scholarly journals E-cigarette constituents propylene glycol and vegetable glycerin decrease glucose uptake and its metabolism in airway epithelial cells in vitro

2020 ◽  
Vol 319 (6) ◽  
pp. L957-L967
Author(s):  
M. Woodall ◽  
J. Jacob ◽  
K. K. Kalsi ◽  
V. Schroeder ◽  
E. Davis ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Glucose Uptake ◽  
Cell Volume ◽  
Propylene Glycol ◽  
Glucose Transport ◽  
Airway Epithelial Cells ◽  
Apical Surface ◽  
Airway Epithelial ◽  
Solute Permeability ◽  
Short Term Exposure

Electronic nicotine delivery systems, or e-cigarettes, utilize a liquid solution that normally contains propylene glycol (PG) and vegetable glycerin (VG) to generate vapor and act as a carrier for nicotine and flavorings. Evidence indicated these “carriers” reduced growth and survival of epithelial cells including those of the airway. We hypothesized that 3% PG or PG mixed with VG (3% PG/VG, 55:45) inhibited glucose uptake in human airway epithelial cells as a first step to reducing airway cell survival. Exposure of H441 or human bronchiolar epithelial cells (HBECs) to PG and PG/VG (30–60 min) inhibited glucose uptake and mitochondrial ATP synthesis. PG/VG inhibited glycolysis. PG/VG and mannitol reduced cell volume and height of air-liquid interface cultures. Mannitol, but not PG/VG, increased phosphorylation of p38 MAPK. PG/VG reduced transepithelial electrical resistance, which was associated with increased transepithelial solute permeability. PG/VG decreased fluorescence recovery after photobleaching of green fluorescent protein-linked glucose transporters GLUT1 and GLUT10, indicating that glucose transport function was compromised. Puffing PG/VG vapor onto the apical surface of primary HBECs for 10 min to mimic the effect of e-cigarette smoking also reduced glucose transport. In conclusion, short-term exposure to PG/VG, key components of e-cigarettes, decreased glucose transport and metabolism in airway cells. We propose that this was a result of PG/VG reduced cell volume and membrane fluidity, with further consequences on epithelial barrier function. Taking these results together, we suggest these factors contribute to reduced defensive properties of the epithelium. We propose that repeated/chronic exposure to these agents are likely to contribute to airway damage in e-cigarette users.

scholarly journals Activity and inhibition of prostasin and matriptase on apical and basolateral surfaces of human airway epithelial cells

2012 ◽  
Vol 303 (2) ◽  
pp. L97-L106 ◽  
Author(s):  
Shilpa Nimishakavi ◽  
Marina Besprozvannaya ◽  
Wilfred W. Raymond ◽  
Charles S. Craik ◽  
Dieter C. Gruenert ◽  
...  
Keyword(s):  
Cystic Fibrosis ◽  
Epithelial Cells ◽  
Cell Surface ◽  
Selective Inhibition ◽  
Airway Epithelial Cells ◽  
Bronchial Epithelial Cells ◽  
Apical Surface ◽  
Airway Epithelial ◽  
Wild Type ◽  
Bronchial Epithelial

Prostasin is a membrane-anchored protease expressed in airway epithelium, where it stimulates salt and water uptake by cleaving the epithelial Na+ channel (ENaC). Prostasin is activated by another transmembrane tryptic protease, matriptase. Because ENaC-mediated dehydration contributes to cystic fibrosis (CF), prostasin and matriptase are potential therapeutic targets, but their catalytic competence on airway epithelial surfaces has been unclear. Seeking tools for exploring sites and modulation of activity, we used recombinant prostasin and matriptase to identify substrate t-butyloxycarbonyl-l-Gln-Ala-Arg-4-nitroanilide (QAR-4NA), which allowed direct assay of proteases in living cells. Comparisons of bronchial epithelial cells (CFBE41o−) with and without functioning cystic fibrosis transmembrane conductance regulator (CFTR) revealed similar levels of apical and basolateral aprotinin-inhibitable activity. Although recombinant matriptase was more active than prostasin in hydrolyzing QAR-4NA, cell surface activity resisted matriptase-selective inhibition, suggesting that prostasin dominates. Surface biotinylation revealed similar expression of matriptase and prostasin in epithelial cells expressing wild-type vs. ΔF508-mutated CFTR. However, the ratio of mature to inactive proprostasin suggested surface enrichment of active enzyme. Although small amounts of matriptase and prostasin were shed spontaneously, prostasin anchored to the cell surface by glycosylphosphatidylinositol was the major contributor to observed QAR-4NA-hydrolyzing activity. For example, the apical surface of wild-type CFBE41o− epithelial cells express 22% of total, extractable, aprotinin-inhibitable, QAR-4NA-hydrolyzing activity and 16% of prostasin immunoreactivity. In conclusion, prostasin is present, mature and active on the apical surface of wild-type and CF bronchial epithelial cells, where it can be targeted for inhibition via the airway lumen.

