Model Of Allergic Airway Disease Research Articles

Pam3CSK4 Induces B Cells to Release FasL+ Killer Exosomes That Suppress Allergic Asthma

Abstract We have previously shown that mouse B lymphocytes can express Fas ligand (FasL) on their cell surfaces and can kill peptide antigen-specific T helper cells. In humans, FasL protein produced by human B cells was exclusively found intracellularly on the surface of exosomes. In the current study we determined that mouse B cells are a major source of FasL+ exosomes, as well as contributing significantly to the Granzme B+ exosome pool. The major B cell subsets in the spleen and lungs that produced FasL and Granzyme B were marked by CD9 and/or CD107a expression. We tested a panel of toll-like receptor ligands in combination with anti-IgM and CD40 ligand for their ability to activate killer function in purified mouse B cells. The TLR2 ligand Pam3CSK4 was a potent inducer of surface FasL expression and FasL+ exosome release from B cells. Using a mouse model of OVA-specific allergic airways disease, we showed that transfer of Pam3CSK4-induced B cell-derived exosomes diminished total numbers of eosinophils and CD4+ T cells in the bronchoalveolar lavage fluid (BAL). Lung TH cell apoptosis was higher in the mice that received exosome transfer. Lung cells produced less IL-4 and IL-5 upon restimulation with OVA in the exosome-treated mice. Effects of Pam3CSK4-induced exosomes on BAL cell infiltration, TH cell apoptosis and cytokine production were all diminished when exosomes were derived from mice with a functional mutation of the FasL gene. These data indicate a potential role for B cell-derived killer exosomes in control of allergic airways disease.

IFNβ inhibits the development of allergen tolerance and is conducive to the development of asthma on subsequent allergen exposure.

Asthma is a chronic disease affecting up to 10% of the Australian population for which medical treatment is solely aimed at relief of symptoms rather than prevention of disease. Evidence from animal and human studies demonstrates a strong link between viral respiratory infections, atopy and the development of asthma. Type I IFNs include IFNα and IFNβ, with subtype expression tailored toward the specific viral infection. We hypothesized that exposure to type I IFNs and allergen may interfere with the healthy response to innocuous airway antigen exposure. In this study, we use an ovalbumin (OVA)-induced BALB/c model of experimental allergic airways disease, where pre-exposure of the airways to OVA is protective against allergen sensitization, leading to allergen tolerance. We investigated airways pre-exposure with OVA and type I IFNs on development of allergic airways disease. We demonstrate restoration of allergic airways disease on pre-exposure with allergen and IFNβ, and not IFNα. Dysfunction in tolerance led to changes in dendritic cell antigen capture/traffic, T-cell and B-cell responses. Furthermore, exposure to IFNβ with ongoing allergen exposure led to the development of hallmark asthma features, including OVA-specific IgE and airways eosinophilia. Data indicate a role for IFNβ in linking viral infection and allergy.

Unique mechanisms of connective tissue growth factor regulation in airway smooth muscle in asthma: Relationship with airway remodelling.

Neovascularization, increased basal membrane thickness and increased airway smooth muscle (ASM) bulk are hallmarks of airway remodelling in asthma. In this study, we examined connective tissue growth factor (CTGF) dysregulation in human lung tissue and animal models of allergic airway disease. Immunohistochemistry revealed that ASM cells from patients with severe asthma (A) exhibited high expression of CTGF, compared to mild and non‐asthmatic (NA) tissues. This finding was replicated in a sheep model of allergic airways disease. In vitro, transforming growth factor (TGF)‐β increased CTGF expression both in NA‐ and A‐ASM cells but the expression was higher in A‐ASM at both the mRNA and protein level as assessed by PCR and Western blot. Transfection of CTGF promoter‐luciferase reporter constructs into NA‐ and A‐ASM cells indicated that no region of the CTGF promoter (−1500 to +200 bp) displayed enhanced activity in the presence of TGF‐β. However, in silico analysis of the CTGF promoter suggested that distant transcription factor binding sites may influence CTGF promoter activation by TGF‐β in ASM cells. The discord between promoter activity and mRNA expression was also explained, in part, by differential post‐transcriptional regulation in A‐ASM cells due to enhanced mRNA stability for CTGF. In patients, higher CTGF gene expression in bronchial biopsies was correlated with increased basement membrane thickness indicating that the enhanced CTGF expression in A‐ASM may contribute to airway remodelling in asthma.

