2001 Progress Report: Pro-inflammatory and the Pro-oxidative Effects of Diesel Exhaust Particulate in Vivo and in Vitro

EPA Grant Number: R827352C002
Subproject: this is subproject number 002 , established and managed by the Center Director under grant R827352
(EPA does not fund or establish subprojects; EPA awards and manages the overall grant for this center).

Center: Southern California Particle Center and Supersite
Center Director: Froines, John R.
Title: Pro-inflammatory and the Pro-oxidative Effects of Diesel Exhaust Particulate in Vivo and in Vitro
Investigators: Nel, Andre E.
Current Investigators: Nel, Andre E. , Cho, Arthur K. , Froines, John R. , Li, Ning , Sioutas, Constantinos
Institution: University of California - Los Angeles
Current Institution: University of California - Los Angeles , University of Southern California
EPA Project Officer: Chung, Serena
Project Period: June 1, 1999 through May 31, 2005 (Extended to May 31, 2006)
Project Period Covered by this Report: June 1, 2000 through May 31, 2001
RFA: Airborne Particulate Matter (PM) Centers (1999) RFA Text |  Recipients Lists
Research Category: Air Quality and Air Toxics , Particulate Matter , Air

Objective:

A central hypothesis of the Southern California Particle Center and Supersite (SCPCS) is that organic constituents associated with particulate matter (PM), including quinones, other organic compounds (polycyclic aromatic hydrocarbons (PAHs), nitro-PAHs, and aldehydes/ketones), and metals, are capable of generating reactive oxygen species (ROS) and acting as electrophilic agents. They have a central role in allergic airway disease including asthma and cardiovascular effects through their ability to generate oxidative stress, inflammation, and immunomodulating effects in the lungs and airways.

The two mechanisms of oxidative stress and electrophilic addition result in impairment and damage to respiratory and cardiac functions. Catalytic action by quinones and related organic compounds and metals represents a key pathway in the toxicity of PM. Quinones produce futile redox cycles as long as reducing equivalents are available, thus enabling continual production of O2-, H2O2, and OH radical. Macrophages and epithelial cells are the principal targets of ROS and the electrophilic agents. We propose that the mechanistic features of the toxicity include activation of NF-kB and AP-1 pathways leading to cytokine and chemokine production and subsequent inflammatory responses, effects on mitochondrial function and apoptosis (see Figure 1). In addition, epidemiologic studies have suggested an association of ambient PM such as PM2.5 with cardiopulmonary diseases and mortality. The 9,10-phenanthroquinone (9,10-PQ) is a potent inhibitor of the neuronal form of nitric oxide synthase (NOS). The 9,10-PQ also inhibits the endothelial form of NOS, which plays a critical role in vascular tone, thereby causing the suppression of nitric oxide (NO)-dependent vasorelaxation of the aorta and a significant increase in blood pressure in rats. Therefore, quinones and other compounds producing ROS (e.g., nitro-PAHs) may contribute to diseases related to vascular dysfunction caused by exposure to urban air particles. In addition to the production of ROS, quinones, PAHs, nitro-PAHs, and related compounds also may undergo electrophilic addition to macromolecules producing complementary toxicity.

Figure 1. Mechanistic Features of Toxicity

Figure 1. Mechanistic Features of Toxicity

Within this mechanistic context we have defined four dosimetric models that represent hypothetical exposure mechanisms to enable quinones, metals, and other organic agents to produce toxicity. These models, outlined in Figure 2, include: (1) the organic compounds and metals that become bioavailable through dissolution and cellular uptake; (2) macrophages and epithelial cells that phagocytize particles and generate ROS and electrophilic chemistry in the intracellular milieu; (3) particles that come in contact with cells followed by dissolution and uptake; and (4) particles that are adsorbed onto cells with subsequent generation of ROS and toxicity. In this latter case, reducing equivalents are required to facilitate the process; elemental carbon may play a role as an electron transfer medium. These hypotheses and mechanistic considerations translate into three related studies:

