evolved to help counter metazoan parasites. Allergens (IgE-antigens) are present in only a small minority of protein families and known IgE targets in helminths belong to these same families (e.g., EF-hand proteins, tropomyosin, and PR-1 proteins). During some helminth infection, especially with the well adapted hookworm, the Th2 response is moderated by parasite-expressed molecules. This has been associated with reduced allergy in helminth endemic areas and worm infection or products have been proposed as treatments for allergic conditions. However, some infections (especially Ascaris) are associated with increased allergy and this has been linked to cross-reactivity between worm proteins (e.g., tropomyosins) and highly similar molecules in dust-mites and insects. The overlap between allergy and helminth infection is best illustrated in Anisakis simplex, a nematode that when consumed in under-cooked fish can be both an infective helminth and a food allergen. Nearly 20 molecular allergens have been isolated from this species, including tropomyosin (Ani s 3) and the EF-hand protein, Ani s troponin. In this review, we highlight aspects of the biology and biochemistry of helminths that may have influenced the evolution of the IgE response. We compare dominant IgE-antigens in worms with clinically important environmental allergens and suggest that arrays of such molecules will provide important information on anti-worm immunity as well as allergy. PMID:24592267

  12. Local IgE production in nonatopic nasal polyposis.

    LENUS (Irish Health Repository)

    Sheahan, Patrick

    2012-02-01

    INTRODUCTION: Chronic rhinosinusitis with nasal polyposis (CRSwNP) represents an eosinophilic T-helper 2 inflammatory response. Local production of IgE within nasal polyps (NPs) has been demonstrated, suggesting a role for local IgE in the pathogenesis of NP in atopic CRS patients. We hypothesized that local IgE specific to inhalant allergens may also play a role in the genesis of NP in nonatopic CRS patients. METHODS: Sinus and inferior turbinate tissue was obtained from nonatopic CRSwNP patients (n = 7), chronic rhinosinusitis without nasal polyps (CRSsNP) patients (n = 15), and healthy controls (n = 9) at the time of surgery. ImmunoCAP analysis (Phadia AB, Portage, MI) for 14 common inhalant antigens was performed on tissue homogenates to determine the antigen-specific response. RESULTS: Total IgE levels did not differ in sinus or turbinate tissue between CRSwNP, CRSsNP, or control patients. CRSwNP sinus tissue had higher levels of specific IgE for cockroach and plantain (p = .03) than other groups and elevated Alternaria IgE levels when compared with CRSsNP sinus tissue (p < .05). No significant differences were found for any of the other antigen-specific IgE levels. Fifty-seven percent of CRSwNP polyps demonstrated a polyclonal IgE response, whereas the other 43% had no demonstrable antigen-specific IgE. In contrast, only 17% of CRSsNP patients demonstrated a polyclonal response within sinus tissue, whereas 67% had no detectable antigen-specific IgE. There was no significant difference in levels of IgE in inferior turbinate tissue between the groups (p > .05). CONCLUSIONS: Localized mucosal IgE specific to common inhalant allergens appears to play a role in a subset of CRSwNP patients without evidence of systemic atopy.

  13. Further characterization of a high affinity thyrotropin binding site on the rat thyrotropin receptor which is an epitope for blocking antibodies from idiopathic myxedema patients but not thyroid stimulating antibodies from Graves' patients.

    Science.gov (United States)

    Kosugi, S; Ban, T; Akamizu, T; Kohn, L D

    1991-10-31

    Cysteine 390 of the rat thyrotropin (TSH) receptor, when mutated to serine, results in a receptor with a reduced ability of TSH to bind and increase cAMP levels but a preserved ability of thyroid stimulating autoantibodies (TSAbs) from hyperthyroid Graves' patients to increase cAMP levels. The ability of receptor autoantibodies from hypothyroid patients with idiopathic myxedema to inhibit the TSAb activity which is preserved is, however, like TSH binding, significantly reduced. Cysteine 390, together with tyrosine 385, thus appears to be an important determinant in a high affinity TSH binding site which is an epitope for receptor autoantibodies which block TSH or TSAb action and cause hypothyroidism rather than TSAbs which increase cAMP levels and are associated with hyperthyroidism. Threonine 388 and aspartic acid 403 may contribute to this ligand interaction site. PMID:1719963

  14. A Theoretical Study of the Relationships between Electronic Structure and CB1 and CB2 Cannabinoid Receptor Binding Affinity in a Group of 1-Aryl-5-(1-H-pyrrol-1-yl-1-H-pyrazole-3-carboxamides

    Directory of Open Access Journals (Sweden)

    Francisco Salgado-Valdés

    2014-01-01

    Full Text Available We report the results of a search for model-based relationships between hCB1 and hCB2 receptor binding affinity and molecular structure for a group of 1-aryl-5-(1-H-pyrrol-1-yl-1-H-pyrazole-3-carboxamides. The wave functions and local atomic reactivity indices were obtained at the B3LYP/6-31G(d,p levels of theory with full geometry optimization. Interaction pharmacophores were generated for both receptors. The main conclusions of this work are as follows. (1 We obtained statistically significant equations relating the variation of hCB1 and hCB2 receptor binding affinities with the variation of definite sets of local atomic reactivity indices. (2 The interaction of the molecules with the hCB1 and hCB2 receptors seems to be highly complex and mainly orbital controlled. (3 The interaction mechanisms seem to be different for each type of receptor. This study, contrarily to the statistically backed ones, is able to provide a microscopic insight of the mechanisms involved in the binding process.

  15. Pulmonary Administration of GW0742, a High-Affinity Peroxisome Proliferator-Activated Receptor Agonist, Repairs Collapsed Alveoli in an Elastase-Induced Mouse Model of Emphysema.

    Science.gov (United States)

    Ozawa, Chihiro; Horiguchi, Michiko; Akita, Tomomi; Oiso, Yuki; Abe, Kaori; Motomura, Tomoki; Yamashita, Chikamasa

    2016-01-01

    Pulmonary emphysema is a disease in which lung alveoli are irreversibly damaged, thus compromising lung function. Our previous study revealed that all-trans-retinoic acid (ATRA) induces the differentiation of human lung alveolar epithelial type 2 progenitor cells and repairs the alveoli of emphysema model mice. ATRA also reportedly has the ability to activate peroxisome proliferator-activated receptor (PPAR) β/δ. A selective PPARβ/δ ligand has been reported to induce the differentiation of human keratinocytes during wound repair. Here, we demonstrate that treatment using a high-affinity PPARβ/δ agonist, GW0742, reverses the lung tissue damage induced by elastase in emphysema-model mice and improves respiratory function. Mice treated with elastase, which collapsed their alveoli, were then treated with either 10% dimethyl sulfoxide (DMSO) in saline (control group) or GW0742 (1.0 mg/kg twice a week) by pulmonary administration. Treatment with GW0742 for 2 weeks increased the in vivo expression of surfactant proteins A and D, which are known alveolar type II epithelial cell markers. GW0742 treatment also shortened the average distance between alveolar walls in the lungs of emphysema model mice, compared with a control group treated with 10% DMSO in saline. Treatment with GW0742 for 3 weeks also improved tissue elastance (cm H2O/mL), as well as the ratio of the forced expiratory volume in the first 0.05 s to the forced vital capacity (FEV 0.05/FVC). In each of these experiments, GW0742 treatment reversed the damage caused by elastase. In conclusion, PPARβ/δ agonists are potential therapeutic agents for pulmonary emphysema. PMID:27150147

  16. Structure-Guided Design of a High-Affinity Platelet Integrin αIIbβ3 Receptor Antagonist That Disrupts Mg2+ Binding to the MIDAS

    Science.gov (United States)

    Zhu, Jieqing; Choi, Won-Seok; McCoy, Joshua G.; Negri, Ana; Zhu, Jianghai; Naini, Sarasija; Li, Jihong; Shen, Min; Huang, Wenwei; Bougie, Daniel; Rasmussen, Mark; Aster, Richard; Thomas, Craig J.; Filizola, Marta; Springer, Timothy A.; Coller, Barry S.

    2012-01-01

    An integrin found on platelets, αIIbβ3 mediates platelet aggregation, and αIIbβ3 antagonists are effective antithrombotic agents in the clinic. Ligands bind to integrins in part by coordinating a magnesium ion (Mg2+) located in the β subunit metal ion–dependent adhesion site (MIDAS). Drugs patterned on the integrin ligand sequence Arg-Gly-Asp have a basic moiety that binds the αIIb subunit and a carboxyl group that coordinates the MIDAS Mg2+ in the β3 subunits. They induce conformational changes in the β3 subunit that may have negative consequences such as exposing previously hidden epitopes and inducing the active conformation of the receptor. We recently reported an inhibitor of αIIbβ3 (RUC-1) that binds exclusively to the αIIb subunit; here, we report the structure-based design and synthesis of RUC-2, a RUC-1 derivative with a ~100-fold higher affinity. RUC-2 does not induce major conformational changes in β3 as judged by monoclonal antibody binding, light scattering, gel chromatography, electron microscopy, and a receptor priming assay. X-ray crystallography of the RUC-2–IIbβ3 headpiece complex in 1 mM calcium ion (Ca2+)/5 mM Mg2+ at 2.6 Å revealed that RUC-2 binds to αIIb the way RUC-1 does, but in addition, it binds to the β3 MIDAS residue glutamic acid 220, thus displacing Mg2+ from the MIDAS. When the Mg2+ concentration was increased to 20 mM, however, Mg2+ was identified in the MIDAS and RUC-2 was absent. RUC-2′s ability to inhibit ligand binding and platelet aggregation was diminished by increasing the Mg2+ concentration. Thus, RUC-2 inhibits ligand binding by a mechanism different from that of all other αIIbβ3 antagonists and may offer advantages as a therapeutic agent. PMID:22422993

  17. Site-specific conjugation of a lanthanide chelator and its effects on the chemical synthesis and receptor binding affinity of human relaxin-2 hormone.

    Science.gov (United States)

    Shabanpoor, Fazel; Bathgate, Ross A D; Belgi, Alessia; Chan, Linda J; Nair, Vinojini B; Wade, John D; Hossain, Mohammed Akhter

    2012-04-01

    Diethylenetriamine pentaacetic acid (DTPA) is a popular chelator agent for enabling the labeling of peptides for their use in structure-activity relationship study and biodistribution analysis. Solid phase peptide synthesis was employed to couple this commercially available chelator at the N-terminus of either the A-chain or B-chain of H2 relaxin. The coupling of the DTPA chelator at the N-terminus of the B-chain and subsequent loading of a lanthanide (europium) ion into the chelator led to a labeled peptide (Eu-DTPA-(B)-H2) in low yield and having very poor water solubility. On the other hand, coupling of the DTPA and loading of Eu at the N-terminus of the A-chain led to a water-soluble peptide (Eu-DTPA-(A)-H2) with a significantly improved final yield. The conjugation of the DTPA chelator at the N-terminus of the A-chain did not have any impact on the secondary structure of the peptide determined by circular dichroism spectroscopy (CD). On the other hand, it was not possible to determine the secondary structure of Eu-DTPA-(B)-H2 because of its insolubility in phosphate buffer. The B-chain labeled peptide Eu-DTPA-(B)-H2 required solubilization in DMSO prior to carrying out binding assays, and showed lower affinity for binding to H2 relaxin receptor, RXFP1, compared to the water-soluble A-chain labeled peptide Eu-DTPA-(A)-H2. The mono-Eu-DTPA labeled A-chain peptide, Eu-DTPA-(A)-H2, thus can be used as a valuable probe to study ligand-receptor interactions of therapeutically important H2 relaxin analogs. Our results show that it is critical to choose an approriate site for incorporating chelators such as DTPA. Otherwise, the bulky size of the chelator, depending on the site of incorporation, can affect yield, solubility, structure and pharmacological profile of the peptide. PMID:22425984

  18. Providing affinity

    DEFF Research Database (Denmark)

    Guglielmi, Michel; Johannesen, Hl

    , Essex, Hertfordshire, Norfolk and Suffolk. Research found that there was a lack of identity or sense of belonging and nothing anchoring people to the region as a whole. Common affinity is somehow forced to the people of East England and thereby we came to the conclusion that a single landmark or a...... a sense of belonging to people sharing deterritorialized synchronic experiences. But at the same time, the immersion experience is highly low tech and desperately analog, mainly based on fabulation, cartoons, and mushrooms growing in local forests. It ultimately appeals to the experienced sense of...

  19. IgA is a more potent inducer of NADPH oxidase activation and degranulation in blood eosinophils than IgE.

    Science.gov (United States)

    Pleass, Richard J; Lang, Mark L; Kerr, Michael A; Woof, Jenny M

    2007-02-01

    Human eosinophils can mediate both beneficial and detrimental responses in parasitic and allergic diseases. Binding of aggregated immunoglobulin to Fc receptors on eosinophils mediates important defence processes, including generation of activated oxygen species resulting from NADPH oxidase activation, and eosinophil peroxidase release following degranulation. The abilities of a matched set of IgA, IgG and IgE antibodies to elicit such responses in blood-derived eosinophils were compared using a chemiluminescence assay. IgA and IgG, but not IgE, were found to trigger NADPH oxidase activation and degranulation in eosinophils. This non-responsiveness to IgE did not result from receptor blockade by endogenous IgE since no blood-derived IgE was detectable on freshly isolated eosinophils. Moreover, while cross-linking of FcalphaRI by specific mAbs triggered NADPH oxidase activation and degranulation in blood-derived eosinophils, equivalent cross-linking of FcvarepsilonRI or FcvarepsilonRII did not elicit such responses. Therefore IgA is more potent at eliciting activated oxygen species release and degranulation in eosinophils than IgE, suggesting that the importance of IgA in eosinophil activation in immune defence and allergy may have been underestimated. PMID:16777227

  20. IgE versus IgG4 epitopes of the peanut allergen Ara h 1 in patients with severe allergy

    DEFF Research Database (Denmark)

    Bøgh, Katrine Lindholm; Nielsen, H.; Eiwegger, T.;

    2014-01-01

    : This study demonstrated the usefulness of the phage-display technology in distinguishing between the epitope pattern of IgE and IgG4, giving detailed information on fine specificity and affinity. Competitive immuno-screening of phage-display random peptide libraries could be a future valuable tool to...... study the balance and dynamics of the IgE and IgG4 epitope recognition repertoire and provide a diagnostic tool giving information on the associated allergic phenotype. (C) 2013 Elsevier Ltd. All rights reserved....

  1. Triggers of IgE class switching and allergy development

    DEFF Research Database (Denmark)

    Poulsen, Lars K.; Hummelshoj, Lone

    2007-01-01

    The prevalence of immunoglobulin E (IgE)-mediated allergic diseases has been increasing for the last four decades. In this review determinants for an increased IgE synthesis are discussed on both an epidemiological and on an immunological level with special emphasis on the differentiation of the ...... the need for more knowledge on preventable causes of IgE- and allergy development....

  2. Synthesis, in vitro validation and in vivo pharmacokinetics of [125I]N-[2-(4-iodophenyl)ethyl]-N-methyl-2-(1-piperidinyl) ethylamine: A high-affinity ligand for imaging sigma receptor positive tumors

    International Nuclear Information System (INIS)

    N-[2-(4-iodophenyl)ethyl]-N-methyl-2-(1-piperidinyl)ethylamine, IPEMP, and the corresponding bromo derivative, BrPEMP, have been synthesized and characterized. Both BrPEMP and IPEMP were evaluated for sigma-1 and sigma-2 subtype receptor affinities and found to possess very high affinities for both receptor subtypes. The precursor for radioiodination n-tributylstannylphenylethylpiperidinylethylamine was prepared from its bromo derivative by palladium-catalyzed stannylation reaction. Radioiodinated 4-[125I]PEMP was readily prepared in high yields and high specific activity by oxidative iododestannylation reaction using chloramine-T as oxidizing agent. Sites labeled by 4-[125I]PEMP in guinea pig brain membranes showed high affinity for BD1008, haloperidol, and (+)-pentazocine (Ki = 5.06 ± 0.40, 32.6 ± 2.75, and 48.1 ± 8.60 nM, respectively), which is consistent with sigma receptor pharmacology. Competition binding studies of 4-[125I]PEMP in melanoma (A375) and MCF-7 breast cancer cells showed a high affinity, dose-dependent inhibition of binding with known sigma ligand N-[2-(3,4-dichlorophenyl)ethyl]-N-methyl-2-(1-pyrrolidinyl) ethylamine, BD1008 (Ki = 5, 11 nM, respectively), supporting the labeling of sigma sites in these cells. Haloperidol, however showed a weaker (Ki 100-200 nM) affinity for the sites labeled by 4-[125I]PEMP in these cells. Biodistribution studies of 4-[125I]PEMP in rats showed a fast clearance of this radiopharmaceutical from blood, liver, lung, and other organs. A co-injection of 4-IPEMP with 4-[125I]PEMP resulted in 37%, 69%, and 35% decrease in activity in liver, kidney, and brain (organs possessing sigma receptors), respectively at 1-h postinjection. These results suggest that 4-[125I]PEMP is a promising radiopharmaceutical for pursuing further studies in animal models with tumors

  3. Serum IgE levels in patients with intracranial tumors

    Directory of Open Access Journals (Sweden)

    George A Alexiou

    2015-03-01

    Full Text Available Aim: Several epidemiological studies have shown an inverse correlation between allergy and brain cancer. The purpose of this study was to compare the serum IgE levels between patients with gliomas and nonglial tumors and their possible prognostic role. Methods: A total of 84 patients with intracranial tumors were included in this study. At clinical presentation, estimation of serum IgE levels was assessed by nephelometry. Detailed information regarding the history of allergies was collected by interview. Results: Of the 84 cases, 42 were gliomas, 23 were meningiomas, 16 were metastases and 3 were primary central nervous system lymphomas. Patients with high-grade glioma had lower IgE levels than patients with low-grade glioma. Patients with glioma and meningioma had statistical significant lower serum IgE levels than patients with metastases. Patients with glioblastoma with serum IgE levels greater than 24 U/mL had a better survival. Conclusion: Patients with glioma and meningioma had lower IgE levels than patients with metastatic lesions. A prognostic role of serum IgE levels was found in glioblastoma. Further studies in larger patient series are required in order to verify our preliminary observations.

  4. Characterization of the binding of a novel nonxanthine adenosine antagonist radioligand, ( sup 3 H)CGS 15943, to multiple affinity states of the adenosine A1 receptor in the rat cortex

    Energy Technology Data Exchange (ETDEWEB)

    Jarvis, M.F.; Williams, M.; Do, U.H.; Sills, M.A. (CIBA-GEIGY Corp., Summit, NJ (USA))

    1991-01-01

    The triazoloquinazoline CGS 15943 is the first reported nonxanthine adenosine antagonist that has high affinity for brain adenosine receptors. In the present study, the binding of (3H) CGS 15943 to recognition sites in rat cortical membranes was characterized. Saturation experiments revealed that (3H)CGS 15943 labeled a single class of recognition sites with high affinity and limited capacity. Competition studies revealed that the binding of (3H)CGS 15943 was consistent with the labeling of brain adenosine A1 receptors. Adenosine agonists inhibited 1 nM (3H)CGS 15943 binding with the following order of activity N6-cyclopentyladenosine (IC50 = 15 nM) greater than 2-chloroadenosine greater than (R)-N6-phenylisopropyladenosine greater than 5'-N6-ethylcarboxamidoadenosine greater than (S)N6-phenylisopropyladenosine greater than CGS 21680 greater than CV 1808 (IC50 greater than 10,000 nM). The potency order for adenosine antagonists was CGS 15943 (IC50 = 5 nM) greater than 8-phenyltheophylline greater than 1,3-dipropyl-8-(4-amino-2-chloro)phenylxanthine greater than 1,3-diethyl-8-phenylxanthine greater than theophylline = caffeine (IC50 greater than 10,000 nM). Antagonist inhibition curves were steep and best described by a one-site binding model. In contrast, adenosine A1 agonist competition curves were shallow, as indicated by Hill coefficients less than unity. Computer analysis revealed that these inhibition curves were best described by a two-site binding model. Agonist competition curves generated in the presence of 1 mM GTP resulted in a rightward shift and steepening of the inhibition-concentration curves, whereas antagonist binding was not altered in the presence of GTP. The complex binding interactions found with adenosine agonists indicate that (3H)CGS 15943 labels both high and low affinity components of the adenosine A1 receptor in the rat cortex.

  5. The rise and fall of IgE

    OpenAIRE

    Vernersson, Molly

    2002-01-01

    Immunoglobulin E (IgE) occurs exclusively in mammals and is one of five immunoglobulin (Ig) classes found in man. Unlike other isotypes, IgE is best known for its pathological effects, whereas its physiological role remains somewhat elusive. To trace the emergence of IgE and other post-switch isotypes we have studied Ig expression in two monotreme species, the duck-billed platypus (Ornithorhynchus anatinus) and the short-beaked echidna (Tachyglossus aculeatus), leading to the cloning of Ig...

  6. Evaluation of an in vitro method for the measurement of specific IgE antibody responses: the rat basophilic leukemia (RBL) cell assay

    International Nuclear Information System (INIS)

    The evaluation of allergenic potential is a key parameter in the safety assessment of novel proteins, including those expressed in genetically modified crops and foodstuffs. The majority of allergic reactions to food proteins are immediate type hypersensitivity reactions in which the principal biological effector is IgE antibody; the accurate measurement of specific IgE antibody is therefore a critical factor in experimental systems designed to characterize protein allergenic potential. Due to the presence of much higher concentrations of other immunoglobulin isotypes, the assessment of specific serum IgE antibody poses substantial technical challenges. We have examined the utility of the rat basophilic leukemia (RBL) cell line for the measurement of murine IgE responses. RBL cells were sensitized with mouse monoclonal anti-dinitrophenyl (DNP) IgE antibody and challenged with DNP-albumin conjugates with various hapten substitution ratios (SR). Polyclonal anti-OVA IgE antisera were also assessed for activity in the RBL assay. Results were compared with titers measured in homologous passive cutaneous anaphylaxis (PCA) assay. Marked degranulation of RBL cells was induced by conjugates with SRs of between 16 and 32, whereas conjugates with lower SRs (of 10 or 3) failed to elicit significant serotonin release. All conjugates were able to induce mast cell degranulation in vivo in a PCA assay. Anti-OVA antisera with PCA titers of 1/32 to 1/64 failed to stimulate RBL cell degranulation, whereas high titer antibody (1/2048 to 1/4096 by PCA) induced a positive RBL cell response. Successful stimulation of RBL cell degranulation requires not only appropriate epitope densities but also high affinity antibody. These data indicate that this assay is inappropriate for the routine analysis of specific polyclonal IgE antibody responses such as those that are induced by exposure to complex protein allergens

  7. [Analysis of the impact of heparin on the affinity of high mobility group box-1 protein and the receptor of advanced glycation end products by surface plasmon resonance technology].

    Science.gov (United States)

    Ling, Yan; Wang, Chun-You; Yang, Zhi-Yong

    2009-11-01

    To investigate the affinity constants of heparin with high mobility group protein 1(HMGB1) and HMGB1 with the receptor of advanced glycation end products (RAGE) and to analyze the impact of heparin on the affinity of HMGB1 and RAGE, the standard BIAcore amine coupling chemistry protocol using EDC and NHS was employed for immobilizing. Surface plasmon resonance biosensor technology was used to detect the affinity constants of heparin/HMGB1, HMGB1/RAGE and heparin/ RAGE. Binding analysis was used to investigate the impact of heparin on the affinity of HMGB1 and RAGE. After the immobilization, 9 000 and 5 000 RU rise of HMGB1 and RAGE respectively were obtained. These meant that the immobilized values of HMGB1 and RAGE were about 9 and 5 ng x mm(-2) respectively. The kinetic constants were k(a) = 1.78 x 10(5) L x mol(-1) x s(-1), kd = 8.02 x 10(-4) s(-1), and the affinity constants were KA = 2.22 x 10(8) L x mol(-1), the equilibrium dissociation constant K(D) = 4.5 x 10(-9) mol x L(-1) for heparin and HMGB1; while the kinetic constants were k(a) = 1.85 x 10(3) L x mol(-1) x s(-1), k(d) = 1.81 x 10(-4) s(-1), K(A) = 1.02 x 10(7) L x mol(-1), K(D) = 9.77 x 10(-8) mol x L(-1) for HMGB1 and RAGE; there was very low affinity of heparin with RAGE. The highest concentration of 10 000 u x L(-1) of heparin in this experiment did not reach the saturation with HMGB1. After 50 mg x L(-1) of HMGB1 was mixed with heparin of 50, 100, 1 000, 10 000 u x L(-1), the combining amount of HMGB1 and RAGE declined from 100 to 50 RU. But there were no significant differences between different concentrations of heparin. It was concluded that heparin can combine with HMGB1 and affect the affinity of HMGB1/RAGE. In addition, this impact was not in a dose-dependent manner. PMID:20101991

  8. An improved resolution structure of the human β common receptor involved in IL-3, IL-5 and GM-CSF signalling which gives better definition of the high-affinity binding epitope

    International Nuclear Information System (INIS)

    Data to a maximum of 2.7 Å resolution have allowed a more accurate model of the extracellular domains of the β common cytokine receptor to be determined. Structural details of the membrane-distal BC and FG loops of domain 4 and loops AB and EF of domain 1, which have been implicated in high-affinity binding, are much better defined, revealing a more compact functional epitope. X-ray diffraction has been used to produce and refine a model of the extracellular domains of the β common cytokine receptor. A minor improvement in resolution has resulted in improved electron-density maps, which have given a clearer indication of the position and stabilization of the key residues Tyr15, Phe79, Tyr347, His349, Ile350 and Tyr403 in the elbow region between domain 1 and domain 4 of the dimer-related molecule

  9. Clinical Manifestations of Hyper IgE Syndromes

    Directory of Open Access Journals (Sweden)

    Alexandra F. Freeman

    2010-01-01

    Full Text Available Over the last 4 years, three genetic etiologies of hyper IgE syndromes have been identified: STAT3, DOCK8, and Tyk2. All of these hyper IgE syndromes are characterized by eczema, sinopulmonary infections, and greatly elevated serum IgE. However, each has distinct clinical manifestations. Mutations in STAT3 cause autosomal dominant HIES (Job’s syndrome, which is unique in its diversity of connective tissue, skeletal, and vascular abnormalities. DOCK8 deficiency is characterized by severe cutaneous viral infections such as warts, and a predisposition to malignancies at a young age. Only one individual has been identified with a hyper IgE phenotype associated with Tyk2 deficiency, which is characterized by nontuberculous mycobacterial infection. The identification of these genetic etiologies is leading to advances in understanding the pathogenesis of these syndromes with the goal of improving treatment.

  10. Redesigning Protein Cavities as a Strategy for Increasing Affinity in Protein-Protein Interaction: Interferon-gamma Receptor 1 as a Model

    Czech Academy of Sciences Publication Activity Database

    Černý, Jiří; Biedermannová, Lada; Mikulecký, Pavel; Zahradník, Jiří; Charnavets, Tatsiana; Šebo, Peter; Schneider, Bohdan

    2015-01-01

    Roč. 716945, 28 April 2015 (2015). ISSN 2314-6133 R&D Projects: GA MŠk(CZ) ED1.1.00/02.0109; GA ČR GAP305/10/2184 Institutional support: RVO:86652036 Keywords : CONFORMATIONAL ENTROPY * MOLECULAR RECOGNITION * BINDING-AFFINITY Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 1.579, year: 2014

  11. Specific IgE response in patients with brucellosis.

    OpenAIRE

    Araj, G F; Lulu, A. R.; Khateeb, M. I.; Haj, M

    1990-01-01

    In the search to find discriminative serological markers to differentiate between patients with acute brucellosis and those with chronic brucellosis, an enzyme-linked immunosorbent assay (ELISA) was used to determine and compare the brucella-specific IgE response in 80 sera from patients with acute brucellosis, 37 sera from patients with chronic brucellosis, 26 sera from patients with positive blood cultures for bacteria other than brucella and 51 sera from healthy controls. The IgE findings ...

  12. Cyclic Lactam Hybrid α-MSH/Agouti-Related Protein (AGRP) Analogues with Nanomolar Range Binding Affinities at the Human Melanocortin Receptors

    OpenAIRE

    Mayorov, Alexander V.; Cai, Minying; Palmer, Erin S.; Tanaka, Dustin K.; Cain, James P.; Dedek, Matthew M.; Tan, Bahar; Trivedi, Dev; Victor J. Hruby

    2011-01-01

    A novel hybrid melanocortin pharmacophore was designed based on the topographical similarities between the pharmacophores of Agouti related protein (AGRP) an endogenous melanocortin antagonist, and α-melanocyte-stimulating hormone (α-MSH), an endogenous melanocortin agonist. When employed in two different 23-membered macrocyclic lactam peptide templates, the designed hybrid AGRP/MSH pharmacophore yielded non-competitive ligands with nanomolar range binding affinities. The topography-based pha...

  13. Identification of three adjacent amino acids of interleukin-2 receptor beta chain which control the affinity and the specificity of the interaction with interleukin-2.

    OpenAIRE

    Imler, J L; Miyajima, A; Zurawski, G

    1992-01-01

    The beta chain of the interleukin-2 (IL-2) receptor (IL-2R beta) and the interleukin-3 (IL-3) binding protein AIC2A are members of the family of cytokine receptors, which also includes the receptors for growth hormone (GHR) and prolactin. A four amino acid sequence of AIC2A has recently been shown to be critical for IL-3 binding. We analyze here the function of the analogous sequence of human IL-2R beta and identify three amino acids, Ser132, His133 and Tyr134, which play a critical role in I...

  14. Affine functors and duality

    OpenAIRE

    J. Navarro; Sancho, C.; Sancho, P.

    2009-01-01

    A functor of sets $\\mathbb X$ over the category of $K$-commutative algebras is said to be an affine functor if its functor of functions, $\\mathbb A_{\\mathbb X}$, is reflexive and $\\mathbb X=\\Spec \\mathbb A_{\\mathbb X}$. We prove that affine functors are equal to a direct limit of affine schemes and that affine schemes, formal schemes, the completion of affine schemes along a closed subscheme, etc., are affine functors. Endowing an affine functor $\\mathbb X$ with a functor of monoids structure...

  15. Antihistamines suppress upregulation of histidine decarboxylase gene expression with potencies different from their binding affinities for histamine H1 receptor in toluene 2,4-diisocyanate-sensitized rats.

    Science.gov (United States)

    Mizuguchi, Hiroyuki; Das, Asish K; Maeyama, Kazutaka; Dev, Shrabanti; Shahriar, Masum; Kitamura, Yoshiaki; Takeda, Noriaki; Fukui, Hiroyuki

    2016-04-01

    Antihistamines inhibit histamine signaling by blocking histamine H1 receptor (H1R) or suppressing H1R signaling as inverse agonists. The H1R gene is upregulated in patients with pollinosis, and its expression level is correlated with the severity of nasal symptoms. Here, we show that antihistamine suppressed upregulation of histidine decarboxylase (HDC) mRNA expression in patients with pollinosis, and its expression level was correlated with that of H1R mRNA. Certain antihistamines, including mepyramine and diphenhydramine, suppress toluene-2,4-diisocyanate (TDI)-induced upregulation of HDC gene expression and increase HDC activity in TDI-sensitized rats. However, d-chlorpheniramine did not demonstrate any effect. The potencies of antihistamine suppressive effects on HDC mRNA elevation were different from their H1R receptor binding affinities. In TDI-sensitized rats, the potencies of antihistamine inhibitory effects on sneezing in the early phase were related to H1R binding. In contrast, the potencies of their inhibitory effects on sneezing in the late phase were correlated with those of suppressive effects on HDC mRNA elevation. Data suggest that in addition to the antihistaminic and inverse agonistic activities, certain antihistamines possess additional properties unrelated to receptor binding and alleviate nasal symptoms in the late phase by inhibiting synthesis and release of histamine by suppressing HDC gene transcription. PMID:26980430

  16. Familial Hypercholesterolemia: EVIDENCE FOR A NEWLY RECOGNIZED MUTATION DETERMINING INCREASED FIBROBLAST RECEPTOR AFFINITY BUT DECREASED CAPACITY FOR LOW DENSITY LIPOPROTEIN IN TWO SIBLINGS

    OpenAIRE

    Ostlund, Richard E.; Levy, Richard A.; Witztum, Joseph L.; Schonfeld, Gustav

    1982-01-01

    Cultured skin fibroblasts were obtained from two siblings with classic clinical features of homozygous familial hypercholesterolemia. Plasma cholesterol values were 970 and 802 mg/100 ml in the siblings, 332 mg/100 ml in the mother, and 426 mg/100 ml in the father. Fibroblast receptor-specific capacity for binding and degradation of 125I-low density lipoprotein (LDL) at 37°C was 11% of normal, consistent with the diagnosis of “homozygous LDL receptor-defective” hypercholesterolemia, a disorde...

