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Long-term tobacco smokers with chronic obstructive pulmonary disease (COPD) or chronic bronchitis display an excessive accumulation of neutrophils in the airways; an inflammation that responds poorly to established therapy. Thus, there is a need to identify new molecular targets for the development of effective therapy. Here, we hypothesized that the neutrophil-mobilizing cytokine interleukin (IL)-26 (IL-26) is involved in airway inflammation amongst long-term tobacco smokers with or without COPD, chronic bronchitis or colonization by pathogenic bacteria. By analyzing bronchoalveolar lavage (BAL), bronchail wash (BW) and induced sputum (IS) samples, we found increased extracellular IL-26 protein in the airways of long-term smokers in vivo without further increase amongst those with clinically stable COPD. In human alveolar macrophages (AM) in vitro, the exposure to water-soluble tobacco smoke components (WTC) enhanced IL-26 gene and protein. In this cell model, the same exposure increased gene expression of the IL-26 receptor complex (IL10R2 and IL20R1) and nuclear factor κ B (NF-κB); a proven regulator of IL-26 production. In the same cell model, recombinant human IL-26 in vitro caused a concentration-dependent increase in the gene expression of NF-κB and several pro-inflammatory cytokines. In the long-term smokers, we also observed that extracellular IL-26 protein in BAL samples correlates with measures of lung function, tobacco load, and several markers of neutrophil accumulation. Extracellular IL-26 was further increased in long-term smokers with exacerbations of COPD (IS samples), with chronic bronchitis (BAL samples ) or with colonization by pathogenic bacteria (IS and BW samples). Thus, IL-26 in the airways emerges as a promising target for improving the understanding of the pathogenic mechanisms behind several pulmonary morbidities in long-term tobacco smokers.

Interleukin (IL)-26 is abundant in human airways and this cytokine is involved in the local immune response to a bacterial stimulus in vivo. Specifically, local exposure to the toll-like receptor (TLR) 4 agonist endotoxin does increase IL-26 in human airways and this cytokine potentiates chemotactic responses in human neutrophils. In addition to T-helper (Th) 17 cells, alveolar macrophages can produce IL-26, but it remains unknown whether this cytokine can also be produced in the airway mucosa per se in response to a viral stimulus. Here, we evaluated whether this is the case using primary bronchial epithelial cells from the airway epithelium in vitro and explored the signaling mechanisms involved, including the modulatory effects of additional Th17 cytokines. Finally, we assessed IL-26 and its archetype signaling responses in healthy human airways in vivo. We found increased transcription and release of IL-26 protein after stimulation with the viral-related double stranded (ds) RNA polyinosinic-polycytidylic acid (poly-IC) and showed that this IL-26 release involved mitogen-activated protein (MAP) kinases and nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB). The release of IL-26 in response to a viral stimulus was modulated by additional Th17 cytokines. Moreover, there was transcription of IL26 mRNA and expression of the protein in epithelial cells of bronchial brush and tissue biopsies respectively after harvest in vivo. In addition, the extracellular IL-26 protein concentrations in bronchoalveolar lavage (BAL) samples did correlate with increased epithelial cell transcription of an archetype intracellular signaling molecule downstream of the IL-26-receptor complex, STAT1, in the bronchial brush biopsies. Thus, our study suggests that viral stimulation causes the production of IL-26 in lining epithelial cells of human airways, structural cells that constitute a critical immune barrier and that this production is modulated by Th17 cytokines.

There is little information on mucins versus potential regulatory factors in the peripheral airway lumen of long-term smokers with (LTS+) and without (LTS−) chronic obstructive pulmonary disease (COPD). We explored these matters in bronchoalveolar lavage (BAL) samples from two study materials, both including LTS+ and LTS− with a very similar historic exposure to tobacco smoke, and healthy non-smokers (HNSs; n=4–20/group). Utilizing slot blot and immunodetection of processed (filtered and centrifuged), as well as unprocessed BAL samples from one of the materials, we compared the quantity and fraction of large complexes of mucins. All LTS displayed an enhanced (median) level of MUC5AC compared with HNS. LTS− displayed a higher level of large MUC5AC complexes than HNS while LTS+ displayed a similar trend. In all LTS, total MUC5AC correlated with blood leukocytes, BAL neutrophil elastase and net gelatinase activity. Large mucin complexes accounted for most MUC5B, without clear group differences. In all LTS, total MUC5B correlated with total MUC5AC and local bacteria. In the same groups, large MUC5B complexes correlated with serum cotinine. MUC1 was increased and correlated with BAL leukocytes in all LTS whereas MUC2 was very low and without clear group differences. Thus, the main part of MUC5AC and MUC5B is present as large complexes in the peripheral airway lumen and historic as well as current exposure to tobacco smoke emerge as potential regulatory factors, regardless of COPD per se. Bacteria, leukocytes and proteinases also constitute potential regulatory factors, of interest for future therapeutic strategies.

