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PubMed1978

In vitro histamine and serotonin release by radiographic contrast media (RCM). Complement-dependent and -independent release reaction and changes in ultrastructure of human blood cells.

Peripheral leucocytes and platelets from twenty healthy volunteers were incubated in vitro with radiographic contrast media (diatrizoate-Hypaque, iothalamate-Conray and iodipamide-Cholografin) under varying conditions. All radiographic contrast media (RCM) were able to induce histamine release from peripheral leucocytes and the release reaction was dose-dependent. There were individual differences in the sensitivity of leucocytes to different RCM. The highest values (up to 80% histamine release) were found with high concentrations (0.07-0.3 M) of diatrizoate. The addition of normal human serum (NHS) to the reaction mixture led to a further increase in histamine release (P is less than 0.01), probably due to complement activation. The mechanism seems to be mediated by proteins of the alternative pathway, because serum depleted of complement components (factor B, factor D, properdin) did not show this synergistic effect. IgG-depleted serum, however, was able to show the augmented release reaction. Washed platelets incubated with RCM released serotonin in a dose- and time-dependent reaction. The most powerful serotonin-releasing RCM was found to be iodipamide, which produced a release reaction in all people investigated at concentrations of 0.04-0.09 M, while diatrizoate and iothalamate were effective only in half of the tested individuals at high concentrations (0.3 and 0.2 M respectively). The addition of plasma proteins to the reaction mixture inhibited the serotonin release quantitatively. There was no difference in inhibitory potency between autologous and heterologous plasma or serum; sera depleted of various complement components showed similar effects as NHS. The serotonin release was not due to platelet lysis, as determined by the concentration of lactic dehydrogenase present in the supernatant during serotonin release. Incubation of the leucocytes with RCM produced ultrastructural changes, including degranulation of basophils, aggregation of platelets and infiltration of the aggregates by polymorphonuclear leucocytes. The most prominent changes were observed when complement was present in the reaction mixture.

Basophils↗

PubMed1978

Cutaneous basophil anaphylaxis. Immediate vasopermeability increases and anaphylactic degranulation of basophils at delayed hypersensitivity reactions challenged with additional antigen.

Many delayed-type reactions contain large infiltrates of basophils whose function is unknown. We have studied these cutaneous basophil hypersensitivity (CBH) reactions in guinea-pigs to ascertain whether basophils that are recruited to delayed reaction sites could be triggered for immediate reactivity. We compared 24 h CBH reactions with nearby skin for immediate hypersensitivity by challenging each site with small amounts of antigen. CBH sites had augmented immediate increases in vascular permeability detected by extravasation of Evan's blue dye. The ability to elicit this augmented anaphylactic phenomenon correlated with the local presence of basophils, and light microscopy at CBH reactions 15 min after antigen challenge showed a 50% decline in basophil counts. Electron microscopy showed that progressive anaphylactic-type degranulation of local basophils occurred within minutes following reintroduction of antigen. There was fusion of vacuoles containing granules, exocytosis of granules, and dissolution of granules, without ultrastructural disruption of cellular integrity. These results establish that basophils in CBH reactions can be triggered with soluble antigen to undergo anaphylactic degranulation, with the immediate release of vasoactive mediators. We have termed this phenomenon 'cutaneous basophil anaphylaxis'. Thus, one function of basophils at sites of delayed hypersensitivity may be to provide the potential for augmented, local, immediate anaphylactic reactivity.

Anaphylaxis↗

PubMed1990

Detection of histamine release inhibitory factor- and histamine releasing factor-like activities in bronchoalveolar lavage fluids.

Histamine releasing factors (HRF) are a group of cytokines that cause degranulation of basophils and mast cells. Recently we have described a histamine release inhibitory factor (HRIF) that inhibits HRF-induced histamine release from basophils and mast cells. The objective of this study was to investigate the presence of these cytokines in bronchoalveolar lavage (BAL) fluid from normal subjects. We found that BAL fluids from 12 to 17 volunteers contained a dialyzable (molecular weight cutoff 3500) factor that inhibited basophil histamine release by HRF, anti-IgE, concanavalin A, and N-formyl-methionyl-leucyl-phenylalanine (FMLP). In addition, BAL fluids from 83% of the tested donors contained a nondialyzable inhibitor that blocked HRF-induced histamine release from basophils. The molecular weight of this inhibitor was estimated to be 20 to 30 and 8 to 10 kD by Sephadex G-50 chromatography and TSK 2000 size-exclusion HPLC. None of the unconcentrated BAL fluids showed any HRF activity on initial screening using basophils from allergic subjects. However, when the BAL fluids were concentrated, all BAL samples that were tested (N = 10) demonstrated significant HRF activity. The molecular weight of BAL HRF has been estimated to be in the range of 15 to 25 kD by size-exclusion HPLC, similar to the HRF synthesized by mononuclear cells. Thus we have demonstrated the presence of both HRF and HRIF in the BAL fluids. We speculate that these cytokines may be involved in the local regulation of basophil and mast cell activation.

