Late-phase Airway Research Articles

The Impact of Genetic variation at TNF-α -308 G/A on their serum production and severity of Asthma disease

Asthma is a chronic respiratory disease leading to morbidity and impaired quality of life worldwide. TNF-α cytokine responsible for the smooth muscle activation and late-phase airway hyper responsiveness. So this study aimed to investigate TNF-α polymorphisms situated at positions –308 G/A with their serum level and asthma severity. A case–control study included 40 asthmatic patients 16 females and 24 males and matched with age and gender 40 healthy subjects as a control group. Blood samples were collected from both groups, genomic DNA was extracted from peripheral blood for further molecular study. The study appeared that TNF-α-308 G/A significantly (P<0.05) high frequencies of homozygous AA, and A allele carrier in asthmatic patients, in contrast homozygous GG genotype and G allele appear at low frequency with patients. The data showed that genetic variants of the TNF-α affect the TNF-α production and associated with the severity of asthma, AA and AG genotype associated with high serum level (58.2 pg/ml and 49.5 pg/ml) respectively and GG genotype associated with low serum level (24.06pg/ml).The study confirmed the distinct role of TNF-α in cases of asthma compared to healthy controls with significantly higher level among patients, mean serum level for patients and control were (50.3 and 5.88 ) pg/ml respectively. Also, mean serum levels of TNF-α were significantly higher in asthmatic patients that suffering from allergic rhinitis(65.39 pg/ml) compared to non-rhinitis patients (34.97pg/ml). In conclusion, TNF-α-308 GG, genotype contribute to the predisposition of asthma and there was significant association among genotype of this gene and level of cytokine as well as development of the disease. Also observed an association between this gene and serum level of cytokine. AA, AG genotype and allergic rhinitis consider a risk factor of asthma and progress the disease.

Bronchial Allergen Challenge of Patients with Atopic Asthma Triggers an Alarmin (IL-33, TSLP, and IL-25) Response in the Airways Epithelium and Submucosa

The alarmin cytokines IL-25, IL-33, and thymic stromal lymphopoietin (TSLP) play a critical role in asthma pathogenesis by inducing mucosal Th2-type cytokine production. Although environmental exposure to aeroallergens has been proposed as an alarmin trigger in asthma, there has been no systematic parallel study of the effects of allergen exposure on the expression of these cytokines in the airways of human asthmatics. Using single and sequential double immunohistochemistry, we evaluated the numbers and phenotypes of IL-25-, IL-33-, and TSLP-immunoreactive cells in sections of bronchial biopsies from mild atopic asthmatics (n = 16) before and 24 h after allergen inhalational challenge. Allergen challenge highly increased expression of baseline immunoreactivity for IL-25, IL-33, and TSLP, both in the bronchial epithelium and submucosa (p < 0.001), to a degree that correlated with the extent of the late phase of airway obstruction. Aside from epithelial cells, the principal source of immunoreactivity for all three alarmins, TSLP, and IL-33 immunoreactivity colocalized principally with endothelial cells and mast cells, neutrophils, and fibroblasts, whereas IL-25 immunoreactivity colocalized principally with eosinophils as well as endothelial cells, mast cells, and fibroblasts. The data implicate that allergen challenge directly increases airway alarmin expression in atopic asthmatics to a degree correlating with increase late-phase airway obstruction, affirming these molecules as potential molecular targets for the inhibition of allergen-induced airway inflammation and obstruction.

P-STAT6, PU.1, and NF-κB are involved in allergen-induced late-phase airway inflammation in asthma patients.