Background: Patients with COPD have increased risk for metabolic syndrome (MetS) and type 2 diabetes (T2D). Increased systemic levels of the chemokine IL-8 are found in MetS and T2D, just like in COPD. Aims and Objectives: To characterize glucose tolerance in smokers with and without COPD in relation to IL-8 and neutrophils, locally and systemically. Methods: We utilized a human study material (n= 5-11 in each group) including healthy never-smokers (controls), smokers without (median [range] pack-years: 36.5 [21-51]) and with (pack-years: 37.5 [20-51]) COPD. Spirometry was utilized for the diagnosis of COPD (GOLD2011 stage I-III). The glucose concentration in plasma was determined after fasting and 120 min after oral glucose tolerance test (OGTT, 75 g). The concentration of IL-8-protein was determined (U-Plex assay) in plasma and in BAL-fluid; the latter not for smokers with COPD GOLD stage III. Results: Smokers with COPD had lower fasting-glucose (p=0.01; n=6-8), greater increase of glucose after OGTT (p=0.024; n=6-8) and elevated IL-8 in BAL-fluid (p=0.03, n=5-8) compared to controls. There was a trend towards negative correlation between IL-8 in BAL-fluid (p=0.051; r=-0.51; n=15) and fasting-glucose among all smokers. Plasma IL-8 was higher in smokers without than in those with COPD (p=0.008; n=5-10) and correlated (p=0.023; r=0.58; n=15) with fasting-glucose among all smokers. Blood neutrophils were higher in smokers with COPD than in controls (p=0.04; n=5-8) and correlated (p=0.017; r=0.97, n=5) with fasting-glucose among all smokers. Conclusions: Systemic and local signs of neutrophil mobilization are associated with altered glucose tolerance in smokers with COPD.

Britt-Marie Sundblad,1,* Jie Ji,1,* Bettina Levänen,1 Klara Midander,2 Anneli Julander,2 Kjell Larsson,1 Lena Palmberg,1 Anders Lindén1 1Unit for Lung and Airway Research, 2Unit for Occupational and Environmental Dermatology, Institute of Environmental Medicine, Karolinska Institutet, Stockholm, Sweden *These authors contributed equally tothis work Abstract: Tobacco contains cadmium, and this metal has been attributed a causative role in pulmonary emphysema among smokers, although extracellular cadmium has not to date been quantified in the bronchoalveolar space of tobacco smokers with or without COPD. We determined whether cadmium is enhanced in the bronchoalveolar space of long-term tobacco smokers with or without COPD invivo, its association with inflammation, and its effect on chemokine release in macrophage-like cells in vitro. Bronchoalveolar lavage (BAL), sputum, and blood samples were collected from current, long-term smokers with and without COPD and from healthy nonsmokers. Cadmium concentrations were determined in cell-free BAL fluid using inductively coupled plasma mass spectrometry. Blood monocyte-derived macrophages were exposed to cadmium chloride invitro. Depending upon the type of sample, molecular markers of inflammation were quantified either as protein (enzyme-linked immunosorbent assay) or as mRNA (real-time polymerase chain reaction). Cadmium concentrations were markedly increased in cell-free BAL fluid of smokers compared to that of nonsmokers (n=19–29; P

