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Respiratory disease, characterized by changes in the cells of the lung, can affect molecular phenotype of cells and the intercellular interactions, resulting in a disbalance in the relative proportions of individual cell types. Understanding these changes is essential to understand the pathophysiology of lung disease. Conventional 'bulk' RNA-sequencing (RNA-seq), analyzing the entire transcriptome of the tissue sample, provides information about average expression levels of each gene in the mixed cell population; whereas it does not consider the cellular heterogeneity in samples composed of more than one cell type 1 . Single-cell RNA-seq (scRNA-seq) assesses the transcriptome of a complex biological sample with single-cell resolution, allowing identification of the relative frequency of discrete cell-types and analysis of their transcriptomes 1 . Nevertheless, analyzing the transcriptomic signature in large numbers of patients by scRNA-Seq is currently limited by its high costs. Mast cells are key regulatory cells driving the inflammatory process in asthma2 . Since they can be quantified by immunohistochemical staining for validation purposes, we used mast cells as an example of a rare cell population to assess the validity of our deconvolution approach. Recently, a number of bulk RNA-seq deconvolution methods have become available 3 , for instance of two deconvolution methods, namely support vector regression (SVR) 4 , the machine-learning method implemented in CYBERSORT, and Non-Negative Least Square (NNLS) 5 , using a matrix of cell-type selective genes identified with AutoGeneSc 6 . Both approaches are designed to estimate relative proportion of the main, common cell types present in the sample. When we used these methods to estimate the number of mast cells, we found a poor correlation with the number of mast cells stained by immunohistochemistry in the biopsies, suggesting the CIBERSORT and NNLS are less reliable in the case of rare cell types. We explored the possibility to use scRNA-Seq data from small numbers of subjects to specifically interrogate the relative cell type frequency of a rare cell population in a bulk RNA-Seq dataset obtained from a large asthma cohort.

A candidate‐based inducible eQTL study was performed in occasional smokers, identifying rs3212855 flanking the FPR as novel cigarette smoke‐induced eQTL. In addition, rs3212855 was associated with lower lung function. Lastly, CRISPR‐Cas9‐downregulated FPR1 alveolar epithelial cells were protected against the harmful effects of cigarette smoke on epithelial repair., Background and objective Cigarette smoking is one of the most prevalent causes of preventable deaths worldwide, leading to chronic diseases, including chronic obstructive pulmonary disease (COPD). Cigarette smoke is known to induce significant transcriptional modifications throughout the respiratory tract. However, it is largely unknown how genetic profiles influence the smoking‐related transcriptional changes and how changes in gene expression translate into altered alveolar epithelial repair responses. Methods We performed a candidate‐based acute cigarette smoke‐induced eQTL study, investigating the association between SNP and differential gene expression of FPR family members in bronchial epithelial cells isolated 24 h after smoking and after 48 h without smoking. The effects FPR1 on lung epithelial integrity and repair upon damage in the presence and absence of cigarette smoke were studied in CRISPR‐Cas9‐generated lung epithelial knockout cells. Results One significant (FDR 2‐fold change in gene expression. The minor allele of rs3212855 was associated with significantly higher gene expression of FPR1, FPR2 and FPR3 upon smoking. Importantly, the minor allele of rs3212855 was also associated with lower lung function. Alveolar epithelial FPR1 knockout cells were protected against CSE‐induced reduction in repair capacity upon wounding. Conclusion We identified a novel smoking‐related inducible eQTL that is associated with a smoke‐induced increase in the expression of FPR1, FPR2 and FPR3, and with lowered lung function. in vitro FPR1 down‐regulation protects against smoke‐induced reduction in lung epithelial repair.

Background and objective Bronchoscopic lung volume reduction using endobronchial valves (EBV) is an effective new treatment option for severe emphysema patients without interlobar collateral ventilation. The objective of this study was to perform an economic evaluation including the costs and cost-effectiveness of EBV treatment compared with standard medical care (SoC) from the hospital perspective in the short term and long term. Methods For the short-term evaluation, incremental cost-effectiveness ratios (ICER) were calculated based on the 6-month end point data from the STELVIO randomized trial. For the long-term evaluation, a Markov simulation model was constructed based on STELVIO and literature. The clinical outcome data were quality-adjusted life-years (QALY) based on the EuroQol5-Dimensions (EQ5D) questionnaire, the 6-min walking distance (6MWD) and the St George's Respiratory Questionnaire (SGRQ). Results The mean difference between the EBV group and controls was €16 721/patient. In the short-term (6 months), costs per additional QALY was €205 129, the ICER for 6MWD was €160 and for SGRQ was €1241. In the long term, the resulting cost-effectiveness ratios indicate additional costs of €39 000 per QALY gained with a 5-year time horizon and €21 500 per QALY gained at 10 years. In comparison, historical costs per additional QALY 1 year after the coil treatment are €738 400, 5 years after lung volume reduction surgery are €48 415 and 15 years after double-lung transplantation are €29 410. Conclusion The positive clinical effects of EBV treatment are associated with increased costs compared with SoC. Our results suggest that the EBV treatment has a favourable cost-effectiveness profile, also when compared with other treatment modalities for this patient group.

