scholarly journals Autocrine Acetylcholine, Induced by IL-17A via NFκB and ERK1/2 Pathway Activation, Promotes MUC5AC and IL-8 Synthesis in Bronchial Epithelial Cells

2016 ◽  
Vol 2016 ◽  
pp. 1-16 ◽  
Author(s):  
Angela Marina Montalbano ◽  
Giusy Daniela Albano ◽  
Anna Bonanno ◽  
Loredana Riccobono ◽  
Caterina Di Sano ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Anticholinergic Drug ◽  
Airway Epithelial Cells ◽  
Bronchial Epithelial Cells ◽  
Choline Uptake ◽  
Airway Epithelial ◽  
Related Activity ◽  
Bronchial Epithelial ◽  
Pathway Activation

IL-17A is overexpressed in the lung during acute neutrophilic inflammation. Acetylcholine (ACh) increases IL-8 and Muc5AC production in airway epithelial cells. We aimed to characterize the involvement of nonneuronal components of cholinergic system on IL-8 and Muc5AC production in bronchial epithelial cells stimulated with IL-17A. Bronchial epithelial cells were stimulated with recombinant human IL-17A (rhIL-17A) to evaluate the ChAT expression, the ACh binding and production, the IL-8 release, and the Muc5AC production. Furthermore, the effectiveness of PD098,059 (inhibitor of MAPKK activation), Bay11-7082 (inhibitor of IkBαphosphorylation), Hemicholinium-3 (HCh-3) (choline uptake blocker), and Tiotropium bromide (Spiriva®) (anticholinergic drug) was tested in ourin vitromodel. We showed that rhIL-17A increased the expression of ChAT, the levels of ACh binding and production, and the IL-8 and Muc5AC production in stimulated bronchial epithelial cells compared with untreated cells. The pretreatment of the cells with PD098,059 and Bay11-7082 decreased the ChAT expression and the ACh production/binding, while HCh-3 and Tiotropium decreased the IL-8 and Muc5AC synthesis in bronchial epithelial cells stimulated with rhIL-17A. IL-17A is involved in the IL-8 and Muc5AC production promoting, via NFκB and ERK1/2 pathway activation, the synthesis of ChAT, and the related activity of autocrine ACh in bronchial epithelial cells.

β-Adrenergic agonist modulation of monocyte adhesion to airway epithelial cells in vitro

2000 ◽  
Vol 278 (1) ◽  
pp. L139-L147 ◽  
Author(s):  
Debra J. Romberger ◽  
Peggy Heires ◽  
Stephen I. Rennard ◽  
Todd A. Wyatt
Keyword(s):  
Cell Adhesion ◽  
Epithelial Cells ◽  
Airway Epithelial Cells ◽  
Bronchial Epithelial Cells ◽  
Airway Epithelial ◽  
Human Bronchial Epithelial Cells ◽  
Adrenergic Agonist ◽  
Bronchial Epithelial

β-Adrenergic agonists are commonly used in the treatment of obstructive airway diseases and are known to modulate cAMP-dependent processes of airway epithelial cells. However, little is known regarding the ability of cAMP-dependent mechanisms to influence cell-cell interactions within the airway. Thus we investigated the role of the β-adrenergic agonist isoproterenol in modulating the ability of human bronchial epithelial cells to support the adhesion of THP-1 cells, a monocyte/macrophage cell line, in vitro. We demonstrated that pretreatment of human bronchial epithelial cells (HBECs) with 10 μM isoproterenol or 100 μM salbutamol augments the adhesion of fluorescently labeled THP-1 cells to HBEC monolayers by ∼40–60%. The increase in THP-1 cell adhesion occurred with 10 min of isoproterenol pretreatment of HBECs and gradually declined but persisted with up to 24 h of isoproterenol exposure. Exposure of THP-1 cells to isoproterenol or salbutamol before the adhesion assays did not result in an increase in adhesion to HBECs, suggesting that the isoproterenol modulation was primarily via changes in epithelial cells. A specific protein kinase A inhibitor, KT-5720, inhibited subsequent isoproterenol augmentation of THP-1 cell adhesion, further supporting the role of cAMP-dependent mechanisms in modulating THP-1 cell adhesion to HBECs.