Chronic obstructive pulmonary disease (COPD) is a leading cause of mortality; however, the role of inflammatory mediators in its pathobiology remains unclear. The aim of this study was to investigate the influence of gender in COPD on lipid mediator levels.Bronchoalveolar lavage fluid (BALF) and serum were obtained from healthy never-smokers, smokers and COPD patients (Global Initiative for Chronic Obstructive Lung Disease stage I–II/A–B) (n=114). 94 lipid mediators derived from the cytochrome-P450, lipoxygenase, and cyclooxygenase pathways were analysed by liquid chromatography-mass spectrometry.Multivariate modelling identified a 9-lipid panel in BALF that classified female smokers with COPD from healthy female smokers (p=6×10−6). No differences were observed for the corresponding male population (p=1.0). These findings were replicated in an independent cohort with 92% accuracy (p=0.005). The strongest drivers were the cytochrome P450-derived epoxide products of linoleic acid (leukotoxins) and their corresponding soluble epoxide hydrolase (sEH)-derived products (leukotoxin-diols). These species correlated with lung function (r=0.87; p=0.0009) and mRNA levels of enzymes putatively involved in their biosynthesis (r=0.96; p=0.003). Leukotoxin levels correlated with goblet cell abundance (r=0.72; p=0.028).These findings suggest a mechanism by which goblet cell-associated cytochrome-P450 and sEH activity produce elevated leukotoxin-diol levels, which play a putative role in the clinical manifestations of COPD in a female-dominated disease sub-phenotype.

Background: IL-26 is an IL-10 family cytokine with pro- and anti-inflammatory effects. In a pilot study, sputum IL-26 concentrations correlated with worsening asthma severity amongst subjects with non-allergic asthma. Objective: To determine whether systemic IL-26 concentrations are increased in children sensitized to dog allergen in vivo and whether it correlates with markers of asthma severity and with the Th17 cytokine, IL-17A. Methods: Serum was obtained from a cohort of dog allergen-sensitized children (n = 60), with non-sensitized children (n = 17) as control subjects. Self-reported clinical history, including medication use and validated symptom-based questionnaire scores, was recorded. IL-26 and IL-17A concentrations were measured by ELISA. Results: Serum IL-26 and IL-17A concentrations were elevated in allergen-sensitized subjects, and IL-26 correlated positively with IL-17A. IL-26 concentrations did not differ significantly between allergen-sensitized subjects with and without asthma or eczema. However, IL-26 correlated positively with increasing Asthma Control Test (ACT) scores and negatively with inhaled corticosteroid dose amongst subjects with asthma. Moreover, subjects with asthma requiring ≥1 course of oral corticosteroids in the preceding 12 months demonstrated a decline in systemic IL-26 concentrations. Conclusion: Systemic IL-26 and IL-17A concentrations are increased in allergen-sensitized individuals compared to those without allergic sensitization, and systemic IL-26 may reflect a non-specific atopic state. Amongst asthmatic children, IL-26 correlates with improved asthma control, suggesting that systemic IL-26 may be associated with a protective effect in asthma.

Cadmium in tobacco smoke may contribute to the development of pulmonary emphysema. However, there is poor understanding of the mechanisms behind the pathogenic role of cadmium in this and other smoking-related lung diseases. The traditional focus on the total body burden of cadmium, estimated through analysis of urine, may not fully reflect the local burden of cadmium, since it is inhaled by smokers. Thus, assessing the local accumulation of cadmium in the lungs appears more relevant, given that there is tissue-specific retention of cadmium.In this review, we outline the principal sources of cadmium exposure and the clinical effects of occupational exposure. In addition, we review evidence on local cadmium and its association with alterations in innate immunity in tobacco smokers. Moreover, we scrutinise the data on cadmium as a cause of lung disease in translational models.We conclude that cadmium may contribute to smoking-related lung diseases, possibly via an altered redox balance and by making macrophages dysfunctional. However, there is a need for new studies on local cadmium levels and their relation to pathology in long-term tobacco smokers, as well as for more in-depth studies on cellular and molecular mechanisms, to elucidate the importance of cadmium in smoking-related lung diseases.

