Search

Rationale: Chronic inflammation plays an important role in alveolar tissue damage in emphysema, but the underlying immune alterations and cellular interactions are incompletely understood. Objectives: To explore disease-specific pulmonary immune cell alterations and cellular interactions in emphysema. Methods: We used single-cell mass cytometry (CyTOF) to compare the immune compartment in alveolar tissue from 15 patients with severe emphysema and 5 control subjects. Imaging mass cytometry (IMC) was applied to identify altered cell-cell interactions in alveolar tissue from patients with emphysema ( n  = 12) compared with control subjects ( n  = 8). Measurements and Main Results: We observed higher percentages of central memory CD4 T cells in combination with lower proportions of effector memory CD4 T cells in emphysema. In addition, proportions of cytotoxic central memory CD8 T cells and CD127 + CD27 + CD69 - T cells were higher in emphysema, the latter potentially reflecting an influx of circulating lymphocytes into the lungs. Central memory CD8 T cells, isolated from alveolar tissue from patients with emphysema, exhibited an IFN-γ response upon anti-CD3 and anti-CD28 activation. Proportions of CD1c + dendritic cells, expressing migratory and costimulatory markers, were higher in emphysema. Importantly, IMC enabled us to visualize increased spatial colocalization of CD1c + dendritic cells and CD8 T cells in emphysema in situ . Conclusions: Using CyTOF, we characterized the alterations of the immune cell signature in alveolar tissue from patients with chronic obstructive pulmonary disease stage III or IV emphysema versus control lung tissue. These data contribute to a better understanding of the pathogenesis of emphysema and highlight the feasibility of interrogating the immune cell signature using CyTOF and IMC in human lung tissue. Clinical trial registered with www.clinicaltrials.gov (NCT04918706).

FEV(1) is fundamental to the diagnosis and staging of chronic obstructive pulmonary disease. In emphysema, airflow obstruction usually coexists with impairment of gas exchange, but discordance is not infrequent. We hypothesized that variations in the distribution of emphysema would be associated with functional differences and therefore account for discordant physiology. We used quantitative computed tomography to assess emphysema severity and distribution in 119 subjects with alpha1-antitrypsin deficiency (PiZ phenotype) and grouped them according to distribution pattern. In the 102 subjects with emphysema, 65 had a predominantly basal pattern ("basal"), but 37 (36%) had greater involvement of the upper regions ("apical"). Subjects from each group were matched for total volume of emphysema and age, and matched pairs analysis was used to relate emphysema distribution to clinical phenotype. Basal distribution was associated with greater impairment of FEV(1) (mean difference, 9.9% predicted; 95% confidence interval, 3.8 to 16.0; p = 0.002) but less impairment of gas exchange (Pa(O(2)) mean difference, 0.5 kPa, 0.03 to 0.1; p = 0.016) and alveolar-arterial oxygen gradient (mean difference, 0.7 kPa; 0.2 to 1.2; p = 0.007) than the apical distribution. Emphysema distribution correlated with physiologic discordance (r = -0.409, p < 0.001). The use of single physiologic parameters as a surrogate measure of emphysema severity may introduce systematic bias in the staging of subjects with emphysema.

The fundamental importance of calibration for any measuring device is indisputable, but computed tomography (CT) calibration in longitudinal lung densitometry studies is largely unexplored. Although the validity of CT as a measure of emphysema has been confirmed in cross-sectional studies, there are limited data on long-term reproducibility, and this is critically important for validating its use as an outcome measure in therapeutic trials. A general understanding of the strengths and pitfalls of CT densitometry is critical for physicians reviewing the published literature using this methodology. In our study of 57 patients with alpha-1 antitrypsin deficiency (phenotype PiZ), progression of voxel index determined from three successive annual scans acquired with a fully calibrated scanner was intimately associated with changes in CT air densitometry, sampled from patient images. Images were therefore reanalyzed, using a correction technique validated in phantom studies that adjusted for changes in measured air density, and the reliability of the voxel index as a measure of emphysema progression was improved. Comparison of adjusted voxel index thresholds indicated the optimum threshold was -950 Hounsfield units. Internal air calibration is therefore critical in longitudinal and multicenter lung densitometry studies of emphysema and incorporation of a correction factor is essential for quantitative image analysis.

Chronic obstructive pulmonary disease (COPD) is defined by progressive, irreversible airflow limitation and an inflammatory response of the lungs, usually to cigarette smoke. However, COPD is a heterogeneous disease in terms of clinical, physiologic, and pathologic presentation. We aimed to evaluate whether airflow limitation, airway responsiveness, and airway inflammation are separate entities underlying the pathophysiology of COPD by using factor analysis. A total of 114 patients (99 males/15 females, age 62 +/- 8 years, 42 pack-years smoking, no inhaled or oral steroids > 6 months) with irreversible airflow limitation (postbronchodilator FEV(1) 63 +/- 9% predicted, FEV(1)/inspiratory vital capacity [IVC] 48 +/- 9%) and symptoms of chronic bronchitis or dyspnea were studied in a cross-sectional design. Postbronchodilator FEV(1) and FEV(1)/IVC, reversibility to inhaled beta(2)-agonists, diffusing capacity, provocative concentration of methacholine required to produce a 20% drop in FEV(1), total serum IgE, exhaled nitric oxide, and induced sputum cell counts (% eosinophils, % neutrophils) were collected. Factor analysis yielded 4 separate factors that accounted for 63.6% of the total variance. Factor 1 was comprised of FEV(1), FEV(1)/IVC, and residual volume/total lung capacity. Factor 2 included reversibility, IgE, provocative concentration of methacholine required to produce a 20% drop in FEV(1,) and diffusing capacity. Factor 3 contained exhaled nitric oxide and factor 4 included sputum % neutrophils and % eosinophils. We conclude that airflow limitation, airway inflammation, and features commonly associated with asthma are separate and largely independent factors in the pathophysiology of COPD.