House dust mite induced allergic airway disease is attenuated in CD11ccreIL-4R\u03b1\u2212/l\xb0x mice

The precise mechanisms leading to development of T helper type (Th)2-driven allergic responses are unknown. We aimed to determine how IL-4 receptor alpha (IL-4Rα) signaling on CD11c+ cells influences allergen-induced Th2 responses in mice. CD11ccreIL-4Rα−/l°x mice, deficient in IL-4Rα on dendritic cells and alveolar macrophages, were compared to IL-4Rα−/l°x littermate controls in models of allergic airway disease induced by OVA/alum, OVA alone or house dust mite. Cytokine responses, eosinophil and neutrophil infiltration into the lungs, airway hyperreactivity and mucus hypersecretion were evaluated after allergen challenge. In the OVA/alum model, CD11ccreIL-4Rα−/lox mice had similar airway hyperreactivity, eosinophil infiltration, Th2-type cytokine production and mucus hypersecretion to littermate controls. When alum was omitted during sensitization, CD11ccreIL-4Rα−/lox mice had similar airway hyperreactivity and mucus secretion but reduced Th2-type cytokine production and eosinophils, suggesting alum overrides the requirement for IL-4Rα signaling on CD11c+ cells in enhancing Th2-type responses. In the house dust mite model, CD11ccreIL-4Rα−/lox mice showed similar mucus secretion, but reduced Th2 responses, eosinophils, neutrophils and airway hyperreactivity, unlike previously tested LysMcreIL-4Rα−/lox mice, which lack IL-4Rα on alveolar macrophages but not on dendritic cells. Therefore, our results indicate that IL-4Rα signaling on dendritic cells promotes allergen-induced Th2 responses and eosinophil infiltration into the lung.

The independent effects of vitamin D deficiency and house dust mite exposure on lung function are sex-specific

Vitamin D deficiency is increasing around the world and has been associated with the development of asthma. This study aims to evaluate the effect of dietary vitamin D deficiency at different life stages on lung function using a murine model of allergic airways disease. BALB/c mice were challenged intranasally with HDM or saline alone for 10 days. Twenty four hours after the last challenge, mice were anesthetized and lung function was measured using the forced oscillation technique (FOT). Mice were euthanized for assessment of inflammation in the bronchoalveolar lavage (BAL) and total collagen content in lung homogenates by ELISA. Vitamin D deficiency impaired lung function in both male and female mice, increasing tissue damping and elastance, however had no effect on HDM induced inflammation. The impact of vitamin D deficiency was more evident in females. HDM also decreased airway distensibility, but only in females and this response was not altered by vitamin D deficiency. Our data suggest that vitamin D deficiency and HDM exposure have independent effects on lung mechanics and that females are more susceptible to these effects. Vitamin D deficiency may exacerbate lung function deficits by having a direct, but independent, effect on parenchymal mechanics.

