Susumu Hosoi

Urinary leukotriene E4 after exercise challenge in children with asthma

To assess the role of sulfidopeptide leukotrienes in the pathogenesis of exercise-induced asthma (EIA), the urinary levels of leukotriene E4 (LTE4), a metabolite of LTC4 and LTD4, were measured by RIA before and after exercise in 13 children with EIA and 10 healthy children. Mass spectrometry was used to confirm the presence of LTE4 in urine and the specificity of the RIA. There was no significant difference in the urinary LTE4 levels before exercise between the children with asthma and healthy children (109 [21 to 265] versus 122 [45 to 156] pg/mg of creatinine; median and range). Urinary LTE4 levels increased significantly after exercise in the children with EIA (from 109 [21 to 265] to 196 [40 to 655] pg/mg of creatinine; median and range; p less than 0.05) but not in the healthy children. The children with asthma demonstrated no significant correlation between the LTE4 level after exercise and the degree of bronchoconstriction, as revealed by the maximal percent fall in the peak expiratory flow rate. Taken together with a recent study that pretreatment with a potent and selective LTD4 antagonist markedly attenuated EIA, our findings suggest that sulfidopeptide leukotrienes may play some role in the pathogenesis of this type of asthma with other factors also being involved in determining the overall airway response.

Obesity and the prevalence of allergic diseases in schoolchildren

Although the association between obesity and bronchial asthma (BA) has been gaining more attention, few studies have been conducted concerning the relationship between obesity and other allergic diseases. The objective of this study was to determine whether and how childhood obesity is associated with allergic diseases other than BA, such as atopic dermatitis (AD), allergic rhinitis (AR), allergic conjunctivitis (AC), and either AR or AC (AR/AC). A questionnaire was administered to the parents of 50,086 Japanese schoolchildren. Associations between childhood obesity and the various allergic diseases were evaluated by univariate and multivariate logistic models. Significant associations were found between higher body mass index (BMI) and AD (p = 0.03), and lower BMI and AC (p < 0.0001), and AR/AC (p < 0.0001). There was a significantly higher prevalence of BA in girls with obesity (p = 0.009) than in those without obesity. Significantly lower prevalence of AC (p = 0.01) and AR/AC (p = 0.002) among children with obesity, and AR (p = 0.04) and AR/AC (p = 0.0004) among boys with obesity were observed than those without obesity. Those who were obese and had AD were significantly more likely to have severe symptoms (p = 0.01). Overall, childhood obesity has positive associations with BA prevalence and AD severity, whereas it has negative associations with AR and AC prevalence, especially among boys. Changes in the immunologic balance accompanied by obesity might have different effects on each type of allergic disease. Exploring the mechanisms by which childhood obesity affects allergic status should lead to new management options for childhood allergy.

Tyrosine phosphorylation and its possible role in superoxide production by human neutrophils stimulated with FMLP and IgG

Superoxide production by human neutrophils stimulated with FMLP and soluble aggregated human IgG were inhibited in a dose dependent manner by two kinds of tyrosine kinase inhibitors, erbstatin and genistein. Superoxide production stimulated with surface bound IgG, however, was scarcely inhibited by either inhibitor. Protein tyrosine phosphorylation studies with immunoblotting revealed specific tyrosine phosphorylation of a 40 Kd protein by soluble aggregated and surface bound IgG, and that of a 39 Kd protein, as well as the 40 Kd protein, by FMLP. These were all inhibited by the tyrosine kinase inhibitors. These data suggest that superoxide production induced by FMLP and soluble aggregated IgG are, at least in part, tyrosine kinase dependent, but the tyrosine kinases and/or substrates of tyrosine kinases involved may be different. In addition, tyrosine kinase independent pathways are also suggested to be involved in superoxide production by stimulation with surface bound IgG.

C8/119S Mutation of Major Mite Allergen Derf-2 Leads to Degenerate Secondary Structure and Molecular Polymerization and Induces Potent and Exclusive Th1 Cell Differentiation

Hyposensitization therapy for atopic diseases has been conducted for decades but suffered from many problems including anaphylactic reactions. We previously developed a mutant protein of the major mite allergen Derf-2, C8/119S, which showed reduced binding to IgE. The C8/119S mutant was shown to exhibit more efficient hyposensitizing effect than Derf-2 in the animal model of allergic bronchial asthma. In the present study, we indicate that C8/119S exhibits markedly augmented immunogenicity for the proliferation of Derf-2-specific human T cells and T cell clones irrespective of the epitope specificity as compared with Derf-2. Furthermore, C8/119S has induced potent and almost exclusive differentiation of Th1 cells from the peripheral blood of atopic patients in vitro. Neither Ag dosage effect nor absence of B cell-mediated Ag presentation could fully account for these effects. C8/119S has been indicated to lose the characteristic β-barrel structure as judged by circular dichroism spectroscopic analysis and to polymerize solubly in physiological condition. Heating of Derf-2 also caused less stable molecular aggregation, but it hardly affected the secondary structure and failed to induce such a polarity toward the Th1 cell differentiation. These results have indicated that the degenerate secondary structure of C8/119S leading to stable molecular polymerization is primarily responsible for the marked increase in T cell-immunogenicity and the induction of exclusive Th1 cell differentiation in atopic patients. It has been suggested strongly that the recombinant C8/119S protein can provide an effective Ag with the least risk of anaphylaxis for allergen immunotherapy against house dust mite in human.

