Glucocorticoids inhibit degranulation of mast cells in allergic asthma via nongenomic mechanism

Background:  Glucocorticoids (GCs) are the most potent anti-inflammatory agents available for allergic diseases including asthma, which are routinely believed to need several hours to take effect through regulating gene expression. Our previous report had shown that GCs could inhibit allergic asthma within 10 min, which the classical mechanism could not explain.
Objective:  To confirm the existence and verify the sites of GCs' rapid action, we investigated nongenomic effects of GCs on degranulation of mast cells in allergic asthma.
Methods:  The GCs' rapid action on airway mast cells deregulations was evaluated in the allergic asthma model of guinea pigs by the computer-assisted morphometry. Using whole-cell patch clamp and fluorometric assay, we examined GCs' nongenomic effect on IgE-mediated exocytosis and histamine release of rat basophilic leukaemia-2H3 mast cells. Employing the flash photolysis technique, we studied the role of Ca²⁺ signal in the GCs' nongenomic effect.
Results:  Inhaled GCs significantly inhibited airway mast cells degranulation in the allergic asthma model of guinea pigs within 10 min. In vitro , GCs could rapidly inhibit IgE-mediated exocytosis and histamine release of mast cells, and neither GC nuclear receptor antagonist nor protein synthesis inhibitor could block the rapid action. We further demonstrated that GCs' nongenomic effect was not through direct action on secretory machinery, but was mediated by a reduction in the [Ca²⁺]_i elevation.
Conclusions:  The study suggested for the first time that nongenomic pathway was involved in GCs' rapid inhibition on allergic asthma, and raised the possibility of new therapeutic strategies for allergic diseases including asthma.     

Histamine- and serotonin-releasing activity of the supernatants from guinea pig spleen cell cultures

The action of supernatants from cultivated in vitro guinea pig spleen cells on the mast cells of guinea pigs, rats and hamsters was studied. It was found that supernatants from guinea pig spleen cell cultures are potent to release histamine from mast cells of the examined populations in a dose-dependent fashion. Histamine release from heterologous mast cells (especially from rat pleural mast cells) was significantly higher than that from homologous mesenteric mast cells. It was also demonstrated (on rat mast cells) that guinea pig spleen cell supernatants possessed not only histamine but 5-hydroxytryptamine releasing activity as well. Rat pleural mast cells were more sensitive and released more serotonin after challenge with spleen cell supernatants than peritoneal cells.     

Anti IgE-induced asthma in guinea pigs; a new model for asthma. Physiological and pathological studies

We established an experimental animal model for IgE-mediated asthma by the use of intravenous injection of rabbit anti guinea pig IgE in normal guinea pigs. The degree of asthmatic airway obstruction was quantitatively estimated by Mead's apparatus which enabled us to use unanesthetized guinea pigs. With the injection of anti IgE, only an early asthmatic response was observed. Inhalation of anti IgE did not trigger asthma. Histo-pathological changes in the lung obtained immediately after the injection of anti IgE resembled those reported in human asthma. This system may be used for pathological and pharmacological studies of the IgE-dependent early asthmatic response.     

Experimental asthma in guinea pigs revisited

Experimental asthma in guinea pigs, as defined in 1937 by Kallós and Pagel, and the pertinent literature are reviewed. The relation between guinea pig asthma and human asthma bronchiale is established. The underlying mechanism in both processes is the misdirected (aberrant) formation of anaphylactic or reaginic (IgE) antibody against innocuous antigens, leading at renewed exposure to the antigen to release of mast cell mediators (such as histamine, SRS-A, eosinotactic peptides, PAF), bronchoconstriction, vasodilation and eosinophilic inflammation at the sites of antigen-antibody reactions in the tissues of the respiratory organs.     

Kidney mast cells, IgE and release of inflammatory mediators capable of altering renal haemodynamics

Collagenase-dispersed human, guinea pig and rat kidney cells, actively or passively sensitized with IgE or IgG subclasses, released histamine, leukotriene C4 and thromboxane B2 on challenge with the specific antigen or antisera to mast-cell-sensitizing immunoglobulins. A similar reaction occurred in the isolated perfused kidney of sensitized guinea pigs, which additionally showed vasoconstriction. Mast cells were identified in cell suspensions and tissue sections. These findings provide support for the possible role of IgE and mast cells in renal pathophysiology.     

