Paracrine stimulation of surfactant secretion by extracellular ATP in response to mechanical deformation

2005 ◽  
Vol 289 (3) ◽  
pp. L489-L496 ◽  
Author(s):  
Anand S. Patel ◽  
David Reigada ◽  
Claire H. Mitchell ◽  
Sandra R. Bates ◽  
Susan S. Margulies ◽  
...  
Keyword(s):  
Mechanical Deformation ◽  
Extracellular Atp ◽  
Alveolar Epithelial Cells ◽  
Luciferase Assay ◽  
Alveolar Type ◽  
Type I ◽  
Lipid Secretion ◽  
Surfactant Secretion ◽  
Surfactant Lipid ◽  
Stimulation Of

We developed a heterologous system to study the effect of mechanical deformation on alveolar epithelial cells. First, isolated primary rat alveolar type II (ATII) cells were plated onto silastic substrata coated with fibronectin and maintained in culture under conditions where they become alveolar type I-like (ATI) cells. This was followed by a second set of ATII cells labeled with the nontransferable, vital fluorescent stain 5-chloromethylfluorescein diacetate to distinguish them from ATI cells. By morphometric analysis, equibiaxial deformation (stretch) of the silastic substratum induced comparable changes in cell surface area for both ATII and ATI cells. Surfactant lipid secretion was measured using cells metabolically labeled with [3H]choline. In response to 21% tonic stretch for 15 min, ATII cells seeded with ATI cells secreted nearly threefold more surfactant lipid compared with ATII cells seeded alone. ATI cells did not secrete lipid in response to stretch. The enhanced lipid secretion by ATII plus ATI cocultures was inhibited by treatment with apyrase and adenosine deaminase, suggesting that ATP release by ATI cells enhanced surfactant lipid secretion at 21% stretch. This was confirmed using a luciferase assay where, in response to 21% stretch, ATI cells released fourfold more ATP than ATII cells. Because ATI cells release significantly more ATP at a lower level of stretch than ATII cells, this supports the hypothesis that ATI cells are mechanosensors in the lung and that paracrine stimulation of ATII cells by extracellular ATP released from ATI cells plays a role in regulating surfactant secretion.

scholarly journals Mechanical stretch activates piezo1 in caveolae of alveolar type I cells to trigger ATP release and paracrine stimulation of surfactant secretion from alveolar type II cells

2020 ◽  
Vol 34 (9) ◽  
pp. 12785-12804 ◽  
Author(s):  
Kathrin Diem ◽  
Michael Fauler ◽  
Giorgio Fois ◽  
Andreas Hellmann ◽  
Natalie Winokurow ◽  
...  
Keyword(s):  
Atp Release ◽  
Mechanical Stretch ◽  
Alveolar Type ◽  
Type I ◽  
Type Ii Cells ◽  
Type Ii ◽  
Alveolar Type Ii Cells ◽  
Surfactant Secretion ◽  
I Cells ◽  
Stimulation Of

scholarly journals Lipopolysaccharide Inhibits Alpha Epithelial Sodium Channel Expression via MiR-124-5p in Alveolar Type 2  Epithelial Cells

2020 ◽  
Vol 2020 ◽  
pp. 1-9 ◽  
Author(s):  
Yan Ding ◽  
Yong Cui ◽  
Zhiyu Zhou ◽  
Yapeng Hou ◽  
Xining Pang ◽  
...  
Keyword(s):  
Acute Lung Injury ◽  
Epithelial Cells ◽  
Lung Injury ◽  
Sodium Channel ◽  
Target Genes ◽  
Cell Model ◽  
Epithelial Sodium Channel ◽  
Alveolar Epithelial Cells ◽  
Luciferase Assay ◽  
Alveolar Type

