Mucins and their O-Glycans from human bronchial epithelial cell cultures

Holmén, Jessica M.; Karlsson, Niclas G.; Abdullah, Lubna H.; Randell, Scott H.; Sheehan, John K.; Hansson, Gunnar C.; Davis, C. William

doi:10.1152/ajplung.00108.2004

Cited by 80 publications

(76 citation statements)

References 57 publications

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“…3A), suggesting that the concentration of mucus, rather than the source, was the critical variable. These data are consistent with previous data demonstrating no differences in the biochemical properties between the CF vs. normal mucins, the major component of mucus (20).…”

supporting

confidence: 93%

A physical linkage between cystic fibrosis airway surface dehydration and Pseudomonas aeruginosa biofilms

Matsui¹,

Wagner

Hill³

et al. 2006

Proc. Natl. Acad. Sci. U.S.A.

217

228

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A vexing problem in cystic fibrosis (CF) pathogenesis has been to explain the high prevalence of Pseudomonas aeruginosa biofilms in CF airways. We speculated that airway surface liquid (ASL) hyperabsorption generates a concentrated airway mucus that interacts with P. aeruginosa to promote biofilms. To model CF vs. normal airway infections, normal (2.5% solids) and CF-like concentrated (8% solids) mucus were prepared, placed in flat chambers, and infected with an Ϸ5 ؋ 10 3 strain PAO1 P. aeruginosa. Although bacteria grew to 10 10 cfu/ml in both mucus concentrations, macrocolony formation was detected only in the CF-like (8% solids) mucus. Biophysical and functional measurements revealed that concentrated mucus exhibited properties that restrict bacterial motility and small molecule diffusion, resulting in high local bacterial densities with high autoinducer concentrations. These properties also rendered secondary forms of antimicrobial defense, e.g., lactoferrin, ineffective in preventing biofilm formation in a CF-like mucus environment. These data link airway surface liquid hyperabsorption to the high incidence of P. aeruginosa biofilms in CF via changes in the hydration-dependent physical-chemical properties of mucus and suggest that the thickened mucus gel model will be useful to develop therapies of P. aeruginosa biofilms in CF airways.mucus ͉ rheology C ystic fibrosis (CF) lung disease reflects the chronic bacterial infection of intrapulmonary airways with Pseudomonas aeruginosa biofilms (1, 2). P. aeruginosa biofilms have mostly been studied in flow chambers (3-7) that are good models for the biofilms that form on venous or urethral catheters under conditions of high flow rates and relatively high oxygen tensions. However, biofilms in CF airways form in thickened (concentrated) mucus gels that are relatively hypoxic and adherent to airway surfaces (see Fig. 1A) (8, 9). Recently, Sriramulu et al. (10) investigated the role autoregulators and amino acids play in the tightness of biofilm formation in a mucus stimulant of constant hydration. For the present study, we designed a culture system to investigate whether dehydration of the CF mucus environment may predispose to P. aeruginosa biofilm formation.A key aspect of the model was the selection of mucus concentrations that mimic normal and CF mucus. Although measurements of mucus concentration, i.e., the percentage of solids content, from CF subjects before infection have not been reported, estimates from cell cultures (11) and sputum (12-18) suggest that CF mucus is at least three or four times more concentrated than normal. Therefore, mucus was obtained from well differentiated human airway cultures and was isotonically concentrated (1) to produce mucus of normal (2.5% solids wt/wt) and CF-like (8%) concentrations (Fig. 1B). Aliquots of each were deposited in (i) an open chamber to model a mucus plaque adherent to CF airway surfaces with a residual lumen allowing airflow and (ii) a closed chamber to mimic a mucus plug occluding a CF airway (Fig. 1 A). P....

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supporting

confidence: 93%

A physical linkage between cystic fibrosis airway surface dehydration and Pseudomonas aeruginosa biofilms

Matsui¹,

Wagner

Hill³

et al. 2006

Proc. Natl. Acad. Sci. U.S.A.

217

228

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“…This channel has been linked to the regulation of mucus by controlling the production and/or release of secreted proteins such as Muc5ac, a major component of mucus found in the lung. 49 Recent studies have shown that by blocking the production and/or activity of mClca3 (or the human counterpart hClca1) inhibited the production of mucus, whereas overexpression of hClca1 enhanced mucus production. 42 These studies suggest that mClca3 plays a pivotal role in the production and/or regulation of mucus although the mechanics of this relationship have not been elucidated.…”

Section: Discussionmentioning

confidence: 99%

Gene Expression Profiles Reveal Increased mClca3 (Gob5) Expression and Mucin Production in a Murine Model of Asbestos-Induced Fibrogenesis

Sabo‐Attwood

Ramos-Niño

Bond

et al. 2005

The American Journal of Pathology

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To elucidate genes important in development or repair of asbestos-induced lung diseases, gene expression was examined in mice after inhalation of chrysotile asbestos for 3, 9, and 40 days. We identified changes in the expression of genes linked to proliferation (cyclin B2, CDC20, and CDC28 protein kinase regulatory subunit 2), inflammation (CCL9, CCL6, complement component 1, chitinase3-like 3, TNF superfamily member 10, and IL-1B), and matrix remodeling (MMP12, MMP3, integrin ␣X, and cathepsins K, Z, B, and S). The most highly induced gene at all time points was mclca3 (gob5), a putative calcium-activated chloride channel involved in the regulation of mucus production and/or secretion. Using histochemistry, we demonstrated accumulation of mucus and increased mClca3 protein in the bronchiolar epithelium of asbestos-exposed mice at all time points but peaking at 9 days. Cytokine levels (interleukin-1␤, interleukin-4, interleukin-6) in bronchoalveolar lavage fluid also increased at 9 days, suggesting Th2-mediated immunity may play a role in asbestosinduced mucus production. In contrast, levels of cathepsin K, a potent elastase, increased between 3 and 40 days at both the mRNA and protein levels, localizing primarily in CD45-positive leukocytes and interstitial cells. Identification of genes involved in lung injury and remodeling after asbestos exposure could aid in defining mechanisms of airborne particulate-

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“…Pediatric BAL fluid, sputum, and cell culture mucus samples were loaded at a constant volume (usually 30 μl, undiluted) onto a 0.7% agarose gel in a 1X Tris-acetate-EDTA (TAE) buffer with 1% SDS and electrophoresed in a horizontal gel apparatus as previously described (42). After electrophoresis, the gel was reduced in 10 mM DTT for 20 minutes prior to vacuum transfer onto a nitrocellulose membrane.…”

Section: Immunological and Physical Measurements Of Mucin Properties mentioning

confidence: 99%

Cystic fibrosis airway secretions exhibit mucin hyperconcentration and increased osmotic pressure

Henderson¹,

Ehré²,

Button³

et al. 2014

J. Clin. Invest.

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300

278

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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.

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Mucins and their O-Glycans from human bronchial epithelial cell cultures

Cited by 80 publications

References 57 publications

A physical linkage between cystic fibrosis airway surface dehydration and Pseudomonas aeruginosa biofilms

A physical linkage between cystic fibrosis airway surface dehydration and Pseudomonas aeruginosa biofilms

Gene Expression Profiles Reveal Increased mClca3 (Gob5) Expression and Mucin Production in a Murine Model of Asbestos-Induced Fibrogenesis

Cystic fibrosis airway secretions exhibit mucin hyperconcentration and increased osmotic pressure

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