scholarly journals The FOXA1 transcriptional network coordinates key functions of primary human airway epithelial cells

2020 ◽  
Vol 319 (1) ◽  
pp. L126-L136
Author(s):  
Alekh Paranjapye ◽  
Michael J. Mutolo ◽  
Jey Sabith Ebron ◽  
Shih-Hsing Leir ◽  
Ann Harris
Keyword(s):  
Transcription Factors ◽  
Epithelial Cells ◽  
Airway Epithelium ◽  
Epithelial To Mesenchymal Transition ◽  
Airway Epithelial Cells ◽  
Airway Epithelial ◽  
Mesenchymal Transition ◽  
Human Airway ◽  
Human Airway Epithelium ◽  
Two Factors

The differentiated functions of the human airway epithelium are coordinated by a complex network of transcription factors. These include the pioneer factors Forkhead box A1 and A2 (FOXA1 and FOXA2), which are well studied in several tissues, but their role in airway epithelial cells is poorly characterized. Here, we define the cistrome of FOXA1 and FOXA2 in primary human bronchial epithelial (HBE) cells by chromatin immunoprecipitation with deep-sequencing (ChIP-seq). Next, siRNA-mediated depletion of each factor is used to investigate their transcriptome by RNA-seq. We found that, as predicted from their DNA-binding motifs, genome-wide occupancy of the two factors showed substantial overlap; however, their global impact on gene expression differed. FOXA1 is an abundant transcript in HBE cells, while FOXA2 is expressed at low levels, and both these factors likely exhibit autoregulation and cross-regulation. FOXA1 regulated loci are involved in cell adhesion and the maintenance of epithelial cell identity, particularly through repression of genes associated with epithelial to mesenchymal transition (EMT). FOXA1 also directly targets other transcription factors with a known role in the airway epithelium such as SAM-pointed domain-containing Ets-like factor (SPDEF). The intersection of the cistrome and transcriptome for FOXA1 revealed enrichment of genes involved in epithelial development and tissue morphogenesis. Moreover, depletion of FOXA1 was shown to reduce the transepithelial resistance of HBE cells, confirming the role of this factor in maintaining epithelial barrier integrity.

scholarly journals Somatic cell hemoglobin modulates nitrogen oxide metabolism in the human airway epithelium

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Nadzeya Marozkina ◽  
Laura Smith ◽  
Yi Zhao ◽  
Joe Zein ◽  
James F. Chmiel ◽  
...  
Keyword(s):  
Gene Expression ◽  
Epithelial Cells ◽  
Nitrogen Oxides ◽  
Airway Epithelium ◽  
Airflow Obstruction ◽  
Airway Epithelial Cells ◽  
Airway Epithelial ◽  
Human Airway ◽  
Human Airway Epithelium ◽  
Human Airway Epithelial Cells

AbstractEndothelial hemoglobin (Hb)α regulates endothelial nitric oxide synthase (eNOS) biochemistry. We hypothesized that Hb could also be expressed and biochemically active in the ciliated human airway epithelium. Primary human airway epithelial cells, cultured at air–liquid interface (ALI), were obtained by clinical airway brushings or from explanted lungs. Human airway Hb mRNA data were from publically available databases; or from RT-PCR. Hb proteins were identified by immunoprecipitation, immunoblot, immunohistochemistry, immunofluorescence and liquid chromatography- mass spectrometry. Viral vectors were used to alter Hbβ expression. Heme and nitrogen oxides were measured colorimetrically. Hb mRNA was expressed in human ciliated epithelial cells. Heme proteins (Hbα, β, and δ) were detected in ALI cultures by several methods. Higher levels of airway epithelial Hbβ gene expression were associated with lower FEV1 in asthma. Both Hbβ knockdown and overexpression affected cell morphology. Hbβ and eNOS were apically colocalized. Binding heme with CO decreased extracellular accumulation of nitrogen oxides. Human airway epithelial cells express Hb. Higher levels of Hbβ gene expression were associated with airflow obstruction. Hbβ and eNOS were colocalized in ciliated cells, and heme affected oxidation of the NOS product. Epithelial Hb expression may be relevant to human airways diseases.

