Instillation Of Allergen Research Articles

Lung parenchymal and airway changes on CT imaging following allergen challenge and bronchoalveolar lavage in atopic and asthmatic subjects.

Computed tomography (CT) imaging findings in the lungs in the setting of an acute allergic response and following bronchoalveolar lavage (BAL) are not well established. Our goals are to characterize the pulmonary CT findings of acute allergic response in both asthmatic and non-asthmatic subjects and, secondarily, to characterize the pulmonary imaging findings following BAL. In this prospective observational (cohort) study, we identified atopic, asthmatic (AA) and atopic, non-asthmatic (ANA) subjects. CT of the chest was performed following BAL and instillation of an allergen (AL) and of an inert diluent (DL). Two radiologists analyzed the CT examinations for airway and parenchymal changes. We had a cohort of 20 atopic subjects (AA=10, ANA=10; F=11, M=9; median age: 23.5 years, range: 18-48 years). Compared to diluent instillation and BAL, allergen instillation resulted in more significant bronchial wall thickening (AL=70%, DL=0%, BAL=0%, P<0.01), consolidations (AL=55%, DL=0%, BAL=15%, P<0.05), and septal thickening (AL=35%, DL=0%, BAL=0%, P<0.01). When present, consolidations tended to be more common in asthmatic subjects compared to non-asthmatics following instillation of the allergen, although this did not reach statistical significance (AA=80% vs. ANA=30%; P=0.07). BAL, on the other hand, resulted in more ground-glass opacities (BAL=15/20, 75% vs. AL=2/20, 10%, vs. DL=0/20, 0%; P<0.01). Acute allergic response in the lungs can result in significant bronchial wall thickening, septal thickening, and consolidations in those with atopy, particularly those with asthma. Localized ground-glass opacities may be expected following BAL, and care should be taken so as to not misinterpret these as significant pathology.

Allergen challenge during halothane compared to isoflurane anesthesia induces a more potent peripheral lung response

Allergen instillation in anaesthetized vs. awake animals results in increased distribution of allergen in the lung. Halothane is a more potent bronchodilator of the small airways than isoflurane. As small airways contribute to asthma pathogenesis, we questioned whether intranasal challenge under halothane vs. isoflurane anesthesia would lead to an increase in allergen deposition in the lung periphery and, consequently, an enhanced allergic response. C57Bl/6 mice were sensitized twice and repeatedly challenged with ovalbumin (OA) under halothane or isoflurane anesthesia. After OA-challenge, in vivo lung function was measured and BAL performed. Peribronchial and peripheral inflammation, cytokine mRNA production and collagen deposition were assessed. Airway hyperresponsiveness, BAL eosinophilia, peripheral lung inflammation, IL-5 mRNA production and collagen deposition were significantly increased in halothane OA-challenged compared to isoflurane OA-challenged mice. Airway challenge induced a higher level of airway hyperresponsiveness, inflammation and remodeling under halothane than isoflurane anesthesia in a murine model of asthma. These differences may be due to increased allergen deposition in the small airways.

Glutathione S‐transferase P1 Ile105Val polymorphism modulates allergen‐induced airway inflammation in human atopic asthmatics in vivo

