Allergen Challenge Research Articles

Maternal allergic inflammation in rats impacts the offspring perinatal neuroimmune milieu and the development of social play, locomotor behavior, and cognitive flexibility

Maternal systemic inflammation increases risk for neurodevelopmental disorders like autism, ADHD, and schizophrenia in offspring. Notably, these disorders are male-biased. Studies have implicated immune system dysfunction in the etiology of these disorders, and rodent models of maternal immune activation provide useful tools to examine mechanisms of sex-dependent effects on brain development, immunity, and behavior. Here, we employed an allergen-induced model of maternal inflammation in rats to characterize levels of mast cells and microglia in the perinatal period in male and female offspring, as well as social, emotional, and cognitive behaviors throughout the lifespan. Adult female rats were sensitized to ovalbumin (OVA), bred, and challenged intranasally on gestational day 15 of pregnancy with OVA or saline. Allergic inflammation upregulated microglia in the fetal brain, increased mast cell number in the hippocampus on the day of birth, and conferred region-, time- and sex- specific changes in microglia measures. Additionally, offspring of OVA-exposed mothers subsequently exhibited abnormal social behavior, hyperlocomotion, and reduced cognitive flexibility. These data demonstrate the long-term effects of maternal allergic challenge on offspring development and provide a basis for understanding neurodevelopmental disorders linked to maternal systemic inflammation in humans.

Systematic evaluation of allergic phenotypes of rhinitis in children and adolescents.

Three allergic phenotypes of rhinitis have been described in adults: allergic rhinitis (AR), local allergic rhinitis (LAR), and dual allergic rhinitis (DAR, coexistence of AR and LAR). Nevertheless, most centers follow a diagnostic approach only based on skin prick test and serum allergen-specific IgE (collectively called atopy tests, AT). This approach prevents the recognition of LAR and DAR, the diagnosis of which requires a nasal allergen challenge (NAC). Here, we investigate the existence of LAR and DAR phenotypes in children and adolescents, and the misdiagnosis rate associated with a work-up exclusively based on AT. Clinical data were obtained during physician-conducted interviews, and AT and NAC were systematically performed in 5- to 18-year-old patients with chronic rhinitis. The misdiagnosis rate was defined as the proportion of cases where AT and NAC results were discordant. A total of 173 patients (mean age 15.1years, 39.9% male) completed the study. AR (positive AT and NAC), LAR (negative AT and positive NAC), DAR (positive AT and NAC for some allergens and negative AT and positive NAC for other allergens), and non-allergic rhinitis (negative NAC) were diagnosed in 45.7%, 24.9%, 11.6%, and 17.9% of individuals, respectively. The clinical profile was comparable among allergic phenotypes, but allergic patients had a significantly earlier rhinitis onset, higher conjunctivitis prevalence, and more severe disease than NAR individuals. A diagnostic work-up exclusively based on AT misclassified 37.6% of patients. LAR and DAR represent relevant differential diagnosis in pediatric rhinitis. NAC increases the diagnostic accuracy of clinical algorithms for rhinitis in children and adolescents.

Phase 3 Randomized Study of Efficacy and Safety of a Dexamethasone Intracanalicular Insert in Patients With Allergic Conjunctivitis