BackgroundPrevious in vitro and animal studies demonstrated that transcription factors p-STAT6 and PU.1 are required to induce interleukin (IL)-9 secretion by T helper (Th) 9 cells. It is believed that n factor-kappaB (NF-κB) plays a role in eosinophil survival. The importance of these transcription factors in the pathogenesis of allergic asthma (AA) in humans is poorly understood. We evaluated p-STAT6 and PU.1 expression in peripheral blood Th9 cells and NF-κB expression in eosinophils during late-phase airway inflammation in AA patients.MethodsNineteen adults with AA and 14 adult healthy individuals (HI) were examined. Peripheral blood collected 24 h before (baseline) and 24 h after bronchial allergen challenge. CD4+ cells and eosinophils were isolated by high-density gradient centrifugation and magnetic separation. The percentage of Th9 cells and apoptotic eosinophils was estimated by flow cytometry. p-STAT6 and PU.1 expression was expressed as mean fluorescence intensity (MFI) in Th9 cells. NF-κB levels were expressed as MFI in peripheral blood eosinophils. Serum IL-9 and IL-5 levels were determined by enzyme-linked immunosorbent assay.ResultsAt baseline, MFI of p-STAT6 and PU.1 in peripheral blood Th9 cells and MFI of NF-κB in eosinophils and, serum IL-5 and IL-9 levels were greater in AA patients (P < 0.05). Decreased eosinophil apoptosis was seen in the AA group compared with HI (P < 0.05). MFI of p-STAT6, PU.1, and NF-κB and serum levels of IL-5 and IL-9 were increased in the AA group 24 h after challenge compared with baseline (P < 0.05). In the AA group, a correlation between serum IL-9 and Th9 cells (r = 0.7, P = 0.001) and MFI of PU.1 (r = 0.6, P = 0.01) 24 h after bronchial allergen challenge was observed. A correlation between Th9 cells and MFI of p-STAT6 (r = 0.45, P = 0.03) as well as MFI of PU.1 (r = 0.5, P = 0.02) 24 h after challenge was only observed in AA patients. A correlation between the MFI of NF-κB and eosinophil apoptosis was observed in AA patients 24 h before (r = −0.46, P = 0.02) and after (r = −0.5, P = 0.02) challenge.Discussionsp-STAT6 and PU.1 may be associated with Th9 cells and IL-9 production, whereas NF-κB and IL-5 may be associated with reduced eosinophil apoptosis in allergen-induced late-phase airway inflammation.Trial registrationClinicalTrials.gov NCT02214303Electronic supplementary materialThe online version of this article (doi:10.1186/s12890-015-0119-7) contains supplementary material, which is available to authorized users.

Functional activity of peripheral blood eosinophils in allergen-induced late-phase airway inflammation in asthma patients.

ObjectiveWe aimed to investigate peripheral blood eosinophil chemotaxis, generation of spontaneous reactive oxygen species (ROS), and apoptosis in patients with allergic asthma after bronchial allergen challenge.Material and methodsA total of 18 patients with allergic asthma (AA), 14 with allergic rhinitis (AR), and 10 healthy subjects (HS) underwent bronchial challenge with a specific allergen extract. Eosinophils from peripheral blood were isolated 24 h before as well as 7 and 24 h after bronchial allergen challenge. Chemotaxis, spontaneous ROS production in eosinophils, and apoptosis were analyzed by flow cytometry. Serum and induced sputum IL-5 levels were measured by ELISA; the cell count in sputum was analyzed by the May-Grünwald-Giemsa method.ResultsBefore bronchial allergen challenge, peripheral blood eosinophil chemotaxis, spontaneous ROS production was enhanced and eosinophil apoptosis was reduced in the patients with AA as compared with AR patients and HS (P < 0.05). Meanwhile, eosinophil chemotaxis and ROS generation markedly increased in the patients with AA 7 h and 24 h after challenge compared with other groups and baseline values (P < 0.05). The percentage of apoptotic eosinophils in the patients with AA decreased at 7 h as well as 24 h after challenge when compared with other groups and the baseline values (P < 0.05). There was a significant correlation between the migrated peripheral blood eosinophil count and the sputum eosinophil count (Rs = 0.89, P < 0.0001) and the sputum IL-5 level (Rs = 0.68, P = 0.002) at 24 h after bronchial challenge only in the patients with AA. Furthermore, the percentage of peripheral blood apoptotic eosinophils significantly correlated with eosinophil count in sputum (Rs = −0.53, P = 0.02), and ROS production correlated with the serum IL-5 levels (Rs = 0.71, P = 0.01).ConclusionDuring allergen-induced late-phase airway inflammation, peripheral blood eosinophils demonstrated further alterations of their functional activity manifested by enhanced spontaneous ROS production, increased chemotaxis, and diminished apoptosis in patients with AA.

