Allergen Inhalation Challenge Research Articles

Controlled human exposures to diesel exhaust or particle-depleted diesel exhaust with allergen modulates transcriptomic responses in the lung

The evidence associating traffic-related air pollution (TRAP) with allergic asthma is growing, but the underlying mechanisms for this association remain unclear. The airway epithelium is the primary tissue exposed to TRAP, hence understanding its interactions with TRAP and allergen is important. Diesel exhaust (DE), a paradigm of TRAP, consists of particulate matter (PM) and gases. Modern diesel engines often have catalytic diesel particulate filters to reduce PM output, but these may increase gaseous concentrations, and their benefits on human health cannot be assumed. We conducted a randomized, double-blinded, crossover study using our unique in vivo human exposure system to investigate the effects of DE and allergen co-exposure, with or without particle depletion as a proxy for catalytic diesel particulate filters, on the airway epithelial transcriptome. Participants were exposed for 2 h before an allergen inhalation challenge, with each receiving filtered air and saline (FA-S), filtered air and allergen (FA-A), DE and allergen (DE-A), or particle-depleted DE and allergen (PDDE-A), over four different occasions, each separated by a 4-week washout period. Endobronchial brushings were collected 48 h after each exposure, and total RNA was sequenced. Differentially expressed genes (DEGs) were identified using DESeq2, followed by GO enrichment and pathway analysis. FA-A, DE-A, and PDDE-A exposures significantly modulated genes relative to FA-S, with 462 unique DEGs identified. FA-A uniquely modulated the highest number (↑178, ↓155), followed by DE-A (↑44, ↓23), and then PDDE-A exposure (↑15, ↓2); 6 DEGs (↑4, ↓2) were modulated by all three conditions. Exposure to PDDE-A resulted in modulation of 285 DEGs compared to DE-A exposure, further revealing 26 biological process GO terms, including “cellular response to chemokine” and “inflammatory response”. The transcriptional epithelial response to diesel exhaust and allergen co-exposure is enriched in inflammatory mediators, the pattern of which is altered upon particle depletion.

Neuromedin-U Mediates Rapid Activation of Airway Group 2 Innate Lymphoid Cells in Mild Asthma.

Rationale: In asthma, sputum group 2 innate lymphoid cells (ILC2s) are activated within 7 hours after allergen challenge. Neuroimmune interactions mediate rapid host responses at mucosal interfaces. In murine models of asthma, lung ILC2s colocalize to sensory neuronal termini expressing the neuropeptide neuromedin U (NMU), which stimulates type 2 (T2) cytokine secretion by ILC2s, with additive effects to alarmins invitro. Objectives: To investigate the effect of the NMU/NMUR1 (NMU receptor 1) axis on early activation of ILC2s in asthma. Methods: Subjects with mild asthma (n = 8) were enrolled in a diluent-controlled allergen inhalation challenge study. Sputum ILC2 expression of NMUR1 and T2 cytokines was enumerated by flow cytometry, and airway NMU levels were assessed by ELISA. This was compared with samples from subjects with moderate to severe asthma (n = 9). Flow sort-purified and ex vivo-expanded ILC2s were used for functional assays and transcriptomic analyses. Measurements and Main Results: Significant increases in sputum ILC2s expressing NMUR1 were detected 7 hours after allergen versus diluent challenge whereby the majority of NMUR1+ ILC2s expressed IL-5/IL-13. Sputum NMUR1+ ILC2 counts were significantly greater in mild versus moderate to severe asthma, and NMUR1+ ILC2s correlated inversely with the dose of inhaled corticosteroid in the latter group. Coculturing with alarmins upregulated NMUR1 in ILC2s, which was attenuated by dexamethasone. NMU-stimulated T2 cytokine expression by ILC2s, maximal at 6 hours, was abrogated by dexamethasone or specific signaling inhibitors for mitogen-activated protein kinase 1/2 and phosphoinositol 3-kinase but not the IL-33 signaling moiety MyD88 invitro. Conclusions: The NMU/NMUR1 axis stimulates rapid effects on ILC2s and may be an important early activator of these cells in eosinophilic inflammatory responses in asthma.