Glutathione S-transferase P1 is a Phase II cytoprotective and detoxifying enzyme that is widely expressed in human airways. The glutathione S-transferase P1 Ile105Val polymorphism has been linked with atopic disorders and asthma. Yet, little remains known about the regulation of allergic inflammation by glutathione S-transferase P1 in human asthmatics. To establish the effect of the glutathione S-transferase P1 Ile105Val polymorphism on allergen-induced airway inflammation and oxidant stress, and non-specific bronchial hyperresponsiveness to methacholine and reactivity to specific allergen in mild human atopic asthmatics in vivo. Five Val(105)/Val(105) , twelve Val(105)/Ile(105) and twenty Ile(105)/Ile(105) mild atopic asthmatics underwent methacholine challenge, inhaled allergen challenge and endobronchial allergen provocation through a bronchoscope. A panel of inflammatory cytokines and chemokines, F2 -isoprostanes and isofuranes, markers of oxidative stress, thromboxane B2 and immunoglobulin E were measured in bronchoalveolar lavage fluid at baseline and 24 h after allergen instillation. Asthmatics with glutathione S-transferase P1 Val(105)/Val(105) compared with asthmatics with the glutathione S-transferase P1 Val(105)/Ile(105) and Ile(105)/Ile(105) had greater generation of acute phase cytokines (TNF-α, IL-6, CXCL8), IL-12, CCL11, thromboxane B2 and immunoglobulin E at 24 h after local allergen challenge. The GSTP1 genotype had no effect on airway hyperresponsiveness to methacholine and the reactivity to specific allergen. The glutathione S-transferase P1 Ile105Val polymorphism markedly modifies allergen-provoked airway inflammation in atopic asthmatics in vivo. Modulation of the biochemical milieu in response to allergen provides a mechanistic explanation for regulatory effects of glutathione S-transferase P1 polymorphism on airway pathophysiology, and may guide improvement of future therapeutic methods in human atopic asthmatics. These findings must me confirmed in a larger study population of asthmatics with various ethnicities.

Asthmatic Airway Neutrophilia after Allergen Challenge Is Associated with the Glutathione S-Transferase M1 Genotype

Asthma is a heterogeneous lung disorder characterized by airway inflammation and airway dysfunction, manifesting as hyperresponsiveness and obstruction. Glutathione S-transferase M1 (GSTM1) is a multifunctional phase II enzyme and regulator of stress-activated cellular signaling relevant to asthma pathobiology. A common homozygous deletion polymorphism of the GSTM1 gene eliminates enzyme activity. To determine the effect of GSTM1 on airway inflammation and reactivity in adults with established atopic asthma in vivo. Nineteen GSTM1 wild-type and eighteen GSTM1-null individuals with mild atopic asthma underwent methacholine and inhaled allergen challenges, and endobronchial allergen provocations through a bronchoscope. The influx of inflammatory cells, panels of cytokines and chemokines linked to asthmatic inflammation, F(2)-isoprostanes (markers of oxidative stress), and IgE were measured in bronchoalveolar lavage fluid at baseline and 24 hours after allergen instillation. Individuals with asthma with the GSTM1 wild-type genotype had greater baseline and allergen-provoked airway neutrophilia and concentrations of myeloperoxidase than GSTM1-null patients. In contrast, the eosinophilic inflammation was unaffected by GSTM1. The allergen-stimulated generation of acute-stress and proneutrophilic mediators, tumor necrosis factor-α, CXCL-8, IL-1β, and IL-6, was also greater in the GSTM1 wild-type patients. Moreover, post-allergen airway concentrations of IgE and neutrophil-generated mediators, matrix metalloproteinase-9, B-cell activating factor, transforming growth factor-β1, and elastase were higher in GSTM1 wild-type individuals with asthma. Total airway IgE correlated with B-cell activating factor concentrations. In contrast, levels of F(2)-isoprostane were comparable in both groups. Finally, GSTM1 wild-type individuals with asthma required lower threshold concentrations of allergen to produce bronchoconstriction. The functional GSTM1 genotype promotes neutrophilic airway inflammation in humans with atopic asthma in vivo.

55 Ocular Signs and Symptoms Elicited by a Naturalistic Allergen Challenge in an Environmental Exposure Chamber Model Versus a Direct Allergen Instillation Model