  17. The high affinity ligand binding conformation of the nuclear 1,25-dihydroxyvitamin D3 receptor is functionally linked to the transactivation domain 2 (AF-2).

    OpenAIRE

    Nayeri, S; Kahlen, J P; Carlberg, C

    1996-01-01

    The nuclear receptor for 1,25-dihydroxyvitamin D3 (VD), VDR, is a transcription factor that mediates all genomic actions of the hormone. The activation of VDR by ligand induces a conformational change within its ligand binding domain (LBD). Due to the lack of a crystal structure analysis, biochemical methods have to be applied in order to investigate the details of this receptor-ligand interaction. The limited protease digestion assay can be used as a tool for the determination of a functiona...

  18. Discovery of the First Small-Molecule Opioid Pan Antagonist with Nanomolar Affinity at Mu, Delta, Kappa, and Nociceptin Opioid Receptors

    OpenAIRE

    Zaveri, Nurulain T.; Journigan, V. Blair; Polgar, Willma E.

    2015-01-01

    The trans-(3R,4R)-dimethyl-4-(3-hydroxyphenyl)piperidine scaffold is a known pharmacophore for mu opioid (MOP), kappa opioid (KOP), and delta opioid (DOP) receptor antagonists; however, it has not been explored in nociceptin opioid (NOP/ORL-1) receptor ligands. We recently found that the selective KOP antagonist JDTic, (3R)-7-hydroxy-N-((1S)-1-{[(3R,4R)-4-(3-hydroxyphenyl)-3,4-dimethyl-1-piperidinyl]methyl}-2-methylpropyl)-1,2,3,4-tetrahydro-3-isoquinolinecarboxamide, containing this opioid a...

  19. Presence of IgE cells in human placenta is independent of malaria infection or chorioamnionitis

    DEFF Research Database (Denmark)

    Rindsjö, E; Hulthén Varli, I; Ofori, M F;

    2006-01-01

    We have shown previously that numerous IgE(+) macrophage-like cells are present in the villous stroma of full term placenta and that there was no difference in the amount of IgE(+) cells between allergic and non-allergic mothers. The presence of such an abundant number of IgE(+) cells in the...... from Ghana with and without malaria parasites. The immunohistochemical staining pattern for IgE looked similar to our previous study, with the IgE located on Hofbauer-like cells. We could not find any difference in the amount or distribution of IgE(+) cells between malaria-infected and non...

  20. Clinical analysis of high serum IgE in autoimmune pancreatitis

    Institute of Scientific and Technical Information of China (English)

    Kenji; Hirano; Minoru; Tada; Hiroyuki; Isayama; Kazumichi; Kawakubo; Hiroshi; Yagioka; Takashi; Sasaki; Hirofumi; Kogure; Yousuke; Nakai; Naoki; Sasahira; Takeshi; Tsujino; Nobuo; Toda; Kazuhiko; Koike

    2010-01-01

    AIM: To clarify the clinical significance of high serum IgE in autoimmune pancreatitis (AIP). METHODS: Forty-two AIP patients, whose IgE was measured before steroid treatment, were analyzed. To evaluate the relationship between IgE levels and the disease activity of AIP, we examined (1) Frequency of high IgE (> 170 IU/mL) and concomitant allergic dis-eases requiring treatment; (2) Correlations between IgG, IgG4, and IgE; (3) Relationship between the presence of extrapancreatic lesions and IgE; (4) Re-lation...

  1. IGES data exchange between dissimilar CAD-CAM systems

    Energy Technology Data Exchange (ETDEWEB)

    Theilen, D.F.

    1982-06-01

    Sandia National Laboratories and Bendix Kansas City have successfully transferred geometrical model data by using a neutral graphics format, Initial Graphics Exchange Specification (IGES). Data was exchanged between two dissimilar CAD-CAM systems without the need for special translators. The exchanged data was used to fabricate a mechanical part. Design information developed on a CAD-CAM system was translated to IGES with a commercial preprocessor at Sandia. A second vendor's postprocessor translated the design information from IGES to the CAD-CAM system at Bendix. The Bendix CAD-CAM system was used to generate the numerical control tape from the exchange data. The tape was used to fabricate the part. A production print from Sandia was used during final inspection to verify part accuracy. Project results indicated that both vendor-furnished translators correctly transferred the geometric model of points, lines and arcs.

  2. IGES data exchange between dissimilar CAD-CAM systems

    Energy Technology Data Exchange (ETDEWEB)

    Theilen, D.F.; Jones, J.F.

    1982-04-01

    Sandia National Laboratories, Livermore (SNLL), and Bendix Kansas City have successfully transferred geometric model data by using a neutral graphics format, Initial Graphics Exchange Specification (IGES). Data was exchanged between two dissimilar CAD-CAM systems without the need for special translators. The exchanged data was used to fabricate a mechanical part. Design information developed on a CAD-CAM system was translated to IGES with a commercial preprocessor at Sandia. A second vendor's postprocessor translated the design information from IGES to the CAD-CAM system at Bendix. The Bendix system was used to generate the numerical control tape from the exchanged data. The tape was used to fabricate the part. A production print from Sandia was used during final inspection to verify part accuracy. Project results indicated that both vendor-furnished translators correctly transferred the three-dimensional geometric model of points, lines, and arcs.

  3. Engineered α4β2 nicotinic acetylcholine receptors as models for measuring agonist binding and effect at the orthosteric low-affinity α4-α4 interface

    DEFF Research Database (Denmark)

    Ahring, Philip K.; Olsen, Jeppe A.; Nielsen, Elsebet O.;

    2015-01-01

    2)(2) receptors. However, standard saturation binding experiments with [H-3]epibatidine did not reveal biphasic binding under the conditions utilized. Therefore, an engineered beta 2 construct (beta 2(HQT)), which converts the beta(-) face to resemble that of an alpha 4(-) face, was utilized...

  4. Mutation-induced quisqualic acid and ibotenic acid affinity at the metabotropic glutamate receptor subtype 4: ligand selectivity results from a synergy of several amino acid residues

    DEFF Research Database (Denmark)

    Hermit, Mette B; Greenwood, Jeremy R; Bräuner-Osborne, Hans

    2004-01-01

    The metabotropic glutamate receptors (mGluRs) are key modulators of excitatory neurotransmission in the central nervous system. The eight mGluR subtypes are seven trans-membrane-spanning proteins that possess a large extracellular amino-terminal domain in which the endogenous ligand binding pocke...

  5. Crystal Structure of Human Interferon-[lamda]1 in Complex with Its High-Affinity Receptor Interferon-[lamda]R1

    Energy Technology Data Exchange (ETDEWEB)

    Miknis, Zachary; Magracheva, Eugenia; Li, Wei; Zdanov, Alexander; Kotenko, Sergei V.; Wlodawer, Alexander (NJMS); (NCI)

    2010-12-01

    Interferon (IFN)-{lambda}1 [also known as interleukin (IL)-29] belongs to the recently discovered group of type III IFNs. All type III IFNs initiate signaling processes through formation of specific heterodimeric receptor complexes consisting of IFN-{lambda}R1 and IL-10R2. We have determined the structure of human IFN-{lambda}1 complexed with human IFN-{lambda}R1, a receptor unique to type III IFNs. The overall structure of IFN-{lambda}1 is topologically similar to the structure of IL-10 and other members of the IL-10 family of cytokines. IFN-{lambda}R1 consists of two distinct domains having fibronectin type III topology. The ligand-receptor interface includes helix A, loop AB, and helix F on the IFN site, as well as loops primarily from the N-terminal domain and inter-domain hinge region of IFN-{lambda}R1. Composition and architecture of the interface that includes only a few direct hydrogen bonds support an idea that long-range ionic interactions between ligand and receptor govern the process of initial recognition of the molecules while hydrophobic interactions finalize it.

  6. Radiosynthesis and in vitro validation of 3H-NS14492 as a novel high affinity alpha7 nicotinic receptor radioligand

    DEFF Research Database (Denmark)

    Magnussen, Janus H.; Ettrup, Anders; Donat, Cornelius K.;

    2015-01-01

    /mg protein. This binding assay further revealed the Ki rank order for a number of alpha 7 nicotinic receptor agonists, and positive allosteric modulators (PAMs). Further, we saw increased binding of 3H-NS14492 to pig frontal cortex membranes when co-incubated with PNU-120596, a type II PAM. Taken together...

  7. Affinity-Based Screening of Tetravalent Peptides Identifies Subtype-Selective Neutralizers of Shiga Toxin 2d, a Highly Virulent Subtype, by Targeting a Unique Amino Acid Involved in Its Receptor Recognition.

    Science.gov (United States)

    Mitsui, Takaaki; Watanabe-Takahashi, Miho; Shimizu, Eiko; Zhang, Baihao; Funamoto, Satoru; Yamasaki, Shinji; Nishikawa, Kiyotaka

    2016-09-01

    Shiga toxin (Stx), a major virulence factor of enterohemorrhagic Escherichia coli (EHEC), can be classified into two subgroups, Stx1 and Stx2, each consisting of various closely related subtypes. Stx2 subtypes Stx2a and Stx2d are highly virulent and linked with serious human disorders, such as acute encephalopathy and hemolytic-uremic syndrome. Through affinity-based screening of a tetravalent peptide library, we previously developed peptide neutralizers of Stx2a in which the structure was optimized to bind to the B-subunit pentamer. In this study, we identified Stx2d-selective neutralizers by targeting Asn16 of the B subunit, an amino acid unique to Stx2d that plays an essential role in receptor binding. We synthesized a series of tetravalent peptides on a cellulose membrane in which the core structure was exactly the same as that of peptides in the tetravalent library. A total of nine candidate motifs were selected to synthesize tetravalent forms of the peptides by screening two series of the tetravalent peptides. Five of the tetravalent peptides effectively inhibited the cytotoxicity of Stx2a and Stx2d, and notably, two of the peptides selectively inhibited Stx2d. These two tetravalent peptides bound to the Stx2d B subunit with high affinity dependent on Asn16. The mechanism of binding to the Stx2d B subunit differed from that of binding to Stx2a in that the peptides covered a relatively wide region of the receptor-binding surface. Thus, this highly optimized screening technique enables the development of subtype-selective neutralizers, which may lead to more sophisticated treatments of infections by Stx-producing EHEC. PMID:27382021

  8. Canonical factorization of the quotient morphism for an affine $\\mathbb{G}_a$-variety

    OpenAIRE

    Freudenburg, Gene

    2016-01-01

    Working over a ground field of characteristic zero, this paper studies the quotient morphism $\\pi :X\\to Y$ for an affine $\\mathbb{G}_a$-variety $X$ with affine quotient $Y$. We show that the degree modules associated to the $\\mathbb{G}_a$-action give a uniquely determined sequence of dominant $\\mathbb{G}_a$-equivariant morphisms, $X=X_r\\to X_{r-1}\\to\\cdots\\to X_1\\to X_0=Y$, where $X_i$ is an affine variety and $X_{i+1}\\to X_i$ is birational for each $i\\ge 1$. This is the canonical factorizati...

  9. Crystallization and preliminary X-ray structural studies of a high-affinity CD8αα co-receptor to pMHC

    Energy Technology Data Exchange (ETDEWEB)

    Cole, David K. [Nuffield Department of Clinical Medicine, John Radcliffe Hospital, Oxford University, Oxford OX3 9DU (United Kingdom); Rizkallah, Pierre J., E-mail: p.j.rizkallah@dl.ac.uk [CCLRC Daresbury Laboratory, Warrington, Cheshire WA4 4AD (United Kingdom); Sami, Malkit; Lissin, Nikolai M.; Gao, Feng [Avidex Ltd, 57c Milton Park, Abingdon, Oxon OX14 4RX (United Kingdom); Bell, John I. [Nuffield Department of Clinical Medicine, John Radcliffe Hospital, Oxford University, Oxford OX3 9DU (United Kingdom); Boulter, Jonathan M. [Medical Biochemistry and Immunology, Henry Wellcome Building, University of Wales College of Medicine, Heath Park, Cardiff CF14 4XN,Wales (United Kingdom); Glick, Meir [Novartis Pharmaceuticals, One Health Plaza, East Hanover, NJ 07936 (United States); Vuidepot, Anne-Lise; Jakobsen, Bent K., E-mail: p.j.rizkallah@dl.ac.uk [Avidex Ltd, 57c Milton Park, Abingdon, Oxon OX14 4RX (United Kingdom); Gao, George F. [Nuffield Department of Clinical Medicine, John Radcliffe Hospital, Oxford University, Oxford OX3 9DU (United Kingdom)

    2005-03-01

    A high-affinity mutant CD8 (haCD8) has been developed with the aim of developing a therapeutic immunosuppressor. In order to fully understand the nature of the haCD8 interaction, this protein was crystallized using the sitting-drop vapour-diffusion method. The class I CD8 positive T-cell response is involved in a number of conditions in which artificial down-regulation and control would be therapeutically beneficial. Such conditions include a number of autoimmune diseases and graft rejection in transplant patients. Although the CD8 T-cell response is dominated by the TCR–pMHC interaction, activation of T cells is in most cases also dependent on a number of associated signalling molecules. Previous work has demonstrated the ability of one such molecule (CD8) to act as an antagonist to T-cell activation if added in soluble form. Therefore, a high-affinity mutant CD8 (haCD8) has been developed with the aim of developing a therapeutic immunosuppressor. In order to fully understand the nature of the haCD8 interaction, this protein was crystallized using the sitting-drop vapour-diffusion method. Single haCD8 crystals were cryocooled and used for data collection. These crystals belonged to space group P6{sub 4}22 (assumed by similarity to the wild type), with unit-cell parameters a = 101.08, c = 56.54 Å. V{sub M} calculations indicated one molecule per asymmetric unit. A 2 Å data set was collected and the structure is currently being determined using molecular replacement.

  10. Interleukin-21-dependent modulation of T cell antigen receptor reactivity towards low affinity peptide ligands in autoreactive CD8(+) T lymphocytes.

    Science.gov (United States)

    Bobbala, Diwakar; Orkhis, Sakina; Kandhi, Rajani; Ramanathan, Sheela; Ilangumaran, Subburaj

    2016-09-01

    IL-21 promotes autoimmune type-1 diabetes (T1D) in NOD mice by facilitating CD4(+) T cell help to CD8(+) T cells. IL-21 also enables autoreactive CD8(+) T cells to respond to weak TCR ligands and induce T1D. Here, we assessed whether IL-21 is essential for T1D induction in a mouse model where the disease can occur independently of CD4 help. In this model, which expresses lymphocytic choriomeningitis virus (LCMV) glycoprotein (GP) antigen under the rat insulin promoter (RIP-GP), LCMV infection activates CD8(+) T cells reactive to the GP-derived GP33 peptide that attack pancreatic islets and cause T1D. We show that IL-21 deficiency in RIP-GP mice did not impair T1D induction by LCMV expressing the wildtype GP33 peptide. Surprisingly, LCMV-L6F, expressing a weak peptide mimic of GP33, induced T1D more efficiently in Il21(-/-)RIP-GP mice than in controls. However, LCMV-C4Y expressing a very weak peptide mimic of GP33 did not induce T1D in Il21(-/-) mice, but T cells from the infected mice caused disease in lymphopenic RIP-GP mice upon adoptive transfer. Using Nur77(GFP) reporter mice, we show that CD8(+) T cells from Il21(-/-) mice expressing the GP33-specific transgenic P14 TCR showed increased reactivity towards low affinity TCR ligands. Collectively, our findings show that IL-21 is not always required for T1D induction by autoreactive CD8(+) T cells, and suggest that IL-21 may play an important role in regulating CD8(+) T cell reactivity towards low affinity TCR ligands. PMID:27300756

  11. Toward Selective Drug Development for the Human 5-Hydroxytryptamine 1E Receptor: A Comparison of 5-Hydroxytryptamine 1E and 1F Receptor Structure-Affinity RelationshipsS⃞

    OpenAIRE

    Klein, Michael T.; Dukat, Małgorzata; Glennon, Richard A.; Teitler, Milt

    2011-01-01

    The 5-hydroxytryptamine (5-HT) 1E receptor is highly expressed in the human frontal cortex and hippocampus, and this distribution suggests the function of 5-HT1E receptors might be linked to memory. To test this hypothesis, behavioral experiments are needed. Because rats and mice lack a 5-HT1E receptor gene, knockout strategies cannot be used to elucidate this receptor's functions. Thus, selective pharmacological tools must be developed. The tryptamine-related agonist BRL54443 [5-hydroxy-3-(1...

  12. β-Lactoglobulin mutant Lys69Asn has attenuated IgE and increased retinol binding activity.

    Science.gov (United States)

    Taheri-Kafrani, Asghar; Tavakkoli Koupaie, Neda; Haertlé, Thomas

    2015-10-20

    β-Lactoglobulin (β-LG) is a member of lipocalin superfamily of transporters for small hydrophobic molecules such as retinoids, fatty acids, drugs, and vitamins. β-LG also is one of the major allergens in milk. Despite a lot of research on decreasing cow's milk allergenicity, the effects of mutations of β-LG on recognition by IgE from cow's milk allergy (CMA) patients have not been investigated. We describe here the expression in the yeast Pichia pastoris of a mutant bovine β-LG, in which lysine at position 69, in the main epitopes of the protein, was changed into asparagine (Lys69Asn). The purity and native like folded structure of the recombinant Lys69Asn β-LG was confirmed by HPLC, SDS-PAGE, mass spectrometry and circular dichroism. Lys69Asn β-LG has a fourfold stronger affinity than the wild-type protein for retinol, palmitic acid, and resveratrol, as determined by quenching of the intrinsic tryptophan fluorescence. At the same time the Lys69Asn mutant had a 9 fold attenuated, compared with the wild-type, affinity for IgE of sera from patients suffering from cow's milk allergy, whereas no difference could be detected between mutant and wild-type for binding of the IgGs of four monoclonal antibodies. The results of this study demonstrated the significant role of Lys69 residue on the binding and immuoreactivity properties of β-LG. PMID:26281976

  13. IgE vs IgG4 epitopes of the peanut allergen Ara h 1 in patients with severe allergy

    DEFF Research Database (Denmark)

    Bøgh, Katrine Lindholm; Nielsen, H.; Eiwegger, T.; Madsen, Charlotte Bernhard; Mills, E. N. C.; Szépfalusi, Z.; Roggen, E. L.

    Background: Development and maintenance of tolerance to food allergens may be associated with increased levels of specific IgG4. It has been suggested that co-localisation of IgG4 and IgE binding epitopes may be of great significance for the tolerance, where IgG4 may act by blocking IgE binding to...... the allergen. However, recent studies have demonstrated the very importance of the IgG4-epitope affinity for the blocking ability. Studies comparing IgE and IgG4 binding epitopes mainly focus on the identification of linear epitopes. Peanut allergy is one of the most severe and persistent forms of...... food allergy. The importance of conformational epitopes, of the major peanut allergen Ara h 1, has been demonstrated. The aim of this study was to compare Ara h 1-specific epitope patterns for IgE and IgG4 in patients with severe peanut allergy applying a method suitable to identify both linear and...

  14. The Duration of Nicotine Withdrawal-Associated Deficits in Contextual Fear Conditioning Parallels Changes in Hippocampal High Affinity Nicotinic Acetylcholine Receptor Upregulation

    OpenAIRE

    Gould, Thomas J.; Portugal, George S.; André, Jessica M.; Tadman, Matthew P.; Marks, Michael J.; Kenney, Justin W.; YILDIRIM, Emre; Adoff, Michael

    2012-01-01

    A predominant symptom of nicotine withdrawal is cognitive deficits, yet understanding of the neural basis for these deficits is limited. Withdrawal from chronic nicotine disrupts contextual learning in mice and this deficit is mediated by direct effects of nicotine in the hippocampus. Chronic nicotine treatment upregulates nicotinic acetylcholine receptors (nAChR); however, it is unknown whether upregulation is related to the observed withdawal-induced cognitive deficits. If a relationship be...

  15. Dual Affine invariant points

    OpenAIRE

    Meyer, Mathieu; Schuett, Carsten; Werner, Elisabeth M.

    2013-01-01

    An affine invariant point on the class of convex bodies in R^n, endowed with the Hausdorff metric, is a continuous map p which is invariant under one-to-one affine transformations A on R^n, that is, p(A(K))=A(p(K)). We define here the new notion of dual affine point q of an affine invariant point p by the formula q(K^{p(K)})=p(K) for every convex body K, where K^{p(K)} denotes the polar of K with respect to p(K). We investigate which affine invariant points do have a dual point, whether this ...

  16. The NKG2D Ligands RAE-1δ and RAE-1ε Differ with Respect to Their Receptor Affinity, Expression Profiles and Transcriptional Regulation

    OpenAIRE

    Cédile, Oriane; Popa, Natalia; Pollet-Villard, Frédéric; Garmy, Nicolas; Ibrahim, El Chérif; Boucraut, José

    2010-01-01

    Background RAE-1 is a ligand of the activating receptor NKG2D expressed by NK cells, NKT, γδT and some CD8+T lymphocytes. RAE-1 is overexpressed in tumor cell lines and its expression is induced after viral infection and genotoxic stress. We have recently demonstrated that RAE-1 is expressed in the adult subventricular zone (SVZ) from C57BL/6 mice. RAE-1 is also expressed in vitro by neural stem/progenitor cells (NSPCs) and plays a non-immune role in cell proliferation. The C57BL/6 mouse geno...

  17. Structure-activity relationships of N-substituted 4-(trifluoromethoxy)benzamidines with affinity for GluN2B-containing NMDA receptors.

    Science.gov (United States)

    Beinat, Corinne; Banister, Samuel D; Hoban, Jane; Tsanaktsidis, John; Metaxas, Athanasios; Windhorst, Albert D; Kassiou, Michael

    2014-02-01

    GluN2B subtype-selective NMDA antagonists represent promising therapeutic targets for the symptomatic treatment of multiple CNS pathologies. A series of N-benzyl substituted benzamidines were synthesised and the benzyl ring was further replaced with various polycyclic moieties. Compounds were evaluated for activity at GluN2B containing NMDA receptors where analogues 9, 12, 16 and 18 were the most potent of the series, replacement of the benzyl ring with polycycles resulted in a complete loss of activity. PMID:24412068

  18. G-protein-coupled receptor affinity prediction based on the use of a profiling dataset: QSAR design, synthesis, and experimental validation.

    Science.gov (United States)

    Rolland, Catherine; Gozalbes, Rafael; Nicolaï, Eric; Paugam, Marie-France; Coussy, Laurent; Barbosa, Frédérique; Horvath, Dragos; Revah, Frédéric

    2005-10-20

    A QSAR model accounting for "average" G-protein-coupled receptor (GPCR) binding was built from a large set of experimental standardized binding data (1939 compounds systematically tested over 40 different GPCRs) and applied to the design of a library of "GPCR-predicted" compounds. Three hundred and sixty of these compounds were randomly selected and tested in 21 GPCR binding assays. Positives were defined by their ability to inhibit by more than 70% the binding of reference compounds at 10 microM. A 5.5-fold enrichment in positives was observed when comparing the "GPCR-predicted" compounds with 600 randomly selected compounds predicted as "non-GPCR" from a general collection. The model was efficient in predicting strongest binders, since enrichment was greater for higher cutoffs. Significant enrichment was also observed for peptidic GPCRs and receptors not included to develop the QSAR model, suggesting the usefulness of the model to design ligands binding with newly identified GPCRs, including orphan ones. PMID:16220973

  19. The NKG2D ligands RAE-1δ and RAE-1ε differ with respect to their receptor affinity, expression profiles and transcriptional regulation

    DEFF Research Database (Denmark)

    Cédile, Oriane; Popa, Natalia; Pollet-Villard, Frédéric;

    2010-01-01

    subventricular zone (SVZ) from C57BL/6 mice. RAE-1 is also expressed in vitro by neural stem/progenitor cells (NSPCs) and plays a non-immune role in cell proliferation. The C57BL/6 mouse genome contains two rae-1 genes, rae-1δ and rae-1ε encoding two different proteins. The goals of this study are first to......RAE-1 is a ligand of the activating receptor NKG2D expressed by NK cells, NKT, γδT and some CD8(+)T lymphocytes. RAE-1 is overexpressed in tumor cell lines and its expression is induced after viral infection and genotoxic stress. We have recently demonstrated that RAE-1 is expressed in the adult...

  20. IgE antibody responses in schistosomiasis measured by a radioallergosorbent test

    International Nuclear Information System (INIS)

    Helminth infections are associated with the production of unusually high concentrations of circulating IgE antibody. Assays for IgE antibodies should provide useful approaches for the study of protective immunity and may also be of use in serodiagnosis of diseases induced by helminths. The radioallergosorbent test is carried out by attaching the antigen or allergen to an insoluble supportive material, allowing the IgE antibodies in the test serum to react with excess bound antigen, and then estimating the IgE antibody bound by its reaction with 125I-labelled goat anti-human IgE antibody

  1. Allergen-specific IgG antibodies purified from mite-allergic patients sera block the IgE recognition of Dermatophagoides pteronyssinus antigens: an in vitro study.

    Science.gov (United States)

    Siman, Isabella Lima; de Aquino, Lais Martins; Ynoue, Leandro Hideki; Miranda, Juliana Silva; Pajuaba, Ana Claudia Arantes Marquez; Cunha-Júnior, Jair Pereira; Silva, Deise Aparecida Oliveira; Taketomi, Ernesto Akio

    2013-01-01

    One of the purposes of specific immunotherapy (SIT) is to modulate humoral immune response against allergens with significant increases in allergen-specific IgG levels, commonly associated with blocking activity. The present study investigated in vitro blocking activity of allergen-specific IgG antibodies on IgE reactivity to Dermatophagoides pteronyssinus (Dpt) in sera from atopic patients. Dpt-specific IgG antibodies were purified by ammonium sulfate precipitation followed by protein-G affinity chromatography. Purity was checked by SDS-PAGE and immunoreactivity by slot-blot and immunoblot assays. The blocking activity was evaluated by inhibition ELISA. The electrophoretic profile of the ammonium sulfate precipitated fraction showed strongly stained bands in ligand fraction after chromatography, compatible with molecular weight of human whole IgG molecule. The purity degree was confirmed by detecting strong immunoreactivity to IgG, negligible to IgA, and no reactivity to IgE and IgM. Dpt-specific IgG fraction was capable of significantly reducing levels of IgE anti-Dpt, resulting in 35%-51% inhibition of IgE reactivity to Dpt in atopic patients sera. This study showed that allergen-specific IgG antibodies purified from mite-allergic patients sera block the IgE recognition of Dermatophagoides pteronyssinus antigens. This approach reinforces that intermittent measurement of serum allergen-specific IgG antibodies will be an important objective laboratorial parameter that will help specialists to follow their patients under SIT. PMID:24069042

  2. Allergen-Specific IgG Antibodies Purified from Mite-Allergic Patients Sera Block the IgE Recognition of Dermatophagoides pteronyssinus Antigens: An In Vitro Study

    Directory of Open Access Journals (Sweden)

    Isabella Lima Siman

    2013-01-01

    Full Text Available One of the purposes of specific immunotherapy (SIT is to modulate humoral immune response against allergens with significant increases in allergen-specific IgG levels, commonly associated with blocking activity. The present study investigated in vitro blocking activity of allergen-specific IgG antibodies on IgE reactivity to Dermatophagoides pteronyssinus (Dpt in sera from atopic patients. Dpt-specific IgG antibodies were purified by ammonium sulfate precipitation followed by protein-G affinity chromatography. Purity was checked by SDS-PAGE and immunoreactivity by slot-blot and immunoblot assays. The blocking activity was evaluated by inhibition ELISA. The electrophoretic profile of the ammonium sulfate precipitated fraction showed strongly stained bands in ligand fraction after chromatography, compatible with molecular weight of human whole IgG molecule. The purity degree was confirmed by detecting strong immunoreactivity to IgG, negligible to IgA, and no reactivity to IgE and IgM. Dpt-specific IgG fraction was capable of significantly reducing levels of IgE anti-Dpt, resulting in 35%–51% inhibition of IgE reactivity to Dpt in atopic patients sera. This study showed that allergen-specific IgG antibodies purified from mite-allergic patients sera block the IgE recognition of Dermatophagoides pteronyssinus antigens. This approach reinforces that intermittent measurement of serum allergen-specific IgG antibodies will be an important objective laboratorial parameter that will help specialists to follow their patients under SIT.

  3. Affine Grassmann codes

    DEFF Research Database (Denmark)

    Høholdt, Tom; Beelen, Peter; Ghorpade, Sudhir Ramakant

    2010-01-01

    We consider a new class of linear codes, called affine Grassmann codes. These can be viewed as a variant of generalized Reed-Muller codes and are closely related to Grassmann codes.We determine the length, dimension, and the minimum distance of any affine Grassmann code. Moreover, we show that...... affine Grassmann codes have a large automorphism group and determine the number of minimum weight codewords....

  4. Open conformers of HLA-F are high-affinity ligands of the activating NK-cell receptor KIR3DS1.

    Science.gov (United States)

    Garcia-Beltran, Wilfredo F; Hölzemer, Angelique; Martrus, Gloria; Chung, Amy W; Pacheco, Yovana; Simoneau, Camille R; Rucevic, Marijana; Lamothe-Molina, Pedro A; Pertel, Thomas; Kim, Tae-Eun; Dugan, Haley; Alter, Galit; Dechanet-Merville, Julie; Jost, Stephanie; Carrington, Mary; Altfeld, Marcus

    2016-09-01

    The activating natural killer (NK)-cell receptor KIR3DS1 has been linked to the outcome of various human diseases, including delayed progression of disease caused by human immunodeficiency virus type 1 (HIV-1), yet a ligand that would account for its biological effects has remained unknown. We screened 100 HLA class I proteins and found that KIR3DS1 bound to HLA-F, a result we confirmed biochemically and functionally. Primary human KIR3DS1(+) NK cells degranulated and produced antiviral cytokines after encountering HLA-F and inhibited HIV-1 replication in vitro. Activation of CD4(+) T cells triggered the transcription and surface expression of HLA-F mRNA and HLA-F protein, respectively, and induced binding of KIR3DS1. HIV-1 infection further increased the transcription of HLA-F mRNA but decreased the binding of KIR3DS1, indicative of a mechanism for evading recognition by KIR3DS1(+) NK cells. Thus, we have established HLA-F as a ligand of KIR3DS1 and have demonstrated cell-context-dependent expression of HLA-F that might explain the widespread influence of KIR3DS1 in human disease. PMID:27455421

  5. Detection of IgE insulin antibody with radioallergosorbent test

    International Nuclear Information System (INIS)

    An in vitro method for detecting IgE insulin antibody using the principle of the radioallergosorbent test (RAST) is described. In six patients with insulin allergy, the RAST values were higher than in normal persons or insulin-treated diabetics without insulin allergy. No differences were observed between normal persons and insulin-treated diabetics without insulin allergy. Moreover, it was observed that in one patient treated with highly purified insulin, there was a gradual decrease of RAST value parallel to the radioinsulin binding activity and clinical allergic symptoms. The RAST value of insulin is slightly inhibited by non-IgE antibodies and is, therefore, a semiquantitative value. However, the RAST is simple to perform and reproducible; it is therefore very useful in the detection of IgE insulin antibodies. (orig.)

  6. Therapeutic effects of antigen affinity-purified polyclonal anti-receptor of advanced glycation end-product (RAGE) antibodies on cholestasis-induced liver injury in rats.

