Protective Effects of Resveratrol Against Airway Hyperreactivity, Oxidative Stress, and Lung Inflammation in a Rat Pup Model of Bronchopulmonary Dysplasia.

Oxygen therapy provides an important treatment for preterm and low-birth-weight neonates, however, it has been shown that prolonged exposure to high levels of oxygen (hyperoxia) is one of the factors contributing to the development of bronchopulmonary dysplasia (BPD) by inducing lung injury and airway hyperreactivity. There is no effective therapy against the adverse effects of hyperoxia. Therefore, this study was undertaken to test the hypothesis that natural phytoalexin resveratrol will overcome hyperoxia-induced airway hyperreactivity, oxidative stress, and lung inflammation. Newborn rats were exposed to hyperoxia (fraction of inspired oxygen - FiO2>95 % O2) or ambient air (AA) for seven days. Resveratrol was supplemented either in vivo (30 mg·kg-1·day-1) by intraperitoneal administration or in vitro to the tracheal preparations in an organ bath (100 mikroM). Contractile and relaxant responses were studied in tracheal smooth muscle (TSM) using the in vitro organ bath system. To explain the involvement of nitric oxide in the mechanisms of the protective effect of resveratrol against hyperoxia, a nitric oxide synthase inhibitor - Nomega-nitro-L-arginine methyl ester (L-NAME), was administered in some sets of experiments. The superoxide dismutase (SOD) and glutathione peroxidase (GPx) activities and the tumor necrosis factor-alpha (TNF-alpha) and interleukin-1beta (IL-1beta) levels in the lungs were determined. Resveratrol significantly reduced contraction and restored the impaired relaxation of hyperoxia-exposed TSM (p<0.001). L-NAME reduced the inhibitory effect of resveratrol on TSM contractility, as well as its promotion relaxant effect (p<0.01). Resveratrol preserved the SOD and GPx activities and decreased the expression of TNF-alpha and IL-1beta in hyperoxic animals. The findings of this study demonstrate the protective effect of resveratrol against hyperoxia-induced airway hyperreactivity and lung damage and suggest that resveratrol might serve as a therapy to prevent the adverse effects of neonatal hyperoxia. Keywords: Bronchopulmonary dysplasia, Hyperoxia, Airway hyperreactivity, Resveratrol, Pro-inflammatory cytokines.

OGR1-dependent regulation of the allergen-induced asthma phenotype.

The proton-sensing receptor, ovarian cancer G protein-coupled receptor (OGR1), has been shown to be expressed in airway smooth muscle (ASM) cells and is capable of promoting ASM contraction in response to decreased extracellular pH. OGR1 knockout (OGR1KO) mice are reported to be resistant to the asthma features induced by inhaled allergen. We recently described certain benzodiazepines as OGR1 activators capable of mediating both procontractile and prorelaxant signaling in ASM cells. Here we assess the effect of treatment with the benzodiazepines lorazepam or sulazepam on the asthma phenotype in wild-type (WT) and OGR1KO mice subjected to inhaled house dust mite (HDM; Dermatophagoides pteronyssius) challenge for 3 wk. In contrast to previously published reports, both WT and OGR1KO mice developed significant allergen-induced lung inflammation and airway hyperresponsiveness (AHR). In WT mice, treatment with sulazepam (a Gs-biased OGR1 agonist), but not lorazepam (a balanced OGR1 agonist), prevented allergen-induced AHR, although neither drug inhibited lung inflammation. The protection from development of AHR conferred by sulazepam was absent in OGR1KO mice. Treatment of WT mice with sulazepam also resulted in significant inhibition of HDM-induced collagen accumulation in the lung tissue. These findings suggest that OGR1 expression is not a requirement for development of the allergen-induced asthma phenotype, but OGR1 can be targeted by the Gs-biased OGR1 agonist sulazepam (but not the balanced agonist lorazepam) to protect from allergen-induced AHR, possibly mediated via suppression of chronic bronchoconstriction and airway remodeling in the absence of effects on airway inflammation.

Cycloastragenol alleviates airway inflammation in asthmatic mice by inhibiting autophagy.

