Defective postsecretory maturation of MUC5B mucin in cystic fibrosis airways
Lubna H. Abdullah, Jessica R. Evans, T. Tiffany Wang, Amina A. Ford, Alexander M. Makhov, Kristine Nguyen, Raymond D. Coakley, Jack D. Griffith, C. William Davis, Stephen T. Ballard, Mehmet Kesimer
Lubna H. Abdullah, Jessica R. Evans, T. Tiffany Wang, Amina A. Ford, Alexander M. Makhov, Kristine Nguyen, Raymond D. Coakley, Jack D. Griffith, C. William Davis, Stephen T. Ballard, Mehmet Kesimer
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Research Article Pulmonology

Defective postsecretory maturation of MUC5B mucin in cystic fibrosis airways

Abstract

In cystic fibrosis (CF), airway mucus becomes thick and viscous, and its clearance from the airways is impaired. The gel-forming mucins undergo an ordered “unpacking/maturation” process after granular release that requires an optimum postsecretory environment, including hydration and pH. We hypothesized that this unpacking process is compromised in the CF lung due to abnormal transepithelial fluid transport that reduces airway surface hydration and alters ionic composition. Using human tracheobronchial epithelial cells derived from non-CF and CF donors and mucus samples from human subjects and domestic pigs, we investigated the process of postsecretory mucin unfolding/maturation, how these processes are defective in CF airways, and the probable mechanism underlying defective unfolding. First, we found that mucins released into a normal lung environment transform from a compact granular form to a linear form. Second, we demonstrated that this maturation process is defective in the CF airway environment. Finally, we demonstrated that independent of HCO3− and pH levels, airway surface dehydration was the major determinant of this abnormal unfolding process. This defective unfolding/maturation process after granular release suggests that the CF extracellular environment is ion/water depleted and likely contributes to abnormal mucus properties in CF airways prior to infection and inflammation.

Authors

Lubna H. Abdullah, Jessica R. Evans, T. Tiffany Wang, Amina A. Ford, Alexander M. Makhov, Kristine Nguyen, Raymond D. Coakley, Jack D. Griffith, C. William Davis, Stephen T. Ballard, Mehmet Kesimer

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Figure 1

Distribution of the molecular forms of MUC5B following secretion from non-CF and CF HTBE cell cultures.

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Distribution of the molecular forms of MUC5B following secretion from no...
After washing the apical surface, HTBE cells were stimulated apically with 100 μM ATPγS to induce mucin secretion. Apical secretions were collected in PBS at 2 minutes and 240 minutes after ATP challenge and immediately added to 4 M GuHCl to preserve the molecular forms of mucin. Secretions were then subjected to rate zonal centrifugation by a 6–8 M GuHCl gradient to separate the molecular forms of MUC5B. (A and B) At 2 minutes, in the ATPγS-stimulated non-CF and CF HTBE cell secretions, MUC5B has similar profiles and is found predominantly in the high-density “compact” region. (B) However, the MUC5B profile in the CF cultures shows no progressive shift from the high-density region to the low-density region, as is observed in the non-CF cultures, suggesting the MUC5B stays predominately in a compact form in the CF cultures (**P = 0.01). (C) The distribution of the molecular forms of MUC5B in “accumulated” secretions in non-CF and CF cultures. Unstimulated basal secretions that accumulated on the surface were subjected to the density gradient. The MUC5B profiles indicated that the mucins recovered from the accumulated (72 hours) mucus from non-CF cultures resolved predominately in the low-density region as the linear form, whereas the majority of the MUC5B from CF HTBE cell cultures was found in the same high-density region as the compact form (*P = 0.01). SD values are indicated by vertical bars. Unpaired, 2-tailed t test was used to determine statistical significance.