Artificial nanoparticles accumulate a protein corona layer in biological fluids, which significantly influences their bioactivity. As nanosized obligate intracellular parasites, viruses share many biophysical properties with artificial nanoparticles in extracellular environments and here we show that respiratory syncytial virus (RSV) and herpes simplex virus type 1 (HSV-1) accumulate a rich and distinctive protein corona in different biological fluids. Moreover, we show that corona pre-coating differentially affects viral infectivity and immune cell activation. In addition, we demonstrate that viruses bind amyloidogenic peptides in their corona and catalyze amyloid formation via surface-assisted heterogeneous nucleation. Importantly, we show that HSV-1 catalyzes the aggregation of the amyloid β-peptide (Aβ42), a major constituent of amyloid plaques in Alzheimer’s disease, in vitro and in animal models. Our results highlight the viral protein corona as an acquired structural layer that is critical for viral–host interactions and illustrate a mechanistic convergence between viral and amyloid pathologies., The protein corona around artificial nanoparticles is known to influence activity and biological fate, the formation around viruses is less well understood. Here, the authors observe the formation of protein corona on viruses and study the effects this corona has on viral infectivity and on amyloid protein assembly.

Background: The IL-36 cytokine family is a group of pro-inflammatory cytokines produced by structural and myeloid-derived cells in vitro , and in in vivo experimental models, such as psoriasis and pneumonia. Objective: We examined whether IL-36 signaling is enhanced in long-term smokers with and without COPD and whether IL-36 receptor agonists contribute to a pro-inflammatory milieu in the lung. Methods: Bronchoalveolar lavage (BAL) fluid and plasma from a cohort of nonsmokers (n = 8), healthy smokers (n = 10), and smokers with COPD/CB (n = 5) were sonicated. Primary mouse lung macrophages, fibroblasts, and alveolar epithelial cells (AEC) were stimulated with either heat-killed K. pneumoniae (Kp) or recombinant IL-36α or –γ. We quantified mRNA (qRT-PCR) from murine primary cells, as well as proteins (ELISA) in BAL fluid and plasma. Results: Lung macrophages, fibroblasts, and AEC express IL-36α and –γ in response to Kp. Treatment of macrophages with IL-36α or –γ triggered induction of CXC and CC chemokines, and TNFα in a dose-dependent fashion. IL-36α and -γ protein levels were enhanced in BAL fluid from smokers with and without COPD, whereas IL-36β, IL-36RA, and IL-38 were undetectable. Plasma levels of IL-36α but not –γ protein were elevated in smokers with and without COPD. The BAL levels of IL-36α protein correlate negatively with FEV1 and FEV1/FVC. Conclusion: IL-36α and -γ are produced by lung cells in vitro and are increased in smokers with and without COPD in vivo, correlating with a decline in lung function and induction of pro-inflammatory chemokines. This suggests that IL-36 agonists may play a pro-inflammatory role in the lungs of long-term smokers.

Chronic obstructive pulmonary disease (COPD) is a heterogeneous disease and a leading cause of mortality and morbidity worldwide. The aim of this study was to investigate the sex dependency of circulating metabolic profiles in COPD. Serum from healthy never-smokers (healthy), smokers with normal lung function (smokers), and smokers with COPD (COPD; Global Initiative for Chronic Obstructive Lung Disease stages I–II/A–B) from the Karolinska COSMIC cohort (n=116) was analysed using our nontargeted liquid chromatography–high resolution mass spectrometry metabolomics platform. Pathway analyses revealed that several altered metabolites are involved in oxidative stress. Supervised multivariate modelling showed significant classification of smokers from COPD (p=2.8×10−7). Sex stratification indicated that the separation was driven by females (p=2.4×10−7) relative to males (p=4.0×10−4). Significantly altered metabolites were confirmed quantitatively using targeted metabolomics. Multivariate modelling of targeted metabolomics data confirmed enhanced metabolic dysregulation in females with COPD (p=3.0×10−3) relative to males (p=0.10). The autotaxin products lysoPA (16:0) and lysoPA (18:2) correlated with lung function (forced expiratory volume in 1 s) in males with COPD (r=0.86; p, Oxidative stress and the autotaxin–lysoPA axis evidence sex-associated metabotypes in the serum of COPD patients http://ow.ly/kAeE309MpdI