Plasmacytoid dendritic cells drive acute asthma exacerbations

Although acute exacerbations, mostly triggered by viruses, account for the majority of hospitalizations in asthmatic patients, there is still very little known about the pathophysiologic mechanisms involved. Plasmacytoid dendritic cells (pDCs), prominent cells of antiviral immunity, exhibit proinflammatory or tolerogenic functions depending on the context, yet their involvement in asthma exacerbations remains unexplored. We sought to investigate the role of pDCs in allergic airway inflammation and acute asthma exacerbations. Animal models of allergic airway disease (AAD) and virus-induced AAD exacerbations were used to dissect pDC function invivo and unwind the potential mechanisms involved. Sputum from asthmatic patients with stable disease or acute exacerbations was further studied to determine the presence of pDCs and correlation with inflammation. pDCs were key mediators of the immunoinflammatory cascade that drives asthma exacerbations. In animal models of AAD and rhinovirus-induced AAD exacerbations, pDCs were recruited to the lung during inflammation and migrated to the draining lymph nodes to boost TH2-mediated effector responses. Accordingly, pDC depletion after allergen challenge or during rhinovirus infection abrogated exacerbation of inflammation and disease. Central to this process was IL-25, which was induced by allergen challenge or rhinovirus infection and conditioned pDCs for proinflammatory function. Consistently, in asthmatic patients pDC numbers were markedly increased during exacerbations and correlated with the severity of inflammation and the risk for asthma attacks. Our studies uncover a previously unsuspected role of pDCs in asthma exacerbations with potential diagnostic and prognostic implications. They also propose the therapeutic targeting of pDCs and IL-25 for the treatment of acute asthma.

Intranasal administration of mesenchymoangioblast‐derived mesenchymal stem cells abrogates airway fibrosis and airway hyperresponsiveness associated with chronic allergic airways disease

Structural changes known as airway remodeling (AWR) characterize chronic/severe asthma and contribute to lung dysfunction. Thus, we assessed the in vivo efficacy of induced pluripotent stem cell and mesenchymoangioblast-derived mesenchymal stem cells (MCA-MSCs) on AWR in a murine model of chronic allergic airways disease (AAD)/asthma. Female Balb/c mice were subjected to a 9-wk model of ovalbumin (Ova)-induced chronic AAD and treated intravenously or intranasally with MCA-MSCs from weeks 9 to 11. Changes in airway inflammation (AI), AWR, and airway hyperresponsiveness (AHR) were assessed. Ova-injured mice presented with AI, goblet cell metaplasia, epithelial thickening, increased airway TGF-β1 levels, subepithelial myofibroblast and collagen accumulation, total lung collagen concentration, and AHR (all P < 0.001 vs. uninjured control group). Apart from epithelial thickness, all other parameters measured were significantly, although not totally, decreased by intravenous delivery of MCA-MSCs to Ova-injured mice. In comparison, intranasal delivery of MCA-MSCs to Ova-injured mice significantly decreased all parameters measured (all P < 0.05 vs. Ova group) and, most notably, normalized aberrant airway TGF-β1 levels, airway/lung fibrosis, and AHR to values measured in uninjured animals. MCA-MSCs also increased collagen-degrading gelatinase levels. Hence, direct delivery of MCA-MSCs offers great therapeutic benefit for the AWR and AHR associated with chronic AAD.-Royce, S. G., Rele, S., Broughton, B. R. S., Kelly, K., Samuel, C. S. Intranasal administration of mesenchymoangioblast-derived mesenchymal stem cells abrogates airway fibrosis and airway hyperresponsiveness associated with chronic allergic airways disease.

Dendritic cells conditioned by activin A–induced regulatory T cells exhibit enhanced tolerogenic properties and protect against experimental asthma

Previously, we demonstrated that regulatory T(Treg) cells induced by the cytokine activin-A suppress TH2-mediated allergic responses and linked airway disease. Still, the effects of activin-A-induced regulatory T (Act-A-iTreg) cells on the regulation of dendritic cell (DC)-driven allergic inflammation remain elusive. Here we investigated whether Act-A-iTreg cells can modulate DC responses and endow them with enhanced tolerogenic functions. Using adoptive cell transfer studies in mouse models of allergic airway disease, we examined the effects of Act-A-iTreg cells on DC phenotype, maturation status, and TH2 cell priming potential. Genome-wide gene expression profiling characterized the transcriptional networks induced in tolerogenic DCs by Act-A-iTreg cells. The ability of DCs conditioned by Act-A-iTreg cells (Act-A-iTreg cell-modified DCs) to protect against experimental asthma, and the mechanisms involved were also explored. Act-A-iTreg cell-modified DCs exhibited a significantly impaired capacity to uptake allergen and stimulate naive and TH2 effector responses on allergen stimulation invivo accompanied by markedly attenuated inflammatory cytokine release in response to LPS. Gene-profiling studies revealed that Act-A-iTreg cells dampened crucial TH2-skewing transcriptional networks in DCs. Administration of Act-A-iTreg cell-modified DCs ameliorated cardinal asthma manifestations in preventive and therapeutic protocols through generation of strongly suppressive forkhead box P3+ Treg cells. Finally, programed death protein 1/programmed death ligand 1 signaling pathways were essential in potentiating the generation of DCs with tolerogenic properties by Act-A-iTreg cells. Our studies reveal that Act-A-iTreg cells instruct the generation of a highly effective immunoregulatory circuit encompassing tolerogenic DCs and forkhead box P3+ Treg cells that could be targeted for the design of novel immunotherapiesfor allergic disorders.