BackgroundDirect-instillation ocular models are well established for eliciting allergic responses in research and clinical testing. This study compared direct ocular instillation of allergen to a more naturalistic airborne allergen exposure.MethodsThirteen subjects with histories of ragweed allergy and positive skin prick responses attended screening, dose-finding, dose, confirmation, and analysis study visits. For conjunctival allergen provocation testing (CAPT), 1 drop of ragweed allergen was administered to each eye, at the lowest possible subject-specific concentration between 1.6 and 100 protein nitrogen units per 25 μl drop. For environmental exposure chamber (EEC) testing, subjects were exposed to continual airborne ragweed pollen at 3500 ± 500 particles/m3. Symptoms of itching and tearing were self-assessed on diary cards by subjects. Signs of hyperemia, swelling, and mucous discharge were assessed by clinicians. Assessment time points started at 30 minutes before exposure and continued through 180 minutes after exposure.ResultsAt baseline, there were minimal signs and symptoms. Maximum mean hyperemia with CAPT was 2.3 ± 0.6 units (between moderate and severe) and with EEC was 1.9 ± 0.5 units (approximately moderate); these maxima occurred after 30 minutes with CAPT (rapid spike) and after 180 minutes with EEC (gradual increase). Mean swelling was <1 unit out of 4 units at all times (CAPT and EEC), and mucous discharge was observed in only 1 subject during the study (with CAPT). Maximum mean itching with both CAPT and EEC was 2.8 ± 1.0 units (approximately severe), but this maximum occurred after 20 minutes with CAPT (rapid spike) and after 180 minutes with EEC (gradual increase). Maximum mean tearing with CAPT was 1.2 ± 0.7 units (approximately mild) and with EEC was 1.6 ± 0.6 units (between mild and moderate); these maxima occurred after 15 minutes with EEC (rapid spike) and after 120 minutes with EEC (gradual increase).ConclusionsThe time courses of allergic signs and symptoms differed between CAPT and EEC models; however, both models evoked similar maximum response levels. This demonstrates that the EEC model is a useful challenge model for mimicking natural airborne ocular allergen exposure.

Pulmonary Endotoxin Tolerance Protects against Cockroach Allergen-Induced Asthma-Like Inflammation in a Mouse Model

Background: Compounds which activate the innate immune system, such as lipopolysaccharide, are significant components of ambient air, and extremely difficult to remove from the environment. It is currently unclear how prior inhalation of endotoxin affects allergen sensitization. We examined whether lung-specific endotoxin tolerance induction prior to sensitization can modulate the response to allergen. Methods: Endotoxin tolerance was induced by repeated intratracheal exposure to endotoxin. All mice were then sensitized and challenged by direct intratracheal instillation of cockroach allergen. Results: After allergen sensitization and challenge, endotoxin tolerant mice had significantly decreased airways hyperresponsiveness to methacholine challenge, which was confirmed by invasive lung function tests. Decreased goblet cell hyperplasia and mucus production were also found by histological assessment. Tolerant mice were protected from airway eosinophilia through the mechanism of reduced CCL11 and CCL24. Interestingly, endotoxin tolerant mice had only a modest reduction in cockroach-specific IgE; however, total IgE was significantly reduced. Conclusions: These data show that induction of endotoxin tolerance prior to sensitization protects against the hallmark features of asthma-like inflammation, and that transient modulation of innate immunity can have long-lasting effects on adaptive responses.

Vitamin E prevents NRF2 suppression by allergens in asthmatic alveolar macrophages in vivo

Asthma is a chronic inflammatory airway disease associated with increased generation of reactive oxidant species and disturbed antioxidant defenses. NRF2 is the master transcription factor that regulates the expression of Phase II antioxidant and detoxifying enzymes. Disruption of NRF2 augments oxidative stress and inflammation in a mouse model of asthma, suggesting a protective role for NRF2 in the lungs in vivo. Yet, little is known about the regulation and function of NRF2 in human asthmatics. Using segmental allergen challenge, a well-established experimental model of IgE-mediated asthma exacerbation in human atopic asthmatics, we investigated the effects of a specific allergen and the modulatory role of vitamin E on NRF2 and a NRF2-target gene, superoxide dismutase, in alveolar macrophages recovered from the airways at 24h after allergen instillation in vivo. Allergen-provoked airway inflammation in sensitive asthmatics caused a profound inhibition of macrophage NRF2 activity and superoxide dismutase, rendering them incapable of responding to the NRF2 inducers. Prolonged treatment with high doses of the antioxidant vitamin E lessened this allergen-induced drop in alveolar macrophage NRF2. These results are the first to demonstrate that NRF2 expression in human asthmatics is compromised upon allergen challenge but can be rescued by vitamin E in vivo.