© 2018 The Authors. Journal of Cellular and Molecular Medicine published by John Wiley & Sons Ltd and Foundation for Cellular and Molecular Medicine. Neovascularization, increased basal membrane thickness and increased airway smooth muscle (ASM) bulk are hallmarks of airway remodelling in asthma. In this study, we examined connective tissue growth factor (CTGF) dysregulation in human lung tissue and animal models of allergic airway disease. Immunohistochemistry revealed that ASM cells from patients with severe asthma (A) exhibited high expression of CTGF, compared to mild and non-asthmatic (NA) tissues. This finding was replicated in a sheep model of allergic airways disease. In vitro, transforming growth factor (TGF)-β increased CTGF expression both in NA- and A-ASM cells but the expression was higher in A-ASM at both the mRNA and protein level as assessed by PCR and Western blot. Transfection of CTGF promoter-luciferase reporter constructs into NA- and A-ASM cells indicated that no region of the CTGF promoter (−1500 to +200 bp) displayed enhanced activity in the presence of TGF-β. However, in silico analysis of the CTGF promoter suggested that distant transcription factor binding sites may influence CTGF promoter activation by TGF-β in ASM cells. The discord between promoter activity and mRNA expression was also explained, in part, by differential post-transcriptional regulation in A-ASM cells due to enhanced mRNA stability for CTGF. In patients, higher CTGF gene expression in bronchial biopsies was correlated with increased basement membrane thickness indicating that the enhanced CTGF expression in A-ASM may contribute to airway remodelling in asthma.

BACKGROUND AND OBJECTIVE: Target lobar volume reduction (TLVR) is an important efficacy outcome measure for bronchoscopic lung volume reduction (BLVR) treatment using one-way endobronchial valves (EBV) in patients with severe emphysema. The commonly used cut-off value for TLVR that expresses a perceivable clinical benefit is -350 mL. However, a scientifically determined minimal important difference (MID) for TLVR never has been published. The objective of the present study was to determine the MID for TLVR on HRCT in patients who were treated with EBV. METHODS: A total of 318 patients with severe emphysema from two BLVR trials were analysed. Anchor-based methods were used to define the TLVR MID at 6 months follow-up. Forced expiratory volume in 1 s (FEV1 ), residual volume (RV) and St. George's Respiratory Questionnaire (SGRQ) were used as anchors. RESULTS: The calculated TLVR MID with each anchor was: FEV1 -587 mL, RV -534 mL and SGRQ -560 mL. The combined MID (average of the three anchor-based MIDs) was -563 mL. CONCLUSION: Using the anchor-based method, we established a TLVR MID of -563 mL in patients with severe emphysema at 6 months follow-up after EBV treatment. This value can be useful for both interpreting the results from trials and clinical practice, as well as for designing future studies on lung volume reduction.

In this article, we review the dynamic role of state and nonstate actors in governance. We first discuss the main arguments for and against the state being the main actor in governance in recent literature. Then, we review some of the literature about the changing role of state and nonstate actors in response to the 2007–08 global financial crisis from 2011 to 2015. The two themes under examination are, first, more control over financial markets and second, austerity measures. They illustrate different trajectories of governance that go beyond the now well-established New Public Management paradigm of public sector reforms. Our review shows that no single actor provides the best mode of governance for all circumstances. Instead, governance is hybrid and dynamic. The mode of governance is dependent on the circumstances under which an actor is more capable of interacting with other actors to provide public services.

We conducted an international study to evaluate practices in the diagnosis and management of patients with chronic thromboembolic pulmonary hypertension (CTEPH) globally across different regions. Between August and October 2012, CTEPH-treating physicians completed a 15-minute online questionnaire and provided patient record data for their 2–5 most recent patients with CTEPH. Overall, 496 physicians (Europe: 260; United States: 152; Argentina: 52; Japan: 32) completed the questionnaire and provided patient record data for 1,748 patients. The proportion of physicians who described themselves as working in or affiliated with a specialized pulmonary hypertension (PH) center ranged from 38% in France and Italy to 83% in the United States. A large proportion of patients did not undergo ventilation/perfusion scanning (46%–67%) or right heart catheterization (24%–57%) for the diagnosis of CTEPH. Referral rates for pulmonary endarterectomy evaluation ranged from 25% in Japan to 44% in Europe, with higher referral rates in PH centers; the main reasons for lack of referral were that surgery was not considered unless medical treatment was failing and patient refusal. Other variations in management included greater use of phosphodiesterase 5 inhibitors in the United States than in Europe and Japan and greater use of combination treatment in the United States than in Europe. Physicians’ perceptions of their treatment strategy were generally consistent with patient record data. Results from this study, which includes a global aspect of CTEPH care, demonstrate not only regional differences in CTEPH management but, more importantly, considerable nonadherence to the diagnosis and treatment guidelines for CTEPH, even in PH centers.