Cycloastragenol (CAG), a secondary metabolite from the roots of Astragalus zahlbruckneri, has been reported to exert anti­inflammatory effects in heart, skin and liver diseases. However, its role in asthma remains unclear. The present study aimed to investigate the effect of CAG on airway inflammation in an ovalbumin (OVA)­induced mouse asthma model. The current study evaluated the lung function and levels of inflammation and autophagy via measurement of airway hyperresponsiveness (AHR), lung histology examination, inflammatory cytokine measurement and western blotting, amongst other techniques. The results demonstrated that CAG attenuated OVA­induced AHR in vivo. In addition, the total number of leukocytes and eosinophils, as well as the secretion of inflammatory cytokines, including interleukin (IL)­5, IL­13 and immunoglobulin E were diminished in bronchoalveolar lavage fluid of the OVA­induced murine asthma model. Histological analysis revealed that CAG suppressed inflammatory cell infiltration and goblet cell secretion. Notably, based on molecular docking simulation, CAG was demonstrated to bind to the active site of autophagy­related gene 4­microtubule­associated proteins light chain 3 complex, which explains the reduced autophagic flux in asthma caused by CAG. The expression levels of proteins associated with autophagy pathways were inhibited following treatment with CAG. Taken together, the results of the present study suggest that CAG exerts an anti­inflammatory effect in asthma, and its role may be associated with the inhibition of autophagy in lung cells.

Identification of an ATP/P2X7/mast cell pathway mediating ozone-induced bronchial hyperresponsiveness.

Ozone is a highly reactive environmental pollutant with well-recognized adverse effects on lung health. Bronchial hyperresponsiveness (BHR) is one consequence of ozone exposure, particularly for individuals with underlying lung disease. Our data demonstrated that ozone induced substantial ATP release from human airway epithelia in vitro and into the airways of mice in vivo and that ATP served as a potent inducer of mast cell degranulation and BHR, acting through P2X7 receptors on mast cells. Both mast cell-deficient and P2X7 receptor-deficient (P2X7-/-) mice demonstrated markedly attenuated BHR to ozone. Reconstitution of mast cell-deficient mice with WT mast cells and P2X7-/- mast cells restored ozone-induced BHR. Despite equal numbers of mast cells in reconstituted mouse lungs, mice reconstituted with P2X7-/- mast cells demonstrated significantly less robust BHR than mice reconstituted with WT mast cells. These results support a model where P2X7 on mast cells and other cell types contribute to ozone-induced BHR.

DM-induced Hypermethylation of IR and IGF1R attenuates mast cell activation and airway responsiveness in rats.

Diabetes has been reported to modulate the airway smooth muscle reactivity and lead to attenuation of allergic inflammatory response in the lungs. In this study, we aimed to study the effect of insulin on cell activation and airway responsiveness in patients with diabetes mellitus (DM). The airway contraction in rat model groups including a non-DM group, a non-DM+INDUCTION group, a DM+INDUCTION group and a DM+INDUCTION+INSULIN group was measured to observe the effect of insulin on airway responsiveness. Radioenzymatic assay was conducted to measure the levels of histamine, and ELISA assay was conducted to measure bronchial levels of interleukin (IL)-1b, tumour necrosis factor (TNF)-a, cytokine-induced neutrophil chemoattractant (CINC)-1, P-selectin and ß-hexosaminidase. The tension in the main and intrapulmonary bronchi of DM+INDUCTION rats was lower than that of the non-DM+INDUCTION rats, whereas the treatment of insulin partly restored the normal airway responsiveness to OA in DM rats. The release of histamine was remarkably suppressed in DM+INDUCTION rats but was recovered by the insulin treatment. Also, OA significantly increased the levels of IL-1b, TNF-a, CINC-1 and P-selectin in non-DM rats, whereas insulin treatment in DM+INDUCTION rats partly restored the normal levels of IL-1b, TNF-a, CINC-1 and P-selectin in DM rats. Moreover, the expression of IR and IGF1R was evidently suppressed in DM rats, with the methylation of both IR and IGF1R promoters was aggravated in DM rats. Therefore, we demonstrated that DM-induced hypermethylation inhibited mast cell activation and airway responsiveness, which could be reversed by insulin treatment.

Attenuation of relaxing response induced by pituitary adenylate cyclase-activating polypeptide in bronchial smooth muscle of experimental asthma.

