Publications by authors named "Alen Faiz"

58 Publications

Determinants of Lung Fissure Completeness.

Am J Respir Crit Care Med 2021 Jun 14. Epub 2021 Jun 14.

University Medical Center Groningen, Pulmonary diseases, Groningen, Netherlands.

Rationale: New advanced bronchoscopic treatment options for patients with severe COPD have led to increased interest for COPD phenotyping, including fissure completeness.

Objectives: We investigated clinical, environmental, and genetic factors contributing to fissure completeness in patients with and without COPD.

Methods: We used data of 9926 participants of the COPDGene study who underwent chest computed tomography (CT). Fissure completeness was calculated from CT scans following quantitative CT analysis at baseline and five-year follow-up. The clinical and environmental factors sex, race, smoking, COPD, emphysema, maternal smoking during pregnancy and maternal COPD were tested for impact on fissure completeness. Genome-wide association analyses were performed separately in non-Hispanic whites and African-Americans.

Measurements And Main Results: African-Americans had significant higher fissure completeness than non-Hispanic whites for all three fissures (p<0.001). There was no change in fissure completeness between baseline and five-year follow-up. For all fissures, No clinically relevant differences in fissure completeness were found for other clinical or environmental factors, including COPD severity. Rs2173623, rs264866, rs2407284, rs7310342, rs4904145, rs6504172, and rs7209556 showed genome-wide significant associations with fissure completeness in non-Hispanic whites. In African-Americans, rs264866, rs4904145 and rs6504172 were identified as significant associations. Rs2173623, rs6504172, and rs7209556 lead to WNT5A and HOXB antisense RNA expression, which play an important role during embryogenesis.

Conclusions: Fissure completeness is genetically determined and not dependent on age, sex, smoking status, the presence and severity of COPD including exacerbation frequency, maternal smoking during pregnancy, or maternal COPD.
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http://dx.doi.org/10.1164/rccm.202102-0260OCDOI Listing
June 2021

expression in asthmatic epithelium depends on intronic methylation and ZNF263 binding.

ERJ Open Res 2021 Apr 7;7(2). Epub 2021 Jun 7.

Division of Paediatric Pneumology and Allergology, University Medical Centre Schleswig-Holstein, Airway Research Centre North, member of the German Centre for Lung Research (DZL), Lübeck, Germany.

Background: Reduction of COL4A3, one of the six isoforms of collagen 4, in asthmatic airways results in increased inflammation and angiogenesis, implicating it as a central part of asthma pathogenesis. However, to date, the path underlying these diminished COL4A3 levels has been elusive. This study investigated a possible mechanism underlying the reduction of COL4A3 expression.

Methods: Bronchial biopsies of 76 patients with asthma and 83 controls were subjected to RNA-sequencing and DNA methylation bead arrays to identify expression and methylation changes. The binding of ZNF263 was analysed by chromatin-immunoprecipitation sequencing coupled with quantitative (q)PCR. Effects of silencing, using small interfering RNA, on the expression were studied using qPCR.

Results: expression was significantly reduced in bronchial biopsies compared to healthy controls, whereas DNA methylation levels at cg11797365 were increased. expression levels were significantly low in asthmatics without inhaled corticosteroid (ICS) use, whereas the expression was not statistically different between asthmatics using ICS and controls. Methylation levels at cg11797365 were increased upon consecutive rhinovirus infections.

Conclusion: Our data indicate an epigenetic modification as a contributing factor for the loss of expression in asthmatic airway epithelium.
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http://dx.doi.org/10.1183/23120541.00802-2020DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8181658PMC
April 2021

Assessing the Anti-Inflammatory Activity of the Anxiolytic Drug Buspirone Using CRISPR-Cas9 Gene Editing in LPS-Stimulated BV-2 Microglial Cells.

Cells 2021 May 25;10(6). Epub 2021 May 25.

Laboratory of Cellular and Molecular Neuroscience (LCMN), School of Life Sciences, Faculty of Science, University of Technology Sydney, Sydney, NSW 2007, Australia.

Buspirone is an anxiolytic drug with robust serotonin receptor 1A (Htr1a) agonist activities. However, evidence has demonstrated that this drug also targets the dopamine D3 receptor (Drd3), where it acts as a potent antagonist. In vivo, Drd3 blockade is neuroprotective and reduces inflammation in models of Parkinson's disease. To test if buspirone also elicited anti-inflammatory activities in vitro, we generated stable Drd3 and Htr1a BV2 microglial cell lines using CRISPR-Cas9 technology and then tested the effects of buspirone after lipopolysaccharide (LPS) challenge. We found that LPS exposure had no effect on cell viability, except in Htr1a cells, where viability was reduced ( < 0.001). Drug treatment reduced viability in Drd3 cells, but not in WT or Htr1a cells. Buspirone counteracted LPS-induced NO release, NOS2, IL-1β and TNF-α gene expression in WT cells, whereas it exerted limited effects in Drd3 or Htr1a microglia. In summary, our findings indicate that buspirone attenuates microglial polarization after LPS challenge. These results also highlight some major effects of Drd3 or Htr1a genetic ablation on microglial biology, raising important questions on the complex role of neurotransmitters in regulating microglia functions.
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http://dx.doi.org/10.3390/cells10061312DOI Listing
May 2021

Current-Smoking alters Gene Expression and DNA Methylation in the Nasal Epithelium of Asthmatics.

Am J Respir Cell Mol Biol 2021 May 14. Epub 2021 May 14.

University of Technology Sydney, 1994, Respiratory Bioinformatics and Molecular Biology (RBMB), School of Life Sciences, Sydney, New South Wales, Australia.

