RESUMO
Immunoglobulin E (IgE) has been shown to play a critical role in the allergic late-phase reaction, which is marked by intense leukocyte infiltration and edema. In this study we assessed the allergic pleural inflammation triggered by intrapleural (i.pl.) challenge in sensitized rats. We examined pleural effluent from actively sensitized rats following anti-IgE monoclonal antibody (mAb) (MARE-1) provocation for protein exudation, neutrophil as well as eosinophil accumulation. Inflammatory changes triggered by antigen after passive sensitization with IgE mAb was also assessed for comparison. Total serum level of IgE was found to be about threefold increased 7-8 days post-active sensitization, remaining augmented for at least 30 days. Increased levels of peritoneal leukocyte-bound IgE and serum IgE with specificity to ovalbumin were also detected. Nevertheless, the anti-IgE challenge in 14-day actively sensitized was shown to be a weak stimulus of neutrophil and eosinophil accumulation, despite being able to cause intense protein extravasation. Similarly, antigen challenge of IgE-passively sensitized rats caused protein leakage that was comparable to that induced by anti-IgE mAb in actively sensitized rats but led to a much lower neutrophil/eosinophil infiltration. Also, blockade of complement with recombinant human soluble C receptor-1 (sCR1) treatment prevented actively sensitized rats from reacting to antigen with neutrophil and eosinophil recruitment without modifying protein extravasation. These data suggest that IgE and complement-mediated mechanisms probably account for the exudation and leukocyte infiltration that is characteristic of the pleural inflammatory response observed in actively sensitized rats.
Assuntos
Imunoglobulina E/imunologia , Pleurisia/imunologia , Proteínas/imunologia , Cloreto de Alumínio , Compostos de Alumínio , Animais , Anticorpos Monoclonais/imunologia , Cloretos , Eosinófilos/efeitos dos fármacos , Feminino , Imunoglobulina E/sangue , Contagem de Leucócitos/efeitos dos fármacos , Masculino , Neutrófilos/efeitos dos fármacos , Ovalbumina/imunologia , Proteínas/metabolismo , Ratos , Ratos Wistar , Receptores de Complemento/imunologiaRESUMO
The identity of the histamine-potentiating activity detected in the rat anaphylactic pleural washing was investigated. Wistar rats of both sexes, weighing 150-200 g, were sensitized by injecting subcutaneously (sc) a mixture of ovalbumin and Al(OH)3 14 days before allergen challenge. In sensitized rats, intrapleural (ipl) injection of ovalbumin (12 micrograms/cavity) caused an intense protein exudation. A single ipl administration of compound 48/80 (12 micrograms/cavity) exhausted the resident mast cell population and turned the pleural cavity hyporeactive to the allergen challenge performed 5 days later. Allergen-induced exudation occurred in parallel to a dramatic decrease in the amount of cell-stored histamine (from 9.6 +/- 1.4 (N = 8) to 1.3 +/- 0.1 (N = 6) micrograms/cavity, P < 0.001) in the pleural fluid within 10 min. The anaphylactic cell-free pleural washing obtained at this time, as well as histamine at a concentration equivalent to that stored in pleural mast cells (10 micrograms/cavity), did not induce pleural exudation when injected into normal rats. In contrast, the combined administration of histamine and anaphylactic pleural washing led to remarkable pleural exudation, comparable to that obtained with a high dose of histamine (200 micrograms/cavity) alone. It is noteworthy that the anaphylactic washing from compound 48/80-pretreated rats failed to synergize with histamine. Also, synergism was not reproduced when recipient rats were pretreated with methysergide (50 micrograms/cavity). Consistently, serotonin (5 micrograms/cavity) acted synergistically with histamine (10 micrograms/cavity), producing a greater exudative response than observed with the sum of the effects of each vasoactive amine alone. The results indicate that serotonin accounts for the histamine-potentiating activity noted in the anaphylactic pleural washing, confirming that the synergistic interaction between these vasoactive amines plays a critical role in the rat allergic pleurisy.