Variants in transient receptor potential channels and toll-like receptors modify airway responses to allergen and air pollution: a randomized controlled response human exposure study

BackgroundEnvironmental co-exposure to allergen and traffic-related air pollution is common globally and contributes to the exacerbation of respiratory diseases. Individual responses to environmental insults remain variable due to gene-environment interactions.ObjectiveThis study examined whether single nucleotide polymorphisms (SNPs) in lung cell surface receptor genes modifies lung function change and immune cell recruitment in allergen-sensitized individuals exposed to diesel exhaust (DE) and allergen.MethodsIn this randomized, double-blinded, four-arm, crossover study, 13 allergen-sensitized participants underwent allergen inhalation challenge following a 2-hour exposure to DE, particle-depleted diesel exhaust (PDDE) or filtered air (FA). Lung function tests and bronchoscopic sample collection were performed up to 48 h after exposures. Transient receptor potential channel (TRPA1 and TRPV1) and toll-like receptor (TLR2 and TLR4) risk alleles were used to construct an unweighted genetic risk score (GRS). Exposure-by-GRS interactions were tested using mixed-effects models.ResultsIn participants with high GRS, allergen exposure was associated with an increase in airway hyperresponsiveness (AHR) when co-exposed to PDDE (p = 0.03) but not FA or DE. FA and PDDE also were associated with a relative increase in macrophages and decrease in lymphocytes in bronchoalveolar lavage.ConclusionsTRPs and TLRs variants are associated with increased AHR and altered immune cellularity in allergen-exposed individuals. This effect is blunted by DE exposure, suggesting greater influence of unmeasured gene variants as primary meditators of a particulate-rich co-exposure.Trial registrationThe study was registered with ClinicalTrials.gov on December 20, 2013 (NCT02017431).

Inhaled anti-TSLP antibody fragment, ecleralimab, blocks responses to allergen in mild asthma.

Thymic stromal lymphopoietin (TSLP) is a key upstream regulator driving allergic inflammatory responses. We evaluated the efficacy and safety of ecleralimab, a potent inhaled neutralising antibody fragment against human TSLP, using allergen inhalation challenge (AIC) in subjects with mild atopic asthma. This was a 12-week, randomised, double-blind, placebo-controlled, parallel-design, multicentre allergen bronchoprovocation study conducted at 10 centres across Canada and Germany. Subjects aged 18-60 years with stable mild atopic asthma were randomised (1:1) to receive 4 mg once-daily inhaled ecleralimab or placebo. Primary end-points were the allergen-induced change in forced expiratory volume in 1 s (FEV1) during the late asthmatic response (LAR) measured by area under the curve (AUC3-7h) and maximum percentage decrease (LAR%) on day 84, and the safety of ecleralimab. Allergen-induced early asthmatic response (EAR), sputum eosinophils and fractional exhaled nitric oxide (F ENO) were secondary and exploratory end-points. 28 subjects were randomised to ecleralimab (n=15) or placebo (n=13). On day 84, ecleralimab significantly attenuated LAR AUC3-7h by 64% (p=0.008), LAR% by 48% (p=0.029), and allergen-induced sputum eosinophils by 64% at 7 h (p=0.011) and by 52% at 24 h (p=0.047) post-challenge. Ecleralimab also numerically reduced EAR AUC0-2h (p=0.097) and EAR% (p=0.105). F ENO levels were significantly reduced from baseline throughout the study (p<0.05), except at 24 h post-allergen (day 43 and day 85). Overall, ecleralimab was safe and well tolerated. Ecleralimab significantly attenuated allergen-induced bronchoconstriction and airway inflammation, and was safe in subjects with mild atopic asthma.

Neutrophil phenotypes in bronchial airways differentiate single from dual responding allergic asthmatics.

