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Additional file 1: Text and Figures S1–S5. Supplementary text for the methods (study phenotype, quality control, imputation, significance and suggestive thresholds, annotation), results (power to detect associations, annotation of rs142755000), and discussion (suggestive associations) as well as related references. Figure S1. a: COPDGene XWAS in Stratified Populations All Subjects. b: COPDGene XWAS in Stratified Populations Males. c: COPDGene XWAS in Stratified Populations Females. Figure S2. Meta-analysis XWAS in Stratified Population. Figure S3. Meta-analysis Locus Plots of Top Suggested Associations in COPD Related Phenotypes. Figure S4. Meta-analysis Quantile–Quantile and Manhattan Plots. Figure S5. Sex differential edge weights connecting the transcription factor POU3F4 in GTEx Lung Tissue.

Additional file 1: Table 1. Baseline characteristics and demographics by treatment and Week 16 fibrinogen quartile. Table 2. Baseline characteristics and demographics by treatment and Week 16 fibrinogen 3.5 g/L threshold. Table 3. COPD exacerbation history by treatment group and by withdrawal status at Week 16 (ITT population). Table 4. Analysis of fibrinogen levels at Week 16 by treatment group. Table 5. Incidence and rates of AESIs by fibrinogen quartile. Figure 1. Rate of on-treatment moderate/severe exacerbations from Week 16 by continuous fibrinogen level at Week 16; (A) moderate exacerbations; (B) moderate/severe exacerbations; (C) severe exacerbations. Figure 2. Time-to-first on-treatment COPD exacerbation from Week 16 by Week 16 fibrinogen 3.5 g/L threshold: (A) moderate exacerbations; (B) moderate/severe exacerbations; (C) severe exacerbations.

PURPOSE: To correct the intensity difference of static background signal between bright blood images and dark blood images in subtractive non-contrast-enhanced MR angiography using robust regression, thereby improving static background signal suppression on subtracted angiograms. METHODS: Robust regression (RR), using iteratively reweighted least squares, is used to calculate the regression coefficient of background tissues from a scatter plot showing the voxel intensity of bright blood images versus dark blood images. The weighting function is based on either the Euclidean distance from the estimated regression line or the deviation angle. Results from RR using the deviation angle (RRDA), conventional RR using the Euclidean distance, and ordinary leastsquares regression were compared with reference values determined manually by two observers. Performance was evaluated over studies using different sequences, including 36 thoracic flow-sensitive dephasing data sets, 13 iliac flow-sensitive dephasing data sets, and 26 femoral fresh blood imaging data sets. RESULTS: RR deviation angle achieved robust and accurate performance in all types of images, with small bias, small mean absolute error, and high-correlation coefficients with reference values. Background tissues, such as muscle, veins, and bladder, were suppressed while the vascular signal was preserved. Euclidean distance gave good performance for thoracic and iliac flow-sensitive dephasing, but could not suppress background tissues in femoral fresh blood imaging. Ordinary least squares regression was sensitive to outliers and overestimated regression coefficients in thoracic flow-sensitive dephasing. CONCLUSION: Weighted subtraction using RR was able to acquire the regression coefficients of background signal and improve background suppression of subtractive non-contrast-enhanced MR angiography techniques. RR deviation angle has the most robust and accurate overall performance among three regression methods.

PURPOSE: To evaluate the performance of acceleration-dependent vascular anatomy for non-contrast-enhanced MR venography (ADVANCE-MRV) in femoral veins and to investigate whether venous signal uniformity can be improved by applying multiple acquisitions with different flow suppressions or multiple flow suppressions in 1 acquisition. METHODS: The ADVANCE-MRV method uses flow-sensitized modules to acquire a dark-artery image set and a dark-artery vein set, which are subsequently subtracted. Ten healthy volunteers were imaged using the ADVANCE-MRV sequence with improved venous suppression uniformity in the dark-artery vein images achieved by applying multiple flow suppressions in the same acquisition or by combining multiple images acquired with different flow suppressions. The performance of the improved technique was also evaluated in 13 patients with lower-limb deep venous thrombosis. RESULTS: Multiple-preparation and multiple-acquisition approaches all improved venous signal uniformity and reduced the signal void artifacts observed in the original ADVANCE-MRV images. The multiple-acquisition approaches achieved excellent blood signal uniformity and intensity, albeit at the cost of an increase in the total acquisition time. The double-preparation approach demonstrated good performance in all measurements, providing a good compromise between signal uniformity and acquisition time. The blood signal spatial variation and its variation using different gradient amplitudes were reduced by 20% and 29%. All patient images showed uniform and bright venous signal in nonoccluded sections of vein. CONCLUSION: The enhanced ADVANCE-MRV methods substantially improved signal uniformity in healthy volunteers and patients with known deep venous thrombosis. The double-preparation approach gave good-quality femoral vein images, providing improved venous signal uniformity without increasing acquisition time in comparison to the original sequence.

Additional file 1: Table S1. ECLIPSE Sample Demographics. Descriptive values for COPD patients (N = 114) in ECLIPSE with gene expression data by Cachectic (N = 9) and Non-Cachectic (N = 105) co-morbid status. Unless otherwise noted values denote mean (SD). Table S2. Gene set enrichment analysis of 23 significantly (FDR p-value

Additional file 1: Supplementary Table 1. SMQs, sub-SMQs, and Preferred Terms for on-treatment CVAESI that were reported in the IMPACT study Supplementary Table 2. Summary of on-treatment CVAESI by baseline CV risk factors (ITT population)

Funder: Ministero dell'Istruzione, dell'Università e della Ricerca Dipartimenti di Eccellenza 2018-2022 grant to the Molecular Medicine Department (University of Pavia), The availability of a genetic model organism with which to study key molecular events underlying amyloidogenesis is crucial for elucidating the mechanism of the disease and the exploration of new therapeutic avenues. The natural human variant of β2-microglobulin (D76N β2-m) is associated with a fatal familial form of systemic amyloidosis. Hitherto, no animal model has been available for studying in vivo the pathogenicity of this protein. We have established a transgenic C. elegans line, expressing the human D76N β2-m variant. Using the INVertebrate Automated Phenotyping Platform (INVAPP) and the algorithm Paragon, we were able to detect growth and motility impairment in D76N β2-m expressing worms. We also demonstrated the specificity of the β2-m variant in determining the pathological phenotype by rescuing the wild type phenotype when β2-m expression was inhibited by RNA interference (RNAi). Using this model, we have confirmed the efficacy of doxycycline, an inhibitor of the aggregation of amyloidogenic proteins, in rescuing the phenotype. In future, this C. elegans model, in conjunction with the INVAPP/Paragon system, offers the prospect of high-throughput chemical screening in the search for new drug candidates.