scholarly journals Glycated Albumin Triggers an Inflammatory Response in the Human Airway Epithelium and Causes an Increase in Ciliary Beat Frequency

2021 ◽  
Vol 12 ◽  
Author(s):  
Moira L. Aitken ◽  
Ranjani Somayaji ◽  
Thomas R. Hinds ◽  
Maricela Pier ◽  
Karla Droguett ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Airway Epithelium ◽  
Glycated Albumin ◽  
Beat Frequency ◽  
Ciliary Beat Frequency ◽  
Airway Epithelial Cells ◽  
Airway Epithelial ◽  
Human Airway ◽  
Human Airway Epithelium ◽  
Ciliary Beat

The role of inflammation in airway epithelial cells and its regulation are important in several respiratory diseases. When disease is present, the barrier between the pulmonary circulation and the airway epithelium is damaged, allowing serum proteins to enter the airways. We identified that human glycated albumin (GA) is a molecule in human serum that triggers an inflammatory response in human airway epithelial cultures. We observed that single-donor human serum induced IL-8 secretion from primary human airway epithelial cells and from a cystic fibrosis airway cell line (CF1-16) in a dose-dependent manner. IL-8 secretion from airway epithelial cells was time dependent and rapidly increased in the first 4 h of incubation. Stimulation with GA promoted epithelial cells to secrete IL-8, and this increase was blocked by the anti-GA antibody. The IL-8 secretion induced by serum GA was 10–50-fold more potent than TNFα or LPS stimulation. GA also has a functional effect on airway epithelial cells in vitro, increasing ciliary beat frequency. Our results demonstrate that the serum molecule GA is pro-inflammatory and triggers host defense responses including increases in IL-8 secretion and ciliary beat frequency in the human airway epithelium. Although the binding site of GA has not yet been described, it is possible that GA could bind to the receptor for advanced glycated end products (RAGE), known to be expressed in the airway epithelium; however, further experiments are needed to identify the mechanism involved. We highlight a possible role for GA in airway inflammation.

Alpha-adrenergic regulation of Na-Cl cotransport in human airway epithelium

1989 ◽  
Vol 257 (2) ◽  
pp. L125-L129 ◽  
Author(s):  
C. M. Liedtke
Keyword(s):  
Epithelial Cells ◽  
Airway Epithelium ◽  
Initial Rate ◽  
Airway Epithelial Cells ◽  
Airway Epithelial ◽  
Adrenergic Agonist ◽  
Beta Adrenergic ◽  
Human Airway ◽  
Cl Transport ◽  
Human Airway Epithelium

The demonstration of abnormal beta-adrenergic and cAMP-modulated apical Cl- channels in cystic fibrosis (CF) airway epithelial cells suggests that other transporters, which are required for Cl- secretion, may also be abnormally regulated. A basolateral cotransporter was investigated by determining the initial rate of 36Cl efflux from cells isolated from CF nasal polyps or trachea and non-CF trachea. Cells were preequilibrated with radioactive tracer at 25 degrees C, and tracer transport was initiated by 10-fold dilution of an aliquot of cells in radioisotope-free medium. The initial rate of Cl transport was calculated from the linear portion of the efflux curves. In CF and non-CF cells, base-line Cl- transport was not blocked by furosemide but was stimulated twofold by l-epinephrine in Ca2+-deficient and Ca2+-replete transport medium. In both types of cells, furosemide blocked 70 and 77%, respectively, of the stimulated Cl- transport. Prazosin, an alpha 1-adrenergic antagonist, blocked the effects of l-epinephrine and methoxamine, an alpha 1-adrenergic agonist, stimulated prazosin- and furosemide-sensitive Cl transport. Ionomycin mimicked the effects of l-epinephrine. l-Isoproterenol, a beta-adrenergic agonist, did not affect Cl transport. The results of this study indicate an alpha 1-adrenergic stimulation of furosemide-sensitive Cl transport in human airway epithelium that functions normally in CF airway epithelial cells. The transport mechanism is probably a Na-Cl or Na-K-2Cl cotransport located in the basolateral membrane and requires elevated intracellular Ca2+ for activation.