Current-smoking contributes to worsened asthma prognosis, more severe symptoms and limits the beneficial effects of corticosteroids. As the nasal epithelium can reflect smoking-induced changes in the lower airways, it is a relevant source to investigate changes in gene expression and DNA methylation. This study explores gene expression and DNA methylation changes in current and ex-smokers with asthma. Matched gene expression and epigenome-wide DNA methylation samples collected from nasal brushings of 55 patients enrolled in a clinical trial investigation of current and ex-smoker asthma patients were analysed. Differential gene expression and DNA methylation analyses were conducted comparing current- vs ex-smokers. Expression quantitative trait methylation (eQTM) analysis was completed to explore smoking relevant genes by CpG sites that differ between current and ex-smokers. To investigate the relevance of the smoking-associated DNA methylation changes for the lower airways, significant CpG sites were explored in bronchial biopsies from patients who had stopped smoking. 809 genes and 18,814 CpG sites were differentially associated with current-smoking in the nose. The cis-eQTM analysis uncovered 171 CpG sites whose methylation status associated with smoking-related gene expression, including AHRR, ALDH3A1, CYP1A1 and CYP1B1. Methylation status of CpG sites altered by current-smoking reversed with one-year smoking cessation. We confirm current-smoking alters epigenetic patterns and affects gene expression in the nasal epithelium of asthma patients, which is partially reversible in bronhcial biopsies after smoking cessation. We demonstrate the ability to discern molecular changes in the nasal epithelium, presenting this as a tool in future investigations into disease-relevant effects of tobacco smoke.
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http://dx.doi.org/10.1165/rcmb.2020-0553OCDOI Listing
May 2021

Bronchial gene expression signature associated with rate of subsequent FEV decline in individuals with and at risk of COPD.

Thorax 2021 May 10. Epub 2021 May 10.

Division of Computational Biomedicine, Boston University School of Medicine, Boston, Massachusetts, USA

Background: COPD is characterised by progressive lung function decline. Leveraging prior work demonstrating bronchial airway COPD-associated gene expression alterations, we sought to determine if there are alterations associated with differences in the rate of FEV decline.

Methods: We examined gene expression among ever smokers with and without COPD who at baseline had bronchial brushings profiled by Affymetrix microarrays and had longitudinal lung function measurements (n=134; mean follow-up=6.38±2.48 years). Gene expression profiles associated with the rate of FEV decline were identified by linear modelling.

Results: Expression differences in 171 genes were associated with rate of FEV decline (false discovery rate <0.05). The FEV decline signature was replicated in an independent dataset of bronchial biopsies from patients with COPD (n=46; p=0.018; mean follow-up=6.76±1.32 years). Genes elevated in individuals with more rapid FEV decline are significantly enriched among the genes altered by modulation of XBP1 in two independent datasets (Gene Set Enrichment Analysis (GSEA) p<0.05) and are enriched in mucin-related genes (GSEA p<0.05).

Conclusion: We have identified and replicated an airway gene expression signature associated with the rate of FEV decline. Aspects of this signature are related to increased expression of XBP1-regulated genes, a transcription factor involved in the unfolded protein response, and genes related to mucin production. Collectively, these data suggest that molecular processes related to the rate of FEV decline can be detected in airway epithelium, identify a possible indicator of FEV decline and make it possible to detect, in an early phase, ever smokers with and without COPD most at risk of rapid FEV decline.
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http://dx.doi.org/10.1136/thoraxjnl-2019-214476DOI Listing
May 2021

Acute cigarette smoke-induced eQTL affects formyl peptide receptor expression and lung function.

Respirology 2021 Mar 19;26(3):233-240. Epub 2020 Oct 19.

Respiratory Bioinformatics and Molecular Biology, University of Technology Sydney, Sydney, NSW, Australia.

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 < 0.05) inducible eQTL (rs3212855) was identified that induced a >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.
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http://dx.doi.org/10.1111/resp.13960DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7983955PMC
March 2021

Identifying a nasal gene expression signature associated with hyperinflation and treatment response in severe COPD.

Sci Rep 2020 10 15;10(1):17415. Epub 2020 Oct 15.

Department of Pulmonology, University of Groningen, University Medical Center Groningen, Hanzeplein 1, 9700 RB, Groningen, The Netherlands.

Hyperinflation contributes to dyspnea intensity in COPD. Little is known about the molecular mechanisms underlying hyperinflation and how inhaled corticosteroids (ICS) affect this important aspect of COPD pathophysiology. To investigate the effect of ICS/long-acting β-agonist (LABA) treatment on both lung function measures of hyperinflation, and the nasal epithelial gene-expression profile in severe COPD. 117 patients were screened and 60 COPD patients entered a 1-month run-in period on low-dose ICS/LABA budesonide/formoterol (BUD/F) 200/6 one inhalation b.i.d. Patients were then randomly assigned to 3-month treatment with either a high dose BDP/F 100/6 two inhalations b.i.d. (n = 31) or BUD/F 200/6 two inhalations b.i.d. (n = 29). Lung function measurements and nasal epithelial gene-expression were assessed before and after 3-month treatment and validated in independent datasets. After 3-month ICS/LABA treatment, residual volume (RV)/total lung capacity (TLC)% predicted was reduced compared to baseline (p < 0.05). We identified a nasal gene-expression signature at screening that associated with higher RV/TLC% predicted values. This signature, decreased by ICS/LABA treatment was enriched for genes associated with increased p53 mediated apoptosis was replicated in bronchial biopsies of COPD patients. Finally, this signature was increased in COPD patients compared to controls in nasal, bronchial and small airways brushings. Short-term ICS/LABA treatment improves RV/TLC% predicted in severe COPD. Furthermore, it decreases the expression of genes involved in the signal transduction by the p53 class mediator, which is a replicable COPD gene expression signature in the upper and lower airways.Trial registration: ClinicalTrials.gov registration number NCT01351792 (registration date May 11, 2011), ClinicalTrials.gov registration number NCT00848406 (registration date February 20, 2009), ClinicalTrials.gov registration number NCT00158847 (registration date September 12, 2005).
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http://dx.doi.org/10.1038/s41598-020-72551-0DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7562702PMC
October 2020

Genetic regulation of gene expression of MIF family members in lung tissue.