Bettina Levänen,1 Pernilla Glader,2 Barbro Dahlén,3,4 Bo Billing,4 Ingemar Qvarfordt,2 Lena Palmberg,1 Kjell Larsson,1 Anders Lindén1,2,4 1Unit for Lung and Airway Research, Institute of Environmental Medicine, Karolinska Institutet, Stockholm, 2Department of Internal Medicine & Clinical Nutrition, Institute of Medicine, Sahlgrenska Academy at the University of Gothenburg, Gothenburg, 3Centre for Allergy Research, Institute of Environmental Medicine, Karolinska Institutet, Stockholm, 4Lung Allergy Clinic, Karolinska University Hospital, Stockholm, Sweden Abstract: There is excessive accumulation of neutrophils in the airways in chronic obstructive pulmonary disease (COPD) but the underlying mechanisms remain poorly understood. It is known that extracellular cytokine signaling via interleukin (IL)-17A contributes to neutrophil accumulation in the airways but nothing is known about the impact of tobacco smoking on extracellular signaling via IL-17A. Here, we characterized the impact of tobacco smoking on extracellular cytokine signaling via IL-17A in the peripheral airways in long-term smokers with and without COPD and in occasional smokers before and after short-term exposure to tobacco smoke. We quantified concentrations of IL-17A protein in cell-free bronchoalveolar lavage (BAL) fluid samples (Immuno-quantitative PCR) and cytotoxic T-cells (immunoreactivity for CD8+ and CD3+) in bronchial biopsies. Matrix metalloproteinase-8 and human beta defensin 2 proteins were also quantified (enzyme-linked immunosorbent assay) in the BAL samples. The concentrations of IL-17A in BAL fluid were higher in long-term smokers without COPD compared with nonsmoking healthy controls, whereas those with COPD did not differ significantly from either of the other groups. Short-term exposure to tobacco smoke did not induce sustained alterations in these concentrations in occasional smokers. Long-term smokers displayed higher concentrations of IL-17A than did occasional smokers. Moreover, these concentrations correlated with CD8+ and CD3+ cells in biopsies among long-term smokers with COPD. In healthy nonsmokers, BAL concentrations of matrix metalloproteinase-8 and IL-17A correlated, whereas this was not the case in the pooled group of long-term smokers with and without COPD. In contrast, BAL concentrations of human beta defensin 2 and IL-17A correlated in all study groups. This study implies that long-term but not short-term exposure to tobacco smoke increases extracellular cytokine signaling via IL-17A in the peripheral airways. In the smokers with COPD, this signaling may involve cytotoxic T-cells. Long-term exposure to tobacco smoke leads to a disturbed association of extracellular IL-17A signaling and matrix metalloproteinase-8, of potential importance for the coordination of antibacterial activity. Keywords: BAL, COPD, IL-17, tobacco, long-term smoking, occasional smoking

Our objective is to classify smoking- and COPD disease status by network-based multi-9omics data fusion analysis. We hypothesize that bridging and integration of multi-molecular level data will provide improved power for classification of smoking and COPD diagnosis. 7 9Omics data sets from the Karolinska COSMIC study were. Feature selections and a network-based multi-9omics data fusion and clustering of subjects analysis was performed. We found that the prediction power (Normalized Mutual Information, NMI between predicted clusters and known groups) was increased with the number of (complementary) 9omics data blocks used. The best predictor for all four groups (NMI=0.85) was achieved by integration of proteome, mRNA transcriptome data from BAL cells, combined with metabolomics and exosomal miRNA profiling from BAL fluid, including 276 significant COPD-influenced features (p The results indicate that there is both comprehensive and redundant information within multi-9omics datasets. The predictive power will be highly increased by integration of comprehensive multi-9omics data. Even for dominant factors smoking, the predictive power will be significantly improved by multi-9omics fusion than single 9omics. Further expansion of these methods to allow sub-phenotyping of at-risk smokers and COPD patients may lead to improved techniques for early diagnosis of COPD. The predictive power (NMI) is increased with the number of complementary 9OMICs.

In this pilot study, we examined associations between local interleukin (IL)-26, disease severity and biomarkers of Th2-mediated inflammation in a well-defined cohort of pediatric patients (14 years median age, 41 % females) with controlled (n = 28) or uncontrolled (n = 48) asthma. Sputum IL-26 protein concentrations (ELISA) reflected disease control in patients without local (low exhaled nitric oxide) or systemic (low blood eosinophils) signs of eosinophilic inflammation. Moreover, sputum-IL-26 concentrations correlated with those of blood neutrophils. Our study indicates that IL-26 is a potential biomarker of disease severity in pediatric asthma without signs of Th2-mediated inflammation.

