Increased circulating β2-adrenergic receptor autoantibodies are associated with smoking-related emphysema

Abstract

Smoking is a dominant risk factor for chronic obstructive pulmonary disease (COPD) and emphysema, but not every smoker develops emphysema. Immune responses in smokers vary. Some autoantibodies have been shown to contribute to the development of emphysema in smokers. β₂-adrenergic receptors (β₂-ARs) are important targets in COPD therapy. β₂-adrenergic receptor autoantibodies (β₂-AAbs), which may directly affect β₂-ARs, were shown to be increased in rats with passive-smoking-induced emphysema in our current preliminary studies. Using cigarette-smoke exposure (CS-exposure) and active-immune (via injections of β₂-AR second extracellular loop peptides) rat models, we found that CS-exposed rats showed higher serum β₂-AAb levels than control rats before alveolar airspaces became enlarged. Active-immune rats showed increased serum β₂-AAb levels, and exhibited alveolar airspace destruction. CS-exposed-active-immune treated rats showed more extensive alveolar airspace destruction than rats undergoing CS-exposure alone. In our current clinical studies, we showed that plasma β₂-AAb levels were positively correlated with the RV/TLC (residual volume/total lung capacity) ratio (r = 0.455, p < 0.001) and RV%pred (residual volume/residual volume predicted percentage, r = 0.454, p < 0.001) in 50 smokers; smokers with higher plasma β₂-AAb levels exhibited worse alveolar airspace destruction. We suggest that increased circulating β₂-AAbs are associated with smoking-related emphysema.

Figure 1. Passive-smoking rats showed higher serum β₂-AAb levels than control rats before alveolar airspaces became enlarged.

(A) Levels of β₂-AAb in serum samples of CS-exposed rats and control rats were assessed at different time points. SE-ELISAs were performed after rats were exposed to clean air or cigarette smoke for 4, 8, 12, or 16 weeks. (B) Statistical analyses of mean linear intercept (MLI) measurements of lung sections after rats were exposed to clean air or cigarette smoke for 8 or 16 weeks. (C) Sections of lungs from a control rat and from a CS-exposed rat sacrificed at the end of the 8^th or 16^th week. Haematoxylin-eosin staining, scale bar = 200 μm. (D) and (E) Lower and higher groups refer to CS-exposed rats with relatively lower and higher serum β₂-AAb levels, respectively, and results of the statistical analysis of serum β₂-AAb levels and MLI values of the two groups were shown in panel (D) and (E) respectively. *p < 0.05, **p < 0.01, ***p < 0.001; Control group vs Smoke group or Lower group vs Higher group (n = 18 for the Control group and Smoke group at the 4^th or 8^th week in panel A, n = 12 for the Control group and Smoke group at the 12 ^th or 16^th week in panel A; n = 6 for the Control group and Smoke group at the 8^th week in panel B, n = 12 for the Control group and Smoke group at the 16^th week in panel B; n = 6 for the Lower group and Higher group at the 16^th week in panels D and E).

Figure 2. Measurements after control or passive-smoking rats were immunized or not with β₂-AR ECL_II peptides for 16 weeks.

(A) SE-ELISAs were performed to detect the levels of β₂-AAbs in the serum samples of rats from the four groups: Control group (Control), Active-Immune group (Immune), Passive-smoking group (Smoking), and Passive-smoking-active-immune group (Smoking + Immune). Analyses were conducted between the Control group and Immune group/Smoking group as well as between the Smoking group and Smoking + Immune group. (B) Lung-section MLI measurements statistical results from the Control group and Immune group as well as from Smoking group and Smoking + Immune group. (C) Section of lung from a control rat showing normal alveolar structures and sections from rats in the other three groups showing enlarged airspaces. Haematoxylin-eosin staining, scale bar = 200 μm. (D) Statistical analysis of lung function parameters (RV (residual volume), TLC (total lung capacity) and RV/TLC (residual volume/total lung capacity) ratio) between the Control group and Immune group/Smoking group as well as between the Smoking group and Smoking + Immune group. *p < 0.05, **p < 0.01, ***p < 0.001; Control group vs Immune group/Smoking group (n = 7). ^###p < 0.001 and ^#p < 0.05; Smoke group vs Smoke + Immune group (n = 7), ns means no significant difference.

Figure 3. Correlation analysis between the plasma β₂-AAb level and the lung function parameters.

Plasma β₂-AAb level were positively correlated with RV% pred (A) and RV/TLC ratio(B) and negatively correlated with FEV₁% pred(C) and FEV₁/FVC(D) ratio in 50 smokers. RV% pred (residual volume/residual volume predicted percentage), RV/TLC (residual volume/total lung capacity), FEV₁% pred (percent of forced expiratory volume in 1 second), FEV₁/FVC (forced vital capacity). Pearson or Spearman correlation analysis, n = 50. The hollow dots represent smokers without COPD, and the solid dots represent those with COPD.

Figure 4. Smokers/COPD patients with relatively higher plasma β₂-AAb levels exhibited worse LAA%.

Fifty smokers were divided into low-β₂-AAb and high-β₂-AAb groups. (A) and (B) show one slice of chest CT scans from a low-β₂-AAb smoker (A) and a high-β₂-AAb smoker (B); the blue regions in the slices indicate the low attenuation area under −950 HU. (C) and (D) show the distributions (median with interquartile range are shown) and results of the statistical analysis of the LAA% comparing low-β₂-AAb and high-β₂-AAb smokers (C) and COPD patients (D). (n = 25 for low-β₂-AAb and high-β₂-AAb smokers, n = 11 for low-β₂-AAb COPD patients and n = 18 for high-β₂-AAb COPD patients). HU: Hounsfield units. LAA%: the percentage of low attenuation area under −950 HU in the chest CT scans.