PM10-exposed macrophages stimulate a proinflammatory response in lung epithelial cells via TNF-α

2002 ◽  
Vol 282 (2) ◽  
pp. L237-L248 ◽  
Author(s):  
L. A. Jiménez ◽  
E. M. Drost ◽  
P. S. Gilmour ◽  
I. Rahman ◽  
F. Antonicelli ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Inflammatory Response ◽  
A549 Cells ◽  
Conditioned Media ◽  
Lung Epithelial Cells ◽  
Mrna Levels ◽  
Protein Levels ◽  
Proinflammatory Mediators ◽  
Tnf Α ◽  
Lung Epithelial

There is now considerable evidence for an association between the levels of particulate air pollution [particulate matter <10 μm in aerodynamic diameter (PM10)] and various adverse health endpoints. The release of proinflammatory mediators from PM10-exposed macrophages may be important in stimulating cytokine release from lung epithelial cells, thus amplifying the inflammatory response. A549 cells were treated with conditioned media from monocyte-derived macrophages stimulated with PM10, titanium dioxide (TiO2), or ultrafine TiO2. We demonstrate that only conditioned media from PM10-stimulated macrophages significantly increased nuclear factor-κB and activator protein-1 DNA binding, enhanced interleukin-8 (IL-8) mRNA levels as assessed by RT-PCR, and augmented IL-8 protein levels, over untreated controls. Furthermore, PM10-conditioned media also caused transactivation of IL-8 as determined by an IL-8-chloramphenicol acetyl transferase reporter. Analysis of these conditioned media revealed marked increases in tumor necrosis factor-α (TNF-α) and protein levels and enhanced chemotactic activity for neutrophils. Preincubation of conditioned media with TNF-α-neutralizing antibodies significantly reduced IL-8 production. These data suggest that PM10-activated macrophages may amplify the inflammatory response by enhancing IL-8 release from lung epithelial cells, in part, via elaboration of TNF-α.

Dual mechanism forMycobacterium tuberculosiscytotoxicity on lung epithelial cells

2012 ◽  
Vol 58 (7) ◽  
pp. 909-916 ◽  
Author(s):  
Jorge Castro-Garza ◽  
W. Edward Swords ◽  
Russell K. Karls ◽  
Frederick D. Quinn
Keyword(s):  
Epithelial Cell ◽  
Epithelial Cells ◽  
Neutralizing Antibodies ◽  
Cytotoxic Effect ◽  
Alveolar Epithelial Cell ◽  
A549 Cells ◽  
Lung Epithelial Cells ◽  
Necrosis Factor Alpha ◽  
Tnf Α ◽  
Lung Epithelial

Mycobacterium tuberculosis strains CDC1551 and Erdman were used to assess cytotoxicity in infected A549 human alveolar epithelial cell monolayers. Strain CDC1551 was found to induce qualitatively greater disruption of A549 monolayers than was strain Erdman, although total intracellular and cell-associated bacterial growth rates over the course of the infections were not significantly different. Cell-free culture supernatants from human monocytic cells infected with either of the 2 M. tuberculosis strains produced a cytotoxic effect on A549 cells, correlating with the amount of tumor necrosis factor alpha (TNF-α) released by the infected monocytes. The addition of TNF-α-neutralizing antibodies to the supernatants from infected monocyte cultures did prevent the induction of a cytotoxic effect on A549 cells overlaid with this mixture but did not prevent the death of epithelial cells when added prior to infection with M. tuberculosis bacilli. Thus, these data agree with previous observations that lung epithelial cells infected with M. tuberculosis bacilli are rapidly killed in vitro. In addition, the data indicate that some of the observed epithelial cell killing may be collateral damage; the result of TNF-α released from M. tuberculosis-infected monocytes.

