2007 Progress Report: Human Clinical Studies of Concentrated Ambient Ultrafine and Fine ParticlesEPA Grant Number: R832415C003
Subproject: this is subproject number 003 , established and managed by the Center Director under grant R832415
(EPA does not fund or establish subprojects; EPA awards and manages the overall grant for this center).
Center: Rochester PM Center
Center Director: Oberdörster, Günter
Title: Human Clinical Studies of Concentrated Ambient Ultrafine and Fine Particles
Investigators: Frampton, Mark W. , Utell, Mark J.
Current Investigators: Frampton, Mark W. , Zareba, Wojciech , Utell, Mark J. , Oakes, David , Phipps, Richard , Gelein, Robert
Institution: University of Rochester
EPA Project Officer: Chung, Serena
Project Period: October 1, 2005 through September 30, 2010 (Extended to September 30, 2012)
Project Period Covered by this Report: October 1, 2006 through September 30,2007
RFA: Particulate Matter Research Centers (2004) RFA Text | Recipients Lists
Research Category: Human Health , Air
The overall objective of our current and planned studies is to determine the pulmonary and cardiovascular effects of exposure to ultrafine and fine particulate matter (PM). The clinical studies in healthy humans and susceptible individuals with diabetes proposed in this research core focus on the effects of ambient ultrafine and fine particles on three major determinants of adverse cardiac events: 1) blood coagulation induced by effects on platelets and circulating microparticles; 2) cardiac output; and 3) cardiac rhythm and repolarization.
Our overall hypothesis is that inhalation of ambient PM causes small but measurable changes in coagulation and cardiovascular function that help explain the cardiovascular effects of PM exposure. We further hypothesize that the cardiovascular effects are determined by the ability of PM to generate reactive oxygen and nitrogen species, and are more pronounced in subjects with type 2 diabetes. Inhaled ultrafine particles increase the burden of reactive oxygen species to the endothelium. Endothelial activation and vasoconstriction increase platelet adherence and release of thromboxane, activate and prolong the transit time of blood leukocytes, and deplete vascular nitric oxide (NO). Particles may also have direct effects on platelets and leukocytes. Vascular injury triggers release of procoagulant microparticles into the blood, and initiation of coagulation. In collaboration with the Vascular and Inflammation Facility Core, we measure the effects of inhaled ambient fine PM on platelet number, phenotype, and function, and quantitate intravascular microparticles derived from platelets and endothelial cells. In collaboration with the Cardiac Core, we use noninvasive monitoring methods to measure exposure effects on cardiac output, rhythm, and repolarization. These studies take advantage of and extend upon our project funded by the National Institute of Environmental Health Sciences and the EPA, “Ultrafine Particle-Induced Oxidative Stress”, which focuses on effects on vascular function and NO.
- Human clinical exposures to concentrated ambient ultrafine particles
- Inhalation of carbon UFP in diabetics
- Ultrafine Particles Activate Platelets In Vitro
The Harvard ultrafine particle concentrator has been installed in a dedicated room in the Kornberg Medical Research Building at the University of Rochester Medical Center, and is fully operational. We have completed construction of a negative-pressure inhalation chamber, which is housed within our new exposure facility. We have initiated human clinical exposures to concentrated ambient ultrafine particles, using the concentrator and the exposure chamber. Installation and maintenance of the concentrator and exposure system requires close collaboration with Dr. Oberdörster, Research Core #1, and the Aerosol Generation & Analysis Core.
We are now conducting a clinical study of healthy never-smoking subjects inhaling concentrated ambient UFP using the concentrator. This study is co-funded by an NIEHS RO1 grant and this EPA center grant. Subjects are equal numbers of males and females admitted to the Clinical Research Center the day prior to exposure, to minimize the potential influence of outdoor pollutant exposure in the hours prior to the experimental exposure. Physiologic measurements are made the day prior to exposure, and them 0, 3.5, 21, and 45 hours after exposure. Measurements include blood sampling for flow cytometry and soluble markers of inflammation and coagulation, pulmonary function testing, diffusing capacity for carbon monoxide, and flow-mediated dilatation of the forearm. The EPA Center is funding continuous cardiac monitoring, noninvasive measurement of cardiac output, and newly developed methods for analysis of platelet activation and circulating microparticles, using flow cytometry.
Figure 1. Change in platelet expression of CD40L (CD154) following exposure to air (circles) or UFP (squares).
So far 9 subjects are enrolled in the study, and 6 subjects have completed both exposures. We plan to study a total of 20 subjects, 10 males and 10 females, age 30 to 60 years. Exposures have been successful from a technical standpoint, and no subject has experienced symptoms or problems with the exposures. Results are not yet available, because the study is double-blinded.
We have now completed the exposures for our study of the effects of inhalation of ultrafine carbon particles in subjects with diabetes. Diabetics have vascular endothelial dysfunction which may increase their risk for adverse cardiovascular effects from air borne particles. Type 2 diabetics, age 30-60, without clinical cardiovascular disease, were exposed to filtered air or 50 μg/m3 carbon UFP (count median diameter 30 nm, GSD 1.8) by mouthpiece for two hours in a randomized double-blinded cross-over study. Exposures were separated by at least two weeks. Nineteen subjects completed the study.
