Mouse Model Of Airway Disease Research Articles

Optogenetic Control of Airway Cholinergic Neurons In Vivo.

Dysregulation of airway nerves leads to airway hyperreactivity, a hallmark of asthma. Although changes to nerve density and phenotype have been described in asthma, the relevance of these changes to nerve function has not been investigated due to anatomical limitations where afferent and efferent nerves run in the same nerve trunk, making it difficult to assess their independent contributions. We developed a unique and accessible system to activate specific airway nerves to investigate their function in mouse models of airway disease. We describe a method to specifically activate cholinergic neurons using light, resulting in immediate, measurable increases in airway inflation pressure and decreases in heart rate. Expression of light-activated channelrhodopsin 2 in these neurons is governed by Cre expression under the endogenous choline acetyltransferase promoter, and we describe a method to decrease variability in channelrhodopsin expression in future experiments. Optogenetic activation of specific subsets of airway neurons will be useful for studying the functional relevance of other observed changes, such as changes to nerve morphology and protein expression, across many airway diseases, and may be used to study the function of subpopulations of autonomic neurons in lungs and other organs.

IL13 activates autophagy to regulate secretion in airway epithelial cells

ABSTRACTCytokine modulation of autophagy is increasingly recognized in disease pathogenesis, and current concepts suggest that type 1 cytokines activate autophagy, whereas type 2 cytokines are inhibitory. However, this paradigm derives primarily from studies of immune cells and is poorly characterized in tissue cells, including sentinel epithelial cells that regulate the immune response. In particular, the type 2 cytokine IL13 (interleukin 13) drives the formation of airway goblet cells that secrete excess mucus as a characteristic feature of airway disease, but whether this process is influenced by autophagy was undefined. Here we use a mouse model of airway disease in which IL33 (interleukin 33) stimulation leads to IL13-dependent formation of airway goblet cells as tracked by levels of mucin MUC5AC (mucin 5AC, oligomeric mucus/gel forming), and we show that these cells manifest a block in mucus secretion in autophagy gene Atg16l1-deficient mice compared to wild-type control mice. Similarly, primary-culture human tracheal epithelial cells treated with IL13 to stimulate mucus formation also exhibit a block in MUC5AC secretion in cells depleted of autophagy gene ATG5 (autophagy-related 5) or ATG14 (autophagy-related 14) compared to nondepleted control cells. Our findings indicate that autophagy is essential for airway mucus secretion in a type 2, IL13-dependent immune disease process and thereby provide a novel therapeutic strategy for attenuating airway obstruction in hypersecretory inflammatory diseases such as asthma, chronic obstructive pulmonary disease, and cystic fibrosis lung disease. Taken together, these observations suggest that the regulation of autophagy by Th2 cytokines is cell-context dependent.

Protease inhibitor reduces airway response and underlying inflammation in cockroach allergen-induced murine model.

Protease(s) enhances airway inflammation and allergic cascade. In the present study, effect of a serine protease inhibitor was evaluated in mouse model of airway disease. Mice were sensitized with cockroach extract (CE) or Per a 10 and treated with 4-(2-aminoethyl) benzenesulfonyl fluoride hydrochloride (AEBSF) 1 h before or after challenge to measure airway response. Mice were euthanized to collect bronchoalveolar lavage fluid (BALF), blood, and lung to evaluate inflammation. AEBSF treatment significantly reduced the AHR in allergen-challenged mice in dose-dependent manner (p≤ 0.01). IgE (p≤0.05) and Th2 cytokines (p≤0.05) were significantly reduced in treated mice. AEBSF treatment lowered total cell (p≤0.05), eosinophil (p≤0.05), and neutrophil (p≤0.05) in BALF and lung tissue. Oxidative stress parameters were impaired on treatment in allergen-challenged mice (p≤0.05). AEBSF had therapeutic effect in allergen-induced airway resistance and underling inflammation and had potential for combination or as add-on therapy for respiratory diseases.

Intranasal administration of a combination of choline chloride, vitamin C, and selenium attenuates the allergic effect in a mouse model of airway disease

