The high-affinity receptor for immunoglobulin E. Prospects for the therapy of immediate hypersensitivity reactions.
Explore the source record for details and available documents.
Animals↗
Biomedical subjects
H Metzger
Publications and source records attributed to H Metzger.
Translocation of protein kinase C in rat basophilic leukemic cells induced by phorbol ester or by aggregation of IgE receptors.
Rat basophilic leukemic cells contain protein kinase C (PKC), 96 +/- 1% of which is located in the cytosol in the resting state. Phorbol ester (PMA), synergistically with calcium ionophore (A23187), caused 55% of the total PKC activity to associate rapidly with membranes where it remained for at least 20 min. When IgE-loaded cells were activated by Ag, maximally 30% of the cytosolic activity associated with membranes within 15 to 30 s, but most of this returned to the cytosol by 2 min. The small amount (3%) of PKC activity that remained associated with the membranes did so for at least 20 min but only if aggregation of the receptors was maintained. PKC translocation correlated with aggregation of receptors both at 30 s and at 10 min. However, only the translocation at 10 min and not that at 30 s correlated with receptor-induced exocytosis. In the absence of extracellular calcium (no exocytosis is observed), translocation at 30 s was diminished by 30% and at 10 min was completely absent. Cells depleted of PKC by 18-h treatment with PMA failed to degranulate in response to PMA and A23187 but responded partially (35%) when receptors were aggregated. We conclude that translocation of PKC is an early event that follows aggregation of IgE receptors but may not be essential for mediating the exocytotic mechanism induced by these receptors.
Animals↗
Studies with a monoclonal antibody to the beta subunit of the receptor with high affinity for immunoglobulin E.
The receptor with high affinity for IgE consists of a tetrameric complex of polypeptides, one of which (alpha), contains the binding site for IgE. The function of the other chains--a single beta and two disulfide-linked gamma chains--is unknown. We report the cloning of a murine hybridoma that secretes an IgG1 antibody which specifically reacts with the beta subunit. Studies with this monoclonal antibody show that the subunit stoichiometry of the receptor is unaffected by the presence or absence of bound IgE. We also found that under certain conditions where the alpha beta gamma 2 complex dissociates, beta remains attached to the dimer of gamma chains, indicating that these chains contact each other in the native receptor. In rat basophilic leukemia cells--a neoplastic line of mucosal-type mast cells--all of the beta subunits expressed by the cells appeared to be associated with the high affinity receptor. However, in at least one cell line which has no high affinity receptors--a putative rat lymphoma line--beta or beta-like polypeptides were also expressed.
Animals↗
Molecular aspects of receptors and binding factors for IgE.
Explore the source record for details and available documents.
Antigens, Differentiation, B-Lymphocyte↗
Isolation and characterization of cDNAs coding for the beta subunit of the high-affinity receptor for immunoglobulin E.
Among receptors that bind the Fc region of immunoglobulins ("Fc receptors"), only the one with high affinity for immunoglobulin E (IgE) is known to consist of more than a single polypeptide. In addition to the IgE-binding alpha chain, the receptor contains a single beta chain and two, disulfide-linked, gamma chains. From a cDNA library of a rat mucosal mast cell tumor, from which we recently cloned cDNAs coding for the alpha chain, we have now isolated cDNAs coding for the beta subunit. In vitro transcription-translation of the cDNA directed the synthesis of a polypeptide reactive with two distinctive anti-beta monoclonal antibodies and whose molecular weight was identical to that of authentic beta chains. Polyclonal antibodies to beta peptides expressed in Escherichia coli reacted with intact receptors and isolated beta chains. The gene encodes a protein of 243 residues with no leader sequence. A hydropathicity plot suggests that the polypeptide crosses the plasma membrane four times. The epitope recognized by one of the monoclonal antibodies was localized to the NH2 terminus; that by the other was localized to the COOH terminus. Since those antibodies react with membranes and not with intact cells, we suggest that both ends of the beta subunit are cytoplasmic. RNA transfer blots at high stringency failed to reveal mRNA for beta chains in a variety of cells (in particular, monocytes) that do not contain the high-affinity receptor for IgE.
Amino Acid Sequence↗
How antibodies work: focus on Fc receptors.
