Erythromycin increases bactericidal activity of surface liquid in human airway epithelial cells

2005 ◽  
Vol 289 (4) ◽  
pp. L565-L573 ◽  
Author(s):  
Kota Ishizawa ◽  
Tomoko Suzuki ◽  
Mutsuo Yamaya ◽  
Yu Xia Jia ◽  
Seiichi Kobayashi ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Bactericidal Activity ◽  
Culture Medium ◽  
Airway Epithelial Cells ◽  
Airway Surface Liquid ◽  
Airway Epithelial ◽  
Tracheal Epithelial Cells ◽  
Tracheal Epithelial ◽  
Surface Liquid ◽  
Number Of Bacteria

Macrolide antibiotics have clinical benefits in patients with diffuse panbronchiolitis and in patients with cystic fibrosis. Although many mechanisms have been proposed, the precise mechanisms are still uncertain. We examined the effects of erythromycin on bactericidal activity of airway surface liquid secreted by cultured human tracheal epithelial cells. Airway surface liquid was collected by washing the surface of human tracheal epithelial cells with a sodium solution (40 meq/l). Methicillin-resistant Staphylococcus aureus and Pseudomonas aeruginosa were incubated with airway surface liquid, and the number of surviving bacteria was examined. The number of bacteria in airway surface liquid from the cells cultured in medium alone was significantly lower than that in the sodium solution. Furthermore, the number of bacteria in airway surface liquid from the cells treated with erythromycin was significantly lower than that in airway surface liquid from the cells treated with solvent alone. The production of mRNA and protein of human β-defensin-1 and human β-defensin-2 was significantly increased by erythromycin. Bactericidal activity of airway surface liquid was observed at low concentrations (40 meq/l) of sodium but not at higher concentrations (≥80 meq/l). Airway surface liquid did not contain significant amounts of antibiotics supplemented in the culture medium. Erythromycin at the levels in airway surface liquid and in culture medium did not inhibit bacterial growth. These results suggest that erythromycin may increase bactericidal activity of airway surface liquid in human airway epithelial cells through human β-defensins production and reduce susceptibility of the airway to bacterial infection.

Paraoxonase-2 deficiency enhancesPseudomonas aeruginosaquorum sensing in murine tracheal epithelia

2007 ◽  
Vol 292 (4) ◽  
pp. L852-L860 ◽  
Author(s):  
David A. Stoltz ◽  
Egon A. Ozer ◽  
Carey J. Ng ◽  
Janet M. Yu ◽  
Srinivasa T. Reddy ◽  
...  
Keyword(s):  
Epithelial Cell ◽  
Epithelial Cells ◽  
Quorum Sensing ◽  
Airway Epithelial Cell ◽  
Airway Epithelial Cells ◽  
Airway Epithelial ◽  
Airway Epithelia ◽  
Tracheal Epithelial Cells ◽  
Human Airway ◽  
Tracheal Epithelial

Pseudomonas aeruginosa is an important cause of nosocomial infections and is frequently present in the airways of cystic fibrosis patients. Quorum sensing mediates P. aeruginosa's virulence and biofilm formation through density-dependent interbacterial signaling with autoinducers. N-3-oxododecanoyl homoserine lactone (3OC12-HSL) is the major autoinducer in P. aeruginosa. We have previously shown that human airway epithelia and paraoxonases (PONs) degrade 3OC12-HSL. This study investigated the role of PON1, PON2, and PON3 in airway epithelial cell inactivation of 3OC12-HSL. All three PONs were present in murine tracheal epithelial cells, with PON2 and PON3 expressed at the highest levels. Lysates of tracheal epithelial cells from PON2, but not PON1 or PON3, knockout mice had impaired 3OC12-HSL inactivation compared with wild-type mice. In contrast, PON1-, PON2-, or PON3-targeted deletions did not affect 3OC12-HSL degradation by intact epithelia. Overexpression of PON2 enhanced 3OC12-HSL degradation by human airway epithelial cell lysates but not by intact epithelia. Finally, using a quorum-sensing reporter strain of P. aeruginosa, we found that quorum sensing was enhanced in PON2-deficient airway epithelia. In summary, these results show that loss of PON2 impairs 3OC12-HSL degradation by airway epithelial cells and suggests that diffusion of 3OC12-HSL into the airway cells can be the rate-limiting step for degradation of the molecule.