    Science.gov (United States)

    Xia, Peng; Deng, Qing; Gao, Jin; Yu, Xiaolan; Zhang, Yang; Li, Jingjing; Guan, Wen; Hu, Jianjun; Tan, Quanhui; Zhou, Liang; Han, Wei; Yuan, Yunsheng; Yu, Yan

    2016-05-15

    Cholestasis leads to acute hepatic injury, fibrosis/cirrhosis, inflammation, and duct proliferation. We investigated whether blocking receptor of advanced glycation end-products (RAGE) with polyclonal anti-RAGE antibodies (anti-RAGE) could regulate acute liver injury and fibrosis in a rat bile duct ligation (BDL) model. Male Wister rats received 0.5mg/kg rabbit anti-RAGE or an equal amount of rabbit IgG by subcutaneous injection twice a week after BDL. Samples of liver tissue and peripheral blood were collected at 14 days after BDL. Serum biochemistry and histology were used to analyze the degree of liver injury. Quantitative real-time PCR (qPCR) and immunohistochemical staining were used to further analyze liver injury. Anti-RAGE improved the gross appearance of the liver and the rat survival rate. Liver tissue histology and relevant serum biochemistry indicated that anti-RAGE attenuated liver necrosis, inflammation, liver fibrosis, and duct proliferation in the BDL model. qPCR and western blotting showed significant reductions in interleukin-1β expression levels in the liver by treatment with anti-RAGE. Anti-RAGE also significantly reduced the mRNA levels of α1(1) collagen (Col1α1) and cholesterol 7α-hydroxylase, and the ratio of tissue inhibitor of matrix metalloproteinase-1 to matrix metalloproteinases (MMPs) in the liver. In addition, anti-RAGE regulated the transcriptional level of Col1α1 and MMP-9 in transforming growth factor-β-induced activated LX-2 cells in vitro. Anti-RAGE was found to inhibit hepatic stellate cell proliferation in vivo and in vitro. Therefore, anti-RAGE can protect the liver from injury induced by BDL in rats. PMID:26970185

  7. Application Protocol, Initial Graphics Exchange Specification (IGES), Layered Electrical Product

    Energy Technology Data Exchange (ETDEWEB)

    O`Connell, L.J. [ed.

    1994-12-01

    An application protocol is an information systems engineering view of a specific product The view represents an agreement on the generic activities needed to design and fabricate the product the agreement on the information needed to support those activities, and the specific constructs of a product data standard for use in transferring some or all of the information required. This application protocol describes the data for electrical and electronic products in terms of a product description standard called the Initial Graphics Exchange Specification (IGES). More specifically, the Layered Electrical Product IGES Application Protocol (AP) specifies the mechanisms for defining and exchanging computer-models and their associated data for those products which have been designed in two dimensional geometry so as to be produced as a series of layers in IGES format The AP defines the appropriateness of the data items for describing the geometry of the various parts of a product (shape and location), the connectivity, and the processing and material characteristics. Excluded is the behavioral requirements which the product was intended to satisfy, except as those requirements have been recorded as design rules or product testing requirements.

  8. Cooperative Interactions of Oligosaccharide and Peptide Moieties of a Glycopeptide Derived from IgE with Galectin-9.

    Science.gov (United States)

    Nakakita, Shin-Ichi; Itoh, Aiko; Nakakita, Yukari; Nonaka, Yasuhiro; Ogawa, Takashi; Nakamura, Takanori; Nishi, Nozomu

    2016-01-01

    We previously showed that galectin-9 suppresses degranulation of mast cells through protein-glycan interaction with IgE. To elucidate the mechanism of the interaction in detail, we focused on identification and structural analysis of IgE glycans responsible for the galectin-9-induced suppression using mouse monoclonal IgE (TIB-141). TIB-141 in combination with the antigen induced degranulation of RBL-2H3 cells, which was almost completely inhibited by human and mouse galectin-9. Sequential digestion of TIB-141 with lysyl endopeptidase and trypsin resulted in the identification of a glycopeptide (H-Lys13-Try3; 48 amino acid residues) with a single N-linked oligosaccharide near the N terminus capable of neutralizing the effect of galectin-9 and another glycopeptide with two N-linked oligosaccharides (H-Lys13-Try1; 16 amino acid residues) having lower activity. Enzymatic elimination of the oligosaccharide chain from H-Lys13-Try3 and H-Lys13-Try1 completely abolished the activity. Removal of the C-terminal 38 amino acid residues of H-Lys13-Try3 with glutamyl endopeptidase, however, also resulted in loss of the activity. We determined the structures of N-linked oligosaccharides of H-Lys13-Try1. The galectin-9-binding fraction of pyridylaminated oligosaccharides contained asialo- and monosialylated bi/tri-antennary complex type oligosaccharides with a core fucose residue. The structures of the oligosaccharides were consistent with the sugar-binding specificity of galectin-9, whereas the nonbinding fraction contained monosialylated and disialylated biantennary complex type oligosaccharides with a core fucose residue. Although the oligosaccharides linked to H-Lys13-Try3 could not be fully characterized, these results indicate the possibility that cooperative binding of oligosaccharide and neighboring polypeptide structures of TIB-141 to galectin-9 affects the overall affinity and specificity of the IgE-lectin interaction. PMID:26582205

  9. BAY 50-4798, a novel, high-affinity receptor-specific recombinant interleukin-2 analog, induces dose-dependent increases in CD25 expression and proliferation among unstimulated, human peripheral blood mononuclear cells in vitro.

    Science.gov (United States)

    Matthews, Lynn; Chapman, Sherita; Ramchandani, Meena S; Lane, H Clifford; Davey, Richard T; Sereti, Irini

    2004-12-01

    Interleukin-2 administration induces CD4 T cell expansion in HIV-infected patients, however, toxicity can limit dosing. BAY 50-4798 is a recombinant IL-2 analog with >1000-fold specificity for the high-affinity IL-2 receptor. The effects of this compound on unstimulated human PBMC were evaluated. PBMC from HIV(-) and HIV(+) donors were cultured in vitro with incremental doses of BAY 50-4798 or aldesleukin. CD25 expression and proliferation were evaluated with flow cytometry. Cytokine levels were measured by ELISA in culture supernatants. BAY 50-4798 induced dose-dependent increases in CD25 expression and proliferation of T cells, NK, and B cells and showed selectivity for CD4 T cells expressing CD25. Induction of pro-inflammatory cytokines was also dose-dependent and was observed at the concentrations of BAY 50-4798 with the highest biologic activity. These data suggest that BAY 50-4798 can induce proliferation of unstimulated T cells but loss of T cell selectivity and induction of pro-inflammatory cytokines occur at concentrations exerting the highest biologic activity. PMID:15507389

  10. Synthesis, structure activity relationship, radiolabeling and preclinical evaluation of high affinity ligands for the ion channel of the N-methyl-d-aspartate receptor as potential imaging probes for positron emission tomography.

    Science.gov (United States)

    Klein, Pieter J; Christiaans, Johannes A M; Metaxas, Athanasios; Schuit, Robert C; Lammertsma, Adriaan A; van Berckel, Bart N M; Windhorst, Albert D

    2015-03-01

    The N-methyl-d-aspartate receptor (NMDAr) is involved in many neurological and psychiatric disorders including Alzheimer's disease and schizophrenia. Currently, it is not possible to assess NMDAr availability in vivo. The purpose of this study was to develop a positron emission tomography (PET) ligand for the NMDAr ion channel. A series of di- and tri-N-substituted diarylguanidines was synthesized. In addition, in vitro binding affinity for the NMDAr ion channel in rat forebrain membrane fractions was assessed. Compounds 10, 11 and 32 were radiolabeled with either carbon-11 or fluorine-18. Ligands [(11)C]10 and [(18)F]32 were evaluated ex vivo in B6C3 mice. Biodistribution studies showed higher uptake of [(11)C]10 and [(18)F]32 in forebrain regions compared with cerebellum. In addition, for [(11)C]10 54% and for [(18)F]32 70% of activity in the brain at 60min was due to intact tracer. Pre-treatment with MK-801 (0.6mg·kg(-1), ip) slightly decreased uptake in NMDAr-specific regions for [(18)F]32, but not for [(11)C]10. As such [(18)F]32 has the best characteristics as a PET tracer for the ion channel of the NMDAr. PMID:25648682

  11. The peripheral benzodiazepine receptor ligand PK11195 binds with high affinity to the acute phase reactant α1-acid glycoprotein: implications for the use of the ligand as a CNS inflammatory marker

    International Nuclear Information System (INIS)

    The peripheral benzodiazepine receptor ligand PK11195 has been used as an in vivo marker of neuroinflammation in positron emission tomography studies in man. One of the methodological issues surrounding the use of the ligand in these studies is the highly variable kinetic behavior of [11C]PK11195 in plasma. We therefore undertook a study to measure the binding of [3H]PK11195 to whole human blood and found a low level of binding to blood cells but extensive binding to plasma proteins. Binding assays using [3H]PK11195 and purified human plasma proteins demonstrated a strong binding to α1-acid glycoprotein (AGP) and a much weaker interaction with albumin. Immunodepletion of AGP from plasma resulted in the loss of plasma [3H]PK11195 binding demonstrating: (i) the specificity of the interaction and (ii) that AGP is the major plasma protein to which PK11195 binds with high affinity. PK11195 was able to displace fluorescein-dexamethasone from AGP with IC50 of 11C]PK11195 to the brain parenchyma in diseases with blood brain barrier breakdown. Finally, local synthesis of AGP at the site of brain injury may contribute the pattern of [11C]PK11195 binding observed in neuroinflammatory diseases

  12. Affine and Projective Geometry

    CERN Document Server

    Bennett, M K

    1995-01-01

    An important new perspective on AFFINE AND PROJECTIVE GEOMETRY. This innovative book treats math majors and math education students to a fresh look at affine and projective geometry from algebraic, synthetic, and lattice theoretic points of view. Affine and Projective Geometry comes complete with ninety illustrations, and numerous examples and exercises, covering material for two semesters of upper-level undergraduate mathematics. The first part of the book deals with the correlation between synthetic geometry and linear algebra. In the second part, geometry is used to introduce lattice theory

  13. IgE sensitization against food allergens : Natural history, relation to airway inflammation and asthma

    OpenAIRE

    Patelis, Antonios

    2015-01-01

    Background: According to recent studies in children, IgE sensitization not only against perennial allergens, but also against food allergens, is related to asthma risk and increased airway inflammation. During the last decade, a new technique for IgE determination based on allergen components has become available, but its use in epidemiological studies has been limited. Aims: To investigate the relationship between the pattern of IgE sensitization to allergen components and the prevalence of ...

  14. Helminth-Induced IgE and Protection Against Allergic Disorders

    OpenAIRE

    Hamid, Firdaus; Amoah, Abena S.; van Ree, Ronald; Yazdanbakhsh, Maria

    2015-01-01

    The immune response against helminths and allergens is generally characterized by high levels of IgE and increased numbers of Th2 cells, eosinophils, and mast cells, yet the clinical outcome with respect to immediate hypersensitivity and inflammation is clearly not the same. High levels of IgE are seen to allergens during helminth infections; however, these IgE responses do not translate into allergy symptoms. This chapter summarizes the evidence of the association between helminth infection...

  15. Helminth-induced IgE and protection against allergic disorders

    OpenAIRE

    Firdaus

    2015-01-01

    he immune response against helminths and allergens is generally characterized by high levels of IgE and increased numbers of Th2 cells, eosinophils, and mast cells, yet the clinical outcome with respect to immediate hypersensitivity and inflammation is clearly not the same. High levels of IgE are seen to allergens during helminth infections; however, these IgE responses do not translate into allergy symptoms. This chapter summarizes the evidence of the association between helminth infections ...

  16. Affine dynamics with torsion

    Energy Technology Data Exchange (ETDEWEB)

    Gueltekin, Kemal [Izmir Institute of Technology, Department of Physics, Izmir (Turkey)

    2016-03-15

    In this study, we give a thorough analysis of a general affine gravity with torsion. After a brief exposition of the affine gravities considered by Eddington and Schroedinger, we construct and analyze different affine gravities based on the determinants of the Ricci tensor, the torsion tensor, the Riemann tensor, and their combinations. In each case we reduce equations of motion to their simplest forms and give a detailed analysis of their solutions. Our analyses lead to the construction of the affine connection in terms of the curvature and torsion tensors. Our solutions of the dynamical equations show that the curvature tensors at different points are correlated via non-local, exponential rescaling factors determined by the torsion tensor. (orig.)

  17. Study of IL-10 and IgE levels in the serum of patients with asthma

    International Nuclear Information System (INIS)

    Using ELISA and IRMA respectively detected IL-10 and IgE levels in the serum of 50 patients with acute asthma and 48 healthy persons. Results show IL-10 levels in the serum of patients with asthma is obviously lower than that of healthy persons (P<0.01), but IgE levels in them are obviously higher than that of healthy persons (P<0.01), there is negative correlation between IL-10 and IgE (r=-0.18, P<0.01). Suggesting that IL-10 is insufficient and IgE is high in patients with asthma may be one of important causes of asthma

  18. Affine and degenerate affine BMW algebras: Actions on tensor space

    CERN Document Server

    Daugherty, Zajj; Virk, Rahbar

    2012-01-01

    The affine and degenerate affine Birman-Murakami-Wenzl (BMW) algebras arise naturally in the context of Schur-Weyl duality for orthogonal and symplectic quantum groups and Lie algebras, respectively. Cyclotomic BMW algebras, affine and cyclotomic Hecke algebras, and their degenerate versions are quotients. In this paper we explain how the affine and degenerate affine BMW algebras are tantalizers (tensor power centralizer algebras) by defining actions of the affine braid group and the degenerate affine braid algebra on tensor space and showing that, in important cases, these actions induce actions of the affine and degenerate affine BMW algebras. We then exploit the connection to quantum groups and Lie algebras to determine universal parameters for the affine and degenerate affine BMW algebras. Finally, we show that the universal parameters are central elements--the higher Casimir elements for orthogonal and symplectic enveloping algebras and quantum groups.

  19. IgE mediates killing of intracellular Toxoplasma gondii by human macrophages through CD23-dependent, interleukin-10 sensitive pathway.

    Directory of Open Access Journals (Sweden)

    Ioannis Vouldoukis

    Full Text Available BACKGROUND: In addition to helminthic infections, elevated serum IgE levels were observed in many protozoal infections, while their contribution during immune response to these pathogens remained unclear. As IgE/antigen immune complexes (IgE-IC bind to human cells through FcεRI or FcεRII/CD23 surface molecules, the present study aimed to identify which functional receptor may be involved in IgE-IC interaction with human macrophages, the major effector cell during parasite infection. METHODOLOGY/PRINCIPAL FINDINGS: Human monocyte-derived macrophages were infected with Toxoplasma gondii before being incubated with IgE-IC. IgE receptors were then identified using appropriate blocking antibodies. The activation of cells and parasiticidal activity were evaluated by mediator quantification and direct counting of infected macrophages. RNAs were extracted and cell supernatants were also collected for their content in tumor necrosis factor (TNF-α, interleukin-10 (IL-10 and nitrites. Sera from symptomatic infected patients were also tested for their content of IgE, IL-10 and nitrites, and compared to values found in healthy donors. Results showed that IgE-IC induced intracellular elimination of parasites by human macrophages. IgE-mediated effect was FcεRI-independent, but required cross-linking of surface FcεRII/CD23, cell activation and the generation of nitric oxide (NO. Although TNF-α was shown to be produced during cell activation, this cytokine had minor contribution in this phenomenon while endogenous and exogenous IL-10 down-regulated parasite killing. Inverse relationship was found between IL-10 and NO expression by infected human macrophages at both mRNA and mediator levels. The relationship between these in vitro data and in vivo levels of various factors in T. gondii infected patients supports the involvement of CD23 antigen and IL-10 expression in disease control. CONCLUSION: Thus, IgE may be considered as immune mediator during

  20. High affinity dopamine D2 receptor radioligands. 2. ( sup 125 I)epidepride, a potent and specific radioligand for the characterization of striatal and extrastriatal dopamine D2 receptors

    Energy Technology Data Exchange (ETDEWEB)

    Kessler, R.M.; Ansari, M.S.; Schmidt, D.E.; de Paulis, T.; Clanton, J.A.; Manning, R.G.; Gillespie, D. (Vanderbilt Univ., Nashville, TN (United States)); Innis, R.; Al-Tikriti, M. (Yale Univ., New Haven, CT (United States))

    1991-01-01

    Epidepride, (S)-N-((1-ethyl-2-pyrrolidinyl)methyl)-5-iodo-2,3-dimethoxybenzamide, the iodine analogue of isoremoxipride (FLB 457), was found to be a very potent dopamine D2 receptor antagonist. Optimal in vitro binding required incubation at 25C for 4 h at pH 7.4 in a buffer containing 120 mM NaCl, 5 mM KCl, 2 mM CaCl{sub 2} and 1 nM MgCl{sub 2}. Scatchard analysis of in vitro binding to striatal, medical frontal cortical, hippocampal and cerebellar membranes revealed a K{sub D} of 24 pM in all regions, with Bmax's of 36.7, 1.04, 0.85, and 0.37 pmol/g tissue, respectively. The Hill coefficients ranged from 0.91-1.00 in all four regions. The IC{sub 50}'s for inhibition of ({sup 125}I)epidepride binding to striatal, medial frontal cortical, and hippocampal membranes for SCH 23390, SKF 83566, serotonin, ketanserin, mianserin, naloxone, QNB, prasozin, clonidine, alprenolol, and norepinephrine ranged from 1 {mu}M to >10 {mu}M. Partial displacement of ({sup 125}I)epidepride by nanomolar concentrations of clonidine was noted in the frontal cortex and hippocampus, but not in the striatum. Scatchard analysis of epidepride binding to {alpha}{sub 2} noradrenergic receptors in the frontal cortex and hippocampus revealed an apparent K{sub D} of 9 nM. At an epidepride concentration equal to the K{sub D} for the D2 receptor, i.e., 25 pM, no striatal {alpha}{sub 2} binding was seen and only 7% of the specific epidepride binding in the cortex or hippocampus was due to binding at the {alpha}{sub 2} site. Correlation of inhibition of ({sup 3}H)spiperone and ({sup 125}I)epidepride binding to striatal membranes by a variety of D2 ligands revealed a correlation coefficient of 0.99, indicating that epidepride labels a D2 site.

  1. Structure-guided design of a high-affinity platelet integrin αIIbβ3 receptor antagonist that disrupts Mg²⁺ binding to the MIDAS.

    Science.gov (United States)

    Zhu, Jieqing; Choi, Won-Seok; McCoy, Joshua G; Negri, Ana; Zhu, Jianghai; Naini, Sarasija; Li, Jihong; Shen, Min; Huang, Wenwei; Bougie, Daniel; Rasmussen, Mark; Aster, Richard; Thomas, Craig J; Filizola, Marta; Springer, Timothy A; Coller, Barry S

    2012-03-14

    An integrin found on platelets, α(IIb)β(3) mediates platelet aggregation, and α(IIb)β(3) antagonists are effective antithrombotic agents in the clinic. Ligands bind to integrins in part by coordinating a magnesium ion (Mg(2+)) located in the β subunit metal ion-dependent adhesion site (MIDAS). Drugs patterned on the integrin ligand sequence Arg-Gly-Asp have a basic moiety that binds the α(IIb) subunit and a carboxyl group that coordinates the MIDAS Mg(2+) in the β(3) subunits. They induce conformational changes in the β(3) subunit that may have negative consequences such as exposing previously hidden epitopes and inducing the active conformation of the receptor. We recently reported an inhibitor of α(IIb)β(3) (RUC-1) that binds exclusively to the α(IIb) subunit; here, we report the structure-based design and synthesis of RUC-2, a RUC-1 derivative with a ~100-fold higher affinity. RUC-2 does not induce major conformational changes in β(3) as judged by monoclonal antibody binding, light scattering, gel chromatography, electron microscopy, and a receptor priming assay. X-ray crystallography of the RUC-2-α(IIb)β(3) headpiece complex in 1 mM calcium ion (Ca(2+))/5 mM Mg(2+) at 2.6 Å revealed that RUC-2 binds to α(IIb) the way RUC-1 does, but in addition, it binds to the β(3) MIDAS residue glutamic acid 220, thus displacing Mg(2+) from the MIDAS. When the Mg(2+) concentration was increased to 20 mM, however, Mg(2+) was identified in the MIDAS and RUC-2 was absent. RUC-2's ability to inhibit ligand binding and platelet aggregation was diminished by increasing the Mg(2+) concentration. Thus, RUC-2 inhibits ligand binding by a mechanism different from that of all other α(IIb)β(3) antagonists and may offer advantages as a therapeutic agent. PMID:22422993

  2. Receptor-targeted metalloradiopharmaceuticals. Final technical report

    International Nuclear Information System (INIS)

    Copper (II) and platinum (II) coordination complexes were prepared and characterized. These complexes were designed to afford structural homology with steroidal and non-steroidal estrogens for possible use as receptor-targeted radiopharmaceuticals. While weak affinity for the estrogen receptor was detectable, none would appear to have sufficient receptor-affinity for estrogen-receptor-targeted imaging or therapy

  3. Prediction of Elevated Cord Blood IgE Levels by Maternal IgE Levels, and the Neonate's Gender and Gestational Age.

    Directory of Open Access Journals (Sweden)

    Chieh-An Liu

    2003-08-01

    Full Text Available Background: Elevation of cord blood immunoglobulin E (IgE is used to predict childhoodatopy. In an effort to catch such problems at an earlier time, we sought toidentify the impact of parental atopy as well as gestational age, the baby'sgender, and the season of delivery on cord blood IgE levels.Methods: The allergic history of parents was collected during pregnancy. Blood samplesfrom parents were collected in the third trimester. Cord blood was collectedimmediately after birth. Total and specific IgE levels were determinedusing the Pharmacia CAP system.Results: In total, 437 core family blood samples were collected. Male babies had asignificantly higher IgE level (0.535

  4. Control of IgE and IgGl antibody production in mice

    International Nuclear Information System (INIS)

    The production of IgE and IgCl was studied in untreated, thymectomized, splenectomized, anti-thymocyte serum-treated, or sublethally X-irradiated mice. Dinitrophenyl, Ascaris, and ovalbumin were used as antigens, and aluminum hydroxide was used as adjuvant. A suppression of IgE production was observed in adult thymectomized mice, although the kinetic pattern of the antibody response was the same as in control animals. IgGl antibody production was not affected by thymectomy. Splenectomy did not change either IgE or IgGl production. A single dose of rabbit antithymocyte serum (ATS) given 8 days after immunization inhibited IgE antibody production. The effect of ATS was dose dependent and also varied with the amount of antigen used, the immune response to high doses being more susceptible to the effect of ATS. No alteration in IgGl production was caused by ATS even when IgE antibody formation was completely inhibited. When preceding immunization, sublethal irradiation enhanced IgE antibody formation and partially suppressed IgGl production; applied after immunization, irradiation caused an enhancement of IgE production which was inversely proportional to the interval elapsed between the two procedures. On the other hand, the IgGl antibody production was fairly resistant to the same treatment. The results suggest a clearcut separation between the mechanisms regulating IgE and IgGl production in mice

  5. Rat liver insulin receptor

    International Nuclear Information System (INIS)

    Using insulin affinity chromatography, the authors have isolated highly purified insulin receptor from rat liver. When evaluated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis under reducing conditions, the rat liver receptor contained the M/sub r/ 125,000 α-subunit, the M/sub r/ 90,000 β-subunit, and varying proportions of the M/sub r/ 45,000 β'-subunit. The specific insulin binding of the purified receptor was 25-30 μg of 125I-insulin/mg of protein, and the receptor underwent insulin-dependent autophosphorylation. Rat liver and human placental receptors differ from each other in several functional aspects: (1) the adsorption-desorption behavior from four insulin affinity columns indicated that the rat liver receptor binds less firmly to immobilized ligands; (2) the 125I-insulin binding affinity of the rat liver receptor is lower than that of the placental receptor; (3) partial reduction of the rat liver receptor with dithiothreitol increases its insulin binding affinity whereas the binding affinity of the placental receptor is unchanged; (4) at optimal insulin concentration, rat liver receptor autophosphorylation is stimulated 25-50-fold whereas the placental receptor is stimulated only 4-6-fold. Conversion of the β-subunit to β' by proteolysis is a major problem that occurs during exposure of the receptor to the pH 5.0 buffer used to elute the insulin affinity column. Proteolytic destruction and the accompanying loss of insulin-dependent autophosphorylation can be substantially reduced by proteolysis inhibitors. In summary, rat liver and human placental receptors differ functionally in both α- and β-subunits. Insulin binding to the α-subunit of the purified rat liver receptor communicates a signal that activates the β-subunit; however, major proteolytic destruction of the β-subunit does not affect insulin binding to the α-subunit

  6. Essential role of Id2 in negative regulation of IgE class switching.

    Science.gov (United States)

    Sugai, Manabu; Gonda, Hiroyuki; Kusunoki, Takashi; Katakai, Tomoya; Yokota, Yoshifumi; Shimizu, Akira

    2003-01-01

    Serum concentrations of immunoglobulin E (IgE) in normal circumstances are kept much lower than those of other Ig isotypes to avoid allergic reactions. B cells lacking Id2 have increased E2A activity, which leads to specific enhancement of germline transcription of the immunoglobulin epsilon locus. As a consequence, Id2-deficient B cells undergo class switch recombination (CSR) to IgE at a much higher frequency than wild-type B cells. In contrast, Id2 is induced in wild-type B cells by transforming growth factor-beta1 (TGF-beta1) and suppresses IgE CSR. Our results provide evidence for the inhibitory and selective role of Id2 in IgE CSR in response to TGF-beta1. Id2 might act as molecular safeguard to suppress IgE CSR to prevent serious complications such as allergic hypersensitivity during the normal course of immune responses. PMID:12483209

  7. Helminth-induced IgE and protection against allergic disorders.

    Science.gov (United States)

    Hamid, Firdaus; Amoah, Abena S; van Ree, Ronald; Yazdanbakhsh, Maria

    2015-01-01

    The immune response against helminths and allergens is generally characterized by high levels of IgE and increased numbers of Th2 cells, eosinophils, and mast cells, yet the clinical outcome with respect to immediate hypersensitivity and inflammation is clearly not the same. High levels of IgE are seen to allergens during helminth infections; however, these IgE responses do not translate into allergy symptoms. This chapter summarizes the evidence of the association between helminth infections and allergic disorders. It discusses how helminth infection can lead to IgE cross-reactivity with allergens and how this IgE has poor biological activity. This information is important for developing new diagnostic methods and treatments for allergic disorders in low-to-middle-income countries. PMID:25553796

  8. Antibody affinity maturation

    DEFF Research Database (Denmark)

    Skjødt, Mette Louise

    Yeast surface display is an effective tool for antibody affinity maturation because yeast can be used as an all-in-one workhorse to assemble, display and screen diversified antibody libraries. By employing the natural ability of yeast Saccharomyces cerevisiae to efficiently recombine multiple DNA...

  9. receptores

    Directory of Open Access Journals (Sweden)

    Salete Regina Daronco Benetti

    2006-01-01

    Full Text Available Se trata de un estudio etnográfico, que tuvo lo objetivo de interpretar el sistema de conocimiento y del significado atribuidos a la sangre referente a la transfusión sanguínea por los donadores y receptores de un banco de sangre. Para la colecta de las informaciones se observaron los participantes y la entrevista etnográfica se realizó el análisis de dominio, taxonómicos y temáticos. Los dominios culturales fueron: la sangre es vida: fuente de vida y alimento valioso; creencias religiosas: fuentes simbólicas de apoyos; donación sanguínea: un gesto colaborador que exige cuidarse, gratifica y trae felicidad; donación sanguínea: fuente simbólica de inseguridad; estar enfermo es una condición para realizar transfusión sanguínea; transfusión sanguínea: esperanza de vida; Creencias populares: transfusión sanguínea como riesgo para la salud; donadores de sangre: personas benditas; donar y recibir sangre: como significado de felicidad. Temática: “líquido precioso que origina, sostiene, modifica la vida, provoca miedo e inseguridad”.

  10. Nanoparticles rapidly assess specific IgE in plasma

    International Nuclear Information System (INIS)

    Allergy is the sixth leading cause of chronic disease in the world. This study demonstrates the feasibility of detecting allergy indicators in human plasma, noninvasively, at the point of care and with a comparable efficiency and reduced turnaround time compared with the gold standard. Peanut allergy was utilized as a model due to its widespread occurrence among the US population and fatality if not treated. The detection procedure utilized magnetic nanoparticles that were coated with an allergen layer (peanut protein extract). Peanut immunoglobulin E (IgE) was detected in concentrations close to the minimum detection range of CAP assay. The results were obtained in minutes compared with the CAP assay which requires more than 3 h. (paper)

  11. Surface-modified magnetic colloids for affinity adsorption of immunoglobulins

    Energy Technology Data Exchange (ETDEWEB)

    Martins, Fernanda [School of Chemical Engineering, State University of Campinas, C.P. 6066, 13083-970 Campinas-SP (Brazil); Pinho, Samantha C. [School of Chemical Engineering, State University of Campinas, C.P. 6066, 13083-970 Campinas-SP (Brazil)], E-mail: samantha@usp.br; Zollner, Terezinha C.A. [School of Chemical Engineering, State University of Campinas, C.P. 6066, 13083-970 Campinas-SP (Brazil); Zollner, Ricardo L. [School of Medical Sciences, State University of Campinas, Campinas-SP (Brazil)], E-mail: zollner@unicamp.br; Cuyper, Marcel de [Interdisciplinary Research Centre, Katholieke Universiteit Leuven-Campus Kortrijk, B-8500 Kortrijk (Belgium)], E-mail: Marcel.DeCuyper@kulak.ac.be; Santana, Maria Helena A. [School of Chemical Engineering, State University of Campinas, C.P. 6066, 13083-970 Campinas-SP (Brazil)], E-mail: lena@feq.unicamp.br

    2008-07-15

    This work describes the preparation, characterization and in vitro adsorption tests of surface-modified magnetoliposomes for affinity binding of (i) anticardiolipin (isotype G) antibodies and (ii) specific isotype E antibodies generated by hypersensitivity reactions in humans with respiratory allergy. In the first case, cardiolipin embedded in the bilayer of magnetoliposomes was used as specific ligand. In the second case, antigenic proteins present in an extract of Dermatophagoids pteronyssinus and Blomia tropicalis mites were covalently coupled on the surface of magnetoliposomes via a diglycolic spacer arm, and used as specific ligands for IgE. Antibody adsorption was performed in a high-gradient magnetophoresis system, using either sera of healthy individuals or a pool of sera from autoimmune or allergic patients. The selectivity and capacity of the system were quantified by a frontal analysis in a capillary column, and by constructing breakthrough curves. The results show that the highest yield and selectivity were obtained if the ligand was extended into the aqueous layer surrounding the magnetoliposome surface. A 100% selectivity was obtained for adsorption of specific IgE, and 8% for IgG. These results demonstrate the potentialities of both types of surface-modified magnetic biocolloids in the field of in vitro diagnosis tests for allergic or autoimmune conditions.

  12. Surface-modified magnetic colloids for affinity adsorption of immunoglobulins

    International Nuclear Information System (INIS)

    This work describes the preparation, characterization and in vitro adsorption tests of surface-modified magnetoliposomes for affinity binding of (i) anticardiolipin (isotype G) antibodies and (ii) specific isotype E antibodies generated by hypersensitivity reactions in humans with respiratory allergy. In the first case, cardiolipin embedded in the bilayer of magnetoliposomes was used as specific ligand. In the second case, antigenic proteins present in an extract of Dermatophagoids pteronyssinus and Blomia tropicalis mites were covalently coupled on the surface of magnetoliposomes via a diglycolic spacer arm, and used as specific ligands for IgE. Antibody adsorption was performed in a high-gradient magnetophoresis system, using either sera of healthy individuals or a pool of sera from autoimmune or allergic patients. The selectivity and capacity of the system were quantified by a frontal analysis in a capillary column, and by constructing breakthrough curves. The results show that the highest yield and selectivity were obtained if the ligand was extended into the aqueous layer surrounding the magnetoliposome surface. A 100% selectivity was obtained for adsorption of specific IgE, and 8% for IgG. These results demonstrate the potentialities of both types of surface-modified magnetic biocolloids in the field of in vitro diagnosis tests for allergic or autoimmune conditions

  13. Surface-modified magnetic colloids for affinity adsorption of immunoglobulins

    Science.gov (United States)

    Martins, Fernanda; Pinho, Samantha C.; Zollner, Terezinha C. A.; Zollner, Ricardo L.; de Cuyper, Marcel; Santana, Maria Helena A.