It is increasingly appreciated that the part of an antibody not involved in the binding of antigen--the Fc region--plays an important biological role. It activates a variety of receptors not only on so-called effector cells such as macrophages and granulocytes, but also on lymphocytes, and it can thereby modulate the immune response itself. Over the past 2 years much new information has been gained about the structure of such receptors, in large part through molecular genetics. In this review we describe the structure and some aspects of the function of the most complicated of the cellular Fc receptors so far identified: the receptor with high affinity for immunoglobulin E (IgE) on mast cells. The structure of its IgE-binding chain is strikingly similar to the corresponding polypeptide of an immunoglobulin G receptor. Like the latter and like a receptor that binds polymeric immunoglobulin, segments of the protein resemble immunoglobulin sequences. It is surprising that other IgE-binding proteins that putatively serve related functions have completely different structures.
Animals↗
A substituted thieno[3.4-d]imidazole versus substituted benzimidazoles as H+, K+-ATPase inhibitors.
S 3337, 2-(2-ethylaminobenzylsulfinyl)-5,6-dimethoxybenzimidazole, and S 1924, 2-(5-methyl-2-picolylsulfinyl)-1H-thieno[3.4-d]imidazole, are members of new classes of H+, K+-ATPase inhibitors. Their effects on H+, K+-ATPase and 14C-aminopyrine uptake in gastric glands were studied as well as in vivo in pylorus-ligated rats, stomach-lumen-perfused rats and Heidenhain pouch dogs. Their inhibitory effects were compared with the effect of omeprazole. In pylorus-ligated rats the two compounds showed a similar effectiveness as omeprazole. In stomach-lumen-perfused rats and in particular in Heidenhain pouch dogs, S 3337 was clearly less effective than omeprazole, while S 1924 was similarly effective in all in vivo models and in the H+, K+-ATPase assay as omeprazole. The difference in potency between S 1924 and omeprazole on 14C-aminopyrine uptake in gastric glands can be explained by the lower pKa value of S 1924 (3.4) than that of omeprazole (4.0). Additionally, this study shows that there was no correlation between the effects in rats, particularly in pylorus-ligated rats, and in dogs for the H+, K+-ATPase inhibitors tested. It is concluded from this study that substituted thieno[3.4-d]imidazoles represent a new class of potent gastric acid inhibitors.
Adenosine Triphosphatases↗
Calcium-independent phosphoinositide breakdown in rat basophilic leukemia cells. Evidence for an early rise in inositol 1,4,5-trisphosphate which precedes the rise in other inositol phosphates and in cytoplasmic calcium.
Aggregation of the receptor with high affinity for immunoglobulin E (IgE) in rat basophilic leukemia cells leads to a calcium-dependent and a calcium-independent hydrolysis of phosphoinositides. The increase in the levels of inositol phosphates induced in the absence of calcium is only 25% of that observed with 1 mM Ca2+. The inositol phosphates reach a new steady state level 2 min after stimulation in EGTA, whereas with calcium they continue to increase up to 15 min. A similar response is observed when the receptors are aggregated due to the interaction of bound IgE with antigen or with anit-IgE, or by the binding of IgE cross-linked chemically. The antigen-mediated response is inhibited by hapten and disruption of such antigen-antibody aggregates late after stimulation leads to a rapid decline in the levels of the inositol phosphates to basal values. Separation of the inositol phosphates by Dowex columns shows that there is a fast rise in inositol trisphosphate which peaks at 15 s and slowly declines to a lower plateau within 2 min. Analysis by high pressure liquid chromatography reveals a 5-fold increase in the levels of inositol 1,4,5-trisphosphate in less than 10 s after stimulation, which precedes any major change in the other inositol phosphates. Aggregation of the receptor in the absence of external calcium induces a transient increase in cytoplasmic calcium which reaches a maximum of approximately 25 nM over basal levels after activation. The onset of the rise in Ca2+ lags after the initial rise in the inositol 1,4,5-trisphosphate.
Animals↗
A cDNA presumptively coding for the alpha subunit of the receptor with high affinity for immunoglobulin E.