  • Evaluation of the airborne concentrations of different classes of organic compounds, including quinones, nitroaromatics, and carbonyl compounds in ultrafine, fine, and coarse particles collected from diesel engines, freeways, source (source-site with no direct impact from freeways and no demonstrable products of atmospheric chemistry)/receptor (site where products of atmospheric chemistry are important), and Children’s Health Study (CHS) sites in the Los Angeles Basin (LAB) to determine whether the agents are present in measurable concentrations, their distribution as a function of size, their temporal and spatial characteristics, and their relation to toxicity and health effects.
  • Development and application of in vitro biological assays to quantitatively assess the toxicity of PM samples. Emphasis is on the use of yeast preparations, macrophages, and epithelial cell lines.
  • Establishment of a murine asthma model that reflects the adjuvant effects of diesel exhaust particles (DEP) and concentrated ambient particles (CAPs) in the lung and whose mode of action is consistent with injury by ROS and electrophilic agents.

Figure 2. Exposure Models

Figure 2. Exposure Models

Progress Summary:

In vitro studies in macrophages revealed a range of biological responses to oxidative stress generation by DEP and CAPs. At low DEP extract concentrations, there is activation of the antioxidant element (AE), which leads to the induction of a sensitive oxidative stress protein, heme oxygenase (HO-1). HO-1 is an antioxidant enzyme and is responsible for carbon monoxide (CO) production in the exhaled air. CO is one of the most sensitive clinical markers for airway inflammation in human subjects exposed to DEP. At higher doses of DEP chemicals, macrophages and epithelial cells produce cytokines and chemokines. This likely forms the basis for the pro-inflammatory effects of DEP in the lung. These pro-inflammatory effects are mediated through activation of specific signaling pathways, including activation of the MAP kinase and the NF - κB cascades. Finally, extract doses greater than 25 μg/mL lead to cytotoxicity, which is mediated by a mitochondrial pathway. Epithelial cytotoxicity may contribute to airway hyperreactivity in asthmatics. Assays were used to assess the toxicity of concentrated coarse (C) and fine plus ultrafine (F + UF) PM. F + UF particles induce oxidative stress (HO-1 expression) more readily than C particles. This effect correlates with a higher CO and PAH content of the F + UF particles. All considered, these findings suggest that the generation of oxidative stress leads to a stratified cellular response, which can vary from protective to pro-inflammatory or cytotoxic depending on the level of oxidative stress.

We experimented with a variety of murine exposure protocols to develop an asthma model. The hypothesis is that ovalbumin (OVA) by itself will lead to immune tolerance, but co-administered DEP will induce allergic sensitization. To date, we achieved the best results with a 10-day inhalation protocol in which aerosolized OVA and DEP were used. This model clearly demonstrated that DEP co-administration enhances OVA-specific IgE and IgG1 production. This model also was used to show that interference in oxidative stress by thiol antioxidants will inhibit the adjuvant effect of DEP in IgE and IgG response induction. We have had some preliminary success with a nasal challenge model, in which DEP is co-administered with OVA to induce allergic inflammation and airway eosinophilia in mice.

Future Activities:

In the future, we will apply the murine asthma model to study CAPs effects. The goal of these studies is to establish an animal model that responds to organic DEP and CAPs components with allergic airway inflammation. We will introduce the Buxco box in these studies to measure airway hyperreactivity (AHR). This will allow us to use intervention approaches, such as the use of antioxidants to interfere in the AHR. Also, we will test the effect of gene knockout in these animals to obtain an idea of the role of antioxidant and PAH detoxification pathways in the lung.

The in vitro toxicity studies will continue with CAPs collections by a liquid impinger to elucidate the toxicity of coarse, fine, and ultrafine particles collected at freeways. Also, we will collect particles at various source/receptor sites to study the in vitro toxicity of organic compounds that are formed via atmospheric transformation in the LAB. We will continue to develop highly sensitive in vitro assays to test relevant mechanistic hypotheses and make use of genetically altered biological preparations to assess specific mechanistic features of the PM toxicity. We already have striking results with the use of yeast preparations deficient in superoxide dismustase (SOD).

