Immunochemical cross-reactivity between albumin and solid-phase adsorbed histamine

For production of an antibody against histamine, this was coupled to human serum albumin (HSA) and used for immunization of rabbits. To test the antiserum, an immunoradiometric assay was developed comprising solid-phase bound histamine, antisera and radiolabelled protein A. Titration and inhibition experiments revealed that histamine adsorbed onto a solid-phase could bind the antiserum. However, neither free histamine nor histamine coupled to unrelated carriers could inhibit the binding of antiserum to the solid-phase histamine. Cross-reactivity was demonstrated between HSA and solid-phase bound histamine, as the immunoradiometric assay was inhibited by HSA. This unexpected cross-reactivity was established, as a commercially available antiserum with specificity to HSA without histamine also bound to the solid-phase bound histamine. It is suggested that preparations of antibodies against histamine are tested for this possible cross-reactivity.     

Histamine activates chloride conductance in motor neurons of the lobster cardiac ganglion.

1. Individual motor neurons of the lobster cardiac ganglion were voltage clamped with two microelectrodes. Superfusion of histamine evoked a concentration-dependent membrane current. The mean effective concentration (EC50) for the concentration-effect relationship was 28 microM. 2. The amplitude and polarity of the histamine-activated current depended on intracellular and extracellular Cl- concentration. The membrane potential at which the current polarity reversed was a function of the Cl- equilibrium potential. 3. The histamine-activated Cl- conductance was voltage dependent, increasing with depolarization. As a consequence, the histamine-evoked current showed outward rectification. 4. We conclude that histamine activates a Cl- conductance with biophysical properties similar to the crustacean Cl- conductance activated by gamma-aminobutyric acid (GABA) and to the histamine responses described in lobster olfactory and stomatogastric neurons. 5. The response to histamine was competitively inhibited (IC50 = 7 microM) by cimetidine, an H2 subtype inhibitor in mammals. Ranitidine, pyrilamine, chlorpheniramine, diphenhydramine, and cyproheptadine were 50-100 times less potent than cimetidine. Tubocurarine, a Cl- channel blocker, blocked with an IC50 of 20 microM, but picrotoxin did not begin to inhibit the histamine response until concentrations exceeded 0.1 mM. 6. These results suggest that the response cannot easily be classified with the use of the pharmacological categories developed in mammals. Like the Cl(-)-dependent responses to various neurotransmitters in a number of invertebrates, the histamine response in the lobster cardiac ganglion was inhibited by tubocurarine. 7. Both GABA and histamine had similar effects on the motor neurons, but only GABA inhibited pacemaker bursts. In this respect, GABA more resembles the endogenous inhibitory postsynaptic potential.(ABSTRACT TRUNCATED AT 250 WORDS)     

Histamine suppresses neopterin production in the human myelomonocytoma cell line THP-1

Histamine, an important inflammatory mediator in allergic diseases and asthma, was reported to have modulatory effects on T cells by down-regulating Th1-type cell cytokines like interleukin 2 (IL-2) and interferon-gamma (IFN-gamma). In this study we examined the effect of histamine and the histamine-receptor antagonists cimetidine and diphenhydramine on the production of neopterin after stimulation with IFN-gamma in the myelomonocytoma cell line THP-1. Increasing concentrations of histamine markedly suppressed IFN-gamma induced neopterin formation. Simultaneous preincubation of THP-1 cells with histamine, IFN-gamma and different concentrations of the H(2)-receptor antagonist cimetidine showed a clear antagonizing effect on neopterin formation. In contrast, the H(1)-receptor antagonist diphenhydramine was not able to abrogate the suppressive effect of histamine on neopterin production. Our results suggest, that histamine may be a potent inhibitor of effects or mechanisms induced by IFN-gamma in monocytes/macrophages. Cimetidine, and possibly other H(2)-receptor antagonists, may reverse down-regulatory actions of endogenously formed histamine on activated monocytic cells.     

