4-Octyl Itaconate Alleviates Airway Eosinophilic Inflammation by Suppressing Chemokines and Eosinophil Development.

4-Octyl itaconate (4-OI) is a derivative of the Krebs cycle-derived metabolite itaconate and displays an array of antimicrobial and anti-inflammatory properties through modifying cysteine residues within protein targets. We have found that 4-OI significantly reduces the production of eosinophil-targeted chemokines in a variety of cell types, including M1 and M2 macrophages, Th2 cells, and A549 respiratory epithelial cells. Notably, the suppression of these chemokines in M1 macrophages was found to be NRF2-dependent. In addition, 4-OI can interfere with IL-5 signaling and directly affect eosinophil differentiation. In a model of eosinophilic airway inflammation in BALB/c mice, 4-OI alleviated airway resistance and reduced eosinophil recruitment to the lungs. Our findings suggest that itaconate derivatives could be promising therapeutic agents for the treatment of eosinophilic asthma.

Landscape fire smoke airway exposure impairs respiratory and cardiac function and worsens experimental asthma.

BACKGROUND: Millions of people are exposed to landscape fire smoke (LFS) globally, and inhalation of LFS particulate matter (PM) is associated with poor respiratory and cardiovascular outcomes. However, how LFS affects respiratory and cardiovascular function is less well understood. OBJECTIVE: We aimed to characterize the pathophysiologic effects of representative LFS airway exposure on respiratory and cardiac function and on asthma outcomes. METHODS: LFS was generated using a customized combustion chamber. In 8-week-old female BALB/c mice, low (25 µg/m3, 24-hour equivalent) or moderate (100 µg/m3, 24-hour equivalent) concentrations of LFS PM (10 µm and below [PM10]) were administered daily for 3 (short-term) and 14 (long-term) days in the presence and absence of experimental asthma. Lung inflammation, gene expression, structural changes, and lung function were assessed. In 8-week-old male C57BL/6 mice, low concentrations of LFS PM10 were administered for 3 days. Cardiac function and gene expression were assessed. RESULTS: Short- and long-term LFS PM10 airway exposure increased airway hyperresponsiveness and induced steroid insensitivity in experimental asthma, independent of significant changes in airway inflammation. Long-term LFS PM10 airway exposure also decreased gas diffusion. Short-term LFS PM10 airway exposure decreased cardiac function and expression of gene changes relating to oxidative stress and cardiovascular pathologies. CONCLUSIONS: We characterized significant detrimental effects of physiologically relevant concentrations and durations of LFS PM10 airway exposure on lung and heart function. Our study provides a platform for assessment of mechanisms that underpin LFS PM10 airway exposure on respiratory and cardiovascular disease outcomes.

Smoking and tetramer tryptase accelerate intervertebral disc degeneration by inducing METTL14-mediated DIXDC1 m6 modification.

Although cigarette smoking (CS) and low back pain (LBP) are common worldwide, their correlations and the mechanisms of action remain unclear. We have shown that excessive activation of mast cells (MCs) and their proteases play key roles in CS-associated diseases, like asthma, chronic obstructive pulmonary disease (COPD), blood coagulation, and lung cancer. Previous studies have also shown that MCs and their proteases induce degenerative musculoskeletal disease. By using a custom-designed smoke-exposure mouse system, we demonstrated that CS results in intervertebral disc (IVD) degeneration and release of MC-restricted tetramer tryptases (TTs) in the IVDs. TTs were found to regulate the expression of methyltransferase 14 (METTL14) at the epigenetic level by inducing N6-methyladenosine (m6A) deposition in the 3' untranslated region (UTR) of the transcript that encodes dishevelled-axin (DIX) domain-containing 1 (DIXDC1). That reaction increases the mRNA stability and expression of Dixdc1. DIXDC1 functionally interacts with disrupted in schizophrenia 1 (DISC1) to accelerate the degeneration and senescence of nucleus pulposus (NP) cells by activating a canonical Wnt pathway. Our study demonstrates the association between CS, MC-derived TTs, and LBP. These findings raise the possibility that METTL14-medicated DIXDC1 m6A modification could serve as a potential therapeutic target to block the development of degeneration of the NP in LBP patients.

RIPK1 kinase-dependent inflammation and cell death contribute to the pathogenesis of COPD.