ICAM-1-independent adhesion of neutrophils to phorbol ester-stimulated human airway epithelial cells

1999 ◽  
Vol 277 (3) ◽  
pp. L465-L471 ◽  
Author(s):  
Alessandro Celi ◽  
Silvana Cianchetti ◽  
Stefano Petruzzelli ◽  
Stefano Carnevali ◽  
Filomena Baliva ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Airway Epithelial Cells ◽  
Bronchial Epithelial Cells ◽  
Neutrophil Adhesion ◽  
Late Time ◽  
Airway Epithelial ◽  
Bronchial Epithelial ◽  
Human Airway ◽  
Stimulation Of ◽  
Human Airway Epithelial Cells

Intercellular adhesion molecule-1 (ICAM-1) is the only inducible adhesion receptor for neutrophils identified in bronchial epithelial cells. We stimulated human airway epithelial cells with various agonists to evaluate whether ICAM-1-independent adhesion mechanisms could be elicited. Phorbol 12-myristate 13-acetate (PMA) stimulation of cells of the alveolar cell line A549 caused a rapid, significant increase in neutrophil adhesion from 11 ± 3 to 49 ± 7% (SE). A significant increase from 17 ± 4 to 39 ± 6% was also observed for neutrophil adhesion to PMA-stimulated human bronchial epithelial cells in primary culture. Although ICAM-1 expression was upregulated by PMA at late time points, it was not affected at 10 min when neutrophil adhesion was already clearly enhanced. Antibodies to ICAM-1 had no effect on neutrophil adhesion. In contrast, antibodies to the leukocyte integrin β-chain CD18 totally inhibited the adhesion of neutrophils to PMA-stimulated epithelial cells. These results demonstrate that PMA stimulation of human airway epithelial cells causes an increase in neutrophil adhesion that is not dependent on ICAM-1 upregulation.

Resting and Cytokine-Stimulated Human Small Airway Epithelial Cells Recognize and Engulf Apoptotic Eosinophils

Blood ◽  
1999 ◽  
Vol 94 (8) ◽  
pp. 2827-2835 ◽  
Author(s):  
Garry M. Walsh ◽  
Darren W. Sexton ◽  
Morgan G. Blaylock ◽  
Catherine M. Convery
Keyword(s):  
Epithelial Cells ◽  
Interleukin 1 ◽  
Airway Epithelial Cells ◽  
Small Airway ◽  
Airway Epithelial ◽  
Human Bronchial Epithelial Cells ◽  
Bronchial Epithelial ◽  
Small Airway Epithelial Cells ◽  
Necrosis Factor

Eosinophils, which are prominent cells in asthmatic inflammation, undergo apoptosis and are recognized and engulfed by phagocytic macrophages in vitro. We have examined the ability of human small airway epithelial cells (SAEC) to recognize and ingest apoptotic human eosinophils. Cultured SAEC ingested apoptotic eosinophils but not freshly isolated eosinophils or opsonized erythrocytes. The ability of SAEC to ingest apoptotic eosinophils was enhanced by interleukin-1 (IL-1) or tumor necrosis factor  (TNF) in a time- and concentration-dependent fashion. IL-1 was found to be more potent than TNF and each was optimal at 10−10 mol/L, with a significant (P < .05) effect observed at 1 hour postcytokine incubation that was maximal at 5 hours. IL-1 stimulation not only increased the number of SAEC engulfing apoptotic eosinophils, but also enhanced their capacity for ingestion. The amino sugars glucosamine, n-acetyl glucosamine, and galactosamine significantly inhibited uptake of apoptotic eosinophils by both resting and IL-1–stimulated SAEC, in contrast to the parent sugars glucose, galactose, mannose, and fucose. Incubation of apoptotic eosinophils with the tetrapeptide RGDS, but not RGES, significantly inhibited their uptake by both resting and IL-1–stimulated SAEC, as did monoclonal antibody against vβ3 and CD36. Thus, SAEC recognize apoptotic eosinophils via lectin- and integrin-dependent mechanisms. These data demonstrate a novel function for human bronchial epithelial cells that might represent an important mechanism in the resolution of eosinophil-induced asthmatic inflammation.

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 Development and validation of an open-source, disposable, 3D-printed in vitro environmental exposure system for Transwell® culture inserts

2020 ◽  
Author(s):  
Abiram Chandiramohan ◽  
Mohammedhossein Dabaghi ◽  
Jennifer A. Aguiar ◽  
Nicholas Tiessen ◽  
Mary Stewart ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Airway Epithelial Cells ◽  
Airway Epithelial ◽  
Exposure System ◽  
Bronchial Epithelial ◽  
Human Airway ◽  
3D Printed ◽  
Air Liquid Interface ◽  
Whole Smoke