17-­o tipo T limfocitų pagalbininkų reikšmė vėlyvosios fazės kvėpavimo takų uždegimui sergant alergine astma

2014 metų rugpjūčio 27 d. Lietuvos sveikatos mokslų universiteto Medicinos akademijos (LSMU MA) viešame Medicinos mokslo krypties tarybos posėdyje Ieva Bajoriūnienė apgynė daktaro mokslo laipsnio disertaciją 17-o tipo T limfocitų pagalbininkų vaidmuo sergant alergine astma. Mokslinis vadovas – prof. dr. Kęstutis Malakauskas. Disertacija rengta 2009–2014 metais Lietuvos sveikatos mokslų universiteto Medicinos akademijos Pulmonologijos ir imunologijos klinikoje.

Sputum neutrophil count after bronchial allergen challenge is related to peripheral blood neutrophil chemotaxis in asthma patients.

The aim of this study was to estimate relations between sputum neutrophilia and the chemotactic activity of peripheral blood neutrophils after the bronchial allergen challenge in asthma patients. Fifteen patients with allergic asthma (AA), 13 patients with allergic rhinitis (AR), all sensitized to Dermatophagoides pteronyssinus, and 8 healthy subjects (HS) underwent bronchial challenge with D. pteronyssinus. Sputum and peripheral blood collection were performed 24h before, 7 and 24h after the bronchial challenge. Cell counts were determined by the May-Grünwald-Giemsa method. Neutrophil chemotaxis was analyzed by a flow cytometer; IL-8 levels were measured by ELISA. Sputum neutrophil count and peripheral blood neutrophil chemotaxis of patients with AA were greater 7 and 24h after the challenge compared with the baseline values and patients with AR and HS (P<0.05). Moreover, a significant correlation was found between the neutrophil count in sputum and IL-8 levels, and the chemotactic activity of peripheral blood neutrophils 24h after the bronchial challenge only the patients with AA (P<0.05). Increased sputum neutrophil count was found to be associated with an enhanced chemotactic activity of peripheral blood neutrophils during allergen-induced late-phase airway inflammation in patients with allergic asthma.

IL-17A Promotes the Exacerbation of IL-33–Induced Airway Hyperresponsiveness by Enhancing Neutrophilic Inflammation via CXCR2 Signaling in Mice

Neutrophilic airway inflammation is a hallmark of patients with severe asthma. Although we have reported that both IL-33 and IL-17A contributed to IgE-mediated neutrophilic inflammation in mice, the relationship remains unclear. In this article, we examined how IL-17A modifies IL-33-induced neutrophilic inflammation and airway hyperresponsiveness (AHR). IL-33 was intratracheally administered to BALB/c mice on days 0-2; furthermore, on day 7, the effect of the combination of IL-33 and IL-17A was evaluated. Compared with IL-33 or IL-17A alone, the combination exacerbated neutrophilic inflammation and AHR, associated with more increased levels of lung glutamic acid-leucine-arginine(+) CXC chemokines, including CXCL1, CXCL2, and CXCL5, and infiltration by alveolar macrophages expressing CXCR2. Treatment with anti-CXCR2 mAb or depletion of alveolar macrophages repressed neutrophilic inflammation and AHR; in addition, depletion of neutrophils suppressed AHR. These findings prompted us to examine the role of CXCR2 in IgE-sensitized mice; a single treatment with anti-CXCR2 mAb in the seventh Ag challenge inhibited late-phase airway obstruction, AHR, and neutrophilic inflammation. In addition to inhibition, multiple treatments during the fourth to seventh challenge attenuated early-phase airway obstruction, eosinophilic inflammation, and goblet cell hyperplasia associated with the reduction of Th2 cytokine production, including IL-4, IL-5, and IL-13. Collectively, IL-33 cooperated with IL-17A to exacerbate AHR by enhancing neutrophilic inflammation via CXCR2 signaling; furthermore, CXCR2 signaling derived Th2 responses. We thus suggest the underlying mechanisms of IL-33 and IL-17A in allergic asthma and CXCR2 as potential therapeutic targets for the disease.