Rational: Improving the understanding of pulmonary host defense against bacteria in humans is vital for the development of effective therapy. Aim: To characterize the involvement of the presumed Th17 cytokine interleukin (IL)-26 in antibacterial host defense of the lungs. Methods: Intra-bronchial exposure of healthy volunteers to the TLR-4 agonist endotoxin and vehicle was performed during bronchoscopy and bronchoalveolar lavage (BAL) samples were harvested. Intracellular IL-26 was detected by immunocytochemistry and -cytofluorescence. This IL-26 was also detected by flow cytometry, as was its receptor complex. Cytokines and phosphorylated STAT1 plus -3 were quantified by ELISA. Gene expression was analyzed by real-time PCR and neutrophil migration was assessed in vitro. Results: Extracellular IL-26 was detected in BAL samples without prior exposure in vivo and was markedly increased after endotoxin exposure. Alveolar macrophages (AM), CD4 and 8 T-cells constituted cellular sources. Recombinant IL-26 potentiated chemotaxis induced by IL-8 and fMLP but decreased chemokinesis in neutrophils. The IL-26 receptor complex was detected in neutrophils and IL-26 decreased phosphorylated STAT3 in these cells. IL-26 increased gene expression of its receptor complex and STAT1 plus -3. Finally, IL-26 increased the release of neutrophil-mobilizing cytokines in BAL cells. Conclusion: Alveolar macrophages,stimulated through TLR-4 produce IL-26 that acts on neutrophil receptors and focuses their mobilization towards bacteria and accumulated immune cells in human lungs. This involvement of IL-26 in host defense in human lungs warrants further evaluation in local infections and inflammatory disorders.

Background: COPD is a leading cause of mortality and morbidity; however, the role of inflammatory mediators in its pathobiology remains unclear. Aim: To investigate the influence of gender in COPD on lipid mediators levels in the bronchoalveolar compartment and serum. Methods: Bronchoalveolar lavage fluid (BALF) and serum were obtained from healthy never-smokers, smokers with normal lung function, and COPD patients (GOLD I-II/A-B) (n=114). Ninety-four lipid mediators derived from the cytochrome P450, lipoxygenase, and cyclooxygenase pathways were analyzed by liquid chromatography-mass spectrometry. Results: Multivariate modeling identified a 9-lipid panel in BALF that classified female smokers with COPD from healthy female smokers (p=6x10-6). No differences were observed for the corresponding male population (p=1.0). Findings were replicated in an independent cohort with 92% accuracy (p=0.005). The strongest drivers were the cytochrome P450-derived epoxide products of linoleic acid (leukotoxins) and their corresponding soluble epoxide hydrolase (sEH)-derived products (leukotoxin-diols). These lipids correlated with lung function in the female COPD group (r=0.87; p=0.0009), as well as mRNA levels of enzymes involved in their biosynthesis (r=0.96; p=0.003). Leukotoxin levels correlated with goblet cell abundance in female COPD patients (r=0.72; p=0.028). Conclusions: Findings suggest a mechanism by which goblet cell-associated cytochrome P4509s in combination with sEH activity produce elevated leukotoxin-diol levels, which play a putative role in the clinical manifestations of COPD in a female-dominated disease sub-phenotype. Data support the inhibition of sEH as a therapeutic target for treatment of COPD.