Histamine receptors in brain vessels of guinea-pig: in-vitro pharmacology and ligand binding

The subtype of histamine receptors in brain vessels of guinea-pig has been characterized by ligand binding and in-vitro pharmacology using selective antagonists. In the basilar artery histamine caused a concentration-related contraction with an EC50 of 1.6 +/- 0.3 microM. H1-receptor blockade with mepyramine and chlorpheniramine caused a displacement to the right of the histamine concentration-response curve with an apparent KD of 0.4 and 4.6 nM respectively, whereas H2-receptor blockade with cimetidine was without effect. Histamine did not induce any dilatory responses of vessels procontracted by 60 mM potassium-containing buffer in the presence or absence of histamine antagonists. Ligand-binding studies with [3H]mepyramine yielded a KD value of 5.5 nM in pial vessel membranes and 1.7 nM in the choroid plexus, confirming the presence of H1-receptors. Nimodipine caused a concentration-related blockade of histamine-induced contractions. Omission of Ca2+ from the extracellular medium for 30 min reduced the contractile responses to histamine in the basilar artery by 96%. Subsequent addition of Ca2+ caused concentration-related contractions which were inhibited by nimodipine. Thus, the histamine H1-receptor activation in guinea-pig basilar artery is coupled to dihydropyridine-sensitive Ca2+ channels.     

A specific enzyme-linked immunosorbent assay for definition of the IgG antibody response to disulphide-conjugated D-penicillamine in the rabbit

An enzyme-linked immunosorbent assay (ELISA) has been developed for unambiguous detection of antibodies against the sulphydryl drug D-penicillamine (PA) and its disulphide-conjugated metabolites. Disulphide-linked PA human serum albumin (PA-HSA) conjugates for use as coating antigens were prepared by a range of procedures employing oxidation with either potassium ferricyanide (0.1 M) or cupric sulphate (5 ppm). A satisfactory degree of conjugation was achieved by both oxidative procedures. Hapten density and antigenicity were increased when urea-denatured rather than non-denatured HSA was used. In 2 out of 3 rabbits, a specific IgG anti-PA response was detected following monthly injection of PA keyhole limpet haemocyanin (PA-KLH) in Freund's complete adjuvant. In the third rabbit, any anti-PA activity was obscured by a high level of binding to HSA. The anti-PA response was slow to develop in the 2 responder rabbits (requiring four injections) and was of low intensity (antibody titres less than 6,000). In contrast, the IgG antibody response to the structurally related drug captopril (CP), administered under identical conditions, was rapid in onset and of greater intensity (titres greater than 6,000 after one injection of CP-KLH). The hapten specificity of the IgG anti-PA-HSA antisera was defined by ELISA inhibition assays. Binding of IgG to PA-HSA was inhibited by PA, PA disulphide, PA cysteine and disulphide-linked PA-HSA conjugates, but not by PA acetone (thiazolidine ring-linked PA), CP, or unconjugated HSA. The inhibitory preparations were inactive in unrelated ELISAs.(ABSTRACT TRUNCATED AT 250 WORDS)     

Inhibition of renal alkaline phosphatase by cimetidine

Alkaline phosphatase (ALP) belongs to hydrolase group of enzymes. It is responsible for removing phosphate groups from many types of molecules, including nucleotides and proteins. Cimetidine (trade name Tagamet) is an antagonist of histamine H2-receptor that inhibits the production of gastric acid. Cimetidine is used for the treatment of gastrointestinal diseases. In this study the inhibitory effect of cimetidine on mouse renal ALP activity was investigated. Our results showed that cimetidine can inhibit ALP by uncompetitive inhibition. In the absence of inhibitor the V(max) and K(m) of the enzyme were found to be 13.7 mmol/mg prot.min and 0.25 mM, respectively. Both the Vmax and Km of the enzyme decreased with increasing cimetidine concentrations (0- 1.2 mM). The Ki and IC(50) of cimetidine were determined to be about 0.5 mM and 0.52 mM, respectively.     

Inhibition of bone resorption in tissue culture by H1-receptor antagonists

Mouse bone culture studies show that several representative H1-receptor antagonists, promethazine hydrochloride, pyrilamine maleate, tripelennamine hydrochloride, and diphenhydramine hydrochloride, inhibit parathyroid extract-stimulated bone resorption. The H2-receptor antagonists, metiamide and cimetidine, are ineffective. In view of the finding that histamine and histamine agonists did not stimulate bone resorption, it is unlikely that histamine receptors are involved in mediating parathyroid extract-stimulated bone resorption. Because H1-receptor antagonists bind to phospholipids and have been shown to influence membrane structure and function, it is suggested that they inhibit bone resorption by a mechanism that depends on their membrane-stabilizing effect.     