Contribution of IL-1β and TNF-α to the initiation of the peripheral lung response to atmospheric particulates (PM10)

2004 ◽  
Vol 287 (1) ◽  
pp. L176-L183 ◽  
Author(s):  
Hiroshi Ishii ◽  
Takeshi Fujii ◽  
James C. Hogg ◽  
Shizu Hayashi ◽  
Hiroshi Mukae ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
A549 Cells ◽  
Lung Epithelial Cells ◽  
Vegf Mrna ◽  
Atmospheric Particulates ◽  
Proinflammatory Mediators ◽  
Acute Response ◽  
Chemotactic Protein ◽  
Tnf Α ◽  
Macrophage Colony

Alveolar macrophages (AM) play a key role in clearing atmospheric particulates from the lung surface and stimulating epithelial cells to produce proinflammatory mediators. The present study examines the role of “acute response” cytokines TNF-α and IL-1β released by AM exposed to ambient particulate matter with a diameter of <10 μm (PM10) in amplifying the proinflammatory mediator expression by A549 cells and human bronchial epithelial cells (HBEC). The results showed that supernatants from human AM incubated 24 h with PM10(100 μg/ml) contained more TNF-α, IL-1β, granulocyte-macrophage colony stimulating factor, IL-6, and IL-8 than nonexposed AM supernatants. The 3-h treatment of A549 cells with PM10-exposed AM supernatants increased TNF-α, IL-1β, IL-8, regulated on activation normal T-cells expressed and secreted (RANTES), and leukemia inhibitory factor mRNA compared with the treatment with nonexposed AM supernatants and, compared with untreated A549 cells, additionally increased ICAM-1 and monocyte chemotactic protein-1 mRNA. Preincubating PM10-exposed AM supernatants with anti-IL-1β antibodies reduced all the above mediators as well as VEGF mRNA expression ( P < 0.05), while anti-TNF-α antibodies were less effective ( P > 0.05), and the combination of the two antibodies most effective. When HBEC were treated similarly, anti-TNF-α antibodies had the greatest effect. In A549 cells PM10-exposed AM supernatants increased NF-κB, activator protein (AP)-1 and specificity protein 1 binding, while anti-TNF-α and anti-IL-1β antibodies reduced NF-κB and AP-1 binding. We conclude that AM-derived TNF-α and IL-1β provide a major stimulus for the production of proinflammatory mediators by lung epithelial cells and that their relative importance may depend on the type of epithelial cell target.

The effects of PM10 particles and oxidative stress on macrophages and lung epithelial cells: modulating effects of calcium-signaling antagonists

2007 ◽  
Vol 292 (6) ◽  
pp. L1444-L1451 ◽  
Author(s):  
David M. Brown ◽  
Laura Hutchison ◽  
Kenneth Donaldson ◽  
Vicki Stone
Keyword(s):  
Oxidative Stress ◽  
Calcium Antagonist ◽  
Epithelial Cells ◽  
Calcium Signaling ◽  
Mononuclear Cells ◽  
Human Peripheral Blood ◽  
A549 Cells ◽  
Lung Epithelial Cells ◽  
Tnf Α ◽  
Lung Epithelial

We have previously examined the ability of air pollution particles (PM10) to promote release of the proinflammatory cytokine tumor necrosis factor-α (TNF-α) from human peripheral blood mononuclear cells and demonstrated a role for calcium as a signaling molecule in this process. We have now studied the ability of oxidative stress induced by a synthetic oxidant tert-butyl hydroperoxide (tBHP) to induce TNF-α production via calcium signaling in the mouse macrophage cell line (J774). The oxidant tBHP significantly increased intracellular calcium and the release of TNF-α in J774 cells, an effect that was reduced to control levels by inhibition of calcium signaling with verapamil, BAPTA-AM, and W-7. This study also investigated interactions between PM10-treated macrophages and epithelial cells by using conditioned medium (CM) from PM10-treated mononuclear cells to stimulate the release of the neutrophil chemoattractant chemokine IL-8 from A549 lung epithelial cells. TNF-α protein release was demonstrated in human mononuclear cells after PM10 treatment, an effect that was inhibited by calcium antagonists. Treatment of A549 cells with monocyte/PM10 CM produced increased IL-8 release that was reduced with CM from monocyte/PM10/calcium antagonist treatments. The expression of ICAM-1 was increased after incubation with CM from monocyte/PM10 treatment, and this increase was prevented by treatment with CM from monocyte/PM10/calcium antagonist. These data demonstrate a link between oxidative stress, calcium, and inflammatory mediator production in macrophages and lung epithelial cells.