Pulmonary allergic reactions impair systemic vascular relaxation in ragweed sensitive mice

Asthma is often associated with cardiovascular complications, and recent observations in animal models indicate that induction of pulmonary allergic inflammation increases susceptibility of the myocardium to ischemia and reperfusion injury. In this study, we used a murine model of allergen sensitization in which aspiration of allergen induces pulmonary and systemic inflammation, to test the hypothesis that pulmonary exposure to allergen alters vascular relaxation responses. BALB/C mice were sensitized by intraperitoneal injection of ragweed and challenged by intratracheal instillation of allergen. Airway hyperreactivity and pulmonary inflammation were confirmed, and endothelium-dependent and -independent reactivity of thoracic aorta rings were evaluated. Ragweed sensitization and challenge induced airway hyperreactivity to methacholine and pulmonary inflammation, but did not affect constrictor responses of the aortic rings to phenylephrine and K + depolarization. In contrast, maximal relaxation of aortic rings to acetylcholine and sodium nitroprusside decreased from 87.6 ± 3.9% and 97.7 ± 1.2% to 32 ± 4% and 51 ± 6%, respectively ( p < 0.05). The sensitivity to acetylcholine was likewise reduced (EC 50 = 0.26 ± 0.05 μM vs. 1.09 ± 0.16 μM, p < 0.001). The results demonstrate that induction of allergic pulmonary inflammation in mice depresses endothelium-dependent and -independent vascular relaxation, which can contribute to cardiovascular complications associated with allergic inflammation.

Immunomodulatory Effects of CpG Oligodeoxynucleotides on House Dust Mite-Induced Airway Inflammation in Mice

Background: CpG oligodeoxynucleotides (CpG ODNs) are reported to protect against airway eosinophilia and hyperresponsiveness in animal models of asthma. However, little is known about the effects of CpG ODNs on house dust mites, one of the most common environmental allergens, causing allergic asthma. In the present study, we evaluated the immunomodulatory effects of CpG ODNs on the development of house dust mite-induced airway inflammation and remodeling in mice. Methods: Mice were instilled with Dermatophagoides farinae (Der f) into the trachea 8 times without any additional adjuvants. 48 h after the final allergen instillation, the airway responsiveness to acetylcholine (Ach) was measured, and bronchoalveolar lavage (BAL) and histopathological examination were carried out. CpG ODNs were instilled into the trachea mixed with Der f at the first allergen instillation. Results: Repeated instillation of Der f induced increases in airway responsiveness to Ach, the numbers of inflammatory cells, the levels of T-helper type 2 cytokines and transforming growth factor-β₁ in the BAL fluid. Furthermore, goblet cell hyperplasia, the thickness of the epithelium and subepithelial fibrosis were observed. The simultaneous instillation of CpG ODNs with Der f at the first allergen instillation showed significant inhibition of these parameters dose dependently. Conclusions: Our results demonstrate that CpG ODNs have inhibitory effects on Der f-induced airway hyperresponsiveness and eosinophilia, as well as airway remodeling, and that CpG ODNs can be a therapeutic approach for the treatment of house dust mite-induced asthma.

Mediator measurements after daily instillation of allergen: Increased IL-5 and decreased IFN-gamma

This study was designed to measure symptoms, IL-4, IL-5, IFN-gamma, and eosinophilic cationic protein (ECP) in nasal secretions from subjects experiencing an artificial allergy season and to look for evidence of priming. Clinically relevant allergen was administered intranasally out of season to 12 asymptomatic individuals with seasonal allergic rhinitis. These individuals were then randomized to receive allergen or saline daily for the next 7 days. Nasal secretions and scrapings of nasal epithelium were obtained at baseline (day 1), 24 hours after the initial allergen administration (day 2), and 24 hours after the last instillation of allergen or saline (day 9). Nasal symptom scores (p < 0.0002), IL-5 mRNA (p = 0.03), and ECP (p < 0.02) increased after receiving the first challenge (day 2 compared with day 1). In the six subjects randomized to receive seven sequential daily challenges with allergen, symptom scores remained elevated (p < 0.02), IL-5 protein increased (p = 0.02), and IFN-gamma (p = 0.02) levels decreased (day 9 compared with day 1). In the six subjects randomized to receive seven sequential daily challenges with placebo, symptom scores, IL-5, and IFN-gamma levels were not significantly different (day 9 compared with day 1). Compared with the findings at day 2 (n = 12), the treated subjects (n = 6) had no further increase in symptoms but did show a further increase in IL-5 (p = 0.01) and a decrease in IFN-gamma (p = 0.02) at day 9. Daily instillation of moderate doses of allergen intranasally is characterized by persistent symptoms, elevation of IL-5, and reduced levels of IFN-gamma.