Assuntos
Histamina/imunologia , Derrame Pleural/imunologia , Serotonina/imunologia , Anafilaxia/imunologia , Animais , Feminino , Masculino , Pleura/imunologia , Derrame Pleural/patologia , Ratos , Ratos WistarAssuntos
Imunoglobulina E/biossíntese , Idiótipos de Imunoglobulinas/imunologia , Alanina/imunologia , Animais , Especificidade de Anticorpos , Biopolímeros , Reações Cruzadas , Grânulos Citoplasmáticos , Dinitrobenzenos/imunologia , Epitopos , Feminino , Glutamatos/imunologia , Imunoglobulina G/imunologia , Masculino , Mastócitos/imunologia , Camundongos , Camundongos Endogâmicos , Tirosina/imunologiaAssuntos
Anticorpos , Histamina/análise , Mastócitos/análise , Anafilaxia Cutânea Passiva , Animais , Antígenos , Feminino , Temperatura Alta , Imunoglobulina E , Técnicas In Vitro , Masculino , CamundongosRESUMO
Kinetics of primary and booster-specific and total IgE responses to distinct antigenic stimuli were studied in two inbred rat strains, Brown-Norway (BN) and Lewis, and one outbred, Sprague-Dawley (SD). The rats were immunized three or four times at intervals varying between 15 and 22 days by subcutaneous injections of 10 microgram ovalbumin, keyhole limpet haemocyanin (KLH) or bovine serum albumin (BSA) mixed with 10 mg aluminium hydroxide gel. IgE antibodies were measured in sera by PCA titres. High responses were obtained in BN rats (PCA titres about 10,000 after booster) and low responses in Lewis and SD rats. Positive booster responses were obtained in the three strains. Peritoneal mast cells collected from the three strains after immunization could degranulate on in vitro addition of specific antigen. In contrast, BN mast cells were bad receptors while Lewis and SD mast cells were good receptors for in vitro passive sensitization by mouse IgE antibodies. Total serum IgE was assayed by an in vitro competitive inhibition bioassay (CIB). The values before immunization were higher in BN (1-4 microgram/ml) than in Lewis (less than 0.25 microgram/ml) or SD rats (0.6 microgram/ml). After immunization, a striking increase could be observed in BN rats (up to 170 microgram/ml). There was no parallel between total IgE and IgE antibody levels at different times after immunization.
Assuntos
Antígenos/imunologia , Imunoglobulina E/biossíntese , Animais , Imunização Secundária , Cinética , Masculino , Mastócitos/imunologia , Anafilaxia Cutânea Passiva , Ratos , Ratos Endogâmicos Lew , Ratos Endogâmicos , Fatores de TempoRESUMO
The IgE antibody response was studied in DBA/2 mice; the mice were pretreated orally with albumin extracts from seeds of Jack fruit (Jackalbumin) and subsequently immunized subcutaneously with Jackalbumin mixed with ovalbumin (OA) and a synthetic adjuvant, muramyl dipeptide (MDP). The allergenicity of Jackalbumin was evaluated by its capacity to induce a specific IgE response which was measured by passive cutaneous anaphylaxis (PCA) and by degranulation of washed peritoneal mast cells following antigen challenge (Jackalbumin or OA). Antibody against the crude extracts and anti-lectin (FIISP) IgE responses were also tested by PCA. After being fed eight doses of 1 mg Jackalbumin, DBA/2 mice became immunized: i.e. specific IgE antibody responses were observed and the peritoneal mast cells became sensitized. An increase in IgE response was verified in mice that were pre-fed and subsequently immunized. The results indicated that: the albumin extracts from Jack seeds, containing lectins, can be allergenic by the oral route; multiple oral doses with these extracts can induce an enhancement of the IgE response on subsequent subcutaneous immunization; antigenically, Jackalbumin does not seem to cross-react with OA; the lectins contained in the albumin fraction from Artocarpus seeds were also shown to be allergenic; the IgE titres showed an inverse correlation to the degree of purification of the lectin used for PCA challenge.
Assuntos
Albuminas/imunologia , Imunoglobulina E/biossíntese , Proteínas de Plantas/imunologia , Albuminas/administração & dosagem , Animais , Feminino , Lectinas/imunologia , Mastócitos/imunologia , Camundongos , Camundongos Endogâmicos DBA , Ovalbumina/imunologia , Anafilaxia Cutânea Passiva , Lectinas de Plantas , SementesRESUMO
Possibility of inhibition of an efficient in vitro IgE-sensitization system was studied. The sentization of mouse peritoneal mast cells with an anti-ovalbumin IgE-rich fraction of serum, as tested by ovalbumin-induced degranulation, was inhibited by previous incubation with antisera of another or of no specificity. Fractionation and other experiments showed that the inhibiting activity correlated with IgE content. IgGl did not seem to have an effect. Sensitization was also inhibited by rat myeloma IgE, 50 ng giving a 50 per cent inhibition. Plots of the logarithms of rat and mouse IgE concentration vs their inhibitory effect on sensitization gave two parallel linear curves, indicating that mouse and rat IgE compete for the same receptor sites. It was thus possible to use this system as a sensitive bioassay for both mouse and rat IgE levels and, by comparing inhibition by mouse IgE to that by a known rat IgE standard, to obtain not only relative data but absolute mouse IgE levels. This, and also a better discrimination of IgE doses, was the major advantage of this bioassay in relation to the equally sensitive anti-IgE degranulation tests.
