MiR449 protects airway cilia and healthy lung ageing

Abstract

Airway cilia dysfunction, either acquired or inborn, results in respiratory diseases e.g. chronic bronchitis. The miR449 cluster is strongly expressed in airway epithelia. Here we show that miR449 knockout (KO) mice develop spontaneous COPD as do wildtypes exposed to cigarette smoke for 6 months, with increased BAL and M2A macrophages. Bacterial challenge leads to increased MMP9 and MMP12 in miR449KO, thus explaining matrix degradation. However, young, naive miR449KO mice do not display any sign of emphysema suggesting that it is a secondary phenotype. miR449KO mice display reduced mucociliary clearance and cilia ultrastructure defects already at young age, which develop into reduced cilia in ageing mice, hinting at defective maintenance of ciliary structures. We thus assessed ciliated regeneration of lower and upper respiratory epithelia respectively by applying naphthalene to monitor regeneration in vivo and isolating tracheal basal cells to monitor differentiation in vitro. Both models resulted in shorter cilia and less ciliated cells in miR449KO. Epithelial regeneration requires a balance between proliferation and differentiation, and this is coupled to ciliary disassembly and re-assembly. Surprisingly, ciliary assembly was unchanged but ciliary disassembly significantly increased in miR449KO. Bioinformatics and omics showed that miR449 indeed directly targets the ciliary disassembly pathway at several nodes: 1) the potassium channel regulating calcium influx encoded by KCNH1, 2) the calcium-sensitive Aurora kinase A, and 3) the cilia tubulin deacetylase HDAC6. As microRNAs usually fine-tunes pathways by hitting them several times, this strongly suggests that miR449 protects airway cilia by reducing ciliary disassembly.