Comparison of the action of calcium antagonists on basophil histamine release

The inhibitory capacity of calcium antagonists on basophil histamine release was examined in allergic patients and in controls. All dihydropyridines tested (nifedipine, nimodipine, nitrendipine, nicardipine, felodipine) dose-dependently inhibited anti-IgE- and A23187-induced release with an order of potency of felodipine greater than nicardipine greater than nifedipine = nimodipine = nitrendipine. Only the inhibition induced by felodipine and nicardipine on anti-IgE-induced release could be counteracted by increasing extracellular calcium. Diltiazem, not belonging to the dihydropyridines, was a weak inhibitor. A combination of felodipine and verapamil in low concentrations exerted a synergistic inhibitory effect on histamine release, whereas this was not the case with other combinations of antagonists. The results suggest differences in the mode of action of the 1.4-dihydropyridines. This might be of significance in the search for calcium antagonists suitable in the treatment of allergic diseases.     

Influence of Neuraminidase and N-Acetylneuraminic Acid on Basophil Histamine Release in Vitro

Since N-acetylneuraminic acid (NANA) in cell membrane glucocalyx mediates or modulates a variety of actions, such as mediator release, we examined a possible modulating role of this amino sugar in histamine release from human basophil leukocytes. Removal of NANA from the cell membrane by the enzyme neuraminidase caused a dose-dependent histamine release. Removal of smaller amounts of NANA enhanced histamine release induced by anti-IgE, Concanavalin A and the calcium ionophore A23187, and reduced the interval between addition of antigen and initiation of histamine release. Pretreatment with free NANA had the opposite effects, i.e. a diminished and delayed maximal histamine release. The hypothesis that NANA in the cell membrane modulates the cellular response to stimulation was further substantiated by demonstrating that the altered response was reflected by a change in the sensitivity of the cell to extracellular calcium. NANA in the cell membrane glucocalyx thus seems to modulate the basophil response to stimulation by modulating transmembraneous calcium transport.     

Factors determining spontaneous histamine release from human basophils purified with Percoll gradients and Dynabeads

Identification of factors influencing histamine release from purified and cultured basophil leukocytes is important for proper interpretation of results obtained on histamine release. This paper describes factors that influence spontaneous histamine secretion from human basophil leukocytes purified on Percoll gradients, followed by negative selection with Dynabeads. Anti-IgE and recombinant human interleukin-3 were used as model stimulants, and the purified basophil leukocytes were stimulated for 10 min and 6 h. The effect of the following conditions was examined: Percoll temperature, cell-suspension density, and serum in the media. The results showed that low Percoll temperature, high cell-suspension density, and the presence of serum in the media decreased spontaneous histamine release and increased maximal net histamine release upon stimulation.     

Regulation of histamine release from human basophil leucocytes: role of H1, H2 and H3 receptors

A novel class of histamine receptors (H3), controlling histamine synthesis and release, was described in rat and human brain and peripheral nerve endings. The present study was undertaken to evaluate whether H3 receptors contribute to the regulation of histamine release from human basophils. Basophil leucocytes were incubated with a H3 antagonist (thioperamide; concentrations ranging from 1 nM to 10 microM) or with a H3 ((R)alpha methyl-histamine; concentrations ranging from 1 to 100 mM), and subsequently were stimulated with optimal doses of anti-IgE and formyl-methionyl-leucyl-phenyl-alanine (f-met peptide). No significant modifications of histamine release were observed after incubation either with the H3 agonist or with the H3 antagonist. By contrast, a H2 antagonist (cimetidine; concentrations ranging from 1 to 100 microM) exerted a dose-dependent enhancing effect on anti-IgE- and, to a lesser extent, on f-met peptide-induced histamine release. A H1 antihistamine (chlorpheniramine; concentrations ranging from 100 nM to 1 microM), at the highest concentration employed, displayed an inhibitory activity on IgE-dependent and IgE-independent histamine release. Exogenous histamine was shown to exert a dose-dependent inhibitory effect on two-staged anti-IgE-induced histamine release. Taken as a whole, these results suggest that H3 receptors are not involved in the regulation of histamine release from human basophils; by contrast, H2 receptors participate in controlling histamine release from human basophils, as previously demonstrated by other authors.     