The biologic activity of mast cell granules. V. The effects of antihistamine treatment on rat cutaneous early- and late-phase allergic reactions

Mast cell-dependent late-phase allergic reactions (LPR) as sequelae of immediate hypersensitivity responses (IR) occur in both human and rat skin; thus the rat has served as a useful model to investigate the pathogenesis of cutaneous LPR. To analyze the roles that histamine might play in the generation of rat LPR, the effects of H1 and/or H2 antihistamines on both LPR and antecedent blueing responses (IR) were investigated. Systemically administered diphenhydramine and cimetidine, alone or in combination, reduced blueing reactions to histamine. However, blueing responses to anti-IgE were only partially abrogated by antihistamine treatment with diphenhydramine alone or the combination of antihistamines. Diphenhydramine treatment alone partially inhibited the histologic intensity of LPR in a dose-dependent manner. Although cimetidine treatment alone had no inhibitory effect, it potentiated the diphenhydramine-induced inhibition of LPR. The inhibitory action of antihistamine treatment was apparent only in reactions elicited by anti-IgE or mast cell granules containing histamine, since LPR caused by histamine-free mast cell granules were not affected by antihistamines. This observation suggests that the inhibitory effect of antihistamines on LPR was the result of a specific blockade of histamine receptors rather than the result of a nonspecific suppressive effect. Our findings demonstrate that cutaneous inflammation generated as a result of mast cell degranulation can be significantly reduced by treatment with H1 and H2 histamine receptor antagonists.     

Binding of [3H]cimetidine to rat brain tissue

Binding of [³H]cimetidine to rat brain tissue was investigated, and a saturable binding with dissociation constant 0.22±0.05 M found. This binding is inhibited by a range of imidazole-derived histamine H₂-receptor antagonists, but not by a number of non-imidazole H₂-receptor antagonists. It is concluded that the [³H]cimetidine binding site in rat brain tissue that is labelled in these experiments is not the histamine H₂-receptor.     

Immunologic cross-reactivity between different albumin-bound isocyanates

Sera of six workers with conclusive evidence for IgE-mediated sensitization to isocyanates were used for evaluation of immunologic cross-reactivities among eight different isocyanate-protein conjugates. In all cases RAST and/or skin-test investigations revealed the presence of IgE antibodies reacting specifically with HSA conjugated with those isocyanates to which workers were exposed as well as with other isocyanates with which they had not been in contact. By the RAST inhibition technique, moderate to strong mutual cross-reactivities between all tested isocyanate-HSA conjugates--even between aromatic and aliphatic ones--could be demonstrated in tests with five sera. The magnitudes of cross-reactivities differed, however, from one patient to another. One serum contained IgE antibodies that were almost completely specific to TDI-HSA; with this serum only weak cross-reactivities with other isocyanate conjugates could be demonstrated. These results indicate the predominance of closely related antigenic determinants in HSA conjugated with different isocyanates. The common antibody-binding regions are obviously recognized to different extents by antibodies of clinically sensitized workers, indicating individual differences in specificities and avidities of antibody populations. Nearly complete lack of IgE binding of ovalbumin-bound TDI in RAST and RAST inhibition indicates carrier-specific antigenicity of isocyanate-protein conjugates. In addition, since unmodified HSA did not bind IgE, antigenic determinants of the conjugates studied should be predominantly formed by the isocyanate-protein bond regions and concurrently by neighboring amino acid residues of the HSA molecule.     

Analysis of 3H-histamine interaction with lymphocytes: receptor binding or uptake?

The immunoregulatory role of histamine is presumably mediated by specific receptors on the plasma membrane of lymphocytes. However, using murine spleen cells and a whole cell assay commonly applied in hormone receptor studies, specific histamine receptors with an affinity higher than that of non-specific binding could not be identified. Nevertheless, approximately 30% of the totally bound histamine was undissociable over a range of added histamine concentrations (9 X 10(-6)-1 X 10(-2) M). Lectin stimulation of spleen cells caused an additional two-fold increase of undissociable histamine. The H1 receptor antagonist, diphenhydramine, blocked histamine uptake, whereas the H2 receptor antagonist, cimetidine, had no effect. Binding experiments carried out at 4 degrees C demonstrated that the amount of undissociable histamine was much reduced. Even at 4 degrees C, evidence for specific membrane associated histamine receptor could not be obtained. It was therefore concluded that lymphocytes take up histamine by an energy-dependent mechanism inhibitable by diphenhydramine but not cimetidine, and that the usual hormone receptor methodology did not allow the identification of specific membrane associated histamine receptors.     