Sci Rep 2020 10 12;10(1):16980. Epub 2020 Oct 12.

Department of Molecular Pharmacology, Groningen Research Institute of Pharmacy (GRIP), University of Groningen, Groningen, The Netherlands.

Macrophage migration inhibitory factor (MIF) is a cytokine found to be associated with chronic obstructive pulmonary disease (COPD). However, there is no consensus on how MIF levels differ in COPD compared to control conditions and there are no reports on MIF expression in lung tissue. Here we studied gene expression of members of the MIF family MIF, D-Dopachrome Tautomerase (DDT) and DDT-like (DDTL) in a lung tissue dataset with 1087 subjects and identified single nucleotide polymorphisms (SNPs) regulating their gene expression. We found higher MIF and DDT expression in COPD patients compared to non-COPD subjects and found 71 SNPs significantly influencing gene expression of MIF and DDTL. Furthermore, the platform used to measure MIF (microarray or RNAseq) was found to influence the splice variants detected and subsequently the direction of the SNP effects on MIF expression. Among the SNPs found to regulate MIF expression, the major LD block identified was linked to rs5844572, a SNP previously found to be associated with lower diffusion capacity in COPD. This suggests that MIF may be contributing to the pathogenesis of COPD, as SNPs that influence MIF expression are also associated with symptoms of COPD. Our study shows that MIF levels are affected not only by disease but also by genetic diversity (i.e. SNPs). Since none of our significant eSNPs for MIF or DDTL have been described in GWAS for COPD or lung function, MIF expression in COPD patients is more likely a consequence of disease-related factors rather than a cause of the disease.
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http://dx.doi.org/10.1038/s41598-020-74121-wDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7552402PMC
October 2020

Gene signatures from scRNA-seq accurately quantify mast cells in biopsies in asthma.

Clin Exp Allergy 2020 12 23;50(12):1428-1431. Epub 2020 Sep 23.

Department of Pulmonology, University Medical Center Groningen, University of Groningen, Groningen, The Netherlands.

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http://dx.doi.org/10.1111/cea.13732DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7756890PMC
December 2020

Determinants of SARS-CoV-2 receptor gene expression in upper and lower airways.

medRxiv 2020 Sep 2. Epub 2020 Sep 2.

The recent outbreak of the severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2), which causes coronavirus disease 2019 (COVID-19), has led to a worldwide pandemic. One week after initial symptoms develop, a subset of patients progresses to severe disease, with high mortality and limited treatment options. To design novel interventions aimed at preventing spread of the virus and reducing progression to severe disease, detailed knowledge of the cell types and regulating factors driving cellular entry is urgently needed. Here we assess the expression patterns in genes required for COVID-19 entry into cells and replication, and their regulation by genetic, epigenetic and environmental factors, throughout the respiratory tract using samples collected from the upper (nasal) and lower airways (bronchi). Matched samples from the upper and lower airways show a clear increased expression of these genes in the nose compared to the bronchi and parenchyma. Cellular deconvolution indicates a clear association of these genes with the proportion of secretory epithelial cells. Smoking status was found to increase the majority of COVID-19 related genes including ACE2 and TMPRSS2 but only in the lower airways, which was associated with a significant increase in the predicted proportion of goblet cells in bronchial samples of current smokers. Both acute and second hand smoke were found to increase ACE2 expression in the bronchus. Inhaled corticosteroids decrease ACE2 expression in the lower airways. No significant effect of genetics on ACE2 expression was observed, but a strong association of DNA- methylation with ACE2 and TMPRSS2- mRNA expression was identified in the bronchus.
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http://dx.doi.org/10.1101/2020.08.31.20169946DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7480059PMC
September 2020

RAGE and TLR4 differentially regulate airway hyperresponsiveness: Implications for COPD.

Allergy 2021 04 15;76(4):1123-1135. Epub 2020 Oct 15.

Graduate School of Health, Faculty of Health, The University of Technology Sydney, Ultimo, NSW, Australia.

Background: The receptor for advanced glycation end products (RAGE) and Toll-like receptor 4 (TLR4) is implicated in COPD. Although these receptors share common ligands and signalling pathways, it is not known whether they act in concert to drive pathological processes in COPD. We examined the impact of RAGE and/or TLR4 gene deficiency in a mouse model of COPD and also determined whether expression of these receptors correlates with airway neutrophilia and airway hyperresponsiveness (AHR) in COPD patients.

Methods: We measured airway inflammation and AHR in wild-type, RAGE , TLR4 and TLR4 RAGE mice following acute exposure to cigarette smoke (CS). We also examined the impact of smoking status on AGER (encodes RAGE) and TLR4 bronchial gene expression in patients with and without COPD. Finally, we determined whether expression of these receptors correlates with airway neutrophilia and AHR in COPD patients.

Results: RAGE mice were protected against CS-induced neutrophilia and AHR. In contrast, TLR4 mice were not protected against CS-induced neutrophilia and had more severe CS-induced AHR. TLR4 RAGE mice were not protected against CS-induced neutrophilia but were partially protected against CS-induced mediator release and AHR. Current smoking was associated with significantly lower AGER and TLR4 expression irrespective of COPD status, possibly reflecting negative feedback regulation. However, consistent with preclinical findings, AGER expression correlated with higher sputum neutrophil counts and more severe AHR in COPD patients. TLR4 expression did not correlate with neutrophilic inflammation or AHR.