The role of the presumed Th17 cytokine IL-26 in antibacterial host defense of the lungs is not known.To characterize the role of IL-26 in antibacterial host defense of human lungs.Intrabronchial exposure of healthy volunteers to endotoxin and vehicle was performed during bronchoscopy and bronchoalveolar lavage (BAL) samples were harvested. Intracellular IL-26 was detected using immunocytochemistry and immunocytofluorescence. This IL-26 was also detected using flow cytometry, as was its receptor complex. Cytokines and phosphorylated signal transducer and activator of transcription (STAT) 1 plus STAT3 were quantified using ELISA. Gene expression was analyzed by real-time polymerase chain reaction and neutrophil migration was assessed in vitro.Extracellular IL-26 was detected in BAL samples without prior exposure in vivo and was markedly increased after endotoxin exposure. Alveolar macrophages displayed gene expression for, contained, and released IL-26. Th and cytotoxic T cells also contained IL-26. In the BAL samples, IL-26 concentrations and innate effector cells displayed a correlation. Recombinant IL-26 potentiated neutrophil chemotaxis induced by IL-8 and fMLP but decreased chemokinesis for neutrophils. Myeloperoxidase in conditioned media from neutrophils was decreased. The IL-26 receptor complex was detected in neutrophils and IL-26 decreased phosphorylated STAT3 in these cells. In BAL and bronchial epithelial cells, IL-26 increased gene expression of the IL-26 receptor complex and STAT1 plus STAT3. Finally, IL-26 increased the release of neutrophil-mobilizing cytokines in BAL but not in epithelial cells.This study implies that alveolar macrophages produce IL-26, which stimulates receptors on neutrophils and focuses their mobilization toward bacteria and accumulated immune cells in human lungs.

BackgroundAsthma is characterized by increased airway narrowing in response to nonspecific stimuli. The disorder is influenced by both environmental and genetic factors. Exosomes are nanosized vesicles of endosomal origin released from inflammatory and epithelial cells that have been implicated in asthma. In this study we characterized the microRNA (miRNA) content of exosomes in healthy control subjects and patients with mild intermittent asthma both at unprovoked baseline and in response to environmental challenge.ObjectiveTo investigate alterations in bronchoalveolar lavage fluid (BALF) exosomal miRNA profiles due to asthma, and following subway air exposure.MethodsExosomes were isolated from BALF from healthy control subjects (n=10) and patients with mild intermittent asthma (n=10) after subway and control exposures. Exosomal RNA was analyzed by using microarrays containing probes for 894 human miRNAs, and selected findings were validated with quantitative RT-PCR. Results were analyzed by using multivariate modeling.ResultsThe presence of miRNAs was confirmed in exosomes from BALF of both asthmatic patients and healthy control subjects. Significant differences in BALF exosomal miRNA was detected for 24 miRNAs with a subset of 16 miRNAs, including members of the let-7 and miRNA-200 families, providing robust classification of patients with mild nonsymptomatic asthma from healthy subjects with 72% cross-validated predictive power (Q(2)=0.72). In contrast, subway exposure did not cause any significant alterations in miRNA profiles.ConclusionThese studies demonstrate substantial differences in exosomal miRNA profiles between healthy subjects and patients with unprovoked, mild, stable asthma. These changes might be important in the inflammatory response leading to bronchial hyperresponsiveness and asthma.

Background Asthma is a chronic inflammatory lung disease that causes significant morbidity and mortality worldwide. Air pollutants such as particulate matter (PM) and oxidants are important factors in causing exacerbations in asthmatics, and the source and composition of pollutants greatly affects pathological implications. Objectives This randomized crossover study investigated responses of the respiratory system to Stockholm subway air in asthmatics and healthy individuals. Eicosanoids and other oxylipins were quantified in the distal lung to provide a measure of shifts in lipid mediators in association with exposure to subway air relative to ambient air. Methods Sixty-four oxylipins representing the cyclooxygenase (COX), lipoxygenase (LOX) and cytochrome P450 (CYP) metabolic pathways were screened using liquid chromatography-tandem mass spectrometry (LC-MS/MS) of bronchoalveolar lavage (BAL)-fluid. Validations through immunocytochemistry staining of BAL-cells were performed for 15-LOX-1, COX-1, COX-2 and peroxisome proliferator-activated receptor gamma (PPARγ). Multivariate statistics were employed to interrogate acquired oxylipin and immunocytochemistry data in combination with patient clinical information. Results Asthmatics and healthy individuals exhibited divergent oxylipin profiles following exposure to ambient and subway air. Significant changes were observed in 8 metabolites of linoleic- and α-linolenic acid synthesized via the 15-LOX pathway, and of the COX product prostaglandin E2 (PGE2). Oxylipin levels were increased in healthy individuals following exposure to subway air, whereas asthmatics evidenced decreases or no change. Conclusions Several of the altered oxylipins have known or suspected bronchoprotective or anti-inflammatory effects, suggesting a possible reduced anti-inflammatory response in asthmatics following exposure to subway air. These observations may have ramifications for sensitive subpopulations in urban areas.