Antigen-induced bronchopulmonary alterations in the guinea pig: a new model of passive sensitization mediated by mouse IgE antibodies

A selective model to study the IgE-mediated anaphylactic bronchoconstriction (BC) in the guinea pig is needed, since human asthma involves mainly this class of antibody. However, most procedures presently available for passive homologous or active sensitization lead to responses which are mediated both by IgE and IgG antibodies. In this study, we developed an anaphylactic model in which guinea pigs are passively sensitized with mouse ascitic fluid containing dinitrophenol (DNP)-specific IgE antibodies. Challenge of sensitized animals with DNP coupled to bovine serum albumin evokes a bronchoconstrictor response that is maximal 5 h after sensitization. The resulting anaphylactic BC is not blocked by the H1 histamine antagonist mepyramine, by the peptido-leukotriene antagonist FLP 55712 nor by the platelet-activating factor antagonist BN 52021 alone. However, when the sensitized animals are pretreated with the three drugs in combination, significantly reduced BC was observed upon challenge with the antigen. This latter result indicates that IgE-dependent BC involves the participation of different mediators, a characteristic shared in common with allergic asthma in human.     

Immunoglobulin-G-dependent stimulation of guinea pig lung mast cells and macrophages

Alveolar macrophages and mast cells isolated from guinea pig lung were passively sensitized with IgG1, IgG2, or serum obtained from guinea pigs actively sensitized with ovalbumin. The release of histamine by mast cells and of thromboxane A2 by alveolar macrophages upon ovalbumin challenge indicated that both antibodies and serum were capable of sensitizing these cells with similar effectiveness. Heating the scrum at 56°C for 4 h to inactivate IgE did not modify the antigen-dependent response of lung cells. These results suggest a predominant role for IgG in the allergic response of the guinea pig through the activation of different cell types such as lung mast cells and alveolar macrophages.     

The sensitivity of guinea pig mesenteric and pulmonary mast cells to some histamine releasers

The mesenteric and pulmonary mast cells of guinea pigs were obtained by enzymatic dispersion of the tissues with the enzyme collagenase. The guinea pig mast cells obtained by this method were morphologically intact, as judged by light microscopy. The mast cells from lung and mesentery of actively sensitized guinea pigs released histamine in a dose-dependent fashion after challenge with specific antigen. Both pulmonary and mesenteric mast cells of nonsensitized animals were unresponsive to the action of compound 48/80 and concanavalin A. Both populations of mast cells responded to polymyxin B, but the magnitude of histamine release was low. These cells responded strongly to the ionophore A23187; the mesenteric mast cells were markedly more reactive than pulmonary mast cells.     

IgG2 antibodies block IgE antibody-induced asthma in guinea pigs

In order to examine the blocking activity of IgG2 antibodies to guinea pig for IgE antibodies-induced guinea pig asthma, experiments were carried out as follows. Guinea pigs were passively sensitized intravenously with guinea pig serum containing IgE antibodies to ovalbumin (OA). 8 days after sensitization, IgG2 purified from guinea pigs hyperimmunized with OA was intravenously injected. One hour later, the guinea pigs were challenged by inhalation of OA solution. Asthma attacks were not observed in the guinea pigs, whereas the attacks were observed in guinea pigs passively sensitized with the IgE antibodies but injected IgG2 fraction from normal guinea pigs 1 h before inhalation. These observations suggested that IgG antibodies that increased after immunotherapy might block asthma caused by inhalation of allergens in humans.     

Aspergillus oryzae lectin induces anaphylactoid oedema and mast cell activation through its interaction with fucose of mast cell-bound non-specific IgE

We investigated whether Aspergillus oryzae lectin (AOL), a fucose-specific lectin, induces anaphylactoid reactions and mast cell activation. The injection of AOL into footpads of mice produced a dose-related acute paw oedema. The AOL-induced oedema was attenuated by predose of histamine H1 receptor blocker or pretreatment of the lectin with fucose before injection and was not observed in SCID and mast cell-deficient WBB6F1-W/Wv mice. These results suggested that the AOL-induced anaphylactoid reaction was mediated by histamine released from mast cells. In addition, the activation of mast cells was seemed to be induced by the crosslinking of IgE on the cell surface following the binding of AOL to fucose residues in IgE. Consistent with the in vivo results, AOL induced the degranulation of the rat mast cell line RBL2H3 sensitized with monoclonal IgE. As AOL induced the increase in intracellular Ca(2+) concentration of IgE-sensitized RBL2H3 cells as well as antigen stimulation, AOL could input signals from FcεRI. The degranulation of IgE-sensitized RBL2H3 cells by AOL was diminished by pretreatment of AOL with fucose. Defucosylated IgE did not induce degranulation of RBL2H3 cells in response to AOL stimulation, in spite of its ability to induce degranulation by antigen stimulation as intact IgE. These results indicated that AOL bound to fucose residue of IgE causing antigen-independent IgE-mediated mast cell activation and anaphylactoid reactions in vitro and in vivo, respectively. AOL bound to human IgE as well as to mouse IgE, suggesting the possible implication of AOL in the allergic response to Aspergillus oryzae in humans.     