The purpose of this study was to evaluate the efficacy and safety of a dexamethasone intracanalicular ocular insert for the treatment of allergic conjunctivitis. Multicenter, randomized, double-masked, placebo-controlled, Phase 3 clinical trial. Subjects with allergic conjunctivitis were randomized 1:1 to receive a dexamethasone insert or a placebo insert in both eyes and were evaluated using a modified version of the conjunctival allergen challenge (CAC) model. After inserts were placed in office, a series of 4 closely spaced post-insertion CACs were conducted at weeks 1, 2, and 4 across approximately 30 days. Primary efficacy endpoints, assessed at week-1 CAC-day 8, were reported by subjects of ocular itching at 3, 5, and 7 minutes post CAC and investigator-evaluated conjunctival redness at 7, 15, and 20 minutes post CAC. For the primary endpoints, dexamethasone inserts showed statistically significantly lower mean ocular itching scores than placebo at all time points (P <.001), with differences favoring dexamethasone inserts over placebo (0.86, 0.98, and 0.96 units at 3, 5, and 7 minutes, respectively) and statistically significantly lower conjunctival redness scores at 20 minutes (P <.05) but not at 7 or 15 minutes (P ≥.05). Results also showed statistically significantly less itching and conjunctival redness at 31 and 29 of 33 other time points, respectively (P <.05). There were no serious adverse events; 1 subject had elevated intraocular pressure in both eyes. Data presented in this study demonstrate the potential for a single, physician-administered dexamethasone intracanalicular insert to provide relief of ocular itching for up to 4 weeks in subjects with allergic conjunctivitis, while maintaining a favorable safety profile.

RNA demethylation by FTO stabilizes the FOXJ1 mRNA for proper motile ciliogenesis.

Adenosine N6-methylation (m6A) is one of the most pervasive mRNA modifications, and yet the physiological significance of m6A removal (demethylation) remains elusive. Here, we report that the m6A demethylase FTO functions as a conserved regulator of motile ciliogenesis. Mechanistically, FTO demethylates and thereby stabilizes the mRNA that encodes the master ciliary transcription factor FOXJ1. Depletion of Fto in Xenopus laevis embryos caused widespread motile cilia defects, and Foxj1 was identified as one of the major phenocritical targets. In primary human airway epithelium, FTO depletion also led to FOXJ1 mRNA destabilization and a severe loss of ciliated cells with an increase of neighboring goblet cells. Consistently, Fto knockout mice showed strong asthma-like phenotypes upon allergen challenge, a result owing to defective ciliated cells in the airway epithelium. Altogether, our study reveals a conserved role of the FTO-FOXJ1 axis in embryonic and homeostatic motile ciliogenesis.

Development of nasal allergen challenge with cockroach in children with asthma.

Nasal allergen challenge (NAC) could be a means to assess indication and/or an outcome of allergen-specific therapies, particularly for perennial allergens. NACs are not commonly conducted in children with asthma, and cockroach NACs are not well established. This study's objective was to identify a range of German cockroach extract doses that induce nasal symptoms and to assess the safety of cockroach NAC in children with asthma. Ten adults (18-37years) followed by 25 children (8-14years) with well-controlled, persistent asthma and cockroach sensitization underwent NAC with diluent followed by up to 8 escalating doses of cockroach extract (0.00381-11.9µg/mL Bla g 1). NAC outcome was determined by Total Nasal Symptom Score (TNSS) and/or sneeze score. Cockroach allergen-induced T-cell activation and IL-5 production were measured in peripheral blood mononuclear cells. 67% (6/9) of adults and 68% (17/25) of children had a positive NAC at a median response dose of 0.120µg/mL [IQR 0.0380-0.379µg/mL] of Bla g 1. Additionally, three children responded to diluent alone and did not receive any cockroach extract. Overall, 32% (11/34) were positive with sneezes alone, 15% (5/34) with TNSS alone, and 21% (7/34) with both criteria. At baseline, NAC responders had higher cockroach-specific IgE (P=.03), lower cockroach-specific IgG/IgE ratios (children, P=.002), and increased cockroach-specific IL-5-producing T lymphocytes (P=.045). The NAC was well tolerated. We report the methodology of NAC development for children with persistent asthma and cockroach sensitization. This NAC could be considered a tool to confirm clinically relevant sensitization and to assess responses in therapeutic studies.