BACKGROUND: Receptor-interacting protein kinase 1 (RIPK1) is a key mediator of regulated cell death (including apoptosis and necroptosis) and inflammation, both drivers of COPD pathogenesis. We aimed to define the contribution of RIPK1 kinase-dependent cell death and inflammation in the pathogenesis of COPD. METHODS: We assessed RIPK1 expression in single-cell RNA sequencing (RNA-seq) data from human and mouse lungs, and validated RIPK1 levels in lung tissue of COPD patients via immunohistochemistry. Next, we assessed the consequences of genetic and pharmacological inhibition of RIPK1 kinase activity in experimental COPD, using Ripk1 S25D/S25D kinase-deficient mice and the RIPK1 kinase inhibitor GSK'547. RESULTS: RIPK1 expression increased in alveolar type 1 (AT1), AT2, ciliated and neuroendocrine cells in human COPD. RIPK1 protein levels were significantly increased in airway epithelium of COPD patients compared with never-smokers and smokers without airflow limitation. In mice, exposure to cigarette smoke (CS) increased Ripk1 expression similarly in AT2 cells, and further in alveolar macrophages and T-cells. Genetic and/or pharmacological inhibition of RIPK1 kinase activity significantly attenuated airway inflammation upon acute and subacute CS exposure, as well as airway remodelling, emphysema, and apoptotic and necroptotic cell death upon chronic CS exposure. Similarly, pharmacological RIPK1 kinase inhibition significantly attenuated elastase-induced emphysema and lung function decline. Finally, RNA-seq on lung tissue of CS-exposed mice revealed downregulation of cell death and inflammatory pathways upon pharmacological RIPK1 kinase inhibition. CONCLUSIONS: RIPK1 kinase inhibition is protective in experimental models of COPD and may represent a novel promising therapeutic approach.

Airway and parenchyma transcriptomics in a house dust mite model of experimental asthma.

Lung transcriptomics studies in asthma have provided valuable information in the whole lung context, however, deciphering the individual contributions of the airway and parenchyma in disease pathogenesis may expedite the development of novel targeted treatment strategies. In this study, we performed transcriptomics on the airway and parenchyma using a house dust mite (HDM)-induced model of experimental asthma that replicates key features of the human disease. HDM exposure increased the expression of 3,255 genes, of which 212 were uniquely increased in the airways, 856 uniquely increased in the parenchyma, and 2187 commonly increased in both compartments. Further interrogation of these genes using a combination of network and transcription factor enrichment analyses identified several transcription factors that regulate airway and/or parenchymal gene expression, including transcription factor EC (TFEC), transcription factor PU.1 (SPI1), H2.0-like homeobox (HLX), metal response element binding transcription factor-1 (MTF1) and E74-like factor 4 (ets domain transcription factor, ELF4) involved in controlling innate immune responses. We next assessed the effects of inhibiting lung SPI1 responses using commercially available DB1976 and DB2313 on key disease outcomes. We found that both compounds had no protective effects on airway inflammation, however DB2313 (8 mg/kg) decreased mucus secreting cell number, and both DB2313 (1 mg/kg) and DB1976 (2.5 mg/kg and 1 mg/kg) reduced small airway collagen deposition. Significantly, both compounds decreased airway hyperresponsiveness. This study demonstrates that SPI1 is important in HDM-induced experimental asthma and that its pharmacological inhibition reduces HDM-induced airway collagen deposition and hyperresponsiveness.

Characterization and inhibition of inflammasome responses in severe and non-severe asthma.

BACKGROUND: Increased airway NLRP3 inflammasome-mediated IL-1ß responses may underpin severe neutrophilic asthma. However, whether increased inflammasome activation is unique to severe asthma, is a common feature of immune cells in all inflammatory types of severe asthma, and whether inflammasome activation can be therapeutically targeted in patients, remains unknown. OBJECTIVE: To investigate the activation and inhibition of inflammasome-mediated IL-1ß responses in immune cells from patients with asthma. METHODS: Peripheral blood mononuclear cells (PBMCs) were isolated from patients with non-severe (n = 59) and severe (n = 36 stable, n = 17 exacerbating) asthma and healthy subjects (n = 39). PBMCs were stimulated with nigericin or lipopolysaccharide (LPS) alone, or in combination (LPS + nigericin), with or without the NLRP3 inhibitor MCC950, and the effects on IL-1ß release were assessed. RESULTS: PBMCs from patients with non-severe or severe asthma produced more IL-1ß in response to nigericin than those from healthy subjects. PBMCs from patients with severe asthma released more IL-1ß in response to LPS + nigericin than those from non-severe asthma. Inflammasome-induced IL-1ß release from PBMCs from patients with severe asthma was not increased during exacerbation compared to when stable. Inflammasome-induced IL-1ß release was not different between male and female, or obese and non-obese patients and correlated with eosinophil and neutrophil numbers in the airways. MCC950 effectively suppressed LPS-, nigericin-, and LPS + nigericin-induced IL-1ß release from PBMCs from all groups. CONCLUSION: An increased ability for inflammasome priming and/or activation is a common feature of systemic immune cells in both severe and non-severe asthma, highlighting inflammasome inhibition as a universal therapy for different subtypes of disease.