We conducted an international study to evaluate practices in the diagnosis and management of pulmonary arterial hypertension (PAH) globally across different geographic regions. Between July and October 2012, PAH-treating physicians completed a 15-minute online questionnaire and provided patient record data for their 3 or 5 most recent patients with PAH. Overall, 560 physicians (Europe: 278; United States: 160; Argentina: 53; Japan: 69) completed the questionnaire and provided data for 2,618 patients. The proportion of physicians who described themselves as working in or affiliated with a specialized pulmonary hypertension center ranged from 13% in Argentina to 74% in the United States. At the time of diagnosis, patients’ New York Heart Association functional class differed significantly between regions. At the time of last assessment, functional class had improved overall, and differences between regions had largely disappeared. A large proportion of patients did not undergo right heart catheterization for the diagnosis of PAH (Europe: 7%–21%; United States: 21%; Japan: 19%; Argentina: 51%). Variations in management included greater use of phosphodiesterase 5 inhibitors in the United States than in Europe and Japan and greater use of triple or greater combination therapy in Japan than in other regions. Results from this study, which includes a global aspect of PAH care, demonstrate that there are significant differences in PAH management between regions and low adherence to guidelines recommending right heart catheterization for the diagnosis of PAH.

New Zealand ephemeral wetlands are ecologically important, containing up to 12% of threatened native plant species and frequently exhibiting conspicuous cyanobacterial growth. In such environments, cyanobacteria and associated heterotrophs can influence primary production and nutrient cycling. Wetland communities, including bacteria, can be altered by increased nitrate and phosphate due to agricultural practices. We have characterized cyanobacteria from the Wairepo Kettleholes Conservation Area and their associated bacteria. Use of 16S rRNA amplicon sequencing identified several operational taxonomic units (OTUs) representing filamentous heterocystous and non-heterocystous cyanobacterial taxa. One Nostoc OTU that formed macroscopic colonies dominated the cyanobacterial community. A diverse bacterial community was associated with the Nostoc colonies, including a core microbiome of 39 OTUs. Identity of the core microbiome associated with macroscopic Nostoc colonies was not changed by the addition of nutrients. One OTU was highly represented in all Nostoc colonies (27.6%-42.6% of reads) and phylogenetic analyses identified this OTU as belonging to the genus Sphingomonas. Scanning electron microscopy showed the absence of heterotrophic bacteria within the Nostoc colony but revealed a diverse community associated with the colonies on the external surface.

This study sought to determine the prevalence of coronary artery-pulmonary artery collaterals in patients with chronic thromboembolic pulmonary hypertension (CTEPH) and to correlate their presence with the degree of clot burden. CTEPH is a treatable cause of severe pulmonary hypertension and right heart failure. Bronchopulmonary collateral vessels have been used as a supplementary diagnostic and prognostic tool for this disease. Coronary artery-pulmonary artery collaterals in this population have not been described. The coronary angiograms of 300 consecutive patients with CTEPH evaluated for pulmonary thromboendarterectomy (PTE) between January 1, 2007, and May 1, 2014, were examined. Of these patients, 259 (50% male; mean age, 58.3 ± 10.6 years) had cineangiographic images deemed adequate to definitively assess for the presence of coronary artery-pulmonary artery collaterals and were included in the final analyses. Pulmonary angiogram reports were reviewed for extent of pulmonary artery obstruction. The coronary angiograms of 259 age- and sex-matched control patients were also examined. Among 259 CTEPH patients with definitive imaging, 34 coronary artery-pulmonary artery collaterals were found in 28 patients (10.8%), versus 1 coronary artery-pulmonary artery collateral among control subjects (0.4%; P < 0.001). Compared with CTEPH patients without collaterals, patients with collaterals had a significantly higher prevalence of total occlusion of their right or left main pulmonary artery (P < 0.001) or lobar arteries (P < 0.001). In conclusion, the prevalence of coronary artery-pulmonary artery collaterals in CTEPH patients undergoing coronary angiography for possible PTE is approximately 11%. These vessels are associated with more severe pulmonary artery occlusion.

Currently, to our knowledge, there are no chronic obstructive pulmonary disease (COPD)-specific formulas to predict the maximum workload of an incremental cycle ergometer test. The aim of the study was to investigate different prediction models including COPD-specific variables of maximum workload in 113 mild to very severe COPD patients. This study shows that simple measures like forced expiratory volume in 1 s, chair-stand test and modified Medical Research Council dyspnoea score may improve the accuracy of the predicted maximum workload in COPD patients.