Afisi S Ismaila,1,2 Nancy Risebrough,3 Melanie Schroeder,4 Dhvani Shah,5 Alan Martin,6 Emma C Goodall,7 Kerigo Ndirangu,5 Gerard Criner,8 Mark Dransfield,9 David MG Halpin,10 MeiLan K Han,11 David A Lomas12 1Value Evidence and Outcomes, GlaxoSmithKline plc, Collegeville, PA, USA; 2Department of Health Research Methods, Evidence and Impact, McMaster University, Hamilton, ON, Canada; 3ICON Health Economics, ICON plc, Toronto, ON, Canada; 4Value Evidence and Outcomes, GlaxoSmithKline plc, Brentford, UK; 5ICON Health Economics, ICON plc, New York, NY, USA; 6Value Evidence and Outcomes, GlaxoSmithKline plc, Uxbridge, UK; 7Health Economics and Outcomes Research, GlaxoSmithKline plc, Mississauga, ON, Canada; 8Lewis Katz School of Medicine at Temple University, Philadelphia, PA, USA; 9Division of Pulmonary, Allergy, and Critical Care Medicine, Lung Health Center, University of Alabama at Birmingham, Birmingham, AL, USA; 10Department of Respiratory Medicine, Royal Devon and Exeter Hospital, Exeter, UK; 11University of Michigan, Pulmonary and Critical Care, Ann Arbor, MI, USA; 12UCL Respiratory, University College London, London, UKCorrespondence: Afisi S IsmailaValue Evidence and Outcomes, GlaxoSmithKline plc, 1250 South Collegeville Road, Collegeville, PA 19426-0989, USATel +1 919 315 8229Email afisi.s.ismaila@gsk.comBackground: We assessed the cost-effectiveness of single-inhaler fluticasone furoate (FF)/umeclidinium (UMEC)/vilanterol (VI) versus FF/VI or UMEC/VI from a Canadian public healthcare perspective, incorporating data from the IMPACT trial in chronic obstructive pulmonary disease (COPD) (NCT02164513).Methods: Baseline inputs and treatment effects from IMPACT were populated into the validated GALAXY-COPD disease progression model. Canadian unit costs and drug costs (Canadian dollars [C$], 2017) were applied to healthcare resource utilization and treatments. Future costs and health outcomes were discounted at 1.5% annually. Analyses were probabilistic, and outputs included exacerbation rates, costs, and life years (LYs) and quality-adjusted life years (QALYs) gained.Results: Compared with FF/VI and UMEC/VI over a lifetime horizon, the analyses predicted that treatment with FF/UMEC/VI resulted in fewer moderate and severe exacerbations, more LYs and more QALYs gained, with a small incremental cost. The base-case incremental cost-effectiveness ratio (ICER) per QALY gained was C$18,989 (95% confidence interval [CI]: C$14,665, C$25,753) versus FF/VI and C$13,776 (95% CI: C$9787, C$19,448) versus UMEC/VI. FF/UMEC/VI remained cost-effective versus both FF/VI and UMEC/VI in all sensitivity analyses, including in scenario analyses that considered different intervention and comparator discontinuation rates, and treatment effects for subsequent therapy.Conclusion: Treatment with FF/UMEC/VI was predicted to improve outcomes and be a cost-effective treatment option for patients with symptomatic COPD and a history of exacerbations compared with FF/VI or UMEC/VI, in Canada.Keywords: chronic obstructive pulmonary disease, cost-effectiveness, single-inhaler triple therapy, quality-adjusted life years, Canada

Chronic obstructive pulmonary disease (COPD) is the leading cause of respiratory mortality worldwide. Genetic risk loci provide new insights into disease pathogenesis. We performed a genome-wide association study in 35,735 cases and 222,076 controls from the UK Biobank and additional studies from the International COPD Genetics Consortium. We identified 82 loci associated with P < 5 × 10-8; 47 of these were previously described in association with either COPD or population-based measures of lung function. Of the remaining 35 new loci, 13 were associated with lung function in 79,055 individuals from the SpiroMeta consortium. Using gene expression and regulation data, we identified functional enrichment of COPD risk loci in lung tissue, smooth muscle, and several lung cell types. We found 14 COPD loci shared with either asthma or pulmonary fibrosis. COPD genetic risk loci clustered into groups based on associations with quantitative imaging features and comorbidities. Our analyses provide further support for the genetic susceptibility and heterogeneity of COPD.

This study introduces the use of 'error-category mapping' in the interpretation of pharmacokinetic (PK) model parameter results derived from dynamic contrast-enhanced (DCE-) MRI data. Eleven patients with metastatic renal cell carcinoma were enrolled in a multiparametric study of the treatment effects of bevacizumab. For the purposes of the present analysis, DCE-MRI data from two identical pre-treatment examinations were analysed by application of the extended Tofts model (eTM), using in turn a model arterial input function (AIF), an individually-measured AIF and a sample-average AIF. PK model parameter maps were calculated. Errors in the signal-to-gadolinium concentration ([Gd]) conversion process and the model-fitting process itself were assigned to category codes on a voxel-by-voxel basis, thereby forming a colour-coded 'error-category map' for each imaged slice. These maps were found to be repeatable between patient visits and showed that the eTM converged adequately in the majority of voxels in all the tumours studied. However, the maps also clearly indicated sub-regions of low Gd uptake and of non-convergence of the model in nearly all tumours. The non-physical condition ve ≥ 1 was the most frequently indicated error category and appeared sensitive to the form of AIF used. This simple method for visualisation of errors in DCE-MRI could be used as a routine quality-control technique and also has the potential to reveal otherwise hidden patterns of failure in PK model applications.