Th17 response to Dermatophagoides pteronyssinus is related to late-phase airway and systemic inflammation in allergic asthma

Th17 cells may play a role in the development of late-phase allergen-induced airway and systemic inflammation in allergic asthma, although the mechanisms involved remain to be elucidated. A total of 36 subjects were enrolled into the study: 15 allergic asthma patients with early asthmatic reaction (n=7) or dual asthmatic reaction (n=8) developed to inhaled D. pteronyssinus, 13 patients with allergic rhinitis, and 8 healthy subjects. Peripheral blood and induced sputum were collected 24h before as well as 7h and 24h after a bronchial challenge with D. pteronyssinus. Th17 cells were analyzed by FACS; IL-17 levels were determined by ELISA. At baseline, the percentage of peripheral blood Th17 cells and serum and sputum IL-17 levels were significantly higher in all groups of studied patients compared with those of healthy subjects. After the bronchial challenge, there was a significant increase in the percentage of peripheral blood Th17 cells and in serum and sputum IL-17 levels in rhinitis and asthma patients compared with their baseline values, particularly in allergic asthma patients with the dual asthmatic reaction. Positive correlations were found between the percentage of Th17 cells and IL-17 levels in serum (Rs=0.649; P=0.009) as well in sputum (Rs=0.583; P=0.022) in allergic asthma patients 24h after the bronchial challenge. The Th17 response is associated with the development of late-phase airway and systemic inflammation after the inhalation of D. pteronyssinus in patients with allergic asthma.

Roles of basophils and mast cells infiltrating the lung by multiple antigen challenges in asthmatic responses of mice

Mast cell hyperplasia has been observed in the lungs of mice with experimental asthma, but few reports have studied basophils. Here, we attempted to discriminate and quantify mast cells and basophils in the lungs in a murine asthma model, determine if both cells were increased by multiple antigen challenges and assess the roles of those cells in asthmatic responses. Sensitized Balb/c mice were intratracheally challenged with ovalbumin four times. Mast cells and basophils in enzymatically digested lung tissue were detected by flow cytometry. An anti-FcεRI monoclonal antibody, MAR-1, was i.p. administered during the multiple challenges. The numbers of both mast cells (IgE(+) C-kit(+) ) and basophils (IgE(+) C-kit(-) CD49b(+) ) increased in the lungs after three challenges. Treatment with MAR-1 completely abolished the increases; however, a late-phase increase in specific airway resistance (sRaw), and airway eosinophilia and neutrophilia were not affected by the treatment, although the early-phase increase in sRaw was suppressed. MAR-1 reduced antigen-induced airway IL-4 production. Basophils infiltrating the lung clearly produced IL-4 after antigen stimulation in vitro; however, histamine and murine mast cell protease 1 were not increased in the serum after the challenge, indicating that mast cell activation was not evoked. Both mast cells and basophils infiltrated the lungs by multiple intratracheal antigen challenges in sensitized mice. Neither mast cells nor basophils were involved in late-phase airway obstruction, although early-phase obstruction was mediated by basophils. Targeting basophils in asthma therapy may be useful for an early asthmatic response.

Erratum to: Peripheral Blood Neutrophil Activity During Dermatophagoides pteronyssinus-Induced Late-Phase Airway Inflammation in Patients with Allergic Rhinitis and Asthma

Erratum to: Peripheral Blood Neutrophil Activity During Dermatophagoides pteronyssinus-Induced Late-Phase Airway Inflammation in Patients with Allergic Rhinitis and Asthma

Complement C3a-Induced IL-17 Plays a Critical Role in an IgE-Mediated Late-Phase Asthmatic Response and Airway Hyperresponsiveness via Neutrophilic Inflammation in Mice