Airway and parenchymal transcriptomics in a novel model of asthma and COPD overlap.

BACKGROUND: Asthma and chronic obstructive pulmonary disease (COPD) are common chronic respiratory diseases, and some patients have overlapping disease features, termed asthma-COPD overlap (ACO). Patients characterized with ACO have increased disease severity; however, the mechanisms driving this have not been widely studied. OBJECTIVES: This study sought to characterize the phenotypic and transcriptomic features of experimental ACO in mice induced by chronic house dust mite antigen and cigarette smoke exposure. METHODS: Female BALB/c mice were chronically exposed to house dust mite antigen for 11 weeks to induce experimental asthma, cigarette smoke for 8 weeks to induce experimental COPD, or both concurrently to induce experimental ACO. Lung inflammation, structural changes, and lung function were assessed. RNA-sequencing was performed on separated airway and parenchyma lung tissues to assess transcriptional changes. Validation of a novel upstream driver SPI1 in experimental ACO was assessed using the pharmacological SPI1 inhibitor, DB2313. RESULTS: Experimental ACO recapitulated features of both asthma and COPD, with mixed pulmonary eosinophilic/neutrophilic inflammation, small airway collagen deposition, and increased airway hyperresponsiveness. Transcriptomic analysis identified common and distinct dysregulated gene clusters in airway and parenchyma samples in experimental asthma, COPD, and ACO. Upstream driver analysis revealed increased expression of the transcription factor Spi1. Pharmacological inhibition of SPI1 using DB2313, reduced airway remodeling and airway hyperresponsiveness in experimental ACO. CONCLUSIONS: A new experimental model of ACO featuring chronic dual exposures to house dust mite and cigarette smoke mimics key disease features observed in patients with ACO and revealed novel disease mechanisms, including upregulation of SPI1, that are amenable to therapy.

Relationship between type 2 cytokine and inflammasome responses in obesity-associated asthma.

BACKGROUND: Obesity is a risk factor for asthma, and obese asthmatic individuals are more likely to have severe, steroid-insensitive disease. How obesity affects the pathogenesis and severity of asthma is poorly understood. Roles for increased inflammasome-mediated neutrophilic responses, type 2 immunity, and eosinophilic inflammation have been described. OBJECTIVE: We investigated how obesity affects the pathogenesis and severity of asthma and identified effective therapies for obesity-associated disease. METHODS: We assessed associations between body mass index and inflammasome responses with type 2 (T2) immune responses in the sputum of 25 subjects with asthma. Functional roles for NLR family, pyrin domain-containing (NLRP) 3 inflammasome and T2 cytokine responses in driving key features of disease were examined in experimental high-fat diet-induced obesity and asthma. RESULTS: Body mass index and inflammasome responses positively correlated with increased IL-5 and IL-13 expression as well as C-C chemokine receptor type 3 expression in the sputum of subjects with asthma. High-fat diet-induced obesity resulted in steroid-insensitive airway hyperresponsiveness in both the presence and absence of experimental asthma. High-fat diet-induced obesity was also associated with increased NLRP3 inflammasome responses and eosinophilic inflammation in airway tissue, but not lumen, in experimental asthma. Inhibition of NLRP3 inflammasome responses reduced steroid-insensitive airway hyperresponsiveness but had no effect on IL-5 or IL-13 responses in experimental asthma. Depletion of IL-5 and IL-13 reduced obesity-induced NLRP3 inflammasome responses and steroid-insensitive airway hyperresponsiveness in experimental asthma. CONCLUSION: We found a relationship between T2 cytokine and NLRP3 inflammasome responses in obesity-associated asthma, highlighting the potential utility of T2 cytokine-targeted biologics and inflammasome inhibitors.

Aim2 suppresses cigarette smoke-induced neutrophil recruitment, neutrophil caspase-1 activation and anti-Ly6G-mediated neutrophil depletion.