Bronchomotor tone is regulated by contraction and relaxation of airway smooth muscle (ASM). A weakened ASM relaxation might be a cause of airway hyperresponsiveness (AHR), a characteristic feature of bronchial asthma. Pituitary adenylyl cyclase-activating polypeptide (PACAP) is known as a mediator that causes ASM relaxation. To date, whether or not the PACAP responsiveness is changed in asthmatic ASM is unknown. The current study examined the hypothesis that relaxation induced by PACAP is reduced in bronchial smooth muscle (BSM) of allergic asthma. The ovalbumin (OA)-sensitized mice were repeatedly challenged with aerosolized OA to induce asthmatic reaction. Twenty-four hours after the last antigen challenge, the main bronchial smooth muscle (BSM) tissues were isolated. Tension study showed a BSM hyperresponsiveness to acetylcholine in the OA-challenged mice. Both quantitative RT-PCR and immunoblot analyses revealed a significant decrease in PAC1 receptor expression in BSMs of the diseased mice. Accordingly, in the antigen-challenged group, the PACAP-induced PAC1 receptor-mediated BSM relaxation was significantly attenuated, whereas the relaxation induced by vasoactive intestinal polypeptide was not changed. These findings suggest that the relaxation induced by PACAP is impaired in BSMs of experimental asthma due to a downregulation of its binding partner PAC1 receptor. Impaired BSM responsiveness to PACAP might contribute to the AHR in asthma.

Leukotriene D4 role in allergic asthma pathogenesis from cellular and therapeutic perspectives.

Asthma is a chronic inflammatory and allergic disease that is mainly characterized by reversible airway obstruction and bronchial hyperresponsiveness. The incidence of asthma is increasing with more than 350 million people worldwide are affected. Up to now, there is no therapeutic option for asthma and most of the prescribed drugs aim to ameliorate the symptoms of the disease especially during the acute exacerbations after trigger exposure. Asthma is a heterogonous disease that involves interactions between inflammatory mediators and cellular components within the disease microenvironment including inflammatory and structural cells. Cysteinyl leukotrienes (cys-LTs) are inflammatory lipid mediators that have potent roles in asthma pathogenesis. CysLTs consisting of LTC4, LTD4, and LTE4 are mainly secreted by leukocytes and act through three main G-protein coupled receptors (CysLT1R, CysLT2R, and CysLT3R). LTD4 is the most potent bronchoconstrictor which gives it the priority to be discussed in detail in this review. LTD4 binds with high affinity to CysLT1R and many studies showed that using CysLT1R antagonists such as montelukast has a beneficial effect for asthmatics especially in corticosteroid refractory cases. Since asthma is a heterogeneous inflammatory disease of many cell types involved in the disease pathogenies and LTD4 has a special role in inflammation and bronchoconstriction, this review highlights the role of LTD4 on each cellular component in asthma and the benefits of using CysLT1R antagonists in ameliorating LTD4-induced effects.

HDAC8 inhibitor attenuates airway responses to antigen stimulus through synchronously suppressing galectin-3 expression and reducing macrophage-2 polarization.

BACKGROUND: This study was to investigate of the mechanism by which histone deacetylase (HDAC) 8 inhibitor ameliorated airway hyperresponsiveness (AHR) and allergic airway inflammation. METHODS: Mice were sensitized and then treated with budesonide (BUD) or PCI-34051 (PCI) prior to exposing to normal saline (NS) or ovalbumin (OVA). The raw264.7 cells were treated with interleukin (IL)-4 and PCI or shRNA alone. Repetitive measurements of enhanced pause (Penh) were executed by increasing concentrations of acetyl-ß-methacholine chloride (0 - 50 mg/ml). Cells in bronchoalveolar lavage fluid (BALF) and pathological changes of lungs were examined, respectively. The expression levels of HDAC8, Galecitn (Gal)-3, CD68, CD86, CD163, Arg1 and NOS2 in lungs were measured. Co-regulation of HDAC8 and Gal-3 proteins was observed by immunofluorescence staining and co-immunoprecipitation assay (Co-IP). RESULTS: Significant increases in Penh and IL-4 level were detected with a large inflammatory infiltrate, comprised predominantly of macrophages and eosinophils, into the BALF in OVA-exposed lungs. HDAC8, Gal-3, CD68, CD86, CD163, Arg1 and NOS2 proteins were over-expressed with the significant changes in the Arg1 and NOS2 mRNA levels in the lungs and the IL-4-treated cells. PCI intervention obviously reduced the counts of CD163+ cells. Furthermore, Gal-3 knockdown suppressed Arg1 expression in the cells. Immunofluorescence staining displayed simultaneous changes in HDAC8 and Gal-3 expression in the investigated samples. Treatment with PCI resulted in synchronous reduction of HDAC8 and Gal-3 expression in the Co-IP complexes. CONCLUSIONS: The HDAC8 inhibitor ameliorates AHR and airway inflammation in animal model of allergic asthma through reducing HDAC8-Gal-3 interaction and M2 macrophage polarization.