scholarly journals Respiratory Syncytial Virus Infection of Human Airway Epithelial Cells Is Polarized, Specific to Ciliated Cells, and without Obvious Cytopathology

2002 ◽  
Vol 76 (11) ◽  
pp. 5654-5666 ◽  
Author(s):  
Liqun Zhang ◽  
Mark E. Peeples ◽  
Richard C. Boucher ◽  
Peter L. Collins ◽  
Raymond J. Pickles
Keyword(s):  
Gene Transfer ◽  
Epithelial Cell ◽  
Epithelial Cells ◽  
Airway Epithelium ◽  
Airway Epithelial Cells ◽  
Apical Surface ◽  
Airway Epithelial ◽  
Ciliated Cells ◽  
Transfer Vectors ◽  
Syncytial Virus

ABSTRACT Gene therapy for cystic fibrosis (CF) lung disease requires efficient gene transfer to airway epithelial cells after intralumenal delivery. Most gene transfer vectors so far tested have not provided the efficiency required. Although human respiratory syncytial virus (RSV), a common respiratory virus, is known to infect the respiratory epithelium, the mechanism of infection and the epithelial cell type targeted by RSV have not been determined. We have utilized human primary airway epithelial cell cultures that generate a well-differentiated pseudostratified mucociliary epithelium to investigate whether RSV infects airway epithelium via the lumenal (apical) surface. A recombinant RSV expressing green fluorescent protein (rgRSV) infected epithelial cell cultures with high gene transfer efficiency when applied to the apical surface but not after basolateral inoculation. Analyses of the cell types infected by RSV revealed that lumenal columnar cells, specifically ciliated epithelial cells, were targeted by RSV and that cultures became susceptible to infection as they differentiated into a ciliated phenotype. In addition to infection of ciliated cells via the apical membrane, RSV was shed exclusively from the apical surface and spread to neighboring ciliated cells by the motion of the cilial beat. Gross histological examination of cultures infected with RSV revealed no evidence of obvious cytopathology, suggesting that RSV infection in the absence of an immune response can be tolerated for >3 months. Therefore, rgRSV efficiently transduced the airway epithelium via the lumenal surface and specifically targeted ciliated airway epithelial cells. Since rgRSV appears to breach the lumenal barriers encountered by other gene transfer vectors in the airway, this virus may be a good candidate for the development of a gene transfer vector for CF lung disease.

scholarly journals Insulin signaling via the PI3-kinase/Akt pathway regulates airway glucose uptake and barrier function in a CFTR-dependent manner

2017 ◽  
Vol 312 (5) ◽  
pp. L688-L702 ◽  
Author(s):  
Samuel A. Molina ◽  
Hannah K. Moriarty ◽  
Daniel T. Infield ◽  
Barry R. Imhoff ◽  
Rachel J. Vance ◽  
...  
Keyword(s):  
Cystic Fibrosis ◽  
Epithelial Cells ◽  
Small Molecules ◽  
Glucose Uptake ◽  
Glucose Transporter ◽  
Airway Epithelial Cells ◽  
Airway Epithelial ◽  
Airway Epithelia ◽  
Human Airway ◽  
Airway Cells

