Airway Eosinophilia on Bronchoalveolar Lavage and the Risk of Exacerbations in COPD.

The associations between airway eosinophilia, measured in sputum or peripheral blood, and acute exacerbations of chronic obstructive pulmonary disease (AECOPD) are inconsistent. We therefore aimed to determine the association between eosinophilia in bronchoalveolar lavage (BAL) fluid and AECOPD in a clinical cohort. We analyzed differential cell counts from baseline BAL fluid in participants in the DISARM clinical trial (Clinicaltrials.gov #NCT02833480) and classified participants by the presence or absence of BAL eosinophilia (>1% of total leukocytes). We determined the association between BAL eosinophilia and AECOPD over 1 year of follow-up using negative binomial regression and Cox proportional hazards test. N = 63 participants were randomized, and N = 57 had BAL differential cell counts available. Participants with BAL eosinophilia (N = 21) had a significantly increased rate of acute exacerbations (unadjusted incidence rate ratio (IRR) 2.0, p = 0.048; adjusted IRR 2.24, p = 0.04) and a trend toward greater probability of acute exacerbation (unadjusted hazard ratio (HR) 1.74, p = 0.13; adjusted HR 2.3, p = 0.1) in the year of follow-up compared to participants without BAL eosinophilia (N = 36). These associations were not observed for BAL neutrophilia (N = 41 participants), BAL lymphocytosis (N = 27 participants) or peripheral blood eosinophilia at various threshold definitions (2%, N = 37; 3%, N = 27; 4%, N = 16). BAL may therefore be a sensitive marker of eosinophilic inflammation in the distal lung and may be of benefit for risk stratification or biomarker-guided therapy in COPD.

Inhaled Corticosteroids Selectively Alter the Microbiome and Host Transcriptome in the Small Airways of Patients with Chronic Obstructive Pulmonary Disease.

BACKGROUND: Patients with chronic obstructive pulmonary disease (COPD) are commonly treated with inhaled corticosteroid/long-acting ß2-agonist combination therapy. While previous studies have investigated the host-microbiome interactions in COPD, the effects of specific steroid formulations on this complex cross-talk remain obscure. METHODS: We collected and evaluated data from the Study to Investigate the Differential Effects of Inhaled Symbicort and Advair on Lung Microbiota (DISARM), a randomized controlled trial. Bronchoscopy was performed on COPD patients before and after treatment with salmeterol/fluticasone, formoterol/budesonide or formoterol-only. Bronchial brush samples were processed for microbial 16S rRNA gene sequencing and host mRNA sequencing. Longitudinal changes in the microbiome at a community, phylum and genus level were correlated with changes in host gene expression using a Spearman's rank correlation test. FINDINGS: In COPD patients treated with salmeterol/fluticasone, the expression levels of 676 host genes were significantly correlated to changes in the alpha diversity of the small airways. At a genus level, the expression levels of 122 host genes were significantly related to changes in the relative abundance of Haemophilus. Gene enrichment analyses revealed the enrichment of pathways and biological processes related to innate and adaptive immunity and inflammation. None of these changes were evident in patients treated with formoterol/budesonide or formoterol alone. INTERPRETATION: Changes in the microbiome following salmeterol/fluticasone treatment are related to alterations in the host transcriptome in the small airways of patients with COPD. These data may provide insights into why some COPD patients treated with inhaled corticosteroids may be at an increased risk for airway infection, including pneumonia. FUNDING: The Canadian Institute of Health Research, the British Columbia Lung Association, and an investigator-initiated grant from AstraZeneca.

Altered Polarization and Impaired Phagocytic Activity of Lung Macrophages in People With Human Immunodeficiency Virus and Chronic Obstructive Pulmonary Disease.

BACKGROUND: People with human immunodeficiency virus (PWH) have an increased risk of developing chronic obstructive pulmonary disease (COPD). METHODS: We phenotyped lung macrophages in 4 subgroups-M1 (CD40+CD163-), M2 (CD40-CD163+), double positives (CD40+CD163+), and double negatives and (CD40-CD163-)-and we determined their phagocytic capacity in PWH with and without COPD. RESULTS: People with human immunodeficiency virus with COPD have more double-negative macrophages (84.1%) versus PWH without (54.3%) versus controls (23.9%) (P=.004) and reduced phagocytosis (P=.012). Double-negative macrophages had the worst phagocytic capacity (P<.001). CONCLUSIONS: People with human immunodeficiency virus with COPD have an abundance of nonpolarized macrophages, which have poor phagocytic capacity and therefore predispose PWH to increased risk of disease progression.