Cysteinyl Leukotriene Synthesis via Phospholipase A2 Group IV Mediates Exercise-induced Bronchoconstriction and Airway Remodeling.

It is well known that the prevalence of asthma is higher in athletes, including Olympic athletes, than in the general population. In this study, we analyzed the mechanism of exercise-induced bronchoconstriction by using animal models of athlete asthma. Mice were made to exercise on a treadmill for a total duration of 1 week, 3 weeks, or 5 weeks. We analyzed airway responsiveness, BAL fluid, lung homogenates, and tissue histology for each period. In mice that were treated (i.e., the treatment model), treatments were administered from the fourth to the fifth week. We also collected induced sputum from human athletes with asthma and analyzed the supernatants. Airway responsiveness to methacholine was enhanced with repeated exercise stimulation, although the cell composition in BAL fluid did not change. Exercise induced hypertrophy of airway smooth muscle and subepithelial collagen deposition. Cysteinyl-leukotriene (Cys-LT) levels were significantly increased with exercise duration. Montelukast treatment significantly reduced airway hyperresponsiveness (AHR) and airway remodeling. Expression of PLA2G4 (phospholipase A2 group IV) and leukotriene C4 synthase in the airway epithelium was upregulated in the exercise model, and inhibition of PLA2 ameliorated AHR and airway remodeling, with associated lower levels of Cys-LTs. The levels of Cys-LTs in sputum from athletes did not differ between those with and without sputum eosinophilia. These data suggest that AHR and airway remodeling were caused by repeated and strenuous exercise. Cys-LTs from the airway epithelium, but not inflammatory cells, may play an important role in this mouse model.

Lipoxin A4 reduces house dust mite and TNFα-induced hyperreactivity in the mouse trachea.

Lipoxin A4 (LXA4) is considered a specialised pro-resolving mediator that decreases inflammation: however, pro-inflammatory effects have been described in the airways. Here, we investigated whether LXA4 could influence airway hyperreactivity induced in mouse trachea by house dust mite extract (HDM) or TNFα. Intranasal instillation of HDM caused a serotonin (5-HT) mediated airway hyperreactivity ex vivo (Emax: 78.1 ± 16.2 % versus control 12.8 ± 1.0 %) that was reduced by LXA4 installation one hour prior to HDM (Emax: 49.9 ± 11.4 %). Also, in isolated tracheal segments cultured for four days, HDM induced a hyperreactivity (Emax: 33.2 ± 3.1 % versus control 9.0 ± 0.7 %) that was decreased by LXA4 (Emax: 18.7 ± 1.5 %). One part of the HDM-induced hyperreactivity could be inhibited by the TNFα-inhibitor etanercept. TNFα-induced upregulation of 5-HT responses (Emax: 51.3 ± 1.2 % versus control 13.9 ± 0.5 %) was decreased by 10-1000 nM LXA4. In precontracted tracheal segments, LXA4 had no relaxing effect. Overall, LXA4 was able to decrease airway hyperreactivity induced by both HDM and TNFα, thus having a sub-acute anti-inflammatory effect in airway inflammation.

Myxopyrum serratulum ameliorates airway inflammation in LPS-stimulated RAW 264.7 macrophages and OVA-induced murine model of allergic asthma.

