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Open Access Research

Nitric oxide and superoxide mediate diesel particle effects in cytokine-treated mice and murine lung epithelial cells — implications for susceptibility to traffic-related air pollution

Nicholas D Manzo15*, Adriana J LaGier23, Ralph Slade3, Allen D Ledbetter4, Judy H Richards3 and Janice A Dye3

Author Affiliations

1 Department of Molecular Biomedical Sciences, College of Veterinary Medicine, North Carolina State University, Raleigh, NC, 27606, USA

2 Department of Biological Sciences, College of Arts and Sciences, Florida Gulf Coast University, 10501 FGCU Blvd, Fort Myers, FL, 33965, USA

3 Cardiopulmonary and Immunotoxicology Branch (CIB), Environmental Public Health Division, National Health and Environmental Effects Research Laboratory, ORD, U.S. Environmental Protection Agency, Research Triangle Park, NC, 27711, USA

4 Inhalation Toxicology Facilities Branch (ITFB), Environmental Public Health Division, National Health and Environmental Effects Research Laboratory, ORD, U.S. Environmental Protection Agency, Research Triangle Park, NC, 27711, USA

5 Current address: Division of Pulmonary, Allergy and Critical Care, Duke University Medical Center, 2075 MSRBII, 106 Research Drive, DUMC Box 103000, Durham, NC, 27710, USA

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Particle and Fibre Toxicology 2012, 9:43  doi:10.1186/1743-8977-9-43

Published: 15 November 2012

Abstract

Background

Epidemiologic studies associate childhood exposure to traffic-related air pollution with increased respiratory infections and asthmatic and allergic symptoms. The strongest associations between traffic exposure and negative health impacts are observed in individuals with respiratory inflammation. We hypothesized that interactions between nitric oxide (NO), increased during lung inflammatory responses, and reactive oxygen species (ROS), increased as a consequence of traffic exposure ─ played a key role in the increased susceptibility of these at-risk populations to traffic emissions.

Methods

Diesel exhaust particles (DEP) were used as surrogates for traffic particles. Murine lung epithelial (LA-4) cells and BALB/c mice were treated with a cytokine mixture (cytomix: TNFα, IL-1β, and IFNγ) to induce a generic inflammatory state. Cells were exposed to saline or DEP (25 μg/cm2) and examined for differential effects on redox balance and cytotoxicity. Likewise, mice undergoing nose-only inhalation exposure to air or DEP (2 mg/m3 × 4 h/d × 2 d) were assessed for differential effects on lung inflammation, injury, antioxidant levels, and phagocyte ROS production.

Results

Cytomix treatment significantly increased LA-4 cell NO production though iNOS activation. Cytomix +  DEP-exposed cells incurred the greatest intracellular ROS production, with commensurate cytotoxicity, as these cells were unable to maintain redox balance. By contrast, saline + DEP-exposed cells were able to mount effective antioxidant responses. DEP effects were mediated by: (1) increased ROS including superoxide anion (O2˙-), related to increased xanthine dehydrogenase expression and reduced cytosolic superoxide dismutase activity; and (2) increased peroxynitrite generation related to interaction of O2˙- with cytokine-induced NO. Effects were partially reduced by superoxide dismutase (SOD) supplementation or by blocking iNOS induction. In mice, cytomix +  DEP-exposure resulted in greater ROS production in lung phagocytes. Phagocyte and epithelial effects were, by and large, prevented by treatment with FeTMPyP, which accelerates peroxynitrite catalysis.

Conclusions

During inflammation, due to interactions of NO and O2˙-, DEP-exposure was associated with nitrosative stress in surface epithelial cells and resident lung phagocytes. As these cell types work in concert to provide protection against inhaled pathogens and allergens, dysfunction would predispose to development of respiratory infection and allergy. Results provide a mechanism by which individuals with pre-existing respiratory inflammation are at increased risk for exposure to traffic-dominated urban air pollution.

Keywords:
Traffic; Diesel; Particles; Epithelial cells; Phagocytes; Nitric oxide; Peroxynitrite; Redox balance