Histamine-releasing properties of guinea pig cardiac mast cells

The immunization of guinea pigs with OA + Al(OH)3 induced substantial IgG1a and IgG1b antibody response and low, transient IgE response, as examined by PCA test. Cardiac mast cells obtained by enzymatic dispersion method from sensitized animals released histamine in vitro after the challenge with specific antigen (histamine release up to 21%). Cardiac mast cells obtained from nonsensitized guinea pigs were sensitive to the action of ionophore A23187 and polymyxin B only when the agents were used in high concentrations (histamine release up to 25.1% and 21. respectively) and were only slightly responsive to the challenge with Concanavalin A and compound 48/80.     

蛔虫变应原诱发豚鼠哮喘后血液流变学及组织胺动态变化

目的 :为了观察蛔虫变应原致喘豚鼠动物模型后血液流变学变化及外周血组织胺的动态变化 ,丰富变应原激发动物哮喘的基础理论。方法 :把实验豚鼠随机分为阴性对照组、佐剂对照组和阳性激发组。用蛔虫变应原激发致喘豚鼠后 ,观察各实验组在激发哮喘后 1h、 2 4h和 72h的血液流变学和组织胺的动态变化。结果 :由蛔虫变应原引起的过敏性哮喘豚鼠动物模型 ,其血液流变性出现异常 ,阳性激发组的血浆粘度、红细胞聚集指数、全血高切变率还原粘度和组织胺水平均较阴性对照组、佐剂对照组显著性升高 (P <0 .0 1) ;而红细胞压积则无明显变化。结论 :表明用蛔虫变应原致喘豚鼠后 ,能诱发速发型变态反应 ,导致肥大细胞和嗜碱性粒细胞脱颗粒并释放出组织胺。与此同时血液出现高粘滞状态 ,进一步加剧气道局部的缺血、缺氧及炎性反应 ,导致气体交换障碍、肺功能降低。     

Is mast cell histamine involved in the regulation of acid secretion in the guinea pig stomach?

In order to find out whether mast cell histamine is involved in acid secretion in the guinea pig stomach, we tried to answer the following questions. Can the calcium ionophore (CaI), A23187, release enough mast cell histamine to evoke acid secretion? Does electrical stimulation of the vagus nerve (VS) provoke any changes in mast cells in the stomach? In the experiments, an isolated part of guinea pig stomach was used. Preparations from CaI and VS treated groups were examined histologically at the end of experiments. CaI evoked a clear secretory response that was blocked by famotidine. The H2 antagonist also blocked the secretory effect of VS. This result shown that these secretory effects were at least partly mediated by the released histamine. Connective tissue mast cells in CaI treated preparations and in preparations with VS showed significant degranulation. These results support the hypothesis that mast cell histamine could be involved in gastric acid secretion in guinea pig stomach.     

Quercetin inhibits anaphylactic contraction of guinea pig ileum smooth muscle

Certain flavonoids inhibit antigen-induced release of histamine from mast cells and basophils and also inhibit contraction of guinea pig ileum induced by histamine, acetylcholine, and PGE2. We examined the effect of one flavonoid, quercetin, on anaphylactic smooth muscle contraction of ileum from guinea pigs sensitized to egg albumin. Quercetin inhibited both the phasic and tonic components of anaphylactic contraction in a concentration-dependent fashion (IC50 approximately 10 microM). Whether this is primarily an effect on mast cell mediator release or inhibition of mediator effects on smooth muscle has not been established.     

Histamine-releasing activity of lymphocyte supernatants of guinea pig spleen cell cultures

We demonstrated the production of a histamine releasing factor (HRF) by 24-h cultures of guinea pig spleen cells which were stimulated or not with specific antigen (ovalbumin, OA) or mitogen (phytohemagglutinins or concanavalin A). HRF induced the release of histamine from homologous mesenteric mast cells in a dose-dependent fashion. The HRF-induced histamine release was not high compared to the release induced by calcium ionophore A23187, but higher than that induced by compound 48/80, polymyxin B and con canavalin A. The mast cells from sensitized guinea pigs released histamine when challenged with OA. We found that HRF-induced histamine release was additive to that induced by antigen, when both agents were added simultaneously to sensitized mast cells. The phenomenon was most significant when a suboptimal dose of antigen was used. Moreover, we did not observe any differences in the magnitude of HRF-induced histamine release between the mast cells from nonsensitized and sensitized guinea pigs. The time course of histamine release induced by HRF was significantly slower than that with specific antigen (10 min and 45 sec, respectively). Our results may suggest that HRF acts on mast cells through a different not immunological mechanism.     