scholarly journals Airway Surface Liquid Osmolality Measured Using Fluorophore-Encapsulated Liposomes

2001 ◽  
Vol 117 (5) ◽  
pp. 423-430 ◽  
Author(s):  
Sujatha Jayaraman ◽  
Yuanlin Song ◽  
A.S. Verkman
Keyword(s):  
Epithelial Cells ◽  
Water Loss ◽  
Airway Epithelial Cells ◽  
Evaporative Water Loss ◽  
Airway Surface Liquid ◽  
Airway Epithelial ◽  
Evaporative Water ◽  
Dry Air ◽  
Surface Liquid ◽  
Sulforhodamine 101

The airway surface liquid (ASL) is the thin layer of fluid coating the luminal surface of airway epithelial cells at an air interface. Its composition and osmolality are thought to be important in normal airway physiology and in airway diseases such as asthma and cystic fibrosis. The determinants of ASL osmolality include epithelial cell solute and water transport properties, evaporative water loss, and the composition of secreted fluids. We developed a noninvasive approach to measure ASL osmolality using osmotically sensitive 400-nm-diam liposomes composed of phosphatidylcholine/cholesterol/polyethylene glycol-phosphatidylcholine (1:0.3:0.08 molar ratio). Calcein was encapsulated in the liposomes at self-quenching concentrations (30 mM) as a volume-sensitive marker, together with sulforhodamine 101 (2 mM) as a volume-insensitive reference. Liposome calcein/sulforhodamine 101 fluorescence ratios responded rapidly (<0.2 s) and stably to changes in solution osmolality. ASL osmolality was determined from calcein/sulforhodamine 101 fluorescence ratios after addition of microliter quantities of liposome suspensions to the ASL. In bovine airway epithelial cells cultured on porous supports at an air–liquid interface, ASL thickness (by confocal microscopy) was 22 μm and osmolality was 325 ± 12 mOsm. In anesthetized mice in which a transparent window was created in the trachea, ASL thickness was 55 μm and osmolality was 330 ± 36 mOsm. ASL osmolality was not affected by pharmacological inhibition of CFTR in airway cell cultures or by genetic deletion of CFTR in knockout mice. ASL osmolality could be increased substantially to >400 mOsm by exposure of the epithelium to dry air; the data were modeled mathematically using measured rates of osmosis and evaporative water loss. These results establish a ratio imaging method to map osmolality in biological compartments. ASL fluid is approximately isosmolar under normal physiological conditions, but can become hyperosmolar when exposed to dry air, which may induce cough and airway reactivity in some patients.

scholarly journals Acid ceramidase rescues cystic fibrosis mice from pulmonary infections

2020 ◽  
Author(s):  
Katrin Anne Becker ◽  
Rabea Verhaegh ◽  
Hedda-Luise Verhasselt ◽  
Simone Keitsch ◽  
Matthias Soddemann ◽  
...  
Keyword(s):  
Cystic Fibrosis ◽  
Epithelial Cells ◽  
Pathogenic Bacteria ◽  
Airway Epithelial Cells ◽  
Airway Epithelial ◽  
Pulmonary Infections ◽  
Acid Ceramidase ◽  
Tracheal Epithelial Cells ◽  
Infection Susceptibility ◽  
Tracheal Epithelial

Previous studies have shown that sphingosine kills a variety of pathogenic bacteria, including Pseudomonas aeruginosa (P. aeruginosa) and Staphylococcus aureus. Sphingosine concentrations are decreased in airway epithelial cells of cystic fibrosis (CF) mice and this defect has been linked to the infection susceptibility of these mice. Here, we tested whether genetic overexpression of the acid ceramidase rescues cystic fibrosis mice from pulmonary infections with P. aeruginosa. We demonstrate that transgenic overexpression of the acid ceramidase in CF mice corresponds to an overexpression of the acid ceramidase in bronchial and tracheal epithelial cells and normalizes ceramide and sphingosine levels in bronchial and tracheal epithelial cells. In addition, expression of β1-integrin, which is ectopically expressed on the luminal surface of airway epithelial cells in cystic fibrosis mice - an alteration that is very important for mediating pulmonary P. aeruginosa infections of cystic fibrosis, is normalized in cystic fibrosis airways upon overexpression of acid ceramidase. Most importantly, overexpression of acid ceramidase protects cystic fibrosis mice from pulmonary P. aeruginosa infections. Infection of CF mice or CF mice that were inhaled with sphingosine with P. aeruginosa or a P. aeruginosa mutant that is resistant to sphingosine indicate that sphingosine and not a metabolite kills P. aeruginosa upon pulmonary infection. These studies further support the use of acid ceramidase and its metabolite sphingosine as a potential treatment of cystic fibrosis.