Journal Articles:

No journal articles submitted with this report: View all 15 publications for this subproject

Supplemental Keywords:

airborne particulate matter, aerosol, size distribution, particle concentrator, NRC priorities, mechanism, quinones, allergens, bioaerosols, dosimetry, children’s study, indoor exposure, exposure assessment, ultrafine, fine and coarse particles, regional human exposure model, REHEX, asthma, polycyclic aromatic hydrocarbons, PAH, clinical human exposures, source-receptor, measurement error, study design, susceptible populations, geo-code, toxicology, epidemiology, regional modeling, source/receptor analysis, Southern California, Los Angeles Basin, photochemistry, meteorology, trajectory modeling, peroxides, Southern California Particle Center and Supersite, SCPCS, air, geographic area, scientific discipline, health, RFA, susceptibility/sensitive population/genetic susceptibility, biology, risk assessments, genetic susceptibility, health risk assessment, biochemistry, particulate matter, environmental chemistry, mobile sources, state, aerosols, automotive exhaust, epidemiology, California (CA), environmentally caused disease, engine exhaust, environmental hazard exposures, allergen, indoor air, indoor air quality, allergens, particle concentrator, air quality, diesel exhaust, particulate emissions, toxics, human health effects, particulates, sensitive populations, toxicology, diesel exhaust particles, environmental triggers, air pollution, airway disease, atmospheric chemistry, children, automotive emissions, dosimetry, inhaled particles, motor vehicle emissions, asthma triggers, PM characteristics, ambient aerosol, asthma, human exposure, particle transport,, RFA, Health, Scientific Discipline, Air, Geographic Area, HUMAN HEALTH, particulate matter, Environmental Chemistry, Air Pollutants, State, Risk Assessments, Biochemistry, Health Effects, particulates, ambient aerosol, asthma, toxicology, quinones, human health effects, airway disease, allergic airway disease, diesel exhaust particulates, air pollution, PAH, diesel exhaust, particulate exposure, human exposure, toxicity, California (CA), allergens, particle concentrator, airborne urban contaminants, human health risk, aerosols, atmospheric chemistry

Relevant Websites:

http://www.scpcs.ucla.edu Exit

Progress and Final Reports:

Original Abstract
  • 1999
  • 2000
  • 2002 Progress Report
  • 2003 Progress Report
  • 2004
  • Final Report

  • Main Center Abstract and Reports:

    R827352    Southern California Particle Center and Supersite

    Subprojects under this Center: (EPA does not fund or establish subprojects; EPA awards and manages the overall grant for this center).
    R827352C001 The Chemical Toxicology of Particulate Matter
    R827352C002 Pro-inflammatory and the Pro-oxidative Effects of Diesel Exhaust Particulate in Vivo and in Vitro
    R827352C003 Measurement of the “Effective” Surface Area of Ultrafine and Accumulation Mode PM (Pilot Project)
    R827352C004 Effect of Exposure to Freeways with Heavy Diesel Traffic and Gasoline Traffic on Asthma Mouse Model
    R827352C005 Effects of Exposure to Fine and Ultrafine Concentrated Ambient Particles near a Heavily Trafficked Freeway in Geriatric Rats (Pilot Project)
    R827352C006 Relationship Between Ultrafine Particle Size Distribution and Distance From Highways
    R827352C007 Exposure to Vehicular Pollutants and Respiratory Health
    R827352C008 Traffic Density and Human Reproductive Health
    R827352C009 The Role of Quinones, Aldehydes, Polycyclic Aromatic Hydrocarbons, and other Atmospheric Transformation Products on Chronic Health Effects in Children
    R827352C010 Novel Method for Measurement of Acrolein in Aerosols
    R827352C011 Off-Line Sampling of Exhaled Nitric Oxide in Respiratory Health Surveys
    R827352C012 Controlled Human Exposure Studies with Concentrated PM
    R827352C013 Particle Size Distributions of Polycyclic Aromatic Hydrocarbons in the LAB
    R827352C014 Physical and Chemical Characteristics of PM in the LAB (Source Receptor Study)
    R827352C015 Exposure Assessment and Airshed Modeling Applications in Support of SCPC and CHS Projects
    R827352C016 Particle Dosimetry
    R827352C017 Conduct Research and Monitoring That Contributes to a Better Understanding of the Measurement, Sources, Size Distribution, Chemical Composition, Physical State, Spatial and Temporal Variability, and Health Effects of Suspended PM in the Los Angeles Basin (LAB)