Inhibitory Effect of Glucocorticosteroids on Anti-IgE-Induced Histamine Release from Human Basophilic Leukocytes: Evidence for a Dual Mechanism of Action

Anti-IgE-induced histamine release from human leukocytes is inhibited when the cells before challenge are cultured overnight in the presence of glucocorticoids (GCSs). The present report suggests that the GCSs might exert their effect by at least a dual mechanism of action. Histamine release was induced by a suboptimum concentration of anti-IgE. When the release recorded in the presence of the steroid is plotted against the release recorded in its absence, the data points of several experiments fit a regression line characterized by two parameters: its slope and its intercept with the abscissa. Structure-activity examination with selected GCSs indicates that the orders of potency for affecting these two parameters are not identical. Furthermore, pulse experiments suggest that the cells require different times of contact with the steroid to express inhibition according to the two parameters. The removal of adherent cells or platelets did not markedly affect the degree of leukocyte histamine release or its inhibition by a given GCS, suggesting that the steroid interacts directly with the basophil. Finally, steroid-induced inhibition was not affected by the putative phospholipase A2-inhibitor p-bromophenacylbromide (BPB) or the 5-lipoxygenase inhibitor nordihydroguaiaretic acid (NDGA).     

Studies on Hypersensitivity to Bacterial Antigens in Intrinsic Asthma

Twelve children, aged 4 to 14 years, with moderate to severe intrinsic asthma (IA) were studied. Symptom-Score charts were used to confirm the relationship of acute respiratory tract infections to exacerbations of asthma. Hypersensitivity to eight commonly occurring bacteria from the normal flora of the upper respiratory tract was studied by skin test, by crossed immunoelectrophoresis, and by basophil histamine release in vitro, using ultrasonicates of the bacteria as antigens. Skin tests were all negative. All children contained low titers of precipitating antibodies against most of the bacteria, but in this respect they did not differ from normal children. In contrast, release of histamine was induced in leukocytes from the IA children by all, or most sonicates, while such reactions, were less frequent in control children. The pattern of responses indicated an element of specificity. These was no correlation to precipitating antibodies, or to the microbial flora of the children. Positive responses were characterized by low values of maximal histamine release, and by a tendency to fluctuations with time. Because of these fluctuations, and because the IA children and control children were tested on separate occasions, we cannot be certain as to the real difference between these two groups. Our studies do, however, demonstrate that water-soluble constituents of all the bacterial strains tested were capable of causing the release of histamine in vitro, but that this phenomenon is not restricted to IA. The clinical significance of these findings awaits further investigations on the mechanism(s) of release in vitro by such agents.     

Impairment of the inhibitory effect of sodium on basophil histamine release in patients with systemic sclerosis

It has been demonstrated that Na⁺ down-regulates IgE-dependent and IgE-independent histamine release from basophils of normal subjects. The aim of this study was to evaluate whether Na⁺ exerts its inhibitory effect on basophil histamine release in patients with systemic sclerosis (SSc). Peripheral blood leucocytes were stimulated with anti-IgE, n-formyl-methionyl-leucyl-phenylalanine (fMLP) and IL-3 in the presence of high and low Na⁺ concentrations, and histamine release was measured by a fluorometric method. The dose–response curves of histamine release induced by the above stimuli were similar in SSc patients (n = 15) and in normal subjects (n = 39). Na⁺ removal from the extracellular medium and its isosmotic replacement with choline chloride led to a significant increase of anti-IgE-and fMLP-induced histamine release in normal subjects, but not in SSc patients. In the former population, histamine release induced by an optimal dose of anti-IgE (1/5000) was 26.4 ± 3.1% in high Na⁺ and 59.3 ± 3.5% in low Na⁺ (mean ± s.e.m., P< 0.0001), whereas in the latter population mean histamine release was 20.4 ± 5.1% in high Na⁺ and 15.8 ± 2.9% in low Na⁺ (P NS). A similar trend was observed when basophils were stimulated with fMLP. Na⁺ exerted a dose-dependent inhibitory effect on anti-IgE- and fMLP-induced histamine release in normal subjects, but not in SSc patients. IL-3-induced histamine release from basophils of SSc patients was increased in a low-Na⁺ solution, but to a lesser extent when compared with normal controls. Therefore basophils from normal subjects and SSc patients behave in a different way when stimulated in a low-Na⁺ medium. The inhibitory effect of Na⁺ on basophil histamine release is impaired in SSc patients, and this abnormality could contribute to basophil dysfunction.     