IL-15-deficient mice develop enhanced airway allergic responses in a mouse model of allergic airway disease

Abstract Interleukin-15 is a pleiotropic cytokine critical for the development of multiple hematopoietic lineages. Mice lacking IL-15 have selective defects in populations of several pro-allergic immune cells including natural killer (NK) cells, NKT cells, and memory CD8+T cells. We therefore hypothesized that IL-15−/− mice will have reduced inflammatory responses during the development of allergic airway disease (AAD). Here we report that IL-15−/− mice developed enhanced allergic responses in an OVA-induced model of AAD. C57BL/6 wild-type (WT) and IL-15−/− mice were sensitized and challenged with ovalbumin (OVA) and the development of AAD was ascertained. In the absence of IL-15, OVA-challenged mice exhibited enhanced bronchial eosinophilic inflammation, elevated IL-13 production, and severe lung histopathology in comparison with WT mice. In addition, increased numbers of CD4+T and B cells in the spleens and broncholaveolar lavage were also observed. Examination of OVA-challenged IL-15R−/− animals revealed a similar phenotype resulting in enhanced airway eosinophilia compared to WT mice. Adoptive transfer of splenic CD8+T cells from OVA-sensitized WT mice suppressed the enhancement of eosinophilia in IL-15−/− animals to levels observed in WT mice, but had no further effects. These data demonstrate that mice with an endogenous IL-15 deficiency are susceptible to the development of severe, enhanced Th2-mediated AAD. Furthermore, the development of disease as well as allergen-specific Th2 responses occurs despite deficiencies in several IL-15-dependent cell types including NK and NKT cells, suggesting that these cells or their subsets are dispensable for the induction of AAD in IL-15-deficient mice.

Intravenous immunoglobulin G (IVIg) concurrently promotes Th2 and T regulatory cell development while abrogating Th17 cells

Abstract We aimed to further elucidate the mechanisms by which IVIg modulates the balance between Th17, Th2, and Treg. Using an OVA-driven murine model of allergic airway disease exhibiting a mixed Th17/Th2 response, we examined lung inflammation by H &amp; E staining and histology, and performed cellular phenotyping of lung homogenates using flow cytometry. In addition, we assessed cytokine production by intracellular staining and ELISA, and performed co-culture studies. IVIg treatment reduced lung inflammation and diminished the frequency and absolute number of lung Th17 cells, IL-17A production, and neutrophilia. In contrast, the frequency and absolute number of eosinophils and basophils was increased, while IgE production was abrogated in the IVIg group. This was accompanied by a significant increase in IL-13+ and IL-5+ T cells and in IL-5 concentration in the BAL. The frequency of Foxp3+IL10+ + cells within CD4+ T cells (1.43 % ± .061 vs 2.57 % ± .30, p &amp;lt; .01) and IL-10 secreting dendritic cells (0.28 % ± 0.060 vs 1.20 % ± 0.27, p &amp;lt; .05) was augmented. We also observed an increase in the frequency of CD206 expressing macrophages, or M2 macrophages, in the IVIg group (10.81% ± 2.21 vs 19.80 % ± 1.81, p &amp;lt; .01). Using an in vitro DC: T cell co-culture, IVIg-treated DC inhibited IL-17A production by T cells and increased IL-13 production and Treg differentiation. In conclusion, IVIg modulates Th2/Th17/Treg balance by favoring Treg and Th2 cells while inhibiting Th17 cells. Furthermore, IVIg renders both dendritic cells and macrophages tolerogenic.