Increased inflammasome responses are strongly implicated in inflammatory diseases; however, their specific roles are incompletely understood. Therefore, we sought to examine the roles of nucleotide-binding oligomerization domain-like receptor (NLR) family, pyrin domain-containing 3 (NLRP3) and absent in melanoma-2 (AIM2) inflammasomes in cigarette smoke-induced inflammation in a model of experimental chronic obstructive pulmonary disease (COPD). We targeted NLRP3 with the inhibitor MCC950 given prophylactically or therapeutically and examined Aim2-/- mice in cigarette smoke-induced experimental COPD. MCC950 treatment had minimal effects on disease development and/or progression. Aim2-/- mice had increased airway neutrophils with decreased caspase-1 levels, independent of changes in lung neutrophil chemokines. Suppressing neutrophils with anti-Ly6G in experimental COPD in wild-type mice reduced neutrophils in bone marrow, blood and lung. By contrast, anti-Ly6G treatment in Aim2-/- mice with experimental COPD had no effect on neutrophils in bone marrow, partially reduced neutrophils in the blood and had no effect on neutrophils or neutrophil caspase-1 levels in the lungs. These findings identify that following cigarette smoke exposure, Aim2 is important for anti-Ly6G-mediated depletion of neutrophils, suppression of neutrophil recruitment and mediates activation of caspase-1 in neutrophils.

Human ß-defensin-2 suppresses key features of asthma in murine models of allergic airways disease.

BACKGROUND: Asthma is an airway inflammatory disease and a major health problem worldwide. Anti-inflammatory steroids and bronchodilators are the gold-standard therapy for asthma. However, they do not prevent the development of the disease, and critically, a subset of asthmatics are resistant to steroid therapy. OBJECTIVE: To elucidate the therapeutic potential of human ß-defensins (hBD), such as hBD2 mild to moderate and severe asthma. METHODS: We investigated the role of hBD2 in a steroid-sensitive, house dust mite-induced allergic airways disease (AAD) model and a steroid-insensitive model combining ovalbumin-induced AAD with C muridarum (Cmu) respiratory infection. RESULTS: In both models, we demonstrated that therapeutic intranasal application of hBD2 significantly reduced the influx of inflammatory cells into the bronchoalveolar lavage fluid. Furthermore, key type 2 asthma-related cytokines IL-9 and IL-13, as well as additional immunomodulating cytokines, were significantly decreased after administration of hBD2 in the steroid-sensitive model. The suppression of inflammation was associated with improvements in airway physiology and treatment also suppressed airway hyper-responsiveness (AHR) in terms of airway resistance and compliance to methacholine challenge. CONCLUSIONS AND CLINICAL RELEVANCE: These data indicate that hBD2 reduces the hallmark features and has potential as a new therapeutic agent in allergic and especially steroid-resistant asthma.

Critical role for iron accumulation in the pathogenesis of fibrotic lung disease.

Increased iron levels and dysregulated iron homeostasis, or both, occur in several lung diseases. Here, the effects of iron accumulation on the pathogenesis of pulmonary fibrosis and associated lung function decline was investigated using a combination of murine models of iron overload and bleomycin-induced pulmonary fibrosis, primary human lung fibroblasts treated with iron, and histological samples from patients with or without idiopathic pulmonary fibrosis (IPF). Iron levels are significantly increased in iron overloaded transferrin receptor 2 (Tfr2) mutant mice and homeostatic iron regulator (Hfe) gene-deficient mice and this is associated with increases in airway fibrosis and reduced lung function. Furthermore, fibrosis and lung function decline are associated with pulmonary iron accumulation in bleomycin-induced pulmonary fibrosis. In addition, we show that iron accumulation is increased in lung sections from patients with IPF and that human lung fibroblasts show greater proliferation and cytokine and extracellular matrix responses when exposed to increased iron levels. Significantly, we show that intranasal treatment with the iron chelator, deferoxamine (DFO), from the time when pulmonary iron levels accumulate, prevents airway fibrosis and decline in lung function in experimental pulmonary fibrosis. Pulmonary fibrosis is associated with an increase in Tfr1+ macrophages that display altered phenotype in disease, and DFO treatment modified the abundance of these cells. These experimental and clinical data demonstrate that increased accumulation of pulmonary iron plays a key role in the pathogenesis of pulmonary fibrosis and lung function decline. Furthermore, these data highlight the potential for the therapeutic targeting of increased pulmonary iron in the treatment of fibrotic lung diseases such as IPF. © 2020 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

Mechanisms and treatments for severe, steroid-resistant allergic airway disease and asthma.

