Hypertonic Saline Is Effective in the Prevention and Treatment of Mucus Obstruction, but Not Airway Inflammation, in Mice with Chronic Obstructive Lung Disease

Recent evidence suggests that inadequate hydration of airway surfaces is a common mechanism in the pathogenesis of airway mucus obstruction. Inhaled hypertonic saline (HS) induces osmotic water flux, improving hydration of airway surfaces. However, trials in patients with obstructive lung diseases are limited. The aim of this study was to investigate effects of HS on mucus obstruction and airway inflammation in the prevention and treatment of obstructive lung disease in vivo. We, therefore, used the β-epithelial Na+ channel (βENaC)–overexpressing mouse as a model of chronic obstructive lung disease and determined effects of preventive and late therapy with 3% HS and 7% HS on pulmonary mortality, airway mucus obstruction, and inflammation. We found that preventive treatment with 3% HS and 7% HS improved growth, reduced mortality, and reduced mucus obstruction in neonatal βENaC-overexpressing mice. In adult βENaC-overexpressing mice with chronic lung disease, mucus obstruction was significantly reduced by 7% HS, but not by 3% HS. Treatment with HS triggered airway inflammation with elevated keratinocyte chemoattractant levels and neutrophils in airways from wild-type mice, but reduced keratinocyte chemoattractant in chronic neutrophilic inflammation in adult βENaC-overexpressing mice. Our data demonstrate that airway surface rehydration with HS provides an effective preventive and late therapy of mucus obstruction with no consistent effects on inflammation in chronic lung disease. These results suggest that, through mucokinetic effects, HS may be beneficial for patients with a spectrum of obstructive lung diseases, and that additional strategies are required for effective treatment of associated airway inflammation

Hypoxic Epithelial Necrosis Triggers Neutrophilic Inflammation via IL-1 Receptor Signaling in Cystic Fibrosis Lung Disease

Rationale: In many organs, hypoxic cell death triggers sterile neutrophilic inflammation via IL-1R signaling. Although hypoxia is common in airways from patients with cystic fibrosis (CF), its role in neutrophilic inflammation remains unknown. We recently demonstrated that hypoxic epithelial necrosis caused by airway mucus obstruction precedes neutrophilic inflammation in Scnn1b-transgenic (Scnn1b-Tg) mice with CF-like lung disease

Genetic background modulates CFTR-mediated Cl⁻ secretion in airways of wild-type and βENaC-Tg mice.

<p>A–C) Effects of genetic background on transepithelial Cl- secretion were determined by adding bumetanide or CFTR_inh-172 to amiloride-pretreated tracheal tissues from neonatal wild-type (WT) and βENaC-Tg mice on the C57BL/6 and BALB/c background. A,B) Summary of bumetanide-sensitive I_sc (A) and CFTR_inh-172-sensitive I_sc (B) in the presence of amiloride (<i>n</i> = 12–23 mice per group). C) Summary of CFTR_inh-172-sensitive I_sc in the presence of amiloride and cAMP-dependent activation (IBMX/forskolin) (<i>n</i> = 5–11 mice per group). Data are presented as mean ± SEM. ^†<i>P</i><0.05 and ^††<i>P</i><0.01 compared with mice of same genotype on C57BL/6 background.</p

Genetic Deletion of <i>Mmp9</i> Does Not Reduce Airway Inflammation and Structural Lung Damage in Mice with Cystic Fibrosis-like Lung Disease

Elevated levels of matrix metalloprotease 9 (MMP-9) and neutrophil elastase (NE) are associated with bronchiectasis and lung function decline in patients with cystic fibrosis (CF). MMP-9 is a potent extracellular matrix-degrading enzyme which is activated by NE and has been implicated in structural lung damage in CF. However, the role of MMP-9 in the in vivo pathogenesis of CF lung disease is not well understood. Therefore, we used β-epithelial Na+ channel-overexpressing transgenic (βENaC-Tg) mice as a model of CF-like lung disease and determined the effect of genetic deletion of Mmp9 (Mmp9-/-) on key aspects of the pulmonary phenotype. We found that MMP-9 levels were elevated in the lungs of βENaC-Tg mice compared with wild-type littermates. Deletion of Mmp9 had no effect on spontaneous mortality, inflammatory markers in bronchoalveolar lavage, goblet cell metaplasia, mucus hypersecretion and emphysema-like structural lung damage, while it partially reduced mucus obstruction in βENaC-Tg mice. Further, lack of Mmp9 had no effect on increased inspiratory capacity and increased lung compliance in βENaC-Tg mice, whereas both lung function parameters were improved with genetic deletion of NE. We conclude that MMP-9 does not play a major role in the in vivo pathogenesis of CF-like lung disease in mice