Figure S2. Relationship between percent emphysema at baseline and Year 1 with cachexia and weight-loss. (PDF 5 kb)

Table S1. Characteristics of ECLIPSE Study COPD cases stratified by BMI category. Continuous variables (age, FEV1pp) are represented by median (IQR). * indicates Pâ

Figure S3. Relationship between 1-year change in emphysema with cachexia and weight-loss. (PDF 5 kb)

Figure S1. Flow-chart depicting number of participants included in analyses. (PDF 22 kb)

Table S2. Relationships between risk of death from any cause with consensus and weight loss (WL) definitions of cachexia including COPD cases who regained weight in COPD cases from ECLIPSE. Significant p-values are bolded. (DOCX 16 kb)

Why only 20% of smokers develop clinically relevant chronic obstructive pulmonary disease (COPD) was a puzzle for many years. Now, epidemiologic studies point clearly toward a large heritable component. The combination of genome-wide association studies and candidate gene analysis is helping to identify those genetic variants responsible for an individual's susceptibility to developing COPD. In this review, the current data implicating specific loci and genes in the pathogenesis of COPD are examined.

Graphical abstract, Highlights • Modelling liver disease due to α1-antitrypsin deficiency reveals disruption of the ER into isolated cisternae. • Comparative ‘omics’ between diseased hiPSC-derived and wild-type hepatocytes provides insights into disease mechanisms. • AKR1B10 was identified as a putative biomarker for α1-antitrypsin deficiency., Background & Aims α1-Antitrypsin deficiency (A1ATD) is an autosomal recessive disorder caused by mutations in the SERPINA1 gene. Individuals with the Z variant (Gly342Lys) retain polymerised protein in the endoplasmic reticulum (ER) of their hepatocytes, predisposing them to liver disease. The concomitant lack of circulating A1AT also causes lung emphysema. Greater insight into the mechanisms that link protein misfolding to liver injury will facilitate the design of novel therapies. Methods Human-induced pluripotent stem cell (hiPSC)-derived hepatocytes provide a novel approach to interrogate the molecular mechanisms of A1ATD because of their patient-specific genetic architecture and reflection of human physiology. To that end, we utilised patient-specific hiPSC hepatocyte-like cells (ZZ-HLCs) derived from an A1ATD (ZZ) patient, which faithfully recapitulated key aspects of the disease at the molecular and cellular level. Subsequent functional and “omics” comparisons of these cells with their genetically corrected isogenic-line (RR-HLCs) and primary hepatocytes/human tissue enabled identification of new molecular markers and disease signatures. Results Our studies showed that abnormal A1AT polymer processing (immobilised ER components, reduced luminal protein mobility and disrupted ER cisternae) occurred heterogeneously within hepatocyte populations and was associated with disrupted mitochondrial structure, presence of the oncogenic protein AKR1B10 and two upregulated molecular clusters centred on members of inflammatory (IL-18 and Caspase-4) and unfolded protein response (Calnexin and Calreticulin) pathways. These results were validated in a second patient-specific hiPSC line. Conclusions Our data identified novel pathways that potentially link the expression of Z A1AT polymers to liver disease. These findings could help pave the way towards identification of new therapeutic targets for the treatment of A1ATD. Lay summary This study compared the gene expression and protein profiles of healthy liver cells and those affected by the inherited disease α1-antitrypsin deficiency. This approach identified specific factors primarily present in diseased samples which could provide new targets for drug development. This study also demonstrates the interest of using hepatic cells generated from human-induced pluripotent stem cells to model liver disease in vitro for uncovering new mechanisms with clinical relevance.

α1-Antitrypsin is primarily synthesised in the liver, circulates to the lung and protects pulmonary tissues from proteolytic damage. The Z mutant (Glu342Lys) undergoes inactivating conformational change and polymerises. Polymers are retained within the hepatocyte endoplasmic reticulum (ER) in homozygous (PiZZ) individuals, predisposing the individuals to hepatic cirrhosis and emphysema. Latency is an analogous process of inactivating, intra-molecular conformational change and may co-occur with polymerisation. However, the relationship between latency and polymerisation remained unexplored in the absence of a suitable probe. We have developed a novel monoclonal antibody specific for latent α1-antitrypsin and used it in combination with a polymer-specific antibody, to assess the association of both conformers in vitro, in disease and during augmentation therapy. In vitro kinetics analysis showed polymerisation dominated the pathway but latency could be promoted by stabilising monomeric α1-antitrypsin. Polymers were extensively produced in hepatocytes and a cell line expressing Z α1-antitrypsin but the latent protein was not detected despite manipulation of the secretory pathway. However, α1-antitrypsin augmentation therapy contains latent α1-antitrypsin, as did the plasma of 63/274 PiZZ individuals treated with augmentation therapy but 0/264 who were not receiving this medication (p

The heritability of chronic obstructive pulmonary disease (COPD) cannot be fully explained by recognized genetic risk factors identified as achieving genome-wide significance. In addition, the combined contribution of genetic variation to COPD risk has not been fully explored. We sought to determine: (1) whether studies of variants from previous studies of COPD or lung function in a larger sample could identify additional associated variants, particularly for severe COPD; and (2) the impact of genetic risk scores on COPD. We genotyped 3,346 single-nucleotide polymorphisms (SNPs) in 2,588 cases (1,803 severe COPD) and 1,782 control subjects from four cohorts, and performed association testing with COPD, combining these results with existing genotyping data from 6,633 cases (3,497 severe COPD) and 5,704 control subjects. In addition, we developed genetic risk scores from SNPs associated with lung function and COPD and tested their discriminatory power for COPD-related measures. We identified significant associations between SNPs near PPIC (P = 1.28 × 10-8) and PPP4R4/SERPINA1 (P = 1.01 × 10-8) and severe COPD; the latter association may be driven by recognized variants in SERPINA1. Genetic risk scores based on SNPs previously associated with COPD and lung function had a modest ability to discriminate COPD (area under the curve, ∼0.6), and accounted for a mean 0.9-1.9% lower forced expiratory volume in 1 second percent predicted for each additional risk allele. In a large genetic association analysis, we identified associations with severe COPD near PPIC and SERPINA1. A risk score based on combining genetic variants had modest, but significant, effects on risk of COPD and lung function.

