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Oxidant-mediated increases in redox factor-1 nuclear protein and activator protein-1 DNA binding in asbestos-treated
Dawn M Flaherty1, Martha M Monick, A Brent Carter
1Department of Internal Medicine, University of Iowa College of Medicine and Veterans Affairs Medical Center, Iowa City, IA 52243, USA. flahertydm@mail.medicine.uiowa.edu
Abstract:
Alveolar macrophages have been implicated in the pathogenesis of a number of acute and chronic lung disorders. We have previously shown that normal human alveolar macrophages exhibit decreased DNA binding activity of the transcription factor, AP-1, compared with monocytes. Furthermore, this decrease in AP-1 DNA binding appears to be due to a decrease in the redox active protein, redox factor (Ref)-1. Ref-1 is an important redox regulator of a number of transcription factors, including NF-kappaB and AP-1. In this study we evaluated the role of asbestos, a prototypic model of chronic fibrotic lung disease, in Ref-1 expression and activity. We found that incubation with low concentrations of crocidolite asbestos (0.5-1.25 microg/cm(2)) resulted in an increase in nuclear Ref-1 protein after 5 min, with a persistent elevation in protein up to 24 h. Additionally, an increase in nuclear Ref-1 could be induced by treating the cells with an oxidant-generating stimulus (iron loading plus PMA) and inhibited by diphenyleneiodonium chloride, an inhibitor of NADPH oxidase. The asbestos-induced accumulation of nuclear Ref-1 was associated with an increase in AP-1 DNA binding activity. These findings suggest that an exposure associated with fibrotic lung disease, i.e., asbestos, modulates accumulation of nuclear Ref-1 in macrophages, and that this effect is mediated by an oxidant stimulus.
Insights
Asbestos exposure increases nuclear redox factor-1 (Ref-1) protein in lung macrophages, enhancing activator protein-1 (AP-1) DNA binding. This suggests an oxidant mechanism in asbestos-induced fibrotic lung disease pathogenesis.
Area of Science:
- Cell Biology
- Immunology
- Toxicology
Background:
- Alveolar macrophages play a role in lung disorders.
- Reduced activator protein-1 (AP-1) DNA binding in macrophages is linked to decreased redox factor-1 (Ref-1).
- Ref-1 regulates transcription factors like AP-1 and NF-kappaB.
Purpose of the Study:
- To investigate the role of asbestos, a model for fibrotic lung disease, in Ref-1 expression and activity.
- To understand the mechanism behind asbestos-induced changes in Ref-1.
Main Methods:
- Incubation of human alveolar macrophages with crocidolite asbestos.
- Treatment with oxidant-generating stimuli and NADPH oxidase inhibitors.
- Measurement of nuclear Ref-1 protein levels and AP-1 DNA binding activity.
Main Results:
- Low concentrations of asbestos increased nuclear Ref-1 protein within minutes, with sustained elevation up to 24 hours.
- Asbestos-induced Ref-1 accumulation was mediated by an oxidant stimulus, inhibited by diphenyleneiodonium chloride.
- Increased nuclear Ref-1 correlated with enhanced AP-1 DNA binding activity.
Conclusions:
- Asbestos exposure modulates nuclear Ref-1 accumulation in macrophages.
- This modulation is mediated by an oxidant stimulus.
- Findings suggest a role for Ref-1 and oxidative stress in asbestos-related fibrotic lung disease.