Conclusions: Inhibition of RAGE but not TLR4 signalling may protect against airway neutrophilia and AHR in COPD.
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http://dx.doi.org/10.1111/all.14563DOI Listing
April 2021

Phenotypic and functional translation of IL33 genetics in asthma.

J Allergy Clin Immunol 2021 Jan 19;147(1):144-157. Epub 2020 May 19.

Department of Health Sciences, University of Leicester, Leicester, United Kingdom; National Institute for Health Research Leicester Respiratory Biomedical Research Centre, Leicester, United Kingdom.

Background: Asthma is a complex disease with multiple phenotypes that may differ in disease pathobiology and treatment response. IL33 single nucleotide polymorphisms (SNPs) have been reproducibly associated with asthma. IL33 levels are elevated in sputum and bronchial biopsies of patients with asthma. The functional consequences of IL33 asthma SNPs remain unknown.

Objective: This study sought to determine whether IL33 SNPs associate with asthma-related phenotypes and with IL33 expression in lung or bronchial epithelium. This study investigated the effect of increased IL33 expression on human bronchial epithelial cell (HBEC) function.

Methods: Association between IL33 SNPs (Chr9: 5,815,786-6,657,983) and asthma phenotypes (Lifelines/DAG [Dutch Asthma GWAS]/GASP [Genetics of Asthma Severity & Phenotypes] cohorts) and between SNPs and expression (lung tissue, bronchial brushes, HBECs) was done using regression modeling. Lentiviral overexpression was used to study IL33 effects on HBECs.

Results: We found that 161 SNPs spanning the IL33 region associated with 1 or more asthma phenotypes after correction for multiple testing. We report a main independent signal tagged by rs992969 associating with blood eosinophil levels, asthma, and eosinophilic asthma. A second, independent signal tagged by rs4008366 presented modest association with eosinophilic asthma. Neither signal associated with FEV, FEV/forced vital capacity, atopy, and age of asthma onset. The 2 IL33 signals are expression quantitative loci in bronchial brushes and cultured HBECs, but not in lung tissue. IL33 overexpression in vitro resulted in reduced viability and reactive oxygen species-capturing of HBECs, without influencing epithelial cell count, metabolic activity, or barrier function.

Conclusions: We identify IL33 as an epithelial susceptibility gene for eosinophilia and asthma, provide mechanistic insight, and implicate targeting of the IL33 pathway specifically in eosinophilic asthma.
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http://dx.doi.org/10.1016/j.jaci.2020.04.051DOI Listing
January 2021

Phenotypic and functional translation of IL1RL1 locus polymorphisms in lung tissue and asthmatic airway epithelium.

JCI Insight 2020 04 23;5(8). Epub 2020 Apr 23.

Department of Human Development and.

The IL1RL1 (ST2) gene locus is robustly associated with asthma; however, the contribution of single nucleotide polymorphisms (SNPs) in this locus to specific asthma subtypes and the functional mechanisms underlying these associations remain to be defined. We tested for association between IL1RL1 region SNPs and characteristics of asthma as defined by clinical and immunological measures and addressed functional effects of these genetic variants in lung tissue and airway epithelium. Utilizing 4 independent cohorts (Lifelines, Dutch Asthma GWAS [DAG], Genetics of Asthma Severity and Phenotypes [GASP], and Manchester Asthma and Allergy Study [MAAS]) and resequencing data, we identified 3 key signals associated with asthma features. Investigations in lung tissue and primary bronchial epithelial cells identified context-dependent relationships between the signals and IL1RL1 mRNA and soluble protein expression. This was also observed for asthma-associated IL1RL1 nonsynonymous coding TIR domain SNPs. Bronchial epithelial cell cultures from asthma patients, exposed to exacerbation-relevant stimulations, revealed modulatory effects for all 4 signals on IL1RL1 mRNA and/or protein expression, suggesting SNP-environment interactions. The IL1RL1 TIR signaling domain haplotype affected IL-33-driven NF-κB signaling, while not interfering with TLR signaling. In summary, we identify that IL1RL1 genetic signals potentially contribute to severe and eosinophilic phenotypes in asthma, as well as provide initial mechanistic insight, including genetic regulation of IL1RL1 isoform expression and receptor signaling.
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http://dx.doi.org/10.1172/jci.insight.132446DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7205441PMC
April 2020

Gene expression profiling of bronchial brushes is associated with the level of emphysema measured by computed tomography-based parametric response mapping.

Am J Physiol Lung Cell Mol Physiol 2020 06 22;318(6):L1222-L1228. Epub 2020 Apr 22.

Respiratory Bioinformatics and Molecular Biology, School of Life Sciences, University of Technology Sydney, Sydney, New South Wales, Australia.

Parametric response mapping (PRM) is a computed tomography (CT)-based method to phenotype patients with chronic obstructive pulmonary disease (COPD). It is capable of differentiating emphysema-related air trapping with nonemphysematous air trapping (small airway disease), which helps to identify the extent and localization of the disease. Most studies evaluating the gene expression in smokers and COPD patients related this to spirometric measurements, but none have investigated the relationship with CT-based measurements of lung structure. The current study aimed to examine gene expression profiles of brushed bronchial epithelial cells in association with the PRM-defined CT-based measurements of emphysema (PRM) and small airway disease (PRM). Using the Top Institute Pharma (TIP) study cohort (COPD = 12 and asymptomatic smokers = 32), we identified a gene expression signature of bronchial brushings, which was associated with PRM in the lungs. One hundred thirty-three genes were identified to be associated with PRM. Among the most significantly associated genes, is a potent chemokine involved with CD8 T cell activation during inflammation in COPD, indicating that it may play an essential role in the development of emphysema. The PRM signature was then replicated in two independent data sets. Pathway analysis showed that the PRM signature is associated with proinflammatory and notch signaling pathways. Together these findings indicate that airway epithelium may play a role in the development of emphysema and/or may act as a biomarker for the presence of emphysema. In contrast, its role in relation to functional small airways disease is less clear.
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http://dx.doi.org/10.1152/ajplung.00051.2020DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7938773PMC
June 2020

ERS International Congress, Madrid, 2019: highlights from the Basic and Translational Science Assembly.