AbstractAccessible in vitro models recapitulating the human airway that are amenable to study whole cannabis smoke exposure are needed for immunological and toxicological studies that inform public health policy and recreational cannabis use. In the present study, we developed and validated a novel 3D printed In Vitro Exposure System (IVES) that can be directly applied to study the effect of cannabis smoke exposure on primary human bronchial epithelial cells.Using commercially available design software and a 3D printer, we designed a four-chamber Transwell® insert holder for exposures to whole smoke. Software was used to model gas distribution, concentration gradients, velocity profile and shear stress within IVES. Following simulations, primary human bronchial epithelial cells cultured at air-liquid interface on Transwell® inserts were exposed to whole cannabis smoke. Following 24 hours, outcome measurements included cell morphology, epithelial barrier function, lactate dehydrogenase (LDH) levels, cytokine and gene expression.Whole smoke delivered through IVES possesses velocity profiles consistent with uniform gas distribution across the four chambers and complete mixing. Airflow velocity ranged between 1.0-1.5 μm s−1 and generated low shear stresses (<< 1 Pa). Human airway epithelial cells exposed to cannabis smoke using IVES showed changes in cell morphology and disruption of barrier function without significant cytotoxicity. Cannabis smoke elevated IL-1 family cytokines and elevated CYP1A1 and CYP1B1 expression relative to control.IVES represents an accessible, open-source, exposure system that can be used to model varying types of cannabis smoke exposures with human airway epithelial cells grown under air-liquid interface culture conditions.

scholarly journals Expression/Activation of PAR-1 in Airway Epithelial Cells of COPD Patients: Ex Vivo/In Vitro Study

2021 ◽  
Vol 22 (19) ◽  
pp. 10703
Author(s):  
Angela Marina Montalbano ◽  
Giuseppina Chiappara ◽  
Giusy Daniela Albano ◽  
Maria Ferraro ◽  
Caterina Di Sano ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Ex Vivo ◽  
In Vitro Study ◽  
Bronchial Epithelial Cell ◽  
Airway Epithelial Cells ◽  
Epithelial Cell Line ◽  
Airway Epithelial ◽  
Bronchial Epithelial ◽  
Copd Patients

The role of PAR-1 expression and activation was described in epithelial cells from the central and distal airways of COPD patients using an ex vivo/in vitro model. PAR-1 immunoreactivity was studied in epithelial cells from surgical specimens of the central and distal airways of COPD patients and healthy control (HC). Furthermore, PAR-1 expression and activation were measured in both the human bronchial epithelial cell line (16HBE) and normal human bronchial epithelial cells (NHBEs) exposed to cigarette smoke extract (CSE) (10%) or thrombin. Finally, cell proliferation, apoptosis, and IL-8 release were detected in stimulated NHBEs. We identified higher levels of PAR-1 expression/activation in epithelial cells from the central airways of COPD patients than in HC. Active PAR-1 increased in epithelial cells from central and distal airways of COPD, with higher levels in COPD smokers (correlated with pack-years) than in COPD ex-smokers. 16HBE and NHBEs exposed to CSE or thrombin showed increased levels of active PAR-1 (localized in the cytoplasm) than baseline conditions, while NHBEs treated with thrombin or CSE showed increased levels of IL-8 proteins, with an additional effect when used in combination. Smoking habits generate the upregulation of PAR-1 expression/activation in airway epithelial cells, and promoting IL-8 release might affect the recruitment of infiltrating cells in the airways of COPD patients.

scholarly journals Capsular Polysaccharide is a Main Component ofMycoplasma ovipneumoniaein the Pathogen-Induced Toll-Like Receptor-Mediated Inflammatory Responses in Sheep Airway Epithelial Cells

2017 ◽  
Vol 2017 ◽  
pp. 1-15 ◽  
Author(s):  
Zhongjia Jiang ◽  
Fuyang Song ◽  
Yanan Li ◽  
Di Xue ◽  
Guangcun Deng ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Capsular Polysaccharide ◽  
Inflammatory Responses ◽  
Airway Epithelial Cells ◽  
Bronchial Epithelial Cells ◽  
Atypical Pneumonia ◽  
Activator Protein 1 ◽  
Airway Epithelial ◽  
Toll Like Receptor ◽  
Bronchial Epithelial

Mycoplasma ovipneumoniae(M. ovipneumoniae) is characterized as an etiological agent of primary atypical pneumonia that specifically infects sheep and goat. In an attempt to better understand the pathogen-host interaction between the invadingM. ovipneumoniaeand airway epithelial cells, we investigated the host inflammatory responses against capsular polysaccharide (designated as CPS) ofM. ovipneumoniaeusing sheep bronchial epithelial cells cultured in an air-liquid interface (ALI) model. Results showed that CPS derived fromM. ovipneumoniaecould activate toll-like receptor- (TLR-) mediated inflammatory responses, along with an elevated expression of nuclear factor kappa B (NF-κB), activator protein-1 (AP-1), and interferon regulatory factor 3 (IRF3) as well as various inflammatory-associated mediators, representatively including proinflammatory cytokines, such as IL1β, TNFα, and IL8, and anti-inflammatory cytokines such as IL10 and TGFβof TLR signaling cascade. Mechanistically, the CPS-induced inflammation was TLR initiated and was mediated by activations of both MyD88-dependent and MyD88-independent signaling pathways. Of importance, a blockage of CPS with specific antibody led a significant reduction ofM. ovipneumoniae-induced inflammatory responses in sheep bronchial epithelial cells. These results suggested that CPS is a key virulent component ofM. ovipneumoniae, which may play a crucial role in the inflammatory response induced byM. ovipneumoniaeinfections.

