At a GlanceScientific Knowledge on the Subject Emphysema and COPD are likely underpinned by aberrant cell death that leads to airway inflammation and remodelling and emphysema. Apoptotic cell death can be programmed and controlled during apoptosis, whereas necroptosis defined by ruptured cell membranes is considered to be non-inflammatory, whereas necroptosis, a form of regulated necrosis, is highly pro-inflammatory. The roles of these pathways in COPD are poorly understood. What the Study Adds to the FieldWe used combination analysis of human COPD lung tissue, mouse models of experimental COPD, mice deficient in key necroptotic pathway mediators (RIPK3, MLKL) and inhibitors to define the roles of cell death pathways. Necroptotsis signalling is increased in the lungs in human and experimental COPD and correlate with disease severity. Genetic inhibition of necroptosis suppresses airway inflammation and remodelling and emphysema in experimental COPD, while pharmacological caspase inhibition reduces inflammation only.Inhibiting necroptosis may be a new therapeutic approach for COPD.

Angiotensin-converting enzyme 2 (ACE2) has been identified as the cell entry receptor used by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) [1, 2]. Importantly, smokers and patients with COPD are at an increased risk of severe complications and a higher mortality upon SARS-CoV-2 infection [3]. We hypothesised that ACE2 expression is increased in lungs of smokers and patients with COPD, which may at least partially explain their higher risk of a more severe course of coronavirus disease 2019 (COVID-19). Therefore, we aimed to investigate the expression of ACE2 on both mRNA and protein level in a large number of lung tissue specimens of well-phenotyped subjects, including never-smokers, current smokers without airflow limitation, and patients with COPD. In this cross-sectional observational study, we analysed lung tissue specimens from 134 subjects from our large lung tissue biobank at Ghent University Hospital (Ghent, Belgium) and from explant lungs from end-stage COPD patients collected at UZ Gasthuisberg Leuven (Leuven, Belgium). Ex-smoking was defined as smoking cessation for ⩾1 year. COPD severity was defined according to the Global Initiative for Chronic Obstructive Lung Disease (GOLD) classification. Written informed consent was obtained from all subjects, and the study was approved by the medical ethical committees of Ghent University Hospital (2016/0132; 2019/0537) and the University Hospital Gasthuisberg Leuven (S51577). RNA extraction from lung tissue blocks of 120 subjects was performed with the miRNeasy Mini kit (Qiagen, Hilden, Germany). Next, cDNA was prepared with the EvoScript Universal cDNA Master Kit (Roche, Basel, Switzerland), followed by reverse transcriptase (RT)-qPCR analysis for ACE2 and 3 reference genes, as described previously [4, 5]. Sections from formalin-fixed paraffin-embedded lung tissue blocks of 87 subjects were stained for ACE2. After antigen retrieval with citrate buffer (Scytek, West Logan, UT, USA), the slides were incubated with anti-ACE2 antibody (polyclonal rabbit-anti-human, Abcam ab15248). Next, slides were coloured with diaminobenzidine (Dako, Carpinteria, CA, USA) and counterstained with Mayer's haematoxylin (Sigma-Aldrich, St Louis, MO, USA). Quantitative measurements of the ACE2-positive signal in alveolar tissue and bronchial epithelium were performed on images of stained paraffin sections as described previously [6]. Statistical analysis was performed using Sigma Stat software (SPSS 26.0, Chicago, IL, USA) and R3.5.1, using Kruskal-Wallis tests (on all six groups) followed by Mann-Whitney U-tests (for the comparison between two groups), and multivariable linear regression analyses. Using RT-PCR, ACE2 mRNA levels were determined in lung tissue from 120 subjects. ACE2 mRNA expression was significantly higher in the lung tissue of current smokers without airflow limitation and current smokers with COPD (GOLD stages II and III-IV) compared with never-smokers (figure 1a). In addition, ex-smokers without airflow limitation showed significantly lower ACE2 mR...

Increased expression of pulmonary ACE2, the SARS-CoV-2 receptor, could contribute to increased infectivity of COVID-19 in subjects with diabetes, but ACE2 expression has not been studied in lung tissue of subjects with diabetes. We therefore studied ACE2 mRNA and protein expression in lung tissue samples of patients with and without diabetes that were collected between 2002 and 2020 from patients undergoing lobectomy for lung tumors. For RT-PCR analyses, samples from 15 subjects with diabetes were compared to 91 randomly chosen control samples. For immunohistochemical staining, samples from 26 subjects with diabetes were compared to 66 randomly chosen control samples. mRNA expression of ACE2 was measured by quantitative RT-PCR. Protein levels of ACE2 were visualized by immunohistochemistry on paraffin-embedded lung tissue samples and quantified in alveolar and bronchial epithelium. Pulmonary ACE2 mRNA expression was not different between subjects with or without diabetes. In contrast, protein levels of ACE2 were significantly increased in both alveolar tissue and bronchial epithelium of patients with diabetes as compared with control subjects, independent of smoking, COPD, BMI, RAAS-inhibitor use and other potential confounders. To conclude, we show increased bronchial and alveolar ACE2 protein expression in patients with diabetes. Further research is needed to elucidate whether upregulation of ACE2 expression in airways and lungs has consequences on infectivity and clinical outcomes of COVID-19.

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Extravasation of monocytes into tissue and to the site of injury is a fundamental immunological process, which requires rapid responses via post translational modifications (PTM) of proteins. Protein arginine methyltransferase 7 (PRMT7) is an epigenetic factor that has the capacity to mono-methylate histones on arginine residues. Here we show that in chronic obstructive pulmonary disease (COPD) patients, PRMT7 expression is elevated in the lung tissue and localized to the macrophages. In mouse models of COPD, lung fibrosis and skin injury, reduced expression of PRMT7 associates with decreased recruitment of monocytes to the site of injury and hence less severe symptoms. Mechanistically, activation of NF-κB/RelA in monocytes induces PRMT7 transcription and consequential mono-methylation of histones at the regulatory elements of RAP1A, which leads to increased transcription of this gene that is responsible for adhesion and migration of monocytes. Persistent monocyte-derived macrophage accumulation leads to ALOX5 over-expression and accumulation of its metabolite LTB4, which triggers expression of ACSL4 a ferroptosis promoting gene in lung epithelial cells. Conclusively, inhibition of arginine mono-methylation might offer targeted intervention in monocyte-driven inflammatory conditions that lead to extensive tissue damage if left untreated.