scholarly journals Measles Virus Ribonucleoprotein Complexes Rapidly Spread across Well-Differentiated Primary Human Airway Epithelial Cells along F-Actin Rings

mBio ◽  
2019 ◽  
Vol 10 (6) ◽  
Author(s):  
Brajesh K. Singh ◽  
Christian K. Pfaller ◽  
Roberto Cattaneo ◽  
Patrick L. Sinn
Keyword(s):  
Epithelial Cells ◽  
Airway Epithelium ◽  
Measles Virus ◽  
Airway Epithelial Cells ◽  
Airway Epithelial ◽  
Particle Release ◽  
Ribonucleoprotein Complex ◽  
Human Airway ◽  
Well Differentiated ◽  
Actin Rings

ABSTRACT Measles virus (MeV) is a highly contagious human pathogen that continues to be a worldwide health burden. One of the challenges for the study of MeV spread is the identification of model systems that accurately reflect how MeV behaves in humans. For our studies, we use unpassaged, well-differentiated primary cultures of airway epithelial cells from human donor lungs to examine MeV infection and spread. Here, we show that the main components of the MeV ribonucleoprotein complex (RNP), the nucleocapsid and phosphoprotein, colocalize with the apical and circumapical F-actin networks. To better understand how MeV infections spread across the airway epithelium, we generated a recombinant virus incorporating chimeric fluorescent proteins in its RNP complex. By live cell imaging, we observed rapid movement of RNPs along the circumapical F-actin rings of newly infected cells. This strikingly rapid mechanism of horizontal trafficking across epithelia is consistent with the opening of pores between columnar cells by the viral membrane fusion apparatus. Our work provides mechanistic insights into how MeV rapidly spreads through airway epithelial cells, contributing to its extremely contagious nature. IMPORTANCE The ability of viral particles to directly spread cell to cell within the airways without particle release is considered to be highly advantageous to many respiratory viruses. Our previous studies in well-differentiated, primary human airway epithelial cells suggest that measles virus (MeV) spreads cell to cell by eliciting the formation of intercellular membrane pores. Based on a newly generated ribonucleoprotein complex (RNP) “tracker” virus, we document by live-cell microscopy that MeV RNPs move along F-actin rings before entering a new cell. Thus, rather than diffusing through the cytoplasm of a newly infected columnar cell, RNPs take advantage of the cytoskeletal infrastructure to rapidly spread laterally across the human airway epithelium. This results in rapid horizontal spread through the epithelium that does not require particle release.

Regulation of CFTR channels by HCO3−-sensitive soluble adenylyl cyclase in human airway epithelial cells

2005 ◽  
Vol 289 (5) ◽  
pp. C1145-C1151 ◽  
Author(s):  
Yan Wang ◽  
Chak Sum Lam ◽  
Fan Wu ◽  
Wen Wang ◽  
Yuanyuan Duan ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Adenylyl Cyclase ◽  
Airway Epithelial Cells ◽  
Airway Epithelial ◽  
Adenylyl Cyclases ◽  
Open Probability ◽  
Soluble Adenylyl Cyclase ◽  
Human Airway ◽  
Human Airway Epithelium ◽  
Polymerase Chain

CFTR channels conduct HCO3− in addition to Cl− in airway epithelial cells. A defective HCO3−-transporting function of CFTR may underlie the pathogenesis of cystic fibrosis. In the present study, we have investigated whether a HCO3−-sensitive soluble adenylyl cyclase (sAC) is functionally coupled with CFTR and thus forms an autoregulatory mechanism for HCO3− transport in human airway epithelial Calu-3 cells. A reverse transcriptase-polymerase chain reaction showed that transcripts of both full-length and truncated sACs are present in Calu-3 cells. Truncated sAC protein is the predominant, if not the only, isoform expressed in Calu-3 cells. HCO3− stimulated a modest increase in cAMP production, and the increase was sensitive to 2-hydroxyestradiol (2-HE), a sAC inhibitor, but not to SQ22,536, a blocker of conventional transmembrane adenylyl cyclases. These results suggest that sAC is functional in Calu-3 cells. Adding 25 mM HCO3− to the bath stimulated CFTR-mediated whole cell currents in the absence, but not in the presence, of 2-HE. In cell-attached membrane patches, 25 or 50 mM HCO3− in the bath markedly increased the product of channel number and open probability of CFTR, and this activation was attenuated by 2-HE. These findings demonstrate that sAC signaling pathway is involved in the regulation of CFTR function in human airway epithelium and thereby provides a link between the level of intracellular HCO3−/CO2 and the modulation of HCO3−-conductive CFTR function by cAMP/PKA.