Successful treatment of chronic idiopathic urticaria and angioedema with cimetidine alone

We have studied a 50-year-old white man with chronic urticaria and angioedema who has responded to treatment with cimetidine alone for over 2 yr. In a double-blind, placebo-controlled study, cimetidine alone was at least as effective as chlorpheniramine in relief of urticaria and angioedema. Additionally, cimetidine significantly inhibited (p less than 0.01) the wheal response to histamine when it was compared to placebo. The inhibition of wheal response to histamine by cimetidine was significantly higher (p less than 0.05) than chlorpheniramine. The presence of predominantly H2- rather than H1-histamine receptors in the cutaneous blood vessels may be responsible for the therapeutic effects of cimetidine in this patient.     

The Regulatory Effect of Histamine on the in Vitro Synthesis of IgE

The effect of histamine and its H1 and H2 antagonists, chlorpheniramine and cimetidine, on the in vitro, PWM-induced, synthesis of IgG and IgE was studied. Histamine had no effect, and cimetidine had a slight inhibitory action. In contrast, chlorpheniramine induced marked suppression of both IgE and IgG synthesis. This effect could not be attributed to drug-induced cytotoxicity. These results suggest that the modulatory effect of histamine on antibody production involves predominantly H1 receptors.     

Inhibition of tryptase release from human colon mast cells by histamine receptor antagonists

The main objective of this study was to investigate the ability of histamine receptor antagonists to modulate tryptase release from human colon mast cells induced by histamine. Enzymatically dispersed cells from human colon were challenged with histamine in the absence or presence of the histamine receptor antagonists, and the tryptase release was determined. It was found that histamine induced tryptase release from colon mast cells was inhibited by up to approximately 61.5% and 24% by the H1 histamine receptor antagonist terfenadine and the H2 histamine receptor antagonist cimetidine, respectively, when histamine and its antagonists were added to cells at the same time. The H3 histamine receptor antagonist clobenpropit had no effect on histamine induced tryptase release from colon mast cells at all concentrations tested. Preincubation of terfenadine, cimetidine or clobenpropit with cells for 20 minutes before challenging with histamine did not enhance the ability of these antihistamines to inhibit histamine induced tryptase release. Apart from terfenadine at 100 microg/ml, the antagonists themselves did not stimulate tryptase release from colon mast cells following both 15 minutes and 35 minutes incubation periods. It was concluded that H1 and H2 histamine receptor antagonists were able to inhibit histamine induced tryptase release from colon mast cells. This not only added some new data to our hypothesis of self-amplification mechanisms of mast cell degranulation, but also suggested that combining these two types of antihistamine drugs could be useful for the treatment of inflammatory bowel disease (IBD).     

Inhibition of allergic histamine release by azelastine and selected antiallergic drugs from rabbit leukocytes

The ability of azelastine to inhibit allergic histamine release from rabbit mixed leukocytes was studied and compared with selected antiallergic drugs. Azelastine, ketotifen, diphenhydramine, theophylline and disodium cromoglycate (DSCG) produced concentration-dependent inhibition of allergic histamine release from rabbit basophils. The concentrations inhibiting histamine release by 50% (IC50; microM) were as follows: azelastine = 4.5; ketotifen = 9.5; diphenhydramine = 18.9; theophylline = 56.9; DSCG = greater than 1,000. DSCG was added to the cells immediately prior to antigen challenge. All other drugs were preincubated for a period of 10 min prior to antigen challenge. At the IC50 level, azelastine is about 2, 4, 13 and greater than 200 times as effective as ketotifen, diphenhydramine, theophylline and DSCG, respectively. The IC50 of azelastine following 0, 10 and 30 min preincubation were 2.4, 1.9 and 3.5 microM, respectively. These observations showed: (1) azelastine is capable of acting rapidly on basophils and of inhibiting allergic histamine secretion, and (2) the prolongation of the preincubation time of azelastine up to 30 min with rabbit leukocytes did not exhibit any sign of tachyphylaxis (loss of activity). In conclusion, azelastine is a potent inhibitor of allergic histamine secretion from the leukocytes of ragweed-sensitized rabbits.     

Production of a monoclonal antibody to autoantigenic components of human glomerular basement membrane

Histamine (10(-3)-10(-8) M) inhibits PHA-induced proliferation of human peripheral blood lymphocytes (HPBL). Inhibition is detected at low concentrations of PHA but is rarely observed at high PHA concentrations. The histamine type II (H2) receptor agonists dimaprit, impromidine and 4-methylhistamine (4MH) inhibit HPBL proliferation and the H2 antagonist, cimetidine, reverses histamine-induced suppression of HPBL proliferation. Lymphocyte proliferation is also inhibited by the H1 receptor agonists, 2-pyridylethylamine and 2-thiazoylylethylamine, but only at high concentrations (10(-3) and 10(-4) M). The H1 agonist 2-methylhistamine, suppresses PHA-induced proliferation of HPBL in analogous fashion to histamine. This effect is reversed by cimetidine but not by diphenhydramine suggesting that an H2 receptor interaction is involved.     