Mesenchymal stem cells (MSCs) have been a potential strategy in the pretreatment of pulmonary diseases, while the mechanisms of MSCs-conditioned medium (MSCs-CM) involved with microRNAs on the regulation of lung ion transport are seldom reported. We investigated the role of miR-124-5p in lipopolysaccharide-involved epithelial sodium channel (ENaC) dysfunction and explored the potential target of miR-124-5p. We observed the lower expression of miR-124-5p after the administration of MSCs-CM, and the overexpression or inhibition of miR-124-5p regulated epithelial sodium channel α-subunit (α-ENaC) expression at protein levels in mouse alveolar type 2 epithelial (AT2) cells. We confirmed that α-ENaC is one of the target genes of miR-124-5p through dual luciferase assay and Ussing chamber assay revealed that miR-124-5p inhibited amiloride-sensitive currents associated with ENaC activity in intact H441 monolayers. Our results demonstrate that miR-124-5p can decrease the expression and function of α-ENaC in alveolar epithelial cells by targeting the 3′-UTR. The involvement of MSCs-CM in lipopolysaccharide-induced acute lung injury cell model could be related to the downregulation of miR-124-5p on α-ENaC, which may provide a new target for the treatment of acute lung injury.

scholarly journals Transient receptor vanilloid 4 (TRPV4) channels are essential for alveolar epithelial cell function

2019 ◽  
Author(s):  
Jonas Weber ◽  
Yu-Kai Chao ◽  
Martina Kannler ◽  
Gabriela Krasteva-Christ ◽  
Suhasini Rajan ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Barrier Function ◽  
Alveolar Epithelial Cells ◽  
Transient Receptor Potential Vanilloid ◽  
Alveolar Type ◽  
Type I ◽  
Edema Formation ◽  
Genetic Ablation ◽  
Alveolar Epithelial ◽  
Transient Receptor

AbstractIschemia-reperfusion(IR)-induced edema formation can be mimicked ex-vivo in isolated perfused mouse lungs (IPL). Here we show enhanced edema formation in transient receptor potential vanilloid 4 (TRPV4)-deficient (TRPV4-/-) IPL compared to wild-type (WT) controls in response to IR, indicating a protective role of TRPV4 to maintain the alveolar epithelial barrier. By immunohistochemistry, mRNA profiling or electrophysiological analysis we detected TRPV4 in bronchial epithelium, alveolar type I (ATI) and alveolar type II (ATII) cells. Genetic ablation of TRPV4 resulted in reduced expression of aquaporin-5 (AQP-5) channels in ATI as well as decreased production of pro surfactant protein C (pSP-C) in ATII cells. Migration of TRPV4-deficient ATI cells was reduced and cell barrier function was impaired. Moreover, adult TRPV4−/− lungs developed emphysema-like changes and altered lung parameters compared to WT lungs. Therefore, our data highlight novel essential functions of TRPV4 channels in alveolar epithelial cells and in the protection from edema formation.eLife digestTransient receptor potential vanilloid 4 (TRPV4) is a non-selective Ca2+ permeable cation channel expressed in lung endothelium where increased channel activity has been shown to compromise endothelial barrier function. In other tissues however, the channel maintains physiological cell barriers, e.g. in skin, the urogenital tract and the corneal epithelium. In tracheal epithelial cells TRPV4 channels regulate ciliar beat frequency and in alveolar epithelial cells TRPV4 activation by 4α-phorbol esters produced blebs and breaks in lung septa by unknown molecular mechanisms. To understand the channels role in lung function Weber et al. employed ex-vivo isolated perfused mouse lungs (IPL) to mimic ischemia-reperfusion-induced edema as one of the most common and significant causes of morbidity and mortality after lung transplantation in human patients. TRPV4-deficient (TRPV4−/−) IPL developed enhanced edema formation compared to wild-type (WT) controls in response to ischemia and reperfusion, indicating a protective role of TRPV4 to maintain the alveolar epithelial barrier. TRPV4 was detected in bronchial epithelium, alveolar type I (ATI) and alveolar type II (ATII) cells by immunohistochemistry or mRNA profiling. Genetic ablation of TRPV4 resulted in reduced expression and plasma membrane insertion of water conducting aquaporin-5 (AQP-5) channels in ATI cells compared to WT mice. Analysis of isolated primary TRPV4−/− ATII cells revealed a reduced expression of pro surfactant protein-C (pSP-C) a precursor of a protein important for decreasing surface tension and for alveolar fluid homeostasis. Moreover, the TRPV4 activator GSK1016790A induced increases in current densities only in WT but not in TRPV4−/− ATII cells. On a molecular level ablation of TRPV4 induced less Ca2+-mediated nuclear translocation of nuclear factor of activated T-cells (NFAT) to the nucleus, which may be responsible for reduced expression of the identified proteins. Although the ability of TRPV4−/− ATII to differentiate to ATI cells was unchanged, migration of TRPV4-deficient ATI cells was reduced and cell barrier function was impaired. Moreover, TRPV4−/− lungs of adult mice developed significantly larger mean chord lengths and altered lung function compared to WT lungs. The findings of Weber et al. highlights novel essential functions of TRPV4 channels in alveolar epithelial cells and in the protection from edema formation.

