Alveolar type II-like cells release G-CSF as neutrophil chemotactic activity

1998 ◽  
Vol 275 (4) ◽  
pp. L687-L693 ◽  
Author(s):  
Sekiya Koyama ◽  
Etsuro Sato ◽  
Takeshi Masubuchi ◽  
Akemi Takamizawa ◽  
Keishi Kubo ◽  
...  
Keyword(s):  
A549 Cells ◽  
Chemotactic Activity ◽  
Alveolar Epithelial Cells ◽  
Epithelial Cell Line ◽  
Alveolar Type ◽  
Type Ii ◽  
Tnf Α ◽  
Dose Dependent Fashion ◽  
Dose Dependent ◽  
Neutrophil Chemotactic Activity

We evaluated the potential of A549 cells, an alveolar type II epithelial cell line, to release granulocyte colony-stimulating factor (G-CSF), in addition to interleukin (IL)-8 and leukotriene B4, as neutrophil chemotactic activity (NCA). Human recombinant IL-1β stimulated A549 cells to release NCA in a time- and dose-dependent fashion. The released NCA was blocked by mouse anti-human G-CSF polyclonal antibody. Molecular-sieve column chromatography revealed that IL-1β induced the release of a 19- to 20-kDa chemotactic mass that was inhibited by anti-human G-CSF antibody. IL-1β stimulated the release of G-CSF in a dose-dependent fashion, but the time-dependent profile of G-CSF showed that the concentration of G-CSF declined after 48 h. Tumor necrosis factor (TNF)-α, Escherichia coli lipopolysaccharide (LPS), and bradykinin (BK) stimulated A549 cells to release NCA that was inhibited by anti-G-CSF antibody. The release of G-CSF in response to TNF-α, LPS, and BK was significantly increased. The similar concentrations of human recombinant G-CSF (10–1,000 pg/ml) as in the supernatant fluid induced neutrophil chemotaxis. G-CSF mRNA was expressed time and dose dependently at 4 h and declined after 4 h in response to IL-1β as evaluated by RT-PCR. The expression of G-CSF mRNA was also observed by TNF-α, LPS, and BK stimulation. These data suggest that type II alveolar epithelial cells may produce G-CSF as NCA and may participate in the regulation of leukocyte extravasation.

Expression of the tyrosine phosphatase LAR-PTP2 is developmentally regulated in lung epithelia

1994 ◽  
Vol 267 (3) ◽  
pp. L263-L270 ◽  
Author(s):  
D. Rotin ◽  
B. J. Goldstein ◽  
C. A. Fladd
Keyword(s):  
Epithelial Cells ◽  
Lung Development ◽  
Tyrosine Phosphatase ◽  
A549 Cells ◽  
Alveolar Epithelial Cells ◽  
Alveolar Type ◽  
Type Ii ◽  
Adult Type ◽  
Alveolar Epithelial

The role of tyrosine kinases in regulating cell proliferation, differentiation, and development has been well documented. In contrast, little is known about the role of protein tyrosine phosphatases (PTPs) in mammalian development. To identify PTPs that may be involved in lung development, we have isolated (by polymerase chain reaction) from rat fetal alveolar epithelial cells a cDNA fragment which was identified as the recently cloned tyrosine phosphatase LAR-PTP2. Analysis of tissue expression of LAR-PTP2 identified a approximately 7.5-kb message in the lung, which is also expressed weakly in brain, and an alternatively spliced approximately 6.0-kb message (LAR-PTP2B) expressed in brain. In the fetal lung, LAR-PTP2 was preferentially expressed in lung epithelial (but not fibroblast) cells grown briefly in primary culture, and its expression was tightly regulated during lung development, peaking at 20 days of gestational age (term = 22 days), when mature alveolar type II epithelium first appears. Accordingly, immunoblot analysis revealed high expression of endogenous LAR-PTP2 protein in alveolar epithelial cells from 21-day gestation fetuses. LAR-PTP2 was also expressed in lungs of newborn rats, but transcripts (and protein) were barely detectable in adult lungs and in the nonproliferating adult alveolar type II cells. Interestingly, expression was restored in the transformed adult type II-like A549 cells. These results suggest that LAR-PTP2 may play a role in the proliferation and/or differentiation of epithelial cells during lung development.