Involvement of CD11b/CD18 in enhanced neutrophil adhesion by Fc gamma receptor stimulation.

Neutrophils showed a rapid and transient adhesion to immunoglobulin G (IgG)‐coated plates compared with their adhesion to bovine serum albumin (BSA)‐coated plates: the adhesion reached a peak after 15 min of incubation and then gradually returned to almost the basal state in 60 min. The addition of monomeric IgG or anti‐FcγRII monoclonal antibody (mAb) (IV.3) suppressed the increase in adhesion, whereas anti‐FcγRIII mAb (3G8) was hardly effective, indicating that the interaction of FcγR, especially FcγRII, with coated IgG is involved in the process. Adhesion was also blocked by cytochalasin B, suggesting that functional actin filament structures are crucial. Protein kinase inhibitors, erbstatin and genistein, inhibited the adhesion in a dose‐dependent manner. The adhesion was inhibited by anti‐CDllb (Ml/70) and anti‐CD18 (MHM23, TS1/18) mAbs. Moreover, neutrophils from a patient with complete leukocyte adhesion deficiency syndrome did not show increased adhesion to IgG‐coated plates. The adhesion of neutrophils to fibrinogen‐ and BSA‐coated plates was also increased when FcγR was stimulated in the fluid phase with soluble aggregated IgG, which was also inhibited by anti‐CDllb mAb. Stimulation of neutrophil FcγR with soluble aggregated IgG enhanced the expression of CDllb in concert with the enhanced adhesion. These data collectively suggest that stimulation via FcγR evokes a tyrosine kinase‐dependent and actin filament‐dependent intracellular signal that enhances the specific and nonspecific adhesive activity of neutrophils, presumably through the activation of CDllb/CD18. J. Leukoc. Biol. 55: 735–742; 1994.

DiGeorge syndrome with hypogammaglobulinaemia: a patient with excess suppressor T cell activity treated with fetal thymus transplantation

A male infant with DiGeorge syndrome had hypogammaglobulinaemia with a normal number of B cells. CD3(+) T cells were reduced and the CD4(+)/CD8(+) ratio was reversed. Proliferative responses of T cells to mitogens and to allogeneic cells were low. The pokeweed mitogen (PWM)-induced B cell differentiation assay revealed a higher than normal suppressor T cell activity. This suggests that some T cells had differentiated into functionally mature cells resulting in an imbalance of regulatory T cell functions and that excess suppressor activity might play a role in hypogammaglobulinaemia. Fetal thymus transplantation improved both cellular and humoral immunity. The patients susceptibility to viral and bacterial infections, proliferative response of T cells and serum Ig concentration returned to normal. The excess suppressor activity seen before transplantation disappeared. Hypocalcaemia did not improve. These results show that fetal thymus transplantation was effective not only in reconstituting cellular immunity but also in normalizing the imbalance of regulatory T cell functions in this patient with DiGeorge syndrome.

Exercise-induced urinary excretion of leukotriene E4 in children with atopic asthma

ABSTRACT: Urinary levels of leukotriene (LT) E4, a stable end-product of LTC4 and LTD4, were measured before and after exercise in 10 children with severe asthma and seven children with moderate asthma using HPLC and RIA to clarify the relationship of LT to the severity of asthma and to the degree of bronchospasm in exercise-induced asthma. The urinary LTE4 level significantly increased after exercise in the severe asthma group, but not in the moderate asthma group (14.3 ± 14.5 to 24.3 ± 20.6 versus 19.6 ± 12.3 to 17.6 ± 10.8 ng/mmol creatinine, p < 0.05). The urinary LTE4 level increased in 10 patients (eight with severe asthma), and it decreased in seven patients (five with moderate asthma). A significant difference in the degree of bronchospasm after exercise (as shown by the maximal % fall in the peak expiratory flow rate), was seen when patients with increased urinary LTE4 excretion were compared with those with decreased excretion (60.4 ± 17.3 versus 24.1 ± 14.3%, p < 0.01). Our findings suggest that exercise-induced asthma, or at least a subtype of exercise-induced asthma, may partly develop through the release of LTC4.