Radiofrequency therapy improves exercise capacity of mice with emphysema.

Emphysema is a common phenotype of chronic obstructive pulmonary disease (COPD). Although resection of emphysematous tissue can improve lung mechanics, it is invasive and fraught with adverse effects. Meanwhile, radiofrequency (RF) treatment is an extracorporeal method that leads to tissue destruction and remodeling, resulting in "volume reduction" and overall improvement in lung compliance of emphysematous lungs. Whether these changes lead to improved exercise tolerance is unknown. Here, we investigated the effectiveness of RF treatment to improve the exercise capacity of mice with emphysema. Fifty-two mice (7 weeks of age) were used in this experiment. A bilateral emphysema model was created by intratracheally instilling porcine pancreatic elastase (PPE) (1.5U/100 g body weight). RF treatment (0.5 W/ g body weight) was administered extracorporeally 14 days later and mice were sacrificed after another 21 days. The exercise capacity of mice was measured using a treadmill. Treadmill runs were performed just before PPE instillation (baseline), before RF treatment and before sacrifice. Following sacrifice, lung compliance and mean linear intercept (Lm) were measured and fibrosis was assessed using a modified Ashcroft score. There were 3 experimental groups: controls (instilled with saline, n = 12), emphysema (instilled with porcine pancreatic elastase, PPE, n = 11) and emphysema + treatment (instilled with PPE and given RF, n = 9). At endpoint, the maximum velocity of the emphysema + treatment group was significantly higher than that of the emphysema group, indicating improved exercise tolerance (86.29% of baseline vs 61.69% of baseline, p = 0.01). Histological analysis revealed a significant reduction in emphysema as denoted by Lm between the two groups (median 29.60 µm vs 35.68 µm, p = 0.03). The emphysema + treatment group also demonstrated a higher prevalence of lung fibrosis (≧Grade 3) compared with the emphysema group (11.7% vs 5.4%, p < 0.01). No severe adverse events from RF were observed. RF treatment improved the exercise capacity of mice with emphysema. These data highlight the therapeutic potential of RF treatment in improving the functional status of patients with COPD.

Effects of Inhaled Corticosteroid/Long-Acting ß2-Agonist Combination on the Airway Microbiome of Patients with Chronic Obstructive Pulmonary Disease: A Randomized Controlled Clinical Trial (DISARM).

Rationale: Inhaled corticosteroids (ICS) are commonly prescribed with long-acting ß2-agonists (LABA) in chronic obstructive pulmonary disease (COPD). To date, the effects of ICS therapy on the airway microbiome in COPD are unknown. Objectives: To determine the effects of ICS/LABA on the airway microbiome of patients with COPD. Methods: Clinically stable patients with COPD were enrolled into a 4-week run-in period during which ICS was discontinued and all participants were placed on formoterol (Form) 12 µg twice daily (BID). The participants were then randomized to budesonide/formoterol (Bud + Form; 400/12 µg BID), fluticasone/salmeterol (Flu + Salm; 250/50 µg BID), or formoterol only (12 µg BID) for 12 weeks. Participants underwent bronchoscopy before and after the 12-week treatment period. The primary endpoint was the comparison of changes in the airway microbiome over the trial period between the ICS/LABA and LABA-only groups. Measurements and Main Results: Sixty-three participants underwent randomization: Bud + Form (n = 20), Flu + Salm (n = 22), and Form (n = 21) groups; 56 subjects completed all visits. After the treatment period, changes in α-diversity were significantly different across groups, especially between Flu + Salm and Form groups (Δrichness: P = 0.02; ΔShannon index: P = 0.03). Longitudinal differential abundance analyses revealed more pronounced microbial shifts from baseline in the fluticasone (vs. budesonide or formoterol only) group. Conclusions: Fluticasone-based ICS/LABA therapy modifies the airway microbiome in COPD, leading to a relative reduction in α-diversity and a greater number of bacterial taxa changes. These data may have implications in patients who develop pneumonia on ICS. Clinical trial registered with www.clinicaltrials.gov (NCT02833480).