scholarly journals UTP regulation of ion transport in alveolar epithelial cells involves distinct mechanisms

2009 ◽  
Vol 297 (3) ◽  
pp. L439-L454 ◽  
Author(s):  
Chuanxiu Yang ◽  
Lijing Su ◽  
Yang Wang ◽  
Lin Liu
Keyword(s):  
Ion Transport ◽  
Short Circuit ◽  
Alveolar Epithelial Cells ◽  
Alveolar Type ◽  
Channel Blockers ◽  
Type I ◽  
Type Ii Cells ◽  
Type Ii ◽  
Type I Cells ◽  
I Cells

UTP is known to regulate alveolar fluid clearance. However, the relative contribution of alveolar type I cells and type II cells to this process is unknown. In this study, we investigated the effects of UTP on ion transport in type I-like cell (AEC I) and type II-like cell (AEC II) monolayers. Luminal treatment of cell monolayers with UTP increased short-circuit current ( Isc) of AEC II but decreased Isc of AEC I. The Cl− channel blockers NPPB and DIDS inhibited the UTP-induced changes in Isc (Δ Isc) in both types of cells. Amiloride, an inhibitor of epithelial Na+ channels (ENaC), abolished the UTP-induced Δ Isc in AEC I, but not in AEC II. The general blocker of K+ channels, BaCl2, eliminated the UTP-induced Δ Isc in AEC II, but not in AEC I. The intermediate conductance (IKCa) blocker, clofilium, also blocked the UTP effect in AEC II. The signal transduction pathways mediated by UTP were the same in AEC I and AEC II. Furthermore, UTP increased Cl− secretion in AEC II and Cl− absorption in AEC I. Our results suggest that UTP induces opposite changes in Isc in AEC I and AEC II, likely due to the reversed Cl− flux and different contributions of ENaC and IKCa. Our results further imply a new concept that type II cells contribute to UTP-induced fluid secretion and type I cells contribute to UTP-induced fluid absorption in alveoli.

Dopamine activates amiloride-sensitive sodium channels in alveolar type I cells in lung slice preparations

2006 ◽  
Vol 291 (4) ◽  
pp. L610-L618 ◽  
Author(s):  
My N. Helms ◽  
Julie Self ◽  
Hui Fang Bao ◽  
Lauren C. Job ◽  
Lucky Jain ◽  
...  
Keyword(s):  
Receptor Antagonist ◽  
Single Channel ◽  
Sch 23390 ◽  
Alveolar Epithelial Cells ◽  
Alveolar Type ◽  
Type I ◽  
Channel Activity ◽  
Open Probability ◽  
Single Channel Recordings ◽  
Alveolar Epithelial

Active Na+ reabsorption by alveolar epithelial cells generates the driving force used to clear fluids from the air space. Using single-channel methods, we examined epithelial Na+ channel (ENaC) activity of alveolar type I (AT1) cells from live 250- to 300-μm sections of lung tissue, circumventing concerns that protracted cell isolation procedures might compromise the innate transport properties of native lung cells. We used fluorescein-labeled Erythrina crystagalli lectin to positively identify AT1 cells for single-channel patch-clamp analysis. We demonstrated, for the first time, single-channel recordings of highly selective and nonselective amiloride-sensitive ENaC channels (HSC and NSC, respectively) from AT1 cells in situ, with mean conductances of 8.2 ± 2.5 and 22 ± 3.2 pS, respectively. Additionally, 25 nM amiloride in the patch electrode blocked Na+ channel activity in AT1 cells. Immunohistochemical studies demonstrated the presence of dopamine D1 and D2 receptors on the surface of AT1 cells, and single-channel recordings showed that 10 μM dopamine increased Na+ channel activity [product of the number of channels and single-channel open probability ( NPo)] from 0.31 ± 0.19 to 0.60 ± 0.21 ( P < 0.001). The D1 receptor antagonist SCH-23390 (10 μM) blocked the stimulatory effect of dopamine on AT1 cells, but the D2 receptor antagonist sulpiride did not.