Allergic asthmatics with both an early (EAR) and a late allergic reaction (LAR) following allergen exposure are termed 'dual responders' (DR), while 'single responders' (SR) only have an EAR. Mechanisms that differentiate DR from SR are largely unknown, particularly regarding the role and phenotypes of neutrophils. Therefore, we aimed to study neutrophils in DR and SR asthmatics. Thirty-four allergic asthmatics underwent an inhaled allergen challenge, samples were collected before and up to 24h post-challenge. Cell differentials were counted from bronchial lavage, alveolar lavage and blood; and tissue neutrophils were quantified in immune-stained bronchial biopsies. Lavage neutrophil nuclei lobe segmentation was used to classify active (1-4lobes) from suppressive neutrophils (≥5lobes). Levels of transmigration markers: soluble (s)CD62L and interleukin-1Ra, and activity markers: neutrophil elastase (NE), DNA-histone complex and dsDNA were measured in lavage fluid and plasma. Compared with SR at baseline, DR had more neutrophils in their bronchial airways at baseline, both in the lavage (p=.0031) and biopsies (p=.026) and elevated bronchial neutrophils correlated with less antitransmigratory IL-1Ra levels (r=-0.64). DR airways had less suppressive neutrophils and more 3-lobed (active) neutrophils (p=.029) that correlated with more bronchial lavage histone (p=.020) and more plasma NE (p=.0016). Post-challenge, DR released neutrophil extracellular trap factors in the blood earlier and had less pro-transmigratory sCD62L during the late phase (p=.0076) than in SR. DR have a more active airway neutrophil phenotype at baseline and a distinct neutrophil response to allergen challenge that may contribute to the development of an LAR. Therefore, neutrophil activity should be considered during targeted diagnosis and bio-therapeutic development for DR.

Dibutyl phthalate exposure alters T‐cell subsets in blood from allergen‐sensitized volunteers

Phthalates are ubiquitous environmental contaminants associated with allergic disease in epidemiological and animal studies. This investigation aims to support these associations by interrogating systemic immune effects in allergen-sensitized volunteers after controlled indoor air exposure to a known concentration of dibutyl phthalate (DBP). The phthalate-allergen immune response (PAIR) study enrolled 16 allergen-sensitized participants to a double-blinded, randomized, crossover exposure to two conditions (DBP or control air for 3hr), each followed immediately by inhaled allergen challenge. Peripheral blood immune cell composition and activation along with inflammatory mediators were measured before and after exposure. DBP exposure prior to the inhaled allergen challenge increased the percentage of CD4+ T helper cells and decreased the percentage of regulatory T cells (3hr and 20hr post-exposure), while only modest overall effects were observed for inflammatory mediators. The cells and mediators affected by the phthalate exposure were generally not overlapping with the endpoints affected by allergen inhalation alone. Thus, in distinction to our previously published effects on lung function, DBP appears to alter endpoints in peripheral blood that are not necessarily enhanced by allergen alone. Further studies are needed to clarify the role of phthalate-induced systemic effects in disease pathogenesis.

Plasma proteome changes linked to late phase response after inhaled allergen challenge in asthmatics

BackgroundA subset of individuals with allergic asthma develops a late phase response (LPR) to inhaled allergens, which is characterized by a prolonged airway obstruction, airway inflammation and airway hyperresponsiveness. The aim of this study was to identify changes in the plasma proteome and circulating hematopoietic progenitor cells associated with the LPR following inhaled allergen challenge.MethodsSerial plasma samples from asthmatics undergoing inhaled allergen challenge were analyzed by mass spectrometry and immunosorbent assays. Peripheral blood mononuclear cells were analyzed by flow cytometry. Mass spectrometry data were analyzed using a linear regression to model the relationship between airway obstruction during the LPR and plasma proteome changes. Data from immunosorbent assays were analyzed using linear mixed models.ResultsOut of 396 proteins quantified in plasma, 150 showed a statistically significant change 23 h post allergen challenge. Among the most upregulated proteins were three protease inhibitors: alpha-1-antitrypsin, alpha-1-antichymotrypsin and plasma serine protease inhibitor. Altered levels of 13 proteins were associated with the LPR, including increased factor XIII A and decreased von Willebrand factor. No relationship was found between the LPR and changes in the proportions of classical, intermediate, and non-classical monocytes.ConclusionsAllergic reactions to inhaled allergens in asthmatic subjects were associated with changes in a large proportion of the measured plasma proteome, whereof protease inhibitors showed the largest changes, likely to influence the inflammatory response. Many of the proteins altered in relation to the LPR are associated with coagulation, highlighting potential mechanistic targets for future treatments of type-2 asthma.