Relationships between atopy and lung function: results from a sample of one hundred medical students in Japan

BACKGROUND The prevalence of allergic diseases has been increasing dramatically and several studies have shown that atopy is related to asthmatic symptoms and bronchial hyperresponsiveness. OBJECTIVE To observe the relationships between atopic status and asthmatic predisposition (obstructive change in lung function) in apparently healthy young adults in Japan. METHODS A sample of 100 healthy Japanese medical students were subjected to a skin prick test for 11 common aeroallergens and food allergens, and their spirometric lung function was measured. RESULTS Surprisingly, 90% of them showed a positive prick test result for at least one of the 11 allergens tested, and 59% of them showed allergic responses to more than three allergens. The positive rate for Dermatophagoidesfarinae (Der) was the highest (71.0%), followed by house dust (57.0%), Dactylois gloinerata (42.0%), Cryptomeria gromerata (Cry) (40.0%), and cat fur (39.0%). Furthermore, there was no statistical difference in the positive rates for Der and Cry between groups with and without either the present illness or past history of any of the three major allergic diseases: bronchial asthma (BA), atopic dermatitis (AD), or allergic rhinitis (AR). Compared with the positive rates for these aeroallergens, those for food allergens were much lower (4% to 9%). Several lung function parameters, including the levels of FEV1% and %V50 which reflect obstructive pulmonary changes, showed significant negative correlation to the number of skin prick test-positive allergens. The same correlation was observed for groups without either the present illness or past history of BA. CONCLUSION These data suggest that those who are multi-allergic tend to feature subclinical asthma-like changes in their lung functions. Further studies are needed to determine whether this multi-allergic status can lead to future onset of asthma or other allergic diseases.

Comparison of skin prick test with serially diluted wild-type and genetically engineered recombinant Der f2.

BACKGROUND C8/119S, a genetically engineered less allergenic mutant of group II allergen (Der f2) of house dust mite, Dermatophagoidesfarinae, was constructed in order to reduce the risk of anaphylactic reactions of allergen specific immunotherapy. OBJECTIVE To further evaluate, with a larger number of mite-allergic patients, the safety of C8/119S for the treatments of humans. METHOD We tested the dose-dependent responses of 20 mite-allergic volunteers to a skin prick test with C8/119S and wild-type recombinant Der f2 and compared the biologic potentials of these allergens to induce type I allergic reactions. In a separate experiment, we compared IgE binding capacities to C8/119S and to wild-type recombinant Der f2 in individual sera from 34 mite-allergic donors. RESULTS The concentration of C8/119S needed to induce positive skin prick test (SPT) reaction was at least 100 times more than that of recombinant Der f2 in 95% of the volunteers tested. Consistent with this result, IgE binding data showed that 85% of the mite-allergic donors had little or no detectable IgE bound to C8/119S. Our data also shows that a minority of mite-allergic patients responded in a similar manner to both C8/119S and wild-type recombinant Der f2. CONCLUSION Our data confirms that C8/119S is much less allergenic and thus can be used safely for immunotherapy of most of mite-allergic patients. Care should still be taken because, in a minority of patients, C8/119S may cause similar type I allergic reactions as does wild-type recombinant Der f2.

Anti-Human IgE Monoclonal Antibodies Recognizing. Epitopes Related to the Binding Sites of High and Low Affinity IgE Receptors

Anti‐human IgE monoclonal antibodies (mAbs) were produced and eight clones recognizing epitopes on native IgE were selected. Epitopes were mapped by a competitive inhibition enzyme‐linked immunosorbent assay, Western blotting and a multi‐pin peptide technology. Four sites (one each in the Cε1, Cε2, Cε2/Cε3 junction and Cε3) were recognized by the mAbs. The relationship between the four epitopes and the binding sites of high and low affinity IgE receptors (FcεRI and FcεRII, respectively) was studied using a monovalent Fab fragment of each mAb as a binding inhibitor. The IgE‐FcεRII binding was clearly inhibited by the mAb recognizing the Cε2/Cε3 junction, suggesting that FcεRII binds to a rather limited area around the Cε2/Cε3 junction. The IgE‐FcεRI binding, on the other hand, was scarcely inhibited by any single mAb. However, the binding was inhibited when the epitope in Cε2 was blocked simultaneously with that at the Cε2/Cε3 junction or with that in Cε3, indicating that these three distinct epitopes are related to the FcεRI binding sites. When these three epitopes were shown in the stereograph of human IgE, the FcεRI binding area was spread largely on the groove side between Cε2 and Cε3 domains. These results suggest that FcεRI acquires the high affinity through multiple bindings.