IL-23 amplifies the epithelial-mesenchymal transition of mechanically conditioned alveolar epithelial cells in RA-ILD through mTOR/S6 signaling

2021 ◽  
Author(s):  
Chujie Zhang ◽  
Shaohua Wang ◽  
Jessica Lau ◽  
Anja C. Roden ◽  
Eric L. Matteson ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Lung Fibrosis ◽  
Epithelial Mesenchymal Transition ◽  
Alveolar Epithelial Cells ◽  
Alveolar Type ◽  
Type I ◽  
Sequencing Analysis ◽  
Mesenchymal Transition ◽  
Caveolin 1 ◽  
Alveolar Epithelial

Epithelial-mesenchymal transition (EMT) creates an environment facilitating fibrosis following alveolar epithelial cell injury. IL-23 has important roles in chronic autoimmune conditions like rheumatoid arthritis (RA), but its role in the interstitial lung disease that affects RA patients is unclear. This study aimed to determine the pro-fibrogenic role of IL-23 on somatic alveolar type I (ATI) epithelial cells. Primary ATI cells were isolated from rats and cultured on plastic dishes for 1-3 weeks. After prolonged culture (≥14 days) on rigid culture dishes, primary ATI cells gradually acquired a mesenchymal phenotype, identified by decreased expression of caveolin-1, and reorganization of F-actin cytoskeleton, indicating the initiation of EMT by matrix stiffness. To determine how IL-23 promotes EMT in vitro, transitioning ATI cells, cultured on a stiff substrate for ≥14 days were stimulated with IL-23. The EMT phenotype was significantly enhanced by IL-23 which upregulated α-SMA, collagen I/III protein, and decreased caveolin-1. Furthermore, IL-23 significantly promoted cell invasion as well as apoptotic resistance on transitioning ATI cells. Mechanistically, IL-23 induced EMT was mTOR/S6 signaling dependent and reversible by rapamycin. Transcriptional sequencing analysis of human lung fibrosis biopsy tissue revealed key roles for IL-23 in RA-ILD. This result was further validated by significantly upregulated IL-23 expression at the mRNA level in RA-ILD lung sections. Notably, transitioning ATI epithelial cells were abundantly detected in RA-ILD tissue. Taken together, these data support a role for IL-23 in the pathogenesis of RA lung fibrosis by promoting EMT in alveolar epithelial cells through mTOR/S6 signaling.

Expression of LAR-PTP2 in rat lung is confined to proliferating epithelia lining the airways and air sacs

1996 ◽  
Vol 270 (4) ◽  
pp. L566-L576 ◽  
Author(s):  
H. Kim ◽  
H. Yeger ◽  
R. Han ◽  
M. Wallace ◽  
B. Goldstein ◽  
...  
Keyword(s):  
Epithelial Cell ◽  
Tyrosine Phosphatase ◽  
Alveolar Epithelial Cells ◽  
Nuclear Antigen ◽  
Alveolar Type ◽  
Type I ◽  
Proliferating Cells ◽  
Alveolar Epithelial ◽  
Air Sacs ◽  
Epithelial Cell Layer

