SummaryMast cells have long been believed to be the central effector cells in the development of immunoglobulin (Ig)E-dependent anaphylaxis. In this study, we investigated the role of mast cells in IgE-dependent hapten-induced active fatal anaphylaxis using mast cell-deficient WBB6F1-W/W v ( W/W v ) and congenic normal ( ϩ / ϩ ) mice. Although a 5-min delay in shock signs and death were observed in W/W v mice, 100% fatal reactions to penicillin V (Pen V) occurred in both ϩ / ϩ and W/W v mice. Administration of monoclonal anti-IL-4 antibody completely prevented the fatal reactions, and the effect of anti-IL-4 was associated with its suppressive activity on Pen V-specific serum levels of IgE, but not IgG. The platelet-activating factor (PAF) antagonist, BN 50739, completely prevented the fatal reactions in both strains of mice. Our kinetic study revealed, in contrast to no elevation of plasma histamine level in W/W v mice, high levels of PAF in the circulation after challenge in both ϩ / ϩ and W/W v mice, albeit to a lesser degree in the latter case. These data indicate that cells other than mast cells are sufficient to induce an IgE-dependent active fatal anaphylaxis by elaborating PAF, which is the critical mediator for fatal murine anaphylaxis. Thus, mast cells may not contribute importantly to protein-induced anaphylaxis. Some evidence indicates that protein-induced anaphylaxis can be elicited by IgG Abs (9, 10) even in the absence of IgE Abs (11), suggesting that cells other than mast cells that bind IgG Abs elaborate sufficient mediators leading to fatal reactions. Nevertheless, mast cells have long been believed to be the central effector cells in the development of IgE-dependent anaphylaxis. However, the in vivo extent to which the reactions are mast cell-dependent remains to be elucidated due to the lack of a suitable animal model of IgE-dependent anaphylaxis.We have recently developed a murine model of IgEdependent, penicillin V (Pen V)-induced active fatal anaphylaxis (12). The reaction was 100% fatal in C57BL/6 mice and was exclusively IgE dependent, since ( a ) IgE, but not IgG, Abs against Pen V passively sensitized normal mice to develop severe anaphylactic reactions; ( b ) anti-IL-4 mAb completely prevented the fatal reaction; and ( c ) the effect of anti-IL-4 was associated with its suppressive activity on Pen V-specific serum IgE, but not IgG, levels. This model allowed us to investigate the role of mast cells in IgE-dependent anaphylaxis. In this study, the role of mast cells in IgE-dependent Pen V-induced anaphylaxis using genetically mast cell-deficient WBB6F1-W/W v ( W/W v ) and congenic normal ( ϩ / ϩ ) mice was investigated. It was 1 Abbreviations used in this paper: CGG, chicken gammaglobulin; NP, nitrophenol; PAF, platelet-activating factor; PCA, passive cutaneous anaphylaxis; Pen V, penicillin V.
Theileria sergenti were isolated from infected erythrocytes by hypotonic lysis, and soluble merozoite antigens were purified by sonication and differential centrifugation. The preparation contained 29, 34, 35 and 105 kD immuno-dominant poly-peptides. The soluble antigens (0.5 mg/ml) were prepared and fortified with Freund's adjuvant. Five month old naive Korean calves were subcutaneously inoculated with the preparation and a booster dose was administered 4 weeks later. Nine weeks after the booster dose, vaccinates and controls were challenged with a homologous stabilate (5.6 x 10(6) RBC/dose, 40% Parasitemia). All animals were monitored for hematocrit, total erythrocyte count, parasitemia and for the specific antibody by Western immunoblot (WB) and indirect immuno-fluorescent antibody (IFA) test. By 18 weeks after vaccination (6 weeks after the challenge), vaccinated cattle had an average IFA titer of 1:10,240 compared with 1:1,280 of the controls. The vaccinates showed negligible change in hematocrit and total RBC count whereas control animals showed significant (p less than 0.05) hematological changes and associated anemia. After vaccination and challenge, the antibody responses demonstrated that vaccination had induced significant production of antibody to the 29 and 35 kD polypeptides. The latter polypeptide was much more strongly recognized by the vaccinated animals, and thus it may be a potential candidate for the vaccine.
We previously reported that anti-IL-4 mAb (11B11) failed to prevent protein-induced fatal murine anaphylaxis. To investigate the effect of anti-IL-4 on hapten-induced anaphylaxis, a model of murine anaphylaxis induced by antibiotics, penicillin V (Pen V) and cephalothin (CET), was developed, and the effect of anti-IL-4 on the anaphylaxis was observed. Pen V and CET induced 100 and 70 to 90% fatal reactions, respectively, when C57BL/6 mice were sensitized i.p. with 500 microg of antibiotic-OVA conjugate with 2 x 10(9) Bordetella pertussis and 1.0 mg of alum and challenged i.v. with 100 microg of antibiotic-BSA conjugate 14 days later. Serum taken from mice sensitized to Pen V passively sensitized normal mice to develop systemic anaphylaxis, and this ability of the serum was abrogated by heating at 56 degrees C for 2 h or depletion of IgE, but not IgG, Abs. Thus, the antibiotic-induced fatal reaction was an IgE-dependent anaphylactic reaction. Administration of anti-IL-4 at the beginning of sensitization completely prevented the fatal anaphylactic reactions to both Pen V and CET. This effect of anti-IL-4 was associated with its suppressive activity on antibiotic-specific serum IgE, but not IgG, levels. More importantly, anti-IL-4 therapy in previously sensitized mice was also effective in preventing the fatal reactions and rapidly reduced the established IgE levels. This study provides a new animal model of hapten-induced anaphylaxis and indicates that blocking of IL-4 activity may be beneficial in allergic diseases caused by a variety of haptens in which IgE Abs play a major role.
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