Respiratory surfaces are exposed to billions of particulates and pathogens daily. A protective mucus barrier traps and eliminates them via mucociliary clearance (MCC)1,2. However, excessive mucus contributes to transient respiratory infections and to the pathogenesis of numerous respiratory diseases1. MUC5AC and MUC5B are evolutionarily conserved genes that encode structurally related mucin glycoproteins, the principal macromolecules in airway mucus1,3. Genetic variants are linked to diverse lung diseases4-6, but specific roles for MUC5AC and MUC5B in MCC, and the lasting effects of their inhibition, are unknown. Here we show that Muc5b (but not Muc5ac) is required for MCC, for controlling infections in the airways and middle ear, and for maintaining immune homeostasis in the lungs. Muc5b deficiency caused materials to accumulate in upper and lower airways. This defect led to chronic infection by multiple bacterial species, including Staphylococcus aureus, and to inflammation that failed to resolve normally7. Apoptotic macrophages accumulated, phagocytosis was impaired, and IL-23 production was reduced inMuc5b−/− mice. By contrast, in Muc5b transgenic (Tg) mice, macrophage functions improved. Existing dogma defines mucous phenotypes in asthma and chronic obstructive pulmonary disease (COPD) as driven by increased MUC5AC, with MUC5B levels either unaffected or increased in expectorated sputum1,8. However, in many patients, MUC5B production at airway surfaces decreases by as much as 90%9-11. By distinguishing a specific role for Muc5b in MCC, and by determining its impact on bacterial infections and inflammation in mice, our results provide a refined framework for designing targeted therapies to control mucin secretion and restore MCC.

Idiopathic Pulmonary Fibrosis (IPF) is a lethal lung disease with progressive fibrosis and death within 2–3 years of diagnosis. IPF incidence and prevalence rates are increasing annually with few effective treatments available. Inhibition of interleukin 6 (IL-6) results in the attenuation of pulmonary fibrosis in mice. It is unclear whether this is due to blockade of classical signaling, mediated by membrane-bound IL-6 receptor alpha (mIL-6Rα), or trans signaling, mediated by soluble IL-6Rα (sIL-6Rα). Our study assessed the role of sL-6Rα in IPF. We demonstrated elevations of sIL-6Rα in IPF patients and in mice during the onset and progression of fibrosis. We demonstrated that protease-mediated cleavage from lung macrophages was important in production of sL-6Rα. In vivo neutralization of sIL-6Rα attenuated pulmonary fibrosis in mice as seen by reductions in myofibroblasts, fibronectin and collagen in the lung. In vitro activation of IL-6 trans signaling enhanced fibroblast proliferation and extracellular matrix protein production, effects relevant in the progression of pulmonary fibrosis. Together these findings demonstrate that the production of sL-6Rα from macrophages in the diseased lung contributes to IL-6 trans signaling that in turn influences events crucial in pulmonary fibrosis.

Background High altitude is a challenging condition caused by insufficient oxygen (O2) supply. Inability to adjust to hypoxia may lead to pulmonary edema, stroke, cardiovascular dysfunction and even death. Thus, understanding the molecular basis of adaptation to high altitude may reveal novel therapeutics to counteract the detrimental consequences of hypoxia. Methods Using high throughput unbiased metabolomic profiling, we report that the metabolic pathway responsible for production of erythrocyte 2,3-bisphosphoglycerate (2,3-BPG), a negative allosteric regulator of hemoglobin-O2 binding affinity, was significantly induced in 21 healthy humans within two hours of arrival at 5260m, and further increased following 16 days at 5260m. Results This finding led us to uncover discover that plasma adenosine concentrations and soluble CD73 (sCD73) activity rapidly increased at high altitude and were associated with elevated erythrocyte 2,3-BPG levels and O2 releasing capacity. Mouse genetic studies demonstrated that elevated CD73 contributed to hypoxia-induced adenosine accumulation and that elevated adenosine-mediated erythrocyte A2B adenosine receptor (ADORA2B) activation was beneficial by inducing 2,3-BPG production, triggering O2 release to prevent multiple tissue hypoxia, inflammation and pulmonary vascular leakage. Mechanistically, we demonstrated that erythrocyte AMP-activated protein kinase (AMPK) was activated in humans at high altitude and that AMPK is a key protein functioning downstream of ADORA2B, phosphorylating and activating BPG mutase and in this way inducing 2,3-BPG production and O2 release from erythrocytes. Significantly, preclinical studies demonstrated that activation of AMPK enhanced BPG mutase activation, 2,3-BPG production and O2 release capacity in CD73-deficient mice, in erythrocyte specific ADORA2B knockouts, and in wild type mice and in turn reduced tissue hypoxia, and inflammation. Conclusions Altogether, both human and mouse studies reveal novel mechanisms of hypoxia adaptation and potential therapeutic approaches for counteracting hypoxia-induced tissue damage.

Lung fibrosis is the hallmark of the interstitial lung diseases. Alveolar epithelial cell (AEC) injury is akey step that contributes to a profibrotic microenvironment. Fibroblasts and myofibroblasts subsequently accumulate and deposit excessive extracellular matrix. In addition to TGF‐β, the IL‐6 family of cytokines, which signal through STAT‐3, may also contribute to lung fibrosis. In the current manuscript, the extent to which STAT‐3 inhibition decreases lung fibrosis is investigated. Phosphorylated STAT‐3 was elevated in lung biopsies from patients with idiopathic pulmonary fibrosis and bleomycin (BLM)‐induced fibrotic murine lungs. C‐188‐9, a small molecule STAT‐3 inhibitor, decreased pulmonary fibrosis in the intraperitoneal BLM model as assessed by arterial oxygen saturation (control, 84.4 ± 1.3%; C‐188‐9, 94.4 ± 0.8%), histology (Ashcroft score: untreated, 5.4 ± 0.25; C‐188‐9, 3.3 ± 0.14), and attenuated fibrotic markers such as diminished α‐smooth muscle actin, reduced collagen deposition. In addition, C‐188‐9 decreased the expression of epithelial injury markers, including hypoxia‐inducible factor‐1α (HIF‐1α) and plasminogen activator inhibitor‐1 (PAI‐1). In vitro studies show that inhibition of STAT‐3 decreased IL‐6‐ and TGF‐β‐induced expression of multiple genes, including HIF‐1α and PAI‐1, in AECs. Furthermore, C‐188‐9 decreased fibroblast‐to‐myofibroblast differentiation. Finally, TGF‐β stimulation of lung fibroblasts resulted in SMAD2/SMAD3‐dependent phosphorylation of STAT‐3. These findings demonstrate that STAT‐3 contributes to the development of lung fibrosis and suggest that STAT‐3 may be a therapeutic target in pulmonary fibrosis.—Pedroza, M., Le, T. T., Lewis, K., Karmouty‐Quintana, H., To, S., George, A. T., Blackburn, M. R., Tweardy, D. J., Agarwal, S. K. STAT‐3 contributes to pulmonary fibrosis through epithelial injury and fibroblast‐myofibroblast differentiation. FASEB J. 30, 129‐140 (2016). http://www.fasebj.org

Our study highlights important roles for APA in human cancer, including reshaping cellular pathways and regulating specific gene expression, exemplifying the complex interplay between APA and other biological processes and yielding new insights into the action mechanism of cancer drugs.