In vivo and ex vivo inhibitory effects of loratadine on histamine release in patients with allergic rhinitis

Background The aim of this study was to evaluate the in vivo and ex vivo effects of the H.-antagonist loratadine on histamine release.
Methods The study was designed as a double-blind, crossover trial. Ten patients with allergic rhinitis due to Dermatophagoides pteronyssinus were treated with loratadine (10 mg daily p.o.) and with placebo for 1 week, with a 2-week interval between the two treatments. Nasal lavages with saline solution were done before and after challenge with the relevant allergen at the end of treatments with loratadine and placebo. Venous blood was taken after treatments, and basophil histamine release induced by anti-IgE (10 μg/ml), N-formyl-methionyl-leucyl-phenylalanine (fMLP, 1 μM). and Ca²⁺ ionophore A23187 (1 μM) was evaluated by ati automated fluorometric method.
Results Treatment with loratadine attenuated early antigen-tnduced nasal obstruction, rhinorrhea. and itching. Nasal symptoms were accompanied by a significant histamine release in the nasal lavages collected 5 min after stimulation when the patients received placebo (median 4 ng/ml, range 1-28; P<0.05). After treatment with loratadine, histamine release in the 5-min postchallenge lavages was almost abrogated (median 0.5 ng/ml, range 0-3; P<0.01 vs placebo). Median anti-IgE-induced histamine release from basophils was 41.9% (range 27.8-79.2) after placebo and 30.0% (range 1.7-73.3, P < 0.05) after loratadine. Active treatment exerted an inhibitory effect also on basophil histamine release induced by fMLP and Ca²⁺ ionophore A23187.
Conclusions Treatment for 1 week with loratadine reduces allergen-mduced nasal symptoms and inhibits in vivo and ex vivo histamine release in patients with allergic rhinitis.     

Human auto-anti-idiotypic antibodies to mite-specific IgE can degranulate human basophils in vitro

Background Anti-idiotypic antibodies (anti-Ids) to specific IgE antibodies are formed spontaneously during an anti-allergen immune response and can be induced by immunotherapy. Although anti-Ids can down-regulate the production of IgF. antibodies, at least in experimental models, their possible role in the modulation of target cell reactivity remains ill-defined. Objective The capacity of human anti-Ids to modulate the release of histamine was examined in an in vitro system of human basophil degranillation. Anti-Ids were prepared from the serum of six Dermatophagoides pteranyssinus (DP)-hypersensitive patients suffering from atopic dermatitis and who had never been desensitized. Basophils were obtained from the blood of atopic donors. The extent of histaminc release was determined using a fluorometric assay. Results We show that: anti-Ids trigger the release of histamine in an allergen-specific, dose- and IgE-dependent manner; the release is not due to the presence of allergen and/ or anti-IgE antibodies: and that the degranulating activity can be removed by absorption with affinity-purified anti-Dp antibodies of the corresponding patient. Conclusion These results indicate that spontaneously produced human anti-Ids can modulate the reactivity of human basophils.     

Basophil histamine release and lymphocyte proliferation tests in latex contact urticaria

Basophil histamine release and lymphocyte proliferation tests were examined with latex allergen prepared from surgical gloves in 15 patients with latex contact urticaria. The basophil histamine release test (BHRT) yielded positive results in 13/14 (93%) patients, whereas commercial latex RAST was positive in only 9/15 (60%) patients. Lymphocyte proliferation test (LPT) was positive in 3/15 (20%) patients, suggesting that cell-mediated immune reactions may also occur in latex allergy. However, patch tests to latex were negative and neither were epidermal Langerhans cells able to present latex antigen to T lymphocytes in vitro.     