Respiratory allergic disease is an inflammatory condition accompanied by oxidative stress. Supplementation of an anti-inflammatory agent with antioxidants may have a therapeutic effect. In this study, the effects of choline chloride in combination with antioxidants were evaluated via the intranasal route in a mouse model of allergic airway disease. Balb/c mice were sensitized on days 0, 7, and 14 and challenged on days 25–30 with cockroach extract (CE) and with a booster challenge on day 38. They were treated with choline chloride (ChCl; 1mg/kg), vitamin C (Vit C; 308.33mg/kg), and selenium (Se; 1mg/kg) alone or in combination via the intranasal route on days 31, 33, 35, 37, and 39. The mice were sacrificed on day 40 to collect blood, bronchoalveolar lavage fluid, lungs, and spleen. Mice immunized with CE showed a significant increase in airway hyperresponsiveness (AHR), lung inflammation, Th2 cytokines, and the oxidative stress markers intracellular reactive oxygen species and 8-isoprostanes compared to the phosphate-buffered saline control group. A significant decrease was observed in these parameters with all the treatments (p<0.01). The highest decrease was noticed in the ChCl+Vit C+Se-treated group, with AHR decreased to the normal level. This group also showed the highest decrease in airway inflammation (p<0.001), IL-4 and IL-5 (p<0.001), IgE and IgG1 (p<0.001), NF-κB (p<0.001), and 8-isoprostane levels (p<0.001). Glutathione peroxidase activity, which was decreased significantly in CE-immunized mice, was restored to normal levels in this group (p<0.001). IL-10 level was decreased in CE-immunized mice and was restored to normal by combination treatment. The combination treatment induced FOXP3+ cells in splenocyte culture, responsible for the upregulation of IL-10. In conclusion, the combination of choline chloride, vitamin C, and selenium via the intranasal route reduces AHR, inflammation, and oxidative stress, probably by causing IL-10 production by FOXP3+ cells, and possesses therapeutic potential against allergic airway disease.

Trefoil Factor 2 Regulates Airway Remodeling in Animal Models of Asthma

Background. Epithelial denudation and metaplasia are important in the pathogenesis of airway remodeling and asthma. Trefoil factor 2 (TFF2) is a member of a family of peptides involved in protection and healing of the gastrointestinal epithelium but which are also secreted in the airway mucosa. Methods. We investigated the role of TFF2 in airway remodeling by histological and morphometric analysis of lung tissue from TFF2-deficient mice subjected to two relevant animal models of asthma: an ovalbumin model of allergic airways disease and an Aspergillus fumigatus antigen sensitization model. Results. In the ovalbumin model TFF2-deficient mice had increased goblet cell hyperplasia, but not epithelial thickening compared to wild-type (WT) counterparts. In the Aspergillus model TFF2-deficient mice also had increased goblet cell hyperplasia, and epithelial thickness was also increased in the Aspergillus-sensitized mice compared to WT controls. TFF2 deficiency was also associated with increased subepithelial collagen layer thickness. Discussion. The current study demonstrates a role of TFF2 in airway remodeling in mouse models of airway disease. Further studies into the mechanisms of action of TFF2 and its role in asthma are warranted.

A Mouse Model of Airway Disease: Oncostatin M-Induced Pulmonary Eosinophilia, Goblet Cell Hyperplasia, and Airway Hyperresponsiveness Are STAT6 Dependent, and Interstitial Pulmonary Fibrosis Is STAT6 Independent

Oncostatin M (OSM), a pleiotropic cytokine of the gp130 cytokine family, has been implicated in chronic allergic inflammatory and fibrotic disease states associated with tissue eosinophilia. Mouse (m)OSM induces airway eosinophilic inflammation and interstitial pulmonary fibrosis in vivo and regulates STAT6 activation in vitro. To determine the requirement of STAT6 in OSM-induced effects in vivo, we examined wild-type (WT) and STAT6-knockout (STAT6(-/-)) C57BL/6 mouse lung responses to transient ectopic overexpression of mOSM using an adenoviral vector (AdmOSM). Intratracheal AdmOSM elicited persistent eosinophilic lung inflammation that was abolished in STAT6(-/-) mice. AdmOSM also induced pronounced pulmonary remodeling characterized by goblet cell hyperplasia and parenchymal interstitial fibrosis. Goblet cell hyperplasia was STAT6 dependent; however, parenchymal interstitial fibrosis was not. OSM also induced airway hyperresponsiveness in WT mice that was abolished in STAT6(-/-) mice. OSM stimulated an inflammatory signature in the lungs of WT mice that demonstrated STAT6-dependent regulation of Th2 cytokines (IL-4, IL-13), chemokines (eotaxin-1/2, MCP-1, keratinocyte chemoattractant), and extracellular matrix modulators (tissue inhibitor of matrix metalloproteinase-1, matrix metalloproteinase-13), but STAT6-independent regulation of IL-4Rα, total lung collagen, collagen-1A1, -1A2 mRNA, and parenchymal collagen and α smooth muscle actin accumulation. Thus, overexpression of mOSM induces STAT6-dependent pulmonary eosinophilia, mucous/goblet cell hyperplasia, and airway hyperresponsiveness but STAT6-independent mechanisms of lung tissue extracellular matrix accumulation. These results also suggest that eosinophil or neutrophil accumulation in mouse lungs is not required for OSM-induced lung parenchymal collagen deposition and that OSM may have unique roles in the pathogenesis of allergic and fibrotic lung disease.