ETHNOPHARMACOLOGICAL RELEVANCE: Myxopyrum serratulum A. W. Hill. (Oleaceae) is a traditionally used Indian medicinal plant for the treatment of cough, asthma and many other inflammatory diseases. AIM OF THE STUDY: In this study, the protective effects of M. serratulum on airway inflammation was investigated in ovalbumin (OVA)-induced murine model of allergic asthma and lipopolysaccharide (LPS)-stimulated inflammation in RAW 264.7 murine macrophages, and the possible mechanisms were elucidated. MATERIALS AND METHODS: The phytochemicals present in the methanolic leaf extract of M. serratulum (MEMS) were identified by reverse phase high performance liquid chromatography (RP-HPLC) analysis. In vitro anti-inflammatory activity of MEMS were evaluated by estimating the levels of nitric oxide (NO), reactive oxygen species (ROS) and cytokines (IL-1α, IL-1ß, IL-2, IL-4, IL-6, IL-10, IL-12, IL-17A, IFN-γ, TNF-α, G-CSF and GM-CSF) in LPS-stimulated RAW 264.7 macrophages. In vivo anti-asthmatic activity of MEMS was studied using OVA-induced murine model. Airway hyperresponsiveness (AHR), was measured; total and differential cell counts, eosinophil peroxidase (EPO), prostaglandin E2 (PGE2), NO, ROS, and cytokines (IL-4, IL-5 and IL-13), were estimated in bronchoalveolar lavage fluid (BALF). Serum total IgE level was measured; and the histopathological changes of lung tissues were observed. The expressions of cyclooxygenase-2 (COX-2) and inducible nitric oxide synthase (iNOS) in lung tissue homogenates were detected by Western blot. RESULTS: The chromatographic analysis of MEMS identified the presence of gallic acid, protocatechuic acid, catechin, ellagic acid, rutin, p-coumaric acid, quercetin, naringenin and apigenin. MEMS (125 and 250 µg/mL) dose-dependently reduced the levels of NO, ROS and pro-inflammatory cytokines in LPS-stimulated RAW 264.7 macrophages. MEMS (200 and 400 mg/kg, p.o.) significantly (p < 0.05) alleviated AHR; number of inflammatory cells, EPO, PGE2, NO, ROS, and cytokines (IL-4, IL-5 and IL-13) in BALF; serum total IgE and the histopathological changes associated with lung inflammation. Western blot studies showed that MEMS substantially suppressed COX-2 and iNOS protein expressions in the lung tissues of OVA-sensitized/challenged mice. CONCLUSIONS: The present study corroborates for the first time the ameliorative effects of MEMS on airway inflammation by reducing the levels of oxidative stress, pro-inflammatory cytokines and inhibiting COX-2, iNOS protein expressions, thereby validating the ethnopharmacological uses of M. serratulum.

Functional contribution of sphingosine-1-phosphate to airway pathology in cigarette smoke-exposed mice.

BACKGROUND AND PURPOSE: A critical role for sphingosine kinase/sphingosine-1-phosphate (S1P) pathway in the control of airway function has been demonstrated in respiratory diseases. Here, we address S1P contribution in a mouse model of mild chronic obstructive pulmonary disease (COPD). EXPERIMENTAL APPROACH: C57BL/6J mice have been exposed to room air or cigarette smoke up to 11 months and killed at different time points. Functional and molecular studies have been performed. KEY RESULTS: Cigarette smoke caused emphysematous changes throughout the lung parenchyma coupled to a progressive collagen deposition in both peribronchiolar and peribronchial areas. The high and low airways showed an increased reactivity to cholinergic stimulation and α-smooth muscle actin overexpression. Similarly, an increase in airway reactivity and lung resistances following S1P challenge occurred in smoking mice. A high expression of S1P, Sph-K2 , and S1P receptors (S1P2 and S1P3 ) has been detected in the lung of smoking mice. Sphingosine kinases inhibition reversed the increased cholinergic response in airways of smoking mice. CONCLUSIONS AND IMPLICATIONS: S1P signalling up-regulation follows the disease progression in smoking mice and is involved in the development of airway hyperresponsiveness. Our study defines a therapeutic potential for S1P inhibitors in management of airways hyperresponsiveness associated to emphysema in smokers with both asthma and COPD.

Anti-inflammatory Property of Galectin-1 in a Murine Model of Allergic Airway Inflammation.

Galectin-1 (Gal-1) has immunomodulatory activities in various allergic inflammatory disorders, but its potential anti-inflammatory properties on allergic airway diseases have not been confirmed. We explored the pharmacological effects of Gal-1 on the progression of allergic airway inflammation and investigated the underlying mechanism. Female C57BL/6 mice were sensitized on day 0 and challenged with ovalbumin (OVA) on days 14-17 to establish an allergic airway inflammation model. In the challenge phase, a subset of mice was treated intraperitoneally with recombinant Gal-1 (rGal-1) or dexamethasone (Dex). We found that rGal-1 inhibited pulmonary inflammatory cell recruitment, mucus secretion, bronchoalveolar lavage fluid (BALF) inflammatory cell infiltration, and cytokine production. The treatment also suppressed the infiltration of eosinophils into the allergic lung as indicated by decreased expression levels of eotaxin and eosinophil peroxidase (EPX). However, only the expression levels of IL-25, neither IL-33 nor TSLP, were significantly decreased in the lung by rGal-1 treatment. These immunomodulatory effects in the allergic lung were correlated with the activation of extracellular signal-regulated kinase (ERK) signaling pathway and downregulation of endogenous Gal-1. In addition, rGal-1 reduced the plasma concentrations of anti-OVA immunoglobulin E (IgE) and IL-17. Our findings suggest that rGal-1 is an effective therapy for allergic airway inflammation in a murine model and may be a potential pharmacological target for allergic airway inflammatory diseases.