Rat mast cells and a neoplastic analogue such as rat basophilic leukemia (RBL) cells have receptors that have exceptionally high affinity for immunoglobulin E (IgE). When aggregated, these receptors induce cellular degranulation. The alpha chain of the receptor contains the binding site for IgE; the function(s) of the noncovalently associated beta and gamma chains is (are) still undefined. Using a cDNA library constructed from the mRNA of RBL cells, we have isolated a cDNA clone whose sequence predicts a putative 23-residue signal peptide, followed by a sequence that accurately predicts the amino acid composition, the peptide molecular weight, and six peptide sequences (encompassing 59 residues or 26% of the total number) determined for the alpha chain by direct analysis. These findings provide strong evidence that the cDNA codes for the alpha chain, even though expression has not been unambiguously achieved. The sequence suggests that the alpha chain contains a 180-residue extracellular portion with two homologous domains of approximately 35 residues, a 20-residue transmembrane segment containing an aspartic acid, and a 27-residue cytoplasmic portion containing 9 basic amino acids. The sequence shows no homology with the low-affinity receptor for IgE from lymphocytes but over 30% homology with an Fc gamma receptor.
Amino Acid Sequence↗
Further characterization of the subunits of the receptor with high affinity for immunoglobulin E.
The alpha, beta, and gamma 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 alpha 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 alpha and beta. However, these putative homologies were not apparent when tryptic maps of the biosynthetically ([3H]leucine) labeled subunits were analyzed.
Amino Acids↗
Influence of microsurgical reanastomosis of the fallopian tube on luminal pH and PO2 and on the fertilization rate and embryo development in the rabbit oviduct.
The luminal pH and PO2 were measured in rabbit oviducts to evaluate the influence of microsurgery on the environment within the fallopian tube before and after resection of a 1 cm ampullary portion with subsequent tubal reanastomosis. While the oxygen tension showed no difference between the operated oviduct (56 +/- 16 mmHg, n = 8) and the control side (53 +/- 13 mmHg, n = 8), there was a significant pH decrease in the oviducts after microsurgery (7.94 +/- 0.10, n = 8, versus 7.75 +/- 0.16, n = 8, p less than 0.0025). In the second part of the experiment the operated animals were induced to ovulate by hCG injection, and artificial insemination was performed at the same time. The number of fertilized and unfertilized oocytes and the stages of embryonic development (64 h post hCG) were compared between the operated and control oviducts. From the operated oviducts a lower number of ova were recovered and a lower rate of fertilization was observed. The data indicate that microsurgery of the rabbit fallopian tube does not disturb the luminal oxygen tension but may cause a lower pH. Ultimately a reduced rate of fertilization and impaired early development were observed.
Animals↗
[Retroperitoneal intestinal duplication in US and CT. A contribution to its differential diagnosis].
Explore the source record for details and available documents.
Adult↗
[Results of treatment following surgical management of complete acromioclavicular joint dislocation (Tossy III injury). Management using ligament sutures and direct transarticular and indirect extra-articular stabilization].
In the last 5 years 36 patients with complete akromioklavikular separation were treated in 21 cases by ligament suture and Balser-plate, in 15 cases by ligament suture and "Zuggurtung". Good and very good results were found in 85%. Patients in which the protection of the ligament suture was given by application of the Hook-plate had better results than those with "Zuggurtung". The use of the Balser-plate remarks a safe method in protecting the coracoid-clavicular ligament suture. The rate of complications is rare, compared with the patients in the other group. Functional results seem to be better in comparison to other methods of operative treatment.
Acromioclavicular Joint↗
Stimulation of adenylate cyclase by water-soluble analogues of forskolin.