BackgroundThere have been a number of candidate gene association studies of cancer cachexia-related traits, but no genome-wide association study (GWAS) has been published to date. Cachexia presents in patients with a number of complex traits, including both cancer and COPD. The objective of the current investigation was to search for a shared genetic aetiology for change in body mass index (ΔBMI) among cancer and COPD by using GWAS data in the Framingham Heart Study.MethodsA linear mixed effects model accounting for age, sex, and change in smoking status was used to calculate ΔBMI in participants over 40 years of age with three consecutive BMI time points (n = 4162). Four GWAS of ΔBMI using generalized estimating equations were performed among 1085 participants with a cancer diagnosis, 204 with gastrointestinal (GI) cancer, 112 with lung cancer, and 237 with COPD to test for association with 418 365 single-nucleotide polymorphisms (SNPs).ResultsTwo SNPs reached a level of genome-wide significance (P

Serpins are important regulators of proteolytic pathways with an antiprotease activity that involves a conformational transition from a metastable to a hyperstable state. Certain mutations permit the transition to occur in the absence of a protease; when associated with an intermolecular interaction, this yields linear polymers of hyperstable serpin molecules, which accumulate at the site of synthesis. This is the basis of many pathologies termed the serpinopathies. We have previously identified a monoclonal antibody (mAb4B12) that, in single-chain form, blocks α1-antitrypsin (α1-AT) polymerisation in cells. Here, we describe the structural basis for this activity. The mAb4B12 epitope was found to encompass residues Glu32, Glu39 and His43 on helix A and Leu306 on helix I. This is not a region typically associated with the serpin mechanism of conformational change, and correspondingly the epitope was present in all tested structural forms of the protein. Antibody binding rendered β-sheet A — on the opposite face of the molecule — more liable to adopt an ‘open’ state, mediated by changes distal to the breach region and proximal to helix F. The allosteric propagation of induced changes through the molecule was evidenced by an increased rate of peptide incorporation and destabilisation of a preformed serpin–enzyme complex following mAb4B12 binding. These data suggest that prematurely shifting the β-sheet A equilibrium towards the ‘open’ state out of sequence with other changes suppresses polymer formation. This work identifies a region potentially exploitable for a rational design of ligands that is able to dynamically influence α1-AT polymerisation.

BackgroundPulmonary function decline is a major contributor to morbidity and mortality among smokers. Post bronchodilator FEV1 and FEV1/FVC ratio are considered the standard assessment of airflow obstruction. We performed a genome-wide association study (GWAS) in 9919 current and former smokers in the COPDGene study (6659 non-Hispanic Whites [NHW] and 3260 African Americans [AA]) to identify associations with spirometric measures (post-bronchodilator FEV1 and FEV1/FVC). We also conducted meta-analysis of FEV1 and FEV1/FVC GWAS in the COPDGene, ECLIPSE, and GenKOLS cohorts (total n = 13,532).ResultsAmong NHW in the COPDGene cohort, both measures of pulmonary function were significantly associated with SNPs at the 15q25 locus [containing CHRNA3/5, AGPHD1, IREB2, CHRNB4] (lowest p-value = 2.17 × 10(-11)), and FEV1/FVC was associated with a genomic region on chromosome 4 [upstream of HHIP] (lowest p-value = 5.94 × 10(-10)); both regions have been previously associated with COPD. For the meta-analysis, in addition to confirming associations to the regions near CHRNA3/5 and HHIP, genome-wide significant associations were identified for FEV1 on chromosome 1 [TGFB2] (p-value = 8.99 × 10(-9)), 9 [DBH] (p-value = 9.69 × 10(-9)) and 19 [CYP2A6/7] (p-value = 3.49 × 10(-8)) and for FEV1/FVC on chromosome 1 [TGFB2] (p-value = 8.99 × 10(-9)), 4 [FAM13A] (p-value = 3.88 × 10(-12)), 11 [MMP3/12] (p-value = 3.29 × 10(-10)) and 14 [RIN3] (p-value = 5.64 × 10(-9)).ConclusionsIn a large genome-wide association study of lung function in smokers, we found genome-wide significant associations at several previously described loci with lung function or COPD. We additionally identified a novel genome-wide significant locus with FEV1 on chromosome 9 [DBH] in a meta-analysis of three study populations.

α1-antitrypsin deficiency is the most widely recognised genetic disorder causing chronic obstructive pulmonary disease (COPD). Mutant Z α1-antitrypsin expression has previously been linked to intracellular accumulation and polymerisation of this proteinase inhibitor. Subsequently, this has been described to underlie an exaggerated endoplasmic reticulum stress response and enhanced nuclear factor-κB signalling. However, whether monocyte-derived macrophages display the same features remains unknown. Monocytes from homozygous PiZZ α1-antitrypsin deficiency patients and PiMM controls were cultured for 6 days in the presence of granulocyte-macrophage or macrophage colony-stimulating factor to obtain pro- and anti-inflammatory macrophages (mφ-1 and mφ-2, respectively). We first showed that, in contrast to monocytes, pre-stressed mφ-1 and mφ-2 from healthy blood donors display an enhanced endoplasmic reticulum stress response upon a lipopolysaccharide trigger (XBP1 splicing, CHOP, GADD34 and GRP78 mRNA). However, this endoplasmic reticulum stress response did not differ between monocyte-derived macrophages and monocytes from ZZ patients compared to MM controls. Furthermore, these ZZ cells do not secrete higher cytokine levels, and α1-antitrypsin polymers were not detectable by ELISA. These data suggest that monocyte-derived macrophages are not the local source of Z α1-antitrypsin polymers found in the lung and that endoplasmic reticulum stress and pro-inflammatory cytokine release is not altered.

