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2006 Progress Report: Project 2: The Role of Oxidative Stress in PM-induced Adverse Health Effects
EPA Grant Number: R832413C002Subproject: this is subproject number 002 , established and managed by the Center Director under grant R832413
(EPA does not fund or establish subprojects; EPA awards and manages the overall grant for this center).
Center: Southern California Particle Center
Center Director: Froines, John R.
Title: Project 2: The Role of Oxidative Stress in PM-induced Adverse Health Effects
Investigators: Nel, Andre E. , Harkema, Jack , Kleinman, Michael T. , Lusis, Aldons
Institution: Michigan State University , University of California - Irvine , University of California - Los Angeles
Current Institution: University of California - Los Angeles , Michigan State University , University of California - Irvine
EPA Project Officer: Katz, Stacey
Project Period: October 1, 2005 through September 30, 2010
Project Period Covered by this Report: October 1, 2005 through September 30, 2006
Project Amount: Refer to main center abstract for funding details.
RFA: Particulate Matter Research Centers (2004)
Research Category: Particulate Matter
Description:
Objective:The primary objective is to elucidate the mechanism(s) of particulate matter (PM)-induced asthma and atherosclerosis exacerbation. Mechanisms are investigated through in vivo animal studies in a mobile trailer suitable for exposure to ambient PM as well as in vitro studies of tissue culture cells.
Progress Summary:Atherosclerosis Studies on Genetically Susceptible Animals and Endothelial Cell Cultures
Epidemiological studies suggest that exposure to ambient PM constitutes a risk factor for atherosclerosis. Exposure to fine PM (aerodynamic diameter < 2.5 μm, PM2.5) has been shown to promote atherogenesis in apoE null mice. We hypothesized that PM synergizes with known proatherogenic stimuli and mediators in their ability to elicit oxidative stress and promote atherosclerosis, and that most of the pro-inflammatory potential resides in the ultrafine particles (aerodynamic diameter < 0.1 μm, UFP) that are highly enriched for redox cycling PM chemicals. Two animal experiments were conducted on genetically susceptible animals in the mobile trailer at USC to pursue this hypothesis.
ApoE Null Mice. Seven-week old female mice were assigned to four different groups, ~17 mice/group: non-exposed (NE), filtered air (FA), concentrated PM2.5 (FP) and concentrated UFP. NE mice were kept at the UCLA vivarium. FA, FP and UFP mice were housed in a mobile animal laboratory located in downtown Los Angeles, and exposed to FP or UFP for a combined total of 75 hours over a period of 40 days. The following specimens were obtained: blood for plasma lipoproteins and plasma cytokines; aortic root for atherosclerosis assessment; lungs, livers, spleens, kidneys, hearts, and brain for histology and mRNA/protein analysis.
Atherosclerosis was assessed by histological examination of the ascending aorta and expressed as average lesional area (μm2/section). Animals exposed to concentrated (17x) UFP developed significantly greater atherosclerotic lesions (33011 ± 3741, n = 15) as compared with NE (17261 ± 1659, n = 17, p = 0.0002) or FA mice (21362 ± 2864, n = 15, p = 0.02), and a similar trend was observed in comparison to mice exposed to concentrated (13x) FP (26361 ± 2275, n = 16, p = 0.06). Increased systemic oxidative stress was suggested by the significant up-regulation of the p45-NFE2 related factor-2 (Nrf2) and its efferent phase-2 response genes in the liver and aorta.
LDL-Receptor Null Mice. Six week old female LDL null mice were fed a Western Diet for 2 months, beginning one month prior to PM exposures. Approximately 12 animals per group were exposed to FA, fine particles (FP) or ultrafine particles (UFP), for a total of 65 hours, over 13 exposure sessions. Although a Western Diet was quite effective in inducing atherosclerotic lesions in the LDL-receptor knockout animals, particle exposure did not enhance the number of lesions. The preliminary findings suggest that a strong pro-atherosclerotic dietary stimulus can override the particle effects. This is in keeping with the notion that the particles synergize with the lipid effects at lower levels of oxidative stress.
In Vitro. We used microarray technology to study gene expression in a human microvascular endothelial cell line. Diesel exhaust particles (5 μg/ml), a component of ambient UFP, synergized with an oxidized LDL proatherogenic mediator, palmitoyl-arachidonyl-phosphatidyl choline (10, 20 and 40 μg/ml), in the up-regulation of a large number of genes (n = 626). Clustering analysis shows that the responsive genes cluster into a number of important functional groups.
In conclusion, our studies show that exposure to pro-oxidative UFP exacerbates atherosclerosis in apoE null mice and appears to act synergistically with oxidized LDL components in the vascular wall. The above studies should be publication-ready within the next year.
Evidence for NF-κB Activation in Brain Tissue of CAPs-Exposed Animals (in Collaboration with Drs. Campbell and Kleinman at UCI)
Brain samples were obtained from the apoE knockout animals described above. Particle number concentration during exposures ranged from 82,000 to 510,000 (average about 250,000). Brains were harvested approximately 30 hr after the last exposure. Tissue from eight mice per exposure group were analyzed for NF-κB expression by EM SA. The data were analyzed by one-way analysis of variance. The effect of exposure on expression was highly significant (p ≤ 0.001). In Tukey multiple comparison analysis, NF-κB expression in mice exposed to both FP and UFP were significantly (p ≤ 0.02) elevated above purified air-exposed controls. Expression levels in mice exposed to UFP were slightly elevated relative to those in mice exposed to fine particles. The study will be extended after further animal exposures in the coming year.