ERJ Open Res 2020 Jan 2;6(1). Epub 2020 Mar 2.

University of Groningen, University Medical Center Groningen, Depts of Pathology & Medical Biology and Pulmonology, Groningen, The Netherlands.

In this review, the Basic and Translational Sciences Assembly of the European Respiratory Society (ERS) provides an overview of the 2019 ERS International Congress highlights. In particular, we discuss how the novel and very promising technology of single cell sequencing has led to the development of a comprehensive map of the human lung, the lung cell atlas, including the discovery of novel cell types and new insights into cellular trajectories in lung health and disease. Further, we summarise recent insights in the field of respiratory infections, which can aid in a better understanding of the molecular mechanisms underlying these infections in order to develop novel vaccines and improved treatment options. Novel concepts delineating the early origins of lung disease are focused on the effects of pre- and post-natal exposures on neonatal lung development and long-term lung health. Moreover, we discuss how these early life exposures can affect the lung microbiome and respiratory infections. In addition, the importance of metabolomics and mitochondrial function analysis to subphenotype chronic lung disease patients according to their metabolic program is described. Finally, basic and translational respiratory science is rapidly moving forward and this will be beneficial for an advanced molecular understanding of the mechanisms underlying a variety of lung diseases. In the long-term this will aid in the development of novel therapeutic targeting strategies in the field of respiratory medicine.
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http://dx.doi.org/10.1183/23120541.00350-2019DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7049707PMC
January 2020

Sputum microbiome profiling in COPD: beyond singular pathogen detection.

Thorax 2020 04 29;75(4):338-344. Epub 2020 Jan 29.

Department of Pulmonary Diseases, University Medical Center Groningen, University of Groningen, Groningen, the Netherlands.

Culture-independent microbial sequencing techniques have revealed that the respiratory tract harbours a complex microbiome not detectable by conventional culturing methods. The contribution of the microbiome to chronic obstructive pulmonary disease (COPD) pathobiology and the potential for microbiome-based clinical biomarkers in COPD are still in the early phases of investigation. Sputum is an easily obtainable sample and has provided a wealth of information on COPD pathobiology, and thus has been a preferred sample type for microbiome studies. Although the sputum microbiome likely reflects the respiratory microbiome only in part, there is increasing evidence that microbial community structure and diversity are associated with disease severity and clinical outcomes, both in stable COPD and during the exacerbations. Current evidence has been limited to mainly cross-sectional studies using 16S rRNA gene sequencing, attempting to answer the question 'who is there?' Longitudinal studies using standardised protocols are needed to answer outstanding questions including differences between sputum sampling techniques. Further, with advancing technologies, microbiome studies are shifting beyond the examination of the 16S rRNA gene, to include whole metagenome and metatranscriptome sequencing, as well as metabolome characterisation. Despite being technically more challenging, whole-genome profiling and metabolomics can address the questions 'what can they do?' and 'what are they doing?' This review provides an overview of the basic principles of high-throughput microbiome sequencing techniques, current literature on sputum microbiome profiling in COPD, and a discussion of the associated limitations and future perspectives.
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http://dx.doi.org/10.1136/thoraxjnl-2019-214168DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7231454PMC
April 2020

Gene network approach reveals co-expression patterns in nasal and bronchial epithelium.

Sci Rep 2019 11 1;9(1):15835. Epub 2019 Nov 1.

University of Groningen, University Medical Center Groningen, Department of Pulmonology, Groningen, The Netherlands.

Nasal gene expression profiling is a new approach to investigate the airway epithelium as a biomarker to study the activity and treatment responses of obstructive pulmonary diseases. We investigated to what extent gene expression profiling of nasal brushings is similar to that of bronchial brushings. We performed genome wide gene expression profiling on matched nasal and bronchial epithelial brushes from 77 respiratory healthy individuals. To investigate differences and similarities among regulatory modules, network analysis was performed on correlated, differentially expressed and smoking-related genes using Gaussian Graphical Models. Between nasal and bronchial brushes, 619 genes were correlated and 1692 genes were differentially expressed (false discovery rate <0.05, |Fold-change|>2). Network analysis of correlated genes showed pro-inflammatory pathways to be similar between the two locations. Focusing on smoking-related genes, cytochrome-P450 pathway related genes were found to be similar, supporting the concept of a detoxifying response to tobacco exposure throughout the airways. In contrast, cilia-related pathways were decreased in nasal compared to bronchial brushes when focusing on differentially expressed genes. Collectively, while there are substantial differences in gene expression between nasal and bronchial brushes, we also found similarities, especially in the response to the external factors such as smoking.
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http://dx.doi.org/10.1038/s41598-019-50963-xDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6825243PMC
November 2019

Cigarette smoke exposure alters phosphodiesterases in human structural lung cells.

Am J Physiol Lung Cell Mol Physiol 2020 01 30;318(1):L59-L64. Epub 2019 Oct 30.

University of Groningen, Department of Molecular Pharmacology, Groningen, The Netherlands.

