2006 Progress Report: Human Clinical Studies of Concentrated Ambient Ultrafine and Fine Particles

EPA 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. , Gelein, Robert , Oakes, David , Phipps, Richard , Utell, Mark J. , Zareba, Wojciech
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, 2005 through September 30, 2006
RFA: Particulate Matter Research Centers (2004) RFA Text |  Recipients Lists
Research Category: Health Effects , Air

Objective:

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 will 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.

Hypothesis

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. We hypothesize that 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 Core, we will 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 will use noninvasive monitoring methods to measure exposure effects on cardiac output, rhythm, and repolarization. These studies will take advantage of and extend upon our project funded by the National Institute of Environmental Health Sciences and the U.S. Environmental Protection Agency (EPA), “Ultrafine Particle-Induced Oxidative Stress,” which will focus on effects on vascular function and NO.

Progress Summary:

Human Clinical Exposures to Concentrated Ambient Ultrafine Particles

A major goal of this project is to initiate human clinical exposures to concentrated ambient ultrafine particles, using the Harvard ultrafine particle concentrator. As noted elsewhere in this progress report, 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. The concentrator is now fully operational, and its performance has been validated, both in terms of “dry runs” and in a series of animal exposure studies undertaken by Dr. Oberdörster’s laboratory (see the 2006 Annual Report for RD832415C004). The intake to the particle concentrator is positioned adjacent to Elmwood Avenue, a busy thoroughfare adjacent to the Medical Center campus, where emissions are dominated by traffic sources during the morning and afternoon rush hours. The concentrated particle exposures will reflect both traffic and point-source pollution.

We have completed construction of a negative-pressure inhalation chamber, which is housed within our new exposure facility. Subjects will sit within this chamber for the ultrafine particle exposures, which will be delivered via face mask. We have now received approval of the study protocol from the Research Subject Review Board at the University of Rochester, and from the EPA, to initiate studies of concentrated ultrafine particles in healthy human subjects and will initiate recruitment of subjects.

Collaboration With Other EPA PM Centers

There are few laboratories in the world currently involved in human clinical studies of air pollution. In order to maximize these human research resources, we are in the midst of a major effort to develop common protocols and outcome measures across the groups that are doing human clinical studies. We are currently collaborating with Dr. Diane Gold at the Harvard EPA Center and Dr. Robert Devlin of the EPA Human Studies Division. These two laboratories and Rochester are the only EPA particle centers with major programs in human clinical studies. All three centers are developing a common list of blood markers of inflammation and particle effects to be used in all studies. Protocols have been developed here in Rochester to measure circulating microparticles, platelet activation, and platelet aggregates using flow cytometry, and these protocols have been shared with both Harvard and EPA.

Samples of human leukocyte RNA have been sent to Dr. Devlin at EPA for gene expression analysis. Preliminary results are currently being analyzed. Efforts are underway to incorporate collection of both leukocyte RNA and genomic DNA in all human studies for future analyses of gene-environment interaction.

Studies of Ultrafine Carbon Particle Inhalation in Subjects With Diabetes

We are nearing completion 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 airborne particles. Diabetics were exposed to filtered air or 50 μg/m3 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. Seventeen of the planned 20 subjects have now completed the study. A preliminary analysis of the first six subjects completing the study was published in abstract form and presented at the 2006 American Thoracic Society International Conference. The subjects with diabetes show significant reductions in forearm flow-mediated vascular dilatation in comparison with healthy subjects, as expected. Both the pulmonary diffusing capacity for carbon monoxide and forearm flow mediated dilatation decreased with exposure to ultrafine particles compared with clean air exposure, although the differences were not statistically significant by paired t-test. These preliminary results suggest that inhalation of carbon UFP alters pulmonary and systemic endothelial function in people with type 2 diabetes. These effects are similar to effects previously seen in healthy subjects. The findings indicate that diabetics, who already have endothelial dysfunction, show further reduction in endothelial function in response to UFP exposure.

Effects of Carbon Ultrafine Particles on Platelet Activation

Blood platelets have a major role in cardiovascular thrombotic events. Because inhaled ultrafine particles may gain access to the pulmonary endothelium and vascular space, one possible mechanistic link between PM exposure and cardiovascular events is direct effects of particles on platelets. The purpose of this study was to assess the direct effects of UFP and diesel exhaust particles (DEP) on platelet activation in vitro. Elemental carbon UFP (count median diameter 0.025 μm, GSD 1.6) and DEP (mean diameter 1.62 ± 0.01 μm) were added to whole blood obtained from 12 healthy never-smoking subjects, at final concentrations from 0 to 200 μg/ml. Samples were incubated for 30 minutes at 37°C, and platelet expression of CD62P (P-selectin), a marker of platelet activation, was measured by flow cytometry. Small, concentration-related increases in platelet expression of CD62P were seen with both UFP and DEP. These studies suggest that direct platelet activation by particles entering the vascular space could play a role in the cardiovascular effects of particulate matter exposure. These data were published in abstract form and presented at the 2006 American Thoracic Society International Conference.

