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.

Current smoking contributes to worsened asthma prognosis and 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 patients with asthma were analyzed. Differential gene expression and DNA methylation analyses were conducted comparing current smokers with 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. A total of 809 genes and 18,814 CpG sites were differentially associated with current smoking in the nose. The cis-eQTM analysis uncovered 171 CpG sites with a methylation status associated with smoking-related gene expression, including AHRR, ALDH3A1, CYP1A1, and CYP1B1. The methylation status of CpG sites altered by current smoking reversed with 1 year of smoking cessation. We confirm that current smoking alters epigenetic patterns and affects gene expression in the nasal epithelium of patients with asthma, which is partially reversible in bronchial 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.

Background: Despite the well-known detrimental effects of cigarette smoke (CS), little is known about the complex gene expression dynamics in the early stages after exposure. This study aims to investigate early transcriptomic responses following CS exposure of airway epithelial cells in culture and compare these to those found in human CS exposure studies. Methods: Primary bronchial epithelial cells (PBEC) were differentiated at the air–liquid interface (ALI) and exposed to whole CS. Bulk RNA-sequencing was performed at 1 h, 4 h, and 24 h hereafter, followed by differential gene expression analysis. Results were additionally compared to data retrieved from human CS studies. Results: ALI-PBEC gene expression in response to CS was most significantly changed at 4 h after exposure. Early transcriptomic changes (1 h, 4 h post CS exposure) were related to oxidative stress, xenobiotic metabolism, higher expression of immediate early genes and pro-inflammatory pathways (i.e., Nrf2, AP-1, AhR). At 24 h, ferroptosis-associated genes were significantly increased, whereas PRKN, involved in removing dysfunctional mitochondria, was downregulated. Importantly, the transcriptome dynamics of the current study mirrored in-vivo human studies of acute CS exposure, chronic smokers, and inversely mirrored smoking cessation. Conclusion: These findings show that early after CS exposure xenobiotic metabolism and pro-inflammatory pathways were activated, followed by activation of the ferroptosis-related cell death pathway. Moreover, significant overlap between these transcriptomic responses in the in-vitro model and human in-vivo studies was found, with an early response of ciliated cells. These results provide validation for the use of ALI-PBEC cultures to study the human lung epithelial response to inhaled toxicants.

Several studies have found that systemic soluble receptor for advanced glycation end-products (sRAGE) levels are decreased in COPD patients and strongly correlate with lung function decline, emphysema and disease progression [1–6]. Therefore, the COPD Biomarker Qualification Consortium and the Biomarker Development Center recently put sRAGE forward as the most promising biomarker for COPD [7]. sRAGE is anti-inflammatory decoy receptor for the pro-inflammatory innate immune receptor RAGE, which is transcribed from the AGER gene. It was shown that one particular single nucleotide polymorphism (SNP) decreases the circulating levels of sRAGE in COPD patients [3]. This SNP, rs2070600, causes an amino acid change from glycine to serine at the 82nd amino acid of RAGE, increasing the glycation-rate of one of the two glycation sites at the ligand-binding domain and subsequently increases the ligand-binding capacity of RAGE [8]. Recently, we showed that AGER expression was lower in lung tissue from smokers compared with never-smokers, and that smoking of three cigarettes severely decreased the serum levels of sRAGE within 2 h [4]. Furthermore, we showed that smokers display increased alternative splicing of AGER into the endogenous soluble splicing form [9]. Interestingly, we also showed that unlike plasma sRAGE levels, the levels in induced sputum did not correlate with COPD status [5]. To date, no studies have been performed investigating the differences between sRAGE levels in serum versus induced sputum. Here, we measured sRAGE in serum and induced sputum of healthy individuals and investigated if these levels were affected by age, smoking or the presence of the minor allele of rs2070600. Additionally, we studied whether rs2070600 can influence the production of sRAGE by affecting splicing of AGER.

Background and Objective: Smoking disturbs the bronchial‐mucus‐barrier. This study assesses the cellular composition and gene expression shifts of the bronchial‐mucus‐barrier with smoking to understand the mechanism of mucosal damage by cigarette smoke exposure. We explore whether single‐cell‐RNA‐sequencing (scRNA‐seq) based cellular deconvolution (CD) can predict cell‐type composition in RNA‐seq data.Methods: RNA‐seq data of bronchial biopsies from three cohorts were analysed using CD. The cohorts included 56 participants with chronic obstructive pulmonary disease [COPD] (38 smokers; 18 ex‐smokers), 77 participants without COPD (40 never‐smokers; 37 smokers) and 16 participants who stopped smoking for 1 year (11 COPD and 5 non‐COPD‐smokers). Differential gene expression was used to investigate gene expression shifts. The CD‐derived goblet cell ratios were validated by correlating with staining‐derived goblet cell ratios from the COPD cohort. Statistics were done in the R software (false discovery rate p‐value < 0.05).Results: Both CD methods indicate a shift in bronchial‐mucus‐barrier cell composition towards goblet cells in COPD and non‐COPD‐smokers compared to ex‐ and never‐smokers. It shows that the effect was reversible within a year of smoking cessation. A reduction of ciliated and basal cells was observed with current smoking, which resolved following smoking cessation. The expression of mucin and sodium channel (ENaC) genes, but not chloride channel genes, were altered in COPD and current smokers compared to never smokers or ex‐smokers. The goblet cell‐derived staining scores correlate with CD‐derived goblet cell ratios.Conclusion: Smoking alters bronchial‐mucus‐barrier cell composition, transcriptome and increases mucus production. This effect is partly reversible within a year of smoking cessation. CD methodology can predict goblet‐cell percentages from RNA‐seq.