Adult↗

PubMed1997

A cell-based immunobiosensor with engineered molecular recognition--Part I: Design feasibility.

A novel bioelectronic sensor is described in which living immune cells are transformed into unique biotransducer couples by engineering their molecular recognition for preselected antigens of clinical interest. This 'hybrid' biosensor, constructed with mast cells interfaced to a microfabricated thermoelectric device with the use of biomolecular linkages, is capable of detecting antigens in real time by transducing minute heat changes arising from antigen-induced mast cell activation processes. The thermoelectric approach was selected based upon preliminary bioenergetic calculations which indicated that metabolic changes arising from mast cell antigen recognition result in a significant increase in exothermic heat relative to basal metabolic conditions. Experimental studies confirmed that mast cell activation and degranulation can be discriminated theramally from basal metabolic activity. Results obtained from microcalorimetry experiments using cultured mast cells (MC/9) mucosal-like mast cell line), and harvested mast cells (rat peritoneal mast cells) indicated that detectable increases in heat output (-3 +/- 0.5 pW/cell, mean peak output) immediately followed cell activation. The construction of a miniature hybrid immunobiosensor device was made possible by bioelectronic coupling achieved with the use of cellular adhesive proteins that immobilized non-adherent (MC/9) cells as well as adherent (RBL-2H3 rat basophilic leukemia) cells to the thermopile. Results from preliminary tests conducted on a hybrid biosensor prototype validated the design feasibility of a miniature, living cell immunodiagnostic biosensor. Such cell-based hybrid biosensor approaches may greatly extend the capability for selective, rapid, on-site, antigen detection for a wide range of clinically relevant antigens and offer new approaches to in vitro diagnostics.

Animals↗

PubMed1999

Immunological diagnosis of human cystic echinococcosis: utility of discriminant analysis applied to the enzyme-linked immunoelectrotransfer blot.

An enzyme-linked immunoelectrotransfer blot for the diagnosis of human hydatid disease was performed, and the different antibody responses were analyzed by a discriminant analysis. This multivariate technique gave us, first, a selection of the most important responses against Echinococcus granulosus infection and, second, a procedure for the classification of patients into two groups: patients with hydatid disease and patients without a history of hydatid disease. This method was applied to 67 patients, 25 with active hydatid cysts (24 hepatic and 1 pulmonary) and 42 without a history of hydatid disease and was compared with the results obtained by conventional serology: indirect hemagglutination, latex particle agglutination, and basophil degranulation. An immunoelectrotransfer blot coupled to a discriminant analysis was more sensitive than conventional serological diagnosis and detected 100% of patients with an active hepatic hydatid cyst with a specificity of 100%. This method, however, failed to detect an uncomplicated hyaline pulmonary hydatid cyst.

Discriminant Analysis↗

PubMed1975

[Intrauterine sensitization to benzylpenicillin in an experiment on rabbits].

Antibodies of benzylpenicillin in the posterity of various ages of female rabbits were found with reaction of passive hemagglutination (RPHA) and degranulation of the basophiles after their immunization with the antibiotic. This provided an assumption of intrauterine sensitization. Treatment of the sera of the sensitized animals with 2-mercaptoethanol partially decreased their activity in the RPHA which indicated to the presence of the antibodies of classes IgG and IgM.

Age Factors↗

PubMedDate not supplied

Nonspecific changes in immunotherapy with house dust extract.

Despite the possible complications and inadequacies of the immunotherapy of allergic patients with house dust extracts (HDE), this is widely applied in Latin America and is commonly considered to be a cost-effective treatment for allergic diseases. Doubts about its efficacy and mechanism of action prompted us to evaluate the immunological changes that occurred in a group of 99 patients selected for their confirmed allergic sensitivity to house dust, and who showed significant clinical improvement after a minimum of 12 months of immunotherapy with HDE. The favorable clinical response of these patients was associated with some of the mechanisms reported for immunotherapy with pollen or venom allergens, such as increased levels of IgG or IgG4 "blocking" antibodies, and the increased affinity of these antibodies for house dust antigens, or diminished basophil sensitivity to HDE. However, the magnitude of these changes was relatively small, and their biological significance questionable. In contrast, particularly evident was a reduction in the cutaneous reactivity not only to the specific allergen, but also to the nonspecific mast cell degranulator codeine and to histamine. These results suggest that as yet unidentified "nonspecific" mechanisms might contribute to the effect of immunotherapy with extracts as heterogeneous and complex as HDE.

