Asthma exacerbations are triggered by rhinovirus infections. We employed a systems biology approach to delineate upper-airway gene network patterns underlying asthma exacerbation phenotypes in children. Cluster analysis unveiled distinct IRF7 hi versus IRF7 lo molecular phenotypes, the former exhibiting robust upregulation of Th1/type I IFN responses and the latter an alternative signature marked by upregulation of cytokine and growth factor signaling and downregulation of IFN-g. The two phenotypes also produced distinct clinical phenotypes. For IRF7 lo children, symptom duration prior to hospital presentation was more than twice as long from initial symptoms (p = 0.011) and nearly three times as long for cough (p < 0.001), the odds ratio of admission to hospital was increased more than 4-fold (p = 0.018), and time to recurrence was shorter (p = 0.015). In summary, our findings demonstrate that asthma exacerbations in children can be divided into IRF7 hi versus IRF7 lo phenotypes with associated differences in clinical phenotypes.

Viral infections are major drivers of exacerbations and clinical burden in patients with asthma and COPD. IFN-β is a key component of the innate immune response to viral infection. To date studies of inhaled IFN-β treatment have not demonstrated a significant effect on asthma exacerbations. OBJECTIVES: The dynamics of exogenous IFN-β activity were investigated to inform on future clinical indications for this potential anti-viral therapy. METHODS: Monocyte-derived macrophages (MDMs), alveolar macrophages (AMs) and primary bronchial epithelial cells (PBECs) were isolated from healthy controls and COPD patients and infected with influenza virus either prior to or after IFN-β stimulation. Infection levels were measured by % nucleoprotein 1 positive (NP1+) cells using flow cytometry. Viral RNA shedding and interferon stimulated gene expression were measured by qPCR. Production of inflammatory cytokines was measured using MSD. MEASUREMENTS AND MAIN RESULTS: Adding IFN-β to MDMs, AMs and PBECs prior to, but not after, infection reduced %NP1 + cells by 85%, 56% and 66%, respectively (p<0.05). Inhibition of infection lasted for 24h following removal of IFN-β and was maintained albeit reduced up to 1 week in MDMs and 72h in PBECs; 2 this was similar between health and COPD. IFN-β did not induce inflammatory cytokine production by MDMs or PBECs but reduced influenza-induced IL-1β production by PBECs. CONCLUSIONS: In vitro modelling of IFN-β dynamics highlights the potential for intermittent prophylactic doses of exogenous IFN-β to modulate viral infection. This provides important insights to aid the future design of clinical trials of IFN-β in asthma and COPD.

A small-molecule natural product, euodenine A (1), was identified as an agonist of the human TLR4 receptor. Euodenine A was isolated from the leaves of Euodia asteridula (Rutaceae) found in Papua New Guinea and has an unusual U-shaped structure. It was synthesized along with a series of analogues that exhibit potent and selective agonism of the TLR4 receptor. SAR development around the cyclobutane ring resulted in a 10-fold increase in potency. The natural product demonstrated an extracellular site of action, which requires the extracellular domain of TLR4 to stimulate a NF-κB reporter response. 1 is a human-selective agonist that is CD14-independent, and it requires both TLR4 and MD-2 for full efficacy. Testing for immunomodulation in PBMC cells shows the induction of the cytokines IL-8, IL-10, TNF-α, and IL-12p40 as well as suppression of IL-5 from activated PBMCs, indicating that compounds like 1 could modulate the Th2 immune response without causing lung damage.

BackgroundCombination therapy with budesonide and formoterol reduces exacerbations of asthma, which are closely associated with human rhinovirus (RV) infections in both children and adults. These data suggest that budesonide and formoterol inhibit virus-induced inflammatory responses of airway epithelial cells.MethodsTo test this hypothesis, bronchial epithelial (BE) cells were obtained from airway brushings of 8 subjects with moderate-to-severe allergic asthma and 9 with neither asthma nor respiratory allergies. Cultured BE cells were incubated for 24 hours with budesonide (1.77 μM), formoterol (0.1 μM), both, or neither, and then inoculated with RV-16 (5×106 plaque forming units [PFU]/mL). After 24 hours, viral replication (RV RNA), cytokine secretion (CXCL8, CXCL10, TNFα, IFN-β, IL-28) and mRNA expression (CXCL8, CXCL10, TNF, IFNB1, IL28A&B) were analyzed.ResultsRV infection induced CXCL10 protein secretion and IFNB1 and IL28 mRNA expression. Drug treatments significantly inhibited secretion of CXCL10 in mock-infected, but not RV-infected, BE cells, and inhibited secretion of TNFα under both conditions. Neither budesonide nor formoterol, alone or in combination, significantly affected viral replication, nor did they inhibit RV-induced upregulation of IFNB1 and IL28 mRNA. Overall, RV replication was positively related to CXCL10 secretion and induction of IFNB1 and IL28 mRNA, but the positive relationship between RV RNA and CXCL10 secretion was stronger in normal subjects than in subjects with asthma.ConclusionsBudesonide and formoterol can inhibit BE cell inflammatory responses in vitro without interfering with viral replication or production of interferons. These effects could potentially contribute to beneficial effects of budesonide/formoterol combination therapy in preventing RV-induced asthma exacerbations.

