Cystic fibrosis airway secretions exhibit mucin hyperconcentration and increased osmotic pressure
Ashley G. Henderson, … , Richard C. Boucher, Mehmet Kesimer
Ashley G. Henderson, … , Richard C. Boucher, Mehmet Kesimer
Published June 2, 2014
Citation Information: J Clin Invest. 2014;124(7):3047-3060. https://doi.org/10.1172/JCI73469.
View: Text | PDF
Research Article Pulmonology

Cystic fibrosis airway secretions exhibit mucin hyperconcentration and increased osmotic pressure

Abstract

The pathogenesis of mucoinfective lung disease in cystic fibrosis (CF) patients likely involves poor mucus clearance. A recent model of mucus clearance predicts that mucus flow depends on the relative mucin concentration of the mucus layer compared with that of the periciliary layer; however, mucin concentrations have been difficult to measure in CF secretions. Here, we have shown that the concentration of mucin in CF sputum is low when measured by immunologically based techniques, and mass spectrometric analyses of CF mucins revealed mucin cleavage at antibody recognition sites. Using physical size exclusion chromatography/differential refractometry (SEC/dRI) techniques, we determined that mucin concentrations in CF secretions were higher than those in normal secretions. Measurements of partial osmotic pressures revealed that the partial osmotic pressure of CF sputum and the retained mucus in excised CF lungs were substantially greater than the partial osmotic pressure of normal secretions. Our data reveal that mucin concentration cannot be accurately measured immunologically in proteolytically active CF secretions; mucins are hyperconcentrated in CF secretions; and CF secretion osmotic pressures predict mucus layer–dependent osmotic compression of the periciliary liquid layer in CF lungs. Consequently, mucin hypersecretion likely produces mucus stasis, which contributes to key infectious and inflammatory components of CF lung disease.

Authors

Ashley G. Henderson, Camille Ehre, Brian Button, Lubna H. Abdullah, Li-Heng Cai, Margaret W. Leigh, Genevieve C. DeMaria, Hiro Matsui, Scott H. Donaldson, C. William Davis, John K. Sheehan, Richard C. Boucher, Mehmet Kesimer

×

Figure 3

The effect of proteases on mucin epitopes and polymeric structure.

Options: View larger image (or click on image) Download as PowerPoint
The effect of proteases on mucin epitopes and polymeric structure. (A) I...
(A) Isolated MUC5B (100 μg) was incubated with either trypsin (1 μg) or elastase (1 μg) for increasing periods of time (5 minutes to 4 hours). Unreduced aliquots of the samples were subjected to agarose gel electrophoresis and probed with MAN5BIII or EU-MUC5Ba antibodies. MAN5BIII immunoreactivity was completely abolished by the NE within 5 minutes, while trypsin treatment substantially reduced the antibody reactivity. EU-MUC5Ba immunoreactivity was lost within 20 minutes of exposure to both trypsin and elastase. (B) An aliquot from the NE-treated samples was subjected to SEC/MALS/dRI measurements using a Sepharose CL-2B (2 × 5 ml) column to determine molecular weight (dotted lines) and sample concentrations (solid lines), respectively. The total mucin mass under the curve was slightly decreased from 28.4 μg (blue line, PBS control) to 25.6 μg after a 20-minute elastase incubation (green line) and to 24.2 μg after a 60-minute elastase incubation (magenta line). Molecular mass measurements were plotted across the mucin peak: control mucins had an average molecular mass of approximately 38 MDa (blue dotted line) and had an average molecular mass of approximately 44 MDa after 20 (green dotted line) and 60 (magenta dotted line) minutes of elastase incubation, indicating a slight increase in molecular weight.