Sensitive Glass Microfibre-Based Histamine Analysis for Allergy Testing in Washed Blood Cells

The new microfibre method for allergy testing is based on basophil histamine release after challenge with suspected allergens in samples of 50 microliter washed blood cells. Released histamine is bound to microfibres and measured after removal of interfering substances by washing. The microfibre method was compared with the conventional leukocyte histamine release assay in 18 allergic patients tested with 10 different allergens. It was found that the same individuals responded with histamine release to the same allergens in both assays, and the number of responders was almost identical. Also the dose-response curves and the cell sensitivity were almost identical, which further substantiated identity between the results obtained by the new microfibre method and the conventional assay. A comparison between the microfibre method and in vivo provocation tests showed good agreement when comparing the number of positive and negative responses in these test. The new method overcomes the problems in allergy testing, where only small amounts of blood are available and many tests have to be carried out.     

Complexity of Lectin-Mediated Reactions in Bacteria-Induced Histamine Release

We have earlier suggested that bacteria-induced histamine release is caused by different mechanisms, including allergic and non-immunological mechanisms, and that the latter probably depends on lectin-mediated reactions. Two possibilities of lectin-mediated reactions were examined in this study, bacterial surface lectins bind to sugars on the basophil cell membrane leading to histamine release, and the reverse reaction where bacterial aminosugars react with lectins on the basophil cell surface. In the bacterial histamine release caused by the Staph. aureus strain Wood 46 it was possible to demonstrate a reverse reaction, but not a bacterial lectin-mediated reaction. The reaction seems to be complex, as lower concentrations of sugars might potentiate the release of histamine by binding to the target cell or bacteria, while the release is inhibited by higher concentrations.     

Release of histamine from human leukocytes by one preparation of IgG oligomers

In preliminary experiments aimed at investigating the effect of covalently cross-linked human myeloma subclass proteins on histamine release from human leukocytes, we observed one preparation (designated here IgG-HR) made from pooled, purified immunoglobulin G, which consistently released histamine from these cells. Dimers and trimers, but not monomers isolated from columns of Sephadex G-200 and Ultrogel AcA22 following incubation of immunoglobulin G (Nordic Laboratories) with dimethyl suberimidate, released histamine from cells of all donors tested. In contrast, cells from the same donors showed variable responsiveness to dimers of IgE (prepared by similar techniques) or to anti-IgE. IgG-HR failed to release histamine from a "basophil-rich" mononuclear cell preparation depleted of most of the erythrocytes, platelets, neutrophils and eosinophils by centrifugation through a Ficoll-Hypaque cushion. The data suggest that IgG-HR was releasing histamine indirectly from basophils by first interacting with another cell. IgG oligomers prepared from different sources of pooled, purified IgG failed to release histamine. Although we did not have sufficient IgG-HR to adequately define this releasing activity, we feel that the data represent a potentially novel, if rare, mechanism of mediator release involving basophils and another cell.     

Passive sensitization of basophil leukocytes from a non-atopic adult by plasma from allergic children

Basophil leukocytes from a non-atopic donor, who responded well to anti-IgE, were depleted of their native membrane-bound IgE by acid treatment and passively sensitized with plasma containing either Phleum pratense-, Dermatophagoides pteronyssinus- or dog dander- specific IgE obtained from 18 allergic children. Histamine release was then performed on the passively sensitized cells and the results were compared with those of bronchial provocation test (BPT), allergen-specific serum IgE (RAST), skin prick test (SPT), and conventional histamine release test (HR). A high coincidence rate was found between BPT, RAST and histamine release after passive sensitization (HR-PS), but compared to HR, it was lower. This could be because several of the patients had non-responding basophils (i.e. no release after challenge with anti-IgE) in the conventional histamine release assay. The lower rate was not related to a lack of antigen-specific IgE, since after passive sensitization of basophils, anti-IgE and allergen provocation could induce release. It is concluded that plasma from allergic children with non-responding basophil leukocytes contain antigen-specific IgE capable of binding to Fc-receptors on the basophils of a non-atopic donor. In addition it was found that the plasma could change the cell reactivity (maximal histamine release) of the donor cells, since the amount of histamine released varied according to the plasma used for passive sensitization. The lack of histamine release response in some patients could be because their own membrane-bound IgE is unable to induce mediator release or, more likely, activation of one or more of the subcellular steps involved in the release is impaired.     