Cigarette smoke (CS), a highly complex mixture containing more than 4,000 compounds, causes aberrant cell responses leading to tissue damage around the airways and alveoli, which underlies various lung diseases. Phosphodiesterases (PDEs) are a family of enzymes that hydrolyze cyclic nucleotides. PDE inhibition induces bronchodilation, reduces the activation and recruitment of inflammatory cells, and the release of various cytokines. Currently, the selective PDE4 inhibitor roflumilast is an approved add-on treatment for patients with severe chronic obstructive pulmonary disease with chronic bronchitis and a history of frequent exacerbations. Additional selective PDE inhibitors are being tested in preclinical and clinical studies. However, the effect of chronic CS exposure on the expression of PDEs is unknown. Using mRNA isolated from nasal and bronchial brushes and lung tissues of never smokers and current smokers, we compared the gene expression of 25 PDE coding genes. Additionally, the expression and distribution of PDE3A and PDE4D in human lung tissues was examined. This study reveals that chronic CS exposure modulates the expression of various PDE members. Thus, CS exposure may change the levels of intracellular cyclic nucleotides and thereby impact the efficiency of PDE-targeted therapies.
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http://dx.doi.org/10.1152/ajplung.00319.2019DOI Listing
January 2020

Current Smoking is Associated with Decreased Expression of miR-335-5p in Parenchymal Lung Fibroblasts.

Int J Mol Sci 2019 Oct 18;20(20). Epub 2019 Oct 18.

University of Groningen, University Medical Center Groningen, Department of Pathology and Medical Biology, 9713 GZ Groningen, The Netherlands.

Cigarette smoking causes lung inflammation and tissue damage. Lung fibroblasts play a major role in tissue repair. Previous studies have reported smoking-associated changes in fibroblast responses and methylation patterns. Our aim was to identify the effect of current smoking on miRNA expression in primary lung fibroblasts. Small RNA sequencing was performed on lung fibroblasts from nine current and six ex-smokers with normal lung function. MiR-335-5p and miR-335-3p were significantly downregulated in lung fibroblasts from current compared to ex-smokers (false discovery rate (FDR) <0.05). Differential miR-335-5p expression was validated with RT-qPCR (-value = 0.01). The results were validated in lung tissue from current and ex-smokers and in bronchial biopsies from non-diseased smokers and never-smokers (-value <0.05). The methylation pattern of the miR-335 host gene, determined by methylation-specific qPCR, did not differ between current and ex-smokers. To obtain insights into the genes regulated by miR-335-5p in fibroblasts, we overlapped all proven miR-335-5p targets with our previously published miRNA targetome data in lung fibroblasts. This revealed , , and as likely targets of miR-335-5p in lung fibroblasts. Our study indicates that miR-335-5p downregulation due to current smoking may affect its function in lung fibroblasts by targeting , and .
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http://dx.doi.org/10.3390/ijms20205176DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6829537PMC
October 2019

MiR-31-5p: A shared regulator of chronic mucus hypersecretion in asthma and chronic obstructive pulmonary disease.

Allergy 2020 03 23;75(3):703-706. Epub 2019 Oct 23.

Groningen Research Institute for Asthma and COPD, University Medical Centre Groningen, University of Groningen, Groningen, The Netherlands.

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http://dx.doi.org/10.1111/all.14060DOI Listing
March 2020

The pharmacogenomics of inhaled corticosteroids and lung function decline in COPD.

Eur Respir J 2019 12 4;54(6). Epub 2019 Dec 4.

The University of British Columbia Center for Heart Lung Innovation, St Paul's Hospital Vancouver, BC, Canada.

Inhaled corticosteroids (ICS) are widely prescribed for patients with chronic obstructive pulmonary disease (COPD), yet have variable outcomes and adverse reactions, which may be genetically determined. The primary aim of the study was to identify the genetic determinants for forced expiratory volume in 1 s (FEV) changes related to ICS therapy.In the Lung Health Study (LHS)-2, 1116 COPD patients were randomised to the ICS triamcinolone acetonide (n=559) or placebo (n=557) with spirometry performed every 6 months for 3 years. We performed a pharmacogenomic genome-wide association study for the genotype-by-ICS treatment effect on 3 years of FEV changes (estimated as slope) in 802 genotyped LHS-2 participants. Replication was performed in 199 COPD patients randomised to the ICS, fluticasone or placebo.A total of five loci showed genotype-by-ICS interaction at p<5×10; of these, single nucleotide polymorphism (SNP) rs111720447 on chromosome 7 was replicated (discovery p=4.8×10, replication p=5.9×10) with the same direction of interaction effect. ENCODE (Encyclopedia of DNA Elements) data revealed that in glucocorticoid-treated (dexamethasone) A549 alveolar cell line, glucocorticoid receptor binding sites were located near SNP rs111720447. In stratified analyses of LHS-2, genotype at SNP rs111720447 was significantly associated with rate of FEV decline in patients taking ICS (C allele β 56.36 mL·year, 95% CI 29.96-82.76 mL·year) and in patients who were assigned to placebo, although the relationship was weaker and in the opposite direction to that in the ICS group (C allele β -27.57 mL·year, 95% CI -53.27- -1.87 mL·year).The study uncovered genetic factors associated with FEV changes related to ICS in COPD patients, which may provide new insight on the potential biology of steroid responsiveness in COPD.
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http://dx.doi.org/10.1183/13993003.00521-2019DOI Listing
December 2019

AGER expression and alternative splicing in bronchial biopsies of smokers and never smokers.

Respir Res 2019 Apr 10;20(1):70. Epub 2019 Apr 10.

Department of Pathology & Medical Biology, University Medical Center Groningen, University of Groningen, Hanzeplein 1, 9713 GZ, Groningen, The Netherlands.