Aerosol Biologics for the Treatment of Eosinophilic Asthma

Asthma is a heterogeneous chronic airway disease comprising of distinct phenotypes characterized by different immunopathophysiologic pathways, clinical features, disease severity, and response to treatment. The phenotypes of asthma include eosinophilic, neutrophilic, mixed cellularity, and paucigranulocytic asthma. Eosinophilic asthma is principally a T helper type 2 (Th2)-mediated airway disease. However, several other immune and structural cells secrete the cytokines implicated in the pathogenesis of eosinophilic asthma. Innate type 2 lymphoid cells, mast cells, basophils, and eosinophils secrete Th2 cytokines, such as interleukin-4 (IL-4), IL-13, and IL-5. Additionally, airway epithelial cells produce alarmin cytokines, including IL-25, IL-33, and thymic stromal lymphopoietin (TSLP). Alarmins are the key initiators of allergic inflammation at the sentinel mucosal surfaces. Innovative biotherapeutic research has led to the discovery of monoclonal antibodies which target and inhibit the immunopathological effects of the cytokines involved in the pathogenesis of eosinophilic asthma. Parenteral biologics targeting the inciting interleukins, include mepolizumab and reslizumab (anti-IL-5), benralizumab (anti-IL-5Rα), dupilumab (anti-4Rα), and tezelizumab (anti-TSLP). They have been shown to significantly reduce annualized exacerbation rates, improve asthma control, lung function, and quality of life. Currently, there are no pulmonary delivered aerosol biologics for topical treatment of asthma. CSJ117 is a potent neutralizing antibody Fab fragment against TSLP, formulated as a PulmoSol TM engineered powder, and is delivered to the lungs by a dry powder inhaler. Phase 2 placebo-controlled clinical trial evaluated the efficacy and safety of CSJ117. CSJ117 delivered as an inhaler attenuated the late asthmatic response (LAR), and the early asthmatic response (EAR) after allergen inhalation challenge (AIC) at day 84 of treatment. The maximum decrease in FVE1 from pre-AIC were significantly lower in the CSJ117 group compared to placebo (P = 029), during LAR. CSJ117 also significantly reduced fractional exhaled nitric oxide before AIC at day 83; and significantly reduced the allergen-induced increase in % sputum eosinophil count. Pulmonary delivery of biologics directly to the airway mucosal surface has several advantages over parenteral routes, particularly in treating airway diseases such as asthma. Inhaler delivered biologics, such as CSJ117 are innovative and attractive methods of future precision treatment of asthma, and other respiratory diseases.

Peroxisome proliferator-activated receptor gamma gene variants modify human airway and systemic responses to indoor dibutyl phthalate exposure

BackgroundSingle nucleotide polymorphisms (SNPs) of peroxisome proliferator-activated receptor gamma (PPAR-γ; gene: PPARG) and oxidative stress genes are associated with asthma risk. However, whether such variants modulate responses to dibutyl phthalate (DBP), a common plasticizer associated with increased asthma development, remains unknown. The purpose of this study is to investigate how SNPs in PPARG and oxidative stress genes, as represented by two separate genetic risk scores, modify the impact of DBP exposure on lung function and the airway and systemic response after an inhaled allergen challenge.MethodsWe conducted a double-blinded human crossover study with sixteen allergen-sensitized participants exposed for three hours to DBP and control air on distinct occasions separated by a 4-week washout. Each exposure was followed by an allergen inhalation challenge; subsequently, lung function was measured, and blood and bronchoalveolar lavage (BAL) were collected and analyzed for cell counts and allergen-specific immunoglobulin E (IgE). Genetic risk scores for PPAR-γ (P-GRS; weighted sum of PPARG SNPs rs10865710, rs709158, and rs3856806) and oxidative stress (OS-GRS; unweighted sum of 16 SNPs across multiple genes) were developed, and their ability to modify DBP effects were assessed using linear mixed-effects models.ResultsP-GRS and OS-GRS modified DBP effects on allergen-specific IgE in blood at 20 h (interaction effect [95% CI]: 1.43 [1.13 to 1.80], p = 0.005) and 3 h (0.99 [0.98 to 1], p = 0.03), respectively. P-GRS also modified DBP effects on Th2 cells in blood at 3 h (− 25.2 [− 47.7 to − 2.70], p = 0.03) and 20 h (− 39.1 [− 57.9 to − 20.3], p = 0.0005), and Th2 cells in BAL at 24 h (− 4.99 [− 8.97 to − 1.01], p = 0.02). An increasing P-GRS associated with reduced DBP effect on Th2 cells. Neither GRS significantly modified DBP effects on lung function parameters.ConclusionsPPAR-γ variants modulated several airway and systemic immune responses to the ubiquitous chemical plasticizer DBP. Our results suggest that PPAR-γ variants may play a greater role than those in oxidative stress-related genes in airway allergic responses to DBP.Trial registration: This study reports results from The Phthalate-Allergen Immune Response Study that was registered on ClinicalTrials.gov with identification NCT02688478.