Analogues of forskolin that are more soluble in water than forskolin have been synthesized and tested for their ability to interact with adenylate cyclase. These analogues are esterified with various heterocyclic amino acids at the 6 beta-hydroxyl position of forskolin or at the 6 beta-hydroxyl or 7 beta-hydroxyl position of 7-desacetyl forskolin. Analogues were tested for their ability to activate rat brain adenylate cyclase, activate detergent-solubilized rat brain adenylate cyclase, increase cyclic AMP in intact S49 wild-type cells, and inhibit the binding of 3H-forskolin to rat brain membranes. Forskolin activated rat brain adenylate cyclase with an EC50 of 4 microM and increased cyclic AMP in intact S49 cells with an EC50 of 5 microM. Analogues esterified at the 7 beta-hydroxyl position had EC50 values that ranged from 4 microM to 15 microM for activating adenylate cyclase in membranes and solubilized preparations, and for increasing cyclic AMP in S49 cells. Analogues esterified at the 6 beta-hydroxyl position with no acyl group at the 7 beta-hydroxyl position were generally less potent than the corresponding 7-acyl analogues with EC50 values that ranged from 30 microM to 100 microM. Interestingly, the diacyl analogues of forskolin containing an acetate group at the 7 beta-hydroxyl position and esterified with heterocyclic amino acids at the 6 beta-hydroxyl position were very potent at stimulating adenylate cyclase, with EC50 values that ranged from 1 microM to 25 microM. The 7-acyl analogues and the 6,7-diacyl analogues inhibited the binding of 3H-forskolin to rat brain membranes with IC50 values that ranged from 20 microM to 70 microM, while the 6-acyl analogues had much higher IC50 values that ranged from 100 nM to 375 nM. Aqueous solutions of forskolin were also produced by dissolving forskolin in solutions of hydroxypropyl-gamma-cyclodextrin. These aqueous solutions of forskolin were equipotent with alcoholic solutions of forskolin in stimulating adenylate cyclase. In conclusion, water-soluble derivatives of forskolin may be useful for increasing cyclic AMP in broken cell preparations or in intact cell preparations where the presence of organic solvents, which are necessary to solubilize forskolin, are detrimental. Alternatively, aqueous solutions of forskolin can be produced by dissolving forskolin in solutions of hydroxypropyl-gamma-cyclodextrin.
Adenylyl Cyclases↗
Quantitative relationships between aggregation of IgE receptors, generation of intracellular signals, and histamine secretion in rat basophilic leukemia (2H3) cells. Enhanced responses with heavy water.
RBL-2H3 cells (a tumor analog of rat mast cells) have plasma-membrane receptors that bind immunoglobulin E, which when aggregated, initiate degranulation. As in other systems, secretion is preceeded by enhanced hydrolysis of inositol phospholipids and by a rise in intracellular Ca2+. Unlike the responses of many other cells, however, both of these earlier events require extracellular Ca2+. The relationship of these events to each other and to the subsequent secretory process is thus unclear. By exposing cells to covalent oligomers of IgE one can demonstrate substantial increases in secretion of histamine by increasing the concentration and size of the oligomers or by using heavy water (D2O) in the medium. We have used such maneuvers to examine the quantitative relationships between aggregation of the receptors and the breakdown of inositol phospholipids, the increase in cytosolic Ca2+ and secretion. Our principal findings were: all treatments that increased secretion, correspondingly increased the changes that precede degranulation. These early events correlated with the degree of aggregation of the receptors even when the stimulatory conditions resulted in maximal secretion. Although the results were insufficient to prove that the hydrolysis of inositol phospholipids is required for the rise in cytosolic Ca2+, the studies with D2O and other observations supported this view. Since a plasma-membrane ion channel for Ca2+ has been implicated in the IgE-mediated rise in cytosolic Ca2+ in RBL 2H3 cells, this in turn suggests a heretofore undescribed role for hydrolysis of inositol phospholipids.
Animals↗
The influence of hemorrhagic hypotension on spinal cord tissue oxygen tension.
In order to investigate the spinal cord surface PO2 (sPO2) reaction to hypovolemic hypotension, nine female pigs (25 kg bw) were premedicated, anesthetized, intubated and artificially ventilated with N2O:O2 = 3:1. Following laminectomy from L3 to L5, sPO2 was measured on the dorsal side of the exposed spinal cord using six gold cathodes (luminal diameter 15 microns) while MAP (mean +/- SD) was lowered in steps of about 10 Torr by bleeding into a reservoir from 69.3 +/- 10.1 Torr to extreme low values of 13.3 +/- 3.1 Torr. Only a slight decrease of lumbar sPO2 (mean +/- SD) from 33.5 +/- 7.2 Torr to 27.6 +/- 4.8 Torr was evaluated from the data in response to MAP reduction to 50.2 +/- 3.5 Torr. Below that value, a marked proportional decrease of sPO2 and MAP was observed (0.65 Torr/Torr) corresponding to pressure passive flow behavior of the Hagen Poisseuille Law ("loss of autoregulation"). Four to five minutes after start of reinfusion both sPO2 and MAP showed an overshoot with maximum values of 54.0 +/- 11.1 Torr resp. 102.6 +/- 18 Torr. Initial values were approximated about 15 min. later. Histograms plotted from the individual sPO2 values of all pigs and of all the different experimental stages showed signs of severe hypoxia only if MAP was reduced below about 30 Torr. In general, this situation was reversible within reinfusion, only one of the nine pigs did not tolerate hemorrhagic hypoxia induced by MAP reduction to less than 30 Torr for at least 5-10 min. Nevertheless, the experiments showed a considerable circulatory stability in the investigated pigs.