Influenza A Virus Infection Induces Hyperresponsiveness in Human Lung Tissue-Resident and Peripheral Blood NK Cells.

NK cells in the human lung respond to influenza A virus- (IAV-) infected target cells. However, the detailed functional capacity of human lung and peripheral blood NK cells remains to be determined in IAV and other respiratory viral infections. Here, we investigated the effects of IAV infection on human lung and peripheral blood NK cells in vitro and ex vivo following clinical infection. IAV infection of lung- and peripheral blood-derived mononuclear cells in vitro induced NK cell hyperresponsiveness to K562 target cells, including increased degranulation and cytokine production particularly in the CD56brightCD16- subset of NK cells. Furthermore, lung CD16- NK cells showed increased IAV-mediated but target cell-independent activation compared to CD16+ lung NK cells or total NK cells in peripheral blood. IAV infection rendered peripheral blood NK cells responsive toward the normally NK cell-resistant lung epithelial cell line A549, indicating that NK cell activation during IAV infection could contribute to killing of surrounding non-infected epithelial cells. In vivo, peripheral blood CD56dimCD16+ and CD56brightCD16- NK cells were primed during acute IAV infection, and a small subset of CD16-CD49a+CXCR3+ NK cells could be identified, with CD49a and CXCR3 potentially promoting homing to and tissue-retention in the lung during acute infection. Together, we show that IAV respiratory viral infections prime otherwise hyporesponsive lung NK cells, indicating that both CD16+ and CD16- NK cells including CD16-CD49a+ tissue-resident NK cells could contribute to host immunity but possibly also tissue damage in clinical IAV infection.

Neutrophilic inflammation in asthma and defective epithelial translational control.

Neutrophilic inflammation in asthma is associated with interleukin (IL)-17A, corticosteroid-insensitivity and bronchodilator-induced forced expiratory volume in 1 s (FEV1) reversibility. IL-17A synergises with tumour necrosis factor (TNF)-α in the production of the neutrophil chemokine CXCL-8 by primary bronchial epithelial cells (PBECs).We hypothesised that local neutrophilic inflammation in asthma correlates with IL-17A and TNF-α-induced CXCL-8 production by PBECs from asthma patients.PBECs from most asthma patients displayed an exaggerated CXCL-8 production in response to TNF-α and IL-17A, but not to TNF-α alone, and which was also insensitive to corticosteroids. This hyperresponsiveness of PBECs strongly correlated with CXCL-8 levels and neutrophil numbers in bronchoalveolar lavage from the corresponding patients, but not with that of eosinophils. In addition, this hyperresponsiveness also correlated with bronchodilator-induced FEV1 % reversibility. At the molecular level, epithelial hyperresponsiveness was associated with failure of the translational repressor T-cell internal antigen-1 related protein (TiAR) to translocate to the cytoplasm to halt CXCL-8 production, as confirmed by TiAR knockdown. This is in line with the finding that hyperresponsive PBECs also produced enhanced levels of other inflammatory mediators.Hyperresponsive PBECs in asthma patients may underlie neutrophilic and corticosteroid-insensitive inflammation and a reduced FEV1, irrespective of eosinophilic inflammation. Normalising cytoplasmic translocation of TiAR is a potential therapeutic target in neutrophilic, corticosteroid-insensitive asthma.

Comparison of multi-wall carbon nanotube and nitrogen-doped multi-wall carbon nanotube effects on lung function and airway reactivity in rats.