scholarly journals Nrf2 Alleviates TNF-α-Induced Oxidative Stress Damage in Type II Alveolar Epithelial Cells by Down-Regulating the Expression of NOX1

2020 ◽  
Author(s):  
Weijing Wu ◽  
Jiamin Zhang ◽  
Xihua Lian ◽  
Xiaoping Lin ◽  
Xiaoshan Su ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Epithelial Cells ◽  
Binding Site ◽  
Luciferase Activity ◽  
A549 Cells ◽  
Alveolar Epithelial Cells ◽  
Type Ii ◽  
Alveolar Epithelial ◽  
Tnf Α ◽  
Stress Damage

Abstract Objective: To study the roles of Nrf2 in acute lung injury (ALI) pathogenesis by investigating the effects of Nrf2 on regulating oxidative stress damage in TNF-α-induced type II alveolar epithelial cells (T2AECs).Methods: T2AECs were transfected with Nrf2 siRNA and overexpression vectors for six hours before being induced by TNF-α for 24 hours. Subsequently, levels of interleukins (IL-6 and IL-8), reactive oxygen species (ROS), malondialdehyde (MDA), total antioxidation capability (T-AOC), Nrf2, NOX1 and NF-kB were measured. Additionally, potential Nrf2 binding site in NOX1 promoter was predicted by AliBaba2.1 and two recombinant vectors, namely “pGL3-NOX1-1500” and “pGL3-NOX1-1489, were constructed by inserting the sequence of NOX1 promoter in full-length and that in the absence of Nrf2 binding site to pGL3 basic vector. T2AECs were transfected with these vectors prior to TNF-α induction and the luciferase activity was measured.Results: Levels of IL-6, IL-8, ROS and MDA were increased (P＜0.05) while T-AOC was decreased in TNF-α-induced A549 cells after the transfection of Nrf2 siRNA vector (P＜0.05). In contrast, concentrations of IL-6, IL-8, ROS and MDA were decreased (P＜0.05) whereas T-AOC was increased after the transfection of Nrf2 overexpression vector (P＜0.05). NOX1 promoter possesses one Nrf2 binding site. Cells transfected by “pGL3-NOX1-1500” vector had the highest luciferase activity, followed by cells transfected by “pGL3-NOX1-1489” vector and the control cells (P＜0.05).Conclusion: Nrf2 modulates NOX1 expression via binding to its promoter, by which against TNF-α-induced oxidative stress damage in T2AECs. Thus, Nrf2 might be a therapeutic target for ALI.

Proinflammatory response of alveolar epithelial cells is enhanced by alveolar macrophage-produced TNF-α during pulmonary ischemia-reperfusion injury

2007 ◽  
Vol 293 (1) ◽  
pp. L105-L113 ◽  
Author(s):  
Ashish K. Sharma ◽  
Lucas G. Fernandez ◽  
Alaa S. Awad ◽  
Irving L. Kron ◽  
Victor E. Laubach
Keyword(s):  
Epithelial Cells ◽  
Alveolar Macrophage ◽  
Alveolar Macrophages ◽  
Ischemia Reperfusion ◽  
Alveolar Epithelial Cells ◽  
Alveolar Type ◽  
Type Ii Cells ◽  
Type Ii ◽  
Alveolar Epithelial ◽  
Tnf Α

Pulmonary ischemia-reperfusion (IR) injury entails acute activation of alveolar macrophages followed by neutrophil sequestration. Although proinflammatory cytokines and chemokines such as TNF-α and monocyte chemoattractant protein-1 (MCP-1) from macrophages are known to modulate acute IR injury, the contribution of alveolar epithelial cells to IR injury and their intercellular interactions with other cell types such as alveolar macrophages and neutrophils remain unclear. In this study, we tested the hypothesis that following IR, alveolar macrophage-produced TNF-α further induces alveolar epithelial cells to produce key chemokines that could then contribute to subsequent lung injury through the recruitment of neutrophils. Cultured RAW264.7 macrophages and MLE-12 alveolar epithelial cells were subjected to acute hypoxia-reoxygenation (H/R) as an in vitro model of pulmonary IR. H/R (3 h/1 h) significantly induced KC, MCP-1, macrophage inflammatory protein-2 (MIP-2), RANTES, and IL-6 (but not TNF-α) by MLE-12 cells, whereas H/R induced TNF-α, MCP-1, RANTES, MIP-1α, and MIP-2 (but not KC) by RAW264.7 cells. These results were confirmed using primary murine alveolar macrophages and primary alveolar type II cells. Importantly, using macrophage and epithelial coculture methods, the specific production of TNF-α by H/R-exposed RAW264.7 cells significantly induced proinflammatory cytokine/chemokine expression (KC, MCP-1, MIP-2, RANTES, and IL-6) by MLE-12 cells. Collectively, these results demonstrate that alveolar type II cells, in conjunction with alveolar macrophage-produced TNF-α, contribute to the initiation of acute pulmonary IR injury via a proinflammatory cascade. The release of key chemokines, such as KC and MIP-2, by activated type II cells may thus significantly contribute to neutrophil sequestration during IR injury.

