Impaired Antiviral Stress Granule and IFN-ß Enhanceosome Formation Enhances Susceptibility to Influenza Infection in Chronic Obstructive Pulmonary Disease Epithelium.

Chronic obstructive pulmonary disease (COPD) is a serious lung disease that progressively worsens lung function. Those affected are highly susceptible to influenza virus infections that result in exacerbations with exaggerated symptoms with increased mortality. The mechanisms underpinning this increased susceptibility to infection in COPD are unclear. In this study, we show that primary bronchial epithelial cells (pBECs) from subjects with COPD have impaired induction of type I IFN (IFN-ß) and lead to heightened viral replication after influenza viral infection. COPD pBECs have reduced protein levels of protein kinase (PK) R and decreased formation of PKR-mediated antiviral stress granules, which are critical in initiating type I IFN inductions. In addition, reduced protein expression of p300 resulted in decreased activation of IFN regulatory factor 3 and subsequent formation of IFN-ß enhanceosome in COPD pBECs. The decreased p300 induction was the result of enhanced levels of microRNA (miR)-132. Ectopic expression of PKR or miR-132 antagomiR alone failed to restore IFN-ß induction, whereas cotreatment increased antiviral stress granule formation, induction of p300, and IFN-ß in COPD pBECs. This study reveals that decreased induction of both PKR and p300 proteins contribute to impaired induction of IFN-ß in COPD pBECs upon influenza infection.

Airway ß-Defensin-1 Protein Is Elevated in COPD and Severe Asthma.

BACKGROUND: Innate immune antimicrobial peptides, including ß-defensin-1, promote the chemotaxis and activation of several immune cells. The role of ß-defensin-1 in asthma and chronic obstructive pulmonary disease (COPD) remains unclear. METHODS: Induced sputum was collected from healthy controls and individuals with asthma or COPD. ß-defensin-1 protein in sputum supernatant was quantified by ELISA. Biomarker potential was examined using receiver operating characteristic curves. ß-defensin-1 release from primary bronchial epithelial cells (pBECs) was investigated in culture with and without cigarette smoke extract (CSE). RESULTS: Airway ß-defensin-1 protein was elevated in COPD participants compared to asthma participants and healthy controls. Inflammatory phenotype had no effect on ß-defensin-1 levels in asthma or COPD. ß-defensin-1 protein was significantly higher in severe asthma compared to controlled and uncontrolled asthma. ß-defensin-1 protein could predict the presence of COPD from both healthy controls and asthma patients. Exposure of pBECs to CSE decreased ß-defensin-1 production in healthy controls; however in pBECs from COPD participants the level of ß-defensin-1 remanied unchanged. CONCLUSIONS: Elevated ß-defensin-1 protein is a feature of COPD and severe asthma regardless of inflammatory phenotype. ß-defensin-1 production is dysregulated in the epithelium of patients with COPD and may be an effective biomarker and potential therapeutic target.

Toll-like receptor 7 governs interferon and inflammatory responses to rhinovirus and is suppressed by IL-5-induced lung eosinophilia.

BACKGROUND: Asthma exacerbations represent a significant disease burden and are commonly caused by rhinovirus (RV), which is sensed by Toll-like receptors (TLR) such as TLR7. Some asthmatics have impaired interferon (IFN) responses to RV, but the underlying mechanisms of this clinically relevant observation are poorly understood. OBJECTIVES: To investigate the importance of intact TLR7 signalling in vivo during RV exacerbation using mouse models of house dust mite (HDM)-induced allergic airways disease exacerbated by a superimposed RV infection. METHODS: Wild-type and TLR7-deficient (Tlr7(-/-)) BALB/c mice were intranasally sensitised and challenged with HDM prior to infection with RV1B. In some experiments, mice were administered recombinant IFN or adoptively transferred with plasmacytoid dendritic cells (pDC). RESULTS: Allergic Tlr7(-/-) mice displayed impaired IFN release upon RV1B infection, increased virus replication and exaggerated eosinophilic inflammation and airways hyper reactivity. Treatment with exogenous IFN or adoptive transfer of TLR7-competent pDCs blocked these exaggerated inflammatory responses and boosted IFNγ release in the absence of host TLR7 signalling. TLR7 expression in the lungs was suppressed by allergic inflammation and by interleukin (IL)-5-induced eosinophilia in the absence of allergy. Subjects with moderate-to-severe asthma and eosinophilic but not neutrophilic airways inflammation, despite inhaled steroids, showed reduced TLR7 and IFNλ2/3 expression in endobronchial biopsies. Furthermore, TLR7 expression inversely correlated with percentage of sputum eosinophils. CONCLUSIONS: This implicates IL-5-induced airways eosinophilia as a negative regulator of TLR7 expression and antiviral responses, which provides a molecular mechanism underpinning the effect of eosinophil-targeting treatments for the prevention of asthma exacerbations.