    This work describes the preparation, characterization and in vitro adsorption tests of surface-modified magnetoliposomes for affinity binding of (i) anticardiolipin (isotype G) antibodies and (ii) specific isotype E antibodies generated by hypersensitivity reactions in humans with respiratory allergy. In the first case, cardiolipin embedded in the bilayer of magnetoliposomes was used as specific ligand. In the second case, antigenic proteins present in an extract of Dermatophagoids pteronyssinus and Blomia tropicalis mites were covalently coupled on the surface of magnetoliposomes via a diglycolic spacer arm, and used as specific ligands for IgE. Antibody adsorption was performed in a high-gradient magnetophoresis system, using either sera of healthy individuals or a pool of sera from autoimmune or allergic patients. The selectivity and capacity of the system were quantified by a frontal analysis in a capillary column, and by constructing breakthrough curves. The results show that the highest yield and selectivity were obtained if the ligand was extended into the aqueous layer surrounding the magnetoliposome surface. A 100% selectivity was obtained for adsorption of specific IgE, and 8% for IgG. These results demonstrate the potentialities of both types of surface-modified magnetic biocolloids in the field of in vitro diagnosis tests for allergic or autoimmune conditions.

  14. Affine and degenerate affine BMW algebras: The center

    CERN Document Server

    Daugherty, Zajj; Virk, Rahbar

    2011-01-01

    The degenerate affine and affine BMW algebras arise naturally in the context of Schur-Weyl duality for orthogonal and symplectic Lie algebras and quantum groups, respectively. Cyclotomic BMW algebras, affine Hecke algebras, cyclotomic Hecke algebras, and their degenerate versions are quotients. In this paper the theory is unified by treating the orthogonal and symplectic cases simultaneously; we make an exact parallel between the degenerate affine and affine cases via a new algebra which takes the role of the affine braid group for the degenerate setting. A main result of this paper is an identification of the centers of the affine and degenerate affine BMW algebras in terms of rings of symmetric functions which satisfy a "cancellation property" or "wheel condition" (in the degenerate case, a reformulation of a result of Nazarov). Miraculously, these same rings also arise in Schubert calculus, as the cohomology and K-theory of isotropic Grassmanians and symplectic loop Grassmanians. We also establish new inte...

  15. The 11S globulin Sin a 2 from yellow mustard seeds shows IgE cross-reactivity with homologous counterparts from tree nuts and peanut

    Directory of Open Access Journals (Sweden)

    Sirvent Sofía

    2012-12-01

    Full Text Available Abstract Background The 11S globulin Sin a 2 is a marker to predict severity of symptoms in mustard allergic patients. The potential implication of Sin a 2 in cross-reactivity with tree nuts and peanut has not been investigated so far. In this work, we studied at the IgG and IgE level the involvement of the 11S globulin Sin a 2 in cross-reactivity among mustard, tree nuts and peanut. Methods Eleven well-characterized mustard-allergic patients sensitized to Sin a 2 were included in the study. A specific anti-Sin a 2 serum was obtained in rabbit. Skin prick tests (SPT, enzyme-linked immunosorbent assay (ELISA, immunoblotting and IgG or IgE-inhibition immunoblotting experiments using purified Sin a 2, Sin a 1, Sin a 3, mustard, almond, hazelnut, pistachio, walnut or peanut extracts were performed. Results The rabbit anti-Sin a 2 serum showed high affinity and specificity to Sin a 2, which allowed us to demonstrate that Sin a 2 shares IgG epitopes with allergenic 11S globulins from tree nuts (almond, hazelnut, pistachio and walnut but not from peanut. All the patients included in the study had positive skin prick test to tree nuts and/or peanut and we subdivided them into two different groups according to their clinical symptoms after ingestion of such allergenic sources. We showed that 11S globulins contain conserved IgE epitopes involved in cross-reactivity among mustard, tree nuts and peanut as well as species-specific IgE epitopes. Conclusions The allergenic 11S globulin Sin a 2 from mustard is involved in cross-reactivity at the IgE level with tree nuts and peanut. Although the clinical relevance of the cross-reactive IgE epitopes present in 11S globulins needs to be investigated in further detail, our results contribute to improve the diagnosis and management of mustard allergic patients sensitized to Sin a 2.

  16. Hierarchical Affinity Propagation

    CERN Document Server

    Givoni, Inmar; Frey, Brendan J

    2012-01-01

    Affinity propagation is an exemplar-based clustering algorithm that finds a set of data-points that best exemplify the data, and associates each datapoint with one exemplar. We extend affinity propagation in a principled way to solve the hierarchical clustering problem, which arises in a variety of domains including biology, sensor networks and decision making in operational research. We derive an inference algorithm that operates by propagating information up and down the hierarchy, and is efficient despite the high-order potentials required for the graphical model formulation. We demonstrate that our method outperforms greedy techniques that cluster one layer at a time. We show that on an artificial dataset designed to mimic the HIV-strain mutation dynamics, our method outperforms related methods. For real HIV sequences, where the ground truth is not available, we show our method achieves better results, in terms of the underlying objective function, and show the results correspond meaningfully to geographi...

  17. Two loci on chromosome 5 are associated with serum IgE levels in Labrador retrievers.

    Directory of Open Access Journals (Sweden)

    Marta Owczarek-Lipska

    Full Text Available Crosslinking of immunoglobulin E antibodies (IgE bound at the surface of mast cells and subsequent mediator release is considered the most important trigger for allergic reactions. Therefore, the genetic control of IgE levels is studied in the context of allergic diseases, such as asthma, atopic rhinitis, or atopic dermatitis (AD. We performed genome-wide association studies in 161 Labrador Retrievers with regard to total and allergen-specific immunoglobulin E (IgE levels. We identified a genome-wide significant association on CFA 5 with the antigen-specific IgE responsiveness to Acarus siro. We detected a second genome-wide significant association with respect to the antigen-specific IgE responsiveness to Tyrophagus putrescentiae at a different locus on chromosome 5. A. siro and T. putrescentiae both belong to the family Acaridae and represent so-called storage or forage mites. These forage mites are discussed as major allergen sources in canine AD. No obvious candidate gene for the regulation of IgE levels is located under the two association signals. Therefore our studies offer a chance of identifying a novel mechanism controlling the host's IgE response.

  18. IgE epitope proximity determines immune complex shape and effector cell activation capacity

    Science.gov (United States)

    Gieras, Anna; Linhart, Birgit; Roux, Kenneth H.; Dutta, Moumita; Khodoun, Marat; Zafred, Domen; Cabauatan, Clarissa R.; Lupinek, Christian; Weber, Milena; Focke-Tejkl, Margarete; Keller, Walter; Finkelman, Fred D.; Valenta, Rudolf

    2016-01-01

    Background IgE-allergen complexes induce mast cell and basophil activation and thus immediate allergic inflammation. They are also important for IgE-facilitated allergen presentation to T cells by antigen-presenting cells. Objective To investigate whether the proximity of IgE binding sites on an allergen affects immune complex shape and subsequent effector cell activation in vitro and in vivo. Methods We constructed artificial allergens by grafting IgE epitopes in different numbers and proximity onto a scaffold protein. The shape of immune complexes formed between artificial allergens and the corresponding IgE was studied by negative-stain electron microscopy. Allergenic activity was determined using basophil activation assays. Mice were primed with IgE, followed by injection of artificial allergens to evaluate their in vivo allergenic activity. Severity of systemic anaphylaxis was measured by changes in body temperature. Results We could demonstrate simultaneous binding of 4 IgE antibodies in close vicinity to each other. The proximity of IgE binding sites on allergens influenced the shape of the resulting immune complexes and the magnitude of effector cell activation and in vivo inflammation. Conclusions Our results demonstrate that the proximity of IgE epitopes on an allergen affects its allergenic activity. We thus identified a novel mechanism by which IgE-allergen complexes regulate allergic inflammation. This mechanism should be important for allergy and other immune complex–mediated diseases. PMID:26684291

  19. Affinity driven social networks

    Science.gov (United States)

    Ruyú, B.; Kuperman, M. N.

    2007-04-01

    In this work we present a model for evolving networks, where the driven force is related to the social affinity between individuals of a population. In the model, a set of individuals initially arranged on a regular ordered network and thus linked with their closest neighbors are allowed to rearrange their connections according to a dynamics closely related to that of the stable marriage problem. We show that the behavior of some topological properties of the resulting networks follows a non trivial pattern.

  20. Affine General Equilibrium Models

    OpenAIRE

    Bjørn Eraker

    2008-01-01

    No-arbitrage models are extremely flexible modelling tools but often lack economic motivation. This paper describes an equilibrium consumption-based CAPM framework based on Epstein-Zin preferences, which produces analytic pricing formulas for stocks and bonds under the assumption that macro growth rates follow affine processes. This allows the construction of equilibrium pricing formulas while maintaining the same flexibility of state dynamics as in no-arbitrage models. In demonstrating the a...

  1. Gaussian Affine Feature Detector

    OpenAIRE

    Xu, Xiaopeng; Zhang, Xiaochun

    2011-01-01

    A new method is proposed to get image features' geometric information. Using Gaussian as an input signal, a theoretical optimal solution to calculate feature's affine shape is proposed. Based on analytic result of a feature model, the method is different from conventional iterative approaches. From the model, feature's parameters such as position, orientation, background luminance, contrast, area and aspect ratio can be extracted. Tested with synthesized and benchmark data, the method achieve...

  2. Total IgE Distribution in Food Allergy Suspected Patients in Republic of Macedonia (2001-2011

    Directory of Open Access Journals (Sweden)

    Slavica Hristomanova Mitkovska

    2015-04-01

    CONCLUSION: The large number of enrolled patients, a particular strength of this study, revealed that average concentrations of total IgE in men are higher than in women and that total IgE did not decrease with age. On the contrary, increased total IgE levels were found in patients aged 65 and 69 of both genders. We continue our work with analyses of the specific IgE antibodies values toward food and the correlation with total IgE values.

  3. On the Affine Isoperimetric Inequalities

    Indian Academy of Sciences (India)

    Wuyang Yu; Gangsong Leng

    2011-11-01

    We obtain an isoperimetric inequality which estimate the affine invariant -surface area measure on convex bodies. We also establish the reverse version of -Petty projection inequality and an affine isoperimetric inequality of $_{-p}K$.

  4. Contribution of Phadebas IgE test to typification of allergic patients

    International Nuclear Information System (INIS)

    Experience is reported with the examination of serum immunoglobulin E in a control group (n=22), in a group of asthmatics with bacterial allergy (n=25) and in a group of asthmatics with antibody allergy (n=33). In the examination by radioimmunosorbent method (Phadebas IgE Test) the levels of IgE over 500 mU/ml were considered as proof of allergy of the antibody type where other etiological factors could be excluded. A marked immunosuppressive effect of corticoids on the level of IgE is pointed out. The method mentioned is considered as suitable for the objective typification of allergic patients. (author)

  5. SERUM IgE LEVEL IN MICE INFECTED WITH SINGLE DOSES OF ANCYLOSTOMA CANINUM LARVAE

    Directory of Open Access Journals (Sweden)

    V. VIVEKA VARDHANI

    2013-01-01

    Full Text Available The level of IgE in the serum of singly infected mice (group A, 500 dose; B, 1000 dose; group C, 2000 dose wasstudied during murine ancylostomiasis in all the 3 groups of singly infected animals, IgE level began to decreaseon 1st day of infection, reached the lowest value on day 30 and progressively declined from day 1 to 30 ofexperimental period. Statistical analysis indicated a significant decrease in all the 3 groups when compared withcontrols. These studies demonstrated that the larval antigens provoked an immune response resulting into a lowlevel of serum IgE in the mouse system.

  6. Specific IgE Anti-Ascaris in Brazilian Children and Adolescents

    OpenAIRE

    Emanuel S. Sarinho; Medeiros, Décio; Silva, Almerinda; Rizzo, José Ângelo

    2010-01-01

    From an article published by our group by Medeiros et al, we discuss and review the literature on the role of serum specific anti-ascaris IgE in patients with respiratory allergies living in countries where helminthic infestations are common. Medeiros et al conducted a study using 101 patients aged 12 to 21 years with respiratory allergy. Median IgE level was 660 IU/mL. Serum specific anti-ascaris IgE was positive in 73% (74/101) of the individuals, but parasitological stool examination yield...

  7. IgE Mediated Autoallergy against Thyroid Peroxidase – A Novel Pathomechanism of Chronic Spontaneous Urticaria?

    OpenAIRE

    Altrichter, Sabine; Peter, Hans-Jürgen; Pisarevskaja, Dina; Metz, Martin; Martus, Peter; Maurer, Marcus

    2011-01-01

    Background Chronic spontaneous urticaria (csU), which is characterized by recurrent episodes of mast cell-driven wheal and flare-type skin reactions, is often associated with elevated total IgE levels and thyroid autoimmunity. We speculate that some csU patients express IgE autoantibodies against thyroid antigens such as thyroid peroxidase (TPO), which could bind to skin mast cells and induce their activation. Methods We developed and used a site-directed human IgE capture ELISA to quantify I...

  8. Adjoint affine fusion and tadpoles

    OpenAIRE

    Urichuk, Andrew; Walton, Mark A.

    2016-01-01

    We study affine fusion with the adjoint representation. For simple Lie algebras, elementary and universal formulas determine the decomposition of a tensor product of an integrable highest-weight representation with the adjoint representation. Using the (refined) affine depth rule, we prove that equally striking results apply to adjoint affine fusion. For diagonal fusion, a coefficient equals the number of nonzero Dynkin labels of the relevant affine highest weight, minus 1. A nice lattice-pol...

  9. An IgE epitope of Bet v 1 and fagales PR10 proteins as defined by a human monoclonal IgE

    DEFF Research Database (Denmark)

    Hecker, J.; Diethers, A.; Schulz, D.;

    2012-01-01

    formats including IgE. Using variants of Bet v 1, the epitope of the antibody was mapped and extrapolated to other PR10 proteins. RESULTS: The obtained antibodies exhibited pronounced reactivity with Bet v 1, but were not reactive with the homologous PR10 protein Mal d 1. The epitope as defined by the Ig......-reactivities predicted by primary structure analyses of different isoforms and PR10 proteins were verified by allergen chip-based analyses. CONCLUSIONS: The obtained results demonstrate that hybrid IgE repertoires represent a source for human antibodies with genuine paratopes. The IgE-derived information about the Ig......E epitope nature of Bet v 1 and homologues allows for detailed insights into molecular aspects of allergenicity and cross-reactivity within the PR10 protein family....

  10. Gaussian Affine Feature Detector

    CERN Document Server

    Xu, Xiaopeng

    2011-01-01

    A new method is proposed to get image features' geometric information. Using Gaussian as an input signal, a theoretical optimal solution to calculate feature's affine shape is proposed. Based on analytic result of a feature model, the method is different from conventional iterative approaches. From the model, feature's parameters such as position, orientation, background luminance, contrast, area and aspect ratio can be extracted. Tested with synthesized and benchmark data, the method achieves or outperforms existing approaches in term of accuracy, speed and stability. The method can detect small, long or thin objects precisely, and works well under general conditions, such as for low contrast, blurred or noisy images.

  11. Full domain closure of the ligand-binding core of the ionotropic glutamate receptor iGluR5 induced by the high affinity agonist dysiherbaine and the functional antagonist 8,9-dideoxyneodysiherbaine

    DEFF Research Database (Denmark)

    Frydenvang, Karla Andrea; Lash, L Leanne; Naur, Peter;

    2009-01-01

    with the structurally related high-affinity agonist dysiherbaine (DH), and to that of L-glutamate. The pharmacological activity of MSVIII-19 was confirmed in patch-clamp recordings from transfected HEK293 cells, where MSVIII-19 predominantly inhibits iGluR5-2a, with little activation apparent at a high concentration (1...

  12. Serum levels of total IgE and soluble CD23 in bronchial asthma

    Directory of Open Access Journals (Sweden)

    G. Di Lorenzo

    1996-01-01

    Full Text Available The aim of the present study was to compare, during the pollen season, serum levels of total IgE and soluble CD23 (sCD23 from patients with allergic bronchial asthma, with those from healthy subjects. Significantly higher levels of total IgE and sCD23 were found in patients with asthma compared to the control group. Both in normal controls and in asthmatic patients, a significant correlation was shown between the levels of these two molecules. In asthmatic patients, significant correlations were found for both total IgE and sCD23, with lung function measured as bronchial responsiveness to inhaled methacholine. These results suggest that in asthmatic patients, in addition to the study of total serum IgE levels, the assessment of sCD23 serum levels may be helpful in the evaluation of disease activity.

  13. NASA geometry data exchange specification for computational fluid dynamics (NASA IGES)

    Science.gov (United States)

    Blake, Matthew W.; Kerr, Patricia A.; Thorp, Scott A.; Jou, Jin J.

    1994-01-01

    This document specifies a subset of an existing product data exchange specification that is widely used in industry and government. The existing document is called the Initial Graphics Exchange Specification. This document, a subset of IGES, is intended for engineers analyzing product performance using tools such as computational fluid dynamics (CFD) software. This document specifies how to define mathematically and exchange the geometric model of an object. The geometry is represented utilizing nonuniform rational B-splines (NURBS) curves and surfaces. Only surface models are represented; no solid model representation is included. This specification does not include most of the other types of product information available in IGES (e.g., no material properties or surface finish properties) and does not provide all the specific file format details of IGES. The data exchange protocol specified in this document is fully conforming to the American National Standard (ANSI) IGES 5.2.

  14. Local B cells and IgE production in the oesophageal mucosa in eosinophilic oesophagitis

    OpenAIRE

    Vicario, M; Blanchard, C; Stringer, K F; Collins, M.H.; Mingler, M K; Ahrens, A.; Putnam, P E; Abonia, J P; Santos, J.; Rothenberg, M E

    2009-01-01

    Background: Eosinophilic oesophagitis (EO) is an emerging yet increasingly prevalent disorder characterised by a dense and selective eosinophilic infiltration of the oesophageal wall. While EO is considered an atopic disease primarily triggered by food antigens, disparities between standard allergen testing and clinical responses to exclusion diets suggest the participation of distinct antigen-specific immunoglobulin E (IgE) in the pathophysiology of EO. Aim: To find evidence for a local IgE ...

  15. Serum Specific IgE to Thyroid Peroxidase Activates Basophils in Aspirin Intolerant Urticaria

    OpenAIRE

    Shin, Yoo Seob; Suh, Dong-Hyeon; Yang, Eun-Mi; Ye, Young-Min; Park, Hae-Sim

    2015-01-01

    Thyroid antibodies are frequently observed in urticaria patients, but their roles in urticaria are not clearly elucidated. We investigated the role of serum specific IgE to thyroid peroxidase (TPO) in patients with aspirin intolerant acute urticaria (AIAU) and aspirin intolerant chronic urticaria (AICU). We recruited 59 AIAU and 96 AICU patients with 69 normal controls (NC). Serum specific IgE to TPO was measured by manual direct ELISA, and CD203c expressions on basophil with additions of TPO...

  16. Serum IgE reactivity profiling in an asthma affected cohort.

    Directory of Open Access Journals (Sweden)

    Tania Dottorini

    Full Text Available BACKGROUND: Epidemiological evidence indicates that atopic asthma correlates with high serum IgE levels though the contribution of allergen specific IgE to the pathogenesis and the severity of the disease is still unclear. METHODS: We developed a microarray immunoassay containing 103 allergens to study the IgE reactivity profiles of 485 asthmatic and 342 non-asthmatic individuals belonging to families whose members have a documented history of asthma and atopy. We employed k-means clustering, to investigate whether a particular IgE reactivity profile correlated with asthma and other atopic conditions such as rhinitis, conjunctivitis and eczema. RESULTS: Both case-control and parent-to-siblings analyses demonstrated that while the presence of specific IgE against individual allergens correlated poorly with pathological conditions, particular reactivity profiles were significantly associated with asthma (p<10E-09. An artificial neural network (ANN-based algorithm, calibrated with the profile reactivity data, correctly classified as asthmatic or non-asthmatic 78% of the individual examined. Multivariate statistical analysis demonstrated that the familiar relationships of the study population did not affect the observed correlations. CONCLUSIONS: These findings indicate that asthma is a higher-order phenomenon related to patterns of IgE reactivity rather than to single antibody reactions. This notion sheds new light on the pathogenesis of the disease and can be readily employed to distinguish asthmatic and non-asthmatic individuals on the basis of their serum reactivity profile.

  17. Nippostrongylus brasiliensis: radioresistant IgE antibody-forming cells in infected rats

    International Nuclear Information System (INIS)

    In Nippostrongylus brasiliensis-infected rats, anti-N. brasiliensis IgE antibody production was observed at 20 weeks postinfection, long after the worms, as a source of antigen, had been expelled. The persistent IgE production was not abrogated after whole body irradiation (800 R) administered at 12 or 20 weeks, suggesting the participation of radioresistant IgE-forming cells. Help of T cells and recruitment of B memory cells in the irradiated rats seems to be ruled out by the findings that the irradiation completely inhibited the initiation of anti-N. brasiliensis IgE production in rats shortly after the infection with N. brasiliensis or after primary and secondary immunization with N. brasiliensis-antigen. Moreover, clearance of anti-N. brasiliensis IgE antibody from circulation did not seem to be crucially affected by the irradiation. The radioresistant cells forming anti-N. brasiliensis IgE were most productive in mesenteric lymph nodes as compared to other lymph nodes. The recognition of antigens fractionated by chromatography on Sephadex G-200 was the same for IgE-forming cells from rats 12 weeks after infection as for those from 3 weeks after infection. Based on these results, one of the mechanisms of persistent elevation of IgE antibody in the host infected with helminth parasites might be explained by the participation of radioresistant IgE-forming cells

  18. Serological prevalence of anti-latex IgE antibodies in unselected blood donors in Argentina.

    Science.gov (United States)

    Docena, G H; Fossati, C A

    2005-01-01

    The prevalence of specific IgE to natural rubber latex proteins in the general population remains uncertain. The purpose of this study was to determine the prevalence of sera containing specific IgE antibodies to latex proteins using immunoenzymatic methods. A population of 500 unselected adult voluntary blood donors was the source of the sera used in this study. Two different immunoenzymatic methods (EAST and CARLA) were used to analyze the presence of specific IgE antibodies. Confirmation assay was carried out by inhibition ELISA and immunoblotting. Sera from healthy nonatopic individuals were also used as control. Two hundred and twenty five sera showed higher than normal total IgE levels. Of those, three presented latex specific IgE antibodies, which could be inhibited in a dose-response manner with the natural rubber latex and glove extracts. Several latex allergens were recognized by the IgE antibodies from these positive sera. This low seroprevalence (0.66%) indicates that latex hypersensitivity is not an important problem in the general population. We believe that prevention of latex exposure is only necessary in high risk groups of patients. PMID:15864882

  19. Nippostrongylus brasiliensis: radioresistant IgE antibody-forming cells in infected rats

    Energy Technology Data Exchange (ETDEWEB)

    Watanabe, N.; Kobayashi, A.

    1989-02-01

    In Nippostrongylus brasiliensis-infected rats, anti-N. brasiliensis IgE antibody production was observed at 20 weeks postinfection, long after the worms, as a source of antigen, had been expelled. The persistent IgE production was not abrogated after whole body irradiation (800 R) administered at 12 or 20 weeks, suggesting the participation of radioresistant IgE-forming cells. Help of T cells and recruitment of B memory cells in the irradiated rats seems to be ruled out by the findings that the irradiation completely inhibited the initiation of anti-N. brasiliensis IgE production in rats shortly after the infection with N. brasiliensis or after primary and secondary immunization with N. brasiliensis-antigen. Moreover, clearance of anti-N. brasiliensis IgE antibody from circulation did not seem to be crucially affected by the irradiation. The radioresistant cells forming anti-N. brasiliensis IgE were most productive in mesenteric lymph nodes as compared to other lymph nodes. The recognition of antigens fractionated by chromatography on Sephadex G-200 was the same for IgE-forming cells from rats 12 weeks after infection as for those from 3 weeks after infection. Based on these results, one of the mechanisms of persistent elevation of IgE antibody in the host infected with helminth parasites might be explained by the participation of radioresistant IgE-forming cells.

  20. Lipoxin Receptors

    Directory of Open Access Journals (Sweden)

    Mario Romano

    2007-01-01

    Full Text Available Lipoxins (LXs represent a class of arachidonic acid (AA metabolites that carry potent immunoregulatory and anti-inflammatory properties, LXA4 and LXB4 being the main components of this series. LXs are generated by cooperation between 5-lipoxygenase (LO and 12- or 15-LO during cell-cell interactions or by single cell types. LX epimers at carbon 15, the 15-epi-LXs, are formed by aspirin-acetylated cyclooxygenase-2 (COX-2 in cooperation with 5-LO. 15-epi-LXA4 is also termed aspirin-triggered LX (ATL. In vivo studies with stable LX and ATL analogs have established that these eicosanoids possess potent anti-inflammatory activities. A LXA4 receptor has been cloned. It belongs to the family of chemotactic receptors and clusters with formyl peptide receptors on chromosome 19. Therefore, it was initially denominated formyl peptide receptor like 1 (FPRL1. This receptor binds with high affinity and stereoselectivity LXA4 and ATL. It also recognizes a variety of peptides, synthetic, endogenously generated, or disease associated, but with lower affinity compared to LXA4. For this reason, this receptor has been renamed ALX. This review summarizes the current knowledge on ALX expression, signaling, and potential pathophysiological role. The involvement of additional recognition sites in LX bioactions is also discussed.

  1. First-In-Human Study Demonstrating Tumor-Angiogenesis by PET/CT Imaging with 68Ga-NODAGA-THERANOST, a High-Affinity Peptidomimetic for αvβ3 Integrin Receptor Targeting

    OpenAIRE

    Baum, Richard P.; Kulkarni, Harshad R.; Müller, Dirk; Satz, Stanley; Danthi, Narasimhan; Kim, Young-Seung; Brechbiel, Martin W.

    2015-01-01

    68Ga-NODAGA-THERANOST™ is an αvβ3 integrin antagonist and the first radiolabeled peptidomimetic to reach clinical development for targeting integrin receptors. In this first-in-human study, the feasibility of integrin receptor peptidomimetic positron emission tomography/computed tomography (PET/CT) imaging was confirmed in patients with non-small-cell lung cancer and breast cancer.

  2. Affinity Purification of Insulin by Peptide-Ligand Affinity Chromatography

    Institute of Scientific and Technical Information of China (English)

    2007-01-01

    The affinity heptapeptide (HWWWPAS) for insulin, selected from phage display library,was coupled to EAH Sepharose 4B gel and packed to a 1-mL column. The column was used for the affinity purification of insulin from protein mixture and commercial insulin preparation. It was observed that the minor impurity in the commercial insulin was removed by the affinity chromatography. Nearly 40 mg of insulin could be purified with the 1-mL affinity column. The results revealed the high specificity and capacity of the affinity column for insulin purification. Moreover, based on the analysis of the amino acids in the peptide sequence, shorter peptides were designed and synthesized for insulin chromatography. As a result, HWWPS was found to be a good alternative to HWWWPAS, while the other two peptides with three or four amino acids showed weak affinity for insulin. The results indicated that the peptide sequence of HWWWPAS was quite conservative for specific binding of insulin.

  3. An affine framework for analytical mechanics

    OpenAIRE

    Urbanski, Pawel

    2003-01-01

    An affine Cartan calculus is developed. The concepts of special affine bundles and special affine duality are introduced. The canonical isomorphisms, fundamental for Lagrangian and Hamiltonian formulations of the dynamics in the affine setting are proved.

  4. Adjoint affine fusion and tadpoles

    Science.gov (United States)

    Urichuk, Andrew; Walton, Mark A.

    2016-06-01

    We study affine fusion with the adjoint representation. For simple Lie algebras, elementary and universal formulas determine the decomposition of a tensor product of an integrable highest-weight representation with the adjoint representation. Using the (refined) affine depth rule, we prove that equally striking results apply to adjoint affine fusion. For diagonal fusion, a coefficient equals the number of nonzero Dynkin labels of the relevant affine highest weight, minus 1. A nice lattice-polytope interpretation follows and allows the straightforward calculation of the genus-1 1-point adjoint Verlinde dimension, the adjoint affine fusion tadpole. Explicit formulas, (piecewise) polynomial in the level, are written for the adjoint tadpoles of all classical Lie algebras. We show that off-diagonal adjoint affine fusion is obtained from the corresponding tensor product by simply dropping non-dominant representations.

  5. Adjoint affine fusion and tadpoles

    CERN Document Server

    Urichuk, Andrew

    2016-01-01

    We study affine fusion with the adjoint representation. For simple Lie algebras, elementary and universal formulas determine the decomposition of a tensor product of an integrable highest-weight representation with the adjoint representation. Using the (refined) affine depth rule, we prove that equally striking results apply to adjoint affine fusion. For diagonal fusion, a coefficient equals the number of nonzero Dynkin labels of the relevant affine highest weight, minus 1. A nice lattice-polytope interpretation follows, and allows the straightforward calculation of the genus-1 1-point adjoint Verlinde dimension, the adjoint affine fusion tadpole. Explicit formulas, (piecewise) polynomial in the level, are written for the adjoint tadpoles of all classical Lie algebras. We show that off-diagonal adjoint affine fusion is obtained from the corresponding tensor product by simply dropping non-dominant representations.

  6. Expressions of nerve growth factor and its high-affinity receptor, tyrosine kinase A, as well as low-affinity common receptor, p75 neurotrophin receptor, in the lesions of lichen planus and their clinical significance%神经生长因子及其高亲和受体酪氨酸激酶、低亲和公共受体p75NTR在扁平苔藓皮损中的表达及意义

    Institute of Scientific and Technical Information of China (English)

    钱悦; 陈思远; 黄长征; 冯爱平; 褚淑娟

    2014-01-01

    Objective To detect the expressions of nerve growth factor (NGF) and its receptors tyrosine kinase A (TrkA) as well as p75 neurotrophin receptor (p75NTR) in the lesions of lichen planus.Methods Biopsy specimens were collected from the lesions of 32 patients with lichen planus and normal skin of 12 healthy human controls and subjected to paraffin embedding.Immunohistochemical avidin-biotin complex (ABC) method was used to detect the expressions of NGF,TrkA and p75NTR.Results NGF and TrkA,which were located in the cytoplasm of keratinocytes,were strongly or moderately expressed in the lesional skin specimens,but absent or weakly expressed in the normal skin specimens (both P < 0.01).No significant differences were observed in the expression of p75NTR between the lesional and normal skin specimens,or in the expressions of NGF,TrkA or p75NTR among specimens from patients in different age groups,patients of different gender or lesions at different sites (all P > 0.05).There was a positive correlation between the expression of NGF and TrkA in the lesions of lichen planus (R2 =0.535,P < 0.01).Conclusion NGF may play a certain role in the development of lichen planus via its highaffinity receptor TrkA.%目的 检测神经生长因子(NGF)及其受体TrkA、p75NTR在扁平苔藓皮损中的表达.方法 应用免疫组化ABC法检测32例扁平苔藓皮损和12例健康人皮肤石蜡标本NGF及其受体TrkA、p75NTR表达状况.结果 NGF及TrkA在32例扁平苔藓皮损表皮角质形成细胞中均有不同程度的表达(++~+++),表达部位为细胞质,高于健康人皮肤NGF(-~+)及TrkA(-~+)的表达,两组间差异均有统计学意义(P<0.01);而p75NTR的表达两组差异无统计学意义.扁平苔藓皮损中NGF与TrkA表达呈正相关(R2=0.535,P< 0.01).NGF及其受体TrkA、p75NTR在扁平苔藓不同发病年龄、部位以及不同性别患者角质形成细胞中的表达差异均无统计学意义.结论 NGF通过其高亲和受体TrkA在

  7. Measurement of total IgE antibody levels in lacrimal fluid of patients suffering from atopic and non-atopic eye disorders. Evidence for local IgE production in atopic eye disorders?