The supplement for this manuscript contains the following information, tables, and figures. The COPD Foundation funding and a list of the COPDGene and ECLIPSE investigators are given in the supplement. Tables S1â S12 list the top SNPs with a p-value

Why only 20% of smokers develop clinically relevant chronic obstructive pulmonary disease (COPD) was a puzzle for many years. Now, epidemiologic studies point clearly toward a large heritable component. The combination of genome-wide association studies and candidate gene analysis is helping to identify those genetic variants responsible for an individual's susceptibility to developing COPD. In this review, the current data implicating specific loci and genes in the pathogenesis of COPD are examined.

UNLABELLED: Point mutants of alpha1 -antitrypsin (α1AT) form ordered polymers that are retained as inclusions within the endoplasmic reticulum (ER) of hepatocytes in association with neonatal hepatitis, cirrhosis, and hepatocellular carcinoma. These inclusions cause cell damage and predispose to ER stress in the absence of the classical unfolded protein response (UPR). The pathophysiology underlying this ER stress was explored by generating cell models that conditionally express wild-type (WT) α1AT, two mutants that cause polymer-mediated inclusions and liver disease (E342K [the Z allele] and H334D) and a truncated mutant (Null Hong Kong; NHK) that induces classical ER stress and is removed by ER-associated degradation. Expression of the polymeric mutants resulted in gross changes in the ER luminal environment that recapitulated the changes observed in liver sections from individuals with PI*ZZ α1AT deficiency. In contrast, expression of NHK α1AT caused electron lucent dilatation and expansion of the ER throughout the cell. Photobleaching microscopy in live cells demonstrated a decrease in the mobility of soluble luminal proteins in cells that express E342K and H334D α1AT, when compared to those that express WT and NHK α1AT (0.34 ± 0.05, 0.22 ± 0.03, 2.83 ± 0.30, and 2.84 ± 0.55 μm(2) /s, respectively). There was no effect on protein mobility within ER membranes, indicating that cisternal connectivity was not disrupted. Polymer expression alone was insufficient to induce the UPR, but the resulting protein overload rendered cells hypersensitive to ER stress induced by either tunicamycin or glucose depletion. CONCLUSION: Changes in protein diffusion provide an explanation for the cellular consequences of ER protein overload in mutants that cause inclusion body formation and α1AT deficiency.

El presente proyecto tiene como fin la obtención de una estructura para colocar paneles solares sobre tejados de naves industriales. Estas estructuras se rigen por la normativa básica de edificación y por el Código Técnico de Edificación (CTE). Se trata de un proyecto de investigación llevado a cabo por el departamento de Ingeniería de Diseño y Fabricación de la Universidad de Zaragoza. La motivación a la realización de este proyecto se debe al desarrollo de las energías renovables, en especial de la fotovoltaica. Ha aparecido la necesidad de desarrollar estructuras capaces de soportar paneles solares fotovoltaicos y, debido a la nueva normativa que obliga a las nuevas empresas a colocar en sus instalaciones nuevas paneles solares fotovoltaicos o a la instalación de éstos para la obtención de una fuente adicional de ingresos, ha aumentado la utilización de los mismos en las cubiertas de las naves industriales. Es por ello que resulta necesario dimensionar las estructuras portantes de los mismos de forma que sean capaces de soportar las cargas que puedan aparecer y transmitirlas eficazmente a la estructura de la nave. Las herramientas de diseño en ingeniería usadas para el desarrollo de nuevos productos han experimentado recientemente una evolución muy positiva, obteniendo estructuras nuevas óptimas en comparación a las obtenidas por medios tradicionales.

BACKGROUND: There is a need for biomarkers to better characterise individuals with COPD and to aid with the development of therapeutic interventions. A panel of putative blood biomarkers was assessed in a subgroup of the Evaluation of COPD Longitudinally to Identify Surrogate Endpoints (ECLIPSE) cohort. METHODS: Thirty-four blood biomarkers were assessed in 201 subjects with COPD, 37 ex-smoker controls with normal lung function and 37 healthy non-smokers selected from the ECLIPSE cohort. Biomarker repeatability was assessed using baseline and 3-month samples. Intergroup comparisons were made using analysis of variance, repeatability was assessed through Bland-Altman plots, and correlations between biomarkers and clinical characteristics were assessed using Spearman correlation coefficients. RESULTS: Fifteen biomarkers were significantly different in individuals with COPD when compared to former or non-smoker controls. Some biomarkers, including tumor necrosis factor-α and interferon-γ, were measurable in only a minority of subjects whilst others such as C-reactive protein showed wide variability over the 3-month replication period. Fibrinogen was the most repeatable biomarker and exhibited a weak correlation with 6-minute walk distance, exacerbation rate, BODE index and MRC dyspnoea score in COPD subjects. 33% (66/201) of the COPD subjects reported at least 1 exacerbation over the 3 month study with 18% (36/201) reporting the exacerbation within 30 days of the 3-month visit. CRP, fibrinogen interleukin-6 and surfactant protein-D were significantly elevated in those COPD subjects with exacerbations within 30 days of the 3-month visit compared with those individuals that did not exacerbate or whose exacerbations had resolved. CONCLUSIONS: Only a few of the biomarkers assessed may be useful in diagnosis or management of COPD where the diagnosis is based on airflow obstruction (GOLD). Further analysis of more promising biomarkers may reveal utility in subsets of patients. Fibrinogen in particular has emerged as a potentially useful biomarker from this cohort and requires further investigation. TRIAL REGISTRATION: SCO104960, clinicaltrials.gov identifier NCT00292552.

