Involvement of superoxide and nitric oxide on airway inflammation and hyperresponsiveness induced by diesel exhaust particles in mice

doi:10.1016/S0891-5849(98)00073-2

Free Radical Biology and Medicine

Volume 25, Issue 6, October 1998, Pages 635-644

https://doi.org/10.1016/S0891-5849(98)00073-2 Get rights and content

Abstract

We previously demonstrated that chronic intratracheal instillation of diesel exhaust particles (DEP) induces airway inflammation and hyperresponsiveness in the mouse, and that these effects were partially reversed by the administration of superoxide dismutase (SOD). In the present study, we have investigated the involvement of superoxide in DEP-induced airway response by analyzing the localization and activity of two enzymes: (1) a superoxide producer, NADPH cytochrome P-450 reductase (P-450 reductase), and (2) a superoxide scavenger, SOD, in the lungs of the exposed mice and controls. P-450 reductase was detected mainly in ciliated cells and clara cells; its activity was increased by the repeated intratracheal instillation of DEP. While CuZn-SOD and Mn-SOD were also present in the airway epithelium, their activity was significantly decreased following DEP instillation. Exposure to DEP doubled the level of nitric oxide (NO) in the exhaled air. DEP exposure also increased the level of constitutive NO synthase (cNOS) in the airway epithelium and inducible NO synthase (iNOS) in the macrophages. Pretreatment with N-G-monomethyl L-arginine, a nonspecific inhibitor of NO synthase, significantly reduced the airway hyperresponsiveness induced by DEP. These results indicate that superoxide and NO may each contribute to the airway inflammation and hyperresponsiveness induced by the repeated intratracheal instillation of DEP in mice.

Introduction

Pathophysiology of asthma is characterized by airway hyperresponsiveness, infiltration of the airways by inflammatory cells, and the release of numerous mediators from the cells [1]. Among the mediators released, reactive oxygen species such as superoxide (O₂⁻), hydrogen peroxide and hydroxyl radical (·OH) are important [1], [2] in the airway inflammation and hyperresponsiveness [3]. We have previously reported that repeated instillation of diesel exhaust particles (DEP) into the trachea of mice led to inflammation and hyperresponsiveness of the airway, as well as hypersecretion of mucus [4], [5], [6]. Our studies demonstrated that pretreatment of the animals with superoxide dismutase (SOD) suppressed these DEP-induced responses, and that the DEP-induced production of O₂⁻ and radical dot OH resulted from nonenzymatic [5] and enzymatic [6] reactions.

Nitric oxide (NO) is another important mediator that may cause airway inflammation and hyperresponsiveness [7], [8], [9], [10], [11]. This mediator is synthesized from L-arginine by several types of NO synthase in pulmonary cells, including inflammatory, endothelial and airway epithelial cells [7], [8]. The reaction between NO and superoxide leads to the production of the toxic substance, peroxynitrite [12], [13], [14]; however, the role of NO in the progression of DEP-induced airway inflammation and hyperresponsiveness has not yet been established.

Accordingly, we investigated the roles of superoxide and NO in airway inflammation and hyperresponsiveness by analyzing the distribution and activity of several superoxide- and NO-related enzymes in the lungs of mice that had received repeated intratracheal instillations of DEP.

Section snippets

Chemicals and reagents

N-[2-hydroxyethyl] piperazine-N′-[2-ethane sulfonic acid] (Hepes), 3,3-diaminobenzidine tetrahydropyridine (DBA), xanthine, and xanthine oxidase were purchased from Sigma Chemical Co. (St. Louis, MO, USA). NADPH, N-G-monomethyl L-arginine (L-NMMA), and cytochrome c were obtained from Wako Pure Chemical Co. Ltd. (Tokyo, Japan). Tween 80 and polyethylene glycol (PEG) were purchased from Nacalai Tesque, Inc. (Kyoto, Japan). Polyethylene glycol-conjugated Mn-superoxide dismutase from Aspergillus

Assessment of DEP-induced airway inflammation

We first investigated the infiltration of eosinophils and neutrophils into the submucosal layer of the airways in DEP-treated mice. Repeated intratracheal instillation of either 0.1 or 0.2 mg of DEP once a week for 10 weeks resulted in a 7-fold increase in the mean number of eosinophils, compared with saline-treated mice (p< .01, Fig. 1). Pretreatment with PEG-SOD inhibited the DEP-induced infiltration of eosinophils. In contrast, pretreatment with PEG alone did not affect the DEP-induced

Discussion

The present study investigated the roles of O₂⁻ and NO in airway inflammation and hyperresponsiveness induced by exposure to DEP. We analyzed the distribution and activity of O₂⁻ and NO-related enzymes in the lungs of mice that had received repeated intratracheal instillations of DEP.

