2004 Progress Report: Studies of Oxidant Mechanisms

EPA Grant Number: R827353C011
Subproject: this is subproject number 011 , established and managed by the Center Director under grant R827353
(EPA does not fund or establish subprojects; EPA awards and manages the overall grant for this center).

Center: EPA Harvard Center for Ambient Particle Health Effects
Center Director: Koutrakis, Petros
Title: Studies of Oxidant Mechanisms
Investigators: Gonzalez-Flecha, Beatriz
Current Investigators: Godleski, John J. , Gonzalez-Flecha, Beatriz
Institution: Harvard University
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, 2003 through May 31, 2004
RFA: Airborne Particulate Matter (PM) Centers (1999) RFA Text |  Recipients Lists
Research Category: Air Quality and Air Toxics , Particulate Matter , Air


The objective is to investigate the role of PM exposure in increasing oxidative stress associated with significant functional alterations in the heart.

Progress Summary:

Oxidant-Mediated Cardio-Toxicity of Ambient Air Pollutants

During the last year we have used an in vivo model of inhalation exposure to ‘real world’ particles to test the hypothesis that the lung-heart signaling after particle deposition in the lung operates through neural mechanisms. Our data show that CAPs exposure increases cardiac oxidants. PM-induced oxidative stress is mediated by autonomic signals and the resulting oxidative stress is associated with significant functional alterations in the heart (Rhoden, et al., 2005).

We used pharmacological strategies to determine whether oxidants are implicated in PM-dependent cardiac dysfunction and whether PM-induced increase in autonomic stimulation on the heart mediates cardiac oxidative stress and toxicity. Adult Sprague-Dawley rats were exposed to either intra-tracheal instillation of urban air particles (UAP 750 μg) or to inhalation of concentrated ambient particles (CAPs mass concentration 740 ± 300 μg/m3 for 5 hours. Oxidative stress and cardiac function were evaluated immediately after exposure. Instillation of UAP led to significant increases in heart oxidants measured as organ chemiluminescence (UAP: 38 ± 5 cps/cm2, sham: 10 ± 1 cps/cm2) or thiobarbituric acid reactive substances (TBARS, UAP: 76 ± 10, Sham 30 ± 6 nmol/mg protein). Heart rate values were increased immediately after exposure (UAP: 390 ± 20 bpm, sham: 350 ± 10 bpm) and returned to basal levels over the next 30 minutes. Heart rate variability (SDNN) was unchanged immediately after exposure, but significantly increased during the recovery phase (UAP: 3.4 ± 0.2, Sham: 2.4 ± 0.3).

To determine the role of ROS in the development of cardiac malfunction, rats were treated with 50 mg/Kg N-acetylcysteine (NAC) 1 hour prior to UAP instillation or CAPs inhalation. NAC prevented changes in heart rate and SDNN in UAP-exposed rats (340 ± 8 and 2.9 ± 0.3 respectively).

To investigate the role of the autonomic nervous system in PM-induced oxidative stress, rats were given 5 mg/Kg atenolol (β-1 receptor antagonist), 0.30 mg/Kg glycopyrrolate (muscarinic receptor antagonist) or saline immediately before exposure to CAPs aerosols. Both atenolol and glycopyrrolate effectively prevented CAPs-induced cardiac oxidative stress (CLATEN: 11 ± 1 cps/cm2, CLGLYCO: 10 ± 1 cps/cm2, TBARSATEN: 40 ± 6 nmol/mg protein, TBARSGLYCO: 38 ± 6 nmol/mg protein) (Rhoden, et al., 2005).

These data show that PM exposure increases cardiac oxidants via autonomic signals and the resulting oxidative stress is associated with significant functional alterations in the heart. The observed preventive effects of NAC suggest that treatment with low doses of this antioxidant could be used to ameliorate the toxic effects of particulate air pollution.

Ambient Particulate Matter Exhibits Direct and Selective Inhibitory Effects on Oxidative Stress Enzymes

A preliminary mechanistic hypothesis by which ambient air particles have a significant impact on human health is via the induction of pulmonary inflammatory responses mediated through the generation of reactive oxygen species (ROS). The objective of this study was to evaluate whether air particulates interact directly with enzymes involved in oxidative stress responses, which are part of ROS-mediated inflammatory response pathways. We performed enzyme inhibition assays on four enzymes involved in oxidative stress responses (Cu/Zn superoxide dismutase, Mn superoxide dismutase, glutathione peroxidase, and glutathione reductase), in the presence of particles of varying toxicities, and found distinctive inhibition patterns that were both particle and enzyme specific. Results from the assays on Cu/Zn SOD and Mn SOD enzymes are shown in Figure 1. In both cases, the control treatment consisted of the corresponding enzyme at 0.15 U/ml, which falls at the middle of the linear range for the assay, with no particles added. The standard curve for each enzyme was determined from a range of standards according to the assay protocol, using replicate wells in the same microplate to ensure consistency within each experiment. The effects of the particle type and concentration appear to be similar for both SODs. The NIST particles reduced the activity of both enzymes by about 50% when present at 100 μg/ml and by more than 90% at a level of 500 μg/ml. The ROFA particles were clearly more active, eliciting 75% and 100% inhibition of the enzymatic activities when present at 100 μg/ml and 500 μg/ml, respectively. Inert MSHA particles were the least inhibitory and did not reduce the activity of Cu/Zn SOD at 100 μg/ml, but they did show a small but significant inhibitory effect on the activity of Mn SOD (the 95% bootstrap confidence interval of residual Mn SOD activity was [76.3, 88.9]). On the basis of these findings, we suggest a strategy for an enzyme bioassay that could be used to assess the potential of particles to generate ROS-induced responses. This work is detailed in a manuscript submitted for publication (Hatzis, et al., 2005).