Human induced pluripotent stem cells (iPSCs) represent a unique opportunity for regenerative medicine because they offer the prospect of generating unlimited quantities of cells for autologous transplantation, with potential application in treatments for a broad range of disorders. However, the use of human iPSCs in the context of genetically inherited human disease will require the correction of disease-causing mutations in a manner that is fully compatible with clinical applications. The methods currently available, such as homologous recombination, lack the necessary efficiency and also leave residual sequences in the targeted genome. Therefore, the development of new approaches to edit the mammalian genome is a prerequisite to delivering the clinical promise of human iPSCs. Here we show that a combination of zinc finger nucleases (ZFNs) and piggyBac technology in human iPSCs can achieve biallelic correction of a point mutation (Glu342Lys) in the α(1)-antitrypsin (A1AT, also known as SERPINA1) gene that is responsible for α(1)-antitrypsin deficiency. Genetic correction of human iPSCs restored the structure and function of A1AT in subsequently derived liver cells in vitro and in vivo. This approach is significantly more efficient than any other gene-targeting technology that is currently available and crucially prevents contamination of the host genome with residual non-human sequences. Our results provide the first proof of principle, to our knowledge, for the potential of combining human iPSCs with genetic correction to generate clinically relevant cells for autologous cell-based therapies.

Background There have been a number of candidate gene association studies of cancer cachexia-related traits, but no genome-wide association study (GWAS) has been published to date. Cachexia presents in patients with a number of complex traits, including both cancer and COPD. The objective of the current investigation was to search for a shared genetic aetiology for change in body mass index (Delta BMI) among cancer and COPD by using GWAS data in the Framingham Heart Study.Methods A linear mixed effects model accounting for age, sex, and change in smoking status was used to calculate Delta BMI in participants over 40 years of age with three consecutive BMI time points (n = 4162). Four GWAS of Delta BMI using generalized estimating equations were performed among 1085 participants with a cancer diagnosis, 204 with gastrointestinal (GI) cancer, 112 with lung cancer, and 237 with COPD to test for association with 418 365 single-nucleotide polymorphisms (SNPs).Results Two SNPs reached a level of genome-wide significance (P Conclusions In sum, one statistically significant common variant in the DOCK1 gene was associated with Delta BMI in GI cancer and COPD cases providing support for at least partially shared aetiology of Delta BMI in complex diseases.

Our purpose is to investigate the feasibility of imaging tumor metabolism in breast cancer patients using 13C magnetic resonance spectroscopic imaging (MRSI) of hyperpolarized 13C label exchange between injected [1-13C]pyruvate and the endogenous tumor lactate pool. Treatment-naïve breast cancer patients were recruited: four triple-negative grade 3 cancers; two invasive ductal carcinomas that were estrogen and progesterone receptor-positive (ER/PR+) and HER2/neu-negative (HER2-), one grade 2 and one grade 3; and one grade 2 ER/PR+ HER2- invasive lobular carcinoma (ILC). Dynamic 13C MRSI was performed following injection of hyperpolarized [1-13C]pyruvate. Expression of lactate dehydrogenase A (LDHA), which catalyzes 13C label exchange between pyruvate and lactate, hypoxia-inducible factor-1 (HIF1α), and the monocarboxylate transporters MCT1 and MCT4 were quantified using immunohistochemistry and RNA sequencing. We have demonstrated the feasibility and safety of hyperpolarized 13C MRI in early breast cancer. Both intertumoral and intratumoral heterogeneity of the hyperpolarized pyruvate and lactate signals were observed. The lactate-to-pyruvate signal ratio (LAC/PYR) ranged from 0.021 to 0.473 across the tumor subtypes (mean ± SD: 0.145 ± 0.164), and a lactate signal was observed in all of the grade 3 tumors. The LAC/PYR was significantly correlated with tumor volume (R = 0.903, P = 0.005) and MCT 1 (R = 0.85, P = 0.032) and HIF1α expression (R = 0.83, P = 0.043). Imaging of hyperpolarized [1-13C]pyruvate metabolism in breast cancer is feasible and demonstrated significant intertumoral and intratumoral metabolic heterogeneity, where lactate labeling correlated with MCT1 expression and hypoxia.

Human induced pluripotent stem cells (iPSCs) represent a unique opportunity for regenerative medicine because they offer the prospect of generating unlimited quantities of cells for autologous transplantation, with potential application in treatments for a broad range of disorders. However, the use of human iPSCs in the context of genetically inherited human disease will require the correction of disease-causing mutations in a manner that is fully compatible with clinical applications. The methods currently available, such as homologous recombination, lack the necessary efficiency and also leave residual sequences in the targeted genome. Therefore, the development of new approaches to edit the mammalian genome is a prerequisite to delivering the clinical promise of human iPSCs. Here we show that a combination of zinc finger nucleases (ZFNs) and piggyBac technology in human iPSCs can achieve biallelic correction of a point mutation (Glu342Lys) in the α(1)-antitrypsin (A1AT, also known as SERPINA1) gene that is responsible for α(1)-antitrypsin deficiency. Genetic correction of human iPSCs restored the structure and function of A1AT in subsequently derived liver cells in vitro and in vivo. This approach is significantly more efficient than any other gene-targeting technology that is currently available and crucially prevents contamination of the host genome with residual non-human sequences. Our results provide the first proof of principle, to our knowledge, for the potential of combining human iPSCs with genetic correction to generate clinically relevant cells for autologous cell-based therapies.