Adolescent↗

PubMed1997

Involvement of the ras-like GTPase rab3d in RBL-2H3 mast cell exocytosis following stimulation via high affinity IgE receptors (Fc epsilonRI).

Aggregation of high affinity IgE receptors (Fc epsilonRI) expressed on mast cells and basophils is a potent stimulus for the release of inflammatory mediators from cytoplasmic granules. Fc epsilonRI-dependent exocytosis requires activation of protein kinase C and mobilization of calcium from intra- and extracellular stores. However, how these events ultimately regulate the membrane fusion step between cytoplasmic granules and the plasma membrane still remains unclear. In this study, we investigated the role of the small GTPases of the rab3 subfamily in the regulated exocytosis following stimulation of rat basophilic leukemia cells (RBL-2H3). Analysis using reverse-transcriptase-based PCR showed that RBL-2H3 cells expressed rab3a and rab3d isoforms, with a predominance of rab3d at the mRNA level. Investigation of the subcellular distribution using isoform-specific Abs demonstrated that the majority of rab3a was expressed in the cytosol, whereas rab3d was found predominantly in the membrane fraction. To determine whether these proteins play a role in Fc epsilonRI-triggered exocytosis, we established RBL-2H3 transfectants that overexpressed wild-type and expressed GTP-binding mutant forms (N135I) of rab3a and rab3d. Whereas expression of rab3a proteins did not significantly affect degranulation as tested by beta-hexosaminidase release, those of both wild-type and mutant rab3d proteins inhibited degranulation. Calculations of the initial fast and of the second slow release rates showed that they are both inhibited about twofold, suggesting that rab3d interferes with a rate-limiting step in Fc epsilonRI-stimulated exocytosis.

Amino Acid Sequence↗

PubMed1983

Effect of verapamil on allergen-induced asthma in patients with respiratory allergy.

Specific degranulation of mast cells and basophils depends upon transmembrane calcium flow. In the present studies the protective effect of the calcium-antagonist drug verapamil against allergen-induced asthma was assessed in eight atopic patients. The results of this study showed that verapamil, administered both orally or by aerosol, can prevent the specific bronchoconstriction.

Administration, Oral↗

PubMed1989

[Studies on the degranulation and histamine release of purified rat peritoneal mast cells. The inhibitory effect of histamine and other chemicals].

We know that the mast cell or basophil degranulation and the release of chemical mediators such as histamine may play an important role in inducing immediate type allergic reactions. In this experiment, employing purified rat peritoneal mast cells (RPMC) the degranulation pattern of RPMC and percent release of histamine from RPMC by pharmacologic (compound 48/80, kallikrein or peptidoglycan) or allergic (IgE-anti IgE complex) stimuli were examined. The inhibitory effects of catecholamine (isoproterenol), an adenylate cyclase stimulator, methylxanthine (IBMX: 3-isobutyl-1-methyl-xanthine), a phosphodiesterase inhibitor and chemical mediators (histamine, serotonin, bradykinin) on compound 48/80-induced RPMC degranulation and also those of catecholamine, methylxanthine and chemical mediators on IgE-anti IgE complex-induced histamine release from RPMC were tested. Compound 48/80-induced RPMC degranulation was remarkably inhibited not only by treatment with isoproterenol and IBMX, but also by treatment with histamine. It was not, however, inhibited by serotonin or bradykinin. The inhibitory effect of histamine on compound 48/80-induced RPMC degranulation was blocked by pretreatment with H2-antagonist (cimetidine), but not by H1-antagonist (diphenhydramine). The cyclic adenosine 3',5'-monophosphate (cAMP) content of RPMC was significantly increased by pretreatment with histamine as well as isoproterenol and IBMX. These facts suggest that cAMP acts as a regulator or modulator of the RPMC degranulation and the histamine release from mast cells and that the inhibitory effect induced by histamine may be related to the H2-receptor existing on the surface of mast cells.

Animals↗

PubMed2001

Basophils and exercise-induced hypoxemia in extreme athletes.