TNF-α inhibits SP-A gene expression in lung epithelial cells via p38 MAPK

2002 ◽  
Vol 283 (2) ◽  
pp. L418-L427 ◽  
Author(s):  
Olga L. Miakotina ◽  
Jeanne M. Snyder
Keyword(s):  
Gene Expression ◽  
Epithelial Cells ◽  
P38 Mapk ◽  
Mapk Pathway ◽  
Lung Epithelial Cells ◽  
Mitogen Activated Protein Kinase ◽  
Mrna Levels ◽  
Dependent Manner ◽  
Tnf Α ◽  
Lung Epithelial

Surfactant protein A (SP-A), the major lung surfactant-associated protein, mediates local defense against pathogens and modulates inflammation in the alveolus. Tumor necrosis factor (TNF)-α, a proinflammatory cytokine, inhibits SP-A gene expression in lung epithelial cells. Inhibitors of the phosphatidylinositol 3-kinase pathway, i.e., wortmannin, LY-294002, and rapamycin, did not block the inhibitory effects of TNF-α on SP-A mRNA levels. An inhibitor of the p44/42 mitogen-activated protein kinase (MAPK) pathway, PD-98059, was also ineffective. PD-169316 and SB-203580, inhibitors of p38 MAPK, blocked the TNF-α-mediated inhibition of SP-A mRNA levels. TNF-α increased the phosphorylation of p38 MAPK within 15 min. Anisomycin, an activator of p38 MAPK, increased p38 MAPK phosphorylation and decreased SP-A mRNA levels in a dose-dependent manner. Finally, TNF-α increased the phosphorylation of ATF-2, a transcription factor that is a p38 MAPK substrate. We conclude that TNF-α downregulates SP-A gene expression in lung epithelial cells via the p38 MAPK signal transduction pathway.

scholarly journals Ethanol Decreases Inflammatory Response in Human Lung Epithelial Cells by Inhibiting the Canonical NF-kB-Pathway

2017 ◽  
Vol 43 (1) ◽  
pp. 17-30 ◽  
Author(s):  
Katharina Mörs ◽  
Jason-Alexander Hörauf ◽  
Shinwan Kany ◽  
Nils Wagner ◽  
Ramona Sturm ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Inflammatory Response ◽  
Acute Inflammation ◽  
Human Lung ◽  
A549 Cells ◽  
Lung Epithelial Cells ◽  
Neutrophil Adhesion ◽  
Lung Epithelial ◽  
Nuclear Levels ◽  
Human Lung Epithelial Cells