Cigarette smoking is one of the major risk factors for the development of chronic obstructive pulmonary disease (COPD). Evidence is accumulating that Receptor for Advanced Glycation-End products (RAGE)-signaling is a key pathway in the pathophysiology of COPD. To date, it is unknown how smoking affects RAGE expression. In the current study, we investigated the effect of smoking on AGER, the gene encoding RAGE, expression and on alternative splicing of AGER. To this end, we conducted RNA-Seq on bronchial biopsies for asymptomatic smokers (n = 36) and never smokers (n = 40). Total AGER gene expression was accessed using DESeq2, while alternative splicing was investigated by measuring the number of specific split reads spanning exon-exon junctions and the total split reads. One of the major isoforms of RAGE is endogenous soluble (es) RAGE, an anti-inflammatory decoy receptor, making up for approximately 10% of the total amount of soluble (s)RAGE. We found that smokers show decreased total gene expression of AGER in bronchial biopsies, while the relative abundance of the esRAGE isoform is increased. Furthermore, no difference in the serum levels of total sRAGE were observed between smokers and non-smokers. Our data indicates that smoking initiates a protective anti-inflammatory mechanism with decreased expression of the pro-inflammatory gene AGER and increased relative abundance of the anti-inflammatory isoform esRAGE.
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http://dx.doi.org/10.1186/s12931-019-1038-6DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6456980PMC
April 2019

Effect of long-term corticosteroid treatment on microRNA and gene-expression profiles in COPD.

Eur Respir J 2019 04 25;53(4). Epub 2019 Apr 25.

University of Groningen, University Medical Center Groningen, Dept of Pulmonary Diseases, Groningen, The Netherlands.

The aim was to investigate whether microRNA (miRNA) expression is modulated by inhaled corticosteroid (ICS) treatmentWe performed genome-wide miRNA analysis on bronchial biopsies of 69 moderate/severe chronic obstructive pulmonary disease (COPD) patients at baseline and after 6- and 30-month treatment with the ICS fluticasone propionate or placebo. The effect of ICS on miRNA expression was validated in differentiated primary bronchial epithelial cultures, and functional studies were conducted in BEAS-2B cells. MiRNAs affected by ICS and their predicted targets were compared to an independent miRNA dataset of bronchial brushings from COPD patients and healthy controls.Treatment with ICS for both 6 and 30 months significantly altered the expression of four miRNAs, including miR-320d, which was increased during ICS treatment compared with placebo. The ICS-induced increase of miR-320d was confirmed in primary airway epithelial cells. MiR-320d negatively correlated targets were enriched for pro-inflammatory genes and were increased in the bronchial brushes of patients with lower lung function in the independent dataset. Overexpression of miR-320d in BEAS-2B cells dampened cigarette smoke extract-induced pro-inflammatory activity inhibition of nuclear factor-κB.Collectively, we identified miR-320d as a novel mediator of ICS, regulating the pro-inflammatory response of the airway epithelium.
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http://dx.doi.org/10.1183/13993003.01202-2018DOI Listing
April 2019

Limited overlap in significant hits between genome-wide association studies on two airflow obstruction definitions in the same population.

BMC Pulm Med 2019 Mar 7;19(1):58. Epub 2019 Mar 7.

Department of Epidemiology, University of Groningen, University Medical Center Groningen, Hanzeplein 1, 9700, RB, Groningen, The Netherlands.

Background: Airflow obstruction is a hallmark of chronic obstructive pulmonary disease (COPD), and is defined as either the ratio between forced expiratory volume in one second and forced vital capacity (FEV/FVC) < 70% or < lower limit of normal (LLN). This study aimed to assess the overlap between genome-wide association studies (GWAS) on airflow obstruction using these two definitions in the same population stratified by smoking.

Methods: GWASes were performed in the LifeLines Cohort Study for both airflow obstruction definitions in never-smokers (NS = 5071) and ever-smokers (ES = 4855). The FEV/FVC < 70% models were adjusted for sex, age, and height; FEV/FVC < LLN models were not adjusted. Ever-smokers models were additionally adjusted for pack-years and current-smoking. The overlap in significantly associated SNPs between the two definitions and never/ever-smokers was assessed using several p-value thresholds. To quantify the agreement, the Pearson correlation coefficient was calculated between the p-values and ORs. Replication was performed in the Vlagtwedde-Vlaardingen study (NS = 432, ES = 823). The overlapping SNPs with p < 10 were validated in the Vlagtwedde-Vlaardingen and Rotterdam Study cohorts (NS = 1966, ES = 3134) and analysed for expression quantitative trait loci (eQTL) in lung tissue (n = 1087).

Results: In the LifeLines cohort, 96% and 93% of the never- and ever-smokers were classified concordantly based on the two definitions. 26 and 29% of the investigated SNPs were overlapping at p < 0.05 in never- and ever-smokers, respectively. At p < 10 the overlap was 4% and 6% respectively, which could be change findings as shown by simulation studies. The effect estimates of the SNPs of the two definitions correlated strongly, but the p-values showed more variation and correlated only moderately. Similar observations were made in the Vlagtwedde-Vlaardingen study. Two overlapping SNPs in never-smokers (NFYC and FABP7) had the same direction of effect in the validation cohorts and the NFYC SNP was an eQTL for NFYC-AS1. NFYC is a transcription factor that binds to several known COPD genes, and FABP7 may be involved in abnormal pulmonary development.

Conclusions: The definition of airflow obstruction and the population under study may be important determinants of which SNPs are associated with airflow obstruction. The genes FABP7 and NFYC(-AS1) could play a role in airflow obstruction in never-smokers specifically.
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http://dx.doi.org/10.1186/s12890-019-0811-0DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6407273PMC
March 2019

Genetic profiling for disease stratification in chronic obstructive pulmonary disease and asthma.

Curr Opin Pulm Med 2019 05;25(3):317-322

Faculty of Science, University of Technology Sydney, Respiratory Bioinformatics and Molecular Biology, Ultimo, NSW, Australia.