Kristina Andelid,1 Sara Tengvall,1 Anders Andersson,1 Bettina Levänen,2 Karin Christenson,3 Pernilla Jirholt,3 Christina Åhrén,4 Ingemar Qvarfordt,1 Ann Ekberg-Jansson,1 Anders Lindén2 1Department of Internal Medicine and Clinical Nutrition, Institute of Medicine, Sahlgrenska Academy at the University of Gothenburg, Gothenburg, Sweden; 2Unit of Lung and Airway Research, Institute of Environmental Medicine, Karolinska Institutet, Stockholm, Sweden; 3Department of Rheumatology and Inflammation Research, Institute of Medicine, 4Department of Infectious Diseases, Infection ControlUnit, Institute of Biomedicine, Sahlgrenska Academy at the University of Gothenburg, Gothenburg, Sweden Abstract: We examined whether systemic cytokine signaling via interleukin (IL)-17 and growth-related oncogene-α (GRO-α) is impaired in smokers with obstructive pulmonary disease including chronic bronchitis (OPD-CB). We also examined how this systemic cytokine signaling relates to bacterial colonization in the airways of the smokers with OPD-CB. Currently smoking OPD-CB patients (n=60, corresponding to Global initiative for chronic Obstructive Lung Disease [GOLD] stage I–IV) underwent recurrent blood and sputum sampling over 60 weeks, during stable conditions and at exacerbations. We characterized cytokine protein concentrations in blood and bacterial growth in sputum. Asymptomatic smokers (n=10) and never-smokers (n=10) were included as control groups. During stable clinical conditions, the protein concentrations of IL-17 and GRO-α were markedly lower among OPD-CB patients compared with never-smoker controls, whereas the asymptomatic smoker controls displayed intermediate concentrations. Notably, among OPD-CB patients, colonization by opportunistic pathogens was associated with markedly lower IL-17 and GRO-α, compared with colonization by common respiratory pathogens or oropharyngeal flora. During exacerbations in the OPD-CB patients, GRO-α and neutrophil concentrations were increased, whereas protein concentrations and messenger RNA for IL-17 were not detectable in a reproducible manner. In smokers with OPD-CB, systemic cytokine signaling via IL-17 and GRO-α is impaired and this alteration may be linked to colonization by opportunistic pathogens in the airways. Given the potential pathogenic and therapeutic implications, these findings deserve to be validated in new and larger patient cohorts. Keywords: COPD, exacerbation, infection, neutrophil, lung, opportunist&nbsp

Summary Background Sarcoidosis is an inflammatory multisystemic granulomatous disease of unknown aetiology commonly affecting the lungs, and pulmonary fibrosis often develops in chronic sarcoidosis. It has been suggested that Wnt (Wingless/integrated)-signaling has a role in inflammatory and fibrotic processes in the lungs, but its role in sarcoidosis has not been investigated. We hypothesised that Wnts secreted from T cells or other inflammatory cells have a role in the pathogenesis of sarcoidosis. Methods Brush biopsies and bronchoalveolar lavage (BAL) were obtained through bronchoscopy from healthy controls ( n = 18) and patients with sarcoidosis ( n = 48). Semi-quantitative RT-PCR, electrophoretic mobility shift assay (EMSA) and immunocytochemistry were performed to analyse Wnt expression and activation of the Wnt-signal transducer β-catenin. Results Altered expression of Wnt5A, Wnt7A and Wnt7B mRNA in BAL cells was observed, as well as an increased activation of β-catenin, measured by EMSA and confirmed with immunocytochemistry, in resident lung cells from patients with sarcoidosis. More pronounced changes in Wnt expression were seen with advancing disease stage. Thus, by three independent methods, we have found evidence of increased pulmonary Wnt-activation in sarcoidosis. Conclusions In the lungs of patients with sarcoidosis there is a previously unappreciated increased Wnt-signal activation that could contribute to the inflammatory processes.