Prostaglandin E2 Exerts Homeostatic Regulation of Pulmonary Vascular Remodeling in Allergic Airway Inflammation

Nonselective inhibition of PG synthesis augments inflammation in mouse models of airway disease, but the roles of individual PGs are not completely clarified. To investigate the role of PGE(2) in a mouse model of airway inflammation induced by a natural allergen, we used mice lacking the critical terminal synthetic enzyme, microsomal PGE(2) synthase (mPGES)-1. Mice lacking mPGES-1 (ptges(-/-) mice) and wild-type C57BL/6 controls were challenged intranasally with low doses of an extract derived from the house dust mite Dermatophagoides farinae (Der f). The levels of PGE(2) in the bronchoalveolar lavage fluids of Der f-treated ptges(-/-) mice were approximately 80% lower than the levels in wild-type controls. Der f-induced bronchovascular eosinophilia was modestly enhanced in the ptges(-/-) mice. Both Der f-treated strains showed similar increases in serum IgE and IgG1, as well as comparable levels of Th1, Th2, and Th17 cytokine production by Der f-stimulated spleen cells. These findings indicated that mPGES-1-derived PGE(2) was not required for allergen sensitization or development of effector T cell responses. Unexpectedly, the numbers of vascular smooth muscle cells and the thickness of intrapulmonary vessels were both markedly increased in the Der f-treated ptges(-/-) mice. These vascular changes were suppressed by the administration of the stable PGE(2) analog 16, 16-dimethyl PGE(2), or of selective agonists of the E-prostanoid (EP) 1, EP2, and EP3 receptors, respectively, for PGE(2). Thus, mPGES-1 and its product, PGE(2), protect the pulmonary vasculature from remodeling during allergen-induced pulmonary inflammation, and these effects may be mediated by more than one EP receptor.

Phase contrast X-ray imaging for the non-invasive detection of airway surfaces and lumen characteristics in mouse models of airway disease

We seek to establish non-invasive imaging able to detect and measure aspects of the biology and physiology of surface fluids present on airways, in order to develop novel outcome measures able to validate the success of proposed genetic or pharmaceutical therapies for cystic fibrosis (CF) airway disease. Reduction of the thin airway surface liquid (ASL) is thought to be a central pathophysiological process in CF, causing reduced mucociliary clearance that supports ongoing infection and destruction of lung and airways. Current outcome measures in animal models, or humans, are insensitive to the small changes in ASL depth that ought to accompany successful airway therapies. Using phase contrast X-ray imaging (PCXI), we have directly examined the airway surfaces in the nasal airways and tracheas of anaesthetised mice, currently to a resolution of ∼2 μm. We have also achieved high resolution three-dimensional (3D) imaging of the small airways in mice using phase-contrast enhanced computed tomography (PC-CT) to elucidate the structure-function relationships produced by airway disease. As the resolution of these techniques improves they may permit non-invasive monitoring of changes in ASL depth with therapeutic intervention, and the use of 3D airway and imaging in monitoring of lung health and disease. Phase contrast imaging of airway surfaces has promise for diagnostic and monitoring options in animal models of CF, and the potential for future human airway imaging methodologies is also apparent.

Selective Deficiency of PGE2 Inhibits Goblet Cell Metaplasia in a Murine Model of Asthma