Incorporation of multi-wall carbon nanotubes (MWCNT) into materials has raised concerns about their potential hazards to manufacturing workers. In animal models, airway inflammation and lung fibrosis follow aspiration, instillation, and inhalation exposures to MWCNT. However, the effects of MWCNT on pulmonary function, airway reactivity and airway epithelium function following inhalation exposure has not been studied. We investigated whether inhaled MWCNT affects lung resistance (RL) and dynamic compliance (Cdyn), reactivity to inhaled methacholine (MCh), epithelial regulation of airway reactivity to MCh in vitro, and airway epithelial ion transport. Male rats were exposed by whole body inhalation for 6 h to air or aerosolized MWCNT (0.5, 1 or 5 mg/m3) for one or nine days. Eighteen h after 1 d exposure to 5 mg/m3 MWCNT, basal RL was increased and basal Cdyn was decreased; changes did not persist for 7 d. Reactivity to MCh (RL) was increased and Cdyn responses were decreased at 18 h, but not 7 d after exposure to 1 and 5 mg/m3 MWCNT. The effects of i.t.-instilled MWCNT and nitrogen-doped MWCNT (N-MWCNT) on pulmonary function and reactivity to MCh at doses comparable to deposition after inhalation of 5 mg/m3 at 1 d and 0.5, 1, and 5 mg/m3 MWCNT 9 d-exposures were compared. Both nanoparticles increased airway reactivity (RL); N-MWCNT did not affect Cdyn responses. Lung function and airway reactivity are altered following a single MWCNT inhalation and generally subside over time. Given i.t., MWCNT's and N-MWCNT's effects were comparable, but N-MWCNT evoke smaller changes in Cdyn responses.

Farnesyltransferase Inhibition Exacerbates Eosinophilic Inflammation and Airway Hyperreactivity in Mice with Experimental Asthma: The Complex Roles of Ras GTPase and Farnesylpyrophosphate in Type 2 Allergic Inflammation.

Ras, a small GTPase protein, is thought to mediate Th2-dependent eosinophilic inflammation in asthma. Ras requires cell membrane association for its biological activity, and this requires the posttranslational modification of Ras with an isoprenyl group by farnesyltransferase (FTase) or geranylgeranyltransferase (GGTase). We hypothesized that inhibition of FTase using FTase inhibitor (FTI)-277 would attenuate allergic asthma by depleting membrane-associated Ras. We used the OVA mouse model of allergic inflammation and human airway epithelial (HBE1) cells to determine the role of FTase in inflammatory cell recruitment. BALB/c mice were first sensitized then exposed to 1% OVA aerosol or filtered air, and half were injected daily with FTI-277 (20 mg/kg per day). Treatment of mice with FTI-277 had no significant effect on lung membrane-anchored Ras, Ras protein levels, or Ras GTPase activity. In OVA-exposed mice, FTI-277 treatment increased eosinophilic inflammation, goblet cell hyperplasia, and airway hyperreactivity. Human bronchial epithelial (HBE1) cells were pretreated with 5, 10, or 20 µM FTI-277 prior to and during 12 h IL-13 (20 ng/ml) stimulation. In HBE1 cells, FTase inhibition with FTI-277 had no significant effect on IL-13-induced STAT6 phosphorylation, eotaxin-3 peptide secretion, or Ras translocation. However, addition of exogenous FPP unexpectedly augmented IL-13-induced STAT6 phosphorylation and eotaxin-3 secretion from HBE1 cells without affecting Ras translocation. Pharmacological inhibition of FTase exacerbates allergic asthma, suggesting a protective role for FTase or possibly Ras farnesylation. FPP synergistically augments epithelial eotaxin-3 secretion, indicating a novel Ras-independent farnesylation mechanism or direct FPP effect that promotes epithelial eotaxin-3 production in allergic asthma.

Intranasal administration of recombinant progranulin inhibits bronchial smooth muscle hyperresponsiveness in mouse allergic asthma.

Progranulin (PGRN) is a growth factor with multiple biological functions and has been suggested as an endogenous inhibitor of Tumor necrosis factor-α (TNF-α)-mediated signaling. TNF-α is believed to be one of the important mediators of the pathogenesis of asthma, including airway hyperresponsiveness (AHR). In the present study, effects of recombinant PGRN on TNF-α-mediated signaling and antigen-induced hypercontractility were examined in bronchial smooth muscles (BSMs) both in vitro and in vivo. Cultured human BSM cells (hBSMCs) and male BALB/c mice were used. The mice were sensitized and repeatedly challenged with ovalbumin antigen. Animals also received intranasal administrations of recombinant PGRN into the airways 1 h before each antigen inhalation. In hBSMCs, PGRN inhibited both the degradation of IκB-α (an index of NF-κB activation) and the upregulation of RhoA (a contractile machinery-associated protein that contributes to the BSM hyperresponsiveness) induced by TNF-α, indicating that PGRN has an ability to inhibit TNF-α-mediated signaling also in the BSM cells. In BSMs of the repeatedly antigen-challenged mice, an augmented contractile responsiveness to acetylcholine with an upregulation of RhoA was observed: both the events were ameliorated by pretreatments with PGRN intranasally. Interestingly, a significant decrease in PGRN expression was found in the airways of the repeatedly antigen-challenged mice rather than those of control animals. In conclusion, exogenously applied PGRN into the airways ameliorated the antigen-induced BSM hyperresponsiveness, probably by blocking TNF-α-mediated response. Increasing PGRN levels might be a promising therapeutic for AHR in allergic asthma.