Relevance of basophil histamine release changes during venom immunotherapy

We investigated the effect of rush and long-term venom immunotherapy on histamine release parameters in bee venom allergic patients. Ten patients received rush venom immunotherapy, and histamine release data were obtained immediately before and after treatment. 17 patients were assessed by histamine release 24 to 63 months after termination of long-term venom immunotherapy. A control group of 10 non-allergic subjects was included in this study. Histamine released from whole blood was determined in a sensitive radio-enzymatic assay using a single isotope technique. Bee venom phospholipase A-induced histamine release from whole blood proved to be a test procedure of high specificity and sensitivity. Eight of 10 untreated patients and no control subject showed significant antigen-induced histamine release. Results obtained from patients immediately after successful rush venom immunotherapy showed an important decrease (mean 45.9%) of total histamine content of basophil leukocytes in all patients. Antigen-induced maximum histamine release was found to be increased in one, decreased in two and unchanged in seven patients. In patients who received long-term immunotherapy cell sensitivity to phospholipase A was significantly lower than in a group of untreated patients (P less than or equal to 0.002). These results suggest that even years after discontinuation of immunotherapy, histamine release parameters reflect patients' protection from systemic sting reactions as assessed by sting challenges. Histamine depletion of basophils induced by rush immunotherapy may play an important role in patients' protection immediately after termination of the rush regimen.     

Ex vivo pharmacologic modulation of basophil histamine release in asthmatic patients

Histamine is one of a range of mediators which play an important role in asthma, and the "releasability" of basophils has been shown to be upregulated in this disease. In vitro , β₂-agonists and to a lesser extent corticosteroids have been shown to reduce histamine release. The ex vivo effects of salmeterol and inhaled corticosteroids on histamine release were studied in 78 asthmatic patients with variable disease severity and 20 control subjects. Spontaneous and anti-IgE-induced histamine release was measured in all subjects. Fifteen patients were not receiving any form of treatment, 42 were treated with inhaled corticosteroids, and 21 received inhaled corticosteroids and salmeterol. Seven patients treated with inhaled corticosteroids and seven patients treated with inhaled corticosteroids and salmeterol were tested twice to assess the effect of salmeterol on histamine release. Nine patients treated with inhaled corticosteroids were tested before and after 1 month of salmeterol treatment to determine the possible inhibition by salmeterol. Patients who were treated with inhaled corticosteroids and salmeterol showed significantly lower levels of spontaneous histamine release (median: 2.5%) than untreated (5.2%) and inhaled corticosteroids-treated asthmatics (3.4%). No tachyphylaxis to salmeterol was observed when patients were tested twice at a 3-month interval. This study suggests that salmeterol may have an additive anti-inflammatory effect with inhaled corticosteroids, although this hypothesis must be tested by further studies involving cells obtained by bronchoalveolar lavage and studies with bronchial biopsies.     

A new glass microfibre-based histamine analysis for allergy testing in children

A new microfibre method for allergy testing measuring histamine release from human basophil leukocytes is described. Samples of 50 microliter washed blood are challenged with the suspected allergens. Released histamine is bound to microfibres and measured by a spectrofluorometrical method after removal of interfering substances by washing. The microfibre method (HR-MM) was compared to the conventional histamine release assay using the Ficoll-Hypaque gradient method (HR-FH) in 19 allergic children tested with one of three allergens. In addition, a comparison was made between the microfibre method and in vivo provocation tests, i.e. skin prick test (SPT), bronchial provocation test (BPT) and allergen specific serum IgE (RAST). It was found that the same individuals responded with histamine release to the same allergens in both histamine release assays, and the dose-response curves were almost identical. A positive correlation was found between the in vivo and in vitro tests. Thus it is concluded that the new method can provide reproducible, analytically precise (at the nanogram level) histamine release results in pediatric cases where: a positive SPT does not correlate with case history; BPT may be considered too hazardous or inconvenient; confirmation of negative or inconclusive SPT or RAST is needed. In contrast to other histamine release assays it is a convenient diagnostic tool in children since only small amounts of blood are needed and at least 96 tests can be carried out in 2 1/2 h.     

Basophil Histamine Release in Cord Blood Regulatory Role of IgE

Thirty-two cord blood samples were studied for histamine releasing capability by using a sensitive glass microfibre-based histamine analysis. Histamine was obtained after challenge with anti-IgE in 24 of the 32 samples. However, the net release in cord blood was only 25% of that in adult blood and no relationship was found between histamine release response, total IgE in cord plasma, and a family history of atopic diseases. The low histamine release in cord blood seemed to be associated with the immunological IgE receptor complex activation and not with an immature basic cell function, since the calcium ionophore A23187 which bypasses the receptor complex induced identical histamine release curves in cord and adult blood. Furthermore, when comparing the results of passive sensitization of basophils from new-born and adult persons, the new-born basophils possessed a significant fraction of free IgE receptors, whereas in adults most of the receptors were occupied by IgE.     