Cystic fibrosis-related diabetes is the most common comorbidity associated with cystic fibrosis (CF) and correlates with increased rates of lung function decline. Because glucose is a nutrient present in the airways of patients with bacterial airway infections and because insulin controls glucose metabolism, the effect of insulin on CF airway epithelia was investigated to determine the role of insulin receptors and glucose transport in regulating glucose availability in the airway. The response to insulin by human airway epithelial cells was characterized by quantitative PCR, immunoblot, immunofluorescence, and glucose uptake assays. Phosphatidylinositol 3-kinase/protein kinase B (Akt) signaling and cystic fibrosis transmembrane conductance regulator (CFTR) activity were analyzed by pharmacological and immunoblot assays. We found that normal human primary airway epithelial cells expressed glucose transporter 4 and that application of insulin stimulated cytochalasin B-inhibitable glucose uptake, consistent with a requirement for glucose transporter translocation. Application of insulin to normal primary human airway epithelial cells promoted airway barrier function as demonstrated by increased transepithelial electrical resistance and decreased paracellular flux of small molecules. This provides the first demonstration that airway cells express insulin-regulated glucose transporters that act in concert with tight junctions to form an airway glucose barrier. However, insulin failed to increase glucose uptake or decrease paracellular flux of small molecules in human airway epithelia expressing F508del-CFTR. Insulin stimulation of Akt1 and Akt2 signaling in CF airway cells was diminished compared with that observed in airway cells expressing wild-type CFTR. These results indicate that the airway glucose barrier is regulated by insulin and is dysfunctional in CF.

scholarly journals Effects of xenon gas on human airway epithelial cells during hyperoxia and hypothermia

2020 ◽  
Vol 13 (4) ◽  
pp. 469-476
Author(s):  
Y. Zhu ◽  
J.J. Mosko ◽  
A. Chidekel ◽  
M.R. Wolfson ◽  
T.H. Shaffer
Keyword(s):  
Epithelial Cells ◽  
Total Protein ◽  
Cell Function ◽  
Airway Epithelial Cells ◽  
Control Group ◽  
Apical Surface ◽  
Airway Epithelial ◽  
Human Airway ◽  
Immunofluorescence Localization ◽  
Human Airway Epithelial Cells

BACKGROUND: Hypothermia with xenon gas has been used to reduce brain injury and disability rate after perinatal hypoxia-ischemia. We evaluated xenon gas therapy effects in an in vitro model with or without hypothermia on cultured human airway epithelial cells (Calu-3). METHODS: Calu-3 monolayers were grown at an air-liquid interface and exposed to one of the following conditions: 1) 21% FiO2 at 37°C (control); 2) 45% FiO2 and 50% xenon at 37°C; 3) 21% FiO2 and 50% xenon at 32°C; 4) 45% FiO2 and 50% xenon at 32°C for 24 hours. Transepithelial resistance (TER) measurements were performed and apical surface fluids were collected and assayed for total protein, IL-6, and IL-8. Three monolayers were used for immunofluorescence localization of zonula occludens-1 (ZO-1). The data were analyzed by one-way ANOVA. RESULTS: TER decreased at 24 hours in all treatment groups. Xenon with hyperoxia and hypothermia resulted in greatest decrease in TER compared with other groups. Immunofluorescence localization of ZO-1 (XY) showed reduced density of ZO-1 rings and incomplete ring-like staining in the 45% FiO2– 50% xenon group at 32°C compared with other groups. Secretion of total protein was not different among groups. Secretion of IL-6 in 21% FiO2 with xenon group at 32°C was less than that of the control group. The secretion of IL-8 in 45% FiO2 with xenon at 32°C was greater than that of other groups. CONCLUSION: Hyperoxia and hypothermia result in detrimental epithelial cell function and inflammation over 24-hour exposure. Xenon gas did not affect cell function or reduce inflammation.

scholarly journals Interactions of pathogenic and commensal strains of Mannheimia haemolytica with differentiated bovine airway epithelial cells grown at an air-liquid interface