Effect of PM Type and Concentration on the Activity of (a) Cu/ZnSOD and (b) MnSOD (n = 5 per determination).

Figure 1. Effect of PM Type and Concentration on the Activity of (a) Cu/ZnSOD and (b) MnSOD (n = 5 per determination)

Journal Articles on this Report : 2 Displayed | Download in RIS Format

Other subproject views: All 3 publications 3 publications in selected types All 3 journal articles
Other center views: All 200 publications 198 publications in selected types All 197 journal articles
Type Citation Sub Project Document Sources
Journal Article Hatzis C, Godleski JJ, Gonzalez-Flecha B, Wolfson JM, Koutrakis P. Ambient particulate matter exhibits direct inhibitory effects on oxidative stress enzymes. Environmental Science & Technology 2006;40(8):2805-2811. R827353 (Final)
R827353C011 (2004)
R827353C011 (Final)
  • Abstract from PubMed
  • Full-text: Research Gate-Abstract & Full Text
  • Abstract: ACS-Abstract
  • Other: ACS-Full Text PDF
  • Journal Article Rhoden CR, Wellenius GA, Ghelfi E, Lawrence J, Gonzalez-Flecha B. PM-induced cardiac oxidative stress and dysfunction are mediated by autonomic stimulation. Biochimica et Biophysica Acta (BBA)-General Subjects 2005;1725(3):305-313. R827353 (Final)
    R827353C011 (2004)
    R827353C011 (Final)
    R832416 (2008)
  • Abstract from PubMed
  • Abstract: ScienceDirect-Abstract
  • Supplemental Keywords:

    exposure, health effects, susceptibility, metals, public policy, biology, engineering, epidemiology, toxicology, environmental chemistry, monitoring, air pollutants, air pollution, air quality, ambient air, ambient air monitoring, ambient air quality, ambient measurement methods, ambient monitoring, ambient particle health effects, ambient particles, animal inhalation study, assessment of exposure, biological mechanism, biological response, cardiopulmonary, cardiopulmonary response, cardiovascular disease, chemical exposure, children, developmental effects, dosimetry, environmental health hazard, exposure and effects, genetic susceptibility, health risks, human exposure, human health, human health effects, human health risk, human susceptibility, indoor air quality, indoor exposure, inhalation, inhalation toxicology, inhaled particles, lead, measurement methods, particle exposure, particulate exposure, particulates, pulmonary, pulmonary disease, respiratory, respiratory disease, risk assessment, sensitive populations, stratospheric ozone,, RFA, Health, Scientific Discipline, Air, particulate matter, Toxicology, Environmental Chemistry, Epidemiology, Risk Assessments, Environmental Microbiology, Environmental Monitoring, tropospheric ozone, Molecular Biology/Genetics, Biology, Environmental Engineering, ambient air quality, particulates, chemical exposure, human health effects, ambient air monitoring, concentrated ambient particulates (CAPs), ambient air, ambient measurement methods, pulmonary disease, developmental effects, epidemelogy, respiratory disease, air pollution, particle exposure, biological mechanism , human exposure, inhalation, pulmonary, particulate exposure, ambient particle health effects, inhalation toxicology, oxidant mechanisms, measurement methods , cardiovascular disease, human health risk, respiratory, genetic susceptibility

    Progress and Final Reports:

    Original Abstract
  • 1999
  • 2000
  • 2001
  • 2002
  • 2003 Progress Report
  • Final Report

  • Main Center Abstract and Reports:

    R827353    EPA Harvard Center for Ambient Particle Health Effects

    Subprojects under this Center: (EPA does not fund or establish subprojects; EPA awards and manages the overall grant for this center).
    R827353C001 Assessing Human Exposures to Particulate and Gaseous Air Pollutants
    R827353C002 Quantifying Exposure Error and its Effect on Epidemiological Studies
    R827353C003 St. Louis Bus, Steubenville and Atlanta Studies
    R827353C004 Examining Conditions That Predispose Towards Acute Adverse Effects of Particulate Exposures
    R827353C005 Assessing Life-Shortening Associated with Exposure to Particulate Matter
    R827353C006 Investigating Chronic Effects of Exposure to Particulate Matter
    R827353C007 Determining the Effects of Particle Characteristics on Respiratory Health of Children
    R827353C008 Differentiating the Roles of Particle Size, Particle Composition, and Gaseous Co-Pollutants on Cardiac Ischemia
    R827353C009 Assessing Deposition of Ambient Particles in the Lung
    R827353C010 Relating Changes in Blood Viscosity, Other Clotting Parameters, Heart Rate, and Heart Rate Variability to Particulate and Criteria Gas Exposures
    R827353C011 Studies of Oxidant Mechanisms
    R827353C012 Modeling Relationships Between Mobile Source Particle Emissions and Population Exposures
    R827353C013 Toxicological Evaluation of Realistic Emissions of Source Aerosols (TERESA) Study
    R827353C014 Identifying the Physical and Chemical Properties of Particulate Matter Responsible for the Observed Adverse Health Effects
    R827353C015 Research Coordination Core
    R827353C016 Analytical and Facilities Core
    R827353C017 Technology Development and Transfer Core