Hypoxia decreases proteins involved in epithelial electrolyte transport in A549 cells and rat lung

2000 ◽  
Vol 279 (6) ◽  
pp. L1110-L1119 ◽  
Author(s):  
Ralf Wodopia ◽  
Hyun Soo Ko ◽  
Javiera Billian ◽  
Rudolf Wiesner ◽  
Peter Bärtsch ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Lung Tissue ◽  
A549 Cells ◽  
Alveolar Epithelial Cells ◽  
Alveolar Type ◽  
Type Ii Cells ◽  
Type Ii ◽  
Alveolar Type Ii Cells ◽  
Alveolar Epithelial

Fluid reabsorption from alveolar space is driven by active Na reabsorption via epithelial Na channels (ENaCs) and Na-K-ATPase. Both are inhibited by hypoxia. Here we tested whether hypoxia decreases Na transport by decreasing the number of copies of transporters in alveolar epithelial cells and in lungs of hypoxic rats. Membrane fractions were prepared from A549 cells exposed to hypoxia (3% O2) as well as from whole lung tissue and alveolar type II cells from rats exposed to hypoxia. Transport proteins were measured by Western blot analysis. In A549 cells, α1- and β1-Na-K-ATPase, Na/K/2Cl cotransport, and ENaC proteins decreased during hypoxia. In whole lung tissue, α1-Na-K-ATPase and Na/K/2Cl cotransport decreased. α- and β-ENaC mRNAs also decreased in hypoxic lungs. Similar results were seen in alveolar type II cells from hypoxic rats. These results indicate a slow decrease in the amount of Na-transporting proteins in alveolar epithelial cells during exposure to hypoxia that also occurs in vivo in lungs from hypoxic animals. The reduced number of transporters might account for the decreased transport activity and impaired edema clearance in hypoxic lungs.

Pattern of NOS2 and NOS3 mRNA expression in human A549 cells and primary cultured AEC II

2002 ◽  
Vol 282 (4) ◽  
pp. L684-L692 ◽  
Author(s):  
D. V. Pechkovsky ◽  
G. Zissel ◽  
T. Goldmann ◽  
M. Einhaus ◽  
C. Taube ◽  
...  
Keyword(s):  
Nitric Oxide ◽  
Mrna Expression ◽  
Primary Cultures ◽  
A549 Cells ◽  
Alveolar Epithelial Cells ◽  
Alveolar Type ◽  
Mrna Levels ◽  
Type Ii ◽  
Isolated Cells

The human alveolar type II epithelium-like cell line A549 expresses nitric oxide synthase type 2 (NOS2), but not NOS3, and produces nitric oxide (NO) upon appropriate stimulation. However, relatively little is known regarding the NOS2 and NOS3 expression of type II human alveolar epithelial cells (AEC II) in primary culture. We detected NOS3 mRNA in freshly isolated AEC II and after 24 h of culture. NOS3 mRNA levels were much higher in AEC II cultured for 24 h with or without interferon-γ, interleukin-1β, and tumor necrosis factor-α, compared with freshly isolated cells. Cytokine stimulation did not change the NOS3 mRNA expression level in AEC II compared with unstimulated cells. NOS3 protein expression was verified by Western blot, and measuring nitrate/nitrite revealed that the protein is active. In contrast, neither NOS2 mRNA nor protein could be detected in freshly isolated, unstimulated or cytokine-stimulated human AEC II in 24- or 72-h primary cultures, whereas A549 cells expressed NOS2 message and protein upon stimulation with proinflammatory cytokines. In situ hybridization confirmed that AEC II express NOS3, but not NOS2 mRNA in vivo. These data demonstrate that there are significant differences between primary AEC II and A549 cells in NOS mRNA expression pattern.