Allergen-specific IgE plays an essential role in the pathogenesis of allergic asthma. Although there has been increasing evidence suggesting the involvement of IL-17 in the disease, the relationship between IL-17 and IgE-mediated asthmatic responses has not yet been defined. In this study, we attempted to elucidate the contribution of IL-17 to an IgE-mediated late-phase asthmatic response and airway hyperresponsiveness (AHR). BALB/c mice passively sensitized with an OVA-specific IgE mAb were challenged with OVA intratracheally four times. The fourth challenge caused a late-phase increase in airway resistance associated with elevated levels of IL-17(+)CD4(+) cells in the lungs. Multiple treatments with a C3a receptor antagonist or anti-C3a mAb during the challenges inhibited the increase in IL-17(+)CD4(+) cells. Meanwhile, a single treatment with the antagonist or the mAb at the fourth challenge suppressed the late-phase increase in airway resistance, AHR, and infiltration by neutrophils in bronchoalveolar lavage fluid. Because IL-17 production in the lungs was significantly repressed by both treatments, the effect of an anti-IL-17 mAb was examined. The late-phase increase in airway resistance, AHR, and infiltration by neutrophils in bronchoalveolar lavage fluid was inhibited. Furthermore, an anti-Gr-1 mAb had a similar effect. Collectively, we found that IgE mediated the increase of IL-17(+)CD4(+) cells in the lungs caused by repeated Ag challenges via C3a. The mechanisms leading to the IgE-mediated late-phase asthmatic response and AHR are closely associated with neutrophilic inflammation through the production of IL-17 induced by C3a.

Peripheral Blood Neutrophil Activity During Dermatophagoides pteronyssinus-Induced Late-Phase Airway Inflammation in Patients with Allergic Rhinitis and Asthma

Recent investigations suggest that neutrophils may play an important role in the late-phase allergen-induced inflammation in allergic airway diseases. The aim of this study was to evaluate neutrophil chemotaxis, phagocytic activity, and reactive oxygen species (ROS) production in patients with allergic rhinitis and asthma challenged with inhaled Dermatophagoides pteronyssinus. Eighteen patients with allergic rhinitis and 14 with allergic asthma, all sensitized to D. pteronyssinus, as well as 15 healthy individuals underwent bronchial challenge with D. pteronyssinus. Peripheral blood collection and neutrophil isolation were performed 24 h before as well as 7 and 24 h after bronchial challenge. Neutrophils chemotaxis, phagocytic activity, and ROS production were analyzed by flow cytometer. Neutrophil chemotaxis and ROS production were increased, while phagocytic activity was decreased 24 h before challenge in patient groups compared with healthy individuals. After challenge, neutrophil chemotaxis and phagocytic activity increased after 7 and 24 h, when ROS production, only after 24 h. Bronchial allergen challenge had no influence for neutrophils activity in healthy subjects. Activated chemotaxis, phagocytic activity, and ROS production of peripheral blood neutrophils after challenge with D. pteronyssinus reflect an enhanced systemic inflammation in allergic rhinitis and asthma patients with induced late-phase airway inflammation.

Preclinical Evaluation of an Inhibitor of Cytosolic Phospholipase A2α for the Treatment of Asthma

Asthma is a chronic inflammatory lung disease with considerable unmet medical needs for new and effective therapies. Cytosolic phospholipase A(2)α (cPLA(2)α) is the rate-limiting enzyme that is ultimately responsible for the production of eicosanoids implicated in the pathogenesis of asthma. We investigated a novel cPLA(2)α inhibitor, PF-5212372, to establish the potential of this drug as a treatment for asthma. PF-5212372 was a potent inhibitor of cPLA(2)α (7 nM) and was able to inhibit prostaglandin (PG)D(2) and cysteinyl leukotriene release from anti-IgE-stimulated human lung mast cells (0.29 and 0.45 nM, respectively). In a mixed human lung cell population, PF-5212372 was able to inhibit ionomycin-stimulated release of leukotriene B(4), thromboxane A(2), and PGD(2) (2.6, 2.6, and 4.0 nM, respectively) but was significantly less effective against PGE(2) release (>301 nM; p < 0.05). In an in vitro cell retention assay, PF-5212372 retained its potency up to 24 h after being washed off. In a sheep model of allergic inflammation, inhalation of PF-5212372 significantly inhibited late-phase bronchoconstriction (78% inhibition; p < 0.001) and airway hyper-responsiveness (94% inhibition; p < 0.001), and isolated sheep lung mast cell assays confirmed species translation via effective inhibition of PGD(2) release (0.78 nM). Finally, PF-5212372 was assessed for its ability to inhibit the contraction of human bronchi induced by AMP. PF5212372 significantly inhibited AMP-induced contraction of human bronchi (81% inhibition; p < 0.001); this finding, together with the ability of this drug to be effective in a wide range of preclinical asthma models, suggests that inhibition of cPLA(2)α with PF-5212372 may represent a new therapeutic option for the treatment of asthma.