Repeated instillations of Dermatophagoides farinae into the airways can induce Th2-dependent airway hyperresponsiveness, eosinophilia and remodeling in mice : Effect of intratracheal treatment of fluticasone propionate

Dermatophagoides farinae are known to be a common environmental allergen causing allergic asthma; however, little is known about their pathophysiological effect via the allergenicities in vivo. Therefore, we first established a mouse model of asthma induced by repeated instillations of D. farinae. Second, to investigate whether the asthmatic responses are Th2-dependent, we examined the effect of the deficiency of interleukin-4 (IL-4) receptor α chain gene. Finally, we examined the effect of fluticasone propionate on this model. Mice were instilled with D. farinae without additional adjuvants into the trachea 8 times. After the final allergen instillation, the airway responsiveness to acetylcholine was measured, and bronchoalveolar lavage and histological examination were carried out. The instillation of the allergen-induced airway hyperresponsiveness, the accumulation of inflammatory cells and increases in the levels of Th2 cytokines and transforming growth factor-β 1 production in the bronchoalveolar lavage fluid dose dependently. The number of goblet cells in the epithelium and the extent of the fibrotic area beneath the basement membrane were also increased in the morphometric study. In contrast, the defect of IL-4/IL-13 signaling through IL-4 receptor α chain completely abrogated all these responses. Furthermore, the simultaneous instillation of fluticasone propionate with the allergen showed significant inhibition or an inhibitory tendency of these changes. These findings demonstrate that the repetitive intratracheal instillations of D. farinae can induce airway remodeling through Th2-type inflammation, and that fluticasone propionate inhibits D. farinae-induced airway remodeling in mice, and this model would be useful for studying mechanisms involved in the development of allergic asthma.

Mast cell inhibitor R112 is well tolerated and affects prostaglandin D2 but not other mediators, symptoms, or nasal volumes in a nasal challenge model of allergic rhinitis

Understanding of the role played by mast cells in allergic rhinitis (AR) has led to the development of novel therapies. The aim of this study was to determine the safety and tolerability of R112, an inhibitor of the tyrosine kinase Syk, in an allergen challenge model of AR. We also examined the effects of R112 on symptoms, mediator release, and nasal airway volumes. This double-blinded, randomized, placebo-controlled, crossover trial enrolled 20 out-of-season volunteers with AR. One intranasal dose of R112 or vehicle was administered and followed by an allergen challenge. In addition to safety monitoring, symptoms; changes in histamine, tryptase, and prostaglandin D2 (PGD2) content of nasal secretions; and acoustic rhinometry were determined over a 15-minute period. R112 was well tolerated. Adverse events were similar between treatments. Five minutes after allergen instillation, PGD2 was decreased when subjects received R112 compared with vehicle (93.4 +/- 23.0 pg/mL versus 171.6 +/- 23.0 pg/mL; p = 0.03), and this correlated with rhinorrhea (p = 0.05). However, at 10 minutes, changes in PGD2, tryptase, and histamine were not significant (46.8 +/- 9.2 pg/mL versus 68.6 +/- 9.2 pg/mL, p = 0.1; 9.5 +/- 2.7 ng/mL versus 16.6 +/- 2.9 ng/mL, p = 0.09; and 1.5 +/- 1.6 ng/mL versus 3.5 +/- 1.6 ng/mL, p = 0.4). No differences were found in symptoms or in acoustic rhinometry between treatment groups. Single-dose R112 appears safe and significantly reduces PGD2 but not histamine or tryptase release in response to allergen challenge in subjects with AR.

