Proteinase-Activated Receptor-2–Triggered Prostaglandin E2 Release, but Not Cyclooxygenase-2 Upregulation, Requires Activation of the Phosphatidylinositol 3–Kinase / Akt / Nuclear Factor-κB Pathway in Human Alveolar Epithelial Cells

Abstract Objective Acute lung injury (ALI) is characterized by inflammation and oxidative stress. Nuclear factor-kappaB (NF-κB) mediates the expression of various inflammation-related genes, including the NADPH oxidase family. This study aimed to identify the potential regulatory role of NF-κB on NADPH oxidases in tumor necrosis factor-α (TNF-α)-induced oxidative stress in human alveolar epithelial cells. Methods A549 cells were treated with TNF-α for 24 h to establish ALI cell models. RT-PCR, western blot, assessment of oxidative stress, Alibaba 2.1 online analysis, electrophoretic mobility shift assays and luciferase reporter analysis were employed to identify the potential regulatory role of NF-κB on NADPH oxidases in TNF-α-induced oxidative stress in human alveolar epithelial cells. Results The expression of NF-κB/p65 was notably upregulated in TNF-α-stimulated A549 cells. NF-κB knockdown by siRNA significantly inhibited the TNF-α-induced oxidative stress. Moreover, NF-κB/p65 siRNA could inhibit the activation of NOX1, NOX2 and NOX4 mRNA and protein expression in TNF-α-stimulated A549 cells. The next study demonstrated that NF-κB activated the transcription of NOX1 by binding to the -261 to -252 bp (NOX1/κB2, TAAAAATCCC) region of NOX1 promoter in TNF-α-stimulated A549 cells. Conclusion Our data demonstrated that NF-κB can aggravate TNF-α-induced ALI by regulating the oxidative stress response and the expression of NOX1, NOX2 and NOX4. Moreover, NF-κB could promote the NOX1 transcriptional activity via binding its promoter in TNF-α-stimulated A549 cells.

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Mediator generation and signaling events in alveolar epithelial cells attacked byS. aureusα-toxin

AJP Lung Cellular and Molecular Physiology ◽

10.1152/ajplung.00156.2001 ◽

2002 ◽

Vol 282 (2) ◽

pp. L207-L214 ◽

Cited By ~ 45

Author(s):

Frank Rose ◽

Gabriele Dahlem ◽

Bernd Guthmann ◽

Friedrich Grimminger ◽

Ulrich Maus ◽

...

Keyword(s):

Protein Kinase C ◽

Protein Kinase ◽

Epithelial Cells ◽

Nuclear Factor Κb ◽

Alveolar Epithelial Cells ◽

Target Cells ◽

Pyrrolidine Dithiocarbamate ◽

Nuclear Factor ◽

Alveolar Epithelial ◽

Kinase C

Staphylococcus aureus α-toxin is a pore-forming bacterial exotoxin that has been implicated as a significant virulence factor in human staphylococcal diseases. In primary cultures of rat pneumocyte type II cells and the human A549 alveolar epithelial cell line, purified α-toxin provoked rapid-onset phosphatidylinositol (PtdIns) hydrolysis as well as liberation of nitric oxide and the prostanoids PGE2, PGI2, and thromboxane A2. In addition, sustained upregulation of proinflammatory interleukin (IL)-8 mRNA expression and protein secretion occurred. “Priming” with low-dose IL-1β markedly enhanced the IL-8 response to α-toxin, which was then accompanied by IL-6 appearance. The cytokine response was blocked by the intracellular Ca2+-chelating reagent 1,2-bis(2-aminophenoxy)-ethane- N,N,N′ ,N′-tetraacetic acid, the protein kinase C inhibitor bis-indolyl maleimide I, as well as two independent inhibitors of nuclear factor-κB activation, pyrrolidine dithiocarbamate and caffeic acid phenethyl ester. We conclude that alveolar epithelial cells are highly reactive target cells of staphylococcal α-toxin. α-Toxin pore-associated transmembrane Ca2+flux and PtdIns hydrolysis-related signaling with downstream activation of protein kinase C and nuclear translocation of nuclear factor-κB are suggested to represent important underlying mechanisms. Such reactivity of the alveolar epithelial cells may be relevant for pathogenic sequelae in staphylococcal lung disease.

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