The LAR family tyrosine phosphatase LAR-PTP2B (RPTP sigma) was previously shown to be expressed in the central and peripheral nervous system. Here we show that LAR-PTP2, the larger alternatively spliced form of the gene, is expressed in proliferating undifferentiated lung epithelia in a developmentally regulated manner: Using in situ hybridization and parallel immunostaining with proliferating cell nuclear antigen to detect proliferating cells, we demonstrate that LAR-PTP2 is expressed exclusively in the undifferentiated epithelial cell layer lining the bronchi, bronchioles, and air sacs in late fetal development and in the neonatal lung. These cells correspond to Clara and fetal alveolar type II cells, as determined by parallel immunostaining with antibodies to surfactant proteins A and B. LAR-PTP2 expression declined progressively with postnatal development, and by adult stage there was no detectable expression in the airways or in the distal (type I and II) mature nonproliferating alveolar epithelial cells. These results suggest that LAR-PTP2 may be involved in the regulation of epithelial cell proliferation/differentiation during lung development.

scholarly journals Mitochondrial quality control in alveolar epithelial cells damaged by S. aureus pneumonia in mice

2017 ◽  
Vol 313 (4) ◽  
pp. L699-L709 ◽  
Author(s):  
Hagir B. Suliman ◽  
Bryan Kraft ◽  
Raquel Bartz ◽  
Lingye Chen ◽  
Karen E. Welty-Wolf ◽  
...  
Keyword(s):  
Quality Control ◽  
Cell Death ◽  
Cell Survival ◽  
Mitochondrial Biogenesis ◽  
Citrate Synthase ◽  
Mitochondrial Damage ◽  
Alveolar Epithelial Cells ◽  
Alveolar Type ◽  
Type I ◽  
Mitochondrial Quality Control

Mitochondrial damage is often overlooked in acute lung injury (ALI), yet most of the lung’s physiological processes, such as airway tone, mucociliary clearance, ventilation-perfusion (Va/Q) matching, and immune surveillance require aerobic energy provision. Because the cell’s mitochondrial quality control (QC) process regulates the elimination and replacement of damaged mitochondria to maintain cell survival, we serially evaluated mitochondrial biogenesis and mitophagy in the alveolar regions of mice in a validated Staphylococcus aureus pneumonia model. We report that apart from cell lysis by direct contact with microbes, modest epithelial cell death was detected despite significant mitochondrial damage. Cell death by TdT-mediated dUTP nick-end labeling staining occurred on days 1 and 2 postinoculation: apoptosis shown by caspase-3 cleavage was present on days 1 and 2, while necroptosis shown by increased levels of phospho- mixed lineage kinase domain-like protein (MLKL) and receptor-interacting serine/threonine-protein kinase 1 (RIPK1) was present on day 1. Cell death in alveolar type I (AT1) cells assessed by bronchoalveolar lavage fluid receptor for advanced glycation end points (RAGE) levels was high, yet AT2 cell death was limited while both mitochondrial biogenesis and mitophagy were induced. These mitochondrial QC mechanisms were evaluated mainly in AT2 cells by localizing increases in citrate synthase content, increases in nuclear mitochondrial biogenesis regulators nuclear respiratory factor-1 (NRF-1) and peroxisome proliferator-activated receptor-γ coactivator-1α (PGC-1α), and increases in light chain 3B protein (LC3-I)/LC3II ratios. Concomitant changes in p62, Pink 1, and Parkin protein levels indicated activation of mitophagy. By confocal microscopy, mitochondrial biogenesis and mitophagy were often observed on day 1 within the same AT2 cells. These findings imply that mitochondrial QC activation in pneumonia-damaged AT2 cells promotes cell survival in support of alveolar function.

Differential expression of Na-K-ATPase isoforms in rat alveolar epithelial cells

1997 ◽  
Vol 273 (1) ◽  
pp. L246-L255 ◽  
Author(s):  
K. M. Ridge ◽  
D. H. Rutschman ◽  
P. Factor ◽  
A. I. Katz ◽  
A. M. Bertorello ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Alveolar Epithelial Cells ◽  
Cell Phenotype ◽  
Alveolar Type ◽  
Type I ◽  
Binding Studies ◽  
Alveolar Epithelial ◽  
Functional Classes ◽  
Atpase Subunits ◽  
Ouabain Inhibition