Assuntos
Imunoglobulina E/análise , Mastócitos/imunologia , Animais , Anticorpos , Anticorpos Anti-Idiotípicos , Antígenos , Ligação Competitiva , Hemocianinas/imunologia , Imunidade Materno-Adquirida , Imunização , Imunoglobulina G , Técnicas In Vitro , Camundongos , Ovalbumina/imunologia , Anafilaxia Cutânea Passiva , RatosRESUMO
The relations between human IgE and mouse peritoneal mast cells were studied in vitro. Non-heated human IgE sensitizes the mouse mast cells for degranulation on challenge with anti-human IgE. This capacity is lost after heating of human IgE at 56 degrees C. The degranulation only occurs in determined quantitative IgE-anti-IgE relationships, an excess of one or the other reagent inhibiting the reaction. Sensitization is practically instanteneous, and the degranulation is independent on the order of addition of the human IgE and anti-IgE. The human IgE can be removed from mouse peritoneal mast cells by a single washing. The results show that human IgE is unable to bind firmly to mouse peritoneal mast cells in vitro. It seems to induce the formation of biologically active human IgE-anti-human IgE complexes, which act on mouse mast cells and induce their degranulation.
Assuntos
Grânulos Citoplasmáticos/imunologia , Imunoglobulina E/metabolismo , Mastócitos/imunologia , Camundongos/imunologia , Animais , Complexo Antígeno-Anticorpo , Feminino , Temperatura Alta , Humanos , Técnicas In Vitro , Injeções Subcutâneas , Ovalbumina/administração & dosagem , CoelhosRESUMO
The heat inactivation at 56 degrees C of mouse IgE antibodies, measured by their PCA activity, was studied in various experimental conditions. Mouse IgE antibodies are partially protected against heat inactivation when previously diluted in sodium chloride or in phosphate buffer media. The protection is better at a higher dilution and molarity (phosphate 1M) and at pH 7. Heat inactivation is increased by the presence of reducing, alkylating and denaturating agents. Heat lability depends upon the concentration of serum proteins in the medium and is increased in presence of immunoglobulins.
Assuntos
Anticorpos/análise , Temperatura Alta , Imunoglobulina E/metabolismo , Animais , CamundongosRESUMO
The conditions for active sensitization of hamster peritoneal and pleural mast cells and IgE-induced histamine release as well as cell desensitization were defined. Immunization of hamsters with ovalbumin (5 micrograms) in Al/OH/3 gel (5 mg) with several boosters resulted in sensitization of peritoneal and pleural mast cells; in the presence of extracellular Ca++, pH of medium 7.2 and at 37 degrees C these cells released up to 70% of histamine on the challenge with specific antigen. Partial release was observed when the cells were challenged with antigen in the absence of extracellular calcium. The rate of release is high during the first seconds of activation and is complete at 1 min. 30 min preincubation of peritoneal and pleural mast cells in calcium-free conditions (in the presence of 4 mM EDTA) resulted in complete desensitization of cells to subsequent action of antigen in optimal conditions. The present experiments demonstrate, that hamster peritoneal and pleural mast cells can be a useful model system for in vitro studies of the mechanisms of IgE-induced cell activation.
Assuntos
Liberação de Histamina , Imunoglobulina E/imunologia , Mastócitos/metabolismo , Animais , Cálcio/fisiologia , Cricetinae , Feminino , Imunização , Masculino , Mastócitos/imunologia , Mesocricetus , Ovalbumina/imunologiaRESUMO
Allergoids have been used successfully for immunotherapy of allergic disorders. It has appeared to us that the effect of allergoids could be potentiated by their coupling to an immunomodulator. In the present study we show that a conjugate made up of the coupling of ovalbumin through glutaraldehyde action to the C. granulosum-derived immunomodulator P40 is completely devoid of antigenicity and of cross-reactivity with ovalbumin. This conjugate was found to significantly inhibit mast cell degranulation. It also proved to be capable of protecting against the lethal systemic anaphylactic shock sensitized mice. Immunotherapy was performed in patients hypersensitive to either the pollen of Dactylis glomerata or to the house dustmite allergens using the conjugates made of the specific allergens and of the P40. Clinical improvement was observed in a significant percentage of the patients subjected to immunotherapy. Administration of the conjugates did not result in untoward reactions in any of the patients.
