Ion the Prize: Defining the Complexities of Airway Epithelial Cell Ion Transport Functions.
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ABSTRACT:
SUBMITTER: Nguyen JP
PROVIDER: S-EPMC6543751 | biostudies-literature | 2019 Jun
REPOSITORIES: biostudies-literature
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Ion the Prize: Defining the Complexities of Airway Epithelial Cell Ion Transport Functions.
Nguyen Jenny P JP Hirota Jeremy A JA
American journal of respiratory cell and molecular biology 20190601 6
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Project description:Smoke inhalation injury is the leading cause of death in firefighters and victims. Inhaled hot air and toxic smoke are the predominant hazards to the respiratory epithelium. We aimed to analyze the effects of thermal stress and smoke aldehyde on the permeability of the airway epithelial barrier. Transepithelial resistance (RTE) and short-circuit current (ISC) of mouse tracheal epithelial monolayers were digitized by an Ussing chamber setup. Zonula occludens-1 tight junctions were visualized under confocal microscopy. A cell viability test and fluorescein isothiocyanate-dextran assay were performed. Thermal stress (40 °C) decreased RTE in a two-phase manner. Meanwhile, thermal stress increased ISC followed by its decline. Na+ depletion, amiloride (an inhibitor for epithelial Na+ channels [ENaCs]), ouabain (a blocker for Na+/K+-ATPase), and CFTRinh-172 (a blocker of cystic fibrosis transmembrane regulator [CFTR]) altered the responses of RTE and ISC to thermal stress. Steady-state 40 °C increased activity of ENaCs, Na+/K+-ATPase, and CFTR. Acrolein, one of the main oxidative unsaturated aldehydes in fire smoke, eliminated RTE and ISC. Na+ depletion, amiloride, ouabain, and CFTRinh-172 suppressed acrolein-sensitive ISC, but showed activating effects on acrolein-sensitive RTE. Thermal stress or acrolein disrupted zonula occludens-1 tight junctions, increased fluorescein isothiocyanate-dextran permeability but did not cause cell death or detachment. The synergistic effects of thermal stress and acrolein exacerbated the damage to monolayers. In conclusion, the paracellular pathway mediated by the tight junctions and the transcellular pathway mediated by active and passive ion transport pathways contribute to impairment of the airway epithelial barrier caused by thermal stress and acrolein. Graphical abstract Thermal stress and acrolein are two essential determinants for smoke inhalation injury, impairing airway epithelial barrier. Transcellular ion transport pathways via the ENaC, CFTR, and Na/K-ATPase are interrupted by both thermal stress and acrolein, one of the most potent smoke toxins. Heat and acrolein damage the integrity of the airway epithelium through suppressing and relocating the tight junctions.
| S-EPMC7669642 | biostudies-literature
Project description:Quantitation of CFTR function in vitro is commonly performed by acutely stimulating then inhibiting ion transport through CFTR and measuring the resulting changes in transepithelial voltage (Vte) and current (ISC). While this technique is suitable for measuring the maximum functional capacity of CFTR, it may not provide an accurate estimate of in vivo CFTR activity. To test if CFTR-mediated ion transport could be measured in the absence of acute CFTR stimulation, primary airway epithelia were analyzed in an Ussing chamber with treatment of amiloride followed by CFTR(inh)-172 without acute activation of CFTR. Non-CF epithelia demonstrated a decrease in Vte and ISC following exposure to CFTR(inh)-172 and in the absence of forskolin/IBMX (F/I); this decrease is interpreted as a measure of spontaneous CFTR activity present in these epithelia. In F508del/F508del CFTR epithelia, F/I-induced changes in Vte and ISC were ~ fourfold increased after treatment with VX-809/VX-770, while the magnitude of spontaneous CFTR activities were only ~ 1.6-fold increased after VX-809/VX-770 treatment. Method-dependent discrepancies in the responses of other CF epithelia to modulator treatments were observed. These results serve as a proof of concept for the analysis of CFTR modulator responses in vitro in the absence of acute CFTR activation. Future studies will determine the usefulness of this approach in the development of novel CFTR modulator therapies.
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Project description:Inhalation of butter flavoring by workers in the microwave popcorn industry may result in “popcorn workers' lung.” In previous in vivo studies rats exposed for 6 h to vapor from the flavoring agents, diacetyl and 2,3-pentanedione, acquired flavoring concentration-dependent damage of the upper airway epithelium and airway hyporeactivity to inhaled methacholine. Because ion transport is essential for lung fluid balance,we hypothesized that alterations in ion transport may be an early manifestation of butter flavoring-induced toxicity.We developed a system to expose cultured human bronchial/tracheal epithelial cells (NHBEs) to flavoring vapors. NHBEs were exposed for 6 h to diacetyl or 2,3-pentanedione vapors (25 or ≥ 60 ppm) and the effects on short circuit current and transepithelial resistance (Rt) were measured. Immediately after exposure to 25 ppm both flavorings reduced Na+ transport,without affecting Cl- transport or Na+,K+-pump activity. Rt was unaffected. Na+ transport recovered 18 h after exposure. Concentrations (100-360 ppm) of diacetyl and 2,3-pentanedione reported earlier to give rise in vivo to epithelial damage, and 60 ppm, caused death of NHBEs 0 h post-exposure. Analysis of the basolateral medium indicated that NHBEs metabolize diacetyl and 2,3-pentanedione to acetoin and 2-hydroxy-3-pentanone, respectively. The results indicate that ion transport is inhibited transiently in airway epithelial cells by lower concentrations of the flavorings than those that result in morphological changes of the cells in vivo or in vitro.