Allergen-specific conventional immunotherapy decreases immunoglobulin E-mediated basophil histamine releasability

BACKGROUND: Allergen-specific immunotherapy has proven to be clinically effective in the treatment of patients with atopic asthma; however, the mechanisms are still unclear. Several noted immunological changes include an increase of the allergen-specific IgG antibody, a reduction in the allergen-specific IgE antibody subsequent to transient increase, an allergen-specific T cell shift in cytokine production from Th2 to Th1, and a decrease in quantity and activity of basophils and mast cells. OBJECTIVE: To analyse the changes of basophil histamine release in response to IgE-mediated and non-IgE-mediated stimuli before and after conventional house-dust mite immunotherapy in children who suffer from atopic asthma. METHODS: Fourteen Dermatophagoides farinae (Df) sensitive asthmatic children with conventional immunotherapy were examined. Basophil histamine releasability was measured 0 months (just before immunotherapy), 4 months and 9 months after immunotherapy. Basophils were stimulated with Df and goat anti-human IgE antibody as IgE-mediated stimuli; and formyl-Met-Leu-Phe (fMLP) and calcium ionophore A23187 as non-IgE-mediated stimuli. Accordingly, the asthma symptom score was used to assess clinical outcome and the skin test reactivity to Df was measured. RESULTS: In contrast to pre-immunotherapy activity, 4 and 9 months after immunotherapy there were significant decreases in histamine release by Df and by anti-IgE antibody. The histamine release by fMLP and by calcium ionophore showed no significant changes after immunotherapy. Histamine release by Df demonstrated significant correlation to that by anti-IgE antibody and by fMLP, yet there was no observable correlation between histamine release by Df and by calcium ionophore. The asthma symptom score decreased significantly 4 and 9 months after immunotherapy and showed significant correlation with histamine release by Df. The skin test reactivity (allergen/histamine ratio) remained constant 4 months after immunotherapy, but decreased significantly 9 months after immunotherapy. CONCLUSION: Basophils have the potential to play an important role in the early clinical improvement of conventional immunotherapy in children with atopic asthma, which may be a result of the decreased IgE-mediated histamine releasability during immunotherapy.     

Enhanced basophil histamine release and neutrophil chemotactic activity predispose grain dust-induced airway obstruction.

BACKGROUND: The pathogenic mechanism of grain dust (GD)-induced occupational asthma (OA) remains unclear. OBJECTIVE: To understand further the mechanism of GD-induced OA. METHODS: Fifteen employees working in a same GD industry, complaining of work-related respiratory symptoms, were enrolled and were divided into two groups according to the GD-bronchoprovocation test (BPT) result: six positive responders were grouped as group III, nine negative responders as group II and five healthy controls as group I. Serum GD-specific immunoglobulin (Ig)E (sIgE), specific IgG (sIgG) and specific IgG4 (sIgG4) antibodies were detected by enzyme-linked immunosorbent assay. Basophil histamine release was measured by the autofluorometric method, and changes of serum neutrophil chemotactic activity were observed by the Boyden chamber method. RESULTS: For clinical parameters such as degree of airway hyperresponsiveness to methacholine, duration of respiratory symptoms, exposure duration, and prevalences of serum sIgE, sIgG and sIgG4 antibodies, there were no significant differences between group II and III (P > 0.05, respectively). Serum neutrophil chemotactic activity increased significantly at 30 min and decreased at 240 min after the GD-BPT in group III subjects (P < 0.05, respectively), while no significant changes were noted in group II subjects (P > 0.05). Basophil histamine release induced by GD was significantly higher in group III than those of group I or group II (P < 0.05, respectively), while minimal release of anti-IgG4 antibodies was noted in all three groups. CONCLUSIONS: These results suggest that enhanced basophil histamine release and serum neutrophil chemotactic activity might contribute to the development of GD-induced occupational asthma.