Regulation of SLPI and elafin release from bronchial epithelial cells by neutrophil defensins

2000 ◽  
Vol 278 (1) ◽  
pp. L51-L58 ◽  
Author(s):  
Sandra van Wetering ◽  
Abraham C. van der Linden ◽  
Marianne A. J. A. van Sterkenburg ◽  
Willem I. de Boer ◽  
Astrid L. A. Kuijpers ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Proteinase Inhibitor ◽  
Airway Epithelial Cells ◽  
Bronchial Epithelial Cells ◽  
Protein Release ◽  
Mrna Levels ◽  
Main Function ◽  
Airway Epithelial ◽  
Dynamic Regulation ◽  
Bronchial Epithelial

Secretory leukocyte proteinase inhibitor (SLPI) is a serine proteinase inhibitor that is produced locally in the lung by cells of the submucosal bronchial glands and by nonciliated epithelial cells. Its main function appears to be the inhibition of neutrophil elastase (NE). Recently, NE was found to enhance SLPI mRNA levels while decreasing SLPI protein release in airway epithelial cells. Furthermore, glucocorticoids were shown to increase both constitutive and NE-induced SLPI mRNA levels. In addition to NE, stimulated neutrophils also release α-defensins. Defensins are small, antimicrobial polypeptides that are found in high concentrations in purulent secretions of patients with chronic airway inflammation. Like NE, defensins induce interleukin-8 production in airway epithelial cells. This induction is sensitive to inhibition by the glucocorticoid dexamethasone and is prevented in the presence of α1-proteinase inhibitor. The aim of the present study was to investigate the effect of defensins on the production of SLPI and the related NE inhibitor elafin/SKALP in primary bronchial epithelial cells (PBECs). Defensins significantly increase SLPI protein release by PBECs in a time- and dose-dependent fashion without affecting SLPI mRNA synthesis. In the presence of α1-proteinase inhibitor, the defensin-induced SLPI protein release is further enhanced, but no effect was observed on SLPI mRNA levels. Dexamethasone did not affect SLPI protein release from control or defensin-treated PBECs. In addition, we observed a constitutive release of elafin/SKALP by PBECs, but this was not affected by defensins. The present results suggest a role for defensins in the dynamic regulation of the antiproteinase screen in the lung at sites of inflammation.

scholarly journals In Vitro Investigations on Optimizing and Nebulization of IVT-mRNA Formulations for Potential Pulmonary based Alpha-1-Antitrypsin Deficiency Treatment

2021 ◽  
Author(s):  
Shan Guan ◽  
Max Darmstaedter ◽  
Chuanfei Xu ◽  
Joseph Rosenecker
Keyword(s):  
Epithelial Cells ◽  
Transfection Efficiency ◽  
Airway Epithelial Cells ◽  
Bronchial Epithelial Cells ◽  
Human Bronchial Epithelial Cells ◽  
Bronchial Epithelial ◽  
Antitrypsin Deficiency ◽  
Alpha 1 Antitrypsin Deficiency ◽  
Alpha 1 Antitrypsin

Abstract Background: In vitro transcribed (IVT) mRNA has come into focus in recent years as a potential therapeutic approach for the treatment of genetic diseases. The pulmonary delivery of IVT-mRNA encoding alpha-1-antitrypsin (A1AT) is a promising strategy for protein replacement therapy of alpha-1-antitrypsin deficiency (AATD). The nebulized A1AT-mRNA formulations would be a highly acceptable and tolerable remedy for the AATD patients in the future. Method: we first optimized parameters that influencing the transfection efficiency of formulations containing IVT-mRNA and Lipofectamine2000 based on human bronchial epithelial cells transfection. Cell viability was evaluated by performing MTT assay after transfection with different IVT-mRNA lipoplexes. Functional analysis was employed to assess the biological function of A1AT proteins produced from optimized formulations using anti-trypsin assay and anti-elastase assay. Results: Lipoplexes prepared by IVT-mRNA encoding A1AT (A1AT-mRNA) in optimum conditions could successfully transfect human bronchial epithelial cells without significant toxicity. A reduction in transfection efficiency was observed for aerosolized lipoplexes that can be partially overcome by increasing the initial amount of components. A1AT produced from cells transfected by nebulized A1AT-mRNA lipoplexes is functional and could successfully inhibit the enzyme activity of trypsin as well as elastase. Conclusion: Aerosolization of A1AT-mRNA therapeutic constitute a potentially powerful means to transfect airway epithelial cells with the purpose of producing functional A1AT while bringing along the unique advantages of IVT-mRNA.

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