Role of Galectin-3 in the pathophysiology underlying allergic lung inflammation in a tissue inhibitor of metalloproteinases 1 knockout model of murine asthma.

Asthma is a chronic inflammatory respiratory disease characterized by airway inflammation, airway hyperresponsiveness and reversible airway obstruction. Understanding the mechanisms that underlie the various endotypes of asthma could lead to novel and more personalized therapies for individuals with asthma. Using a tissue inhibitor of metalloproteinases 1 (TIMP-1) knockout murine allergic asthma model, we previously showed that TIMP-1 deficiency results in an asthma phenotype, exhibiting airway hyperreactivity, enhanced eosinophilic inflammation and T helper type 2 cytokine gene and protein expression following sensitization with ovalbumin. In the current study, we compared the expression of Galectins and other key cytokines in a murine allergic asthma model using wild-type and TIMP-1 knockout mice. We also examined the effects of Galectin-3 (Gal-3) inhibition on a non-T helper type 2 cytokine interleukin-17 (IL-17) to evaluate the relationship between Gal-3 and the IL-17 axis in allergic asthma. Our results showed a significant increase in Gal-3, IL-17 and transforming growth factor-ß1 gene expression in lung tissue isolated from an allergic asthma murine model using TIMP-1 knockout. Gal-3 gene and protein expression levels were also significantly higher in lung tissue from an allergic asthma murine model using TIMP-1 knockout. Our data show that Gal-3 may regulate the IL-17 axis and play a pivotal role in the modulation of inflammation during experimental allergic asthma.

Airway remodelling and inflammation in asthma are dependent on the extracellular matrix protein fibulin-1c.

Asthma is a chronic inflammatory disease of the airways. It is characterized by allergic airway inflammation, airway remodelling, and airway hyperresponsiveness (AHR). Asthma patients, in particular those with chronic or severe asthma, have airway remodelling that is associated with the accumulation of extracellular matrix (ECM) proteins, such as collagens. Fibulin-1 (Fbln1) is an important ECM protein that stabilizes collagen and other ECM proteins. The level of Fbln1c, one of the four Fbln1 variants, which predominates in both humans and mice, is increased in the serum and airways fluids in asthma but its function is unclear. We show that the level of Fbln1c was increased in the lungs of mice with house dust mite (HDM)-induced chronic allergic airway disease (AAD). Genetic deletion of Fbln1c and therapeutic inhibition of Fbln1c in mice with chronic AAD reduced airway collagen deposition, and protected against AHR. Fbln1c-deficient (Fbln1c-/- ) mice had reduced mucin (MUC) 5 AC levels, but not MUC5B levels, in the airways as compared with wild-type (WT) mice. Fbln1c interacted with fibronectin and periostin that was linked to collagen deposition around the small airways. Fbln1c-/- mice with AAD also had reduced numbers of α-smooth muscle actin-positive cells around the airways and reduced airway contractility as compared with WT mice. After HDM challenge, these mice also had fewer airway inflammatory cells, reduced interleukin (IL)-5, IL-13, IL-33, tumour necrosis factor (TNF) and CXCL1 levels in the lungs, and reduced IL-5, IL-33 and TNF levels in lung-draining lymph nodes. Therapeutic targeting of Fbln1c reduced the numbers of GATA3-positive Th2 cells in the lymph nodes and lungs after chronic HDM challenge. Treatment also reduced the secretion of IL-5 and IL-13 from co-cultured dendritic cells and T cells restimulated with HDM extract. Human epithelial cells cultured with Fbln1c peptide produced more CXCL1 mRNA than medium-treated controls. Our data show that Fbln1c may be a therapeutic target in chronic asthma. Copyright © 2017 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.