Effects of Interleukin-13 Blockade on Allergen-induced Airway Responses in Mild Atopic Asthma

Extensive evidence in animal models supports a role for IL-13 in the pathobiology of asthma. IMA-638 and IMA-026 are fully humanized IgG(1) antibodies that bind to different epitopes and neutralize IL-13 bioactivity. We hypothesized that anti-IL-13 treatment would inhibit allergen-induced late-phase asthmatic responses, airway hyperresponsiveness, and inflammation in subjects with asthma. Fifty-six subjects with mild, atopic asthma were recruited for two double-blind, randomized, placebo-controlled, parallel group trials to compare IMA-638 and IMA-026 IL-13 antibody treatments with placebo treatment. Drug was administered on Days 1 and 8, and allergen challenges were performed on Days 14 and 35. The primary outcome variable was the late-phase area under the curve (AUC), and secondary outcome variables were the early- and late-phase maximum percent fall in FEV(1), early AUC, allergen-induced shift in airway hyperresponsiveness, and sputum eosinophils. The treatment difference with IMA-638 on Day 14 was -19.1 FEV(1) × hour (95% confidence interval: -36.2, -1.9) for the allergen-induced early AUC and -23.8 FEV(1) × hour (95% confidence interval: -46.4, -1.2) for the late AUC (both P < 0.05), but this effect was lost by Day 35. Treatment with IMA-026 did not attenuate the asthmatic responses on Day 14 or Day 35. There was no effect of either antibody on allergen-induced airway hyperresponsiveness or sputum eosinophils. The frequency of adverse events after administration of the IL-13 antibodies was similar to placebo. IL-13 has a role in allergen-induced airway responses in humans. Further study is required to determine whether anti-IL-13 monoclonal antibodies will be beneficial clinically.

Pharmacological Characterization of MK-7246, a Potent and Selective CRTH2 (Chemoattractant Receptor-Homologous Molecule Expressed on T-Helper Type 2 Cells) Antagonist

The chemoattractant receptor-homologous molecule expressed on T-helper type 2 cells (CRTH2) is a G protein-coupled receptor that has been reported to modulate inflammatory responses in various rodent models of asthma, allergic rhinitis and atopic dermatitis. In this study, we describe the biological and pharmacological properties of {(7R)-7-[[(4-fluorophenyl)sulfonyl](methyl)amino]-6,7,8,9-tetrahydropyrido[1,2-a]indol-10-yl}acetic acid (MK-7246), a novel synthetic CRTH2 antagonist. We show that MK-7246 1) has high affinity for the human, monkey, dog, rat, and mouse CRTH2, 2) interacts with CRTH2 in a reversible manner, 3) exhibits high selectivity over all prostanoid receptors as well as 157 other receptors and enzymes, 4) acts as a full antagonist on recombinant and endogenously expressed CRTH2, 5) demonstrates good oral bioavailability and metabolic stability in various animal species, 6) yields ex vivo blockade of CRTH2 on eosinophils in monkeys and sheep, and 7) significantly blocks antigen-induced late-phase bronchoconstriction and airway hyper-responsiveness in sheep. MK-7246 represents a potent and selective tool to further investigate the in vivo function of CRTH2.