Animals↗
The influence of moderate hypothermia on cerebral cortex tissue oxygen tension.
The validity of mechanisms balancing oxygen transport and consumption in hypothermia are difficult to analyse because almost any parameter of the transport system is temperature dependent. Two types of experiments have been performed in the rat cerebral cortex to elucidate this problem: PO2 microelectrode study: 32-37 degrees C body temperature, spontaneous respiration. Measurements within the upper tissue layer of 200-500/microgram thickness (N = 15). PO2 surface electrode study: 26-37 degrees C body temperature, artificial respiration. Investigation of the arachnoidal and pial area (N = 46). Mean tissue PO2 (+/- SD) of neo- and archeocortex declined slightly from 27.8 +/- 14.4 mm Hg at 37 degrees C to 25.5 +/- 15.5 mm Hg at 32 degrees C (p less than 0.05, n = 15). Surface PO2 (sPO2) (+/- SD) in response to hypothermia showed a small decrease from 18.1 +/- 4.5 mm Hg (n = 186) at 37 degrees C to 14.7 +/- 4.3 mm Hg (n = 41) at 26 degrees C (p less than 0.05, N = 46). Despite different initial PO2 values at normothermia both groups of experiments are characterized by the same relatively small hypothermia induced PO2 decrease. Oxygen transport and consumption might be balanced under hypothermia and tissue hypoxia does not exist in the temperature range between 26 degrees and 37 degrees C.
Animals↗
Biochemical and physicochemical determinations in a premyelin fraction obtained by zonal centrifugation in normal mouse and in dysmyelinating mutants (quaking, shiverer, and myelin-deficient).
Myelin and premyelin material denser than myelin were obtained from quaking (Qk), shiverer (Shi), and myelin-deficient (mld) mutant and control mice, using zonal centrifugation on zonal rotor. On these fractions, we performed biochemical analysis (lipids and fatty acid), and, in parallel, we determined the physical structure of membranes by the spin-label method. The hyperfine splitting constant (2 Tll) was used to determine the order of membranes and their rigidity, and frequency of rotation (Vc) was used to measure fluidity. In control mice, the premyelin material contained a lesser amount of sphingolipids than pure myelin, but the relative proportions between hydroxy- and nonhydroxy-cerebroside and sulfatides were similar in the premyelin material and in pure myelin. The premyelin material contained half the alkanes found in the pure myelin and much less very-long-chain fatty acids. The (2 Tll) was lower in the premyelin material, but the (Vc) was similar in myelin and premyelin material. In mutants, the amount of material recovered in the premyelin fraction was reduced in qk, and increased in both shi and mld. The relative amount of sphingolipids were normal in mld, but not in shi mutants, especially in cerebrosides formed with alpha-hydroxylated fatty acids and sulfatides formed with unsubstituted fatty acids. The absolute amounts of sphingolipids were nearly normal in both shi and mld. In the premyelin fraction from qk mutants, both relative and absolute amounts of sphingolipids were drastically altered. In percentage, cerebrosides and sulfatides formed with nonhydroxyfatty acids were dramatically reduced, and, conversely, cerebrosides and sulfatides formed with hydroxyfatty acids were increased. In terms of absolute amount, only cerebrosides and sulfatides formed with nonhydroxyfatty acids were dramatically reduced. In the premyelin fraction, polyunsaturated fatty acids were increased in shi and mld, but decreased in qk. In this mutant, lignoceric (24:0) and nervonic (24:1) acids were drastically reduced. The amount of alkanes in the premylin material from qk and mld was reduced by 50%. The shi fraction was nearly free of alkanes. The maximal apparent coupling constant (hyperfine splitting constant, 2 Tll) was not affected in the mld and qk mutant, but was reduced in the shi mutant premyelin fraction. The Vc was dramatically increased in the qk, slightly decreased in the shi, and close to control in the mld. This work provides additional data on premyelin material prepared in various neurological mutants using continuous gradients in zonal rotor.(ABSTRACT TRUNCATED AT 400 WORDS)
Alkanes↗