scholarly journals In primary airway epithelial cells, the unjamming transition is distinct from the epithelial-to-mesenchymal transition

2020 ◽  
Vol 11 (1) ◽  
Author(s):  
Jennifer A. Mitchel ◽  
Amit Das ◽  
Michael J. O’Sullivan ◽  
Ian T. Stancil ◽  
Stephen J. DeCamp ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Epithelial To Mesenchymal Transition ◽  
Airway Epithelial Cells ◽  
Cellular Migration ◽  
Cell Junctions ◽  
Airway Epithelial ◽  
Human Bronchial Epithelial Cells ◽  
Intact Cells ◽  
Bronchial Epithelial ◽  
Mesenchymal Transition

Abstract The epithelial-to-mesenchymal transition (EMT) and the unjamming transition (UJT) each comprises a gateway to cellular migration, plasticity and remodeling, but the extent to which these core programs are distinct, overlapping, or identical has remained undefined. Here, we triggered partial EMT (pEMT) or UJT in differentiated primary human bronchial epithelial cells. After triggering UJT, cell-cell junctions, apico-basal polarity, and barrier function remain intact, cells elongate and align into cooperative migratory packs, and mesenchymal markers of EMT remain unapparent. After triggering pEMT these and other metrics of UJT versus pEMT diverge. A computational model attributes effects of pEMT mainly to diminished junctional tension but attributes those of UJT mainly to augmented cellular propulsion. Through the actions of UJT and pEMT working independently, sequentially, or interactively, those tissues that are subject to development, injury, or disease become endowed with rich mechanisms for cellular migration, plasticity, self-repair, and regeneration.

scholarly journals Human Airway Epithelial Cell Responses to Neisseria lactamica and Purified Porin via Toll-Like Receptor 2-Dependent Signaling

2010 ◽  
Vol 78 (12) ◽  
pp. 5314-5323 ◽  
Author(s):  
Xiuping Liu ◽  
Lee M. Wetzler ◽  
Laura Oliveira Nascimento ◽  
Paola Massari
Keyword(s):  
Epithelial Cells ◽  
Host Cell ◽  
Airway Epithelium ◽  
Airway Epithelial Cells ◽  
Airway Epithelial ◽  
Toll Like Receptor ◽  
Tlr2 Expression ◽  
Neisseria Lactamica ◽  
Human Airway ◽  
Human Airway Epithelial Cells

ABSTRACT The human airway epithelium is constantly exposed to microbial products from colonizing organisms. Regulation of Toll-like receptor (TLR) expression and specific interactions with bacterial ligands is thought to mitigate exacerbation of inflammatory processes induced by the commensal flora in these cells. The genus Neisseria comprises pathogenic and commensal organisms that colonize the human nasopharynx. Neisseria lactamica is not associated with disease, but N. meningitidis occasionally invades the host, causing meningococcal disease and septicemia. Upon colonization of the airway epithelium, specific host cell receptors interact with numerous Neisseria components, including the PorB porin, at the immediate bacterial-host cell interface. This major outer membrane protein is expressed by all Neisseria strains, regardless of pathogenicity, but its amino acid sequence varies among strains, particularly in the surface-exposed regions. The interaction of Neisseria PorB with TLR2 is essential for driving TLR2/TLR1-dependent cellular responses and is thought to occur via the porin's surface-exposed loop regions. Our studies show that N. lactamica PorB is a TLR2 ligand but its binding specificity for TLR2 is different from that of meningococcal PorB. Furthermore, N. lactamica PorB is a poor inducer of proinflammatory mediators and of TLR2 expression in human airway epithelial cells. These effects are reproduced by whole N. lactamica organisms. Since the responsiveness of human airway epithelial cells to colonizing bacteria is in part regulated via TLR2 expression and signaling, commensal organisms such as N. lactamica would benefit from expressing a product that induces low TLR2-dependent local inflammation, likely delaying or avoiding clearance by the host.

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