A mechanical test of the tenertaxis hypothesis for leukocyte diapedesis.

As part of the immune response, leukocytes can directly transmigrate through the body of endothelial cells or through the gap between adjacent endothelial cells. These are known, respectively, as the transcellular and paracellular route of diapedesis. What determines the usage of one route over the other is unclear. A recently proposed tenertaxis hypothesis claims that leukocytes choose the path with less mechanical resistance against leukocyte protrusions. We examined this hypothesis using numerical simulation of the mechanical resistance during paracellular and transcellular protrusions. By using parameters based on human lung endothelium, our results show that the required force to breach the endothelium through the transcellular route is greater than paracellular route, in agreement with experiments. Moreover, experiments have demonstrated that manipulation of the relative strength between the two routes can make the transcellular route preferable. Our simulations have demonstrated this reversal and thus tentatively confirmed the hypothesis of tenertaxis.

Macrophages with reduced expressions of classical M1 and M2 surface markers in human bronchoalveolar lavage fluid exhibit pro-inflammatory gene signatures.

The classical M1/M2 polarity of macrophages may not be applicable to inflammatory lung diseases including chronic obstructive pulmonary disease (COPD) due to the complex microenvironment in lungs and the plasticity of macrophages. We examined macrophage sub-phenotypes in bronchoalveolar lavage (BAL) fluid in 25 participants with CD40 (a M1 marker) and CD163 (a M2 marker). Of these, we performed RNA-sequencing on each subtype in 10 patients using the Illumina NextSeq 500. Approximately 25% of the macrophages did not harbor classical M1 or M2 surface markers (double negative, DN), and these cells were significantly enriched in COPD patients compared with non-COPD patients (46.7% vs. 14.5%, p < 0.001). 1886 genes were differentially expressed in the DN subtype compared with  all other subtypes at a 10% false discovery rate. The 602 up-regulated genes included 15 mitochondrial genes and were enriched in 86 gene ontology (GO) biological processes including inflammatory responses. Modules associated with cellular functions including oxidative phosphorylation were significantly down-regulated in the DN subtype. Macrophages in the human BAL fluid, which were negative for both M1/M2 surface markers, harbored a gene signature that was pro-inflammatory and suggested dysfunction in cellular homeostasis. These macrophages may contribute to the pathogenesis and manifestations of inflammatory lung diseases such as COPD.

COPD and the risk of poor outcomes in COVID-19: A systematic review and meta-analysis.

BACKGROUND: Patients with chronic obstructive pulmonary disease (COPD) are highly susceptible from respiratory exacerbations from viral respiratory tract infections. However, it is unclear whether they are at increased risk of COVID-19 pneumonia or COVID-19-related mortality. We aimed to determine whether COPD is a risk factor for adverse COVID-19 outcomes including hospitalization, severe COVID-19, or death. METHODS: Following the PRISMA guidelines, we performed a systematic review of COVID-19 clinical studies published between November 1st, 2019 and January 28th, 2021 (PROSPERO ID: CRD42020191491). We included studies that quantified the number of COPD patients, and reported at least one of the following outcomes stratified by COPD status: hospitalization; severe COVID-19; ICU admission; mechanical ventilation; acute respiratory distress syndrome; or mortality. We meta-analyzed the results of individual studies to determine the odds ratio (OR) of these outcomes in patients with COPD compared to those without COPD. FINDINGS: Fifty-nine studies met the inclusion criteria, and underwent data extraction. Most studies were retrospective cohort studies/case series of hospitalized patients. Only four studies examined the effects of COPD on COVID-19 outcomes as their primary endpoint. In aggregate, COPD was associated with increased odds of hospitalization (OR 4.23, 95% confidence interval [CI] 3.65-4.90), ICU admission (OR 1.35, 95% CI 1.02-1.78), and mortality (OR 2.47, 95% CI 2.18-2.79). INTERPRETATION: Having a clinical diagnosis of COPD significantly increases the odds of poor clinical outcomes in patients with COVID-19. COPD patients should thus be considered a high-risk group, and targeted for preventative measures and aggressive treatment for COVID-19 including vaccination.

Abundance of Non-Polarized Lung Macrophages with Poor Phagocytic Function in Chronic Obstructive Pulmonary Disease (COPD).