Figure 2. Change in number of microvesicles expressing TF (CD142) following exposure to air (circles) or UFP (squares).
A preliminary analysis of the findings was published in abstract form and presented at the 2007 American Thoracic Society International Conference. Compared with air exposure, UFP exposure increased platelet expression of CD40 ligand (CD40L) (Figure 1), a marker of platelet activation and a key molecule in the development of atherosclerosis. UFP exposure also increased platelet-associated tissue factor (TF) and increased the number of microparticles expressing TF (Figure 2). Changes occurred most consistently 3.5 hours after exposure, and effects were no longer seen 24 hours after exposure. There were no significant effects of UFP on platelet counts or platelet aggregates. The subjects with diabetes showed significant reductions in forearm flow-mediated vascular dilatation in comparison with healthy subjects, as expected. Both the pulmonary diffusing capacity for carbon monoxide (an indicator of pulmonary vascular function), and forearm flow mediated dilatation (an indicator of systemic vascular function), decreased with UFP exposure to ultrafine particles compared with clean air exposure, but the differences were not statistically significant. We concluded that inhalation of carbon UFP for 2 hours may transiently activate vascular endothelium and/or platelets in subjects with type 2 diabetes. This finding supports the hypothesis that exposure to ambient UFP may increase the potential for vascular thrombosis in patients with severe vascular disease or ulcerated atherosclerotic plaques. We are currently measuring plasma levels of TF and CD40L in collaboration with the Vascular and Inflammation Core, and analyzing the cardiac monitoring data in collaboration with the Cardiac Core. The Biostatics Core is assisting with data analysis. This project also involves close collaboration with Research Core #1 and the Aerosol Generation & Analysis Core for UFP generation and exposure monitoring.
Inhaled UFP have the capability of entering pulmonary vascular endothelial cells and even the blood. One possible mechanism for activation of platelets, thus increasing risk for cardiovascular thrombotic events, may involve direct effects of UFP that have entered the circulation. The purpose of this study was to examine the effects of particles with varying composition, size, and surface area on platelet activation in vitro. The following particles (and count diameters) were studied: copper (0.022 μm), commercial elemental carbon (P90, 0.014 μm), laboratory-generated elemental carbon (lab carbon, 0.030 μm), and diesel exhaust particles (DEP, 1.62 μm). Vortexed particle suspensions were added to whole blood from seven healthy never-smoking subjects and incubated for 30 minutes at 37°C. Platelet expression of CD62P (p-selectin), platelet aggregates, and platelet-leukocyte conjugates were measured by flow cytometry. Dose-dependent increases in the expression of CD62P were most pronounced with copper and lab carbon. Lab carbon also significantly increased platelet aggregates and platelet-leukocyte conjugates. These findings indicate that UFP can activate platelets in vitro, and the activity varies with particle size and composition.
In summary, preliminary studies suggest that inhalation of carbon ultrafine particles may increase the possibility of blood clot formation in people with diabetes who have atherosclerotic vascular disease, through activation of platelets and circulating microparticles. They also suggest that flow cytometry analyses of platelets and microparticles may be sensitive biomarkers of pro-thrombotic effects of air pollution exposure.
In the coming year we expect to complete our study of healthy subjects exposed to ambient concentrated ultrafine particles, and initiate comparable studies in subjects with type 2 diabetes. In addition, we will be completing our analysis and publishing the results of our study of people with type 2 diabetes exposed to ultrafine carbon particles. For the in vitro studies of platelet activation, future experiments will explore the mechanisms for particle activation of platelets, and the role of particle composition and surface area. We are collaborating with Dr. Phipps and the Vascular & Inflammation Core in developing these research strategies.
Journal Articles:No journal articles submitted with this report: View all 57 publications for this subproject
Supplemental Keywords:RFA, Health, PHYSICAL ASPECTS, Scientific Discipline, Air, particulate matter, Health Risk Assessment, Risk Assessments, Physical Processes, atmospheric particulate matter, atmospheric particles, long term exposure, acute cardiovascular effects, airway disease, exposure, human exposure, ambient particle health effects, atmospheric aerosol particles, ultrafine particulate matter, PM, aersol particles, cardiovascular disease
Progress and Final Reports:Original Abstract
Main Center Abstract and Reports:R832415 Rochester PM Center
Subprojects under this Center: (EPA does not fund or establish subprojects; EPA awards and manages the overall grant for this center).
R832415C001 Characterization and Source Apportionment
R832415C002 Epidemiological Studies on Extra Pulmonary Effects of Fresh and Aged Urban Aerosols from Different Sources
R832415C003 Human Clinical Studies of Concentrated Ambient Ultrafine and Fine Particles
R832415C004 Animal models: Cardiovascular Disease, CNS Injury and Ultrafine Particle Biokinetics
R832415C005 Ultrafine Particle Cell Interactions In Vitro: Molecular Mechanisms Leading To Altered Gene Expression in Relation to Particle Composition