Tracheobronchial air-liquid interface cell culture: a model for innate mucosal defense of the upper airways?

Abstract

Human tracheobronchial epithelial cells grown in air-liquid interface culture have emerged as a powerful tool for the study of airway biology. In this study, we have investigated whether this culture system produces “mucus” with a protein composition similar to that of in vivo, induced airway secretions. Previous compositional studies of mucous secretions have greatly underrepresented the contribution of mucins, which are major structural components of normal mucus. To overcome this limitation, we have used a mass spectrometry-based approach centered on prior separation of the mucins from the majority of the other proteins. Using this approach, we have compared the protein composition of apical secretions (AS) from well-differentiated primary human tracheobronchial cells grown at air-liquid interface and human tracheobronchial normal induced sputum (IS). A total of 186 proteins were identified, 134 from AS and 136 from IS; 84 proteins were common to both secretions, with host defense proteins being predominant. The epithelial mucins MUC1, MUC4, and MUC16 and the gel-forming mucins MUC5B and MUC5AC were identified in both secretions. Refractometry showed that the gel-forming mucins were the major contributors by mass to both secretions. When the composition of the IS was corrected for proteins that were most likely derived from saliva, serum, and migratory cells, there was considerable similarity between the two secretions, in particular, in the category of host defense proteins, which includes the mucins. This shows that the primary cell culture system is an important model for study of aspects of innate defense of the upper airways related specifically to mucus consisting solely of airway cell products.

Cystic fibrosis airway secretions exhibit mucin hyperconcentration and increased osmotic pressure

“…Physical and biophysical methods have been developed to measure mucin concentrations by antibody-independent approaches (35). These approaches use size-selective gel filtration techniques to isolate high molecular weight polymers in V o volumes, followed by analysis of the molecular size and concentration of mucins by light scattering and refractometry, respectively.…”

Small Airways in Asthma: Distal is Different

“…There are two different types of mucins, the membrane-tethered mucins like Muc1, Muc4, and MucC16, which contribute to the periciliary layer, and the secreted gel-forming mucins like Muc5AC and Muc5B, which form the upper gel-layer and determine the mucus viscosity [28,[35][36][37]. Whereas Muc5B is produced by both, goblet cells and the submucosal glands, Muc5AC is mainly produced by goblet cells and is widely used as marker for goblet cell metaplasia [38][39][40][41]. Furthermore, increased Muc5AC expression is induced during airway inflammation, whereas Muc5B expression is constitutive and remains unaltered [42,43].…”

Airway Mucins: “Aircraft Carriers” in the Apical Surface Fluid

“…Kesimer et al [18] identified 134 proteins from apical secretions of primary airway epithelial cells, with 84 proteins (62.7%) being common with the proteins identified in vivo human tracheobronchial sputum. Later, Ali et al [19] identified 56 proteins in a "mucin" fraction isolated under physiological conditions from primary human tracheobronchial epithelial cells grown in air/liquid interface, supporting the previous report [11] that airway mucins are tightly associated with various molecules.…”