Severe, steroid-resistant asthma is clinically and economically important since affected individuals do not respond to mainstay corticosteroid treatments for asthma. Patients with this disease experience more frequent exacerbations of asthma, are more likely to be hospitalized, and have a poorer quality of life. Effective therapies are urgently required, however, their development has been hampered by a lack of understanding of the pathological processes that underpin disease. A major obstacle to understanding the processes that drive severe, steroid-resistant asthma is that the several endotypes of the disease have been described that are characterized by different inflammatory and immunological phenotypes. This heterogeneity makes pinpointing processes that drive disease difficult in humans. Clinical studies strongly associate specific respiratory infections with severe, steroid-resistant asthma. In this review, we discuss key findings from our studies where we describe the development of representative experimental models to improve our understanding of the links between infection and severe, steroid-resistant forms of this disease. We also discuss their use in elucidating the mechanisms, and their potential for developing effective therapeutic strategies, for severe, steroid-resistant asthma. Finally, we highlight how the immune mechanisms and therapeutic targets we have identified may be applicable to obesity-or pollution-associated asthma.

Crucial role for lung iron level and regulation in the pathogenesis and severity of asthma.

Accumulating evidence highlights links between iron regulation and respiratory disease. Here, we assessed the relationship between iron levels and regulatory responses in clinical and experimental asthma.We show that cell-free iron levels are reduced in the bronchoalveolar lavage (BAL) supernatant of severe or mild-moderate asthma patients and correlate with lower forced expiratory volume in 1 s (FEV1). Conversely, iron-loaded cell numbers were increased in BAL in these patients and with lower FEV1/forced vital capacity (FVC) ratio. The airway tissue expression of the iron sequestration molecules divalent metal transporter 1 (DMT1) and transferrin receptor 1 (TFR1) are increased in asthma, with TFR1 expression correlating with reduced lung function and increased Type-2 (T2) inflammatory responses in the airways. Furthermore, pulmonary iron levels are increased in a house dust mite (HDM)-induced model of experimental asthma in association with augmented Tfr1 expression in airway tissue, similar to human disease. We show that macrophages are the predominant source of increased Tfr1 and Tfr1+ macrophages have increased Il13 expression. We also show that increased iron levels induce increased pro-inflammatory cytokine and/or extracellular matrix (ECM) responses in human airway smooth muscle (ASM) cells and fibroblasts ex vivo and induce key features of asthma in vivo, including airway hyper-responsiveness (AHR) and fibrosis, and T2 inflammatory responses.Together these complementary clinical and experimental data highlight the importance of altered pulmonary iron levels and regulation in asthma, and the need for a greater focus on the role and potential therapeutic targeting of iron in the pathogenesis and severity of disease.

IL-22 and its receptors are increased in human and experimental COPD and contribute to pathogenesis.

Chronic obstructive pulmonary disease (COPD) is the third leading cause of morbidity and death globally. The lack of effective treatments results from an incomplete understanding of the underlying mechanisms driving COPD pathogenesis.Interleukin (IL)-22 has been implicated in airway inflammation and is increased in COPD patients. However, its roles in the pathogenesis of COPD is poorly understood. Here, we investigated the role of IL-22 in human COPD and in cigarette smoke (CS)-induced experimental COPD.IL-22 and IL-22 receptor mRNA expression and protein levels were increased in COPD patients compared to healthy smoking or non-smoking controls. IL-22 and IL-22 receptor levels were increased in the lungs of mice with experimental COPD compared to controls and the cellular source of IL-22 included CD4+ T-helper cells, γδ T-cells, natural killer T-cells and group 3 innate lymphoid cells. CS-induced pulmonary neutrophils were reduced in IL-22-deficient (Il22 -/-) mice. CS-induced airway remodelling and emphysema-like alveolar enlargement did not occur in Il22 -/- mice. Il22 -/- mice had improved lung function in terms of airway resistance, total lung capacity, inspiratory capacity, forced vital capacity and compliance.These data highlight important roles for IL-22 and its receptors in human COPD and CS-induced experimental COPD.

IL-6 Drives Neutrophil-Mediated Pulmonary Inflammation Associated with Bacteremia in Murine Models of Colitis.