Purpose Of Review: In asthma and chronic obstructive pulmonary disease (COPD), the movement towards genetic profiling with a push towards 'personalized medicine' has been hindered by complex environment--gene interactions and lack of tools to identify clear causal genetic traits. In this review, we will discuss the need for genetic profiling in asthma and COPD, what methods are currently used in the clinics and the recent finding using new sequencing methods.

Recent Findings: Over the past 10-15 years, genome-wide association studies analysis of common variants has provide little in the way of new genetic profiling markers for asthma and COPD. Whole exome/genome sequencing has provided a new method to identify lowly abundant alleles, which might have a much higher impact. Although, low population numbers due to high costs has hindered early studies, recent studies have reached genome wide significance.

Summary: The use of genetic profiling of COPD in the clinic is current limited to the identification of Alpha-1 antitrypsin deficiency, while being absent in asthma. Advances in sequencing technology provide new avenues to identify disease causes or therapy response altering variants that in the short-term will allow for the development of screening procedures for disease to identify patients at risk of developing asthma or COPD.
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http://dx.doi.org/10.1097/MCP.0000000000000568DOI Listing
May 2019

Genome-wide interaction study of gene-by-occupational exposures on respiratory symptoms.

Environ Int 2019 01 16;122:263-269. Epub 2018 Nov 16.

University of Groningen, University Medical Center Groningen, Department of Epidemiology, Groningen, the Netherlands; University of Groningen, University Medical Center Groningen, Groningen Research Institute for Asthma and COPD (GRIAC), Groningen, the Netherlands. Electronic address:

Respiratory symptoms are important indicators of respiratory diseases. Both genetic and environmental factors contribute to respiratory symptoms development but less is known about gene-environment interactions. We aimed to assess interactions between single nucleotide polymorphisms (SNPs) and occupational exposures on respiratory symptoms cough, dyspnea and phlegm. As identification cohort LifeLines I (n = 7976 subjects) was used. Job-specific exposure was estimated using the ALOHA + job exposure matrix. SNP-by-occupational exposure interactions on respiratory symptoms were tested using logistic regression adjusted for gender, age, and current smoking. SNP-by-exposure interactions with a p-value <10 were tested for replication in two independent cohorts: LifeLines II (n = 5260) and the Vlagtwedde-Vlaardingen cohort (n = 1529). The interaction estimates of the replication cohorts were meta-analyzed using PLINK. Replication was achieved when the meta-analysis p-value was <0.05 and the interaction effect had the same direction as in the identification cohort. Additionally, we assessed whether replicated SNPs associated with gene expression by analyzing if they were cis-acting expression quantitative trait loci (eQTL) in lung tissue. In the replication meta-analysis, sixteen out of 477 identified SNP-by-occupational exposure interactions had a p-value <0.05 and 9 of these interactions had the same direction as in the identification cohort. Several identified loci were plausible candidates for respiratory symptoms, such as TMPRSS9, SERPINH1, TOX3, and ARHGAP18. Three replicated SNPs were cis-eQTLs for FCER1A, CHN1, and TIMM13 in lung tissue. Taken together, this genome-wide SNP-by-occupational exposure interaction study in relation to cough, dyspnea, and phlegm identified several suggestive susceptibility genes. Further research should determine if these genes are true susceptibility loci for respiratory symptoms in relation to occupational exposures.
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http://dx.doi.org/10.1016/j.envint.2018.11.017DOI Listing
January 2019

An airway epithelial IL-17A response signature identifies a steroid-unresponsive COPD patient subgroup.

J Clin Invest 2019 01 26;129(1):169-181. Epub 2018 Nov 26.

Department of Medicine, UCSF, San Francisco, California, USA.

Background: Chronic obstructive pulmonary disease (COPD) is a heterogeneous smoking-related disease characterized by airway obstruction and inflammation. This inflammation may persist even after smoking cessation and responds variably to corticosteroids. Personalizing treatment to biologically similar "molecular phenotypes" may improve therapeutic efficacy in COPD. IL-17A is involved in neutrophilic inflammation and corticosteroid resistance, and thus may be particularly important in a COPD molecular phenotype.

Methods: We generated a gene expression signature of IL-17A response in bronchial airway epithelial brushings from smokers with and without COPD (n = 238), and validated it using data from 2 randomized trials of IL-17 blockade in psoriasis. This IL-17 signature was related to clinical and pathologic characteristics in 2 additional human studies of COPD: (a) SPIROMICS (n = 47), which included former and current smokers with COPD, and (b) GLUCOLD (n = 79), in which COPD participants were randomized to placebo or corticosteroids.

Results: The IL-17 signature was associated with an inflammatory profile characteristic of an IL-17 response, including increased airway neutrophils and macrophages. In SPIROMICS the signature was associated with increased airway obstruction and functional small airways disease on quantitative chest CT. In GLUCOLD the signature was associated with decreased response to corticosteroids, irrespective of airway eosinophilic or type 2 inflammation.

Conclusion: These data suggest that a gene signature of IL-17 airway epithelial response distinguishes a biologically, radiographically, and clinically distinct COPD subgroup that may benefit from personalized therapy.

Trial Registration: ClinicalTrials.gov NCT01969344.

Funding: Primary support from the NIH, grants K23HL123778, K12HL11999, U19AI077439, DK072517, U01HL137880, K24HL137013 and R01HL121774 and contracts HHSN268200900013C, HHSN268200900014C, HHSN268200900015C, HHSN268200900016C, HHSN268200900017C, HHSN268200900018C, HHSN268200900019C and HHSN268200900020C.
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http://dx.doi.org/10.1172/JCI121087DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6307967PMC
January 2019