The placental protein syncytin-1 impairs antiviral responses and exaggerates inflammatory responses to influenza.

BACKGROUND: Pregnancy increases susceptibility to influenza. The placenta releases an immunosuppressive endogenous retroviral protein syncytin-1. We hypothesised that exposure of peripheral monocytes (PBMCs) to syncytin-1 would impair responses to H1N1pdm09 influenza. METHODS AND FINDINGS: Recombinant syncytin-1 was produced. PBMCs from non-pregnant women (n=10) were exposed to H1N1pdm09 in the presence and absence of syncytin-1 and compared to responses of PBMCs from pregnant women (n=12). PBMCs were characterised using flow cytometry, release of interferon (IFN)-α, IFN-λ, IFN-γ, IL-10, IL-2, IL-6 and IL-1ß were measured by cytometric bead array or ELISA. Exposure of PBMCs to H1N1pdm09 resulted in the release of IFN-α, (14,787 pg/mL, 95% CI 7311-22,264 pg/mL) IFN-λ (1486 pg/mL, 95% CI 756-2216 pg/mL) and IFN-γ (852 pg/mL, 95% CI 193-1511 pg/mL) after 48 hours. This was significantly impaired in pregnant women (IFN-α; p<0.0001 and IFN-λ; p<0.001). Furthermore, in the presence of syncytin-1, PBMCs demonstrated marked reductions in IFN-α and IFN-λ, while enhanced release of IL-10 as well as IL-6 and IL-1ß. CONCLUSIONS: Our data indicates that a placental derived protein, syncytin-1 may be responsible for the heightened vulnerability of pregnant women to influenza.

Targeting PI3K-p110α Suppresses Influenza Virus Infection in Chronic Obstructive Pulmonary Disease.

RATIONALE: Chronic obstructive pulmonary disease (COPD) and influenza virus infections are major global health issues. Patients with COPD are more susceptible to infection, which exacerbates their condition and increases morbidity and mortality. The mechanisms of increased susceptibility remain poorly understood, and current preventions and treatments have substantial limitations. OBJECTIVES: To characterize the mechanisms of increased susceptibility to influenza virus infection in COPD and the potential for therapeutic targeting. METHODS: We used a combination of primary bronchial epithelial cells (pBECs) from COPD and healthy control subjects, a mouse model of cigarette smoke-induced experimental COPD, and influenza infection. The role of the phosphoinositide-3-kinase (PI3K) pathway was characterized using molecular methods, and its potential for targeting assessed using inhibitors. MEASUREMENTS AND MAIN RESULTS: COPD pBECs were susceptible to increased viral entry and replication. Infected mice with experimental COPD also had more severe infection (increased viral titer and pulmonary inflammation, and compromised lung function). These processes were associated with impaired antiviral immunity, reduced retinoic acid-inducible gene-I, and IFN/cytokine and chemokine responses. Increased PI3K-p110α levels and activity in COPD pBECs and/or mice were responsible for increased infection and reduced antiviral responses. Global PI3K, specific therapeutic p110α inhibitors, or exogenous IFN-ß restored protective antiviral responses, suppressed infection, and improved lung function. CONCLUSIONS: The increased susceptibility of individuals with COPD to influenza likely results from impaired antiviral responses, which are mediated by increased PI3K-p110α activity. This pathway may be targeted therapeutically in COPD, or in healthy individuals, during seasonal or pandemic outbreaks to prevent and/or treat influenza.

Differential injurious effects of ambient and traffic-derived particulate matter on airway epithelial cells.

