Airway Epithelial Heterogeneity and Mucus Plugging in Asthmatic Bronchioles

Rationale: Bronchiolar dysfunction is associated with asthma exacerbations and poor symptom control. However, the molecular pathophysiology of asthmatic bronchiolar disease is poorly defined. Objectives: Test the hypothesis that asthmatic bronchioles exhibit disturbances in epithelial biology that produce MUC5AC-dominated mucus plugs. Methods: Peripheral lung tissues from severe asthmatics, fatal asthmatics (FA), and controls were evaluated with histology, RNA in situ hybridization, and immunohistochemistry. Isolated bronchiolar and bronchial basal cell responses to IL13 were compared in culture. Spatial transcriptomics and multiplex immunophenotyping were performed on excised tissue sections. Measurements and Main Results: In excised tissues, severe and FA bronchiolar epithelia, depleted of distal airway secretory cells (DASCs) and enriched in MUC5AC goblet cells, circumscribed MUC5AC-dominated mucus plugs. In cultured bronchiolar basal cells, IL13 suppressed FOXA2 and DASC gene signatures and upregulated MUC5AC expression. Additional studies in severe and FA excised tissues demonstrated that bronchiolar epithelia were populated by MUC5AC-expressing goblet cell niches heterogeneously distributed within single segments and, indeed, individual bronchioles. Spatial transcriptomics and immuno-proteomics of these MUC5AC-expressing bronchiolar niches identified increased goblet, suprabasal (SERPINB3), and basal cell, juxtaposed to a loss of DASC, gene signatures. MUC5AC-high niche bronchiolar basal cells expressed reduced FOXA2 and elevated type-2 inflammatory (T2) gene signatures. Immune cell distributions surrounding asthmatic bronchioles differed from controls but did not correlate with MUC5AC-high niches. Conclusions: Asthmatic bronchioles exhibit a T2-driven proximalization associated with mucus plugging. MUC5AC-high niches were identified heterogeneously in bronchiolar epithelia independent of immune cell localizations, suggesting asthmatic bronchioles contain cellular niches which perpetuate T2-initiated epithelial remodeling.