| S-EPMC4739639 | biostudies-literature
Project description:PurposeWe sought to identify key variables in cellular architecture and physiology that might explain observed differences in the passive transport properties of small molecule drugs across different airway epithelial cell types.MethodsPropranolol (PR) was selected as a weakly basic, model compound to compare the transport properties of primary (NHBE) vs. tumor-derived (Calu-3) cells. Differentiated on Transwell™ inserts, the architecture of pure vs. mixed cell co-cultures was studied with confocal microscopy followed by quantitative morphometric analysis. Cellular pharmacokinetic modeling was used to identify parameters that differentially affect PR uptake and transport across these two cell types.ResultsPure Calu-3 and NHBE cells possessed different structural and functional properties. Nevertheless, mixed Calu-3 and NHBE cell co-cultures differentiated as stable cell monolayers. After measuring the total mass of PR, the fractional areas covered by Calu-3 and NHBE cells allowed deconvoluting the transport properties of each cell type. Based on the apparent thickness of the unstirred, cell surface aqueous layer, local differences in the extracellular microenvironment explained the measured variations in passive PR uptake and permeation between Calu-3 and NHBE cells.ConclusionMixed cell co-cultures can be used to compare the local effects of the extracellular microenvironment on drug uptake and transport across two epithelial cell types.
| S-EPMC3706189 | biostudies-literature
Project description:Cigarette smoking is associated with chronic obstructive pulmonary disease and chronic bronchitis. Acquired ion transport abnormalities, including cystic fibrosis transmembrane conductance regulator (CFTR) dysfunction, caused by cigarette smoking have been proposed as potential mechanisms for mucus obstruction in chronic bronchitis. Although e-cigarette use is popular and perceived to be safe, whether it harms the airways via mechanisms altering ion transport remains unclear. In the present study, we sought to determine if e-cigarette vapor, like cigarette smoke, has the potential to induce acquired CFTR dysfunction, and to what degree. Electrophysiological methods demonstrated reduced chloride transport caused by vaporized e-cigarette liquid or vegetable glycerin at various exposures (30 min, 57.2% and 14.4% respectively, vs. control; P < 0.0001), but not by unvaporized liquid (60 min, 17.6% vs. untreated), indicating that thermal degradation of these products is required to induce the observed defects. We also observed reduced ATP-dependent responses (-10.8 ± 3.0 vs. -18.8 ± 5.1 μA/cm2 control) and epithelial sodium channel activity (95.8% reduction) in primary human bronchial epithelial cells after 5 minutes, suggesting that exposures dramatically inhibit epithelial ion transport beyond CFTR, even without diminished transepithelial resistance or cytotoxicity. Vaporizing e-cigarette liquid produced reactive aldehydes, including acrolein (shown to induce acquired CFTR dysfunction), as quantified by mass spectrometry, demonstrating that respiratory toxicants in cigarette smoke can also be found in e-cigarette vapor (30 min air, 224.5 ± 15.99; unvaporized liquid, 284.8 ± 35.03; vapor, 54,468 ± 3,908 ng/ml; P < 0.0001). E-cigarettes can induce ion channel dysfunction in airway epithelial cells, partly through acrolein production. These findings indicate a heretofore unknown toxicity of e-cigarette use known to be associated with chronic bronchitis onset and progression, as well as with chronic obstructive pulmonary disease severity.
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Project description:Cystic fibrosis (CF) is a life-limiting genetic disorder caused by loss-of-function mutations in the gene which codes for the CF transmembrane conductance regulator (CFTR) Cl- channel. Loss of Cl- secretion across the apical membrane of airway lining epithelial cells results in dehydration of the airway surface liquid (ASL) layer which impairs mucociliary clearance (MCC), and as a consequence promotes bacterial infection and inflammation of the airways. Interventions that restore airway hydration are known to improve MCC. Here we review the ion channels present at the luminal surface of airway epithelial cells that may be targeted to improve airway hydration and MCC in CF airways.
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Project description:Mucociliary clearance (MCC) in CF lung disease is limited by airway dehydration, leading to persistent bacterial infection and inflammation in the airways. Agents designed to rehydrate the airway mucosa lead to improved MCC. Hyperosmolar agents, such as hypertonic saline and mannitol, create a luminal osmotic gradient, drawing water into the dehydrated ASL. Ion transport modulators function to activate alternative chloride channels and/or to block sodium hyperabsorption that occurs through a dysregulated ENaC channel. Combinations of these therapies may result in a synergistic improvement in airway hydration, and thus, restore MCC. Active ongoing phase II and III trials of new pharmacotherapeutics are covered in this review.
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Project description: Not available
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