Lung Na-K-ATPase has been shown to contribute to vectorial Na+ transport and edema clearance. The alpha 1- and beta 1-Na-K-ATPase subunits have been localized to alveolar type II (ATII) cells, and the alpha 2-Na-K-ATPase has been reported in rat lung homogenates. Expression of Na-K-ATPase alpha 1-, alpha 2-, and beta 1-subunits was investigated in rat ATII cells cultured for 7 days, a period during which they lose their phenotypic markers and differentiate to an alveolar type I (ATI)-like cell phenotype. Differentiation of ATII cells to an ATI-like phenotype resulted in a decrease of alpha 1- and an increase of alpha 2-mRNA and protein abundance without changes in the beta 1-subunit. Thus ATI-like cells exhibited a mixture of alpha 1- and alpha 2-isoforms. Nuclear run-on analysis suggests that these changes were transcriptionally regulated. The existence of the distinct functional classes of Na-K-ATPase in ATII and ATI-like cells was confirmed by ouabain inhibition of Na-K-ATPase activity. Ouabain inhibition of ATII cells was consistent with expression of the alpha 1-isozyme [50% inhibitory concentration (IC50) = 4 x 10(-5) M], whereas, in ATI-like cells, it was consistent with the presence of both alpha 1- and alpha 2-isozymes (IC50 = 9.0 x 10(-5) and 1.5 x 10(-7) M, respectively); [3H]ouabain binding studies corroborated these findings. Our results indicate that, during ATII cell cytodifferentiation with time in culture, there is a shift in isoform composition that may reflect physiological functions of alveolar epithelial cells.

scholarly journals Therapeutic Potential of Cytosolic PLA2 Isoform–Specific Inhibitor Arachidonyl Trifluromethyl Ketone in Cigarette Smoke Condensate–Induced Pathological Conditions in Alveolar Type I and II Epithelial Cells

2018 ◽  
Vol 4 (Supplement 2) ◽  
pp. 221s-221s
Author(s):  
S. Kumar ◽  
S.K. Sharma ◽  
B. Medhi ◽  
K.L. Khanduja
Keyword(s):  
Epithelial Cells ◽  
Cigarette Smoke ◽  
Therapeutic Potential ◽  
Specific Inhibitor ◽  
Alveolar Epithelial Cells ◽  
Alveolar Type ◽  
Type I ◽  
Continuous Exposure ◽  
Alveolar Epithelial ◽  
Cytosolic Pla2

Background: Cigarette smoking is responsible for various lung pathologies including chronic lung inflammation, emphysema, chronic obstructive pulmonary disease (COPD), cancer, and annually causes almost 10 million deaths globally. During smoke exposure, most affected cells are the alveolar epithelial cells where as a repair mechanism, activation of cytosolic phospholipase A2 enzymes takes place. High free radicals and cPLA2 activity due to continuous exposure of smoke exposure leads to elevated levels of secondary metabolites and various pathophysiologic conditions such as chronic inflammation, oxidative stress and cancer. To reduce the burden of chronic inflammation as well as oxidative stress, and higher levels of secondary metabolites whose role is well defined in progression of cancer, we checked the therapeutic potential of cPLA2 inhibitor arachidonyl trifluromethyl ketone (ATK) by pharmacologically targeting the most expressible cPLA2 during continuous exposure of cigarette smoke. Aim: To check the therapeutic potential of cytosolic PLA2 isoform specific inhibitor arachidonyl trifluromethyl ketone in cigarette smoke condensate–induced pathologic conditions in alveolar type I and II epithelial cells. Methods: Effect of cPLA2 inhibitor on CSC-induced cPLA2 activity were checked using colorimetric assay, cell viability using MTT assay, FDA uptake assay using fluorescence microscopy, ROS levels and apoptosis markers through flow cytometry, and ERK levels using ELISA, in both type of alveolar epithelial cells. Results: ATK significantly mimicked CSC-induced cPLA2 activity, free radicals, primary apoptosis, ratio of apoptotic/apoptotic proteins and levels of ERK whereas protected cells from loss of cell viability and membrane integrity. Conclusion: Current observations revealed cPLA2s as a potential therapeutic target and their inhibitor ATK as a potential therapeutic agent in Cigarette smoke induced pathological conditions in alveolar type I and II epithelial cells.

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