    OpenAIRE

    Aalders-Deenstra, V; Kok, P T; Bruynzeel, P L

    1985-01-01

    Total IgE levels in lacrimal fluid of patients suffering from different eye disorders were quantitatively measured by a modification of the paper radio immuno sorbent test (PRIST). The geometric mean values for patients with atopic conjunctivitis, patients with keratoconjunctivitis vernalis, and patients with asthma without conjunctivitis differed significantly from those for control persons and those for patients without atopic conjunctivitis. Besides lacrimal fluid IgE levels, serum IgE lev...

  8. The IgE antibody system: Mature, peripheral B lymphocytes exert regulatory influences on the IgE systems of self-reconstituting, sublethally irradiated mice

    International Nuclear Information System (INIS)

    Previous studies have documented clear biological differences, such as sensitivity to moderate doses of irradiation, between B lymphocytes of the IgE type and B lymphocytes of other immunoglobulin isotypes. The present experiments were originally designed to explore such differences further by comparing the abilities of B lymphocytes derived from various IgE responder phenotypes, which differ among various inbred mouse strains, to reconstitute in a positive way the ability of sublethally irradiated recipient mice (of syngeneic or semisyngeneic type) to mount specific immune responses of the IgE antibody class. This was an important question with regard to delineating fully underlying differences in IgE responder phenotypes among different mouse strains, since heretofore most of the emphasis in experimentally defining such differences has focused on differences in T cell function, rather than B cell function. The experimental approach chosen to address this question seemed logical for two reasons: (1) it was our expectation that following exposure to the dose of irradiation employed (700 rads), individual mice would only slowly repopulate peripheral lymphoid tissues with their own stem cell products, and hence the expression of IgE responsiveness observed could be expected to reflect the responsiveness of the donor B cell population transferred into such recipients; and (2) since recipient mice were carrier-primed one week prior to irradiation in order to create a pool of radioresistant carrier-specific helper T cells, one could expect that this amplified pool of helper T cells would hasten the development of antibody production by the transferred donor B cells

  9. Effect of maternal lifestyle on cord blood IgE factor.

    Science.gov (United States)

    Shirakawa, T; Morimoto, K; Sasaki, S; Taniguchi, K; Motonaga, M; Akahori, W; Akahori, S; Akahori, T; Ohmori, H; Kuroda, E; Okabe, K; Yugari, K; Yamana, M

    1997-06-01

    During recent decades much interest has been focused on the possibility of predicting and preventing atopic diseases during pregnancy. The idea of being able to detect a predisposition early and take suitable environmental measures in order to avoid overt allergy is an attractive position. Elevated cord IgE of around 1.0 IU/ml has been proposed as a predictor in western children. However, there remains no information about the effect of maternal lifestyle during pregnancy on these levels. Total IgE levels were therefore determined using Pharmacia CAP system and PRIST, with sensitivities of 0.01 kU/l and 0.25 kU/l, respectively, from serum samples taken from 1138 Japanese pairs of cord blood and pregnant women responding to a questionnaire regarding 17 health practices, intake of 32 food allergens and 5 environmental factors. Of these, 28 (2.5%) pairs of samples were excluded from further analysis because of high contamination of IgA (> 15.4 mg/ml) in cord blood. Median cord blood IgE was 0.286 kU/l and geometric mean IgE was 66.25 kU/l in maternal sera using CAP system; there was no significant correlation between maternal log (IgE) and cord blood IgE. Similar results were obtained from PRIST, whose correlation with CAP system was significant (r = 0.884, p 400 IU/ml), especially in women aged more than 35 years who are pregnant with a male child. However, maintenance of healthy lifestyles, especially taking proper exercise and sleeping, and avoidance of inhalant allergens during late pregnancy may be a more important strategy for the reduction of cord blood IgE levels. PMID:9258545

  10. Seasonal split influenza vaccine induced IgE sensitization against influenza vaccine.

    Science.gov (United States)

    Nakayama, Tetsuo; Kumagai, Takuji; Nishimura, Naoko; Ozaki, Takao; Okafuji, Teruo; Suzuki, Eitaro; Miyata, Akiko; Okada, Kenji; Ihara, Toshiaki

    2015-11-01

    Although anaphylaxis is an extremely rare vaccine-associated adverse event, it occurred in young children following administration of the 2011/12 seasonal split influenza vaccine, which contained 2-phenoxyethanol as the preservative. These children had high levels of IgE antibodies against influenza vaccine components. We herein investigated why these children were sensitized. One hundred and seventeen series of serum samples were obtained immediately before, and one month after the first and second immunizations with the HA split vaccine of 2011/12. Forty-two sequential serum samples were collected in the acute and convalescent phases (2 and 4 weeks) after natural infection with H1N1 Pdm in 2009. IgE antibodies developed following the vaccination of young children with seasonal split vaccines, whereas no significant IgE response was observed following natural infection with H1N1 Pdm 2009. The prevalence of IgE antibodies was not influenced by outbreaks of H1N1 Pdm. Repeated immunization with the HA split vaccine induced IgE sensitization against the influenza vaccine irrespective of the H1N1, H3N2, or B influenza subtypes. The reasons why anaphylaxis only occurred in recipients of the influenza vaccine containing 2-phenoxyethanol are still being investigated, and the size distribution of antigen particles may have shifted to a slightly larger size. Since the fundamental reason was IgE sensitization, current split formulation for the seasonal influenza vaccine needs to be reconsidered to prevent the induction of IgE sensitization. PMID:26188254

  11. Increased serum IgE concentrations during infection and graft versus host disease after bone marrow transplantation.

    OpenAIRE

    Walker, S. A.; Rogers, T. R.; Perry, D; Hobbs, J R; Riches, P G

    1984-01-01

    Serum IgE concentrations estimated in 25 bone marrow transplant recipients during episodes of infection or graft versus host disease, or both, were raised not only in some patients with acute graft versus host disease but also in many patients with infection. Raised values were not seen in chronic graft versus host disease. The routine estimation of serum IgE in bone marrow transplant recipients had minimal value because of the lack of specificity of the IgE response.

  12. Serological analysis of human IgG and IgE anti-insulin antibodies by solid-phase radioimmunoassays

    International Nuclear Information System (INIS)

    A single solid-phase assay system which is useful for quantitative measurement of both IgG and IgE anti-insulin antibodies in human serum has been developed. Insulin-specific immunoglobulins are absorbed from human serum by excess quantities of insulin-agarose. After washes to remove unbound immunoglobulins, radioiodinated Staph A or rabbit anti-human IgE is added to detect bound IgG or IgE anbitodies, respectively

  13. Effect of Leukotriene B4 on Enhancement of Superoxide Production Evoked by Formyl-Methionyl-Leucyl-Phenylalanine in Myeloid Differentiated HL-60 Cells : Possible Involvement of Intracellular Calcium Influx and High Affinity Receptor for Leukotriene B4

    OpenAIRE

    Harada, Yoshiaki

    1990-01-01

    Exposure of a human leukemic cell line HL-60 to 1 % dimethylsulfoxide (DMSO) for 4 days induced myeloid differentiation. DMSO-differentiated HL-60 cells displayed high and low-affinity binding sites for leukotriene B4 (LTB4). The pretreatment of myeloid differentiated HL-60 cells with 1-10 nM LTB4 enhanced superoxide production evoked by 100 nM formyl-methionyl-leucyl-phenylalanine (fMLP) to 127-137% of the controls stimulated by fMLP alone. A concentration eliciting a half maximal increase (...

  14. Exploring the temporal development of childhood IgE profiles to allergen components

    Directory of Open Access Journals (Sweden)

    Önell Annica

    2012-12-01

    Full Text Available Abstract Background Children often develop allergies that may or not persist into adulthood. Although the different allergic symptoms over time have been well documented, the underlying pattern of sensitization to various proteins and subsequent allergy development is unexplored. The aim was to study the sensitization pattern to allergen components over time from infancy to adulthood in a group of infants with heredity for allergic diseases. Methods IgE profiles were monitored in a group of 67 children from 6 months to 18 years using a microarray chip (ImmunoCAP® ISAC containing 103 allergen components derived from 47 allergen sources. The chip IgE profile was compared with clinical history, skin prick test results and diagnoses (atopic dermatitis, asthma and allergic rhinoconjunctivitis at each time point for each child. Results IgE profiles were unique for each child and showed broad agreement with the results of skin prick tests and doctors’ diagnoses. In addition, close examination of the IgE profiles often revealed early indication of subsequent allergies. IgE profiles also facilitated the examination of cross-reactivity contra co-sensitization, thereby greatly enhancing the possibility for managing patients. Conclusion This explorative description indicates that sensitization pattern to allergen components differs over time as well as among allergic individuals when examined with microarray technology.

  15. Differential induction of total IgE by two Salmonella enterica serotypes

    Directory of Open Access Journals (Sweden)

    Zhanna eKtsoyan

    2015-06-01

    Full Text Available The main goal of this study was to establish how the inflammation caused by infection with two different Salmonella enterica serotypes, S. Typhimurium and S. Enteritidis, may lead to the predisposition to allergy as measured by total IgE level in the blood. Infection by S. Typhimurium did not affect the systemic IgE concentration while in S. Enteritidis-infected patients there was a significant 3.5-fold increase. This effect was especially profound in patients >4 years old, with up to the eight-fold increase above the norm. The degree of dysbiosis in these two infections measured with the comparative counts of cultivated bacteria showed an inverse relationship with the IgE concentration. Earlier we reported the elevated level of IL-17 in patients infected by S. Enteritidis. In the current study a significant correlation was found between the concentrations of IL-17 and IgE suggesting a possible role played by this cytokine in triggering the production of IgE in response to S. Enteritidis infection.

  16. Detection of Trichinella-specific IgE in human Trichinellosis: Creating a new test

    Directory of Open Access Journals (Sweden)

    Dević Marija

    2014-01-01

    Full Text Available Trichinellosis is a parasitic disease of humans caused by the nematode from the genus Trichinella, predominantly Trichinella spiralis (T. spiralis. If Trichinella infection is suspected, based on epidemiological link and clinical criteria within defined period of time, then finding of Trichinella-specific antibodies in the examined sera provides a definitive proof of the infection establishment. Detection of Trichinella-specific IgE that could precede, coincide or follow IgG seroconversion not only confirms the infection existence, but could narrow the time frame in which the infection took place to a year or even less. Since there are no commercially available tests for monitoring the Trichinella-specific IgE presence during the course of the disease, our work was aimed to establish this kind of ELISA test. Specificity and sensitivity of so far described Trichinella-specific IgE ELISAs are not satisfying enough; two major problems are poor discrimination between positive and negative results and cross reactivity with sera of patients with different parasitic diseases. In this study, we have developed Trichinella-specific IgE Capture ELISA that overcomes problems with specificity and sensitivity and enables determination of Trichinella-specific IgE. [Projekat Ministarstva nauke Republike Srbije, br. 173047

  17. Use of Specific IgE and Skin Prick Test to Determine Clinical Reaction Severity.

    Science.gov (United States)

    Ta, Von; Weldon, Brittany; Yu, Grace; Humblet, Olivier; Neale-May, Susan; Nadeau, Kari

    2011-01-01

    AIMS: To determine whether specific IgE and skin prick test correlate better in predicting reaction severity during a double-blinded placebo controlled food challenge (DBPCFC) for egg, milk, and multiple tree nut allergens. STUDY DESIGN: Prospective study. PLACE AND DURATION OF STUDY: Department of Pediatrics, Stanford University School of Medicine, August 2009 and ongoing. METHODOLOGY: We examined the reaction severity of twenty-four subjects to nine possible food allergens: milk, egg, almond, cashew, hazelnut, peanut, sesame, pecan and walnut. Specific IgE and SPT were performed before each DBPCFC. DBPCFC results were classified into mild (1), moderate (2), or severe (3) reactions using a modified Bock's criteria. RESULTS: Twenty four subjects underwent a total of 80 DBPCFC. Eighty percent of all DBPCFCs resulted in a positive reaction. A majority, 71%, were classified as mild. No reactions occurred with a SPT of zero mm while three reactions occurred with a negative specific IgE. All reactions were reversible with medication. CONCLUSION: These data suggest that SPT and specific IgE levels are not associated with reaction severity (p 5mm had a sensitivity of 91% and specificity of 50%. Combining SPT > 5mm and IgE > 7 resulted in a reduced sensitivity of 64%. Unexpectedly, a history of anaphylaxis 70% (n=17) was not predictive of anaphylaxis on challenge 4% (n=2). PMID:22993721

  18. Long Term Persistence of IgE Anti-Varicella Zoster Virus in Pediatric and Adult Serum Post Chicken Pox Infection and after Vaccination with Varicella Virus Vaccine

    OpenAIRE

    Smith-Norowitz, Tamar A; Josekutty, Joby; Silverberg, Jonathan I; Lev-Tov, Hadar; Norowitz, Yitzchok M.; Kohlhoff, Stephan; Nowakowski, Maja; Durkin, Helen G.; Bluth, Martin H

    2009-01-01

    The production of IgE specific to different viruses (HIV-1, Parvovirus B19, RSV), and the ability for IgE anti-HIV-1 to suppress HIV-1 production in vitro, strongly suggest an important role for IgE and/or anti viral specific IgE in viral pathogenesis. Previous studies in our laboratory were the first to report the presence of IgE anti-varicella zoster virus (VZV) in an adolescent patient with shingles. However, the presence and long term persistence of IgE anti VZV antibodies has not been st...

  19. IgE epitopes of intact and digested Ara h 1

    DEFF Research Database (Denmark)

    Bøgh, Katrine Lindholm; Nielsen, H.; Madsen, Charlotte Bernhard; Mills, E.N.C.; Rigby, N.; Eiwegger, T.; Szépfalusi, Z.; Roggen, E.L.

    epitopes have been suggested to be of great importance. ObjectiveThe aim of this study was to identify IgE specific epitopes of intact and digested Ara h 1, and to compare epitope patterns between humans and rats. MethodsSera from five peanut allergic patients and five Brown Norway rats were used to......E, which by far accounted for most of the eluted peptide sequences. Epitope patterns were rather similar for both intact and digested Ara h 1 as well as for humans and rats. ConclusionsIndividual patient specific epitope patterns have been identified for the major allergen Ara h 1. IgE binding epitopes......Background Allergen epitope characterization provides valuable information useful for the understanding of proteins as food allergens. It is believed that IgE epitopes in general are conformational, nevertheless, for food allergens known to sensitize through the gastrointestinal tract linear...

  20. Aspergillus specific IgE estimation by radioallergosorbent technique (RAST) in obstructive airways disease at Agra

    International Nuclear Information System (INIS)

    The radioallergosorbent technique (RAST) was used to measure the levels of Aspergillus specific IgE in 25 normal controls, 25 cases of extrinsic bronchial asthma and 25 cases of allergic broncho-pulmonary aspergillosis with a view to study the clinical role and its correlation with sputum culture, skin sensitivity and severity of airways obstruction. The test was performed using Pharmacia diagnostic kits with antigen derived from Aspergillus fumigatus. Abnormal levels of Aspergillus specific IgE were observed in 84 per cent cases of bronchial asthma but none of the controls. 86.7 per cent of all cases with positive skin test had positive radioallergosorbent test and there was no false positive reaction. There was a positive correlation of Aspergillus specific IgE with skin test positivity and with FEV1/FVC per cent. (author)

  1. Helminths and malaria co-infections are associated with elevated serum IgE

    DEFF Research Database (Denmark)

    Mulu, Andargachew; Kassu, Afework; Legesse, Mengistu;

    2014-01-01

    BACKGROUND: Both helminth and malaria infections result in a highly polarized immune response characterized by IgE production. This study aimed to investigate the total serum IgE profile in vivo as a measure of Th2 immune response in malaria patients with and without helminth co-infection. METHODS......: A cross sectional observational study composed of microscopically confirmed malaria positive (N = 197) and malaria negative (N = 216) apparently healthy controls with and without helminth infection was conducted at Wondo Genet Health Center, Southern Ethiopia. A pre-designed structured format was...... utilized to collect socio-demographic and clinical data of the subjects. Detection and quantification of helminths, malaria parasites and determination of serum IgE levels were carried out following standard procedures. RESULTS: Irrespective of helminth infection, individuals infected by malaria showed...

  2. Domestic exposure to fungi and total serum IgE levels in asthmatic children.

    Science.gov (United States)

    Su, Huey-Jen Jenny; Wu, Pei-Chih; Lei, Huan-Yao; Wang, Jiu-Yao

    2005-08-14

    We measured the number of airborne, viable fungi and house dust mite (HDM) allergen levels in the homes of a group of asthmatic children. Blood samples were drawn and the amounts of total and specific serum IgE were determined. The association between the number of fungal colonies, dust mite allergen exposure, and specific and total IgE was evaluated. The number of viable airborne fungi was high (20,543 CFU/m(3)) in those investigated houses. Der p1 concentrations on child's mattress exceeding 2 microg/g were found in 78.6% of the houses. A quantitative dose-response relationship was demonstrated between the exposure to viable, airborne molds and the amount of total IgE (r = 0.4399 and P = .0249) and the level was further increased in children with coexposure to viable fungi and HDM. PMID:16106103

  3. A Study on Specific IgE Against Candida Albicans in Atopic Dermatitis Patients Referred to Boali Hospital, Sari- Iran

    Directory of Open Access Journals (Sweden)

    R.A. Mohammadpour, Ph.D.

    2007-01-01

    Full Text Available AbstractBackground and purpose: Candida albicans (C. albicans as a micro flora of the human could be responsible for a continuous release of allergen and may be responsible for chronic atopic dermatitis (AD in sensitive patients. Thus, in this study, we analyzed AD patients for total IgE and specific IgE, against C. albicans.Materials and Methods: A total of 120 AD patients (male 52 and female 68 were introduced in this study. The age range varied from 4 months to 60 years (mean about 12.9 years. Serum total IgE was assayed by ELISA kit (RADIM. Solid phase was captured by sandwich ELISA assay, using a micro well format for the determination of serum specific IgE to C. Albicans was used according to the manufacturer’s instructions, (ALerCHEK Allergen specific human IgE.Results: Of the 120 AD patients, 37 subjects (30.8% had total IgE higher than 100 IU/mL, 44 subjects (63.7 % 20-100IU/mL and 39 subjects (32.5% less than 20 IU/mL. 9 (7.5% of the patients had specific IgE against C. albicans. Among the patients who were positive for specific IgE to C. albicans, 6 (66.7% were women.Conclusion: The result of our study on serum total IgE in AD patients is concordant with other studies from different countries. In comparison to other studies, our AD patients showed less frequency of specific IgE against Candida albicans. The explanations for the variation in the results obtained in various studies could be due to the age of patients, severity of disease, difference in the antigen preparation, different methods for IgE analysis and total IgE level.

  4. Structural analysis and molecular modeling of two antitrichosanthin IgE clones from phage antibody library

    Institute of Scientific and Technical Information of China (English)

    LIZONGDONG; YURENYUAN; 等

    1997-01-01

    Recently we constructed a murine IgE phage surface display library and screened out two IgE (Fab) clones with specific binding activity to Trichosanthin (TCS).In this work,the Vε and Vκ genes of the two clones were sequenced and their putative germline gene usages were studied.On the basis of the known 3D structure of Trichosanthin and antibody,molecular modeling was carried out to study the antigen-antibody interaction.The possible antigenic determinant sites on the surface of TCS recognized by both the clones were analyzed,and the reaction forces between TCS and two Fab fragments were also analyzed respectively.

  5. Domestic Exposure to Fungi and Total Serum IgE Levels in Asthmatic Children

    OpenAIRE

    Huey-Jen Jenny Su; Pei-Chih Wu; Huan-Yao Lei; Jiu-Yao Wang

    2005-01-01

    We measured the number of airborne, viable fungi and house dust mite (HDM) allergen levels in the homes of a group of asthmatic children. Blood samples were drawn and the amounts of total and specific serum IgE were determined. The association between the number of fungal colonies, dust mite allergen exposure, and specific and total IgE was evaluated. The number of viable airborne fungi was high (20 543 CFU/m3) in those investigated houses. Der p1 concentrations on child's mattress exceeding ...

  6. Epidemiological survey of anti-flea IgE in dogs in Japan by using an antigen-specific IgE quantitative measurement method

    OpenAIRE

    Ichikawa Y.; Beugnet F.

    2012-01-01

    In Japan, an epidemiological survey was performed in dogs from October to December 2008 by using a quantitative measurement method for antigen-specific IgE towards specific Ctenocephalides felis antigens. 214 dogs from 22 veterinary clinics were included. These clinics were located as follows, from North to South: Hokkaido, Aomori, Fukushima, Tochigi, Saitama, Chiba, Tokyo (Tama-City and Ota-ku), Kanagawa, Gifu, Niigata, Kyoto, Nara, Osaka, Hyogo, Kagawa, Ehime, Hiroshima, Yamaguchi, Fukuoka,...

  7. Relationship of Organizational Structure and Leadership Behavior to Staff Satisfaction in IGE Schools. Technical Report No. 412.

    Science.gov (United States)

    Mendenhall, Diana Reed

    The purposes of this study were to determine the relationship of structural aspects of the school organization and leadership behavior in schools to job satisfaction within Individually Guided Education (IGE) schools, and to determine whether organizational structure or leadership behavior had more impact on job satisfaction in IGE schools. Data…

  8. Influence of magnesium supplementation on insulin receptor affinity in erythrocytes of type 2 diabetes rats%镁补充对2型糖尿病大鼠红细胞胰岛素受体亲和力的影响

    Institute of Scientific and Technical Information of China (English)

    钟文雯; 方福生; 成晓玲; 李春霖

    2013-01-01

    Objective To observe the influence of oral magnesium supplementation on insulin receptor affinity in erythrocytes of type 2 diabetes rats. Method Type 2 diabetes rats were induced by high-fat-diet and intraperitoneal injection of streptozocin, and divided into 4 groups. Magnesium was supplemented in high-fat-diet at the dose of 2000, 1000, 200 and 0 mg/kg,respectively. Normal control rats were fed with ordinary diet. Rats were killed after four weeks. Erythrocytes insulin receptor number, combination constants, combined capacity, fasting insulin level and fasting glucose levels were measured, and insulin resistance index and insulin sensitivity index were calculated. Results High affinity insulin receptor combination constant, combined capacity and receptor number in high dosage group were( 1. 24 ± 0. 47 ) ×109L/mol, ( 1. 26 ± 0. 53 ) × 1014/L and 80.23 ±0.47 respectively, significantly higher than diabetes control group. Insulin resistance index decreased and insulin sensitivity index increased in high dosage group than diabetes control. Conclusion Magnesium supplementation could improve erythrocyte insulin receptor affinity and improve insulin resistance in type 2 diabetic rats.%目的 研究镁补充对2型糖尿病大鼠红细胞胰岛素受体亲和力的影响.方法 将高脂饲料喂饲加链脲佐菌素腹腔注射诱发的2型糖尿病大鼠随机分成糖尿病对照及高、中、低剂量组,饲料中分别加入氧化镁(以镁计)0、2000、1000、200mg/kg,正常对照组喂饲普通饲料,共喂养4周.检测红细胞胰岛素受体结合常数、结合容量、受体数目、空腹血糖及空腹胰岛素等指标,计算胰岛素抵抗指数与敏感指数.结果 高剂量组高亲和力胰岛素受体结合常数、结合容量和受体数目分别为(1.24 ±0.47)×109L/mol、(1.26 ±0.53)×1014/L和80.23±0.47,均高于糖尿病对照组;胰岛素抵抗指数较糖尿病对照组降低而胰岛素敏感指数升高.结论

  9. Infinite transitivity on affine varieties

    OpenAIRE

    Arzhantsev, Ivan; Flenner, Hubert; Kaliman, Shulim; Kutzschebauch, Frank; ZAIDENBERG, MIKHAIL

    2012-01-01

    In this note we survey recent results on automorphisms of affine algebraic varieties, infinitely transitive group actions and flexibility. We present related constructions and examples, and discuss geometric applications and open problems.

  10. Representations of affine Hecke algebras

    CERN Document Server

    Xi, Nanhua

    1994-01-01

    Kazhdan and Lusztig classified the simple modules of an affine Hecke algebra Hq (q E C*) provided that q is not a root of 1 (Invent. Math. 1987). Ginzburg had some very interesting work on affine Hecke algebras. Combining these results simple Hq-modules can be classified provided that the order of q is not too small. These Lecture Notes of N. Xi show that the classification of simple Hq-modules is essentially different from general cases when q is a root of 1 of certain orders. In addition the based rings of affine Weyl groups are shown to be of interest in understanding irreducible representations of affine Hecke algebras. Basic knowledge of abstract algebra is enough to read one third of the book. Some knowledge of K-theory, algebraic group, and Kazhdan-Lusztig cell of Cexeter group is useful for the rest

  11. A Panel of IgG1 b12 Variants with Selectively Diminished or Enhanced Affinity for Fcγ Receptors To Define the Role of Effector Functions in Protection against HIV ▿

    OpenAIRE

    Moldt, Brian; Schultz, Niccole; Dunlop, D. Cameron; Alpert, Michael D.; Harvey, Jackson D.; Evans, David T.; Poignard, Pascal; Hessell, Ann J.; Burton, Dennis R.

    2011-01-01

    Passive transfer of neutralizing antibodies is effective in protecting rhesus macaques against simian/human immunodeficiency virus (SHIV) challenge. In addition to neutralization, effector functions of the crystallizable fragment (Fc) of antibodies are involved in antibody-mediated protection against a number of viruses. We recently showed that interaction between the Fc fragment of the broadly neutralizing antibody IgG1 b12 and cellular Fcγ receptors (FcγRs) plays an important role in protec...

  12. Affine toric SL(2)-embeddings

    International Nuclear Information System (INIS)

    In the theory of affine SL(2)-embeddings, which was constructed in 1973 by Popov, a locally transitive action of the group SL(2) on a normal affine three-dimensional variety X is determined by a pair (p/q,r), where 0GV//T-hat. In the substantiation of this result a key role is played by Cox's construction in toric geometry. Bibliography: 12 titles

  13. The influence of Trichosanthin on the induction of IgE responses to ovalbumin under adjuvant—free condition

    Institute of Scientific and Technical Information of China (English)

    JIYONGYONG; CUIHONGYANG; 等

    1995-01-01

    Trichosanthin(TCS) is a potent allergen in mice.It can reproducibly induce specific IgE responses in C57BL/6J mice without the help of adjuvant alum.TCS can bring out the IgE responses to ovalbumin(OVA),while OVA itself could not induce IgE responses to it .However,TCS only works when OVA immunization is given one day after TCS immunization.Either time lag in OVA immunization,or immunization of both antigens at the same time,or OVA immunization given first,all has no effect on the induction of IgE responses to OVA.Through analysis of the antibody specificity of hybridoma clones,it indicated that specific antibodies to TCS or OVA were secreted by independent B cell clones.The IgE antibldies showed no polyreactivity to different antigens.

  14. Potentiation of IgE responses to third-party antigens mediated by Ascaris suum soluble products.

    Science.gov (United States)

    Lee, T D; McGibbon, A

    1993-01-01

    A reductive approach was used to examine the potentiation of IgE responses by nematode infection. Ascaris homogenized extract, Ascaris pseudocoelomic (body) fluid (ABF) and purified Ascaris allergen (ABA) were tested for their ability to act as protein carriers and as mediators of potentiated IgE responses to third-party (ovalbumin; OVA) responses. All three nematode products were excellent protein carriers for the hapten dinitrophenol and showed significantly better activity in this respect than OVA. Neither ABF nor ABA enhanced the level of the IgE response to the third-party antigen but both prolonged the response markedly. ABF, but not ABA, induced high levels of total circulating IgE when given at the same time as OVA with alum. The data suggest that the enhancement and prolongation of IgE responses by nematodes may be two separate but related activities. PMID:8400897

  15. The changes of IL-8, IL-10, IL-12 and IgE in serum of patients with asthma

    International Nuclear Information System (INIS)

    To evaluate the relationship and the clinical significance between the serum IL-8, IL-10, IL-12 and IgE in patients with asthma, the serum IL-8, IL-10 are measured by radioimmunoassay method and the serum IL-12, IgE by ELISA in 55 patients with asthma. The level of serum IL-8, IgE at stage of episode are significantly higher than that at stage of remission (P<0.01); the level of serum IL-10, IL-12 at stage of episode are significantly lower than that at stage of remission (P<0.01). Linear regression shows that the decrease of IL-12 relate to the increase of IgE. The results suggests that the change of the level of serum IL-8, IL-10, IL-12 and IgE could be a maker for the aggravation of asthma

  16. Changes in IgE- and Antigen-dependent histamine-release in peripheral blood of Schistosoma mansoni-infected Ugandan fishermen after treatment with praziquantel

    Directory of Open Access Journals (Sweden)

    Kazibwe Francis

    2004-04-01

    Full Text Available Abstract Background Parasite-specific IgE levels correlate with human resistance to reinfection with Schistosoma spp. after chemotherapy. Although the role of eosinophils in schistosomiasis has been the focus of a great deal of important research, the involvement of other Fcε receptor-bearing cells, such as mast cells and basophils, has not been investigated in relation to human immunity to schistosomes. Chemotherapy with praziquantel (PZQ kills schistosomes living in an in vivo blood environment rich in IgE, eosinophils and basophils. This releases parasite Ags that have the potential to cross-link cell-bound IgE. However, systemic hypersensitivity reactions are not induced by treatment. Here, we describe the effects of schistosomiasis, and its treatment, on human basophil function by following changes in total cellular histamine and in vitro histamine-release induced by schistosome Ags or anti-IgE, in blood samples from infected Ugandan fishermen, who are continuously exposed to S. mansoni infection, before and 1-day and 21-days after PZQ treatment. Results There was a significant increase in the total cellular histamine in blood samples at 1-day post-treatment, followed by a very significant further increase by 21-days post-treatment. In vitro histamine-release induced by S. mansoni egg (SEA or worm (SWA Ags or anti-IgE antibody, was significantly reduced 1-day post-treatment. The degree of this reduction correlated with pre-treatment infection intensity. Twenty-1-days post-treatment, SEA-induced histamine-release was still significantly lower than at pretreatment. Histamine-release was not correlated to plasma concentrations of total or parasite-specific IgE, nor to specific IgG4 plasma concentrations. Conclusion The biology of human blood basophils is modulated by S. mansoni infection and praziquantel treatment. Infection intensity-dependent suppression of basophil histamine-release, histamine-dependent resistance to infection, and

  17. Determination of insulin specific IgE in serum of diabetic patients by solid-phase radioimmunoassay

    International Nuclear Information System (INIS)

    A solid-phase assay system for quantitative measurement of insulin specific IgE has been developed. Insulin specific IgE and IgG are bound to insulin covalently coupled to Sepharose particles. After a washing procedure which removes unbound immunoglobulins, 125I-anti-human IgE-rabbit globulin is added to the Sepharose to determine the amount of bound IgE. The use of standardized 125I-anti-human-IgE-globulin permits quantitation against a calibration curve of IgE and expression as units/ml. No cross-reactivity of IgG was found. Insulin specific IgE was determined in the sera of diabetic patients. Patients treated with procine or mixed species purified insulin (monocomponent MC) did not differ significantly from a non-diabetic control group, whereas serum samples taken from patients treated with crystallized insulin preparations showed a significantly higher level of insulin specific IgE (p < 0.05). Twenty-four patients with generalized insulin allergy and eight patients with immunological insulin resistance also had considerably higher values of IgE antibodies than the control group (p < 0.001 and p < 0.005, respectively). No correlation was found between the concentration of insulin specific IgE and IgG in individual sera and the level of insulin specific IgE was independent of the total IgE. In all cases of allergy elicited by purified insulin (monocomponent MC), it was ascertained that the diabetic patients in question had received less pure insulin during earlier treatment. (orig.)