Androgen receptor activation alleviates airway hyperresponsiveness, inflammation, and remodeling in a murine model of asthma

Epidemiological studies demonstrate an apparent sex-based difference in the prevalence of asthma, with a higher risk in boys than girls, which is reversed postpuberty, where women become more prone to asthma than men, suggesting a plausible beneficial role for male hormones, especially androgens as a regulator of pathophysiology in asthmatic lungs. Using a murine model of asthma developed with mixed allergen (MA) challenge, we report a significant change in airway hyperresponsiveness (AHR), as demonstrated by increased thickness of epithelial and airway smooth muscle layers and collagen deposition, as well as Th2/Th17-biased inflammation in the airways of non-gonadectomized (non-GDX) and gonadectomized (GDX) male mice. Here, compared with non-GDX mice, MA-induced AHR and inflammatory changes were more prominent in GDX mice. Activation of androgen receptor (AR) using 5α-dihydrotestosterone (5α-DHT, AR agonist) resulted in decreased Th2/Th17 inflammation and remodeling-associated changes, resulting in improved lung function compared with MA alone challenged mice, especially in GDX mice. These changes were not observed with Flutamide (Flut, AR antagonist). Overall, we show that AR exerts a significant and beneficial role in asthma by regulating AHR and inflammation.

Anxiety-like behavior and intestinal microbiota changes as strain-and sex-dependent sequelae of mild food allergy in mouse models of cow’s milk allergy

A number of studies have reported comorbidity of food allergies with various neuropsychiatric disorders, such as anxiety, depression, attention-deficit hyperactivity disorder, and autism. However, inconsistent results across clinical studies have left the association between food allergy and behavioral disorders inconclusive. We postulated that the heterogeneities in genetic background among allergic cohorts affect symptom presentation and severity of food allergy, introducing bias in patient selection criteria toward individuals with overt physical reactions. To understand the influence of genetic background on food allergy symptoms and behavioral changes beyond anaphylaxis, we generated mouse models with mild cow’s milk allergy by sensitizing male and female C57BL/6J and BALB/cJ mice to a bovine whey protein, β-lactoglobulin (BLG; Bos d 5). We compared strain- and sex-dependent differences in their immediate physical reactions to BLG challenge as well as anxiety-like behavior one day after the challenge. While reactions to the allergen challenge were either absent or mild for all groups, a greater number of BLG-sensitized BALB/cJ mice presented visible symptoms and hypothermia compared to C57BL/6J mice. Interestingly, male mice of both strains displayed anxiety-like behavior on an elevated zero maze without exhibiting cognitive impairment with the cross maze test. Further characterization of plasma cytokines/chemokines and fecal microbiota also differentiated strain- and sex-dependent effects of BLG sensitization on immune-mediator levels and bacterial populations, respectively. These results demonstrated that the genetic variables in mouse models of milk allergy influenced immediate physical reactions to the allergen, manifestation of anxiety-like behavior, levels of immune responses, and population shift in gut microbiota. Thus, stratification of allergic cohorts by their symptom presentations and severity may strengthen the link between food allergy and behavioral disorders and identify a population(s) with specific genetic background that have increased susceptibility to allergy-associated behavioral disorders.

Fetomaternal immune cross talk modifies T-cell priming through sustained changes to DC function

Prenatal exposure to infections can modify immune development. These environmental disturbances during early life potentially alter the incidence of inflammatory disorders as well as priming of immune responses. Infection with the helminth Schistosoma mansoni is widely studied for its ability to alter immune responsiveness and is associated with variations in coinfection, allergy, and vaccine efficacy in endemic populations. Exposure to maternal schistosomiasis during early life, even without transmission of infection, can result in priming effects on offspring immune responses to bystander antigenic challenges as related to allergic responsiveness and vaccination, with this article seeking to further clarify the effects and underlying immunologic imprinting. Here, we have combined a model of chronic maternal schistosomiasis infection with a thorough analysis of subsequent offspring immune responses to allergy and vaccination models, including viral challenge and steady-state changes to immune cell compartments. We have demonstrated that maternal schistosomiasis alters CD4+ responses during allergic sensitization and challenge in a skewed IL-4/B-cell-dominant response to antigenic challenge associated with limited inflammatory response. Beyond that, we have uncovered previously unidentified alterations to CD8+ T-cell responses during immunization that are dependent on vaccine formulation and have functional impact on the efficacy of vaccination against viral infection in a murine hepatitis B virus model. In addition to steady-state modifications to CD4+ T-cell polarization and B-cell priming, we have traced these modified CD8+ responses to an altered dendritic cell phenotype sustained into adulthood, providing evidence for complex priming effects imparted by infection via fetomaternal cross talk.