Connexin expression by alveolar epithelial cells is regulated by extracellular matrix

2001 ◽  
Vol 280 (2) ◽  
pp. L191-L202 ◽  
Author(s):  
Yihe Guo ◽  
Cara Martinez-Williams ◽  
Clare E. Yellowley ◽  
Henry J. Donahue ◽  
D. Eugene Rannels
Keyword(s):  
Extracellular Matrix ◽  
Epithelial Cells ◽  
Gap Junction ◽  
Intercellular Communication ◽  
Translational Regulation ◽  
Alveolar Epithelial Cells ◽  
Alveolar Type ◽  
Type Ii ◽  
Gap Junction Intercellular Communication ◽  
Type Ii Cell

Extracellular matrix (ECM) proteins promote attachment, spreading, and differentiation of cultured alveolar type II epithelial cells. The present studies address the hypothesis that the ECM also regulates expression and function of gap junction proteins, connexins, in this cell population. Expression of cellular fibronectin and connexin (Cx) 43 increase in parallel during early type II cell culture as Cx26 expression declines. Gap junction intercellular communication is established over the same interval. Cells plated on a preformed, type II cell-derived, fibronectin-rich ECM demonstrate accelerated formation of gap junction plaques and elevated gap junction intercellular communication. These effects are blocked by antibodies against fibronectin, which cause redistribution of Cx43 protein from the plasma membrane to the cytoplasm. Conversely, cells cultured on a laminin-rich ECM, Matrigel, express low levels of Cx43 but high levels of Cx26, reflecting both transcriptional and translational regulation. Cx26 and Cx43 thus demonstrate reciprocal regulation by ECM constituents.

Fluid transport across cultured rat alveolar epithelial cells: a novel in vitro system

2004 ◽  
Vol 287 (1) ◽  
pp. L104-L110 ◽  
Author(s):  
Xiaohui Fang ◽  
Yuanlin Song ◽  
Rachel Zemans ◽  
Jan Hirsch ◽  
Michael A. Matthay
Keyword(s):  
Epithelial Cells ◽  
Fluid Transport ◽  
Alveolar Epithelial Cells ◽  
Lung Epithelial Cells ◽  
Morphological Evidence ◽  
Alveolar Type ◽  
Type Ii ◽  
In Vitro System ◽  
Alveolar Epithelial

Previous studies have used fluid-instilled lungs to measure net alveolar fluid transport in intact animal and human lungs. However, intact lung studies have two limitations: the contribution of different distal lung epithelial cells cannot be studied separately, and the surface area for fluid absorption can only be approximated. Therefore, we developed a method to measure net vectorial fluid transport in cultured rat alveolar type II cells using an air-liquid interface. The cells were seeded on 0.4-μm microporous inserts in a Transwell system. At 96 h, the transmembrane electrical resistance reached a peak level (1,530 ± 115 Ω·cm2) with morphological evidence of tight junctions. We measured net fluid transport by placing 150 μl of culture medium containing 0.5 μCi of 131I-albumin on the apical side of the polarized cells. Protein permeability across the cell monolayer, as measured by labeled albumin, was 1.17 ± 0.34% over 24 h. The change in concentration of 131I-albumin in the apical fluid was used to determine the net fluid transported across the monolayer over 12 and 24 h. The net basal fluid transport was 0.84 μl·cm−2·h−1. cAMP stimulation with forskolin and IBMX increased fluid transport by 96%. Amiloride inhibited both the basal and stimulated fluid transport. Ouabain inhibited basal fluid transport by 93%. The cultured cells retained alveolar type II-like features based on morphologic studies, including ultrastructural imaging. In conclusion, this novel in vitro system can be used to measure net vectorial fluid transport across cultured, polarized alveolar epithelial cells.

scholarly journals Idebenone has preventative and therapeutic effects on pulmonary fibrosis via preferential suppression of fibroblast activity