The activity of P-450 reductase, as a representative of O₂⁻ producing enzymes, was increased by the DEP-treatment, whereas the activity of SOD as O₂⁻ scavenging enzymes were decreased. The level of NO in the

References (45)

P.J. Barnes
Reactive oxygen species and airway inflammation
Free Radic. Biol. Med.
(1990)
C.J.A. Doelman et al.
Oxygen radicals in lung pathology
Free Radic. Biol. Med.
(1990)
M. Sagai et al.
Biological effects of diesel exhaust particles (DEP). III. Pathogenesis of asthma like symptoms in mice
Free Radic. Biol. Med.
(1996)
M. Sagai et al.
Biological effect of diesel exhaust particles. I. In vitro production of superoxide and in vivo toxicity in mouse
Free Radic. Biol. Med.
(1993)
O.H. Lowry et al.
Protein measurement with the Foline-phenol reagent
J. Biol. Chem.
(1951)
C.H. Williams et al.
Microsomal triphosphopyridine nucleotide-cytochrome c reductase of liver
J. Biol. Chem.
(1962)
J.M. McCord et al.
Superoxide dismutase, an enzymatic function for erythrocuprein (hemocuprein)
J. Biol. Chem.
(1969)
M. Cluzel et al.
Enhanced alveolar cell luminol-dependent chemiluminescence in asthma
J. Allergy Clin. Immunol.
(1987)
F. Engels et al.
Pulmonary macrophages induce deterioration of guinea pig tracheal beta-adrenergic function through release of oxygen radicals
Eur. J. Pharmacol.
(1985)
V. Berlin et al.
Reduction of adriamycin to a semiquinone free radical by NADPH cytochrome P-450 reductase produces DNA cleavage in a reaction mediated by molecular oxygen
J. Biol. Chem.
(1981)

H. Kappus

Overview of enzyme systems involved in bioreduction of drugs and in redox cycling

Biochem. Pharmacol.

(1986)

T. Ichinose et al.

Biological effects of diesel exhaust particles (DEP). II. Acute toxicity of DEP introduced into lung by intratracheal instillation

Toxicology

(1995)

E. Frigas et al.

The eosinophil and pathophysiology of asthma

J. Allergy Clin. Immunol.

(1986)

G.J. Gleich

The eosinophil and bronchial asthmacurrent understanding

J. Allergy Clin. Immunol.

(1990)

S. Horie et al.

Cytokines directly induce degranulation and superoxide production from human eosinophils

J. Allergy Clin. Immunol.

(1996)

S.A. Kharitonov et al.

Increased nitric oxide in exhaled air of asthmatic patients

Lancet

(1994)

M.G. Persson et al.

Single-breath nitric oxide measurements in asthmatic patients and smokers

Lancet

(1994)

P.J. Barnes et al.

Inflammatory mediators and asthma

Pharmacol. Rev.

(1988)

Y. Kumagai et al.

Generation of reactive oxygen species during interaction of diesel exhaust particle components with NADPH-cytochrome P-450 reductase and involvement of the bioactivation in the DNA damage

Free Radic. Biol. Med.

(1996)

B. Gaston et al.

The biology of nitrogen oxides in the airways

Am. J. Respir. Crit. Care Med.

(1994)

P.J. Barnes

Nitric oxide and airways

Eur. J. Respir.

(1993)

P.J. Barnes et al.

Nitric oxide and asthmatic inflammation

Immunol. Today

(1995)

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Original ContributionsInvolvement of superoxide and nitric oxide on airway inflammation and hyperresponsiveness induced by diesel exhaust particles in mice

Abstract

Introduction

Section snippets

Chemicals and reagents

Assessment of DEP-induced airway inflammation

Discussion

Free Radic. Biol. Med.

Free Radic. Biol. Med.

Free Radic. Biol. Med.

Free Radic. Biol. Med.

J. Biol. Chem.

J. Biol. Chem.

J. Biol. Chem.

J. Allergy Clin. Immunol.

Eur. J. Pharmacol.

J. Biol. Chem.

Biochem. Pharmacol.

Toxicology

J. Allergy Clin. Immunol.

J. Allergy Clin. Immunol.

J. Allergy Clin. Immunol.

Lancet

Lancet

Inflammatory mediators and asthma

Pharmacol. Rev.

Generation of reactive oxygen species during interaction of diesel exhaust particle components with NADPH-cytochrome P-450 reductase and involvement of the bioactivation in the DNA damage

Free Radic. Biol. Med.

The biology of nitrogen oxides in the airways

Am. J. Respir. Crit. Care Med.

Nitric oxide and airways

Eur. J. Respir.

Nitric oxide and asthmatic inflammation

Immunol. Today

Original Contributions
Involvement of superoxide and nitric oxide on airway inflammation and hyperresponsiveness induced by diesel exhaust particles in mice