2017 ◽  
Author(s):  
Daniel Cozens ◽  
Erin Sutherland ◽  
Miquel Lauder ◽  
Geraldine Taylor ◽  
Catherine C. Berry ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Tight Junctions ◽  
Airway Epithelial Cells ◽  
Respiratory Epithelium ◽  
Mannheimia Haemolytica ◽  
Sudden Increase ◽  
Apical Surface ◽  
Airway Epithelial ◽  
Upper Respiratory Tract

AbstractMannheimia haemolyticaserotype A2 is a common commensal species present in the nasopharynx of healthy cattle. However, prior to the onset of bovine pneumonic pasteurellosis, there is sudden increase inM. haemolyticaserotype A1 within the upper respiratory tract. The events during this selective proliferation of serotype A1 strains are poorly characterised. In this investigation, a differentiated bovine airway epithelial cell culture was used to study the interactions of A1 and A2 bovine isolates with the respiratory epithelium. This model reproduced the key defences of the airway epithelium, including tight junctions and mucociliary clearance. Although initial adherence of the serotype A1 strains was low, by 12 hours post-infection the bacteria was able to traverse the tight junctions to form foci of infection below the apical surface. The size, density and number of these foci increased with time, as did the cytopathic effects observed in the bovine bronchial epithelial cells. Penetration ofM. haemolyticaA1 into the sub-apical epithelium was shown to be through transcytosis but not paracytosis. Commensal A2 bovine isolates however were not capable of colonising the model to a high degree, and did not penetrate the epithelium following initial adherence at the apical surface. This difference in their ability to colonise the respiratory epithelium may account for the sudden proliferation of serotype A1 in the onset of pneumonia pasteurellosis. The pathogenesis observed was replicated by virulent A2 ovine isolates; however colonisation was 10-fold lower in comparison to bovine A1 strains. This investigation provides new insight into the interactions ofM. haemolyticawith bovine airway epithelial cells which are occurringin vivoduring pneumonia pasteurellosis.

scholarly journals Human Airway Epithelial Cells Sense Pseudomonas aeruginosa Infection via Recognition of Flagellin by Toll-Like Receptor 5

2005 ◽  
Vol 73 (11) ◽  
pp. 7151-7160 ◽  
Author(s):  
Zhe Zhang ◽  
Jean-Pierre Louboutin ◽  
Daniel J. Weiner ◽  
Joanna B. Goldberg ◽  
James M. Wilson
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Epithelial Cells ◽  
Inflammatory Responses ◽  
Airway Epithelial Cells ◽  
Respiratory Pathogen ◽  
Luciferase Reporter ◽  
Apical Surface ◽  
Airway Epithelial ◽  
Human Airway ◽  
Human Airway Epithelial Cells

ABSTRACT Pseudomonas aeruginosa, an opportunistic respiratory pathogen that infects the majority of patients with cystic fibrosis, initiates host inflammatory responses through interaction with airway epithelial cells. The Toll-like receptors (TLRs) are a family of pathogen pattern recognition receptors that play key roles in host innate immunity. In this study we aimed to determine whether TLRs mediate the interaction between P. aeruginosa and airway epithelial cells. Individual murine TLRs (TLR1 to TLR9) and dual combinations of these TLRs that activate an NF-κB-driven luciferase reporter in response to PAO1 were screened in HEK 293 cells. TLR5, TLR2, a combination of TLR1 and TLR2, or a combination of TLR2 and TLR6 responded to PAO1. Another P. aeruginosa strain, strain PAK, activated TLR5 similarly, while the isogenic flagellin-deficient strain PAK/fliC and the flagellum-free bacterium Haemophilus influenzae failed to activate TLR5. Reverse transcription-PCR was used to probe the presence of multiple TLRs (including TLR5) in primary human airway epithelial cells (HAECs). Immunostaining with TLR5 antibodies showed that TLR5 was expressed in HAECs and on the apical surface of the human trachea epithelium. In HAECs, PAO1, PAK, and Burkholderia cepacia, but not flagellin-deficient strain PAK/fliC or a B. cepacia fliC mutant, activated the NF-κB reporter. Dominant negative TLR5 specifically blocked the response to P. aeruginosa but not to the response to lipoteichoic acid, a specific ligand of TLR2. We also determined that MyD88, IRAK, TRAF6, and Toll-interacting protein (Tollip), but not TIRAP, were involved in the TLR-mediated response to P. aeruginosa in HAECs. These findings demonstrate that the airway epithelial receptor TLR5 senses P. aeruginosa through its flagellin protein, which may have an important role in the initiation of the host inflammatory reaction to clear the invading pathogen.