  18. IgE Myeloma: the First Report of Chinese Case and a Review of the Literature

    Institute of Scientific and Technical Information of China (English)

    HOUJian; JIANGHua; HUHuixian; HEJin

    2004-01-01

    To report a case of IgE myeloma and to compare its clinical features with those reported in the literature. Methods: M-component in serum and urine was determined using celluloseacetate membrane electrophoresis, immunofixation electrophoresis and quantification of immunoglobulins. Immunohistochemistry was performed to detect the expression of IgE and its light chain. Results: A monoclonal peak was detected by cellulose-acetate membrane electrophoresis. The monoclonal band showed on immunofixation electrophoresis the following reactive patterns: positive for A antisera and negative for κ, IgG, IgA, IgM and IgD antisera. The serum immunoglobulin concentrations were: IgG 21.6 g/L, IgA1.2 g/L, IgM 2.64 g/L, κ 7.49 g/L, λ 16.0 g/L, κ/A 0.47. The results of immunohistochemistry showed cytoplasmic expression of IgE immunoglobulin and A light chain in the tumor tissue. Conclusion: This is the first case of IgE multiple myeloma reported in China.

  19. Differential induction of total IgE by two Salmonella enterica serotypes

    DEFF Research Database (Denmark)

    Ktsoyan, Zhanna A.; Mkrtchyan, Mkhitar S.; Zakharyan, Magdalina K.; Mnatsakanyan, Armine A.; Arakelova, Karine A.; Gevorgyan, Zaruhi U.; Ktsoyan, Lusntag A.; Sedrakyan, Anahit; Hovhannisyan, Alvard I; Ghazaryan, Karine A.; Boyajyan, Anna S.; Aminov, Rustam

    2015-01-01

    The main goal of this study was to establish how the inflammation caused by infection with two different Salmonella enterica serotypes, S. Typhimurium and S. Enteritidis, may lead to the predisposition to allergy as measured by total IgE level in the blood. Infection by S. Typhimurium did not aff...

  20. Multiple IgE recognition on the major allergen of the Parietaria pollen Par j 2.

    Science.gov (United States)

    Longo, Valeria; Costa, Maria Assunta; Cibella, Fabio; Cuttitta, Giuseppina; La Grutta, Stefania; Colombo, Paolo

    2015-02-01

    The interaction between IgE antibodies and allergens is a key event in triggering an allergic reaction. The characterization of this region provides information of paramount importance for diagnosis and therapy. Par j 2 Lipid Transfer Protein is one of the most important allergens in southern Europe and a well-established marker of sensitization in Parietaria pollen allergy. The main aim of this study was to map the IgE binding regions of this allergen and to study the pattern of reactivity of individual Parietaria-allergic patients. By means of gene fragmentation, six overlapping peptides were expressed in Escherichia coli, and their IgE binding activity was evaluated by immunoblotting in a cohort of 79 Parietaria-allergic patients. Our results showed that Pj-allergic patients display a heterogeneous pattern of IgE binding to the different recombinant fragments, and that patients reacted simultaneously against several protein domains spread all the over the molecule, even in fragments which do not contain structural features resembling the native allergen. Our results reveal the presence of a large number of linear and conformational epitopes on the Par j 2 sequence, which probably explains the high allergenic activity of this allergen. PMID:25284812

  1. Prevalence of latex-specific IgE antibodies in hospital personnel

    NARCIS (Netherlands)

    Kaczmarek, RG; Silverman, BG; Gross, TP; Hamilton, RG; Kessler, E; ArrowsmithLowe, JT; Moore, RM

    1996-01-01

    Background: Rubber latex hypersensitivity is an important concern for health care workers. Purpose: The Center for Devices and Radiological Health, in collaboration with the Consumer Product Safety Commission, conducted a multicenter study of the prevalence of latex-specific IgE antibodies among Uni

  2. High affinity dopamine D2 receptor radioligands. 3. [123I] and [125I]epidepride: In vivo studies in rhesus monkey brain and comparison with in vitro pharmacokinetics in rat brain

    International Nuclear Information System (INIS)

    Studies of [123I]epidepride uptake in rhesus monkey brain were performed using single photon tomography. Striatal uptake peaked at 0.85% of administered dose/g at 107 min post-injection, then declined slowly to 0.70% of administered dose/g at 6 h. Striatal:posterior brain ratios rose from 2 at 25 min to 6.8 at 105 min, to 15 at 4 h and to 58 at 6.4 h. [123I]Epidepride was displaced by haloperidol (0.1 and 1 mg/kg) with a half-life of washout of 55 min. Little displacement of [123I]epidepride was observed following administration of 1 or 2 mg/kg d-amphetamine, respectively, indicating [123I]epidepride is not easily displaced by endogenous dopamine. In vitro equilibrium binding studies with [125I]epidepride using rat striatum revealed a KD of 46 pM and Bmax of 33 pmol/g tissue at 37 degrees C, while at 25 degrees C the KD was 25 pM and the Bmax 32 pmol/g tissue. In vitro kinetic analysis of association and dissociation curves revealed a half-life for receptor dissociation at 37 degrees C of 15 min and 79--90 min at 25 degrees C. Allowing for the temperature difference, there is good correspondence between in vivo and in vitro dissociation kinetics at 25 degrees C. Increasing in vitro incubation temperature from 25 to 37 degrees C caused a 6-fold increase in the dissociation rate, suggesting that there is a change in binding kinetics at the dopamine D2 receptor at 37 degrees C compared to in vivo binding. The results of this study indicate that [123I]epidepride is an excellent radioligand for SPECT studies of the dopamine D2 receptor in man. 34 refs., 4 figs

  3. Density and Affinity of IL-6 Receptors in Human Leukemic Cells%IL-6受体在人白血病细胞膜上的表达及亲和力

    Institute of Scientific and Technical Information of China (English)

    刘爽; 奚永志; 郭斯启; 刘楠; 屠敏; 金荔; 陈兴国; 孔繁华

    2000-01-01

    目的:研究各种白血病细胞膜IL-6R的表达、密度及亲和力,为深入探讨对IL-6/IL-6R系统介导IL-6-PE40外毒素融合蛋白靶向杀伤白血病细胞的敏感性提供可靠依据。方法:采用放射性配基结合实验(RBA)并结合Satchard作图法,系统分析IL-6R在8种极具代表性的人类白血病细胞系U937,HL-60,KG1,TF1,K562,CEM,HUT28和Raji上的表达密度及亲和力,同时应用流式细胞术(FACS)检测IL-6R的α和β亚单位蛋白在这些白血病细胞上的表达。结果:RBA表明,粒、单、红白血病细胞系HL-60,U937,KG1和TF1表面存在着高亲和力IL-6R,平均IL-6R密度/细胞分别为2 502个,2 874个,2 319个及9 329个,而淋系白血病细胞系CEM,HUT28和Raji以及慢粒K562细胞系则未测到IL-6R。FACS显示,HL-60,KG1,U937和TF1均高表达IL-6R的α亚单位蛋白,而CEM,HUT28及K562则不表达IL-6Rα亚单位蛋白;IL-6Rβ亚单位蛋白在U937,KG1,TF1,HUT28及CEM中呈阳性表达,而在HL-60,Raji和K562细胞中则为阴性。结论:鉴于粒、单、红白血病细胞异常高表达IL-6R,而淋系和慢粒白血病呈阴性表达这一事实,提示急非淋白血病可能更适合用重组IL-6-PE40外毒素融合蛋白进行IL-6/IL-6R介导的靶向治疗。%Objective: To make a study of density and affinity of IL-6R in human leukemic cell lines, and discuss the affection of high affinity IL-6R to the targeted treatment of leukemia with IL-6-PE40 fusion protein. Methods: Radial binding assay with scatchard plot and FACS were used to analysis the density and affinity of IL-6R and protein expression of IL-6Rα and β subunits in totally 8 representative human leukemic cell lines. Results: Myelocytie, monocytic and erythrocytic leukemic cell lines U937, HL-60, KG1 and TF1 express high affinity IL-6R, whose average density per cell is 2 502,2 874, 2 319 and 9 329 respectively, however no 125I-IL-6 binding was

  4. HLA-DRB genotype and specific IgE responses in patients with allergies to penicillins

    Institute of Scientific and Technical Information of China (English)

    2006-01-01

    Background Because of the pivotal role of the human leukocyte antigen (HLA) class II molecules in regulating the immune response and their extensive polymorphism, it is not surprising that particular HLA class II alleles have been implicated in susceptibility to allergic diseases and in restriction of the IgE responses to a variety of allergens. We investigated the relationship between HLA-DRB genotype and allergies to various penicillins and explored HLA-DRB restriction of IgE responses to these derivatives of penicillin.Methods Radioallergosorbent test was used to examine 8 kinds of specific IgE antibodies (4 major and 4 minor antigenic determinants) in the sera of 248 patients with an allergy to penicillins and 101 healthy subjects without any allergic reaction. Some (113 patients and 87 healthy control subjects) were chosen from all subjects to type for HLA-DRB alleles by sequence specific primer-polymerase chain reaction.Results Compared with control subjects, a significantly increased frequency of DR9 was present in 77 patients with allergic reactions, with immediate hypersensitive reaction and with urticaria (P = 0.011; P = 0.019; P = 0.005 respectively). Conversely, a significantly decreased frequency of DR14.1 was found in 80 patients with positive IgE antibodies, with immediate reaction and with urticaria compared with control group (P = 0.024; P = 0.038; P = 0.038). A possible excess of HLA-DR17 was found in subjects who were responsive to benzylpenicilloyl compared with those were not (χ2 = 5.134, P = 0.023), and of HLA-DR4 was found in subjects responsive to phenoxomethylpenicillanyl (PVA, χ2 = 4.057, P = 0.044).Conclusion HLA-DRB gene may be involved in allergy to penicillins through modulating specific serum IgE to penicillins.

  5. Characterization of astrocytic and neuronal benzodiazepine receptors

    Energy Technology Data Exchange (ETDEWEB)

    Bender, A.S.

    1988-01-01

    Primary cultures of astrocytes and neurons express benzodiazepine receptors. Neuronal benzodiazepine receptors were of high-affinity, K{sub D} values were 7.5-43 nM and the densities of receptors (B{sub max}) were 924-4131 fmol/mg protein. Astrocytes posses a high-affinity benzodiazepine receptor, K{sub D} values were 6.6-13 nM. The B{sub max} values were 6,033-12,000 fmol/mg protein. The pharmacological profile of the neuronal benzodiazepine receptor was that of the central-type benzodiazepine receptor, where clonazepam has a high-affinity and Ro 5-4864 (4{prime}-chlorodiazepam) has a low-affinity. Whereas astrocytic benzoidazepine receptor was characteristic of the so called peripheral-type benzodiazepine receptors, which shows a high-affinity towards Ro 5-4863, and a low-affinity towards clonazepam. The astrocytic benzodiazepine receptors was functionally correlated with voltage dependent calcium channels, since dihydropyridines and benzodiazepines interacted with ({sup 3}H) diazepam and ({sup 3}H) nitrendipine receptors with the same rank order of potency, showing a statistically significant correlation. No such correlation was observed in neurons.

  6. High affinity dopamine D2 receptor radioligands. 3. [[sup 123]I] and [[sup 125]I]epidepride: In vivo studies in rhesus monkey brain and comparison with in vitro pharmacokinetics in rat brain

    Energy Technology Data Exchange (ETDEWEB)

    Kessler, R.M.; Votaw, J.R.; Schmidt, D.E.; Ansari, M.S.; Holdeman, K.P.; Paulis, T. de; Clanton, J.A.; Pfeffer, R.; Manning, R.G.; Ebert, M.H. (Vanderbilt Univ., Nashville, TN (United States))

    1993-01-01

    Studies of [[sup 123]I]epidepride uptake in rhesus monkey brain were performed using single photon tomography. Striatal uptake peaked at 0.85% of administered dose/g at 107 min post-injection, then declined slowly to 0.70% of administered dose/g at 6 h. Striatal:posterior brain ratios rose from 2 at 25 min to 6.8 at 105 min, to 15 at 4 h and to 58 at 6.4 h. [[sup 123]I]Epidepride was displaced by haloperidol (0.1 and 1 mg/kg) with a half-life of washout of 55 min. Little displacement of [[sup 123]I]epidepride was observed following administration of 1 or 2 mg/kg d-amphetamine, respectively, indicating [[sup 123]I]epidepride is not easily displaced by endogenous dopamine. In vitro equilibrium binding studies with [[sup 125]I]epidepride using rat striatum revealed a K[sub D] of 46 pM and B[sub max] of 33 pmol/g tissue at 37[degrees]C, while at 25[degrees]C the K[sub D] was 25 pM and the B[sub max] 32 pmol/g tissue. In vitro kinetic analysis of association and dissociation curves revealed a half-life for receptor dissociation at 37[degrees]C of 15 min and 79--90 min at 25[degrees]C. Allowing for the temperature difference, there is good correspondence between in vivo and in vitro dissociation kinetics at 25[degrees]C. Increasing in vitro incubation temperature from 25 to 37[degrees]C caused a 6-fold increase in the dissociation rate, suggesting that there is a change in binding kinetics at the dopamine D2 receptor at 37[degrees]C compared to in vivo binding. The results of this study indicate that [[sup 123]I]epidepride is an excellent radioligand for SPECT studies of the dopamine D2 receptor in man. 34 refs., 4 figs.

  7. First-In-Human Study Demonstrating Tumor-Angiogenesis by PET/CT Imaging with 68Ga-NODAGA-THERANOST, a High-Affinity Peptidomimetic for αvβ3 Integrin Receptor Targeting

    Science.gov (United States)

    Baum, Richard P.; Kulkarni, Harshad R.; Müller, Dirk; Danthi, Narasimhan; Kim, Young-Seung; Brechbiel, Martin W.

    2015-01-01

    Abstract 68Ga-NODAGA-THERANOST™ is an αvβ3 integrin antagonist and the first radiolabeled peptidomimetic to reach clinical development for targeting integrin receptors. In this first-in-human study, the feasibility of integrin receptor peptidomimetic positron emission tomography/computed tomography (PET/CT) imaging was confirmed in patients with non-small-cell lung cancer and breast cancer. Methods: Patients underwent PET/CT imaging with 68Ga NODAGA-THERANOST. PET images were analyzed qualitatively and quantitatively and compared to 2-deoxy-2-(18F) fluoro-d-glucose (18F-FDG) findings. Images were obtained 60 minutes postinjection of 300–500 MBq of 68Ga-NODAGA-THERANOST. Results: 68Ga-NODAGA-THERANOST revealed high tumor-to-background ratios (SUVmax=4.8) and uptake at neoangiogenesis sites. Reconstructed fused images distinguished cancers with high malignancy potential and enabled enhanced bone metastasis detection. 18F-FDG-positive lung and lymph node metastases did not show uptake, indicating the absence of neovascularization. Conclusions: 68Ga-NODAGA-THERANOST was found to be safe and effective, exhibiting in this study rapid blood clearance, stability, rapid renal excretion, favorable biodistribution and PK/PD, low irradiation burden (μSv/MBq/μg), and convenient radiolabeling. This radioligand might enable theranostics, that is, a combination of diagnostics followed by the appropriate therapeutics, namely antiangiogenic therapy, image-guided presurgical assessment, treatment response evaluation, prediction of pathologic response, neoadjuvant-peptidomimetic-radiochemotherapy, and personalized medicine strategies. Further clinical trials evaluating 68Ga-NODAGA-THERANOST are warranted. PMID:25945808

  8. Serum IgE and risk of pancreatic cancer in the Prostate, Lung, Colorectal, and Ovarian Cancer Screening Trial (PLCO)

    Science.gov (United States)

    Olson, Sara H.; Hsu, Meier; Wiemels, Joseph L.; Bracci, Paige M.; Zhou, Mi; Patoka, Joseph; Reisacher, William I.; Wang, Julie; Kurtz, Robert C.; Silverman, Debra T.; Stolzenberg-Solomon, Rachael Z.

    2014-01-01

    Epidemiologic studies have consistently found that self-reported allergies are associated with reduced risk of pancreatic cancer. Our aim was to prospectively assess the relationship between serum IgE, a marker of allergy, and risk. This nested case-control study within the Prostate, Lung, Colorectal, and Ovarian Cancer Screening Trial (PLCO) included subjects enrolled in 1994-2001 and followed through 2010. There were 283 cases of pancreatic cancer and 544 controls matched on age, gender, race, and calendar date of blood draw. Using the ImmunoCAP system, we measured total IgE (normal, borderline, elevated), IgE to respiratory allergens, and IgE to food allergens (negative or positive) in serum collected at baseline. Odds ratios (OR) and 95% confidence intervals (CI) were estimated using conditional logistic regression. We assessed interactions with age, gender, smoking, body mass index, and time between randomization and case diagnosis. Overall, there was no association between the IgE measures and risk. We found a statistically significant interaction by baseline age: in those aged >65, elevated risks were observed for borderline total IgE (OR=1.43; 95% CI, 0.88-2.32) and elevated total IgE (OR=1.98; 95% CI, 1.16-3.37) and positive IgE to food allergens (OR=2.83; 95% CI, 1.29-6.20); among participants <65, ORs were <1. Other interactions were not statistically significant. The reduced risk of pancreatic cancer associated with self-reported allergies is not reflected in serum IgE. PMID:24718282

  9. Radioimmunological methods of IgE immunoglobulin determination by the radioimmunosorption test (RIST) and the radioallergosorption test (RAST) in allergic diseases of the upper airways

    International Nuclear Information System (INIS)

    The level of total IgE was determined by the method of radioimmunosorption (RIST) and the specific IgE antibodies by the method of radioallergosorption (RAST). The determinations were carried out in a group of 30 healthy subjects. The serum levels of IgE in the studied group ranged from 10 to 155 u/ml. In no case presence of specific IgE antibodies was demonstrated. Radioimmunological methods of IgE determination are a valuable test in the diagnosis of allergic diseases of upper airways. (author)

  10. [Fractionation of lymphocytes using affinity chromatography with 9 lectins].

    Science.gov (United States)

    de Dios, I; Manso, M; López-Borrasca, A

    1983-12-01

    Lymphocyte subclasses from normal peripheral blood have been fractionated by affinity chromatography with lectins. Concanavalin A (Con A), Lens culinaris lectin (LC), Pisum sativum lectin (PS), Phaseolus vulgaris lectin (PHA), Dolichos biflours lectin (DB), Glicine max lectin (SBA), Ricinus communis lectin (RCA II), Tetragonolobus purpureus lectin (TP) and Triticum vulgaris lectin (WGA), were coupled to Sepharose 6MB, and lymphocytes labelled with 125I were eluted through the chromatographic columns. The binding of lymphocytes to WGA and SBA lectins was 32% and 13% respectively. The binding to the other lectins tested were found to be between 32% and 13%. When solutions of increasing concentrations of specific sugar were added to the columns a progressive elution of bound lymphocytes was observed. These results indicate the existence of a large range of lymphocyte subclasses, with different binding capacity to lectins, which was a function of the receptor number or/and receptor affinity to each lectin. Furthermore, these two parameters were found to vary in each functional population. Even though all the lymphocytes had lectin receptors, T lymphocytes showed higher affinity for Con A, PHA and TP lectins, while B lymphocytes appeared to be more specific for LC, PS, SBA, DB, RCAII and WGA lectins. PMID:6675094

  11. Increased IgE serum levels are unrelated to allergic and parasitic diseases in patients with juvenile systemic lupus erythematosus

    Directory of Open Access Journals (Sweden)

    Bernadete L. Liphaus

    2012-11-01

    Full Text Available OBJECTIVE: The aim of this study was to assess the IgE serum levels in juvenile systemic lupus erythematosus patients and to evaluate possible associations with clinical and laboratory features, disease activity and tissue damage. METHODS: The IgE serum concentrations in 69 consecutive juvenile systemic lupus erythematosus patients were determined by nephelometry. IgG, IgM and IgA concentrations were measured by immunoturbidimetry. All patients were negative for intestinal parasites. Statistical analysis methods included the Mann-Whitney, chi-square and Fisher's exact tests, as well as the Spearman rank correlation coefficient. RESULTS: Increased IgE concentrations above 100 IU/mL were observed in 31/69 (45% juvenile systemic lupus erythematosus patients. The mean IgE concentration was 442.0 ± 163.4 IU/ml (range 3.5-9936.0 IU/ml. Fifteen of the 69 patients had atopic disease, nine patients had severe sepsis and 56 patients presented with nephritis. The mean IgE level in 54 juvenile systemic lupus erythematosus patients without atopic manifestations was 271.6 ± 699.5 IU/ml, and only nine of the 31 (29% patients with high IgE levels had atopic disease. The IgE levels did not statistically differ with respect to the presence of atopic disease, severe sepsis, nephritis, disease activity, or tissue damage. Interestingly, IgE concentrations were inversely correlated with C4 levels (r = -0.25, p = 0.03 and with the SLICC/ACR-DI score (r = -0.34, p = 0.005. The IgE concentration was also found to be directly correlated with IgA levels (r = 0.52, p = 0.03. CONCLUSIONS: The present study demonstrated for the first time that juvenile systemic lupus erythematosus patients have increased IgE serum levels. This increase in IgE levels was not related to allergic or parasitic diseases. Our results are in line with the hypothesis that high IgE levels can be considered a marker of immune dysregulation.

  12. IgE anti Hepatitis B virus surface antigen antibodies detected in serum from inner city asthmatic and non asthmatic children.

    Science.gov (United States)

    Smith-Norowitz, Tamar A; Tam, Elizabeth; Norowitz, Kevin B; Chotikanatis, Kobkul; Weaver, Diana; Durkin, Helen G; Bluth, Martin H; Kohlhoff, Stephan

    2014-04-01

    Viral Hepatitis type B (HBV) is a public health concern, but has not been linked to asthma. Immunoglobulin (Ig) G is involved in HBV immune responses; less is known about IgE antibodies (Abs) against HBV in asthma. Given the importance of HBV, we sought to determine whether HBV vaccine contributes to asthma in children, by stimulating specific IgE production. Total IgE, IgE- or IgG-anti-HBVs Abs were studied in vaccinated pediatric asthmatics and non asthmatics. We found: (1) total IgE was higher in asthmatics; (2) total IgE did not correlate with IgE anti-HBVs; (3) IgE anti-HBVs did correlate with IgG-anti-HBVs in all subjects; (4)IgE- and IgG-HBVs Abs were similar in both groups; (5) IgE- or IgG anti-HBVs Abs did not correlate with age. Our findings indicate that HBV vaccination induces IgE responses in asthmatics and non asthmatics. PMID:24374043

  13. Prediction of allergy from family history and cord blood IgE levels. A follow-up at the age of 5 years. Cord blood IgE. IV

    DEFF Research Database (Denmark)

    Hansen, L G; Halken, S; Høst, A;

    1993-01-01

    with elevated cord blood IgE levels developed allergic disease before 5 years of age (p < 0.01). A cut-off limit of 0.3 kU/l was superior to the originally suggested limit of 0.5 kU/l. A total IgE level > 63 kU/l (geometric mean + 1 SD) at the age of 5 years can be regarded as being an elevated level...

  14. IgE sensitization to bacterial and fungal biopesticides in a cohort of Danish greenhouse workers:

    DEFF Research Database (Denmark)

    Doekes, G.; Larsen, Preben; Sigsgaard, Torben; Bælum, Jesper

    2004-01-01

    BACKGROUND: The use of biopesticides in agriculture may implicate new risks of work-related allergic reactions. METHODS: Sera were tested from the BIOGART project, a longitudinal respiratory health study among >300 Danish greenhouse workers. IgE was measured by enzyme immunoassay (EIA) with...... BT, or to different Verticillium products were also significantly correlated (both r >0.70), whereas IgE anti-BT and anti-Verticillium showed no correlation at all. CONCLUSIONS: Exposure to these microbial biopesticides may confer a risk of IgE-mediated sensitization. In future research there is a...... need to identify allergenic components in the preparations, perform studies on non-exposed controls and analyze the relation between sensitization and health parameters....

  15. Prevalence of IgE antibodies to grain and grain dust in grain elevator workers

    International Nuclear Information System (INIS)

    IgE-mediated allergic reactions have been postulated to contribute to respiratory reactions seen in workers exposed to grain dusts. In an attempt better to define the prevalence of IgE antibodies in workers exposed to grain dusts, we performed the radioallergosorbent test (RAST) on worker sera using both commercial allergens prepared from grain and worksite allergens prepared from grain dust samples collected at the worksite. We found that the two types of reagents identified different populations with respect to the specificity of IgE antibodies present. The RAST assay performed using worksite allergens correlated well with skin test procedures. These results may allow us to gain better understanding of allergy associated with grain dust exposure, and document the utility of the RAST assay in assessment of occupational allergies

  16. Chitin enhances serum IgE in Aspergillus fumigatus induced allergy in mice

    DEFF Research Database (Denmark)

    Dubey, Lalit Kumar; Moeller, Jesper Bonnet; Schlosser, Anders;

    2015-01-01

    Aspergillus fumigatus (A. fumigatus) is a ubiquitous fungus that activates, suppresses or modulates the immune response by changing its cell wall structure and by secreting proteases. In this study, we show that chitin acts as an adjuvant in a murine model of A. fumigatus protease induced allergy....... The mice were immunised intraperitoneally with A. fumigatus culture filtrate antigen either with or without chitin and were subsequently challenged with the culture filtrate antigen intranasally. Alum was used as an adjuvant control. Compared to alum, chitin induced a weaker inflammatory response in...... the lungs, measured as the total cell efflux in BAL, EPO and chitinase production. However, chitin enhanced the total IgE, specific IgE and specific IgG1 production as efficiently as alum. Pre-treatment with chitin but not with alum depressed the concentration of the Th2 cytokines IL-4 and IL-13 in...

  17. The adjuvant effect of phthalate exposure on IgE sensitisation in early childhood

    DEFF Research Database (Denmark)

    Callesen, M.; Weschler, Charles J.; Jensen, T.;

    2012-01-01

    doctor diagnosed allergic diseases (asthma, allergic rhinoconjunctivitis or atopic dermatitis). The same physician conducted a clinical examination of all the 500 children including a structured interview on allergic heredity, clinical and medical history Specific serum-IgE against inhalant and food...... allergens was determined. Samples of settled dust were collected from the children’s bedroom and daycare center for analyses of five phthalates (DEP, DnBP, DiBP, BBzp & DEHP). Phthalate intakes through three different exposure routes were calculated. The diagnosis of allergic disease was based on...... with asthma, allergic rhinoconjunctivitis or atopic dermatitis (P < 0.05, aOR = 2.59) and DEHP exposure (P < 0.05, aOR =3.45). IgE sensitization in children with asthma was associated with DnBP exposure (P < 0.05). IgE sensitization in the cases were associated (P < 0.05) with DnBP and BBzP exposure in...

  18. Construction and diversity analysis of a murine IgE phage surface display library

    Institute of Scientific and Technical Information of China (English)

    LIZONGDONG; MINGYEH

    1997-01-01

    To make further investigation of the IgE antibody repertoire in Trichosanthin (TCS) allergic responses,a murine IgE phage surface display library was constructed (3.0×105 independent clones).We first constructed the Vε cDNA library (4.6×105 independent clones) and Vκ cDNA library (3.0×105 independent clones).Then,the Vε and Vκgene segments were amplified from both libraries by PCR respectively,and assembled into Fab fragment by SOE PCR.The phage library containing Fabs was thus constructed.The diversity of Vε from this library was analyzed and proved.Fab clones with high specificity to TCS have been screened out.

  19. Effect of total lymphoid irradiation on IgE antibody responses in rheumatoid arthritis and systemic lupus erythematosus

    Energy Technology Data Exchange (ETDEWEB)

    Terr, A.I.; Moss, R.B.; Strober, S.

    1987-12-01

    Thirteen patients with rheumatoid arthritis and four patients with systemic lupus erythematosus and nephritis were treated with total lymphoid irradiation because of severe disease refractory to other forms of treatment. Serum samples before and after irradiation were tested for changes in total serum IgE and for changes in specific IgE antibodies to ryegrass pollen, dust mite, cat dander, and Alternaria. There were no statistically significant changes in total or specific IgE from lymphoid irradiation in these patients. The therapy caused a significant decrease in circulating total lymphocyte and Leu-3 (helper/inducer) T-lymphocyte counts. Therefore, reduction in circulating levels of helper/inducer T cells does not appear to influence preexisting levels of IgE antibodies.

  20. Changes of plasma ET and serum IgE levels in children with asthma before and after allergen immunotherapy (AIT)

    International Nuclear Information System (INIS)

    Objective: To investigate the relationship between the changes of plasma ET-1, serum IgE levels and allergen immunotherapy(AIT) in children with asthma. Methods: Plasma levels of ET-1 and serum levels of IgE were measured with radioimmunoassay in 46 children with asthma before and after allergen immunotherapy. Results: In 35 patients benefited from allergen immunotherapy, the plasma levels of ET-1 (59.1 ± 11.7pg/ml) and serum levels of IgE (11.1 ± 3.2IU/ml) were higher significantly than those in controls (P0.05, respectively). Conclusion: These data suggested that the changes of ET-1 and IgE levels after AIT may play an important role in the effective treatment of asthma with AIT. (authors)

  1. Prevalence of IgE antibodies to grain and grain dust in grain elevator workers.

    OpenAIRE

    Lewis, D M; Romeo, P A; Olenchock, S A

    1986-01-01

    IgE-mediated allergic reactions have been postulated to contribute to respiratory reactions seen in workers exposed to grain dusts. In an attempt better to define the prevalence of IgE antibodies in workers exposed to grain dusts, we performed the radioallergosorbent test (RAST) on worker sera using both commercial allergens prepared from grain and worksite allergens prepared from grain dust samples collected at the worksite. We found that the two types of reagents identified different popula...

  2. Respiratory infections in adults with atopic disease and IgE antibodies to common aeroallergens.

    Directory of Open Access Journals (Sweden)

    Aino Rantala

    Full Text Available BACKGROUND: Atopic diseases, including allergic rhinitis, allergic dermatitis and asthma, are common diseases with a prevalence of 30-40% worldwide and are thus of great global public health importance. Allergic inflammation may influence the immunity against infections, so atopic individuals could be susceptible to respiratory infections. No previous population-based study has addressed the relation between atopy and respiratory infections in adulthood. We assessed the relation between atopic disease, specific IgE antibodies and the occurrence of upper and lower respiratory infections in the past 12 months among working-aged adults. METHODS AND FINDINGS: A population-based cross-sectional study of 1008 atopic and non-atopic adults 21-63 years old was conducted. Information on atopic diseases, allergy tests and respiratory infections was collected by a questionnaire. Specific IgE antibodies to common aeroallergens were measured in serum. Adults with atopic disease had a significantly increased risk of lower respiratory tract infections (LRTI; including acute bronchitis and pneumonia with an adjusted risk ratio (RR 2.24 (95% confidence interval [CI] 1.43, 3.52 and upper respiratory tract infections (URTI; including common cold, sinusitis, tonsillitis, and otitis media with an adjusted RR 1.55 (1.14, 2.10. The risk of LRTIs increased with increasing level of specific IgE (linear trend P = 0.059. CONCLUSIONS: This study provides new evidence that working-aged adults with atopic disease experience significantly more LRTIs and URTIs than non-atopics. The occurrence of respiratory infections increased with increasing levels of specific IgE antibodies to common aeroallergens, showing a dose-response pattern with LRTIs. From the clinical point of view it is important to recognize that those with atopies are a risk group for respiratory infections, including more severe LRTIs.

  3. Helminth infection alters IgE responses to allergens structurally related to parasite proteins

    OpenAIRE

    Santiago, Helton da Costa; Flávia L. Ribeiro-Gomes; Bennuru, Sasisekhar; Nutman, Thomas B.

    2014-01-01

    Immunological cross-reactivity between environmental allergens and helminth proteins has been demonstrated, though the clinically-related implications of this cross-reactivity have not been addressed. To investigate the impact of molecular similarity among allergens and cross-reactive homologous helminth proteins in IgE-based serologic assessment of allergic disorders in helminth-infected population, we performed Immunocap™ tests in filarial-infected and non-infected individuals for IgE measu...

  4. Trial of a centralized IgE allergy service to general practitioners in a rural area

    OpenAIRE

    Parker, J H; Moore, R. M. A.; Wilson, R. S. E.

    1981-01-01

    We describe a pilot study of a community IgE service which serves a large rural area and is centred on the biochemistry laboratory and allergy clinic of a district general hospital. The service has proved useful because in many cases it has made attendance at an outpatient department unnecessary. The results appear to be reliable and have provided the general practitioners with additional knowledge of their patients. Other benefits included the investigation of larger numbers of patients, the...

  5. IgE cross-reactivity between house dust mite allergens and Ascaris lumbricoides antigens

    OpenAIRE

    Valmonte, Gardette R.; Cauyan, Gil A.; Ramos, John Donnie A.