Free Feeding of CpG-Oligodeoxynucleotide Particles Prophylactically Attenuates Allergic Airway Inflammation and Hyperresponsiveness in Mice.

CpG-oligodeoxynucleotides (CpG-ODNs) constitute an attractive alternative for asthma treatment. However, very little evidence is available from studies on the oral administration of CpG-ODNs in animals. Previously, we developed acid-resistant particles (named ODNcap) as an oral delivery device for ODNs. Here, we showed that free feeding of an ODNcap-containing feed prophylactically attenuates allergic airway inflammation, hyperresponsiveness, and goblet cell hyperplasia in an ovalbumin-induced asthma model. Using transcriptomics-driven approaches, we demonstrated that injury of pulmonary vein cardiomyocytes accompanies allergen inhalation challenge, but is inhibited by ODNcap feeding. We also showed the participation of an airway antimicrobial peptide (Reg3γ) and fecal microbiota in the ODNcap-mediated effects. Collectively, our findings suggest that daily oral ingestion of ODNcap may provide preventive effects on allergic bronchopulmonary insults via regulation of mechanisms involved in the gut-lung connection.

Peripheral blood intermediate monocyte protease-activated receptor-2 expression increases during asthma exacerbations and after inhalation allergen challenge

BackgroundMyeloid cells, especially dendritic cells and macrophages, play important roles in asthma pathophysiology. Monocytes (Mo) and macrophages express protease-activated receptor-2 (PAR-2), a proinflammatory serine protease receptor implicated in the pathophysiology of allergic airway inflammation. We have revealed that patients with severe asthma and those with a history of frequent asthma exacerbations exhibit increased PAR-2 expression on peripheral blood monocytes. ObjectiveTo determine PAR-2 expression on peripheral blood intermediate monocytes (IMMo) in subjects with increased airway inflammation, either as a result of an asthma exacerbation or after an inhalation allergen challenge. MethodsA total of 16 adults who presented to the emergency department with asthma exacerbations were recruited after giving an informed consent. After 2 weeks, 10 patients returned for follow-up. A total of 11 patients with mild asthma treated only with as-needed bronchodilators were recruited and underwent inhalation allergen challenge after providing an informed consent. Immune cell profiling was performed by whole blood flow cytometry in both groups of patients. ResultsPAR-2 expression in peripheral blood IMMo increased in patients with an asthma exacerbation compared with those with stable disease, but this expression decreased after treatment of the asthma exacerbation. Subjects with mild asthma had an increase in percentages of IMMo expressing PAR-2 after an allergen challenge. Patients who presented to the emergency department had lower dendritic cell and dendritic cell subset numbers in peripheral blood during exacerbation compared with after treatment. ConclusionIncreased PAR-2 expression on Mo during periods of increased airway inflammation may initiate a positive feedback loop leading to systemic inflammatory changes.

Characterizing the early and late asthmatic responses in the allergen inhalation challenge

Allergen inhalation challenges (AICs) are widely used in research to assess airway responses to allergen exposure and predict the therapeutic effectiveness of various medications.1 The characteristics of early asthmatic response (EAR) and late asthmatic response (LAR) are often described in general terms.2-4 Original research findings focus on mean data. We are not aware of any data objectively documenting the responses of individual participants, and it is important to acknowledge that mean data do not accurately reflect the range of individual responses and the relevant safety implications of performing AICs.