Changes in asthma‐like responses after extended removal from exposure to trimellitic anhydride in the Brown Norway rat model

Organic acid anhydride-induced occupational asthma is considered to be IgE-mediated. Airway and skin exposure are the two main routes of sensitization in the work place. Recently we developed an allergic asthmatic Brown Norway rat model sensitized by dermal exposure to trimellitic anhydride (TMA) using an occlusion patch application. The objectives of this study were (1) to develop a model of non-occluded dermal exposure leading to allergic sensitization and (2) to examine the effect of extended removal from exposure on persistence of both specific IgE and TMA aerosol-induced airway responses in this model. TMA powder (4 or 40 mg) was applied, unoccluded, to the skin of rats for 4 h, once/week for 4 weeks. Rats were given a 10-min aerosol challenge to 40 mg/m(3) TMA 2 weeks after the last dermal exposure (day 35). Another group was challenged on day 35 and again 18-24 months later. Respiratory enhanced pause (Penh), pulmonary histopathology and inflammation and specific IgE titres were measured. Rats produced dose-dependent specific IgE titres after exposure and developed early-phase (EAR) and late-phase airway responses (LAR) after airway challenge to TMA aerosol as well as airway eosinophilic inflammation. Specific airway responses were still manifested after a second TMA airway challenge given 18-24 months following the initial airway challenge. While persistent, airway inflammation, specific IgE and EAR were significantly attenuated following the second TMA challenge. LAR remained robust at 18-24 months and was not significantly different from the response on day 35. These results demonstrate the persistence of chemical sensitization and further suggest that IgE is not essential for LAR.

Rethinking the Pathogenesis of Asthma

Asthma research has focused primarily on allergic pathways on the basis that the majority of asthma is associated with atopy, the recruitment of the Th2-type T cell, the cytokines, and the chemokines that are released on exposure to allergens, and the IgE pathway. However, despite considerable investment by industry, targeting these pathways has not resulted in new treatments being developed beyond blockade of cysteinyl leukotrienes and IgE and improvements in inhaled corticosteroids and beta(2)-adrenoceptor bronchodilators. Increasingly, it is recognized that asthma is a heterogeneous disorder, and while important, allergen sensitization is only one component of the disease, with many other environmental and genetic factors playing a role. In addition, these factors act locally on a susceptible airway epithelium that is both structurally and functionally deficient. It may be worthwhile to focus on increasing the resilience of the airways to environmental insults in addition to improving strategies that modify adaptive immunity or suppress inflammation.

Complement C3a Regulates Late Asthmatic Response and Airway Hyperresponsiveness in Mice

Allergic asthma is a chronic inflammatory disorder of the airways characterized by biphasic airway obstruction and airway hyperresponsiveness. In this study, we attempted to elucidate the contribution of the complement C3a to these asthmatic symptoms. BALB/c mice sensitized by i.p. injections of OVA plus alum were challenged with OVA intratracheally four times. The fourth challenge caused a biphasic asthmatic response peaking at 10 min and 3-4 h, as well as airway hyperresponsiveness to methacholine. Histological examination revealed increased expression of C3a receptors in the lung on the fourth challenge. Additionally, the C3 level in serum 4 h after the fourth challenge was significantly reduced compared with that before the challenge. When a C3a receptor antagonist, SB290157, was administered i.p. 30 min before the fourth challenge, the late-phase asthmatic response and airway hyperresponsivness induced by the fourth challenge were significantly inhibited, although the early-phase response was not influenced. In bronchoalveolar lavage fluid, neutrophil infiltration 24 h after the fourth challenge was reduced by the treatment. On the other hand, SB290157 suppressed the increased expression of IL-1beta in the lung in this model, and the intratracheal administration of IL-1beta induced airway obstruction, airway hyperresponsiveness, and neutrophil infiltration in normal mice. These results illustrate that C3a is involved in the development of the late asthmatic response and airway hyperresponsiveness. The mechanism leading to the development of these symptoms may correlate with the recruitment of neutrophils and/or the production of IL-1beta induced by C3a.