2019 ◽  
Vol 5 (1) ◽  
Author(s):  
Toshifumi Sugizaki ◽  
Ken-ichiro Tanaka ◽  
Teita Asano ◽  
Daisuke Kobayashi ◽  
Yuuki Hino ◽  
...  
Keyword(s):  
Pulmonary Fibrosis ◽  
A549 Cells ◽  
Alveolar Epithelial Cells ◽  
Lung Fibroblasts ◽  
Epithelial Cell Line ◽  
Therapeutic Effects ◽  
Synthetic Analogue ◽  
Apoptosis Resistance ◽  
Collagen Production ◽  
Alveolar Epithelial

AbstractAlveolar epithelial injury induced by reactive oxygen species (ROS) and abnormal collagen production by activated fibroblasts (myofibroblasts) is involved in the onset and exacerbation of idiopathic pulmonary fibrosis (IPF). Compared with alveolar epithelial cells, lung fibroblasts, especially myofibroblasts, exhibit an apoptosis-resistance phenotype (apoptosis paradox) that appears to be involved in IPF pathogenesis. Thus, we screened for chemicals eliciting preferential cytotoxicity of LL29 cells (lung fibroblasts from an IPF patient) compared with A549 cells (human lung alveolar epithelial cell line) from medicines already in clinical use. We identified idebenone, a synthetic analogue of coenzyme Q10 (CoQ10, an antioxidant) that has been used clinically as a brain metabolic stimulant. Idebenone induced cell growth inhibition and cell death in LL29 cells at a lower concentration than in A549 cells, a feature that was not observed for other antioxidant molecules (such as CoQ10) and two IPF drugs (pirfenidone and nintedanib). Administration of idebenone prevented bleomycin-induced pulmonary fibrosis and increased pulmonary ROS levels. Importantly, idebenone also improved pulmonary fibrosis and lung function when administered after the development of fibrosis, whereas administration of CoQ10 similarly prevented bleomycin-induced pulmonary fibrosis, but had no effect after its development. Administration of idebenone, but not CoQ10, suppressed bleomycin-induced increases in lung myofibroblasts. In vitro, treatment of LL29 cells with idebenone, but not CoQ10, suppressed TGF-β–induced collagen production. These results suggest that in addition to antioxidant activity, idebenone exerts inhibitory activity on the function of lung fibroblasts, with the former activity being preventative and the latter therapeutic for bleomycin-induced fibrosis. Thus, we propose that idebenone may be more therapeutically beneficial for IPF patients than current treatments.

Secretion of cytokines by rat alveolar epithelial cells: possible regulatory role for SP-A

1994 ◽  
Vol 266 (2) ◽  
pp. L148-L155 ◽  
Author(s):  
H. Blau ◽  
S. Riklis ◽  
V. Kravtsov ◽  
M. Kalina
Keyword(s):  
Epithelial Cells ◽  
Alveolar Epithelial Cells ◽  
Alveolar Type ◽  
Type Ii Cells ◽  
Type Ii ◽  
Alveolar Type Ii Cells ◽  
Long Term Culture ◽  
Alveolar Epithelial ◽  
Gm Csf

Cultured alveolar type II cells and pulmonary epithelial (PE) cells in long-term culture were found to secrete colony-stimulating factors (CSF) into the medium in similar fashion to alveolar macrophages. CSF activity was determined by using the in vitro assay for myeloid progenitor cells [colony-forming units in culture (CFU-C)]. Both lipopolisaccharide (LPS) and interleukin-1 alpha (IL-1 alpha) were found to upregulate the secretion 6.5- to 8-fold from alveolar type II cells and macrophages. However, no stimulatory effect of these factors was observed in PE cells that release CSF into the medium constitutively, possibly due to the conditions of long-term culture. The CSF activity was partially neutralized (70% inhibition) by antibodies against murine granulocyte/macrophage (GM)-CSF and IL-3, thus indicating the presence of both GM-CSF and IL-3-like factors in the CSF. However, the presence of other cytokines in the CSF is highly probable. Surfactant-associated protein A (SP-A), which is known to play a central role in surfactant homeostasis and function, was also found to upregulate secretion of CSF (at concentrations of 0.1-5 micrograms/ml) from alveolar type II cells and macrophages. Control cells such as rat peritoneal macrophages, alveolar fibroblasts, and 3T3/NIH cell line could not be elicited by SP-A to release CSF. The results are discussed in relation to the possible participation of the alveolar epithelial cells in various intercellular signaling networks. Our studies suggest that alveolar type II cells and SP-A may play an important regulatory role in the modulation of immune and inflammatory effector cells within the alveolar space.

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