Assuntos
Alérgenos/administração & dosagem , Antígenos de Bactérias/imunologia , Corynebacterium/imunologia , Dessensibilização Imunológica/métodos , Rinite Alérgica Sazonal/terapia , Alérgenos/imunologia , Animais , Antígenos de Bactérias/administração & dosagem , Células Cultivadas , Reações Cruzadas , Feminino , Humanos , Mastócitos/imunologia , Camundongos , Ácaros/imunologia , Ovalbumina/imunologia , Poaceae/imunologia , Pólen/imunologia , Rinite Alérgica Sazonal/imunologiaRESUMO
Peritoneal mast cells of Syrian hamsters release histamine to the action of concanavalin A (Con A) in dose-dependent fashion. The rate of release was very rapid in the first seconds of cell activation and completed in 60 s after the challenge. Morphological changes concomitant to the lectin treatment, followed by electron microscopy, show that early signs of exocytosis are seen after 10 s. The process starts in peripherally located granules which swell, have a decreased density and form pores by fusion of the cellular membrane and the perigranular membranes. Then it spreads toward the cell interior by fusion of granules and forming intracytoplasmic cavities. Some extruded granules are also observed. Preincubation of lectin with rat IgE or with rat serum induced an inhibition of its histamine releasing action. Immunization increased the Con A-induced histamine release in young but not in older hamsters. An IgE-mediated mechanism is suggested for the parallel ultrastructural changes and histamine release effects induced by Con A on the hamster mast cell.
Assuntos
Concanavalina A/farmacologia , Liberação de Histamina/efeitos dos fármacos , Mastócitos/efeitos dos fármacos , Mastócitos/metabolismo , Animais , Cricetinae , Relação Dose-Resposta a Droga , Exocitose , Feminino , Imunização Passiva , Imunoglobulina E/imunologia , Técnicas In Vitro , Lectinas/imunologia , Mastócitos/citologia , Microscopia Eletrônica , Fatores de TempoRESUMO
L-Leucine methyl ester (Leu-OMe), a lysosomotropic compound, has been found to eliminate several lysosome-rich cellular subtypes and all natural killer cell function from peripheral blood mononuclear cells. In this report, the effect of Leu-OMe on mouse peritoneal mast cells is described. The L-Leu-OMe induced the release of histamine from mouse peritoneal mast cells in a dose-dependent manner (0.25 to 3 mM), while its D-stereoisomer had no effect. L-Leu-OMe displayed also a potent histamine release effect on purified mast cells, indicating a direct effect on mast cells. The monitoring of radioactive chromium release versus histamine release showed that both processes may be unrelated for Leu-OMe concentrations inferior to 1.5 mM. At higher doses, L-Leu-OMe, but not its D-stereoisomer, exerted a potent cytotoxic effect on mast cells. The secretory effect of Leu-OMe was temperature- and energy-dependent. Experiments performed in the absence of extracellular calcium and magnesium demonstrated that these divalent cations were not necessary for the Leu-OMe-induced histamine release, and their deprivation even involved a higher histamine release. The secretory characteristics of the Leu-OMe-induced histamine release appeared to be different from those of the IgE-induced ones. These results support the conclusion that exposure of mouse peritoneal mast cells to high doses of L-Leu-OMe results in killing of these cells, that are new targets of this lysosomotropic agent.
Assuntos
Citotoxicidade Imunológica , Liberação de Histamina , Leucina/análogos & derivados , Mastócitos/efeitos dos fármacos , Animais , Anticorpos , Relação Dose-Resposta a Droga , Imunoglobulina E , Leucina/farmacologia , Mastócitos/imunologia , Camundongos , Camundongos Endogâmicos CBA , Peritônio , TemperaturaRESUMO
Mouse and rat IgE fix firmly to the peritoneal mast cells from the other species, sensitizing them for anaphylactic reaction. Sensitization with IgE can be demonstrated by inducing degranulation either with specific antigens or with corresponding anti-IgE. Sensitization of rat mast cells by mouse IgE antibodies is more easily obtained than that of mouse mast cells by rat IgE antibodies. In this case, anti-IgE-induced degranulation is higher than antigen-induced degranulation. Heterologous sensitization by IgE is time requiring and temperature-dependent. Its kinetics depend upon IgE concentration. Cross-reactions between IgE from one species and anti-IgE from another species have been observed: anti-IgE for one species is able to neutralize PCA reaginic activity of sera from the other species; anti-rat IgE induces degranulation of mouse actively sensitized mast cells. The results suggest strongly that there exists a structural and functional similarity between the IgE molecules from the two species.