This study examined whether the increase in histamine release (%H, i.e., plasma histamine expressed as a percentage of whole blood histamine) associated with exercise-induced hypoxemia (EIH) is related to high training-induced changes in basophil and osmolarity factors in arterial blood. All parameters were measured in 20 endurance athletes, 11 of whom presented an EIH (HT(hyp)) and 9 of whom were nonhypoxemic (HT(nor)), and in 10 untrained control subjects (UT). Measurements were made at rest, at the maximal workload of an incremental exhaustive exercise test, and at the fifth minute of recovery. %H increased during exercise in HT(hyp) (P < 0.01) but did not increase significantly in HT(nor) and UT controls. The results indicated that 1) osmolarity and Na(+) and K(+) concentrations did not differ between the two trained groups and 2) the basophil count and basophil histamine content did not differ among groups. We concluded that the %H increase associated with EIH was not due to a training effect on these parameters. The relatively low increase in histamine content during exercise in HT(hyp) in comparison to HT(nor) (P < 0.05) and UT (P < 0.01) and the low recovery vs. resting basophil count only in HT(hyp) (P < 0.01) suggested an accentuated exercise-induced basophil degranulation in the hypoxemic athletes.

Adult↗

PubMedDate not supplied

[Parameters of polynuclear cell activity in torpid and latent recurrent rheumocarditis in the process of treatment with a high-protein diet and drug preparations].

The in vitro test of direct granulation of basophils affected by specific antigens (cardial tissue, acid-soluble fraction of collagen, streptococcal allergens) was investigated in the course of treating 41 patients (29 women and 12 men aged from 20 to 50 years) suffering from a little active rheumatic fever. The patients received a complex of therapeutic measures including high-protein diet (130-140 g protein) and drugs (1.5-2.0 g acetylsalicylic acid and 15 mg of prednisolone a day). As a result of the treatment all the patients improved and demonstrated pain relief in the heart region, reduced palpitation, abatement of general weakness and fatigue. At the same time there was a decrease in the reactivity of basophilic leukocytes, shown by less number of degranulated cells in vitro, activated with specific antigens.

Adult↗

PubMed2000

The IgE-binding epitopes of rPar j 2, a major allergen of Parietaria judaica pollen, are heterogeneously recognized among allergic subjects.

Pollen allergens are multivalent proteins that cross-link IgE antibodies on mast or basophil cells, inducing secretion of biologic mediators, and resulting in various allergic symptoms. The IgE-binding regions of the Parietaria judaica (Pj) pollen major allergen rPar j 2 were investigated. Twenty-nine single sera from Pj-allergic subjects were tested by Western blot against five recombinant peptides. At least four putative IgE-binding epitopes were identified. The analysis of their diffusion suggested a heterogeneous IgE-binding response. In fact, 75% of the sera reacted with peptide 1-54, 48% with peptide 48-101, 24% with peptide 1-30, 7% with peptide 29-54, and none with peptide 48-76. These five peptides were analyzed with the histamine-release assay. Only peptide 48-101 was capable of inducing degranulation and release of histamine. These results suggest that the recombinant rPar j 2 allergen contains IgE epitopes that are heterogeneously recognized by sensitive patients, and that therefore the therapeutic approach based on the use of haptenic peptides needs a careful evaluation.

Allergens↗

PubMed2004

Hypersensitivity reactions to radiocontrast media: the role of complement activation.

Although intravenous use of radiocontrast media (RCM) for a variety of radiographic procedures is generally safe, clinically significant acute hypersensitivity reactions still occur in a significant percentage of patients. The mechanism of these anaphylactoid, or "pseudoallergic," reactions is complex, involving complement activation, direct degranulation of mast cells and basophils, and modulation of enzymes and proteolytic cascades in plasma. In this review, basic information on different RCMs and their reactogenicity is summarized and updated, and the prevalence, pathomechanism, prediction, prevention, treatment, and economic impact of hypersensitivity reactions are discussed. Particular attention is paid to the in vitro and in vivo evidence supporting complement activation as an underlying cause of RCM reactions.

Complement Activation↗

PubMed2004

Identification of selective basophil chemoattractants in human nasal polyps as insulin-like growth factor-1 and insulin-like growth factor-2.