Cigarette smoking results in oxidative stress and inflammation in the lungs, which are involved in the pathogenesis of chronic obstructive pulmonary disease (COPD). 4-Hydroxy-2-nonenal (4-HNE), a highly reactive diffusible product of lipid peroxidation, is a key mediator of oxidant-induced cell signaling and apoptosis. 4-HNE has a high affinity toward cysteine, histidine, and lysine groups and forms direct protein adducts. We investigated the presence of 4-HNE-modified proteins in lung tissue obtained from subjects with and without COPD. We studied 23 current or ex-smokers with similar smoking histories with COPD (n = 11; FEV(1) < 70% predicted) or without COPD (n = 12; FEV(1) > 84% predicted) who had undergone lung resection. As 4-HNE and transforming growth factor-beta(1) (TGF-beta(1)) can modulate gamma-glutamylcysteine synthetase (gamma-GCS) mRNA levels in lung cells, we assessed the relations between 4-HNE-modified protein levels, FEV(1), gamma-GCS, and TGF-beta(1). 4-HNE-modified protein levels were elevated in airway and alveolar epithelial cells, endothelial cells, and neutrophils in subjects with COPD, compared with the levels in subjects without COPD (p < 0.01). We also observed a significant inverse correlation between the levels of 4-HNE adducts in alveolar epithelium, airway endothelium, and neutrophils and FEV(1) (p < 0.05) and a positive correlation between 4-HNE adducts and TGF-beta(1) protein and mRNA as well as gamma-GCS mRNA levels in airway and alveolar epithelium (p < 0.01). The elevated levels of 4-HNE may play a role in the signaling events in lung inflammation leading to the imbalance of the expression of both proinflammatory mediators and protective antioxidant genes in COPD.

We have investigated whether restoration of the balance between neutrophil elastase and its inhibitor, alpha(1)-antitrypsin, can prevent the progression of pulmonary emphysema in patients with alpha(1)-antitrypsin deficiency. Twenty-six Danish and 30 Dutch ex-smokers with alpha(1)-antitrypsin deficiency of PI*ZZ phenotype and moderate emphysema (FEV(1) between 30% and 80% of predicted) participated in a double-blind trial of alpha(1)-antitrypsin augmentation therapy. The patients were randomized to either alpha(1)-antitrypsin (250 mg/kg) or albumin (625 mg/kg) infusions at 4-wk intervals for at least 3 yr. Self-administered spirometry performed every morning and evening at home showed no significant difference in decline of FEV(1) between treatment and placebo. Each year, the degree of emphysema was quantified by the 15th percentile point of the lung density histogram derived from computed tomography (CT). The loss of lung tissue measured by CT (mean +/- SEM) was 2.6 +/- 0.41 g/L/yr for placebo as compared with 1.5 +/- 0.41 g/L/yr for alpha(1)-antitrypsin infusion (p = 0.07). Power analysis showed that this protective effect would be significant in a similar trial with 130 patients. This is in contrast to calculations based on annual decline of FEV(1) showing that 550 patients would be needed to show a 50% reduction of annual decline. We conclude that lung density measurements by CT may facilitate future randomized clinical trials of investigational drugs for a disease in which little progress in therapy has been made in the past 30 yr.

Chronic airways inflammation is one of the features of chronic obstructive pulmonary disease (COPD). We demonstrated previously that bronchiolar epithelium in COPD contains increased numbers of macrophages and mast cells. Transforming growth factor beta1 (TGF-beta1) may be involved in this influx because it has chemotactic activity for macrophages and mast cells. In this study, we examined expression patterns of TGF-beta1, TGF-beta receptors type I and II (TGF-betaRI and TGF-betaRII) by immunohistochemistry and mRNA in situ hybridization in peripheral lung tissue of 14 current or ex-smokers with COPD (FEV1 < 75%) and 14 without COPD (FEV1 > 84%). In both groups, TGF-beta1 and its receptors are present in airway and alveolar epithelial cells, airway and vascular smooth muscle cells, and tissue and alveolar CD68(+) cells (considered herein to be macrophages). In subjects with COPD, a semiquantitative analysis revealed approximately twofold higher levels of TGF-beta1 mRNA and protein in bronchiolar and alveolar epithelium (p < 0.02) as compared with subjects without COPD. With regard to bronchiolar epithelial cells, we found a significant correlation between TGF-beta1 mRNA and protein expression (r = 0.62; p < 0.002), and between the FEV1 of all subjects together and TGF-beta1 protein (r = -0.60; p < 0.0002) and mRNA (r = -0.67; p < 0. 002) levels. The epithelial expression of TGF-beta1 mRNA and TGF-beta1 protein correlates with the number of intraepithelial macrophages (both: r = 0.44; p < 0.03) whereas intraepithelial mast cell numbers correlate with epithelial TGF-beta1 mRNA expression. These data suggest a role for TGF-beta1 in recruiting macrophages into the airway epithelium in COPD.