Antiallergic activity of nylidrin hydrochloride (RHC 3432-A). I. Effect on release of histamine in vitro from rat peritoneal mast cells, guinea pig lung slices and human basophils

Nylidrin (RHC 3432-A) has been investigated for its antiallergic activity in three in vitro models. Nylidrin was an effective inhibitor of IgE-mediated release of histamine from passively sensitized rat peritoneal mast cells and human basophils, and of IgG1-mediated release of histamine from passively sensitized guinea pig lung slices. The inhibition of the release of histamine by nylidrin in all three models was not antagonized by propranolol, indicating that nylidrin does not inhibit histamine release via stimulation of beta-adrenergic receptors. Isoproterenol and epinephrine were effective as inhibitors of the release of histamine only from guinea pig lung while salbutamol and terbutaline had no effect on immunologic release of histamine in all three models. Detailed comparative studies with disodium cromoglycate (DSCG) indicated that the mechanism of action of nylidrin in the rat mast cell model is different from that of DSCG.     

Change in sensitivity to lysophosphatidylserine of mouse bone marrow-derived mast cells during cultivation with fibroblasts.

Lysophosphatidylserine (lysoPS) is known to enhance IgE-mediated activation of rodent connective tissue mast cells (CTMCs). In the present study, we investigated the effect of lysoPS on degranulation of interleukin-3-dependent mouse bone marrow-derived mucosal mast cells (BMMCs) and of their CTMC-like differentiated cells. In the absence of lysoPS, BMMCs released approximately 20% of their histamine when sensitized with anti-dinitrophenyl (DNP) IgE and challenged with DNP-conjugated antigen. When stimulated in the presence of lysoPS, no appreciable enhancement was observed. On the other hand, histamine release from BMMCs, which had differentiated to CTMC-like cells by co-culture with 3T3 fibroblasts, was enhanced 2- to 3-fold by the addition of lysoPS. The maximum potentiation was observed at 5 x 10(-6) M lysoPS. These results suggest that mast cells might acquire their dependence on exogenous lysoPS during differentiation from mucosal mast cells to CTMC-like cells.     

Description of a hybridoma bank towards Bordetella pertussis toxin and surface antigens

This paper describes the development of a murine bank of monoclonal antibodies against Bordetella pertussis toxin, filamentous hemagglutinin (FHA), pili, lipopolysaccharide (LPS), or outer membrane proteins (OMPs). Subunits S1, S2, S3 of pertussis toxin (PT) bound immunoglobulins and glycoproteins such as fetuin and haptoglobin in an unspecific manner. The specificity of monoclonal antibodies towards subunits S1, S2, S3 or S4 of PT could be demonstrated by using purified immunoglobulins or their Fab2 fragments. A set of FHA-specific monoclonal antibodies could be differentiated on the basis of their binding to the various breakdown products present in FHA preparations. Pili-specific monoclonal antibodies reacted with either native pili or denatured pilin, and both demonstrated serotype specificity. Monoclonal antibodies to Bordetella pertussis OMPs were directed to either the virulent phase-regulated trypsin-sensitive, detergent-extractable OMPs 92 kDa, 32 kDa, and 30 kDa or the non-virulent phase-expressed, not-trypsin sensitive OMPs 38 kDa, 33kDa, and 18 kDa.     

Histological study of mast cells in the actively sensitized guinea pig lung and after challenge: effect of a corticosteroid

Although mast cells are sometimes considered to play a minor role in hypersensitivity reactions, we used a histological method to show the modifications in the guinea pig lung mast cell population in the course of such reactions. We sensitized guinea pigs with ovalbumin and studied the effect of the challenge with and without corticosteroid treatment. We observed that the mast cell count was not modified after sensitization but was decreased after challenge. Twenty-four hours after challenge, the number of mast cells returned to the control value, indicating a renewal of the mast cell pool. A second challenge, 1 week after the first, did not provoke the same mast cell degranulation, suggesting a non-responsiveness to aerosol antigen. Betamethasone dipropionate treatment protected mast cells against challenge: in treated guinea pigs, mast cell degranulation was prevented, and we did not observe any change in mast cell count after challenge. The present study was useful to show an effect of corticosteroids on mast cell degranulation in immediate hypersensitivity reactions in vivo.