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Background: Nearly 60% of birch pollen–allergic patients react exclusively to Bet v 1. With use of the skin blister model, previously only established for installation of crude allergens, we have for the first time characterized the inflammatory response in vivo to recombinant birch pollen allergen, rBet v 1, molecules (rBet v 1 wild type, fragments and trimer). Objective: Our purpose was to examine whether challenge with rBet v 1 derivatives (fragments and trimer) compared with rBet v 1 wild type differs with respect to influx of activated eosinophils and detectable levels of cytokines/chemokines related to allergic inflammation in skin chambers applied to birch pollen–allergic patients. Methods: The skin blister chambers were filled for 2 hours with rBet v 1, the derivatives or PBS and heparin (negative control). The fluids were analyzed after 2 and 8 hours. The number of eosinophils was determined and EG2 and CD69 expression measured by flow cytometry. Cytokines and mediators were analyzed by ELISA and RIA techniques. Results: Comparable numbers of eosinophils were recruited to the chambers challenged with rBet v 1 molecules, but the eosinophils from the rBet v 1 wild-type challenged chambers showed a significantly higher expression of CD69. The levels of eotaxin were similar in all 4 chambers, whereas rBet v 1 wild type induced significantly higher levels of histamine, eosinophil cationic protein, and GM-CSF than the derivatives did. Recombinant Bet v 1 trimer elicited significantly lower levels of IL-4 compared with rBet v1 wild type. Conclusion: Genetically engineered hypoallergenic rBet v 1 derivatives recruited eosinophils analogously with rBet v 1 wild type. However, the derivatives exhibited a lower capacity to activate eosinophils and to release proinflammatory mediators and T helper type 2–derived cytokines. The derivatives may therefore be candidate molecules for specific immunotherapy of birch pollen allergy with reduced risk of inducing allergenic or inflammatory side effects. (J Allergy Clin Immunol 2000;106:101-9.)

The aim of this study was to examine potential differences between healthy and atopic subjects with regard to IgE-mediated cutaneous inflammation. For this purpose, we analyzed histamine, tryptase, leukotriene B4, albumin, eosinophils, and total leukocytes in skin chamber fluid after challenge with anti-human IgE. We also measured gross skin reactivity (wheal, flare, and late-phase reactions), circulating IgE, and eosinophils, as well as the state of eosinophil activation. It was found that despite having more circulating IgE, the skin responsiveness of the atopic subjects did not differ significantly from that of the nonatopic subjects with respect to mediator release, albumin extravasation, or total recruitment of leukocytes. Moreover, the sizes of anti-IgE-induced wheal, flare, and late-phase reactions were very similar in the two groups. On the other hand, significant recruitment of eosinophils during the IgE-mediated reaction was more or less restricted to the atopic group. Yet the recruited eosinophils, of which the majority was in an early state of activation before degranulation, did not seem to contribute significantly to the IgE-mediated delayed skin edema. Furthermore, the eosinophil count in anti-IgE chambers of the atopic subjects did not correlate with any of the other parameters monitored. Thus because the anti-IgE-induced recruitment of eosinophils appeared to be unrelated to factors such as the number of peripheral blood eosinophils, the degree of mast cell activation, the intensity of inflammatory skin changes, and the level of circulating IgE, it is apparent that the mechanisms for and pathophysiologic role of IgE-mediated dermal eosinophil accumulation in atopic subjects require further investigation.

The concept of an obligate association between mast cell activation and development of a late phase reaction (LPR) to various agents in human skin was further elucidated. Skin sites were treated four times at 24 h intervals with non-LPR-inducing doses of the histamine liberator compound 48/80 in 10 volunteers. The previously compound 48/80-challenged sites responded with an approximately 40% attenuated early response (P less than 0.01) and a 70% reduction of the LPR (P less than 0.001) to subsequently injected anti-IgE as compared with simultaneous reactions at control sites. The data suggest that mediators from the cutaneous mast cells are necessary for the final expression of an LPR.