Impaired induction of Slc26a4 promotes respiratory acidosis and severe, steroid-resistant asthma

Abstract CO2 produced by systemic cellular respiration is hydrated into carbonic acid (H2CO3) that dissociates into H+ and HCO3−. These are transported in the plasma to the lungs where HCO3− is converted back into H2CO3 and CO2 that are expelled through breathing. Evidence suggests that dysfunction of the mechanisms that govern these processes may result in the development of respiratory acidosis (RA) and the cardinal features of severe, steroid-resistant (SSR) asthma. Reduced lung function, which occurs in SSR asthma, impairs removal of volatile H2CO3 and CO2, resulting in acid accumulation and increased arterial PaCO2. Patients with severe asthma often develop complications from increased PaCO2, which skews the PaCO2/HCO3− ratio resulting in increased H+ concentration and reduced pH. We developed three mouse models of respiratory infection and ovalbumin-induced SSR allergic airways disease (SSRAAD) that are highly representative of SSR asthma in humans. We used these models to show a role for impaired homeostatic acid-base balance in SSR asthma. All three infections suppress the induction of the expression of the chloride (Cl−)/HCO− pump, Slc26a4, in the airway mucosa in AAD. Importantly, SSRAAD is associated with increased levels of free H+ ions in bronchoalveolar lavage fluid. Administration of Slc26a4-specific siRNA in steroid-sensitive AAD, which mimics the effect of decreased Slc26a4 responses in SSRAAD, induced RA and steroid-resistant airway inflammation and AHR. Importantly, treatment of RA with NaHCO3 during infection-induced SSRAAD suppressed steroid-resistant AHR. Thus, we have identified a previously unrecognised role for deficient Slc26a4 responses that result in the development of RA and the pathogenesis of SSRAAD.

Peripherally Generated Foxp3+ Regulatory T Cells Mediate the Immunomodulatory Effects of IVIg in Allergic Airways Disease.

IVIg is widely used as an immunomodulatory therapy. We have recently demonstrated that IVIg protects against airway hyperresponsiveness (AHR) and inflammation in mouse models of allergic airways disease (AAD), associated with induction of Foxp3+ regulatory T cells (Treg). Using mice carrying a DTR/EGFP transgene under the control of the Foxp3 promoter (DEREG mice), we demonstrate in this study that IVIg generates a de novo population of peripheral Treg (pTreg) in the absence of endogenous Treg. IVIg-generated pTreg were sufficient for inhibition of OVA-induced AHR in an Ag-driven murine model of AAD. In the absence of endogenous Treg, IVIg failed to confer protection against AHR and airway inflammation. Adoptive transfer of purified IVIg-generated pTreg prior to Ag challenge effectively prevented airway inflammation and AHR in an Ag-specific manner. Microarray gene expression profiling of IVIg-generated pTreg revealed upregulation of genes associated with cell cycle, chromatin, cytoskeleton/motility, immunity, and apoptosis. These data demonstrate the importance of Treg in regulating AAD and show that IVIg-generated pTreg are necessary and sufficient for inhibition of allergen-induced AAD. The ability of IVIg to generate pure populations of highly Ag-specific pTreg represents a new avenue to study pTreg, the cross-talk between humoral and cellular immunity, and regulation of the inflammatory response to Ags.

Bromodomain and Extra-Terminal Protein Inhibition Attenuates Neutrophil-dominant Allergic Airway Disease