FcεR1-expressing nociceptors trigger allergic airway inflammation

Lung nociceptor neurons amplify immune cell activity and mucus metaplasia in response to an inhaled allergen challenge in sensitized mice. We sought to identify the cellular mechanisms by which these sensory neurons are activated subsequent to allergen exposure. We used calcium microscopy and electrophysiologic recording to assess whether vagal neurons directly respond to the model allergen ovalbumin (OVA). Next, we generated the first nociceptor-specific FcεR1γ knockdown (TRPV1Cre::FcεR1γfl/fl) mice to assess whether this targeted invalidation would affect the severity of allergic inflammation in response to allergen challenges. Lung-innervating jugular nodose complex ganglion neurons express the high-affinity IgE receptor FcεR1, the levels of which increase in OVA-sensitized mice. FcεR1γ-expressing vagal nociceptor neurons respond directly to OVA complexed with IgE with depolarization, action potential firing, calcium influx, and neuropeptide release. Activation of vagal neurons by IgE-allergen immune complexes, through the release of substance P from their peripheral terminals, directly amplifies TH2 cell influx and polarization in the airways. Allergic airway inflammation is decreased in TRPV1Cre::FcεR1γfl/fl mice and in FcεR1α-/- mice into which bone marrow has been transplanted. Finally, increased invivo circulating levels of IgE following allergen sensitization enhances the responsiveness of FcεR1 to immune complexes in both mouse jugular nodose complex ganglion neurons and human induced pluripotent stem cell-derived nociceptors. Allergen sensitization triggers a feedforward inflammatory loop between IgE-producing plasma cells, FcεR1-expressing vagal sensory neurons, and TH2 cells, which helps to both initiate and amplify allergic airway inflammation. These data highlight a novel target for reducing allergy, namely, FcεR1γ expressed by nociceptors.

Dibutyl Phthalate Augments Allergen-induced Lung Function Decline and Alters Human Airway Immunology. A Randomized Crossover Study.

Rationale: Phthalates are a group of chemicals used in common commercial products. Epidemiological studies suggest that phthalate exposure is associated with development or worsening of allergic diseases such as asthma. However, effects of dibutyl phthalate (DBP) or other phthalates found in high concentrations in indoor air have never been examined in allergic individuals in a controlled exposure setting.Objectives: To investigate the airway effects in humans caused by inhalation of a known concentration of a single phthalate, DBP.Methods: In a randomized crossover study, 16 allergen-sensitized participants were exposed to control air or DBP for 3 hours in an environmental chamber followed immediately by an allergen inhalation challenge. Bronchoalveolar wash and lavage were obtained 24 hours after exposure. Lung function, early allergic response, airway responsiveness, inflammation, immune mediators, and immune cell phenotypes were assessed after DBP exposure.Measurements and Main Results: DBP exposure increased the early allergic response (21.4% decline in FEV1 area under the curve, P = 0.03). Airway responsiveness was increased by 48.1% after DBP exposure in participants without baseline hyperresponsiveness (P = 0.01). DBP increased the recruitment of BAL total macrophages by 4.6% (P = 0.07), whereas the M2 macrophage phenotype increased by 46.9% (P = 0.04). Airway immune mediator levels were modestly affected by DBP.Conclusions: DBP exposure augmented allergen-induced lung function decline, particularly in those without baseline hyperresponsiveness, and exhibited immunomodulatory effects in the airways of allergic individuals. This is the first controlled human exposure study providing biological evidence for phthalate-induced effects in the airways.Clinical trial registered with www.clinicaltrials.gov (NCT02688478).

Effects of myeloid and plasmacytoid dendritic cells on ILC2s in patients with allergic rhinitis