scholarly journals Transepithelial glucose transport and metabolism in H441 human airway epithelial cells

2008 ◽  
Vol 22 (S1) ◽  
Author(s):  
Kameljit Kaur Kalsi ◽  
Owen Fraser ◽  
Yuen‐Li Chung ◽  
Edward Tarelli ◽  
Barbara Philips ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Glucose Transport ◽  
Airway Epithelial Cells ◽  
Airway Epithelial ◽  
Human Airway ◽  
Human Airway Epithelial Cells

scholarly journals Glucose transport in lung airway epithelial cells

2007 ◽  
Vol 21 (5) ◽  
Author(s):  
Kameljit Kaur Kalsi ◽  
Rudulfo A Medina ◽  
Graham A Heap ◽  
Emma H Baker ◽  
Barbara J Philips ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Glucose Transport ◽  
Airway Epithelial Cells ◽  
Airway Epithelial

scholarly journals Regulation of endogenous ENaC functional expression by CFTR and ΔF508-CFTR in airway epithelial cells

2011 ◽  
Vol 300 (1) ◽  
pp. L88-L101 ◽  
Author(s):  
Ronald C. Rubenstein ◽  
Shannon R. Lockwood ◽  
Ellen Lide ◽  
Rebecca Bauer ◽  
Laurence Suaud ◽  
...  
Keyword(s):  
Cystic Fibrosis ◽  
Epithelial Cells ◽  
Functional Expression ◽  
Surface Expression ◽  
Airway Epithelial Cells ◽  
Cell Surface Expression ◽  
Apical Surface ◽  
Airway Epithelial ◽  
Airway Epithelia ◽  
Δf508 Cftr

The functional expression of the epithelial sodium channel (ENaC) appears elevated in cystic fibrosis (CF) airway epithelia, but the mechanism by which this occurs is not clear. We tested the hypothesis that the cystic fibrosis transmembrane conductance regulator (CFTR) alters the trafficking of endogenously expressed human ENaC in the CFBE41o− model of CF bronchial epithelia. Functional expression of ENaC, as defined by amiloride-inhibited short-circuit current ( Isc) in Ussing chambers, was absent under control conditions but present in CFBE41o− parental and ΔF508-CFTR-overexpressing cells after treatment with 1 μM dexamethasone (Dex) for 24 h. The effect of Dex was mimicked by incubation with the glucocorticoid hydrocortisone but not with the mineralocorticoid aldosterone. Application of trypsin to the apical surface to activate uncleaved, “near-silent” ENaC caused an additional increase in amiloride-sensitive Isc in the Dex-treated cells and was without effect in the control cells, suggesting that Dex increased ENaC cell surface expression. In contrast, Dex treatment did not stimulate amiloride-sensitive Isc in CFBE41o− cells that stably express wild-type (wt) CFTR. CFBE41o− wt cells also had reduced expression of α- and γ-ENaC compared with parental and ΔF508-CFTR-overexpressing cells. Furthermore, application of trypsin to the apical surface of Dex-treated CFBE41o− wt cells did not stimulate amiloride-sensitive Isc, suggesting that ENaC remained absent from the surface of these cells even after Dex treatment. We also tested the effect of trafficking-corrected ΔF508-CFTR on ENaC functional expression. Incubation with 1 mM 4-phenylbutyrate synergistically increased Dex-induced ENaC functional expression in ΔF508-CFTR-overexpressing cells. These data support the hypothesis that wt CFTR can regulate the whole cell, functional, and surface expression of endogenous ENaC in airway epithelial cells and that absence of this regulation may foster ENaC hyperactivity in CF airway epithelia.

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