Pharmacological modulation of the immediate and late phase reaction (LPR) to anti-human IgE was further investigated in a double-blind cross-over study. Tranexamic acid (AMCA) 1 g t.i.d. 24 h prior to and following intradermal injection of anti-IgE produced an approximate 40% inhibition of the LPR (P less than 0.01) without affecting the early response as compared with placebo in 10 volunteers. Antagonistic effect on activation of fibrinolysis and possibly the complement system is suggested as a possible mode of action.

In eight atopic subjects wheal and flare responses to intradermally injected horse dander and histamine were determined after pretreatment with 5 mg oral terbutaline or placebo in a double-blind cross-over study. In each individual a dose of allergen was used that produced a flare reaction approximately the size of the ED50 for histamine. Pretreatment with terbutaline was found to attenuate both the wheal and the flare reactions to allergen throughout the observation period of 150 min but only the effect on the wheal response reached statistical significance (P less than 0.01). The responses to histamine were not influenced. In five subjects with cold urticaria, treatment with 2.5 mg terbutaline t.i.d. for a week had no effect on the time period of cold provocation needed to evoke an urticarial lesion. It is concluded that oral treatment with terbutaline may produce an inhibitory action on allergen induced reactions but that this effect is not strong enough to interfere with clinical skin testing and hence the drug need not to be withdrawn prior to such testing.

Wheal and flare reactions as well as a late cutaneous allergic reaction (LCAR) were induced by anti-human IgE in healthy subjects. The effect of the beta 2-adrenoceptor stimulant terbutaline, the histamine-1 (H1) receptor blocking agent mepyramine and the synthetic glucocorticoid betamethasone on these reactions was studied. All administrations were given intradermally (i.d.). The immediate reaction to anti-IgE was inhibited by 3 micrograms terbutaline and 30 micrograms mepyramine (P less than 0.01) whereas 50 micrograms betamethasone had no effects. Terbutaline had no effect on the flare response induced by i.d. injected histamine but a slight effect on whealing. Terbutaline and mepyramine weakly reduced the LCAR throughout the observation period of 24 h (P less than 0.01). In contrast, betamethasone almost completely abolished the LCAR. It is concluded that the two phases of the skin reaction to anti-IgE are interrelated since an inhibition of the early phase was followed by an attenuation of the LCAR. The mechanism of action of steroids seems to differ fundamentally from that of other anti-allergic drugs since inhibition of the early step in the reaction is not essential to the action on the late step. It is further suggested that terbutaline inhibits anti-IgE-mediated cutaneous reactions by inhibition of the mast cell release reaction.

Beta-adrenoceptor stimulating agents possess anti-allergic effects in vitro and in vivo. To study the site of action further 15 atopic subjects were pretreated with 1 microgram terbutaline injected intradermally (i.d.) followed by allergen challenge. Other skin sites challenged without such pretreatment served as controls. At different time intervals the skin was rechallenged with the same allergen, dissimilar allergen, or the histamine liberating agent, compound 48/80. In addition terbutaline was injected i.d. alone followed by allergen challenge at different time intervals to determine the duration of the anti-allergic effect. Pretreatment of the skin with 1 microgram terbutaline inhibited skin reactions to subsequently injected allergen with approximately 80% (flare) and 60% (wheal) (P less than 0.001) as compared with control. The inhibitory effect of terbutaline on the wheal and flare response to allergen was found to last up to 8 h (P less than 0.01). Rechallenge with the same allergen or a dissimilar challenging agent resulted in a reduced skin reaction compared with control challenge (P less than 0.01). Skin challenged with allergen in the presence of terbutaline gave a diminished response on rechallenge with the same allergen 24 h later, whereas rechallenge with an unrelated allergen or compound 48/80 produced a response similar to that of the control. The results suggest that allergen challenge in the presence (or absence) of terbutaline desensitizes the mast cells to further challenge with the same allergen and that terbutaline is capable of preventing mediator depletion of skin mast cells.