Group 2 innate lymphoid cells (ILC2s) were reported to serve a critical role in allergic diseases. Myeloid dendritic cells (mDCs) and plasmacytoid DCs (pDCs) play significant roles in allergic immune response. However, effects of DCs on ILC2s in allergic diseases, especially for patients with allergic rhinitis (AR), remain unclear. We sought to address the roles of mDCs and pDCs in regulating ILC2 function in AR. mDCs and pDCs were cocultured with human PBMCs isolated from patients with AR or ILC2s to measure soluble or intracellular TH2 cytokines, transcription factors, signaling pathways in ILC2s, and the following mechanisms were further investigated. The levels of peripheral IL-33+mDCs, pDCs, and ILC2s were studied in patients under an inhaled allergen challenge. We confirmed the presence of ILC2s, mDCs, and pDCs in the nasal mucosa of patients with AR. Both allogenic and autologous mDCs were found to activate ILC2s from patients with AR to produce TH2 cytokines, and increase the levels of GATA-3 and signal transducer and activator of transcription signaling pathways, in which IL-33-producing mDCs exerted the major role by binding on ST2 on ILC2s. We further identified high levels of IL-33+mDCs and ILC2s in patients with AR under antigen challenge. Activated pDCs inhibited the cytokine production of ILC2s isolated from patients with AR by secretion of IL-6. mDCs promote ILC2 function by the IL-33/ST2 pathway, and activation of pDCs suppresses ILC2 function through IL-6 in patients with AR. Our findings provide new understanding of the interplay between DCs and ILC2s in allergic diseases.

Particle Depletion Does Not Remediate Acute Effects of Traffic-related Air Pollution and Allergen. A Randomized, Double-Blind Crossover Study.

Rationale: Diesel exhaust (DE), an established model of traffic-related air pollution, contributes significantly to the global burden of asthma and may augment the effects of allergen inhalation. Newer diesel particulate-filtering technologies may increase NO2 emissions, raising questions regarding their effectiveness in reducing harm from associated engine output.Objectives: To assess the effects of DE and allergen coexposure on lung function, airway responsiveness, and circulating leukocytes, and determine whether DE particle depletion remediates these effects.Methods: In this randomized, double-blind crossover study, 14 allergen-sensitized participants (9 with airway hyperresponsiveness) underwent inhaled allergen challenge after 2-hour exposures to DE, particle-depleted DE (PDDE), or filtered air. The control condition was inhaled saline after filtered air. Blood sampling and spirometry were performed before and up to 48 hours after exposures. Airway responsiveness was evaluated at 24 hours.Measurements and Main Results: PDDE plus allergen coexposure impaired lung function more than DE plus allergen, particularly in those genetically at risk. DE plus allergen and PDDE plus allergen each increased airway responsiveness in normally responsive participants. DE plus allergen increased blood neutrophils and was associated with persistent eosinophilia at 48 hours. DE and PDDE each increased total peripheral leukocyte counts in a manner affected by participant genotypes. Changes in peripheral leukocytes correlated with lung function decline.Conclusions: Coexposure to DE and allergen impaired lung function, which was worse after particle depletion (which increased NO2). Thus, particulates are not necessarily the sole or main culprit responsible for all harmful effects of DE. Policies and technologies aimed at protecting public health should be scrutinized in that regard.Clinical trial registered with www.clinicaltrials.gov (NCT02017431).

Human BCR analysis of single-sorted, putative IgE+ memory B cells in food allergy

R. Jimenez-Saiz, Y. Ellenbogen and K. Bruton et al. developed a method to identify bona fide IgE+ MBCs in humans, demonstrated their extreme rarity in circulation and cautioned against the clinical utility of their assessment.

Effects of interleukin-6 receptor blockade on allergen-induced airway responses in mild asthmatics.

BackgroundInterleukin (IL)‐6 signalling has been implicated in allergic asthma by animal, genetic association and clinical studies. In this study, we tested the hypothesis that tocilizumab (TCZ), a human monoclonal antibody that blocks IL‐6 signalling, can prevent the development of allergen‐induced bronchoconstriction in humans.MethodsWe performed a randomised, double‐blind, placebo‐controlled study, with eligible participants completing two allergen inhalation challenge tests, conducted before and after treatment with a single dose of TCZ or placebo. The primary efficacy endpoint was the magnitude of the late asthmatic response recorded between 3 and 7 after allergen challenge. The secondary efficacy endpoint was the early asthmatic response, measured 20 min to 2 h after allergen challenge.ResultsA total of 66 patients enrolled between September 2014 and August 2017, when the trial was stopped for futility based on results from an interim analysis. Eleven patients fulfilled all eligibility criteria assessed at baseline and were subsequently randomised to the TCZ (n = 6) or placebo (n = 5) groups. Both the primary and secondary efficacy endpoints were not significantly different between the two groups. Five patients reported adverse events (AEs), three in the TCZ group (11 AEs) and two in the placebo group (four AEs). Only one AE was TCZ‐related (mild neutropenia), and there were no serious AEs. Significant treatment effects were observed for serum levels of C‐reactive protein, IL‐6 and soluble IL‐6R levels.ConclusionIn a small proof‐of‐concept clinical trial, we found no evidence that a single dose of tocilizumab was able to prevent allergen‐induced bronchoconstriction. (Trial registered in the Australian New Zealand Clinical Trials Registry, number ACTRN12614000123640).