Local nitric oxide levels reflect the degree of allergic airway inflammation after segmental allergen challenge in asthmatics

Nitric oxide (NO) levels are increased in the exhaled air of asthmatics. As NO levels correlate with allergic airway inflammation, NO measurement has been suggested for disease monitoring. In patients with asthma, we previously demonstrated that intrabronchial treatment with a natural porcine surfactant enhanced airway inflammation after segmental allergen provocation. We studied whether local levels of NO reflect the degree of allergic airway inflammation following segmental allergen challenge with or without surfactant pretreatment. Segmental NO, as well as nitrite and nitrate in bronchoalveolar lavage (BAL) fluid, was measured before and after segmental challenge with either saline, saline plus allergen, or surfactant plus allergen in 16 patients with asthma and five healthy subjects. The data were compared with inflammatory BAL cells. Segmental NO levels were increased after instillation of saline (p < 0.05), or surfactant plus allergen in asthmatics (p < 0.05), and values were higher after surfactant plus allergen compared to saline challenge. Nitrate BAL levels were not altered after saline challenge but increased after allergen challenge (p < 0.05) and further raised by surfactant (p < 0.05), whereas nitrite levels were not altered by any treatment. Segmental NO and nitrate levels correlated with the degree of eosinophilic airway inflammation, and nitrate levels also correlated with neutrophil and lymphocyte numbers in BAL. In healthy subjects, NO, nitrite, and nitrate were unaffected. Thus, segmental NO and nitrate levels reflect the degree of allergic airway inflammation in patients with asthma. Measurement of both markers can be useful in studies using segmental allergen provocation, to assess local effects of potential immunomodulators.

The role of leukotrienes in allergic rhinitis

To review the role of cysteinyl leukotrienes (cysLTs) in allergic rhinitis and the scientific rationale for therapy with leukotriene receptor antagonists (LTRAs). Relevant basic science and clinical articles were identified by a search of the PubMed database for articles published from 1984 to 2004 using the following keywords: allergic rhinitis; nose; immune response; allergen challenge; leukotrienes C, D, and E; cysteinyl leukotriene; cysteinyl leukotriene receptor; cytokine; leukocyte; montelukast; zafirlukast; and pranlukast. The authors' expert opinion was used to select studies for inclusion in this review. CysLTs are synthesized via 5-lipoxygenase metabolism of arachidonic acid by mast cells and basophils during the early-phase response to antigen and by eosinophils and macrophages during the late phase. The cysLT levels in nasal secretions are elevated after short-term allergen instillation and in allergy season in patients with allergic rhinitis. These lipid mediators act locally and systemically by interacting with receptors, particularly the cysLT1 receptor, on target cells. Evidence derived from topical application of cysLTs in the nose and from the effects of LTRAs indicates that cysLTs contribute to nasal mucous secretion, congestion, and inflammation. CysLTs promote allergic inflammation by enhancing immune responses and the production, adhesion, migration, and survival of inflammatory cells such as eosinophils. They also increase the generation of an array of other proinflammatory mediators, such as cytokines, which in turn increase the production of and receptors for cysLTs. Clinical trials have demonstrated that LTRAs have significant but modest efficacy as single agents but additive efficacy when used with other classes of agents. CysLTs fulfill the criteria for relevant mediators of allergic rhinitis via their diverse effects on immune, inflammatory, and local structural components of disease. By blocking the cysLT1 receptor responsible for most of these effects, LTRAs represent a useful approach to treatment of this important and prevalent disorder.

Le Test de provocation conjonctivale allergénique : recommandations pratiques pour le diagnostic des conjonctivites allergiques. Compte-rendu de la table ronde du Groupe Ophtalmo Allergo (GOA) (Journées parisiennes d’allergie, 10 janvier 2004)