Basophil activation test: Mechanisms and considerations for use in clinical trials and clinical practice.

The basophil activation test (BAT) is a functional assay that measures the degree of degranulation following stimulation with allergen or controls by flow cytometry. It correlates directly with histamine release. From the dose-response curve resulting from BAT in allergic patients, basophil reactivity (%CD63+ basophils) and basophil sensitivity (EC50 or similar) are the main outcomes of the test. BAT takes into account all characteristics of IgE and allergen and thus can be more specific than sensitization tests in the diagnosis of allergic disease. BAT reduces the need for in vivo procedures, such as intradermal tests and allergen challenges, which can cause allergic reactions of unpredictable severity. As it closely reflects the patients' phenotype in most cases, it may be used to support the diagnosis of food, venom and drug allergies and chronic urticaria, to monitor the natural resolution of food allergies and to predict and monitor clinical the response to immunomodulatory treatments, such as allergen-specific immunotherapy and biologicals. Clinical application of BAT requires analytical validation, clinical validation, standardization of procedures and quality assurance to ensure reproducibility and reliability of results. Currently, efforts are ongoing to establish a platform that could be used by laboratories in Europe and in the USA for quality assurance and certification.

Mucosal Nanoemulsion Allergy Vaccine Suppresses Alarmin Expression and Induces Bystander Suppression of Reactivity to Multiple Food Allergens.

We have demonstrated that intranasal immunotherapy with allergens formulated in a nanoemulsion (NE) mucosal adjuvant suppresses Th2/IgE-mediated allergic responses and protects from allergen challenge in murine food allergy models. Protection conferred by this therapy is associated with strong suppression of allergen specific Th2 cellular immunity and increased Th1 cytokines. Here we extend these studies to examine the effect of NE-allergen immunization in mice sensitized to multiple foods. Mice were sensitized to both egg and peanut and then received NE vaccine formulated with either one or both of these allergens. The animals were then subjected to oral challenges with either egg or peanut to assess reactivity. Immunization with NE formulations containing both egg and peanut markedly reduced reactivity after oral allergen challenge with either allergen. Interestingly, mice that received the vaccine containing only peanut also had reduced reactivity to challenge with egg. Protection from oral allergen challenge was achieved despite the persistence of allergen-specific IgE and was associated with strong suppression of both Th2-polarized immune responses, alarmins and type 2 innate lymphoid cells (ILC2). NE-induced bystander suppression of reactivity required IFN-γ and the presence of an allergen in the NE vaccine. These results demonstrate that anaphylactic reactions to food allergens can be suppressed using allergen-specific immunotherapy without having to eliminate allergen-specific IgE and suggests that modulation of Th2 immunity towards one allergen may induce bystander effects that suppress reactivity to other allergens through the induction of IFN-γ and suppression of alarmins in the intestine. In addition, these data suggest that a NE vaccine for a single food allergen may lead to a global suppression of allergic responses to multiple foods.

Platelets Independently Recruit into Asthmatic Lungs and Models of Allergic Inflammation via CCR3.