Diabetes and Blood Leukocyte Production of Reactive Oxygen Species

Blood leukocyte production of reactive oxygen species (ROS) via the NADPH oxidase system is a significant mechanism for host defense against infection, but also a potential source of damaging ROS during inflammatory responses. People with diabetes have increased susceptibility to the adverse effects of PM exposure. One of our overall objectives in these studies is to determine whether PM exposure enhances the vascular burden of ROS in people with diabetes.

We hypothesized that diabetes itself would impair the oxidative burst in peripheral blood leukocytes. Previous studies of the effects of diabetes on leukocyte ROS production have shown inconsistent findings. We therefore measured blood leukocyte production of ROS in healthy subjects and in subjects with type 2 diabetes, using a flow cytometric method. Subjects were 10 never-smokers with stable, uncomplicated type 2 diabetes (5 each gender, weight 103 ± 24 kg, BMI 35 ± 6) and 10 never-smoking healthy subjects (BMI 26 ± 5). Heparinized whole blood was diluted with phosphate buffered saline and then reacted with 1,2,3-dihydrorhodamine. The oxidative burst was measured after stimulation with PMA. Samples, including an unstained, unstimulated sample, were analyzed using flow cytometry. Table 1 below shows ROS production in blood monocytes and neutrophils from the healthy and diabetic subjects.

Table 1. ROS Production in Blood Monocytes and Neutrophils from Health and Diabetic Subjects

Monocytes

Healthy

Diabetic

p value

Constitutive ROS

9195 ± 1541

29881 ± 5386

0.002

Stimulated ROS

41175 ± 6962

58463 ± 8275

0.128

 

 

 

 

Neutrophils

Healthy

Diabetic

p value

Constitutive ROS

2464 ± 146

2484 ± 249

0.944

Stimulated ROS

670673 ± 116461

188945 ± 3125

0.008

Constitutive monocyte production of ROS was increased in diabetics compared with healthy subjects, and stimulated PMN ROS production was markedly reduced in diabetics. There were no significant correlations of ROS production with hemoglobin A1C, high density lipoprotein, or blood glucose levels. We concluded that constitutive monocyte ROS production was increased, and stimulated PMN ROS production was reduced, in subjects with type 2 diabetes compared with healthy subjects. We could not exclude the possibility that the increased BMI in the diabetes group contributed to these differences. However, the data suggest that diabetics have both an increased vascular burden of ROS from circulating monocytes, and reduced oxidative burst in PMN, which increases the risk for infection. These data were presented at the meeting of the International Society for Analytical Cytology, Montreal, Canada, May 2006.

Future Activities:

Now that our study protocol has been approved by both the Research Subjects Review Board of the University of Rochester, and by the EPA, we plan to initiate recruitment of subjects. This study, titled UPCON, will expose 20 healthy human subjects to air and to concentrated ambient ultrafine particles, in a randomized, crossover, double-blind design. We are also continuing studies of the effects of ultrafine particles of varying surface area on platelet activation and aggregation in vitro.


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

Other subproject views: All 57 publications 44 publications in selected types All 41 journal articles
Other center views: All 190 publications 156 publications in selected types All 143 journal articles
Type Citation Sub Project Document Sources
Journal Article Shah AP, Pietropaoli AP, Frasier LM, Speers DM, Chalupa DC, Delehanty JM, Huang L-S, Utell MJ, Frampton MW. Effect of inhaled carbon ultrafine particles on reactive hyperemia in healthy human subjects. Environmental Health Perspectives 2008;116(3):375-380. R832415 (2007)
R832415 (2008)
R832415 (2010)
R832415 (2011)
R832415 (Final)
R832415C003 (2006)
R832415C003 (2010)
R832415C003 (2011)
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  • Abstract from PubMed
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  • Journal Article Stewart JC, Villasmil ML, Frampton MW. Changes in fluorescence intensity of selected leukocyte surface markers following fixation. Cytometry Part A 2007;71A(6):379-385. R832415 (2007)
    R832415 (2008)
    R832415 (2010)
    R832415 (2011)
    R832415 (Final)
    R832415C003 (2006)
    R832415C003 (2010)
    R832415C003 (2011)
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  • Supplemental Keywords:

    human, air pollution, ultrafine particles, clinical studies, microparticles, platelets, diabetes,, RFA, Health, Scientific Discipline, PHYSICAL ASPECTS, Air, particulate matter, Health Risk Assessment, Risk Assessments, Physical Processes, atmospheric particulate matter, acute cardiovascular effects, long term exposure, atmospheric particles, airway disease, exposure, ambient particle health effects, human exposure, ultrafine particulate matter, atmospheric aerosol particles, PM, aersol particles, cardiovascular disease

    Relevant Websites:

    http://www2.envmed.rochester.edu/envmed/PMC/indexPMC.html Exit

    Progress and Final Reports:

    Original Abstract
  • 2007 Progress Report
  • 2008 Progress Report
  • 2009 Progress Report
  • 2010 Progress Report
  • 2011 Progress Report
  • Final Report

  • 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