RationaleInhalation of Prostaglandin E2 (PGE2) inhibits the early and late phase bronchoconstrictor response to allergen in patients with asthma. Nonselective inhibition of prostaglandin synthesis augments inflammation in mouse models of airway disease, but the responsible prostaglandins have not been clarified. We investigated the role of PGE2 in a model of airways inflammation using mice deficient in a critical PGE synthase, microsomal PGE synthase-1 (mPGES-1).MethodsWe treated mPGES-1 null mice with a C57BL/6 background and wild-type controls with intranasal saline or Dermatophagoides farinea-1 (D. farinae) extract twice weekly for 3 weeks. At the end of the protocol, we analyzed histologic, physiologic, and transcriptional responses.ResultsmPGES-1 null mice treated with D. farinae exhibited significantly increased airway hyperreactivity (AHR), slightly augmented pulmonary inflammation, and normal BAL fluid eosinophilia compared to similarly treated wild-type controls. Unexpectedly, mPGES-1 null mice demonstrated 80% decrease in goblet cell metaplasia and mucus-related gene transcripts.ConclusionsLack of mPGES-1 abrogates the development of goblet cell metaplasia, despite enhanced D. farinae-induced AHR. RationaleInhalation of Prostaglandin E2 (PGE2) inhibits the early and late phase bronchoconstrictor response to allergen in patients with asthma. Nonselective inhibition of prostaglandin synthesis augments inflammation in mouse models of airway disease, but the responsible prostaglandins have not been clarified. We investigated the role of PGE2 in a model of airways inflammation using mice deficient in a critical PGE synthase, microsomal PGE synthase-1 (mPGES-1). Inhalation of Prostaglandin E2 (PGE2) inhibits the early and late phase bronchoconstrictor response to allergen in patients with asthma. Nonselective inhibition of prostaglandin synthesis augments inflammation in mouse models of airway disease, but the responsible prostaglandins have not been clarified. We investigated the role of PGE2 in a model of airways inflammation using mice deficient in a critical PGE synthase, microsomal PGE synthase-1 (mPGES-1). MethodsWe treated mPGES-1 null mice with a C57BL/6 background and wild-type controls with intranasal saline or Dermatophagoides farinea-1 (D. farinae) extract twice weekly for 3 weeks. At the end of the protocol, we analyzed histologic, physiologic, and transcriptional responses. We treated mPGES-1 null mice with a C57BL/6 background and wild-type controls with intranasal saline or Dermatophagoides farinea-1 (D. farinae) extract twice weekly for 3 weeks. At the end of the protocol, we analyzed histologic, physiologic, and transcriptional responses. ResultsmPGES-1 null mice treated with D. farinae exhibited significantly increased airway hyperreactivity (AHR), slightly augmented pulmonary inflammation, and normal BAL fluid eosinophilia compared to similarly treated wild-type controls. Unexpectedly, mPGES-1 null mice demonstrated 80% decrease in goblet cell metaplasia and mucus-related gene transcripts. mPGES-1 null mice treated with D. farinae exhibited significantly increased airway hyperreactivity (AHR), slightly augmented pulmonary inflammation, and normal BAL fluid eosinophilia compared to similarly treated wild-type controls. Unexpectedly, mPGES-1 null mice demonstrated 80% decrease in goblet cell metaplasia and mucus-related gene transcripts. ConclusionsLack of mPGES-1 abrogates the development of goblet cell metaplasia, despite enhanced D. farinae-induced AHR. Lack of mPGES-1 abrogates the development of goblet cell metaplasia, despite enhanced D. farinae-induced AHR.

Interactions between leukotriene C4 and interleukin 13 signaling pathways in a mouse model of airway disease.

Abstract Context.—During an asthmatic episode, leukotriene C4 (LTC4) and interleukin 13 (IL-13) are released into the airways and are thought to be central mediators of the asthmatic response. However, little is known about how these molecules interact or affect each other's signaling pathway. Objective.—To determine if the LTC4 and IL-13 signaling pathways interact with each other's pathways. Design.—We examined airway responsiveness, cysteinyl LTs (Cys-LTs), and Cys-LT and IL-13 receptor transcript levels in wild-type mice and in mice that were deficient in γ-glutamyl leukotrienase (an enzyme that converts LTC4 to LTD4), STAT6 (signal transducer and activator of transcription 6 [a critical molecule in IL-13 signaling]), and IL-4Rα (a subunit of the IL-13 receptor). Results.—Wild-type (C57BL/129SvEv) and γ-glutamyl leukotrienase–deficient mice showed increased airway responsiveness after intranasal instillation of IL-13; similar results were observed after intranasal instillation of IL-13 or LTC4 in a se...

Regulatory T cells as a target for induction of immune tolerance in allergy

Reduced activity of regulatory T cells has recently been described in several diseases, including allergy. This concept of an alteration in the balance between the suppression and activation of harmful T helper type 2 immunity in allergy has important potential implications for strategies to prevent and control disease. The past year has seen several important advances in analysing the determinants of this balance and models for inducing tolerance through regulatory T cells, including several different subtypes. These include the recognition that although T helper type 2 responses drive atopic sensitization, the expression of disease involves additional factors, and of a potential role for regulatory T cells in the development of neonatal tolerance. In addition to CD4CD25 regulatory T cells, experimental protocols for the induction of IL-10-producing, transforming growth factor beta and T helper type 1 regulatory T cells have been described in mouse models of airway disease. IL-6 and co-stimulation have been identified as potential determinants of the balance between the suppression and activation of allergic responses. Different strategies for inducing regulatory T cells in animal models of allergic inflammation and an improved understanding of the factors accentuating or reducing suppression have identified important questions for future translational research.