The availability of a genetic model organism with which to study key molecular events underlying amyloidogenesis is crucial for elucidating the mechanism of the disease and the exploration of new therapeutic avenues. The natural human variant of β2-microglobulin (D76N β2-m) is associated with a fatal familial form of systemic amyloidosis. Hitherto, no animal model has been available for studying in vivo the pathogenicity of this protein. We have established a transgenic C. elegans line, expressing the human D76N β2-m variant. Using the INVertebrate Automated Phenotyping Platform (INVAPP) and the algorithm Paragon, we were able to detect growth and motility impairment in D76N β2-m expressing worms. We also demonstrated the specificity of the β2-m variant in determining the pathological phenotype by rescuing the wild type phenotype when β2-m expression was inhibited by RNA interference (RNAi). Using this model, we have confirmed the efficacy of doxycycline, an inhibitor of the aggregation of amyloidogenic proteins, in rescuing the phenotype. In future, this C. elegans model, in conjunction with the INVAPP/Paragon system, offers the prospect of high-throughput chemical screening in the search for new drug candidates.

The human Aβ peptide causes progressive paralysis when expressed in the muscles of the nematode worm, C. elegans. We have exploited this model of Aβ toxicity by carrying out an RNAi screen to identify genes whose reduced expression modifies the severity of this locomotor phenotype. Our initial finding was that none of the human orthologues of these worm genes is identical with the genome-wide significant GWAS genes reported to date (the "white zone"); moreover there was no identity between worm screen hits and the longer list of GWAS genes which included those with borderline levels of significance (the "grey zone"). This indicates that Aβ toxicity should not be considered as equivalent to sporadic AD. To increase the sensitivity of our analysis, we then considered the physical interactors (+1 interactome) of the products of the genes in both the worm and the white+grey zone lists. When we consider these worm and GWAS gene lists we find that 4 of the 60 worm genes have a +1 interactome overlap that is larger than expected by chance. Two of these genes form a chaperonin complex, the third is closely associated with this complex and the fourth gene codes for actin, the major substrate of the same chaperonin.

Chronic obstructive pulmonary disease (COPD) is the leading cause of respiratory mortality worldwide. Genetic risk loci provide new insights into disease pathogenesis. We performed a genome-wide association study in 35,735 cases and 222,076 controls from the UK Biobank and additional studies from the International COPD Genetics Consortium. We identified 82 loci associated with P < 5 × 10-8; 47 of these were previously described in association with either COPD or population-based measures of lung function. Of the remaining 35 new loci, 13 were associated with lung function in 79,055 individuals from the SpiroMeta consortium. Using gene expression and regulation data, we identified functional enrichment of COPD risk loci in lung tissue, smooth muscle, and several lung cell types. We found 14 COPD loci shared with either asthma or pulmonary fibrosis. COPD genetic risk loci clustered into groups based on associations with quantitative imaging features and comorbidities. Our analyses provide further support for the genetic susceptibility and heterogeneity of COPD.

Endoplasmic reticulum (ER) dysfunction might have an important part to play in a range of neurological disorders, including cerebral ischaemia, sleep apnoea, Alzheimer's disease, multiple sclerosis, amyotrophic lateral sclerosis, the prion diseases, and familial encephalopathy with neuroserpin inclusion bodies. Protein misfolding in the ER initiates the well studied unfolded protein response in energy-starved neurons during stroke, which is relevant to the toxic effects of reperfusion. The toxic peptide amyloid β induces ER stress in Alzheimer's disease, which leads to activation of similar pathways, whereas the accumulation of polymeric neuroserpin in the neuronal ER triggers a poorly understood ER-overload response. In other neurological disorders, such as Parkinson's and Huntington's diseases, ER dysfunction is well recognised but the mechanisms by which it contributes to pathogenesis remain unclear. By targeting components of these signalling responses, amelioration of their toxic effects and so the treatment of a range of neurodegenerative disorders might become possible.

The accumulation of β-amyloid (Aβ) peptide in the brain is one of the pathological hallmarks of Alzheimer's disease and is thought to be of primary aetiological significance. In an unbiased genetic screen, we identified puromycin-sensitive aminopeptidase (PSA) as a potent suppressor of Aβ toxicity in a Drosophila model system. We established that coexpression of Drosophila PSA (dPSA) in the flies' brains improved their lifespan, protected against locomotor deficits, and reduced brain Aβ levels by clearing the Aβ plaque-like deposits. However, confocal microscopy and subcellular fractionation of amyloid-expressing 7PA2 cells demonstrated that PSA localizes to the cytoplasm. Therefore, PSA and Aβ are unlikely to be in the same cellular compartment; moreover, when we artificially placed them in the same compartment in flies, we could not detect a direct epistatic interaction. The consequent hypothesis that PSA's suppression of Aβ toxicity is indirect was supported by the finding that Aβ is not a proteolytic substrate for PSA in vitro. Furthermore, we showed that the enzymatic activity of PSA is not required for rescuing Aβ toxicity in neuronal SH-SY5Y cells. We investigated whether the stimulation of autophagy by PSA was responsible for these protective effects. However PSA's promotion of autophagosome fusion with lysosomes required proteolytic activity and so its effect on autophagy is not identical to its protection against Aβ toxicity.

Introductive and objectives. The airflow limitation of COPD results....

Severe α1‐antitrypsin deficiency results from the Z allele (Glu342Lys) that causes the accumulation of homopolymers of mutant α1‐antitrypsin within the endoplasmic reticulum of hepatocytes in association with liver disease. We have used a DNA‐encoded chemical library to undertake a high‐throughput screen to identify small molecules that bind to, and stabilise Z α1‐antitrypsin. The lead compound blocks Z α1‐antitrypsin polymerisation in vitro, reduces intracellular polymerisation and increases the secretion of Z α1‐antitrypsin threefold in an iPSC model of disease. Crystallographic and biophysical analyses demonstrate that GSK716 and related molecules bind to a cryptic binding pocket, negate the local effects of the Z mutation and stabilise the bound state against progression along the polymerisation pathway. Oral dosing of transgenic mice at 100 mg/kg three times a day for 20 days increased the secretion of Z α1‐antitrypsin into the plasma by sevenfold. There was no observable clearance of hepatic inclusions with respect to controls over the same time period. This study provides proof of principle that “mutation ameliorating” small molecules can block the aberrant polymerisation that underlies Z α1‐antitrypsin deficiency., A chemistry campaign has developed a small molecule that stabilises the severe Z deficiency mutant of α1‐antitrypsin. The lead compound binds to a cryptic pocket and blocks the conformational change and pathological polymerisation that underlie α1‐antitrypsin deficiency.