The respiratory tract is normally kept essentially free of bacteria by cilia-mediated mucus transport, but in chronic obstructive pulmonary disease (COPD) and cystic fibrosis (CF), bacteria and mucus accumulates instead. To address the mechanisms behind the mucus accumulation, the proteome of bronchoalveolar lavages from COPD patients and mucus collected in an elastase-induced mouse model of COPD was analyzed, revealing similarities with each other and with the protein content in colonic mucus. Moreover, stratified laminated sheets of mucus were observed in airways from patients with CF and COPD and in elastase-exposed mice. On the other hand, the mucus accumulation in the elastase model was reduced in Muc5b-KO mice. While mucus plugs were removed from airways by washing with hypertonic saline in the elastase model, mucus remained adherent to epithelial cells. Bacteria were trapped on this mucus, whereas, in non-elastase-treated mice, bacteria were found on the epithelial cells. We propose that the adherence of mucus to epithelial cells observed in CF, COPD, and the elastase-induced mouse model of COPD separates bacteria from the surface cells and, thus, protects the respiratory epithelium.

RationaleCommon colds are associated with acute respiratory symptom exacerbations in COPD patients.ObjectiveTo determine exacerbation risk and severity in COPD patients with/without coincident self-reported colds.MethodsGlobal initiative for chronic Obstructive Lung Disease stage I–IV COPD patients electronically transmitted respiratory symptom diaries to research staff daily between December 2006 and April 2009. Respiratory symptom worsening prompted contact by a study nurse and patient assessment to determine if a cold was present or an exacerbation underway. A composite daily symptom score was derived for each subject from diarized symptom data. The exacerbation/cold/virus relation was examined using a Poisson regression model, the relation of colds to respiratory symptom severity using generalized estimating equation models.ResultsDaily diary transmission compliance of >97% enabled detection of all possible exacerbations. Among 262 exacerbations meeting Anthonisen criteria, 218 (83%) had cold-like symptoms present at their inception, but respiratory viruses were detected in only 106 (40%). Within-subject exacerbation risk was 30 times (95% confidence interval [CI]: 20, 47; P<0.001) greater with colds present. Compared to cold- and virus-negative exacerbations (n=57), the mean increase in composite symptom score in those cold and virus positive (n=79) was 0.93 (95% CI: 0.61, 1.25; P<0.001), cold-positive and virus-negative exacerbations (n=100) 0.51 (95% CI: 0.21, 0.81; P<0.001), cold-negative and virus-positive exacerbations (n=26) 0.58 (95% CI: 0.23, 0.94; P<0.001).ConclusionThis study emphasizes the importance of colds in COPD exacerbation risk and severity, even in the absence of virus detection. COPD patients should act promptly when cold symptoms appear to facilitate early intervention for exacerbation prevention or management.

Human rhinovirus (RV) infections are a significant risk factor for exacerbations of asthma and chronic obstructive pulmonary disease. Thus, approaches to prevent RV infection in such patients would give significant benefit. Through RNA interference library screening, we identified lanosterol synthase (LSS), a component of the cholesterol biosynthetic pathway, as a novel regulator of RV replication in primary normal human bronchial epithelial cells. Selective knock down of LSS mRNA with short interfering RNA inhibited RV2 replication in normal human bronchial epithelial cells. Small molecule inhibitors of LSS mimicked the effect of LSS mRNA knockdown in a concentration-dependent manner. We further demonstrated that the antiviral effect is not dependent on a reduction in total cellular cholesterol but requires a 24-hour preincubation with the LSS inhibitor. The rank order of antiviral potency of the LSS inhibitors used was consistent with LSS inhibition potency; however, all compounds showed remarkably higher potency against RV compared with the LSS enzyme potency. We showed that LSS inhibition led to an induction of 24(S),25 epoxycholesterol, an important regulator of the sterol pathway. We also demonstrated that LSS inhibition led to a profound increase in expression of the innate antiviral defense protein, IFN-β. We found LSS to be a novel regulator of RV replication and innate antiviral immunity and identified a potential molecular mechanism for this effect, via induction of 24(S),25 epoxycholesterol. Inhibition of LSS could therefore be a novel therapeutic target for prevention of RV-induced exacerbations.

There is a striking seasonal variation in asthma exacerbations in the United States. Substantial differences between regions of the United States in asthma exacerbation rates cannot readily be explained and invite further investigation.