In asthma, airflow obstruction is thought to result primarily from inflammation-triggered airway smooth muscle (ASM) contraction. However, anti-inflammatory and smooth muscle-relaxing treatments are often temporary or ineffective. Overproduction of the mucin MUC5AC is an additional disease feature that, while strongly associated pathologically, is poorly understood functionally. Here we show that Muc5ac is a central effector of allergic inflammation that is required for airway hyperreactivity (AHR) to methacholine (MCh). In mice bred on two well-characterized strain backgrounds (C57BL/6 and BALB/c) and exposed to two separate allergic stimuli (ovalbumin and Aspergillus extract), genetic removal of Muc5ac abolishes AHR. Residual MCh responses are identical to unchallenged controls, and although inflammation remains intact, heterogeneous mucus occlusion decreases by 74%. Thus, whereas inflammatory effects on ASM alone are insufficient for AHR, Muc5ac-mediated plugging is an essential mechanism. Inhibiting MUC5AC may be effective for treating asthma and other lung diseases where it is also overproduced.

Airway mucus is essential for lung defense, but excessive mucus in asthma obstructs airflow, leading to severe and potentially fatal outcomes. Current asthma treatments have minimal effects on mucus, and the lack of therapeutic options stems from a poor understanding of mucus function and dysfunction at a molecular level and in vivo. Biophysical properties of mucus are controlled by mucin glycoproteins that polymerize covalently via disulfide bonds. Once secreted, mucin glycopolymers can aggregate, form plugs, and block airflow. Here we show that reducing mucin disulfide bonds disrupts mucus in human asthmatics and reverses pathological effects of mucus hypersecretion in a mouse allergic asthma model. In mice, inhaled mucolytic treatment loosens mucus mesh, enhances mucociliary clearance, and abolishes airway hyperreactivity (AHR) to the bronchoprovocative agent methacholine. AHR reversal is directly related to reduced mucus plugging. These findings establish grounds for developing treatments to inhibit effects of mucus hypersecretion in asthma.

VDR activity even in subjects with asthma and with IL-13, highlighting retained functionality. Expression of Class I histone deacetylases 1-3 (HDAC) and overall HDAC activity were lower in IL-13-exposed ASM, but calcitriol enhanced HDAC expression/activity. Conclusions:In asthmatic ASM, Vit D functionality is maintained, allowing calcitriol to reduce the procontractile and proremodeling effects of inflammatory cytokines, particularly IL-13, which is relevant to asthma. These findings highlight a potential role for Vit D in asthma pathogenesis, particularly in the context of airway structure and functional changes early in disease.

Airway mucus is essential for healthy lung defense 1 , but excessive mucus in asthma obstructs airflow, leading to severe and potentially fatal outcomes 2-5 . Current asthma therapies reduce allergic inflammation and relax airway smooth muscle, but treatments are often inadequate due to their minimal effects on mucus obstruction 6,7 . The lack of efficacious mucus-targeted treatments stems from a poor understanding of healthy mucus function and pathological mucus dysfunction at a molecular level. The chief macromolecules in mucus, polymeric mucins, are massive glycoproteins whose sizes and biophysical properties are dictated in part by covalent disulfide bonds that link mucin molecules into assemblies of 10 or more subunits 8 . Once secreted, mucin glycopolymers can aggregate to form plugs that block airflow. Here we show that reducing mucin disulfide bonds depolymerizes mucus in human asthma and reverses pathological effects of mucus hypersecretion in a mouse allergic asthma model. In mice challenged with a fungal allergen, inhaled mucolytic treatment acutely loosened mucus mesh, enhanced mucociliary clearance (MCC), and abolished airway hyperreactivity (AHR) to the bronchoprovocative agent methacholine. AHR reversal was directly related to reduced mucus plugging. Furthermore, protection in mucolytic treated mice was identical to prevention observed in mice lacking Muc5ac, the polymeric mucin required for allergic AHR in murine models 9 . These findings establish grounds for developing novel fast-acting agents to treat mucus hypersecretion in asthma 10,11 . Efficacious mucolytic therapies could be used to directly improve airflow, help resolve inflammation, and enhance the effects of inhaled treatments for asthma and other respiratory conditions 11,12 .With daily exposures to >8,000 liters of air containing billions of particles and potential pathogens, respiratory tissues embody the need for robust host defense. Airway mucus is critical for protection, but poor control of mucus function is central to numerous lung diseases. In patients who die during asthma exacerbations, mucus obstruction is a feature long-recognized by pathologists 13 , with plugging observed in >90% of cases and thus considered a major cause of fatal obstruction 5 . Mucus obstruction is also prominent in non-fatal cases of severe asthma 2 , but effective mucolytic therapies are lacking.

In the lungs, the epithelium is a first line of innate defense. In acute settings, such as infection or particulate exposure, the epithelium is protective. Protection is conferred by the epithelium's role as a physical barrier and by its ability to synthesize proteins that promote defense directly through physical interactions (e.g., mucins and anti-microbial peptides) and indirectly through the production of proteins that regulate inflammation (e.g., cytokines and chemokines). Despite its importance as a first line of host defense, the epithelium is also a significant target and an effector in lung pathologies. Accordingly, to determine the significance and biological mechanisms of genes involved in pulmonary defense, it is important to be able to interrogate the lung epithelium. In mice, this presents challenges related to the cellular location and timing of interventions. Effective genetic strategies for targeting the lung epithelium using tissue-/cell-specific and inducible control have been developed over the past decade. Methods for spatiotemporal targeting of gene expression are described here.