Atopic asthma is a prevalent respiratory disease that is characterized by inflammation, mucus hypersecretion, and airway hyperresponsiveness. The complexity of this heterogeneous disorder has commanded the need to better define asthma phenotypes based on underlying molecular mechanisms of disease. Although classically viewed as a type 2-regulated disease, type 17 helper T (Th17) cells are known to be influential in asthma pathogenesis, predominantly in asthmatics with neutrophilia and severe refractory disease. Bromodomain and extra-terminal domain (BET) chromatin adaptors serve as immunomodulators by directly regulating Th17 responses and Th17-mediated pathology in murine models of autoimmunity and infection. Based on this, we hypothesized that BET proteins may also play an essential role in neutrophil-dominant allergic airway disease. Using a murine model of neutrophil-dominant allergic airway disease, we demonstrate that BET inhibition limits pulmonary inflammation and alters the Th17-related inflammatory milieu in the lungs. In addition, inhibition of BET proteins improved lung function (specifically quasi-static lung compliance and tissue elastance) and reduced mucus production in airways. Overall, these studies show that BET proteins may have a critical role in asthma pathogenesis by altering type 17 inflammation, and thus interfering with BET-dependent chromatin signaling may provide clinical benefits to patients suffering from asthma.

IL-15-deficient mice develop enhanced allergic responses to airway allergen exposure.

Interleukin-15 is a pleiotropic cytokine that is critical for the development and survival of multiple haematopoietic lineages. Mice lacking IL-15 have selective defects in populations of several pro-allergic immune cells including natural killer (NK) cells, NKT cells, and memory CD8+ T cells. We therefore hypothesized that IL-15-/- mice will have reduced inflammatory responses during the development of allergic airway disease (AAD). To determine whether IL-15-/- mice have attenuated allergic responses in a mouse model of AAD. C57BL/6 wild-type (WT) and IL-15-/- mice were sensitized and challenged with ovalbumin (OVA), and the development of AAD was ascertained by examining changes in airway inflammatory responses, Th2 responses, and lung histopathology. Here, we report that IL-15-/- mice developed enhanced allergic responses in an OVA-induced model of AAD. In the absence of IL-15, OVA-challenged mice exhibited enhanced bronchial eosinophilic inflammation, elevated IL-13 production, and severe lung histopathology in comparison with WT mice. In addition, increased numbers of CD4+ T and B cells in the spleens and bronchoalveolar lavage (BAL) were also observed. Examination of OVA-challenged IL-15Rα-/- animals revealed a similar phenotype resulting in enhanced airway eosinophilia compared to WT mice. Adoptive transfer of splenic CD8+ T cells from OVA-sensitized WT mice suppressed the enhancement of eosinophilia in IL-15-/- animals to levels observed in WT mice, but had no further effects. These data demonstrate that mice with an endogenous IL-15 deficiency are susceptible to the development of severe, enhanced Th2-mediated AAD, which can be regulated by CD8+ T cells. Furthermore, the development of disease as well as allergen-specific Th2 responses occurs despite deficiencies in several IL-15-dependent cell types including NK, NKT, and γδ T cells, suggesting that these cells or their subsets are dispensable for the induction of AAD in IL-15-deficient mice.

Aspergillus in chronic lung disease: Modeling what goes on in the airways.

Aspergillus species cause a range of respiratory diseases in humans. While immunocompromised patients are at risk for the development of invasive infection with these opportunistic molds, patients with underlying pulmonary disease can develop chronic airway infection with Aspergillus species. These conditions span a range of inflammatory and allergic diseases including Aspergillus bronchitis, allergic bronchopulmonary aspergillosis, and severe asthma with fungal sensitization. Animal models are invaluable tools for the study of the molecular mechanism underlying the colonization of airways by Aspergillus and the host response to these non-invasive infections. In this review we summarize the state-of-the-art with respect to the available animal models of noninvasive and allergic Aspergillus airway disease; the key findings of host-pathogen interaction studies using these models; and the limitations and future directions that should guide the development and use of models for the study of these important pulmonary conditions.

A pathogenic role for the integrin CD103 in experimental allergic airways disease.