Conjunctival allergen challenge (CAC) is the only way to confirm allergen involvement in the diagnosis of allergic conjunctivitis. Implementation of CAC for diagnostic purposes was the subject of a round table discussion during the last meeting of the Group ophthalmoallergo. The main conclusions of the discussion are reported here. After a brief review of the indications for CAC, the group discussed different aspects of CAC: criteria for the diagnosis of sensitisation; safety considerations; the nature and quality of the allergen extracts used; the possibility of avoiding responsible allergens and drugs; contraindications. A consensus was reached regarding the practice of CAC for the diagnosis of allergic conjunctivitis, the dose escalation protocol for allergen instillation, the clinical as well as the complementary criteria for positivity, and interpretation of the results. The group agreed on the need to consider two distinct patient populations, outpatients and inpatients, according to the CAC technique to be used and the follow-up to be considered. Outpatients usually have benign forms of allergic conjunctivitis (acute and seasonal or chronic) due to common allergens (mainly domestic allergens and aeroallergens). In these patients, CAC can be carried out in a medical environment, with an allergist or an ophthalmologist being present, in which case escalation of the allergen dose instilled in the conjunctivae can be rapid and positivity can be assessed by evaluating the pruritus only. Inpatients, affected mainly by vernal keratoconjunctivitis, will have to undergo a more detailed investigation to identify the triggering factor. In these cases, CAC should be carried out by an ophthalmology service. The clinical and biological criteria for positivity are multiple, and a wider variety of allergens may be tested, using a slower dose escalation. In both types of patients, 20 μL of the allergen are instilled every 30 minutes, and the reaction is assessed 15 minutes after each instillation. Emergency measures, including an oral histamine H1 antagonist, an inhalation bronchodilator, and topical and systemic steroids, must be available. Patients must be observed closely until the procedure is completed. If an ocular reaction occurs, an antihistamine should be instilled locally and the patient followed for an additional two hours. In addition, a topical and an oral antihistamine should be prescribed to be used in case the patient has a conjunctival or systemic reaction after discharge. These national guidelines should help practitioners to standardize CAC. They will be reevaluated in three years.

Eosinophil traffic in the circulation following allergen challenge

Eosinophils contribute to the pathogenesis of asthma and localize to the lung after allergen exposure by uncertain mechanisms. We used intrabronchial instillation of allergen to model the interaction between inhaled allergen and the lung. We measured the number of peripheral blood leukocytes and the expression of VLA-4 (CD49d), Mac-1 (CD11b) and PSGL-1 (CD162) up to 4 h after instillation of allergen into a bronchus of eight atopic asthmatics. For controls, we instilled normal saline into a subset of the asthmatic subjects, and allergen into nonatopic, nonasthmatic subjects. There were changes of total leukocyte number, number of polymorphonuclear leukocytes, lymphocytes, monocytes and eosinophils in all three groups (atopic asthmatics instilled with allergen, atopic asthmatics instilled with saline, nonatopic nonasthmatic subjects instilled with allergen), which were likely related to bronchoscopy. However, the decrease of eosinophils was significant only in the atopic asthmatics instilled with allergen. The remaining eosinophils in the allergen challenged asthmatics were not activated as defined by cell density or change of expression of VLA-4, Mac-1 and PSGL-1. While eosinophils rapidly and specifically leave the circulation after allergen challenge of atopic asthmatics, the remaining circulating eosinophils are not activated.

Mediator measurements during an artificial allergy season: Increased IL-5 and eosinophilic cationic protein (ECP) but no evidence of priming

Rationale The objectives of this study were to measure symptoms, IL4, IL5, IFN-γ, and ECP in subjects experiencing an artificial allergy season and to look for priming by comparing these values with those after a single instillation of allergen. Methods Twelve subjects received nasal instillation of an extract of relevant allergen out of pollen season at a dose that gave moderate symptoms and were then randomized to receive nasal allergen or to receive nasal saline daily over the next 7 days. Scrapings of nasal epithelium and nasal secretions were obtained at baseline, 24 hours after initial allergen administration, and 24 hours after the last instillation of allergen or saline. All subjects kept a log of daily symptoms. Results Subjects receiving nasal allergen extract had an increase in total symptom scores at day 9 compared with baseline (day 1) (p=0.007). IL5 protein was increased in the subjects who received nasal allergen extract compared with the subjects receiving nasal saline instillation (p=0.02); there was a trend towards increase in ECP in the subjects receiving allergen when compared with those receiving placebo ( p=.078). However, symptom scores, IL5 levels, and ECP levels were similar in subjects receiving nasal allergen at days 2 and 9. Conclusions An artificial allergy season is characterized by persistent symptoms and elevations of both IL5 and ECP. When symptom scores and mediator levels at day 2 and day 9 in the active allergen group were compared, we did not detect a priming effect due to daily instillation of allergen.