IL-33 and Its Receptor ST2 after Inhaled Allergen Challenge in Allergic Asthmatics

Background: Previous murine models have demonstrated interleukin (IL)-33 to be an important mediator of type-2 inflammation and to promote airway hyperresponsiveness in allergic asthma. A number of inflammatory cells produce IL-33 and eosinophils express ST2 mRNA. The relationship between IL-33 and eosinophils in allergic asthma, however, remains unclear. Objective: The aim of this work was to evaluate in vitro the effect of allergen inhalation on IL-33 levels and expression of its receptor (ST2L) on eosinophils in allergic asthmatics, and the effect of IL-33 stimulation on eosinophil activity. Methods: Plasma and sputum IL-33, soluble ST2 (sST2) levels, and ST2L expression on eosinophils were measured in 10 healthy controls and 10 allergic asthmatics. Asthmatics underwent allergen and diluent inhalation challenges. Blood and sputum samples were collected to measure IL-33, sST2, and ST2L eosinophil expression before and 24 h after allergen inhalation. Purified blood eosinophils from allergic asthmatics were incubated overnight with IL-33 to assess ST2 and intracellular IL-5 expression. Results: Baseline levels of IL-33 in sputum and sST2 in plasma and sputum were similar in allergic asthmatics compared to healthy controls. In addition, there was no difference in blood or sputum eosinophil ST2L expression in healthy controls versus allergic asthmatics. Eosinophil ST2L expression was significantly increased 24 h postallergen inhalation in allergic asthmatics. In vitro stimulation of human eosinophils with IL-33 and LPS significantly increased eosinophil ST2L expression and IL-33 stimulation increased intracellular IL-5 expression, which was attenuated by treatment with sST2 and ST2 blockade. Conclusion and Clinical Relevance: In mild asthmatics, there was a significant upregulation of ST2 surface expression on eosinophils from blood and sputum following allergen inhalation challenge. In vitro, IL-33 stimulation of eosinophils increases both ST2 membrane expression and IL-5 production. These results support a role for IL-33 in causing allergen-induced eosinophilia. Blockade of IL-33 and ST2 signaling may present a novel therapeutic avenue for asthma treatment.

The effects of a CCR3 inhibitor, AXP1275, on allergen‐induced airway responses in adults with mild‐to‐moderate atopic asthma

CCR3 is the cognate receptor for major human eosinophil chemoattractants from the eotaxin family of proteins that are elevated in asthma and correlate with disease severity. This proof-of-mechanism study examined the effect of AXP1275, an oral, small-molecule inhibitor of CCR3, on airway responses to inhaled allergen challenge. Twenty-one subjects with mild atopic asthma and documented early and late asthmatic responses to an inhaled aeroallergen completed a randomized double-blind cross-over study to compare early and late allergen-induced asthmatic responses, methacholine PC20 , blood and sputum eosinophils and exhaled nitric oxide after 2weeks of treatment with once-daily doses of AXP1275 (50mg) or placebo. There was a significant increase in methacholine PC20 after 12days of AXP1275 treatment compared to placebo (increase of 0.92 doubling doses versus 0.17 doubling doses, P=.01), but this protection was lost post-allergen challenge. There was no effect of AXP1275 on allergen-induced late asthmatic responses, or eosinophils in blood and sputum. The early asthmatic response and exhaled nitric oxide levels were slightly lower with AXP1275, but this did not reach statistical significance. The number of subjects who experienced treatment-emergent adverse events while receiving AXP1275 was comparable placebo. AXP1275 50mg administered daily was safe and well tolerated, and there was no difference in the type, severity or frequency of treatment-emergent adverse events in subjects while receiving AXP1275 compared to placebo. AXP1275 increased the methacholine PC20 ; however, the low and variable exposure to APX1275 over a short treatment period may have contributed to poor efficacy on other outcomes.