Inflammatory bowel disease (IBD) is associated with several immune-mediated extraintestinal manifestations. More than half of all IBD patients have some form of respiratory pathology, most commonly neutrophil-mediated diseases, such as bronchiectasis and chronic bronchitis. Using murine models of colitis, we aimed to identify the immune mechanisms driving pulmonary manifestations of IBD. We found increased neutrophil numbers in lung tissue associated with the pulmonary vasculature in both trinitrobenzenesulfonic acid- and dextran sulfate sodium-induced models of colitis. Analysis of systemic inflammation identified that neutrophilia was associated with bacteremia and pyrexia in animal models of colitis. We further identified IL-6 as a systemic mediator of neutrophil recruitment from the bone marrow of dextran sulfate sodium animals. Functional inhibition of IL-6 led to reduced systemic and pulmonary neutrophilia, but it did not attenuate established colitis pathology. These data suggest that systemic bacteremia and pyrexia drive IL-6 secretion, which is a critical driver for pulmonary manifestation of IBD. Targeting IL-6 may reduce neutrophil-associated extraintestinal manifestations in IBD patients.

Toll-like receptor 2 and 4 have opposing roles in the pathogenesis of cigarette smoke-induced chronic obstructive pulmonary disease.

Chronic obstructive pulmonary disease (COPD) is the third leading cause of morbidity and death and imposes major socioeconomic burdens globally. It is a progressive and disabling condition that severely impairs breathing and lung function. There is a lack of effective treatments for COPD, which is a direct consequence of the poor understanding of the underlying mechanisms involved in driving the pathogenesis of the disease. Toll-like receptor (TLR)2 and TLR4 are implicated in chronic respiratory diseases, including COPD, asthma and pulmonary fibrosis. However, their roles in the pathogenesis of COPD are controversial and conflicting evidence exists. In the current study, we investigated the role of TLR2 and TLR4 using a model of cigarette smoke (CS)-induced experimental COPD that recapitulates the hallmark features of human disease. TLR2, TLR4, and associated coreceptor mRNA expression was increased in the airways in both experimental and human COPD. Compared with wild-type (WT) mice, CS-induced pulmonary inflammation was unaltered in TLR2-deficient ( Tlr2-/-) and TLR4-deficient ( Tlr4-/-) mice. CS-induced airway fibrosis, characterized by increased collagen deposition around small airways, was not altered in Tlr2-/- mice but was attenuated in Tlr4-/- mice compared with CS-exposed WT controls. However, Tlr2-/- mice had increased CS-induced emphysema-like alveolar enlargement, apoptosis, and impaired lung function, while these features were reduced in Tlr4-/- mice compared with CS-exposed WT controls. Taken together, these data highlight the complex roles of TLRs in the pathogenesis of COPD and suggest that activation of TLR2 and/or inhibition of TLR4 may be novel therapeutic strategies for the treatment of COPD.

MicroRNA Profiling Reveals a Role for MicroRNA-218-5p in the Pathogenesis of Chronic Obstructive Pulmonary Disease.

RATIONALE: Aberrant expression of microRNAs (miRNAs) can have a detrimental role in disease pathogenesis. OBJECTIVES: To identify dysregulated miRNAs in lung tissue of patients with chronic obstructive pulmonary disease (COPD). METHODS: We performed miRNA and mRNA profiling using high throughput stem-loop reverse-transcriptase quantitative polymerase chain reaction and mRNA microarray, respectively, on lung tissue of 30 patients (screening cohort) encompassing 8 never-smokers, 10 smokers without airflow limitation, and 12 smokers with COPD. Differential expression of miRNA-218-5p (miR-218-5p) was validated by reverse-transcriptase quantitative polymerase chain reaction in an independent cohort of 71 patients, an in vivo murine model of COPD, and primary human bronchial epithelial cells. Localization of miR-218-5p was assessed by in situ hybridization. In vitro and in vivo perturbation of miR-218-5p combined with RNA sequencing and gene set enrichment analysis was used to elucidate its functional role in COPD pathogenesis. MEASUREMENTS AND MAIN RESULTS: Several miRNAs were differentially expressed among the different patient groups. Interestingly, miR-218-5p was significantly down-regulated in smokers without airflow limitation and in patients with COPD compared with never-smokers. Decreased pulmonary expression of miR-218-5p was validated in an independent validation cohort, in cigarette smoke-exposed mice, and in human bronchial epithelial cells. Importantly, expression of miR-218-5p strongly correlated with airway obstruction. Furthermore, cellular localization of miR-218-5p in human and murine lung revealed highest expression of miR-218-5p in the bronchial airway epithelium. Perturbation experiments with a miR-218-5p mimic or inhibitor demonstrated a protective role of miR-218-5p in cigarette smoke-induced inflammation and COPD. CONCLUSIONS: We highlight a role for miR-218-5p in the pathogenesis of COPD.