Cholinergic responsiveness of the individual airway after allergen instillation in sensitised pigs

Allergen exposure of sensitised lungs produces bronchial hyperresponsiveness in vivo associated with airway inflammation and remodelling. It is unclear if hyperresponsiveness is also present in airways in vitro under similar conditions of drug provocation as carried out in vivo, and at different times after allergen-challenge. This study records responsiveness of individual airway segments to acetylcholine (ACh) in sensitised bronchi after instillation of allergen (ovalbumin, OA). Airway histology and sensitivity and maximum effects to ACh were recorded 1, 24 and 72 h and 1 week after OA. OA-instilled airways exhibited eosinophilia and epithelial proliferation. Physiological recordings showed no change in maximum contractions of airway segments to acetylcholine placed in the airway lumen except at 24 h where they were reduced. In contrast maximum contractions to ACh to the airway adventitia were reduced at all times except 1 week, with the greatest change occurring at 24 h. There were no changes in airway sensitivity to either route of ACh in OA-instilled airways but the difference in sensitivity to adventitial and lumenal ACh was reduced. Results show that allergen does not produce hyperresponsiveness at the airway wall but it may alter an interaction between airway smooth muscle and other structural components of the airway.

Transient neutrophil infiltration after allergen challenge is dependent on specific antibodies and Fc gamma III receptors.

Following allergen challenge of sensitized mice, neutrophils are the first inflammatory cells found in bronchoalveolar lavage (BAL) fluid. To determine the underlying mechanism for their accumulation, mice were sensitized to OVA on days 0 and 14, and received, on day 28, a single intranasal challenge (s.i.n.) with either OVA or ragweed. Eight hours after the s.i.n., BAL fluid was obtained. BALB/c mice sensitized and challenged with OVA showed significantly higher total cell counts and numbers of neutrophils in BAL fluid compared to the OVA-sensitized and ragweed-challenged or nonsensitized mice. Levels of neutrophil chemokines in BAL fluid supernatants were markedly elevated in the sensitized and OVA-challenged mice; Fc epsilon RI-deficient mice showed comparable numbers of neutrophils and neutrophil chemokines in BAL fluid after s.i.n. But in sensitized mice lacking the Fc common gamma-chain and B cell-deficient mice, the number of neutrophils and levels of neutrophil chemokines in BAL fluid were significantly lower. Further, mice lacking the FcgammaRIII did not develop this early neutrophil influx. Neutrophil infiltration could be induced in naive mice following intranasal instillation of allergen combined with allergen-specific IgG1. In addition, macrophages from sensitized mice were stimulated with allergen and activated to produce neutrophil chemokines. These results demonstrate that neutrophil influx after allergen challenge requires prior sensitization, is allergen-specific, is mediated through FcgammaRIII, and is dependent on the presence of Ab.

Role of matrix metalloproteinases in asthma.

Airway inflammation and remodeling are key features of asthma. Matrix metalloproteinases (MMPs) and their inhibitors, tissue inhibitors of metalloproteinases (TIMPs) are thought to contribute to the pathogenesis of asthma via their influence on the function and migration of inflammatory cells as well as matrix deposition and degradation. TIMPs bind MMPs in a 1:1 fashion. Thus, an increase in the molar ratio of MMP/TIMP may favor tissue injury, while the reverse could be associated with increased fibrosis. MMP-9 is the predominant MMP in asthma, and its expression is enhanced when patients have spontaneous exacerbations or in response to local instillation of allergen in the airway. As acute inflammation resolves, MMP-9 levels return toward normal. Interestingly, corticosteroids downregulate MMP and enhance TIMPs. Even though it is clear that enhanced airway inflammation in asthma is associated with increased expression of MMPs, whether specific inhibitors of MMP could reduce airway injury and facilitate orderly healing in asthma is still unknown.