scholarly journals Connexin mimetic peptides reversibly inhibit Ca2+signaling through gap junctions in airway cells

2000 ◽  
Vol 279 (4) ◽  
pp. L623-L630 ◽  
Author(s):  
Scott Boitano ◽  
W. Howard Evans
Keyword(s):  
Epithelial Cells ◽  
Gap Junctions ◽  
Airway Epithelial Cells ◽  
Airway Epithelial ◽  
Gap Junctional Communication ◽  
Tracheal Epithelial Cells ◽  
Junctional Communication ◽  
Mimetic Peptides ◽  
Intracellular Ca ◽  
Tracheal Epithelial

The effect of peptides with sequences derived from connexins, the constituent proteins of gap junctions, on mechanically stimulated intercellular Ca2+signaling in tracheal airway epithelial cells was studied. Three peptides with sequences corresponding to connexin extracellular loop regions reversibly restricted propagation of Ca2+ waves to neighboring cells. Recovery of communication began within 10 min of removal of the peptides, with inhibition totally reversed by 20–40 min. The peptides were shown to be more effective in inhibiting Ca2+ waves than glycyrrhetinic acid or oleamide. Inhibition of intercellular Ca2+ waves by connexin mimetic peptides did not affect the Ca2+ response to extracellular ATP. Although the intracellular Ca2+ response of tracheal epithelial cells to ATP was greatly reduced by either pretreatment with high doses of ATP or application of apyrase, mechanically stimulated intercellular Ca2+ signaling was not affected by these agents. We conclude that connexin mimetic peptides are effective and reversible inhibitors of gap junctional communication of physiologically significant molecules that underlie Ca2+wave propagation in tracheal epithelial cells and propose a potential mechanism for the mode of action of mimetic peptides.

scholarly journals Functional Apical Large Conductance, Ca2+-activated, and Voltage-dependent K+ Channels Are Required for Maintenance of Airway Surface Liquid Volume

2011 ◽  
Vol 286 (22) ◽  
pp. 19830-19839 ◽  
Author(s):  
Dahis Manzanares ◽  
Carlos Gonzalez ◽  
Pedro Ivonnet ◽  
Ren-Shiang Chen ◽  
Monica Valencia-Gattas ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Ion Transport ◽  
Apical Membrane ◽  
Airway Epithelial Cells ◽  
Bk Channels ◽  
Liquid Volume ◽  
Airway Surface Liquid ◽  
Airway Epithelial ◽  
Voltage Dependent ◽  
Surface Liquid

Large conductance, Ca2+-activated, and voltage-dependent K+ (BK) channels control a variety of physiological processes in nervous, muscular, and renal epithelial tissues. In bronchial airway epithelia, extracellular ATP-mediated, apical increases in intracellular Ca2+ are important signals for ion movement through the apical membrane and regulation of water secretion. Although other, mainly basolaterally expressed K+ channels are recognized as modulators of ion transport in airway epithelial cells, the role of BK in this process, especially as a regulator of airway surface liquid volume, has not been examined. Using patch clamp and Ussing chamber approaches, this study reveals that BK channels are present and functional at the apical membrane of airway epithelial cells. BK channels open in response to ATP stimulation at the apical membrane and allow K+ flux to the airway surface liquid, whereas no functional BK channels were found basolaterally. Ion transport modeling supports the notion that apically expressed BK channels are part of an apical loop current, favoring apical Cl− efflux. Importantly, apical BK channels were found to be critical for the maintenance of adequate airway surface liquid volume because continuous inhibition of BK channels or knockdown of KCNMA1, the gene coding for the BK α subunit (KCNMA1), lead to airway surface dehydration and thus periciliary fluid height collapse revealed by low ciliary beat frequency that could be fully rescued by addition of apical fluid. Thus, apical BK channels play an important, previously unrecognized role in maintaining adequate airway surface hydration.

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