Platelet activation and pulmonary recruitment occur in patients with asthma and in animal models of allergic asthma, in which leukocyte infiltration, airway remodeling, and hyperresponsiveness are suppressed by experimental platelet depletion. These observations suggest the importance of platelets to various characteristics of allergic disease, but the mechanisms of platelet migration and location are not understood. The aim of this study was to assess the mechanism of platelet recruitment to extravascular compartments of lungs from patients with asthma and after allergen challenge in mice sensitized to house dust mite (HDM) extract (contains the DerP1 [Dermatophagoides pteronyssinus extract peptidase 1] allergen); in addition, we assessed the role of chemokines in this process. Lung sections were immunohistochemically stained for CD42b+ platelets. Intravital microscopy in allergic mice was used to visualize platelets tagged with an anti–mouse CD49b-PE (phycoerythrin) antibody. Platelet–endothelial interactions were measured in response to HDM (DerP1) exposure in the presence of antagonists to CCR3, CCR4, and CXCR4. Extravascular CD42b+ platelets were detected in the epithelium and submucosa in bronchial biopsy specimens taken from subjects with steroid-naive mild asthma. Platelets were significantly raised in the lung parenchyma from patients with fatal asthma compared with postmortem control-lung tissue. Furthermore, in DerP1-sensitized mice, subsequent HDM exposure induced endothelial rolling, endothelial adhesion, and recruitment of platelets into airway walls, compared with sham-sensitized mice, via a CCR3-dependent mechanism in the absence of aggregation or interactions with leukocytes. Localization of singular, nonaggregated platelets occurs in lungs of patients with asthma. In allergic mice, platelet recruitment occurs via recognized vascular adhesive and migratory events, independently of leukocytes via a CCR3-dependent mechanism.

Increased IL-6 and Potential IL-6 trans-signalling in the airways after an allergen challenge.

In asthma, IL-6 is a potential cause of enhanced inflammation, tissue damage and airway dysfunction. IL-6 signalling is regulated by its receptor, which is composed of two proteins, IL-6R and GP130. In addition to their membrane form, these two proteins may be found as extracellular soluble forms. The interaction of IL-6 with soluble IL-6R (sIL-6R) can trigger IL-6 trans-signalling in cells lacking IL-6R. Conversely, the soluble form of GP130 (sGP130) competes with its membrane form to inhibit IL-6 trans-signalling. We aimed to analyse IL-6 trans-signalling proteins in the airways of subjects after an allergen challenge. We used a model of segmental bronchoprovocation with an allergen (SBP-Ag) in human subjects with allergy. Before and 48h after SBP-Ag, bronchoalveolar lavages (BALs) allowed for the analysis of proteins in BAL fluids (BALFs) by ELISA, and membrane proteins on the surface of BAL cells by flow cytometry. In addition, we performed RNA sequencing (RNA-seq) and used proteomic data to further inform on the expression of the IL-6R subunits by eosinophils, bronchial epithelial cells and lung fibroblasts. Finally, we measured the effect of IL-6 trans-signalling on bronchial fibroblasts, in vitro. IL-6, sIL-6R, sGP130 and the molar ratio of sIL-6R/sGP130 increased in the airways after SBP-Ag, suggesting the potential for enhanced IL-6 trans-signalling activity. BAL lymphocytes, monocytes and eosinophils displayed IL-6R on their surface and were all possible providers of sIL-6R, whereas GP130 was highly expressed in bronchial epithelial cells and lung fibroblasts. Finally, bronchial fibroblasts activated by IL-6 trans-signalling produced enhanced amounts of the chemokine, MCP-1 (CCL2). After a bronchial allergen challenge, we found augmentation of the elements of IL-6 trans-signalling. Allergen-induced IL-6 trans-signalling activity can activate fibroblasts to produce chemokines that can further enhance inflammation and lung dysfunction.

Honing in on the effectors of oxidative stress in the asthmatic lung: oxidised phosphatidylcholines

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Evaluation of House Dust Mite Dose Response in the Environmental Exposure Unit

As a controlled allergen challenge facility, the Environmental Exposure Unit (EEU) is a valuable model to investigate allergic rhinitis (AR). While seasonal allergens have been previously studied in the EEU, here we sought to evaluate the clinical manifestations of house dust mite (HDM)-induced AR in the HDM-EEU.