The integrin CD103 is the α E chain of integrin α E β 7 that is important in the maintenance of intraepithelial lymphocytes and recruitment of T cells and dendritic cells (DC) to mucosal surfaces. The role of CD103 in intestinal immune homeostasis has been well described, however, its role in allergic airway inflammation is less well understood. In this study, we used an ovalbumin (OVA)‐induced, CD103‐knockout (KO) BALB/c mouse model of experimental allergic airways disease (EAAD) to investigate the role of CD103 in disease expression, CD4+ T‐cell activation and DC activation and function in airways and lymph nodes. We found reduced airways hyper‐responsiveness and eosinophil recruitment to airways after aerosol challenge of CD103 KO compared to wild‐type (WT) mice, although CD103 KO mice showed enhanced serum OVA‐specific IgE levels. Following aerosol challenge, total numbers of effector and regulatory CD4+ T‐cell subsets were significantly increased in the airways of WT but not CD103 KO mice, as well as a lack of DC recruitment into the airways in the absence of CD103. While total airway DC numbers, and their in vivo allergen capture activity, were essentially normal in steady‐state CD103 KO mice, migration of allergen‐laden airway DC to draining lymph nodes was disrupted in the absence of CD103 at 24 h after aerosol challenge. These data support a role for CD103 in the pathogenesis of EAAD in BALB/c mice through local control of CD4+ T cell and DC subset recruitment to, and migration from, the airway mucosa during induction of allergic inflammation.

A novel microbe-based treatment that attenuates the inflammatory profile in a mouse model of allergic airway disease.

There is an unmet need for effective new and innovative treatments for asthma. It is becoming increasingly evident that bacterial stimulation can have beneficial effects at attenuating allergic airway disease through immune modulation. Our aim was to test the ability of a novel inactivated microbe-derived therapeutic based on Klebsiella (KB) in a model of allergic airway disease in mice. BALB/c mice were exposed intranasally to house dust mite (HDM) for two weeks. Mice were treated prophylactically via subcutaneous route with either KB or placebo for one week prior to HDM exposure and throughout the two week exposure period. 24 hours after the last exposure, lungs were analysed for inflammatory cell infiltrate, gene expression, cytokine levels, goblet cell metaplasia, and serum was analysed for allergen-specific serum IgE levels. HDM exposed mice developed goblet cell hyperplasia, elevated allergen-specific serum IgE, airway eosinophilia, and a concomitant increase in TH2 cytokines including IL-4, IL-13 and IL-5. Treatment with KB attenuated HDM-mediated airway eosinophilia, total bronchoalveolar lavage (BAL) cell numbers, BAL TH2 cytokine production, and goblet cell metaplasia. Our prophylactic intervention study illustrates the potential of subcutaneous treatment with bacterial derived biologics as a promising approach for allergic airway disease treatment.

Intraepithelial neutrophils in pediatric severe asthma are associated with better lung function

BackgroundNeutrophils and IL-17A have been linked mechanistically in models of allergic airways disease and have been associated with asthma severity. However, their role in pediatric asthma is unknown.ObjectivesWe sought to investigate the role of neutrophils and the IL-17A pathway in mediating pediatric severe therapy-resistant asthma (STRA).MethodsChildren with STRA (n = 51; age, 12.6 years; range, 6-16.3 years) and controls without asthma (n = 15; age, 4.75 years; range, 1.6-16 years) underwent clinically indicated fiberoptic bronchoscopy, bronchoalveolar lavage (BAL), endobronchial brushings, and biopsy. Neutrophils, IL-17A, and IL-17RA–expressing cells and levels of IL-17A and IL-22 were quantified in BAL and biopsies and related to clinical features. Primary bronchial epithelial cells were stimulated with IL-17A and/or IL-22, with and without budesonide.ResultsChildren with STRA had increased intraepithelial neutrophils, which positively correlated with FEV1 %predicted (r = 0.43; P = .008). Neutrophilhigh patients also had better symptom control, despite lower dose maintenance inhaled steroids. Submucosal neutrophils were not increased in children with STRA. Submucosal and epithelial IL-17A–positive cells and BAL IL-17A and IL-22 levels were similar in children with STRA and controls. However, there were significantly more IL-17RA–positive cells in the submucosa and epithelium in children with STRA compared with controls (P = .001). Stimulation of primary bronchial epithelial cells with IL-17A enhanced mRNA expression of IL-17RA and increased release of IL-8, even in the presence of budesonide.ConclusionsA proportion of children with STRA exhibit increased intraepithelial airway neutrophilia that correlated with better lung function. STRA was also characterized by increased airway IL-17RA expression. These data suggest a potential beneficial rather than adverse role for neutrophils in pediatric severe asthma pathophysiology.