Schistosomiasis, a parasite-borne disease, is highly prevalent in Africa and Asia; it is estimated that close to 20 million people worldwide have a severe form of the disease. The chronic form can affect the gastrointestinal system and lead to hepatosplenic disease, and it may cause cardiopulmonary complications, including pulmonary hypertension. The exact pathogenesis of schistosomiasis-associated pulmonary hypertension (Sch-PH) remains unclear, although several mechanisms, including parasitic arterial embolization, pulmonary arteriopathy, and portopulmonary hypertension–like pathophysiology, have been suggested. The immunopathology of the disease is also unclear, although there are similarities with the immunology of idiopathic pulmonary arterial hypertension (PAH). Finally, the treatment of Sch-PH has not been well studied. There is some evidence on treating the underlying infection, with unclear effect on Sch-PH, and advanced PAH therapies are now being suggested, but more studies are needed to confirm their efficacy.

Microfluidics-based nanoLC-MS/MS (chipLC-MS/MS) was used to identify and quantify proteins in epithelial lining fluid (ELF), collected during bronchoscopy from the main bronchi of chronic obstructive pulmonary disease (COPD) patients and healthy controls using microprobes. ELF is a biofluid that is well suited to study pathophysiological processes in the lung, because it contains high concentrations of biologically active molecules. 1D-PAGE followed by in-gel tryptic digestion and chipLC-MS/MS resulted in identification of approximately 300 proteins. A comparative study of ELF from COPD patients and non-COPD controls using chemical stable isotope labeling (iTRAQ®-8Plex) showed that the levels of lactotransferrin, high-mobility group protein B1 (HMGB 1), alpha 1-antichymotrypsin and cofilin-1 differed significantly in ELF from COPD patients and non-COPD controls (p-values < 0.05). These results were reproduced in another, independent set of ELF samples from COPD patients and non-COPD controls and further validated by immunohistochemistry. This study shows the feasibility of performing chipLC-MS/MS and quantitative proteomics in human ELF.

UNLABELLED The ratio of tricuspid regurgitation velocity divided by the velocity-time integral of right ventricular outflow tract pulsed-wave Doppler tracing (TRV/VTI(RVOT) ) has been used to estimate pulmonary vascular resistance (PVR). However, this method has not been validated in chronic thromboembolic pulmonary hypertension (CTEPH). We assessed the utility of TRV/VTI(RVOT) in patients with CTEPH and PVR from 2 to 20 WU. All had right heart catheterization (RHC) within 2 days of echocardiography. TRV/VTI(RVOT) was calculated and RHC-derived pressures, PVR, and cardiac outputs were recorded. Mean pulmonary artery pressure was 47 ± 12 mmHg, cardiac output: 4.2 ± 1.1 L/min, PVR: 9 ± 4 WU, right atrial pressure: 12 ± 6 mmHg. Mean VTI(RVOT) was 13 ± 5 cm; mean TRV was 4.2 ± 0.8 m/s, mean tricuspid regurgitation severity was 2.5 ± 0.8 (1 = trace, 2 = mild, 3 = moderate, 4 = severe). Regression analysis demonstrated a correlation between RHC PVR and TRV/VTI(RVOT) : PVR = 19.4 × (TRV/VTI(RVOT) ) + 2.4 (r = 0.74, P < 0.001). However, Bland-Altman analysis found a poor degree of agreement between echo-derived PVR and RHC PVR. We also studied 28 patients with non-CTEPH pulmonary hypertension. Similar analysis revealed a regression equation of PVR = 20.1 × (TRV/VTIR(RVOT) ) + 0.3 (r = 0.57, P < 0.01). CONCLUSION TRV/VTI(RVOT) is only marginally useful for estimating PVR in CTEPH (r = 0.74). Moreover, the regression equation in CTEPH differs significantly from previous studies in pulmonary hypertension. Reasons for this may include the markedly elevated PVR levels in this population and specific effects on VTI(RVOT) from CTEPH.

Pulmonary hypertension as a result of chronic thromboembolic disease (CTEPH) is potentially curable with pulmonary endarterectomy surgery. Consequently, correctly diagnosing patients with this type of pulmonary hypertension and evaluating these patients with the goal of establishing their candidacy for surgical intervention is of utmost importance. And as advancements in surgical techniques have allowed successful resection of segmental-level chronic thromboembolic disease, the number of CTEPH patients that are deemed suitable surgical candidates has expanded, making it even more important that the evaluation be conducted with greater precision. This article will review a diagnostic approach to patients with suspected chronic thromboembolic disease with an emphasis on the criteria considered in selecting patients for pulmonary endarterectomy surgery.