Submucosal gland secretions in airways from cystic fibrosis patients have normal [Na⁺] and pH but elevated viscosity

Abstract

ASL ͉ chloride ͉ trachea ͉ ratio imaging T he air-facing surface of respiratory airways is lined by a layer of surface epithelial cells whose mucosa is bathed by a thin film of fluid called the airway surface liquid (ASL) (1-5). The airways also contain submucosal glands that secrete fluid and macromolecules onto the ASL (6-9). The submucosal glands contain serous tubules and acini that secrete salt, water, and various antimicrobial proteins. The serous secretions pass through mucous tubules, where viscous glycoproteins are added, and then into a collecting duct and onto the airway surface. Active salt and water secretion by serous epithelial cells is believed to involve Cl Ϫ transport by the cystic fibrosis transmembrane conductance regulator (CFTR) protein (10-12) driving water transport through AQP4 and AQP5 water channels (13-15). Submucosal gland secretions are proposed to be important for generation of ASL fluid and for creation of an environment that inhibits bacterial colonization.Abnormalities in submucosal gland secretions have been proposed to contribute to the airway pathophysiology in cystic fibrosis (CF). CFTR is expressed in serous epithelial cells of submucosal glands more strongly than in other tissues of the airways and lung (16,17 ], pH and viscosity of freshly secreted fluid from human submucosal airway glands to determine: (i) whether the composition of fluid secreted from glands is similar to that of the ASL, and (ii) whether gland fluid composition differs in normal vs. CF human airways. We recently developed fluorescent probes and ratio imaging microscopy methods to measure [Na MethodsAirway Preparations. Fragments of normal and CF human airways were obtained after lung transplantation and consisted of scrap airway trimmings from normal donor lungs and larger tissue fragments from discarded CF lungs. Tissues were placed in cold Physiosol (Abbott) within 30 min after removal for transport to the laboratory and initial dissection. Tissues were then transferred to ice-cold HCO 3 Ϫ -buffered Krebs solution (see below) and continuously gassed with 95% O 2 and 5% CO 2 . Tissues were cut into rectangles of Ϸ2 cm 2 , and a 1.5-to 2-mm thick layer containing the mucosa and submucosa was dissected away from the underlying cartilage. Two different procedures were used to mount the tissues. In the first procedure, tissues were placed over a wire mesh in a tissue culture well with HCO 3 Ϫ -containing media bathing the serosa and left in a cell culture incubator for 30 min. The airway fragment was then mounted by using pins on a sponge soaked in HCO 3 Ϫ -containing Kreb's buffer (120 mM NaCl͞25 mM NaHCO 3 ͞3.3 mM KH 2 PO 4 ͞0.8 mM K 2 HPO 4 ͞1.2 mM MgCl 2 ͞1.2 mM CaCl 2 ͞10 mM glucose, pH 7.4) (on the serosal side) and held in a perfusion chamber with This paper was submitted directly (Track II) to the PNAS office.Abbreviations: CF, cystic fibrosis; ASL, airway surface liquid; BCECF, 2Ј,7Ј-bis-(2-carboxyethyl)-5-(and-6)-carboxy fluorescein; CFTR, cystic fibrosis transmembrane conductance regulator. § ...

Characterization of Wild-Type and ΔF508 Cystic Fibrosis Transmembrane Regulator in Human Respiratory Epithelia

“…Our observations also appear to conflict with ex vivo studies of CF gland preparations that have shown decreased agonist-stimulated glandular secretion rates and a more viscous (increased mucin concentration) secretion, implying that CFTR is involved in regulating glandular fluid volume (Jayaraman et al, 2001;Joo et al, 2002). However, in every functional ex vivo study of normal and CF tissues, all the glandular components (e.g., acini and ducts) and adjacent gland tissue/cells (e.g., muscle cells) were exposed to the pharmacological stimulus, and glandular secretions were analyzed at the duct entrance onto the airway surface (Joo et al, 2002;Verkman et al, 2003;Ballard and Inglis, 2004).…”

Contribution of α7 nicotinic receptor to airway epithelium dysfunction under nicotine exposure

“…Similarly, elevated NaCl concentrations have been reported in CF airway fluids (19). However, it has then been shown that airway-surface liquid in CFTR-null mice is approximately isotonic (20) and that submucosal gland secretions in airways from patients with CF have normal [Na + ], although presenting elevated viscosity (21). Contrary to α7 nAChR, absence of either α5, β2, or β4 nAChR subunit does not impact on airway mucus transport in mice (Fig.…”

Novel Regulation of Cystic Fibrosis Transmembrane Conductance Regulator (CFTR) Channel Gating by External Chloride

“…It has been reported that CF-affected epithelia, particularly in the gastrointestinal tract, secrete fluid with a more acidic pH than normal epithelia (11,12). Whether this is also true for airway surface liquid is still controversial (13,14), but recent measurements of the pH and HCO 3 Ϫ concentration of fluid secreted by polarized human submucosal gland cells, Calu-3, indicate that these cells are capable of secreting substantial amounts of HCO 3 Ϫ (ϳ80 mM) under appropriate stimulation, a process that would be defective in CF (15). An acidic luminal environment affects the physical properties of mucus (16,17) and promotes bacterial binding to mucins (18,19), both of which may have important implications for CF lung disease (20).…”