Effect of histamine antagonists and agonists on IgE production in mice.

This study was undertaken to investigate the effect of histamine, its receptor antagonists and agonists on IgE antibody production in mice. BALB/c mice were immunized intraperitoneally with 1 mg alum plus 30 micrograms of Ag90, the antigen of Japanese occupational asthma. Histamine receptor antagonists were administered before the immunization and simultaneous injection of histamine. The mice were bled 14 days after immunization and anti-Ag90 IgE antibody was obtained. Titers of the antisera were measured by passive cutaneous anaphylaxis reaction in Sprague-Dawley rats. Treatment with histamine only did not affect the level of specific IgE antibody. Administration of H1 antagonist or H2 agonist suppressed IgE production significantly. In contrast, treatment with H2 antagonist or H1 agonist augmented the IgE antibody production. Injection of H1 + H2 antagonists had no effect on the antibody production. These results suggest that histamine suppressed specific IgE production via H2 receptors and enhanced it through H1 receptors in the induction phase of the system.     

CI-922--a novel, potent antiallergic compound--I. Inhibition of mediator release in vitro

CI-922 (3,7-dimethoxy-4-phenyl-N-1H-tetrazol-5-yl-4H-furo[3,2-b]-indole- 2-carboxamide, L-arginine salt) is a novel antiallergy compound which inhibits the release of the inflammatory mediators histamine and leukotriene (LT) from stimulated cells. CI-922 showed potent, effective inhibition of antigen-induced mediator release from human basophils and isolated guinea pig lung. The drug inhibited ragweed or housedust-induced histamine release from basophils of allergic human donors (IC50 = 8.6 microM). The antiallergy agents proxicromil (IC50 = 80 microM) and cromolyn (100 microM) were less potent than CI-922 or inactive, respectively. In fragmented lung from actively sensitized guinea pigs, CI-922 (IC50 = 1.5 microM), blocked the antigen-induced production of LT and was a more potent inhibitor of histamine release (IC50 = 13.4 microM) than proxicromil (IC50 = 72.9 microM), or cromolyn (inactive at 1 mM). CI-922 (IC50 = 0.9 microM) completely inhibited repeated contractions of guinea pig lung strips that were induced by low antigen concentration in the presence of antihistamine (H1). Nordihydroguaiaretic acid (NDGA) (IC50 = 2.8 microM), proxicromil (IC50 = 6.2 microM) and the LT antagonist FPL-55712 (IC50 = 3.3 microM) also were fully effective, but cromolyn (300 microM) was inactive. In other experiments, CI-922 (IC50 = 7.0 microM) inhibited a strong, nonrepeatable lung contraction induced with high antigen concentration (histamine responses blocked), and was six times more potent than FPL-55712. Other investigations in isolated tissue preparations showed CI-922 to be a weak inhibitor of LT or histamine-induced effects with no anticholinergic activity.(ABSTRACT TRUNCATED AT 250 WORDS)     

Carbodiimide or periodate method to prepare peroxidase hydrazide for its use in immunoassay

Peroxidase hydrazides were prepared by conjugating horseradish peroxidase (HRP) to adipic acid dihydrazide (ADH) by carbodiimide or periodate oxidation method. The resulting HRP hydrazides (ADH-HRP) were conjugated to cortisol-21-hemisuccinate (cortisol-21-HS) by forming diimide bonds using the N-hydroxysuccinimide (NHS) carbodiimide mediated reaction. The prepared cortisol-21-HS-ADH-HRP enzyme conjugates were utilized for the development of an enzyme linked immunosorbent assay (ELISA) for direct estimation of cortisol. To the cortisol antibody coated microtiter wells, standard or serum sample (50 microL), along with 100 microL of cortisol-21-HS-ADH-HRP enzyme conjugate (ADH-HRP used is prepared by either carbodiimide or periodate oxidation method), was incubated for 1 hr at 37 degrees C. Bound enzyme activity was measured by using tetramethyl benzidine/hydrogen peroxide (TMB/H202) as substrate. The sensitivity, specificity, and recovery of the assays were found to be identical when ELISAs were employed with cortisol enzyme conjugates prepared by conjugating cortisol-21-HS to HRP hydrazide, made either by the carbodiimide method or periodate oxidation method.