The influence of the time interval between administration of a glucocorticoid, betamethasone, and anti-IgE on the effect of the drug on the early and late phase response (LPR) to anti-IgE was examined in 38 volunteers in a randomized controlled study. The immediate flare and wheal responses to intradermally (i.d.) injected anti-IgE were inhibited by approximately 20 and 30% respectively by a 10 day i.d. pretreatment period with repeated 50 micrograms doses (total 300 micrograms) of betamethasone (P less than 0.01), whereas responses to histamine were not influenced. A single i.d. injection of 50 micrograms betamethasone 24 h prior to anti-IgE challenge attenuated the wheal response (P less than 0.05), whereas injection of the drug 2 h prior to and together with anti-IgE had no influence on the early response. Corresponding LPRs were reduced approximately 30% throughout the observation period of 1-24 h (P less than 0.01). Injecting the drug i.d. 30 min following anti-IgE challenge had no significant influence on the total observation period (1-24 h) of the LPR but antagonized the LPR by 35% at 6-24 h (P less than 0.01). It is concluded that glucocorticoids inhibit IgE-dependent LPRs. When extending the topical pretreatment period with the compounds also the early part of the allergic reaction is attenuated.

beta-Adrenoceptor stimulating agents possess anti-allergic effects in vitro and in vivo. To study the mechanism of action further we compared in 10 atopic subjects the effects on allergen-induced cutaneous reactions of intradermally (i.d.) injected prenalterol (1-10 microgram) terbutaline (2.5-250 ng) and KWD 2131 (100 ng-2.5 microgram), i.e. compounds with preferential actions on beta 1-, beta 2-adrenoceptors and on beta-adrenoceptors not fitting this classification. All injections were given according to a double-blind design. Terbutaline and KWD 2131 produced a dose-dependent inhibition of the skin reactions induced by injecting horse dander allergen 5 min later. Terbutaline was about 20 times as potent as KWD 2131 whereas prenalterol was inactive. Propranolol fully blocked the anti-allergic effect produced by terbutaline. The results suggest that the cutaneous anti-allergic effect of beta-adrenoceptor-stimulating drugs is mediated via activation of beta 2-adrenoceptors, possibly on the mast cell, resulting in inhibition of mediator release.

The aim of this study was to investigate whether ACE-inhibitors could influence bronchial reactivity and interfere with inflammatory skin responses. Ten hypertensive subjects, who had reacted with coughs during ACE-inhibitor therapy, were treated in a double-blind crossover fashion for two weeks with enalapril and with placebo. Enalapril reduced the PC 50 value for histamine and augmented the dermal response. Circulating eosinophilic leukocyte level in venous blood dropped markedly after the histamine bronchoprovocation performed during enalapril treatment. Plasma substance P was reduced after histamine provocation performed during placebo treatment, whereas this reduction was abolished by enalapril. In this study, we have demonstrated ACE-inhibitor-induction of moderately increased bronchial reactivity in subjects with suspected ACE-inhibitor-elicited coughs. It is suggested that coughing during ACE-inhibitor therapy is due to an increased inflammatory state in the airways.

The beta-adrenoceptor stimulating agent terbutaline (2 ng-2 microgram) injected intradermally in eight atopic subjects produced a dose-dependent inhibition of the skin reactions induced by subsequently injected allergen. After injection of 0.5 microgram terbutaline inhibition of the flare and weal responses was demonstrable throughout the observation period of 90 min. The flare response induced by histamine, the histamine liberator compound 48/80 and the proteolytic enzyme trypsin was not inhibited by terbutaline in the doses used, suggesting a selective action of terbutaline on the allergen-induced response. The weal response elicited by histamine and compound 48/80 was slightly reduced by 2 microgram terbutaline. It is suggested that pretreatment of the skin with terbutaline interferes with the ability of the cutaneous mast cells to respond to challenge with allergen and that terbutaline produces this effect in doses lower than those needed to counteract the permeability increasing effect of released mediator substances.