$\textit{Purpose:}$ Hepatic magnetic resonance elastography (MRE) is currently a breath-hold imaging technique. Patients with chronic liver disease can have comorbidities that limit their ability to breath-hold (BH) for the required acquisition time. Our aim was to evaluate whether stiffness measurements obtained from a navigator-triggered MRE acquisition are comparable to standard expiratory breath-hold, inspiratory breath-hold or free-breathing in healthy participants. $\textit{Materials and methods:}$ Twelve healthy participants were imaged using the four methods on a clinical 1.5 T MR system equipped with a product MRE system. Mean liver stiffness, and measurable area of stiffness (with a confidence threshold >95%) were compared between sequences using the concordance correlation coefficient. Repeatability of each sequence between two acquisitions was also assessed. $\textit{Results:}$ The standard BH expiratory technique had high concordance with the navigated technique (r = 0.716), and low concordance with the BH inspiration (r = 0.165) and free-breathing (r = 0.105) techniques. The navigator-triggered technique showed no statistical difference in measurable area of liver or in repeatability compared with the standard expiratory acquisition (p = 0.997 and p = 0.407 respectively). The free-breathing technique produced less measurable liver area and was less repeatable than the alternative techniques. The increase in acquisition time for navigator techniques was 3 min 6 s compared to standard expiratory breath-hold. $\textit{Conclusion:}$ Navigator-based hepatic MRE measurements are comparable to the reference standard expiratory breath-hold acquisition in healthy participants.

Summary Objectives We have developed a new grading system for hip osteoarthritis using clinical computed tomography (CT). This technique was compared with Kellgren and Lawrence (K&L) grading and minimum joint space width (JSW) measurement in digitally reconstructed radiographs (DRRs) from the same CT data. In this paper we evaluate and compare the accuracy and reliability of these measures in the assessment of radiological disease. Design CT imaging of hips from 30 female volunteers aged 66 ± 17 years were used in two reproducibility studies, one testing the reliability of the new system, the other testing K&L grading and minimum JSW measurement in DRRs. Results Intra- and inter-observer reliability was substantial for CT grading according to weighted kappa (0.74 and 0.75 respectively), while intra- and inter-observer reliability was at worst moderate (0.57) and substantial (0.63) respectively for DRR K&L grading. Bland–Altman analysis showed a systematic difference in minimum JSW measurement of 0.82 mm between reviewers, with a least detectable difference of 1.06 mm. The area under the curve from ROC analysis was 0.91 for our CT composite score. Conclusions CT grading of hip osteoarthritis (categorised as none, developing and established) has substantial reliability. Sensitivity was increased when CT features of osteoarthritis were assigned a composite score (0 = none to 7 = severest) that also performed well as a diagnostic test, but at the cost of reliability. Having established feasibility and reliability for this new CT system, sensitivity testing and validation against clinical measures of hip osteoarthritis will now be performed.

To aid the extraction of the portal venous input function (PVIF) from axial dynamic contrast-enhanced CT images of the liver, eliminating the need for full manual outlining of the vessel across time points.A cohort of 20 patients undergoing perfusion CT imaging of the liver was examined. Dynamic images of the liver were reformatted into contiguous thin slices. A region of interest was defined within a transverse section of the portal vein on a single contrast-enhanced image. This region of interest was then computationally projected across all thin slices for all time points to yield a semi-automated PVIF curve. This was compared against the "gold-standard" PVIF curve obtained by conventional manual outlining.Bland-Altman plots of curve characteristics indicated no substantial difference between automated and manual PVIF curves [concordance correlation coefficient in the range (0.66, 0.98)]. No substantial differences were shown by Bland-Altman plots of derived pharmacokinetic parameters when a suitable kinetic model was applied in each case [concordance correlation coefficient in range (0.92, 0.95)].This semi-automated method of extracting the PVIF performed equivalently to a "gold-standard" manual method for assessing liver function. Advances in knowledge: This technique provides a quick, simple and effective solution to the problems incurred by respiration motion and partial volume factors in the determination of the PVIF in liver dynamic contrast-enhanced CT.

Acute renal impairment is common in patients with chronic liver disease, occurring in approximately 19% of hospitalised patients with cirrhosis. A variety of types of renal impairment are recognised. The most important of these is the hepatorenal syndrome, a functional renal impairment due to circulatory and neurohormonal abnormalities that underpin cirrhosis. It is one of the most severe complications of cirrhosis with survival often measured in weeks to months. A variety of treatment options exist with early diagnosis and appropriate treatment providing the best hope for cure. This paper provides a comprehensive and up-to-date review of hepatorenal syndrome and lays out the topic according to the following sections: pathophysiology, historical developments, diagnostic criteria and limitations, epidemiology, precipitating factors, predictors, clinical and laboratory findings, prognosis, treatment options, prophylaxis, and conclusion.

OBJECTIVES: This work evaluates rapid magnetic resonance projection hydrography (PH) based amniotic fluid volume (AFV) estimates against established routine ultrasound single deepest vertical pocket (SDVP) and amniotic fluid index (AFI) measurements, in utero at 28-32 weeks gestation. Manual multi-section planimetry (MSP) based measurement of AFV is used as a proxy reference standard. METHODS: Thirty-five women with a healthy singleton pregnancy (20-41 years) attending routine antenatal ultrasound were recruited. SDVP and AFI were measured using ultrasound, with same day MRI assessing AFV with PH and MSP. The relationships between the respective techniques were assessed using linear regression analysis and Bland-Altman method comparison statistics. RESULTS: When comparing estimated AFV, a highly significant relationship was observed between PH and the reference standard MSP (R(2) = 0.802, p