    2012-01-01

    Background Common antigens between intestinal parasites and environmental allergens may play a role in the modulation of allergic immune responses. There is a growing interest in investigating cross-reactivity between common helminths and dust mites affecting humans, particularly in the tropics. Objective This study examined the cross-reactivity between the human roundworm Ascaris lumbricoides (Al) and three house dust mite (HDM) species. Methods Specific serum IgE levels to HDM species Blomi...

  6. Detection of Trichinella-specific IgE in human Trichinellosis: Creating a new test

    OpenAIRE

    Dević Marija; Gruden-Movsesijan Alisa; Sofronić-Milosavljević Ljiljana

    2014-01-01

    Trichinellosis is a parasitic disease of humans caused by the nematode from the genus Trichinella, predominantly Trichinella spiralis (T. spiralis). If Trichinella infection is suspected, based on epidemiological link and clinical criteria within defined period of time, then finding of Trichinella-specific antibodies in the examined sera provides a definitive proof of the infection establishment. Detection of Trichinella-specific IgE that could precede, coi...

  7. Influence of protein expression system on elicitation of IgE antibody responses: experience with lactoferrin.

    Science.gov (United States)

    Almond, Rachael J; Flanagan, Brian F; Kimber, Ian; Dearman, Rebecca J

    2012-11-15

    With increased interest in genetically modified (GM) crop plants there is an important need to understand the properties that contribute to the ability of such novel proteins to provoke immune and/or allergic responses. One characteristic that may be relevant is glycosylation, particularly as novel expression systems (e.g. bacterial to plant) will impact on the protein glycoprofile. The allergenicity (IgE inducing) and immunogenicity (IgG inducing) properties of wild type native human lactoferrin (NLF) from human milk (hm) and neutrophil granules (n) and a recombinant molecule produced in rice (RLF) have been assessed. These forms of lactoferrin have identical amino acid sequences, but different glycosylation patterns: hmNLF and nNLF have complex glycoprofiles including Lewis (Le)(x) structures, with particularly high levels of Le(x) expressed by nNLF, whereas RLF is simpler and rich in mannose residues. Antibody responses induced in BALB/c strain mice by intraperitoneal exposure to the different forms of lactoferrin were characterised. Immunisation with both forms of NLF stimulated substantial IgG and IgE antibody responses. In contrast, the recombinant molecule was considerably less immunogenic and failed to stimulate detectable IgE, irrespective of endotoxin and iron content. The glycans did not contribute to epitope formation, with equivalent IgE and IgG binding recorded for high titre anti-NLF antisera regardless of whether the immunising NLF or the recombinant molecule were used substrates in the analyses. These data demonstrate that differential glycosylation profiles can have a profound impact on protein allergenicity and immunogenicity, with mannose and Le(x) exhibiting opposing effects. These results have clear relevance for characterising the allergenic hazards of novel proteins in GM crops. PMID:22813905

  8. Relationship between total and specific IgE in patients with asthma from Siberia

    Czech Academy of Sciences Publication Activity Database

    Gusareva, Elena; Ogorodova, L.M.; Chernyak, L.M.; Lipoldová, Marie

    0091-6749, č. 121 (2008), s. 781-781. ISSN 0091-6749 R&D Projects: GA ČR GD310/03/H147; GA MŠk(CZ) LC06009 Grant ostatní: EC(XE) 05-1000004-7761 Institutional research plan: CEZ:AV0Z50520514 Keywords : total and specific IgE * asthma * Russian population Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 9.773, year: 2008

  9. [Round Table: Immunological urticaria mediated by IgE].

    Science.gov (United States)

    Eseverri, J L; Cozzo, M; Castillo, M f; Marín, A

    1999-01-01

    Urticaria is characterized by the appearance of hives and pruritus. Those hives are formed by oedema and vasodilatation and they disappear when they are pressed on. The acute presentation is extremely common and affects between 10 and 20% of the population at a determined moment. In its simplest form, urticaria is envisioned to represent the same sort of wheal-and-flare reaction observed when histamine is injected into the skin. It produces erythema because of capillary vasodilatation, oedema because of increased permeability in capillary and pruritus secondary at local specific receptors stimulation. Angioedema is caused by the same pathologic alterations that occur in the deep dermis and subcutaneous tissue. Thus, an area involved with angioedema has swelling as the prominent manifestation and appearance of the skin itself may be normal. Due to reduced nerve supply in dermis, angioedema is associated with oppression and not pruritus. Immunoallergological study of urticaria and/or angioedema was requested in 133 cases from 648 from the first patient's visits to the surgery. It supposes a 20.52%. The family suspicion of etiology was food in 62 cases, chemical products in 39 cases, other factors (physical, stings, balloons and other manufactured products.) in 7 cases and 25 cases without a direct relation. Out of 100 children diagnosed of allergic urticaria-angioedema 67 was by food; the foods implicated in frequency order were: eggs and nuts, fruit, milk, vegetables, fish and shellfish. In second place, chemical products were responsible of urticaria in 12 children; five of them were positive in diagnosed proof (prick, oral challenge) for penicillin and amoxicillin, both from beta-lactamic group; two of them had and adverse reaction to anesthetic agents; other two cases were after administration of vaccination and due to tetanus toxin; and three cases were due to aspirin, confirmed by oral provocation test. In 10 children the etiological agent was latex. Other

  10. Evaluation of total IgE, CRP and blood count parameters in children with asthma and allergic rhinitis

    OpenAIRE

    Tülübaş, Feti; Gürel, Ahmet; Donma, Mustafa Metin; NALBANTOĞLU, Burçin; TOPÇU, Birol; Mut, Zeynep Deniz

    2013-01-01

    Objective: This study aimed to research retrospectively whether asthma and allergic rhinitis are related to total IgE, C-reactive protein (CRP) and complete blood count parameters. Materials and methods: Files of 443 children who applied to pediatric outpatient clinics of our hospital,aged 2-18 were retrospectively investigated. Patients were grouped into three as asthma (n=179), allergic rhinitis (n= 171) and control group (n= 93). Patients\\' ages, genders, total IgE, CRP and hemogram va...

  11. Sensitization rates of causative allergens for dogs with atopic dermatitis: detection of canine allergen-specific IgE

    OpenAIRE

    Kang, Min-Hee; Kim, Ha-Jung; Jang, Hye-Jin; Park, Hee-Myung

    2014-01-01

    Allergen-specific IgE serology tests became commercially available in the 1980s. Since then these tests have been widely used to diagnose and treat allergic skin diseases. However, the relationship between a positive reaction and disease occurrence has been controversial. The purpose of this study was to evaluate allergens using a serologic allergy test in dogs with atopic dermatitis (AD). Dogs clinically diagnosed with AD (n=101) were tested using an allergen-specific IgE immunoassay. Among ...

  12. Presensitization to Ascaris antigens promotes induction of mite-specific IgE upon mite antigen inhalation in mice

    OpenAIRE

    Mayu Suzuki; Mutsuko Hara; Saori Ichikawa; Seiji Kamijo; Takuya Nakazawa; Hideki Hatanaka; Kazuo Akiyama; Hideoki Ogawa; Ko Okumura; Toshiro Takai

    2016-01-01

    Background: Patients with house dust mite (HDM) allergy or Ascariasis produce serum IgE specific to the antigens of HDM or nematode Ascaris, respectively. Although human IgE cross-reactivity has been reported between HDM and Ascaris antigens, it remains unclear whether it contributes to the pathogenesis of allergic diseases. We herein investigated the induction of cross-reactive antibodies and T cells in mice and effects of airway exposure to HDM antigens after preimmunization with Ascaris an...

  13. SUB-LETHAL INJECTION OF COBRA VENOM DECREASES ADENOSINE DEAMINASE, HISTAMINE, AND IgE IN ORGANS OF MICE

    Directory of Open Access Journals (Sweden)

    B. V. LIPPS

    2002-01-01

    Full Text Available The pharmacological substances adenosine deaminase (ADA, histamine, and IgE are endogenously present in animals. They are implicated in allergy and asthma and detectable in blood serum. This research reports the presence of ADA, histamine, and IgE at varying levels in almost all major organs of mice. This research further reports that intramuscular injection of sub-lethal dose of Naja kaouthia venom disrupted homeostasis and lowered the levels of ADA, histamine, and IgE in the organs of mice. Adult Balb/C male mice were injected with a half lethal dose of the venom. The mice were sacrificed at 2, 8, and 24 hours post-injection and different organs were collected. Organs were homogenized, centrifuged, and the supernatants were assayed for ADA, histamine, and IgE using respective antisera by immunological test enzyme-linked immunosorbent assay (ELISA. Organs from mice injected with PBS served as controls. No major decrease in the levels of ADA, histamine, and IgE was observed after 2 hours of venom injection. However, tremendous decreases in the levels of ADA, histamine, and IgE was observed in organs 24 h post-injection. The highest decrease for ADA was observed in the brain, liver, lung, muscle, and testis; for histamine, in the heart, muscle, lung, and testis; and for IgE, in the bone, heart, lung, muscle, and testis. This is a first-hand investigation showing the effect of envenomation on the pharmacokinetics of ADA, histamine, and IgE in organs.

  14. Ligand-gated chloride channels are receptors for biogenic amines in C. elegans

    OpenAIRE

    Ringstad, Niels; Abe, Namiko; Horvitz, H. Robert

    2009-01-01

    Biogenic amines such as serotonin and dopamine are intercellular signaling molecules that function widely as neurotransmitters and neuromodulators. We have identified in the nematode Caenorhabditis elegans three ligand-gated chloride channels that are receptors for biogenic amines: LGC-53 is a high-affinity dopamine receptor, LGC-55 is a high-affinity tyramine receptor, and LGC-40 is a low-affinity serotonin receptor that is also gated by choline and acetylcholine. lgc-55 mutants are defectiv...

  15. Development of the Abbott MATRIX Aero assay for the measurement of specific IgE.

    Science.gov (United States)

    Lindberg, R E; Anawis, M A; Bailey, M; Mangat, D; Frank, P M; Hrusovsky, I G; Hooyman, L; Putterman, C; Defreese, J D

    1991-01-01

    An enzyme immunoassay has been developed for the quantitation of specific immunoglobulin E (IgE) in human serum to a panel of allergens. The assay system, called the Abbott MATRIX Aero, includes an instrument, reagents and test cell disposables. Each test cell contains fourteen airborne allergens individually localized on a nitrocellulose solid phase. Individual calibration curves for each allergen are established by the manufacturer and included in barcode form with each test kit. Stable factory calibration eliminates the need to establish a calibration curve with each assay run. The instrument automatically incubates, washes, and reads the test cell and prints each result, which ensures assay reproducibility and provides ease-of-use. Analysis of test results shows good agreement with another in vitro assay for specific IgE. The Abbott MATRIX Aero is a sensitive, reproducible and easy-to-use system for the measurement of specific IgE to a panel of fourteen allergens simultaneously using a single, small volume of serum. PMID:1806584

  16. A murine model of allergic bronchopulmonary aspergillosis with elevated eosinophils and IgE.

    Science.gov (United States)

    Kurup, V P; Mauze, S; Choi, H; Seymour, B W; Coffman, R L

    1992-06-15

    A model of allergic bronchopulmonary aspergillosis was developed by exposing BALB/c mice to Aspergillus fumigatus (AF) Ag. Animals immunized intranasally (i.n.) with soluble AF Ag produced low levels of serum IgE compared to animals given alum precipitated AF Ag i.p. The latter treatment also produced higher levels of serum IgG1 and AF-specific IgG1 than soluble AF given i.p. or i.n.. Blood and lung eosinophilia was detected in mice repeatedly exposed to AF by i.n. but not in the groups injected i.p. Particulate AF Ag-induced striking blood and lung eosinophilia and elevated levels of serum IgE in mice preexposed to AF Ag. The results indicate that route of inoculation and physical nature of Ag determine the immune response and can be manipulated to obtain enhanced IgE, eosinophils, or both in the animal model. PMID:1602128

  17. Specific IgE antibodies to reactive dye-albumin conjugates

    International Nuclear Information System (INIS)

    Hypersensitivity to reactive dye powders has been recognised for a number of years, although the extent of sensitisation amongst dye house operatives and the immunochemistry of the dye molecules has not been investigated. The authors have developed a radioallergosorbent test (RAST) to detect specific IgE to reactive dye-human serum albumin (HSA) conjugates. From a total of 19 dye-HSA conjugates, positive RASTs were found in six workers with allergic symptoms associated with dye exposure, while six asymptomatic case-matched controls were negative. Sera with raised total IgE (up to 4300 kU/l) from unexposed workers gave negative results except for two conjugates which gave a weak positive at 4300 kU/l and one which gave weak positives at all concentrations tested (750-4300 kU/l). RAST inhibition studies demonstrated that the antibody was specific for the complete dye-HSA conjugate. Substitution of bovine serum albumin (BSA) for HSA in the conjugate markedly reduced immunoreactivity and free hapten gave lower inhibition than the complete conjugate. Comparison of the dye-HSA RAST with a RAST using dyed discs showed that the latter did not correlate well with symptoms and was influenced by the total IgE concentration. (Auth.)

  18. Brain Abscess and Keratoacanthoma Suggestive of Hyper IgE Syndrome

    Directory of Open Access Journals (Sweden)

    Soheyla Alyasin

    2015-01-01

    Full Text Available Hyper immunoglobulin-E (IgE syndrome is an autosomal immune deficiency disease. It is characterized by an increase in IgE and eosinophil count with both T-cell and B-cell malfunction. Here, we report an 8-year-old boy whose disease started with an unusual skin manifestation. When 6 months old he developed generalized red, nontender nodules and pathologic report of the skin lesion was unremarkable (inflammatory. Then he developed a painless, cold abscess. At the age of 4 years, he developed a seronegative polyarticular arthritis. Another skin biopsy was taken which was in favor of Keratoacanthoma. Laboratory workup for immune deficiency showed high eosinophil count and high level of immunoglobulin-E, due to some diagnostic criteria (NIH sores: 41 in 9-year-olds, he was suggestive of hyper IgE syndrome. At the age of 8, the patient developed an abscess in the left inguinal region. While in hospital, the patient developed generalized tonic colonic convulsion and fever. Brain computed tomography scan revealed an abscess in the right frontal lobe. Subsequently magnetic resonance imaging (MRI of the brain indicated expansion of the existing abscess to contralateral frontal lobe (left side. After evacuating the abscesses and administrating intravenous antibiotic, the patient’s condition improved dramatically and fever stopped.

  19. Serum IgE Antibodies against Hazelnut in Hazelnut Processing Workers

    Directory of Open Access Journals (Sweden)

    Ege Gulec Balbay

    2012-01-01

    Full Text Available Aim. Previous studies have shown a higher sensitization rate to hazelnut in processing workers but no relation was found between the respiratory symptoms in workplace and hazelnut sensitization. Material and Method. To evaluate the association between the hazelnut sensitization and workplace-related respiratory complaints, hazelnut processing workers had undergone a questionnaire included work-related respiratory symptoms, smoking history, pulmonary function testing, and measurement of serum IgE antibodies against hazelnut. Results. This study consisted of 88 hazelnut processing workers (79 females and 9 males, aged 14–59 years (Mean ± SD: years. The mean working duration was months (min: 1–max: 180. Specific IgE against hazelnut allergens was positive in 14 of cases (17.1%. There was no significant difference between the cases with and without specific IgE against hazelnut allergens regarding respiratory symptoms, history of allergy, smoking status and spirometric values. Conclusion. 17.1% of the hazelnut processing workers were seropositive against hazelnut. Being sensitized to hazelnut was not found to be associated with work-related respiratory symptoms in this study. Further studies are needed in hazelnut workers respiratory health to search topics other than asthma.

  20. 棉铃虫中肠氨肽酶 APN4与 Cry1Ac、Cry2Aa 结合能力的比较%Comparison of the binding affinity of the APN4 receptor in Helicoverpa armigera (Hübner) to the Cry1Ac and Cry2Aa insecticidal proteins

    Institute of Scientific and Technical Information of China (English)

    袁向东; 葛朝虹; 肖玉涛; 梁革梅

    2015-01-01

    Objectives] Bt (Bacillus thuringiensis) insecticidal proteins have been widely adopted to control agricultural pests because of their high target specificity. The binding of a Bt insecticidal protein to its specific receptor in the insect midgut plays a key role in the insecticidal action process. Aminopeptidase N (APN) is one of the major Bt protein receptors. To better characterize the molecular mechanism underlying the insecticidal activity of different Bt insecitcidal proteins, and lay the foundation for Bt resistance management and novel Bt insecticidal protein development, we analyzed the binding affinity of aminopeptidase N4 (APN4) to the Cry1Ac and Cry2Aa insecitcidal proteins in Helicoverpa armigera. [Methods] The binding affinity of aminopeptidase N4 (APN4) was assessed using ligand blot analysis and an ELISA binding assay, respectively. [Results] The results show that recombinant APN4 could bind to both Cry1Ac and Cry2A; their respective dissociation constants were 46.7 nmol/L and 26.5 nmol/L. [Conclusion] The results suggest that there was no significant difference in the binding affinity of APN4 to Cry1Ac and Cry2Aa in H. armigera.%【目的】 Bt 杀虫蛋白(Bacillus thuringiensis)具有高度的靶标特异性,已经被广泛用于农业害虫防治。Bt 杀虫蛋白要发挥杀虫活性,必须首先与其受体蛋白结合,氨肽酶 N(Aminopeptidase N)是一类重要的 Bt 受体蛋白。因此,分析该受体与 Bt 杀虫蛋白的结合能力,可为进一步明确不同 Bt 的分子作用机制、Bt 的抗性治理以及新 Bt 的开发应用等提供借鉴。【方法】本文利用 Ligand blot 和 Elisa 方法比较了棉铃虫 Helicoverpa armigera 中肠 APN4(Aminopeptidase N4,APN4)与 Cry1Ac、Cry2Aa 的结合能力。【结果】原核表达的 APN4片段与活化的 Cry1Ac、Cry2Aa 都可以结合,解离常数(Kd)分别是48.59 nmol/L和21.73 nmol/L。【结论】 APN4片段与 Cry1Ac、Cry2Aa 的结合能力

  1. Affine density in wavelet analysis

    CERN Document Server

    Kutyniok, Gitta

    2007-01-01

    In wavelet analysis, irregular wavelet frames have recently come to the forefront of current research due to questions concerning the robustness and stability of wavelet algorithms. A major difficulty in the study of these systems is the highly sensitive interplay between geometric properties of a sequence of time-scale indices and frame properties of the associated wavelet systems. This volume provides the first thorough and comprehensive treatment of irregular wavelet frames by introducing and employing a new notion of affine density as a highly effective tool for examining the geometry of sequences of time-scale indices. Many of the results are new and published for the first time. Topics include: qualitative and quantitative density conditions for existence of irregular wavelet frames, non-existence of irregular co-affine frames, the Nyquist phenomenon for wavelet systems, and approximation properties of irregular wavelet frames.

  2. Blind prediction of host-guest binding affinities: A new SAMPL3 challenge

    OpenAIRE

    Muddana, Hari S.; Varnado, C. Daniel; Bielawski, Christopher W.; Urbach, Adam R.; Isaacs, Lyle; Geballe, Matthew T; Gilson, Michael K.

    2012-01-01

    The computational prediction of protein-ligand binding affinities is of central interest in early-stage drug-discovery, and there is a widely recognized need for improved methods. Low molecular weight receptors and their ligands—i.e. host-guest systems – represent valuable test-beds for such affinity prediction methods, because their small size makes for fast calculations and relatively facile numerical convergence. The SAMPL3 community exercise included the first ever blind prediction challe...

  3. Protein isolation using affinity chromatography

    OpenAIRE

    Besselink, T.

    2012-01-01

    Many product or even waste streams in the food industry contain components that may have potential for e.g. functional foods. These streams are typically large in volume and the components of interest are only present at low concentrations. A robust and highly selective separation process should be developed for efficient isolation of the components. Affinity chromatography is such a selective method. Ligands immobilized to a stationary phase (e.g., a resin or membrane) are used to bind the c...

  4. Inhomogeneous self-affine carpets

    OpenAIRE

    Fraser, Jonathan M.

    2013-01-01

    We investigate the dimension theory of inhomogeneous self-affine carpets. Through the work of Olsen, Snigireva and Fraser, the dimension theory of inhomogeneous self-similar sets is now relatively well-understood, however, almost no progress has been made concerning more general non-conformal inhomogeneous attractors. If a dimension is countably stable, then the results are immediate and so we focus on the upper and lower box dimensions and compute these explicitly for large classes of inhomo...

  5. Steroid hormone receptors in human salivary gland tumours.

    OpenAIRE

    Lamey, P J; Leake, R. E.; Cowan, S K; Soutar, D S; McGregor, I. A.; McGregor, F M

    1987-01-01

    Major salivary gland tumours were studied for the presence of hormone receptors for oestrogen and progesterone. Of the eight salivary gland tumours exhibiting varied histology, none showed high affinity receptors for oestrogen or progesterone. Salivary tissue from four patients with non-neoplastic salivary gland disease was also studied and found not to contain high affinity receptor sites. The absence of hormone receptors in these glands suggests that such tumours are not dependent on endocr...

  6. Alternative affinity tools: more attractive than antibodies?

    NARCIS (Netherlands)

    Ruigrok, V.J.B.; Levisson, M.; Eppink, M.H.M.; Smidt, H.; Oost, van der J.

    2011-01-01

    Antibodies are the most successful affinity tools used today, in both fundamental and applied research (diagnostics, purification and therapeutics). Nonetheless, antibodies do have their limitations, including high production costs and low stability. Alternative affinity tools based on nucleic acids

  7. Prolactin-binding components in rabbit mammary gland: characterization by partial purification and affinity labeling

    International Nuclear Information System (INIS)

    The molecular characteristics of the PRL receptor isolated from rabbit mammary gland microsomes were investigated. Two approaches were employed: 1) affinity purification of PRL receptors and direct electrophoretic analysis, and 2) affinity cross-linking of microsomal receptors with [125I]ovine PRL [(125I]oPRL). PRL receptors were solubilized from mammary microsomes with 3-[(3-cholamidopropyl)dimethylammonio]1-propane sulfonate and purified using an oPRL agarose affinity column. Sodium dodecylsulfate-polyacrylamide gel electrophoresis and silver staining of the gel revealed at least nine bands, including a 32,000 mol wt band which was most intensively labeled with 125I using the chloramine-T method. Covalent labeling of PRL receptors with [125I]oPRL was performed using N-hydroxysuccinimidyl-4-azido benzoate, disuccinimidyl suberate, or ethylene glycol bis (succinimidyl succinate). A single band of 59,000 mol wt was produced by all three cross-linkers when sodium dodecylsulfate-polyacrylamide gel electrophoresis was performed under reducing conditions. Assuming 1:1 binding of hormone and binding subunit and by subtracting the mol wt of [125I]oPRL, which was estimated from the migration distance on the gel, the mol wt of the binding subunit was calculated as 32,000. In the absence of dithiothreitol during electrophoresis, only one major hormone-receptor complex band was observed. The same mol wt binding components were also detected in microsomal fractions of rabbit kidney, ovary, and adrenal. A slightly higher mol wt binding subunit was observed in rat liver microsomes. Rabbit liver microsomes revealed five [125I]oPRL-binding components, three of which were considered to be those of a GH receptor. Moreover, affinity labeling of detergent-solubilized and affinity purified mammary PRL receptors showed a similar major binding subunit

  8. Spectral affinity in protein networks

    Directory of Open Access Journals (Sweden)

    Teng Shang-Hua

    2009-11-01

    Full Text Available Abstract Background Protein-protein interaction (PPI networks enable us to better understand the functional organization of the proteome. We can learn a lot about a particular protein by querying its neighborhood in a PPI network to find proteins with similar function. A spectral approach that considers random walks between nodes of interest is particularly useful in evaluating closeness in PPI networks. Spectral measures of closeness are more robust to noise in the data and are more precise than simpler methods based on edge density and shortest path length. Results We develop a novel affinity measure for pairs of proteins in PPI networks, which uses personalized PageRank, a random walk based method used in context-sensitive search on the Web. Our measure of closeness, which we call PageRank Affinity, is proportional to the number of times the smaller-degree protein is visited in a random walk that restarts at the larger-degree protein. PageRank considers paths of all lengths in a network, therefore PageRank Affinity is a precise measure that is robust to noise in the data. PageRank Affinity is also provably related to cluster co-membership, making it a meaningful measure. In our experiments on protein networks we find that our measure is better at predicting co-complex membership and finding functionally related proteins than other commonly used measures of closeness. Moreover, our experiments indicate that PageRank Affinity is very resilient to noise in the network. In addition, based on our method we build a tool that quickly finds nodes closest to a queried protein in any protein network, and easily scales to much larger biological networks. Conclusion We define a meaningful way to assess the closeness of two proteins in a PPI network, and show that our closeness measure is more biologically significant than other commonly used methods. We also develop a tool, accessible at http://xialab.bu.edu/resources/pnns, that allows the user to

  9. A molecular determinant of phosphoinositide affinity in mammalian TRPV channels.

    Science.gov (United States)

    Velisetty, Phanindra; Borbiro, Istvan; Kasimova, Marina A; Liu, Luyu; Badheka, Doreen; Carnevale, Vincenzo; Rohacs, Tibor

    2016-01-01

    Phosphatidylinositol 4,5-bisphosphate [PI(4,5)P2] is an important cofactor for ion channels. Affinity for this lipid is a major determinant of channel inhibition by depletion of PI(4,5)P2 upon phospholipase C (PLC) activation. Little is known about what determines PI(4,5)P2 affinity in mammalian ion channels. Here we report that two members of the Transient Receptor Potential Vanilloid (TRPV) ion channel family, TRPV5 and TRPV6 lack a positively charged residue in the TM4-TM5 loop that was shown to interact with PI(4,5)P2 in TRPV1, which shows high affinity for this lipid. When this positively charged residue was introduced to either TRPV6 or TRPV5, they displayed markedly higher affinities for PI(4,5)P2, and were largely resistant to inhibition by PI(4,5)P2 depletion. Furthermore, Ca(2+)-induced inactivation of TRPV6 was essentially eliminated in the G488R mutant, showing the importance of PLC-mediated PI(4,5)P2 depletion in this process. Computational modeling shows that the introduced positive charge interacts with PI(4,5)P2 in TRPV6. PMID:27291418

  10. Electrochemical affinity biosensors for detection of mycotoxins: A review.

    Science.gov (United States)

    Vidal, Juan C; Bonel, Laura; Ezquerra, Alba; Hernández, Susana; Bertolín, Juan R; Cubel, Carlota; Castillo, Juan R

    2013-11-15

    This review discusses the current state of electrochemical biosensors in the determination of mycotoxins in foods. Mycotoxins are highly toxic secondary metabolites produced by molds. The acute toxicity of these results in serious human and animal health problems, although it has been only since early 1960s when the first studied aflatoxins were found to be carcinogenic. Mycotoxins affect a broad range of agricultural products, most important cereals and cereal-based foods. A majority of countries, mentioning especially the European Union, have established preventive programs to control contamination and strict laws of the permitted levels in foods. Official methods of analysis of mycotoxins normally requires sophisticated instrumentation, e.g. liquid chromatography with fluorescence or mass detectors, combined with extraction procedures for sample preparation. For about sixteen years, the use of simpler and faster analytical procedures based on affinity biosensors has emerged in scientific literature as a very promising alternative, particularly electrochemical (i.e., amperometric, impedance, potentiometric or conductimetric) affinity biosensors due to their simplicity and sensitivity. Typically, electrochemical biosensors for mycotoxins use specific antibodies or aptamers as affinity ligands, although recombinant antibodies, artificial receptors and molecular imprinted polymers show potential utility. This article deals with recent advances in electrochemical affinity biosensors for mycotoxins and covers complete literature from the first reports about sixteen years ago. PMID:23743326

  11. Regulation of IgE antibody production by serum molecules. I. Serum from complete Freund's adjuvant-immune donors suppresses irradiation-enhanced IgE production in low responder mouse strains

    International Nuclear Information System (INIS)

    Exposure of mice to low doses of x irradiation at or near the time of primary immunization with 2,4-dinitrophenyl (DNP)-Ascaris suum extract (ASC) results in substantial enhancement of IgE anti-DNP antibody responses; the IgG antibody responses of such mice do not increase after such manipulations. This selective enhancement of IgE antibody production occurs in mice of both high and low IgE responder phenotype, although the extent of enhancement compared to unmanipulated control animals is more striking in low IgE responder mice. The studies presented here demonstrate that the irradiation-enhanced IgE antibody responses of low responder SJL and C57BL/6 mice as well as of intermediate responder AKR mice can be effectively suppressed by passive transfer of CFA-immune serum obtained from isologous donor mice. Moreover, adoptive secondary IgE antibody responses in SJL recipients of primed syngeneic spleen cells can be totally abolished by passive transfer of CFA-immune serum or ascitic fluid from CFA-immune mice. The suppressive activity of CFA-immune serum can be diminished or eliminated by exposure of CFA-primed donor mice to low dose x irradiation at an appropriate point during the priming regimen, after a single inoculation of CFA, and before collection of serum. Low dose x irradiation was not effective in eliminating suppressive activity of CFA-induced ascites fluid obtained from donor mice inoculated repeatedly with CFA. In contrast to the capacity of CFA-immune serum from isologous donors to suppress irradiation-enhanced IgE responses of low responder mice, similar sera or ascites fluids were ineffective in suppressing irradiation-enhanced responses of high responder BALB/c or (SJL x BALB/c)F1 hybrid mice

  12. Manifolds with integrable affine shape operator

    Directory of Open Access Journals (Sweden)

    Daniel A. Joaquín

    2005-05-01

    Full Text Available This work establishes the conditions for the existence of vector fields with the property that theirs covariant derivative, with respect to the affine normal connection, be the affine shape operatorS in hypersurfaces. Some results are obtained from this property and, in particular, for some kind of affine decomposable hypersurfaces we explicitely get the actual vector fields.

  13. Effects of nasal corticosteroids on boosts of systemic allergen-specific IgE production induced by nasal allergen exposure.

    Directory of Open Access Journals (Sweden)

    Cornelia Egger

    Full Text Available Allergen exposure via the respiratory tract and in particular via the nasal mucosa boosts systemic allergen-specific IgE production. Intranasal corticosteroids (INCS represent a first line treatment of allergic rhinitis but their effects on this boost of allergen-specific IgE production are unclear.Here we aimed to determine in a double-blind, placebo-controlled study whether therapeutic doses of an INCS preparation, i.e., nasal fluticasone propionate, have effects on boosts of allergen-specific IgE following nasal allergen exposure.Subjects (n = 48 suffering from grass and birch pollen allergy were treated with daily fluticasone propionate or placebo nasal spray for four weeks. After two weeks of treatment, subjects underwent nasal provocation with either birch pollen allergen Bet v 1 or grass pollen allergen Phl p 5. Bet v 1 and Phl p 5-specific IgE, IgG1-4, IgM and IgA levels were measured in serum samples obtained at the time of provocation and one, two, four, six and eight weeks thereafter.Nasal allergen provocation induced a median increase to 141.1% of serum IgE levels to allergens used for provocation but not to control allergens 4 weeks after provocation. There were no significant differences regarding the boosts of allergen-specific IgE between INCS- and placebo-treated subjects.In conclusion, the application of fluticasone propionate had no significant effects on the boosts of systemic allergen-specific IgE production following nasal allergen exposure.http://clinicaltrials.gov/NCT00755066.