Specific inhibition of tissue kallikrein 1 with a human monoclonal antibody reveals a potential role in airway diseases

KLK1 (tissue kallikrein 1) is a member of the tissue kallikrein family of serine proteases and is the primary kinin-generating enzyme in human airways. DX-2300 is a fully human antibody that inhibits KLK1 via a competitive inhibition mechanism (Ki=0.13 nM). No binding of DX-2300 to KLK1 was observed in a surface-plasmon-resonance biosensor assay when KLK1 was complexed to known active-site inhibitors, suggesting that DX-2300 recognizes the KLK1 active site. DX-2300 did not inhibit any of the 21 serine proteases that were each tested at a concentration of 1 microM. We validated the use of DX-2300 for specific KLK1 inhibition by measuring the inhibition of KLK1-like activity in human urine, saliva and bronchoalveolar lavage fluid, which are known to contain active KLK1. In human tracheobronchial epithelial cells grown at the air/liquid interface, DX-2300 blocked oxidative-stress-induced epidermal-growth-factor receptor activation and downstream mucus cell proliferation and hypersecretion, which have been previously shown to be mediated by KLK1. In an allergic sheep model of asthma, DX-2300 inhibited both allergen-induced late-phase bronchoconstriction and airway hyper-responsiveness to carbachol. These studies demonstrate that DX-2300 is a potent and specific inhibitor of KLK1 that is efficacious in in vitro and in vivo models of airway disease.

Montelukast as Add-on Therapy to β-Agonists and Late Airway Response

Rosewich M, Rose MA, Eickmeier O, Travaci M, Kitz R, Zielen S. Eur Respir J. 2007;30(1):56–61 PURPOSE OF THE STUDY. To evaluate montelukast's ability to inhibit the late-phase airway response after allergen exposure in adults with house dust mite–induced asthma. STUDY POPULATION. Thirty-five adults (19 men, 16 women) aged 18 to 31 with mild, stable asthma and sensitivity to house dust mites were included. METHODS. This study was designed as a randomized, double-blind, single-dose, placebo-controlled, crossover trial. Subjects underwent serum testing for antigen-specific immunoglobulin E (IgE) to house dust mite. Sensitized subjects were then examined for airway infections and underwent spirometry and determination of their fraction of nitric oxide in exhaled air (FeNO). They were then challenged with increasing concentrations of inhaled Dermatophagoides farinae via a nebulizer system. Testing was stopped when the patient's forced expiratory volume in 1 second (FEV1) decreased at least 20% (early airway response [EAR]), they experienced significant clinical symptoms, or they received a cumulative dose of 1270 mg. Patients were then given 1 puff of salbutamol (0.1 mg) and either montelukast (10 mg) or a placebo. FEV1 was measured for the following 8 hours with a decrease of at least 20% demonstrating a late airway response (LAR). Formoterol (12 μg) was then given to treat those patients. Subjects returned at least 2 weeks later and were crossed-over into the other group. RESULTS. Of the 35 subjects in the study, 12 showed no significant EAR on 1 or both study days, 11 showed only an EAR, and 12 demonstrated both an EAR and LAR. This last group was analyzed further. The difference in FEV1 from baseline values 3 to 8 hours after challenge was expressed as the area under the FEV1 time-response curve (FEV1-AUC). The FEV1-AUC of patients on placebo was −2.47 ± 1.32 vs −0.768 ± 1.68 for the patients on montelukast (P &amp;lt; .005). Subjects with a dual response had a significantly higher baseline FeNO than those with no response (56.4 vs 21.0 ppb; P &amp;lt; .05). CONCLUSIONS. Montelukast was able to block the late-phase airway response in subjects who responded dually to the allergen challenge. In addition, patients with a higher baseline FeNO seemed more likely to develop an LAR than those with lower ones. REVIEWER COMMENTS. The results of this study implicate a role for montelukast as an “add-on,” therapeutic option for symptomatic relief after allergen exposure in subjects with mild asthma. It also suggests a rapid onset of action that allows for its usage in such a setting. This study supports existing ones that have demonstrated the effect of montelukast within 2 to 3 hours for up to 24 hours. There was also a “trend toward significance” showing subjects on montelukast with higher FEV1 values than those on placebo. However, this study was limited in its sample size and should be expanded to fully elicit montelukast's role in acute exacerbations. It would also be worthwhile to investigate its effects on subjects with moderate-to-severe asthma in the same setting. Last, this study also alluded to the fact that a patient's baseline FeNO may be a predictor of whether he or she develops an LAR. This should also be explored further in expanded studies and in children.