Cystic fibrosis transmembrane conductance regulator (CFTR) inhibition results in mucus accumulation in human airway epithelia Calu-3 cells: Experimental and Machine Learning Studies

Abstract

Porosomes are cup-shaped lipoprotein structures at the cell plasma membrane involved in fractional release of intra-vesicular contents during secretion. At the base of the porosome facing the cell cytoplasm, secretory vesicles dock, fuse and swell, to release intra-vesicular content during secretion. Earlier studies demonstrate the cystic fibrosis trans-membrane conductance regulator (CFTR) associated with the porosome in human airways epithelial Calu-3 mucous-secreting cells, suggesting its possible involvement in porosome-mediated mucus secretion. The current study was undertaken to test this hypothesis. Electron microscopy followed by morphometric analysis using manual and computational machine learning approaches were used to assess changes in secretory vesicle size and content, following stimulation of secretion in the absence and presence of CFTR inhibitors. Results from the study demonstrate that stimulated Calu-3 cells pre-exposed to CFTR inhibitors, demonstrate attenuation of secretory vesicle swelling and the release of mucin. Consequently, accumulation of intracellular mucin is observed in cells exposed to CFTR inhibitors. These results further suggest that mucin secretion from Calu-3 cells involve CFTR both at the secretory vesicle membrane to regulate vesicle volume and hydration, and at the porosome to facilitate mucin hydration and secretion. These new findings progress our understanding of the involvement of CFTR on mucus hydration and secretion, providing critical insights into the etiology of CF disease.