In a search for novel leukocyte chemoattractants at sites of allergic inflammation, we found basophil-selective chemoattractant activity in extracts of human nasal polyps. The extracts were fractionated by reverse phase HPLC, and the resulting fractions were tested for leukocyte-stimulating activity using sensitive shape change assays. The basophil-selective activity detected was not depleted by a poxvirus CC-chemokine-binding protein affinity column. This activity was further purified by HPLC, and proteins in the bioactive fractions were analyzed by tandem electrospray mass spectrometry. Insulin-like growth factor-2 (IGF-2) was identified in these HPLC fractions, and the basophil-stimulating activity was inhibited by an anti-IGF-2-neutralizing Ab. Recombinant IGF-2 induced a substantial shape change response in basophils, but not eosinophils, neutrophils, or monocytes. IGF-2 stimulated chemokinesis of basophils, but not eosinophils or neutrophils, and synergized with eotaxin-1/CCL11 in basophil chemotaxis. IGF-2 also caused up-regulation of basophil CD11b expression and inhibited apoptosis, but did not stimulate degranulation or Ca(2+) flux. Recombinant IGF-1 exhibited similar basophil-selective effects as IGF-2, and both growth factors were detected in nasal polyp extracts by ELISA. This is the first demonstration of chemokinetic factors that increase the motility of basophils, but do not act on other granulocytes or monocytes. IGF-1 and IGF-2 could play a role in the selective recruitment of basophils in vivo.

Amino Acid Sequence↗

PubMed1989

Penicillin G-induced cutaneous anaphylaxis in the guinea pig.

Allergic cutaneous responses were induced by intradermal injection of penicillin G (PCG) and PCG-bovine serum albumin (BSA) conjugates onto the back of guinea pig actively immunized with PCG potassium (25 mg/animal) incorporated in Freund's complete adjuvant. The PCG-induced response was characterized macroscopically by erythema and edema with a maximum level at 24 hrs after elicitation and microscopically by the infiltration with basophils, macrophages and lymphocytes following with neutrophils. In addition, intensity of macrophage-infiltration shared a similar time course change with those of erythema and edema. These suggest that this response is associated to a delayed type hypersensitivity of Jones-Mote type. On the other hand, in the PCG-BSA-induced response the edema with erythema at the early phase was a noticeable observation and this response disappeared within 12 hrs, although the erythema continued by 24 hrs. Microscopically, the degranulation of mast cells and severe infiltration with neutrophils in the early phase and the infiltration of eosinophils in the late phase accompanying the infiltration with monocytes, basophils and lymphocytes were characteristic findings, which suggest that PCG-BSA response is a similar hypersensitivity to an atopic dermatitis. As mentioned above, we confirmed two types of allergic cutaneous responses in the guinea pig immunized with PCG.

Animals↗

PubMed1987

Effects of clonidine on the dermal inflammatory cell response of experimental toxic and allergic contact reactions and intradermal hypersensitivity.

In previous studies, the alpha 2-adrenoceptor agonist clonidine has been shown to suppress the wheal and flare reaction in guinea pigs sensitized to ovalbumin. This phenomenon has been further studied with special reference to effects on the dermal inflammatory cell infiltrate and mast cells. Clonidine lessens the degranulation of mast cells seen in control untreated immediate hypersensitivity reactions. Less neutrophils and eosinophils arrive to the treated reactions. Basophils and mononuclear cells (chiefly lymphocytes) which characterize the late phase of the wheal and flare reaction were not influenced by clonidine. Clonidine had a possible minimal effect on allergic contact (delayed hypersensitivity) reactions. The toxic contact reaction to croton oil (nonspecific cutaneous inflammation) was not affected.

Animals↗

PubMed2005

[Ala12]MCD peptide: a lead peptide to inhibitors of immunoglobulin E binding to mast cell receptors.

An effort was made to discover mast cell degranulating (MCD) peptide analogs that bind with high affinity to mast cell receptors without triggering secretion of histamine or other mediators of the allergic reaction initiated by immunoglobulin E (IgE) after mast cell activation. Such compounds could serve as inhibitors of IgE binding to mast cell receptors. An alanine scan of MCD peptide reported previously showed that the analog [Ala12]MCD was 120-fold less potent in histamine-releasing activity and fivefold more potent in binding affinity to mast cell receptors than the parent MCD peptide. Because this analog showed marginal intrinsic activity and good binding affinity it was subsequently tested in the present study as an IgE inhibitor. In contrast to MCD peptide, [Ala12]MCD showed a 50% inhibition of IgE binding to the Fc epsilon RI alpha mast cell receptor by using rat basophilic leukemia (RBL-2H3) mast cells and fluorescence polarization. Furthermore, in a beta-hexosaminidase secretory assay, the peptide also showed a 50% inhibition of the secretion of this enzyme caused by IgE. An attempt was made to relate structural changes and biologic differences between the [Ala12]MCD analog and the parent MCD peptide. The present results show that [Ala12]MCD may provide a base for designing agents to prevent IgE/Fc epsilon RI alpha interactions and, consequently, allergic conditions.

Alanine↗