  14. Production and immunological analysis of IgE reactive recombinant egg white allergens expressed in Escherichia coli.

    Science.gov (United States)

    Dhanapala, Pathum; Doran, Tim; Tang, Mimi L K; Suphioglu, Cenk

    2015-05-01

    IgE-mediated allergy to chicken egg affects a large number of children and adults worldwide. The current management strategy for egg allergy is strict avoidance, however this is impractical due to the presence of eggs in a range of foods and pharmaceutical products including vaccines. Strict avoidance also poses nutritional disadvantages due to high nutritional value of eggs. Allergen specific immunotherapy is being pursued as a curative treatment, in which an allergic individual is gradually exposed to the allergen to induce tolerance. Use of recombinant proteins for immunotherapy has been beneficial due to the purity of the recombinant proteins compared to natural proteins. In this study, we produced IgE reactive recombinant egg white proteins that can be used for future immunotherapy. Using E. coli as an expression system, we successfully produced recombinant versions of Gal d 1, 2 and 3, that were IgE reactive when tested against a pool of egg allergic patients' sera. The IgE reactivity indicates that these recombinant proteins are capable of eliciting an immune response, thus being potential candidates for immunotherapy. We have, for the first time, attempted to produce recombinant versions of all 4 major egg white allergens in E. coli, and successfully produced 3, with only Gal d 4 showing loss of IgE reactivity in the recombinant version. The results suggest that egg allergy in Australian populations may mainly be due to IgE reactivity to Gal d 3 and 4, while Gal d 1 shows higher IgE reactivity. This is the first report of a collective and comparative immunological analysis of all 4 egg white allergens. The significance of this study is the potential use of the IgE reactive recombinant egg white proteins in immunotherapy to treat egg allergic patients. PMID:25656803

  15. Affinity of four polar neurotransmitters for lipid bilayer membranes

    DEFF Research Database (Denmark)

    Wang, Chunhua; Ye, Fengbin; Valardez, Gustavo F.;

    2011-01-01

    Weak interactions of neurotransmitters and the lipid matrix in the synaptic membrane have been hypothesized to play a role in synaptic transmission of nerve signals, particularly with respect to receptor desensitization (Cantor, R. S. Biochemistry 2003, 42, 11891). The strength of such interactions......, however, was not measured, and this is an obvious impediment for further evaluation and understanding of a possible role for desensitization. We have used dialysis equilibrium to directly measure the net affinity of selected neurotransmitters for lipid membranes and analyzed this affinity data with...... respect to calorimetric measurements and molecular dynamics simulations. We studied an anionic (glutamate), a cationic (acetylcholine), and two zwitterionic (-aminobutyric acid and glycine) neurotransmitters, and membranes of pure dimyristoyl phosphatidylcholine (DMPC), DMPC doped with 10% anionic lipid...

  16. Allergen-induced bronchial hyperreactivity and eosinophilic inflammation occur in the absence of IgE in a mouse model of asthma

    OpenAIRE

    Mehlhop, Paul D.; van de Rijn, Matthijs; Goldberg, Alison B.; Brewer, Joanne P.; Kurup, Viswanath P.; Martin, Thomas R.; Oettgen, Hans C.

    1997-01-01

    In patients with asthma, elevations of IgE correlate both with allergic inflammation of the airways and with bronchial hyperreactivity (BHR). Several investigations, using mouse models of this disease, have indicated a central role for IgE in the pathogenesis of the eosinophilic inflammation as well as in the obstructive airway physiology of BHR. Some diagnostic studies and therapeutic strategies for asthma are based on the putative role of IgE in asthma pathogenesis. Here, we use mice with a...

  17. Long Term Persistence of IgE Anti-Varicella Zoster Virus in Pediatric and Adult Serum Post Chicken Pox Infection and after Vaccination with Varicella Virus Vaccine.

    Science.gov (United States)

    Smith-Norowitz, Tamar A; Josekutty, Joby; Silverberg, Jonathan I; Lev-Tov, Hadar; Norowitz, Yitzchok M; Kohlhoff, Stephan; Nowakowski, Maja; Durkin, Helen G; Bluth, Martin H

    2009-12-01

    The production of IgE specific to different viruses (HIV-1, Parvovirus B19, RSV), and the ability for IgE anti-HIV-1 to suppress HIV-1 production in vitro, strongly suggest an important role for IgE and/or anti viral specific IgE in viral pathogenesis. Previous studies in our laboratory were the first to report the presence of IgE anti-varicella zoster virus (VZV) in an adolescent patient with shingles. However, the presence and long term persistence of IgE anti VZV antibodies has not been studied in adults. The presence of serum IgE in addition to IgE and IgG anti-VZV antibody in sera were studied in children (N=12) (0-16 y/o) and adults (N=9) (32-76 y/o) with either a past history of (wild type) chicken pox (N=7 children, 9 adults) or 5 years after vaccination with varicella zoster (N=2 children) (Varicella virus vaccine live, Oka/Merck), as well as in non-infected subjects (N=3 children). Of the patients who had a positive history of chicken pox 13 of 16 (81%) contained IgE anti-VZV antibodies; they were both serum IgEHi (>100 IU/ml) and IgELo (chicken pox or vaccination did not make either IgE or IgG anti-VZV antibodies. This is the first demonstration of the existence of IgE anti-VZV antibodies, and its long-term persistence in serum of previously infected subjects. Future studies regarding the functional role of anti-viral IgE and its relationship to VZV are warranted. PMID:23675158

  18. IgE in the diagnosis and treatment of allergic disease.

    Science.gov (United States)

    Platts-Mills, Thomas A E; Schuyler, Alexander J; Erwin, Elizabeth A; Commins, Scott P; Woodfolk, Judith A

    2016-06-01

    Traditionally, the concept of allergy implied an abnormal response to an otherwise benign agent (eg, pollen or food), with an easily identifiable relationship between exposure and disease. However, there are syndromes in which the relationship between exposure to the relevant allergen and the "allergic" disease is not clear. In these cases the presence of specific IgE antibodies can play an important role in identifying the relevant allergen and provide a guide to therapy. Good examples include chronic asthma and exposure to perennial indoor allergens and asthma related to fungal infection. Finally, we are increasingly aware of forms of food allergy in which the relationship between exposure and the disease is delayed by 3 to 6 hours or longer. Three forms of food allergy with distinct clinical features are now well recognized. These are (1) anaphylactic sensitivity to peanut, (2) eosinophilic esophagitis related to cow's milk, and (3) delayed anaphylaxis to red meat. In these syndromes the immunology of the response is dramatically different. Peanut and galactose α-1,3-galactose (alpha-gal) are characterized by high- or very high-titer IgE antibodies for Ara h 2 and alpha-gal, respectively. By contrast, eosinophilic esophagitis is characterized by low levels of IgE specific for milk proteins with high- or very high-titer IgG4 to the same proteins. The recent finding is that patients with alpha-gal syndrome do not have detectable IgG4 to the oligosaccharide. Thus the serum results not only identify relevant antigens but also provide a guide to the nature of the immune response. PMID:27264001

  19. [Detection of IgE specific for egg yolk by enzyme immunoassay. Description of a case].

    Science.gov (United States)

    Carrillo Díaz, T; Cuevas Agustín, M; Moneo Goiri, I; Ibáñez Sandín, M D; Ureña Vilardell, V

    1986-01-01

    Food allergy is a common disease in our country, especially affecting atopic children. Egg-white hypersensitivity is frequently found in these patients. However, egg-yolk hypersensitivity is not usually reported in patients with egg allergy. This article describes a young patient with egg-yolk hypersensitivity, a 12 year old female patient with a medical history of contact urticaria, angioedema and severe acute bronchospasm shortly after the intake of small amounts of egg-yolk. All these episodes required treatment in emergency care units because of the severity of the symptoms. The patient did not describe any other food hypersensitivity and remained symptom-free after the intake of boiled or fried egg-white. She had clinical symptoms of grass pollen hypersensitivity and was therefore on specific immunotherapy at the time of the study. The skin prick-tests were positive to grass pollen and egg-yolk and were negative to mites, moulds, animal dander and to the common food tested (milk, fish, peanut, almond and hazel-nut). Total serum IgE was 1.160 UL/ml. The patient had a positive RAST to egg-white (0.0 PRU/ml) as well as to egg-yolk (8.6 PRU/ml). Furthermore, an indirect enzyme immunoassay as well as a reverse enzyme immunoassay also revealed the presence of specific IgE antibodies. The reverse enzyme immunoassay uses microtiter plates as a solid surface. These plates are coated with a monospecific antihuman IgE antibody. Thereafter, the serum samples are incubated overnight in the wells. After several washings, the presence of specific antibodies is revealed by means of a peroxidase conjugated allergen.(ABSTRACT TRUNCATED AT 250 WORDS) PMID:3515886

  20. Rational self-affine tiles

    CERN Document Server

    Steiner, Wolfgang

    2012-01-01

    An integral self-affine tile is the solution of a set equation $\\mathbf{A} \\mathcal{T} = \\bigcup_{d \\in \\mathcal{D}} (\\mathcal{T} + d)$, where $\\mathbf{A}$ is an $n \\times n$ integer matrix and $\\mathcal{D}$ is a finite subset of $\\mathbb{Z}^n$. In the recent decades, these objects and the induced tilings have been studied systematically. We extend this theory to matrices $\\mathbf{A} \\in \\mathbb{Q}^{n \\times n}$. We define rational self-affine tiles as compact subsets of the open subring $\\mathbb{R}^n\\times \\prod_\\mathfrak{p} K_\\mathfrak{p}$ of the ad\\'ele ring $\\mathbb{A}_K$, where the factors of the (finite) product are certain $\\mathfrak{p}$-adic completions of a number field $K$ that is defined in terms of the characteristic polynomial of $\\mathbf{A}$. Employing methods from classical algebraic number theory, Fourier analysis in number fields, and results on zero sets of transfer operators, we establish a general tiling theorem for these tiles. We also associate a second kind of tiles with a rational matr...

  1. The affine quantum gravity programme

    International Nuclear Information System (INIS)

    The central principle of affine quantum gravity is securing and maintaining the strict positivity of the matrix { g-hat ab(x)} composed of the spatial components of the local metric operator. On spectral grounds, canonical commutation relations are incompatible with this principle, and they must be replaced by noncanonical, affine commutation relations. Due to the partial second-class nature of the quantum gravitational constraints, it is advantageous to use the recently developed projection operator method, which treats all quantum constraints on an equal footing. Using this method, enforcement of regularized versions of the gravitational operator constraints is formulated quite naturally by means of a novel and relatively well-defined functional integral involving only the same set of variables that appears in the usual classical formulation. It is anticipated that skills and insight to study this formulation can be developed by studying special, reduced-variable models that still retain some basic characteristics of gravity, specifically a partial second-class constraint operator structure. Although perturbatively nonrenormalizable, gravity may possibly be understood nonperturbatively from a hard-core perspective that has proved valuable for specialized models. Finally, developing a procedure to pass to the genuine physical Hilbert space involves several interconnected steps that require careful coordination

  2. Prevalence of Atopy, Eosinophilia, and IgE Elevation in IgG4-Related Disease

    OpenAIRE

    Torre, Emanuel Della; Mattoo, Hamid; Mahajan, Vinay S; Carruthers, Mollie; Pillai, Shiv; John H. Stone

    2013-01-01

    IgG4-related disease (IgG4-RD) is a fibroinflammatory disorder that can affect virtually every organ system. T helper type 2 responses have been presumed to be pathogenic in this disease and a high proportion of IgG4-RD patients are reported to have longstanding allergies, peripheral blood eosinophilia, and serum IgE elevation. It has therefore been proposed that allergic mechanisms drive IgG4-RD. However, no epidemiological assessment of atopy, peripheral blood eosinophilia, serumIgEconcentr...

  3. The prevalence of allergic disease and IgE antibodies to house dust mite in schoolchildren in Taiwan.

    Science.gov (United States)

    Tang, R B; Tsai, L C; Hwang, H M; Hwang, B; Wu, K G; Hung, M W

    1990-01-01

    The prevalence of positive specific IgE antibodies to house dust mites (Dermatophagoides pteronyssinus; D. farinae) was determined by enzyme-linked immunosorbent assay (ELISA) in 5097 (61%) volunteers of 8345 schoolchildren aged between 7 and 14 yr from two government schools. All of them filled out a questionnaire concerning allergic symptoms. Among them, 412 (8.1%) children showed a positive reaction to at least one of the two mite allergens, the range varying between 5.6 and 11.2% according to the child's age. Boys had higher prevalence of positive mite specific IgE than girls (9.8% vs. 6.4%, P less than 0.01), with the overall male to female ratio 1.5:1. The prevalence of bronchial asthma in boys and girls was 5.3% and 3.3% respectively. The positive mite specific IgE antibody in children with asthma and allergic rhinitis was 52% (103 of 198) and 28.7% (193 of 673) respectively. The mean levels of mite specific IgE were not significantly related to the age of onset and severity of asthmatic symptoms (P greater than 0.1), but were significantly different among subjects with current and past asthma (P less than 0.001). It is suggested that the mite-specific IgE may play a role in the pathogenesis of bronchial asthma in children. PMID:2310980

  4. IgE antibodies to cow allergens and respiratory health in dairy farmers in Denmark and The Netherlands

    DEFF Research Database (Denmark)

    Doekes, Gert; Wouters, I.; de Vries, J.;

    2000-01-01

    study no relation could be found: IgE anti-cow dander proteins were found in 22.5% of the cases and in 26.8% of the controls. The previously reported strong association between anti-cow IgE sensitization and work-related respiratory disease in Finland could not be confirmed in Dutch and Danish dairy...... the prevalence of IgE anti-cow allergens in Dutch and Danish dairy farmers, and the association with common and work-related respiratory health symptoms. In a pilot study, sera from 37 Dutch dairy farmers were tested in an enzyme immunoassay (EIA) for specific IgE against the major cow allergen Bos d2......, and results were compared with their response to questions in a standardized questionnaire on common and work-related health problems. In addition, IgE anti-cow sensitization was assessed in 71 cases and 71 controls from the follow-up phase of the SUS project (a cohort study on respiratory disease in...

  5. Fucoidan prevents Cε germline transcription and NFκB p52 translocation for IgE production in B cells

    International Nuclear Information System (INIS)

    Fucoidan, a dietary fiber contained in seaweed, reduces the increase of antigen-specific IgE in mice exposed to ovalbumin. In this study, we investigated the effect of fucoidan on IgE production and intracellular events in B cells in vitro. Fucoidan inhibited the production of IgE and Cε germline transcription in murine B cells induced by IL-4 (100 ng/ml) and anti-CD40 antibodies (10 μg/ml), whereas it stimulated cell proliferation. A significant effect of fucoidan on IgE production was observed when B cells were stimulated with a higher dose (5 μg/ml) of anti-CD40 antibodies, but not when stimulated with lower doses (1.25, 2.5 μg/ml), regardless of the IL-4 concentrations. Moreover, nuclear translocation of NFκB p52, but neither that of NFκB p65, nor the phosphorylation of JAK1 and STAT6 was reduced by fucoidan. These results suggest that fucoidan inhibited IgE production by preventing the NFκB p52-mediated pathways activated by CD40

  6. AFFINITY OF THE ALLIGATOR ESTROGEN RECEPTOR FOR SERUM PESTICIDE CONTAMINANTS

    Science.gov (United States)

    Top predators, like the American alligator (Alligator mississippiensis) bioaccumulate and biomagnify persistent pollutants, such as organochlorine pesticides. In a recently published study, several pesticides and pesticide metabolites not previously reported in alligator eggs wer...

  7. Fyn kinase controls Fc{epsilon}RI receptor-operated calcium entry necessary for full degranulation in mast cells

    Energy Technology Data Exchange (ETDEWEB)

    Sanchez-Miranda, Elizabeth; Ibarra-Sanchez, Alfredo [Departamento de Farmacobiologia, Centro de Investigacion y de Estudios Avanzados (Cinvestav), Sede Sur, Calzada de los Tenorios 235, Col. Granjas Coapa, CP 14330 Mexico City (Mexico); Gonzalez-Espinosa, Claudia, E-mail: cgonzal@cinvestav.mx [Departamento de Farmacobiologia, Centro de Investigacion y de Estudios Avanzados (Cinvestav), Sede Sur, Calzada de los Tenorios 235, Col. Granjas Coapa, CP 14330 Mexico City (Mexico)

    2010-01-22

    IgE-antigen-dependent crosslinking of the high affinity IgE receptor (Fc{epsilon}RI) on mast cells leads to degranulation, leukotriene synthesis and cytokine production. Calcium (Ca{sup 2+}) mobilization is a sine qua non requisite for degranulation, allowing the rapid secretion of stored pro-inflammatory mediators responsible for allergy symptoms. Fyn is a Src-family kinase that positively controls Fc{epsilon}RI-induced mast cell degranulation. However, our understanding of the mechanism connecting Fyn activation to secretion of pre-synthesized mediators is very limited. We analyzed Fc{epsilon}RI-dependent Ca{sup 2+} mobilization in bone marrow-derived mast cells (BMMCs) differentiated from WT and Fyn -/- knock out mice. Fyn -/- BMMCs showed a marked defect in extracellular Ca{sup 2+} influx after Fc{epsilon}RI crosslinking but not after thapsigargin addition. High concentrations of Gadolinium (Gd{sup 3+}) partially blocked Fc{epsilon}RI-induced Ca{sup 2+} influx in WT cells but, in contrast, completely inhibited Ca{sup 2+} mobilization in Fyn -/- cells. Low concentrations of an inhibitor of the canonical transient receptor potential (TRPC) Ca{sup 2+} channels (2-aminoethoxyphenyl-borane, 2-APB) blocked Fc{epsilon}RI-induced maximal Ca{sup 2+} rise in WT but not in Fyn -/- cells. Ca{sup 2+} entry through Fyn-controlled, 2-APB sensitive channels was found to be important for full degranulation and IL-2 mRNA accumulation in WT cells. Immunoprecipitation assays showed that Fyn kinase interacts with TRPC 3/6/7 channels after IgE-antigen stimulation, but its association is not related to protein tyrosine phosphorylation. Results indicate Fyn kinase mediates the receptor-dependent activation of TRPC channels that contribute to degranulation in Fc{epsilon}RI-stimulated mast cells.

  8. Fyn kinase controls FcεRI receptor-operated calcium entry necessary for full degranulation in mast cells

    International Nuclear Information System (INIS)

    IgE-antigen-dependent crosslinking of the high affinity IgE receptor (FcεRI) on mast cells leads to degranulation, leukotriene synthesis and cytokine production. Calcium (Ca2+) mobilization is a sine qua non requisite for degranulation, allowing the rapid secretion of stored pro-inflammatory mediators responsible for allergy symptoms. Fyn is a Src-family kinase that positively controls FcεRI-induced mast cell degranulation. However, our understanding of the mechanism connecting Fyn activation to secretion of pre-synthesized mediators is very limited. We analyzed FcεRI-dependent Ca2+ mobilization in bone marrow-derived mast cells (BMMCs) differentiated from WT and Fyn -/- knock out mice. Fyn -/- BMMCs showed a marked defect in extracellular Ca2+ influx after FcεRI crosslinking but not after thapsigargin addition. High concentrations of Gadolinium (Gd3+) partially blocked FcεRI-induced Ca2+ influx in WT cells but, in contrast, completely inhibited Ca2+ mobilization in Fyn -/- cells. Low concentrations of an inhibitor of the canonical transient receptor potential (TRPC) Ca2+ channels (2-aminoethoxyphenyl-borane, 2-APB) blocked FcεRI-induced maximal Ca2+ rise in WT but not in Fyn -/- cells. Ca2+ entry through Fyn-controlled, 2-APB sensitive channels was found to be important for full degranulation and IL-2 mRNA accumulation in WT cells. Immunoprecipitation assays showed that Fyn kinase interacts with TRPC 3/6/7 channels after IgE-antigen stimulation, but its association is not related to protein tyrosine phosphorylation. Results indicate Fyn kinase mediates the receptor-dependent activation of TRPC channels that contribute to degranulation in FcεRI-stimulated mast cells.

  9. Beta adrenergic receptors in pigmented ciliary processes.

    OpenAIRE

    Trope, G. E.; Clark, B.

    1982-01-01

    Beta adrenergic receptors from membrane fragments of pigmented sheep eyes were studied and characterised by ligand binding techniques after the removal of melanin. In a representative experiment the beta max (total number of beta receptors) was 394.9 fmol/mg protein. The receptor affinity (Ka) was 440 pM. The potency series of drugs to displace 125I-HYP from the receptors was timolol = (-) propranolol greater than (+) propranolol greater than salbutamol greater than practolol. beta 1 Recepto...

  10. Selective effects of ligands on vitamin D3 receptor- and retinoid X receptor-mediated gene activation in vivo.

    OpenAIRE

    Lemon, B D; Freedman, L P

    1996-01-01

    Steroid/nuclear hormone receptors are ligand-regulated transcription f factors that play key roles in cell regulation, differentiation, and oncogenesis. Many nuclear receptors, including the human 1,25-dihydroxyvitamin D3 receptor (VDR), bind cooperatively to DNA either as homodimers or as heterodimers with the 9-cis retinoic acid (RA) receptor (retinoid X-receptor [RXR]). We have previously reported that the ligands for VDR and RXR can differentially modulate the affinity of the receptors' i...

  11. The Structure-Function Relationships of Complement Receptor Type 2 (CR2; CD21).

    Science.gov (United States)

    Hannan, Jonathan Paul

    2016-01-01

    Human complement receptor type 2 (CR2; CD21) is a surface-associated glycoprotein which binds to a variety of endogenous ligands, including the complement component C3 fragments iC3b, C3dg and C3d, the low-affinity IgE receptor CD23, and the type I cytokine, interferon-alpha. CR2 links the innate complement-mediated immune response to pathogens and foreign antigens with the adaptive immune response by binding to C3d that is covalently attached to targets, and which results in a cell signalling phenomenon that lowers the threshold for B cell activation. Variations or deletions of the CR2 gene in humans, or the Cr2 gene in mice associate with a variety of autoimmune and inflammatory conditions. A number of infectious agents including Epstein-Barr virus (EBV), Human Immunodeficiency Virus (HIV) and prions also bind to CR2 either directly or indirectly by means of C3d-targeted immune complexes. In this review we discuss the interactions that CR2 undertakes with its best characterized ligands C3d, CD23 and the EBV gp350/220 envelope protein. To date only a single physiologically relevant complex of CR2 with one of its ligands, C3d, has been elucidated. By contrast, the interactions with CD23 and EBV gp350/220, while being important from physiologic and disease-associated standpoints, respectively, are only incompletely understood. A detailed knowledge of the structure-function relationships that CR2 undergoes with its ligands is necessary to understand the implications of using recombinant CR2 in therapeutic or imaging agents, or alternatively targeting CR2 to down-regulate the antibody mediated immune response in cases of autoimmunity. PMID:26916158

  12. IL-1, IL-4 production and IgE levels in acute and chronic fasciolosis before and after triclabendazole treatment.

    Science.gov (United States)

    Allam, A F; Osman, M M; el-Sayed, M H; Demian, S R

    2000-12-01

    IL-1 generation by mononuclear phagocytes, IL-4 production by Th2 lymphocytes and IgE levels in serum were measured in eight patients with acute fasciolosis and seven patients in the chronic stage of the disease before and after triclabendazole treatment. Results were compared with those of a control group of ten individuals. The monocytes and lymphocytes from patients with acute and chronic fasciolosis produced significantly lower levels of IL-1 and IL-4 respectively, particularly in the chronic phase of the disease, as compared to the control. A significant increase in IgE level in both acute and chronic fasciolosis was observed. The level was significantly higher in acute as compared to chronic cases. After treatment with triclabendazole IL-1, IL-4 and IgE levels moved towards the control indicating obvious improvement in the immunological responses of the patients. PMID:11198376

  13. Characterization and crystallization of a recombinant IgE Fab fragment in complex with the bovine β-lactoglobulin allergen

    International Nuclear Information System (INIS)

    The high-resolution mass-spectrometric characterization, crystallization and X-ray diffraction studies of a recombinant IgE Fab fragment in complex with bovine β-lactoglobulin are reported. A D1 Fab fragment containing the allergen-binding variable domains of the IgE antibody was characterized by ESI FT–ICR mass spectrometry and crystallized with bovine β-lactoglobulin (BLG) using the hanging-drop vapour-diffusion method at 293 K. X-ray data suitable for structure determination were collected to 2.8 Å resolution using synchrotron radiation. The crystal belonged to the orthorhombic space group P212121, with unit-cell parameters a = 67.0, b = 100.6, c = 168.1 Å. The three-dimensional structure of the D1 Fab fragment–BLG complex will provide the first insight into IgE antibody–allergen interactions at the molecular level

  14. Quantitation of IgE by means of a modified radial immunodiffusion method in comparison with the radioimmunosorbent test (RIST)

    International Nuclear Information System (INIS)

    Serum IgE were quantitied by means of modified radial immunodiffusion technique (RID). To improve visibility of precipitin bands a staining procedure with DOPA was applied. Pretreatment of sera with dextransulfate proved necessary in order to avoid unspecific ringformation in the agargel. In comparison with the RIST it turned out that sera containing less than 500 I.U. IgE/ml did not produce precipitin bands with this method. Sera containing 500-999 I.U. IgE/ml occasionally exhibited positive results with the RID technique, whereas sera with more than 1,000 I.U./ml were regulary positive. In its present form the RID may be used as screening method for sera with higher IgE levels. Within the above mentioned limits the IgE levels calculated by means of the RID-test roughly corresponded the values determined by the RIST. (orig.)

  15. Effect of in vitro irradiation and cell cycle-inhibitory drugs on the spontaneous human IgE synthesis in vitro

    International Nuclear Information System (INIS)

    The in vitro effects of radiation, diterpine forskolin (FK), and hydrocortisone (HC) on the in vitro spontaneous IgE synthesis by peripheral blood B-lymphocytes from atopic patients were investigated. Without affecting cell viability, in vitro irradiation inhibited in a dose-dependent fashion de novo IgE synthesis in vitro by B cells from all patients examined with a mean 40% reduction of in vitro IgE product after treatment with 100 rads. In contrast, the in vitro IgE production by the U266 myeloma cell line was unaffected, even by irradiation with 1600 rads. The addition to B cell cultures from atopic patients of FK consistently resulted in a dose-dependent inhibition of the spontaneous IgE production in vitro. The addition to cultures of 10(-5) and 10(-6) molar concentrations of HC was also usually inhibitory, whereas lower HC concentrations were uneffective or even enhanced the spontaneous in vitro IgE synthesis. When 10(-6) molar concentrations of both HC and FK were combined in culture, a summation inhibitory effect on the spontaneous IgE synthesis was observed. In contrast, neither FK nor HC had inhibitory effect on the in vitro spontaneous IgE synthesis by the U266 myeloma cell line. The spontaneous in vitro IgE synthesis by B cells from patients with Hodgkin's disease, demonstrating high levels of serum IgE, was strongly reduced or virtually abolished after patients underwent total nodal irradiation to prevent the spread of the disease. In addition, the in vitro spontaneous IgE synthesis by B cells from atopic patients was markedly decreased or abolished by in vivo administration of betamethasone

  16. Effect of in vitro irradiation and cell cycle-inhibitory drugs on the spontaneous human IgE synthesis in vitro

    Energy Technology Data Exchange (ETDEWEB)

    Del Prete, G.F.; Vercelli, D.; Tiri, A.; Maggi, E.; Rossi, O.; Romagnani, S.; Ricci, M.

    1987-01-01

    The in vitro effects of radiation, diterpine forskolin (FK), and hydrocortisone (HC) on the in vitro spontaneous IgE synthesis by peripheral blood B-lymphocytes from atopic patients were investigated. Without affecting cell viability, in vitro irradiation inhibited in a dose-dependent fashion de novo IgE synthesis in vitro by B cells from all patients examined with a mean 40% reduction of in vitro IgE product after treatment with 100 rads. In contrast, the in vitro IgE production by the U266 myeloma cell line was unaffected, even by irradiation with 1600 rads. The addition to B cell cultures from atopic patients of FK consistently resulted in a dose-dependent inhibition of the spontaneous IgE production in vitro. The addition to cultures of 10(-5) and 10(-6) molar concentrations of HC was also usually inhibitory, whereas lower HC concentrations were uneffective or even enhanced the spontaneous in vitro IgE synthesis. When 10(-6) molar concentrations of both HC and FK were combined in culture, a summation inhibitory effect on the spontaneous IgE synthesis was observed. In contrast, neither FK nor HC had inhibitory effect on the in vitro spontaneous IgE synthesis by the U266 myeloma cell line. The spontaneous in vitro IgE synthesis by B cells from patients with Hodgkin's disease, demonstrating high levels of serum IgE, was strongly reduced or virtually abolished after patients underwent total nodal irradiation to prevent the spread of the disease. In addition, the in vitro spontaneous IgE synthesis by B cells from atopic patients was markedly decreased or abolished by in vivo administration of betamethasone.

  17. IgE and the role of the Radio-Allergo-Sorbent Test (RAST)

    International Nuclear Information System (INIS)

    In 1966 a new immunoglobulin was found in persons with allergies and in non-typical myeloma proteins. Normally this immunoglobulin E is present only in nanogramms and rests with predilection on the membrane of mast-cells. There is a reaginic-anaphylactic reaction after re-exposure of antigens to the antigen-antibody reaction followed by denudation of mediators of the anaphylactic reaction. With the Radio-Immuno-Sorbent-Test (RIST) the IgE can be quantitatively determined. Elevated IgE-blood levels are typically found in atopic eczema. With the Radio-Allergo-Sorbent-Test (RAST) the allergen specific IgE can be defined. A conformity with appropriate patchtests can be achieved in 60-80% of the cases. In this review advantoses and problem of RIST- and RAST-diagnoses are described. RAST presents a valuable aid in diagnosis of allergies beeing not burdensome and risky, as it is easy to perform and bears no risk to the patients. At the present time, however, patch tests are necessary in the diagnosis of allergies. (orig.)

  18. Reduction in IgE reactivity of Pacific mackerel parvalbumin by heat treatment.

    Science.gov (United States)

    Kubota, Hiroyuki; Kobayashi, Ayako; Kobayashi, Yukihiro; Shiomi, Kazuo; Hamada-Sato, Naoko

    2016-09-01

    Parvalbumin, a major fish allergen, has been reported to be highly thermostable. However, little is known as to whether parvalbumin is stable at more than 100°C. Thermostability of the Pacific mackerel parvalbumin was examined by subjecting heated (20-140°C) muscle extracts to SDS-PAGE, western blotting and ELISA. As judged by SDS-PAGE and western blotting with the anti-parvalbumin antiserum recognizing the primary structure, the parvalbumin was not degraded even under severe heating conditions. However, western blotting analysis with the monoclonal antibody recognizing the stereoscopic structure revealed that the parvalbumin undergoes conformational changes in a heating load-dependent manner. Importantly, the IgE reactivity of the parvalbumin determined by ELISA using patient sera was also reduced in a heating load-dependent manner; complete loss of IgE reactivity was induced by heating at 140°C. This study showed that the allergenicity of the Pacific mackerel parvalbumin is considerably less thermostable than assumed for other fish parvalbumins. PMID:27041301

  19. Conformal field theory on affine Lie groups

    International Nuclear Information System (INIS)

    Working directly on affine Lie groups, we construct several new formulations of the WZW model, the gauged WZW model, and the generic affine-Virasoro action. In one formulation each of these conformal field theories (CFTs) is expressed as a one-dimensional mechanical system whose variables are coordinates on the affine Lie group. When written in terms of the affine group element, this formulation exhibits a two-dimensional WZW term. In another formulation each CFT is written as a two-dimensional field theory, with a three- dimensional WZW term, whose fields are coordinates on the affine group. On the basis of these equivalent formulations, we develop a translation dictionary in which the new formulations on the affine Lie group are understood as mode formulations of the conventional formulations on the Lie group. Using this dictionary, we also express each CFT as a three-dimensional field theory on the Lie group with a four-dimensional WZW term. 36 refs

  20. Conformal field theory on affine Lie groups

    Energy Technology Data Exchange (ETDEWEB)

    Clubok, K.S.

    1996-04-01

    Working directly on affine Lie groups, we construct several new formulations of the WZW model, the gauged WZW model, and the generic affine-Virasoro action. In one formulation each of these conformal field theories (CFTs) is expressed as a one-dimensional mechanical system whose variables are coordinates on the affine Lie group. When written in terms of the affine group element, this formulation exhibits a two-dimensional WZW term. In another formulation each CFT is written as a two-dimensional field theory, with a three- dimensional WZW term, whose fields are coordinates on the affine group. On the basis of these equivalent formulations, we develop a translation dictionary in which the new formulations on the affine Lie group are understood as mode formulations of the conventional formulations on the Lie group. Using this dictionary, we also express each CFT as a three-dimensional field theory on the Lie group with a four-dimensional WZW term. 36 refs.