BACKGROUND AND OBJECTIVE: Exposure to airborne particulate matter (PM) may promote development of childhood asthma and trigger acute exacerbations of existing asthma via injury to airway epithelial cells (AEC). METHODS: We compared the response of AEC to ambient particulates with median aerodynamic diameters of <10 µm or <2.5 µm from the Sydney metropolitan region (Sydney PM10 or PM2.5), to traffic-derived particulates from the exhaust stack of a motorway tunnel or to inert carbon black as a control. RESULTS: Sydney PM10 strongly stimulated messenger RNA expression and secretion of the pro-inflammatory cytokines interleukin 6 (IL-6) and chemokine (C-X-C motif) ligand 1 (CXCL1) by mouse tracheal AEC. In contrast, traffic-derived particulates did not. Similarly, PM10 stimulated expression of IL6, IL8 and IL1B by human AEC. Mass spectrometric analysis showed that PM10 contained much higher levels of elements associated with dusts of geological origin. In contrast, tunnel soot contained much higher levels of various organic compounds, notably including long straight-chain alkanes and diesel-derived polycyclic aromatic hydrocarbons. Sydney PM2.5, as well as PM10 collected during a period including a major dust storm, both of which contained relatively lower levels of iron but similar levels of other crustal elements, did not stimulate expression or secretion of CXCL1 by mouse AEC. CONCLUSIONS: Ambient PM10 is likely to be more important than traffic-derived PM in causing injury to AEC leading to production of pro-inflammatory cytokines. The injurious effects may be related to the presence of iron in the coarse fraction of airborne PM. These findings are likely to be relevant to the pathogenesis of asthma.

Viral and bacterial infection in acute asthma and chronic obstructive pulmonary disease increases the risk of readmission.

BACKGROUND AND OBJECTIVE: Infection is as an important trigger for acute asthma and chronic obstructive pulmonary disease (COPD). The aim of this article was to determine the prevalence and impact of virus and bacterial infections in acute asthma and COPD. METHODS: Subjects were recruited, within 24 h of hospital admission for acute exacerbations of asthma and COPD. Nose/throat swabs and sputum samples were collected and examined by multiplex polymerase chain reaction for respiratory viruses and cultured for bacteria. The primary outcomes were length of stay (LOS) and readmission to hospital within 60 days. RESULTS: A total of 199 subjects were recruited (96 had asthma and 103 COPD) for 235 events (36 re-presented). A virus was detected in 79 subjects (40%), bacteria in 41 (21%), and of these, 18 had both. Rhinovirus A was the most frequently isolated virus. A multivariate analysis was performed to control for confounders. It found that detection of a virus, a virus and bacteria, forced expiratory volume in 1 s (FEV(1)) and a diagnosis of COPD were all independent predictors of prolonged LOS, while risk of readmission within 60 days was increased with virus infection alone, virus and bacterial infection, lower FEV(1) and current smoking. CONCLUSIONS: Virus infection, especially in the presence of chronic bacterial infection, is an important determinant of more severe acute exacerbations in both asthma and COPD, and patients with co-infections are more likely to be readmitted to hospital following their exacerbation.

Critical role of constitutive type I interferon response in bronchial epithelial cell to influenza infection.

Innate antiviral responses in bronchial epithelial cells (BECs) provide the first line of defense against respiratory viral infection and the effectiveness of this response is critically dependent on the type I interferons (IFNs). However the importance of the antiviral responses in BECs during influenza infection is not well understood. We profiled the innate immune response to infection with H3N2 and H5N1 virus using Calu-3 cells and primary BECs to model proximal airway cells. The susceptibility of BECs to influenza infection was not solely dependent on the sialic acid-bearing glycoprotein, and antiviral responses that occurred after viral endocytosis was more important in limiting viral replication. The early antiviral response and apoptosis correlated with the ability to limit viral replication. Both viruses reduced RIG-I associated antiviral responses and subsequent induction of IFN-ß. However it was found that there was constitutive release of IFN-ß by BECs and this was critical in inducing late antiviral signaling via type I IFN receptors, and was crucial in limiting viral infection. This study characterizes anti-influenza virus responses in airway epithelial cells and shows that constitutive IFN-ß release plays a more important role in initiating protective late IFN-stimulated responses during human influenza infection in bronchial epithelial cells.