Background/Aims: Alcohol (ethanol, EtOH) as significant contributor to traumatic injury is linked to suppressed inflammatory response, thereby influencing clinical outcomes. Alcohol-induced immune-suppression during acute inflammation (trauma) was linked to nuclear factor-kappaB (NF-ĸB). Here, we analyzed alcohol`s effects and mechanisms underlying its influence on NF-ĸB-signaling during acute inflammation in human lung epithelial cells. Methods: A549-cells were stimulated with interleukin (IL)-1β, or sera from trauma patients (TP) or healthy volunteers, with positive/negative blood alcohol concentrations (BAC), and subsequently exposed to EtOH (170 Mm, 1h). IL-6-release and neutrophil adhesion to A549 were analyzed. Specific siRNA-NIK mediated downregulation of non-canonical, and IKK-NBD-inhibition of canonical NF-ĸB signaling were performed. Nuclear levels of activated p50 and p52 NF-ĸB-subunits were detected using TransAm ELISA. Results: Both stimuli significantly induced IL-6-release (39.79±4.70 vs. 0.58±0.8 pg/ml) and neutrophil adhesion (132.30±8.80 vs. 100% control, p<0.05) to A549-cells. EtOH significantly decreased IL-6-release (22.90±5.40, p<0.05) and neutrophil adherence vs. controls (105.40±14.5%, p<0.05). IL-1β-induced significant activation of canonical/p50 and non-canonical/p52 pathways. EtOH significantly reduced p50 (34.90±23.70 vs. 197.70±36.43, p<0.05) not p52 activation. Inhibition of canonical pathway was further increased by EtOH (less p50-activation), while p52 remained unaltered. Inhibition of non-canonical pathway was unchanged by EtOH. Conclusion: Here, alcohol`s anti-inflammatory effects are mediated via decreasing nuclear levels of activated p50-subunit and canonical NF-ĸB signaling pathway.

scholarly journals Effect of Alpha-1 Antitrypsin on CFTR Levels in Primary Human Airway Epithelial Cells Grown at the Air-Liquid-Interface

Molecules ◽  
2021 ◽  
Vol 26 (9) ◽  
pp. 2639
Author(s):  
Frauke Stanke ◽  
Sabina Janciauskiene ◽  
Stephanie Tamm ◽  
Sabine Wrenger ◽  
Ellen Luise Raddatz ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Human Lung ◽  
Liquid Interface ◽  
Lung Epithelial Cells ◽  
Western Blots ◽  
Mesenchymal Transition ◽  
Protein Levels ◽  
Lung Epithelial ◽  
Cftr Protein ◽  
Air Liquid Interface

The cystic fibrosis transmembrane conductance regulator (CFTR) gene is influenced by the fundamental cellular processes like epithelial differentiation/polarization, regeneration and epithelial–mesenchymal transition. Defects in CFTR protein levels and/or function lead to decreased airway surface liquid layer facilitating microbial colonization and inflammation. The SERPINA1 gene, encoding alpha1-antitrypsin (AAT) protein, is one of the genes implicated in CF, however it remains unknown whether AAT has any influence on CFTR levels. In this study we assessed CFTR protein levels in primary human lung epithelial cells grown at the air-liquid-interface (ALI) alone or pre-incubated with AAT by Western blots and immunohistochemistry. Histological analysis of ALI inserts revealed CFTR- and AAT-positive cells but no AAT-CFTR co-localization. When 0.5 mg/mL of AAT was added to apical or basolateral compartments of pro-inflammatory activated ALI cultures, CFTR levels increased relative to activated ALIs. This finding suggests that AAT is CFTR-modulating protein, albeit its effects may depend on the concentration and the route of administration. Human lung epithelial ALI cultures provide a useful tool for studies in detail how AAT or other pharmaceuticals affect the levels and activity of CFTR.

Nitric oxide increases IL-8 gene transcription and mRNA stability to enhance IL-8 gene expression in lung epithelial cells

2004 ◽  
Vol 287 (4) ◽  
pp. L764-L773 ◽  
Author(s):  
Loretta Sparkman ◽  
Vijayakumar Boggaram
Keyword(s):  
Gene Expression ◽  
Nitric Oxide ◽  
Protein Kinase ◽  
Epithelial Cells ◽  
Gene Transcription ◽  
Mrna Stability ◽  
Inflammatory Responses ◽  
Lung Epithelial Cells ◽  
Mrna Levels ◽  
Lung Epithelial