Lung macrophages are the key immune effector cells in the pathogenesis of Chronic Obstructive Pulmonary Disease (COPD). Several studies have shown an increase in their numbers in bronchoalveolar lavage fluid (BAL) of subjects with COPD compared to controls, suggesting a pathogenic role in disease initiation and progression. Although reduced lung macrophage phagocytic ability has been previously shown in COPD, the relationship between lung macrophages' phenotypic characteristics and functional properties in COPD is still unclear. (1) Methods: Macrophages harvested from bronchoalveolar lavage (BAL) fluid of subjects with and without COPD (GOLD grades, I-III) were immuno-phenotyped, and their function and gene expression profiles were assessed using targeted assays. (2) Results: BAL macrophages from 18 COPD and 10 (non-COPD) control subjects were evaluated. The majority of macrophages from COPD subjects were non-polarized (negative for both M1 and M2 markers; 77.9%) in contrast to controls (23.9%; p < 0.001). The percentages of these non-polarized macrophages strongly correlated with the severity of COPD (p = 0.006) and current smoking status (p = 0.008). Non-polarized macrophages demonstrated poor phagocytic function in both the control (p = 0.02) and COPD (p < 0.001) subjects. Non-polarized macrophages demonstrated impaired ability to phagocytose Staphylococcus aureus (p < 0.001). They also demonstrated reduced gene expression for CD163, CD40, CCL13 and C1QA&B, which are involved in pathogen recognition and processing and showed an increased gene expression for CXCR4, RAF1, amphiregulin and MAP3K5, which are all involved in promoting the inflammatory response. (3) Conclusions: COPD is associated with an abundance of non-polarized airway macrophages that is related to the severity of COPD. These non-polarized macrophages are predominantly responsible for the poor phagocytic capacity of lung macrophages in COPD, having reduced capacity for pathogen recognition and processing. This could be a key risk factor for COPD exacerbation and could contribute to disease progression.

Widespread Sexual Dimorphism in the Transcriptome of Human Airway Epithelium in Response to Smoking.

Epidemiological studies have shown that female smokers are at higher risk of chronic obstructive pulmonary disease (COPD). Female patients have worse symptoms and health status and increased risk of exacerbations. We determined the differences in the transcriptome of the airway epithelium between males and females, as well the sex-by-smoking interaction. We processed public gene expression data of human airway epithelium into a discovery cohort of 211 subjects (never smokers n = 68; current smokers n = 143) and two replication cohorts of 104 subjects (21 never, 52 current, and 31 former smokers) and 238 subjects (99 current and 139 former smokers. We analyzed gene differential expression with smoking status, sex, and smoking-by-sex interaction and used network approaches for modules' level analyses. We identified and replicated two differentially expressed modules between the sexes in response to smoking with genes located throughout the autosomes and not restricted to sex chromosomes. The two modules were enriched in autophagy (up-regulated in female smokers) and response to virus and type 1 interferon signaling pathways which were down-regulated in female smokers compared to males. The results offer insights into the molecular mechanisms of the sexually dimorphic effect of smoking, potentially enabling a precision medicine approach to smoking related lung diseases.

Measuring and Remediating Nonspecific Modifications of Gold Surfaces Using a Coupled in Situ Electrochemical Fluorescence Microscopic Methodology.

In surface-based biosensors, the nonspecific or undesired adsorption of the probe is an important characteristic that is typically difficult to measure and therefore to control or eliminate. A methodology for measuring and then minimizing or eliminating this problem on gold surfaces, readily applicable to many common surface modifications is presented. Combining electrochemical perturbation and fluorescence microscopy, we show that the potential at which the adsorbed species is removed can be used as an estimate of the strength of the adsorbate-surface interaction. This desorption potential can be easily measured through an increase in fluorescence intensity as the potential is manipulated. Furthermore, this method can be used to evaluate strategies for preventing or removing nonspecific adsorption. This is demonstrated for a wide variety of surface modifications, from strongly chemisorbed monolayers such as thiol self-assembled monolayers (SAMs) to physisorbed monolayers as well as for complex surface structures like peptide and DNA mixed-component SAMs. The use of a coadsorption strategy or small magnitude potential-step cycles was shown to significantly decrease the amount of nonspecifically or noncovalently bound probe, creating better defined surfaces.