Comparison of the Prooxidative Effects of UFP with Commercial Nanoparticles
We compared the cellular effects of ambient UFP with manufactured titanium dioxide (TiO2), carbon black, fullerol and polystyrene (PS) nanoparticles (NP) (Xia, et al., 2006). The study was conducted in a phagocytic cell line (RAW 264.7) that is representative of a lung target for NP. Physicochemical characterization of the NP showed a dramatic change in their state of aggregation, dispersibility and charge during transfer from a buffered aqueous solution to cell culture medium. Particles differed with respect to cellular uptake, subcellular localization and ability to catalyze the production of reactive oxygen species (ROS) under biotic and abiotic conditions. Spontaneous ROS production by different particle types was studied with an ROS quenching assay as well as an NADPH peroxidase bioelectrode platform. Among the particles tested, ambient UFP and cationic PS nanospheres were capable of inducing cellular ROS production, GSH depletion and toxic oxidative stress. This toxicity involves mitochondrial injury through increased calcium uptake and structural organellar damage. Although active under abiotic conditions, TiO2 and fullerol did not induce toxic oxidative stress in cultured cells. While increased TNF-˜production could be seen to accompany UFP-induced oxidant injury, cationic PS nanospheres induced mitochondrial damage and cell death without inflammation. In summary, we demonstrate that ROS generation and oxidative stress is a valid test paradigm to compare NP toxicity. Although not all materials have electronic configurations or surface properties to allow spontaneous ROS generation, particle interactions with cellular components are capable of generating oxidative stress. This study also served as the background for a review article on nanoparticle toxicity (Nel, et al., 2006).
Tracheobronchial Particle Dose Considerations for In Vitro Toxicology Studies
The purpose of this study was to present a method to reconcile particle doses that were used in in vitro toxicology studies with in vivo exposure levels. The focus is on the dose of PM to the tracheobronchial tree of heavily exposed individuals. A review of the factors that influence inhaled particle deposition in environmental exposures led to the identification of scenarios in which greater than average tracheobronchial tree doses occur. Exercising individuals and those with chronic obstructive pulmonary disease not only inhale increased volumes of air, but they also have uneven ventilation that leads to greater than average particle deposition doses per unit of tracheobronchial tree surface area. In addition, deposition hot spots, as occur at airway bifurcations, will greatly increase the particle exposures of target cells in the tracheobronchial tree. Although the exposure of cells in vitro cannot fully replicate the complexity of in vivo exposures, it is possible to calculate toxicologically relevant doses that may determine adverse health effects in potentially sensitive human populations, and when factors that enhance particle doses in vivo are considered, substantial particle doses may be justified for in vitro tissue culture studies that use tracheobronchial target cells, such as epithelial cells and macrophages.
Future Activities:Our studies in the next year will continue to address the role of CAPs in murine atherosclerosis and asthma models. We will also focus on the role of CAPs and pro-oxidative DEP chemicals in perturbing the function of endothelial cells and dendritic cells. The effects on dendritic cells could help to explain the adjuvant role of PM asthma.
Journal Articles on this Report: 2 Displayed | Download in RIS Format
| Other subproject views: | All 17 publications | 10 publications in selected types | All 10 journal articles |
| Other center views: | All 94 publications | 48 publications in selected types | All 47 journal articles |
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Li N, Nel AE. The cellular impacts of diesel exhaust particles: beyond inflammation and death. European Respiratory Journal 2006;27(4):667-668. |
R832413 (2008) R832413C002 (2006) R832413C002 (2008) |
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Xia T, Kovochich M, Brant J, Hotze M, Sempf J, Oberley T, Sioutas C, Yeh JI, Wiesner MR, Nel AE. Comparison of the abilities of ambient and manufactured nanoparticles to induce cellular toxicity according to an oxidative stress paradigm. Nano Letters 2006;6(8):1794-1807. |
R832413 (2008) R832413C001 (2007) R832413C001 (2008) R832413C002 (2006) R832413C002 (2008) R827352 (Final) R827352C002 (Final) R827352C014 (Final) |
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asthma, atherosclerosis, oxidative stress, ambient PM, ambient air, health effects, biology, sensitive populations, human health, animal, PAH,
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Air, Scientific Discipline, Health, RFA, Toxicology, Risk Assessments, Health Risk Assessment, Biochemistry, particulate matter, Ecology and Ecosystems, cardiovascular disease, cardiovascular vulnerability, human health risk, oxidative stress, human health effects, particulates, PM 2.5, air pollution, airway disease, atmospheric particulate matter, vascular dysfunction, airborne particulate matter, human exposure
Relevant Websites:
Progress and Final Reports:
Original Abstract
2007 Progress Report
2008 Progress Report
Main Center Abstract and Reports:
R832413 Southern California Particle Center
Subprojects under this Center:
(EPA does not fund or establish subprojects; EPA awards and manages the overall grant for this center).
R832413C001 Contribution of Primary and Secondary PM Sources to Exposure & Evaluation of Their Relative Toxicity
R832413C002 Project 2: The Role of Oxidative Stress in PM-induced Adverse Health Effects
R832413C003 The Chemical Properties of PM and their Toxicological Implications
R832413C004 Oxidative Stress Responses to PM Exposure in Elderly Individuals With Coronary Heart Disease
R832413C005 Ultrafine Particles on and Near Freeways