COVID-19 has rapidly spread through the world, and is caused by SARS-CoV-2. Patients with chronic obstructive pulmonary disease (COPD) face an increased risk for severe illness from COVID-19 because of angiotensin-converting enzyme 2 (ACE2) upregulation, an entry receptor for SARS-CoV-2. Our group recently revealed that COPD-derived lung fibroblasts express higher level of ACE2, and provide direct evidence that chronic cigarette smoke (CS) exposure significantly increases pulmonary ACE2 protein. CS is the primary risk factor for COPD. ACE2 expression may be regulated at the transcriptional, post-transcriptional and translational levels. Changes in protein expression can be controlled by RNA-binding proteins (RBPs). One of the best studied RBPs is human antigen R (HuR), a ubiquitously expressed RBP, that regulates the stability, localization and/or translation of target mRNAs. To stabilize target mRNA, HuR translocates from the nucleus to the cytoplasm. CS may increases HuR translocation to the cytoplasm, thereby indirectly augmenting cellular protein levels. Therefore, we hypothesized that HuR regulates ACE2 expression in response to CS, thereby accounting for higher ACE2 in COPD-derived cells. Aims: 1)- Evaluate HuR expression and localization in Normal, Smoker and COPD lung tissues;2) Establish that HuR induces ACE2 expression in primary human lung fibroblasts (HLF);3)- Determine if HuR controls ACE2 expression by regulating mRNA stability. Methods: 1)- Multiplex Immunohistochemistry was performed on lung tissue from Normal, Smoker and COPD subjects. The localization of HuR was assessed in HLF exposed to 2% cigarette smoke extract (CSE) for 4h by subcellular fractionation-western blot. 2)- Normal, Smoker and COPD HLF were transfected with either small interfering RNA (siRNA) against HuR or Control siRNA. ACE2 protein was evaluated by western blot. 3)- RNA-binding protein immunoprecipitation-qPCR was used to assess the association of HuR to ACE2 mRNA in Normal, Smoker and COPD HLF. The effect of HuR on the stability of ACE2 mRNA was assessed by using actinomycin D pulse-chase followed by qPCR. Results: 1)- HuR cytoplasmic localization is higher in Smoker and COPD lung tissues. There was an increase in cytoplasmic HuR in HLF exposed to 2%CSE for 4h. 2)- Knockdown of HuR slightly increases ACE2 protein. 3)- HuR associates with ACE2 mRNA in Normal and Smoker HLF. Significance: Our work is the first to highlight the association between ACE2 and HuR in COPD. The regulation of ACE2 by HuR could provide the basis to understand the upregulation of ACE2 expression in COPD HLF and the potential consequence towards COVID-19.

BACKGROUND:Bronchial vascular remodelling may contribute to the severity of airway narrowing through mucosal congestion. Interleukin (IL)-17A is associated with the most severe asthmatic phenotype but whether it might contribute to vascular remodelling is uncertain.OBJECTIVE:To assess vascular remodelling in severe asthma and whether IL-17A directly or indirectly may cause endothelial cell activation and angiogenesis.METHODS:Bronchial vascularization was quantified in asthmatic subjects, COPD and healthy subjects together with the number of IL-17A+ cells as well as the concentration of angiogenic factors in the sputum. The effect of IL-17A on in vitro angiogenesis, cell migration and endothelial permeability was assessed directly on primary human lung microvascular endothelial cells (HMVEC-L) or indirectly with conditioned medium derived from normal bronchial epithelial cells (NHBEC), fibroblasts (NHBF) and airway smooth muscle cells (ASMC) after IL-17A stimulation.RESULTS:Severe asthmatics have increased vascularity compared to the other groups, which correlates positively with the concentrations of angiogenic factors in sputum. Interestingly, we demonstrated that increased bronchial vascularity correlates positively with the number of subepithelial IL-17A+ cells. However IL-17A had no direct effect on HMVEC-L function but it enhanced endothelial tube formation and cell migration through the production of angiogenic factors by NHBE and ASMC.CONCLUSIONS & CLINICAL RELEVANCE:Our results shed light on the role of IL-17A in vascular remodelling, most likely through stimulating the synthesis of other angiogenic factors. Knowledge of these pathways may aid in the identification of new therapeutic targets.

Summary Background Chronic inflammation, typified by increased expression of IL ‐17A, together with airway and parenchymal remodelling are features of chronic lung diseases. Emerging evidence suggests that phenotypic heterogeneity of repair and inflammatory capacities of fibroblasts may contribute to the differential structural changes observed in different regions of the lung. Objective To investigate phenotypic differences in parenchymal and bronchial fibroblasts, either in terms of inflammation and remodelling or the ability of these fibroblasts to respond to IL ‐17A. Methods Four groups of primary fibroblasts were used: normal human bronchial fibroblast ( NHBF ), normal human parenchymal fibroblast ( NHPF ), COPD human bronchial fibroblast ( CHBF ) and COPD human parenchymal fibroblast ( CHPF ). Cytokine and extracellular matrix ( ECM ) expression were measured at baseline and after stimulation with IL ‐17A. Actinomycin D was used to measure cytokine mRNA stability. Results At baseline, we observed higher protein production of IL ‐6 in NHPF than NHBF , but higher levels of IL ‐8 and GRO ‐α in NHBF . IL ‐17A induced a higher expression of GRO ‐α ( CXCL 1) and IL ‐6 in NHPF than in NHBF , and a higher level of IL ‐8 expression in NHBF . IL ‐17A treatment decreased the mRNA stability of IL ‐6 in NHBF when compared with NHPF . CHPF expressed higher protein levels of fibronectin, collagen‐I and collagen‐ III than CHBF , NHBF and NHPF . IL ‐17A increased fibronectin and collagen‐ III protein only in NHPF and collagen‐ III protein production in CHBF and CHPF . Conclusions and Clinical Relevance These findings provide insight into the inflammatory and remodelling processes that may be related to the phenotypic heterogeneity of fibroblasts from airway and parenchymal regions and in their response to IL ‐17A.