Plethysmography Phenotype QTL in Mice Before and After Allergen Sensitization and Challenge

Allergic asthma is common airway disease that is characterized in part by enhanced airway constriction in response to nonspecific stimuli. Genome-wide association studies have identified multiple loci associated with asthma risk in humans, but these studies have not accounted for gene–environment interactions, which are thought to be important factors in asthma. To identify quantitative trait loci (QTL) that regulate responses to a common human allergen, we applied a house dust mite mouse (HDM) model of allergic airway disease (AAD) to 146 incipient lines of the Collaborative Cross (CC) and the CC founder strains. We employed a longitudinal study design in which mice were phenotyped for response to the bronchoconstrictor methacholine both before and after HDM sensitization and challenge using whole body plethysmography (WBP). There was significant variation in methacholine responsiveness due to both strain and HDM treatment, as reflected by changes in the WBP parameter enhanced pause. We also found that distinct QTL regulate baseline [chromosome (Chr) 18] and post-HDM (Chr 19) methacholine responsiveness and that post-HDM airway responsiveness was correlated with other features of AAD. Finally, using invasive measurements of airway mechanics, we tested whether the Chr 19 QTL affects lung resistance per se using C57BL/6J mice and a consomic strain but found that QTL haplotype did not affect lung resistance. We conclude that aspects of baseline and allergen-induced methacholine responsiveness are associated with genetic variation, and that robust detection of airway resistance QTL in genetically diverse mice will be facilitated by direct measurement of airway mechanics.

Differential Expression and Function of PDE8 and PDE4 in Effector T cells: Implications for PDE8 as a Drug Target in Inflammation.

Abolishing the inhibitory signal of intracellular cAMP is a prerequisite for effector T (Teff) cell function. The regulation of cAMP within leukocytes critically depends on its degradation by cyclic nucleotide phosphodiesterases (PDEs). We have previously shown that PDE8A, a PDE isoform with 40–100-fold greater affinity for cAMP than PDE4, is selectively expressed in Teff vs. regulatory T (Treg) cells and controls CD4+ Teff cell adhesion and chemotaxis. Here, we determined PDE8A expression and function in CD4+ Teff cell populations in vivo. Using magnetic bead separation to purify leukocyte populations from the lung draining hilar lymph node (HLN) in a mouse model of ovalbumin-induced allergic airway disease (AAD), we found by Western immunoblot and quantitative (q)RT-PCR that PDE8A protein and gene expression are enhanced in the CD4+ T cell fraction over the course of the acute inflammatory disease and recede at the late tolerant non-inflammatory stage. To evaluate PDE8A as a potential drug target, we compared the selective and combined effects of the recently characterized highly potent PDE8-selective inhibitor PF-04957325 with the PDE4-selective inhibitor piclamilast (PICL). As previously shown, PF-04957325 suppresses T cell adhesion to endothelial cells. In contrast, we found that PICL alone increased firm T cell adhesion to endothelial cells by ~20% and significantly abrogated the inhibitory effect of PF-04957325 on T cell adhesion by over 50% when cells were co-exposed to PICL and PF-04957325. Despite its robust effect on T cell adhesion, PF-04957325 was over two orders of magnitude less efficient than PICL in suppressing polyclonal Teff cell proliferation, and showed no effect on cytokine gene expression in these cells. More importantly, PDE8 inhibition did not suppress proliferation and cytokine production of myelin-antigen reactive proinflammatory Teff cells in vivo and in vitro. Thus, targeting PDE8 through PF-04957325 selectively regulates Teff cell interactions with endothelial cells without marked immunosuppression of proliferation, while PDE4 inhibition has partially opposing effects. Collectively, our data identify PF-04957325 as a novel function-specific tool for the suppression of Teff cell adhesion and indicate that PDE4 and PDE8 play unique and non-redundant roles in the control of Teff cell functions.