A Single-Dose of REGN1908-1909 Reduced Bronchoconstriction in Cat-Allergic Subjects with Mild Asthma for up to 3 months following a controlled cat allergen challenge: A Phase 2, Randomized, Double-Blind, Placebo-Controlled Study

To evaluate the efficacy of a novel anti-Fel d 1 monoclonal antibody cocktail (REGN1908-1909) in reducing acute bronchoconstriction, defined as early asthmatic response (EAR), in cat-allergic subjects with mild asthma (NCT03838731).

Staphylococcus aureus Carriage and Differential Cell Counts in Nasal Lavages of Ragweed-Induced Allergic Rhinitis Using Nasal Allergen Challenge

The impact of Staphylococcus aureus nasal carriage on allergic rhinitis(AR) is essentially unknown. The current study sought to compare the differential cell counts in nasal lavage(NL) between (1) Ragweed(RW)-allergic and non-allergic participants and (2) S. aureus carriers and non-carriers using the nasal allergen challenge(NAC) model.

Accumulation of Allergen-specific CD4+ T cells in the Airways after Segmental Allergen Challenge is Associated with Reduced Expression of T2 Chemoattractant Receptors on Allergen-specific CD4+ T cells in Blood

Accumulation of Allergen-specific CD4+ T cells in the Airways after Segmental Allergen Challenge is Associated with Reduced Expression of T2 Chemoattractant Receptors on Allergen-specific CD4+ T cells in Blood

Changes in lung immune cell infiltrates after electric field treatment in mice

Exogenous electric fields are currently used in human therapy in a number of contexts. Interestingly, electric fields have also been shown to alter migration and function of immune cells, suggesting the potential for electric field-based immune therapy. Little is known as to the effect of electric field treatment (EFT) on the lung. To determine if EFT associates with changes in lung immune cell infiltration, we used a mouse model with varying methods of EFT application and measured cells and soluble mediators using flow cytometry and cytokine/chemokine multiplex. EFT was associated with a transient increase in lung neutrophils and decrease in eosinophils in naïve mice within 2 h of treatment, accompanied by an increase in IL-6 levels. In order to test whether EFT could alter eosinophil/neutrophil recruitment in a relevant disease model, a mouse model of allergic airway inflammation was used. Four EFT doses in allergen-sensitized mice resulted in increased neutrophil and reduced eosinophil infiltrates following allergen challenge, suggesting a durable change in inflammation by EFT. Mice with allergic inflammation were analyzed by flexiVent for measures of lung function. EFT-treated mice had increased inspiratory capacity and other measures of lung function were not diminished. These data suggest EFT may be used to manipulate immune cell infiltration in the lung without affecting lung function.

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.

Integrating Proteomes for Lung Tissues and Lavage Reveals Pathways That Link Responses in Allergen-Challenged Mice

To capture interplay between biological pathways, we analyzed the proteome from matched lung tissues and bronchoalveolar lavage fluid (BALF) of individual allergen-naïve and house dust mite (HDM)-challenged BALB/c mice, a model of allergic asthma. Unbiased label-free liquid chromatography with tandem mass spectrometry (LC-MS/MS) analysis quantified 2675 proteins from tissues and BALF of allergen-naïve and HDM-exposed mice. In comparing the four datasets, we found significantly greater diversity in proteins between lung tissues and BALF than in the changes induced by HDM challenge. The biological pathways enriched after allergen exposure were compartment-dependent. Lung tissues featured innate immune responses and oxidative stress, while BALF most strongly revealed changes in metabolism. We combined lung tissues and BALF proteomes, which principally highlighted oxidation reduction (redox) pathways, a finding influenced chiefly by the lung tissue dataset. Integrating lung and BALF proteomes also uncovered new proteins and biological pathways that may mediate lung tissue and BALF interactions after allergen challenge, for example, B-cell receptor signaling. We demonstrate that enhanced insight is fostered when different biological compartments from the lung are investigated in parallel. Integration of proteomes from lung tissues and BALF compartments reveals new information about protein networks in response to environmental challenge and interaction between intracellular and extracellular processes.