CFTR Regulates Early Pathogenesis of Chronic Obstructive Lung Disease in βENaC-Overexpressing Mice

<div><h3>Background</h3><p>Factors determining the onset and severity of chronic obstructive pulmonary disease remain poorly understood. Previous studies demonstrated that airway surface dehydration in βENaC-overexpressing (βENaC-Tg) mice on a mixed genetic background caused either neonatal mortality or chronic obstructive lung disease suggesting that the onset of lung disease was modulated by the genetic background.</p> <h3>Methods</h3><p>To test this hypothesis, we backcrossed βENaC-Tg mice onto two inbred strains (C57BL/6 and BALB/c) and studied effects of the genetic background on neonatal mortality, airway ion transport and airway morphology. Further, we crossed βENaC-Tg mice with CFTR-deficient mice to validate the role of CFTR in early lung disease.</p> <h3>Results</h3><p>We demonstrate that the C57BL/6 background conferred increased CFTR-mediated Cl⁻ secretion, which was associated with decreased mucus plugging and mortality in neonatal βENaC-Tg C57BL/6 compared to βENaC-Tg BALB/c mice. Conversely, genetic deletion of CFTR increased early mucus obstruction and mortality in βENaC-Tg mice.</p> <h3>Conclusions</h3><p>We conclude that a decrease or absence of CFTR function in airway epithelia aggravates the severity of early airway mucus obstruction and related mortality in βENaC-Tg mice. These results suggest that genetic or environmental factors that reduce CFTR activity may contribute to the onset and severity of chronic obstructive pulmonary disease and that CFTR may serve as a novel therapeutic target.</p> </div

Survival of βENaC-Tg mice is modified by genetic background.

<p>Survival curves of βENaC-Tg mice and wild-type (WT) littermates on mixed (C3H/He x C57BL/6), C57BL/6 and BALB/c backgrounds (n = 36–59 mice per group). *<i>P</i><0.05 and ^†<i>P</i><0.01 (Kaplan-Meier survival analysis).</p

Genetic background modulates airway ion transport in wild-type and βENaC-Tg mice.

<p>A) Representative original recordings of the effects of amiloride and cAMP-dependent activation (IBMX/forskolin) on transepithelial voltage (V_te) and transepithelial resistance (R_te) across freshly excised tracheal tissues from neonatal (3-day-old) wild-type (WT) and βENaC-Tg mice on the C57BL/6 and BALB/c background. R_te was determined from V_te deflections obtained by pulsed current injection. (B–F) Summary of basal equivalent short-circuit current (I_sc) (B), amiloride-sensitive I_sc (C), amiloride-insensitive I_sc (D), cAMP-induced I_sc (E) and UTP-induced I_sc (F) in freshly excised tracheal tissues from neonatal WT and βENaC-Tg mice on the C57BL/6 and BALB/c background. Data are presented as mean ± SEM (<i>n</i> = 8–20 mice per group). *<i>P</i><0.05 and **<i>P</i><0.001 compared with WT mice on same strain background, ^†<i>P</i><0.05 and ^††<i>P</i><0.01 compared with mice of same genotype on C57BL/6 background.</p

Genetic background modifies early airway mucus obstruction and epithelial necrosis in neonatal βENaC-Tg mice.

<p>A) Longitudinal sections of tracheae from neonatal (3-day-old) wild-type (WT) and βENaC-Tg mice on the C57BL/6 and BALB/c background. Sections were stained with Alcian blue–periodic acid Schiff (AB-PAS) to determine the presence of mucus and goblet cells. Scale bars  = 200 µm. B–D) Summary of mucus content, as determined from measuring volume density of AB-PAS-positive material in the tracheal lumen (B), goblet cell numbers (C) and Transcript levels of Muc5b (D) (<i>n</i> = 4–13 mice per group). E) Airway histology from neonatal (3-day-old) WT and βENaC-Tg mice on the C57BL/6 and BALB/c background. Sections were stained with hematoxylin and eosin (H&E) and evaluated for degenerative airway epithelial cells (arrows). Scale bars  = 40 µm (upper panels) and 20 µm (lower panels). F) Summary of airway epithelial necrosis as determined from the number of degenerative epithelial cells per mm of the basement membrane (<i>n</i> = 9–11 mice per group). *<i>P</i><0.001 compared with WT mice on same strain background, ^†<i>P</i><0.05 compared with mice of same genotype on C57BL/6 background.</p