Interleukin (IL)-8, a C-X-C chemokine, is a potent chemoattractant and an activator for neutrophils, T cells, and other immune cells. The airway and respiratory epithelia play important roles in the initiation and modulation of inflammatory responses via production of cytokines and surfactant. The association between elevated levels of nitric oxide (NO) and IL-8 in acute lung injury associated with sepsis, acute respiratory distress syndrome, respiratory syncytial virus infection in infants, and other inflammatory diseases suggested that NO may play important roles in the control of IL-8 gene expression in the lung. We investigated the role of NO in the control of IL-8 gene expression in H441 lung epithelial cells. We found that a variety of NO donors significantly induced IL-8 mRNA levels, and the increase in IL-8 mRNA was associated with an increase in IL-8 protein. NO induction of IL-8 mRNA was due to increases in IL-8 gene transcription and mRNA stability. NO induction of IL-8 mRNA levels was not inhibited by 1H-[1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one and KT-5823, inhibitors of soluble guanylate cyclase and protein kinase G, respectively, and 8-bromo-cGMP did not increase IL-8 mRNA levels. This indicated that NO induces IL-8 mRNA levels independently of changes in the intracellular cGMP levels. NO induction of IL-8 mRNA was significantly reduced by inhibitors of extracellular regulated kinase and protein kinase C. IL-8 induction by NO was also reduced by hydroxyl radical scavengers such as dimethyl sulfoxide and dimethylthiourea, indicating the involvement of hydroxyl radicals in the induction process. NO induction of IL-8 gene expression could be a significant contributing factor in the initiation and induction of inflammatory response in the respiratory epithelium.

scholarly journals PPARγ inhibits airway epithelial cell inflammatory response through a MUC1-dependent mechanism

2012 ◽  
Vol 302 (7) ◽  
pp. L679-L687 ◽  
Author(s):  
Yong Sung Park ◽  
Erik P. Lillehoj ◽  
Kosuke Kato ◽  
Choon Sik Park ◽  
Kwang Chul Kim
Keyword(s):  
Epithelial Cells ◽  
Culture Media ◽  
A549 Cells ◽  
Luciferase Reporter ◽  
Muc1 Expression ◽  
Mrna Levels ◽  
Airway Epithelial ◽  
Pparγ Agonist ◽  
Tnf Α ◽  
Primary Mouse

This study was conducted to examine the relationship between the peroxisome proliferator-associated receptor-γ (PPARγ) and MUC1 mucin, two anti-inflammatory molecules expressed in the airways. Treatment of A549 lung epithelial cells or primary mouse tracheal surface epithelial (MTSE) cells with phorbol 12-myristate 13-acetate (PMA) increased the levels of tumor necrosis factor (TNF)-α in cell culture media compared with cells treated with vehicle alone. Overexpression of MUC1 in A549 cells decreased PMA-stimulated TNF-α levels, whereas deficiency of Muc1 expression in MTSE cells from Muc1 null mice increased PMA-induced TNF-α levels. Treatment of A549 or MTSE cells with the PPARγ agonist troglitazone (TGN) blocked the ability of PMA to stimulate TNF-α levels. However, the effect of TGN required the presence of MUC1/Muc1, since no differences in TNF-α levels were seen between PMA and PMA plus TGN in MUC1/Muc1-deficient cells. Similarly, whereas TGN decreased interleukin-8 (IL-8) levels in culture media of MUC1-expressing A549 cells treated with Pseudomonas aeruginosa strain K (PAK), no differences in IL-8 levels were seen between PAK and PAK plus TGN in MUC1-nonexpressing cells. EMSA confirmed the presence of a PPARγ-binding element in the MUC1 gene promoter. Finally, TGN treatment of A549 cells increased MUC1 promoter activity measured using a MUC1-luciferase reporter gene, augmented MUC1 mRNA levels by quantitative RT-PCR, and enhanced MUC1 protein expression by Western blot analysis. These combined data are consistent with the hypothesis that PPARγ stimulates MUC1/Muc1 expression, thereby blocking PMA/PAK-induced TNF-α/IL-8 production by airway epithelial cells.

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