Biomarkers of Exposure to Particulate Air Pollution in the Czech Republic
Joellen Lewtas1, Blanka Binkova2, Iva Miskova3, Pavel Subrt4, Jan Lenicek4, Radim Sram2
'U.S. Environmental Protection Agency, National Exposure Research Laboratory,
Human Exposure and Atmospheric Sciences Division, OEA-095, 1200-6,h Ave., Seattle, WA
98101
2 Institute of Experimental Medicine, Academy of Sciences of the Czech Republic and Regional
Institute of Hygiene of Central Bohemia, Prague, Czech Republic
3 District Institute of Hygiene, Teplice, Czech Republic
4 Regional Institute of Hygiene, Usti nad Labem, Czech Republic
Send proofs to corresponding author:
Joellen Lewtas, Ph.D.
US EPA, OEA 095
1200-6th Ave.
Seattle, WA 98101
phone: 206-553-1605 fax: 206-553-0119
email address: lewtas.ioellenfgepa.gov
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Key Words: exposure, biomarkers, particulate matter, polycyclic aromatic hydrocarbons, trace
metals, urinary metabolites, DNA adducts, fine particles, air pollution
ACKNOWLEDGMENTS
Many people are responsible for the success of the Teplice Program including the many staff
members of the District Institutes of Hygiene in Teplice and Prachatice, the Czech Hydro-
meteorological Institute and staff in the National Exposure Research Laboratory and the National
Health and Environmental Effects Laboratory of the U.S. EPA. Special thanks goes to
collaborators in the Czech Republic (R. Sram., F. Kot sovec, I. Benes, L.Dobias) and at the U.S.
EPA (R,Williams, D. Walsh, R. Watts, D. Otto, S. Perreault, S. Selevan, R. Stevens, T.
Hartledge, J. Pinto) and ollaborators from other institutions, especially S. Myers from U. of
Louisville in the US. This study was supported by the Czech Ministry of Environment (Teplice
Program) and the US Environmental Protection Agency/US Agency for International
Development.
HUMAN SUBJECT CONSENT
Informed consent was obtained from each of the subjects prior to partcipation in the studies
described in this paper.
DISCLAIMER
This paper has been reviewed in accordance with the US Environmental Protection Agency's
peer and administrative review policies and approved for publication. Mention of trade names or
commercial products does not constitute endorsement or recommendation for use.
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ABSTRACT
The use of biomarkers in the Teplice Program, provided a key tool to relate health
outcomes to individual personal exposures and to provide measures of confounding exposures.
This research program on the health effects of air pollution studied a population living in the
heavily industrialized district of Teplice in Northern Bohemia and compared the exposure and
health of this population to that of a non-industrialized district, Prachatice, in Southern Bohemia,
The studies included characterization of the environmental and personal air pollution exposure,
biomarkers, and studies on reproductive, respiratory, and neurobehavioral effects. Biomarkers
were measured in blood, urine, placenta, and sperm. The biomarkers included measures of
exposure (e.g., urine metabolites and blood metals), dose (e.g., DNA adducts), DNA damage,
genetic and cytogenetic effects, and susceptibility. During winter temperature inversions,
unusually high concentrations of a complex mixture of air pollutants were measured, including
fine particles, genotoxic organic compounds, and toxic trace elements. This population, however,
was also exposed to multiple pollutants via all pathways, and including pollutants resulting from
environmental exposures, occupational exposures, and personal habits (e.g., tobacco and alcohol
use). Longitudinal and repeated measures used individuals as their own control to examine the
influence of environmental exposures as they changed over time and season. Chronic and
seasonal exposures to elevated air pollution in the Teplice District were shown to have serious
adverse respiratory health consequences for children and reproductive effects in adults. Elevated
levels of air pollutants, even for short-term winter inversions resulted in measurable uptake,
metabolism, and excretion of polycyclic aromatic hydrocarbons, increased blood concentrations
of toxic metals, and resulted in DNA damage. Results of the exposure, biomarker, and health
studies indicated that environmental exposure to a complex mixture of air pollutants resulted in
significant elevations in personal exposure, uptake, excretion of pollutants and DNA damage.
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INTRODUCTION
The Northern Bohemia brown coal basin comprises four mining districts located in the
northwestern region of the Czech Republic, including Teplice district. The brown coal (lignite)
in this region is very high in sulfur and low in quality. It is surface-mined from open pits. This
coal is primarily used to produce steam and power for the heavy industrialization in this region.
Coal-fired power plants in this region produced 35% of the electricity utilized in the former
Czechoslovakia, The combustion of this coal combined with heavy industrialization of this region
over the past few decades, has resulted in some of the worst environmental pollution in Europe
(/). This same coal has also been used extensively for local area heating (e.g., homes,
apartments, offices) and local industries (e.g., glass production, chemical manufacturing, and
petrochemical industries). Prior to 1990, air pollution from these sources caused extensive
deforestation. A review of these environmental problems in the former Czechoslovakia was
published by Moldan and Schnoor (I).
The health consequence of environmental pollution in this region has been a major
concern of the public. Although exploratory analysis of data collected prior to 1989 suggested a
higher incidence of cancer, reproductive, and behavioral effects in this region(2), no
comprehensive research had been conducted to address these hypotheses. The Teplice research
program was developed in 1989 to evaluate the short-term and long-term health impact of air
pollution on the population. Teplice, one of the mining districts in Northern Bohemia, was
designated as a model district for investigation of the health effects of air pollution. The district of
Prachatice in Southern Bohemia, which was originally thought to have some of the cleanest air in
the Czech Republic, was selected as a comparison district.
The Teplice Program was initiated by the Czech Ministry of Environment in cooperation
with the Czech Ministry of Health late in 1990 to provide scientifically valid information needed
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to assess environmental health problems in the Northern Bohemian basin area (3). Thereafter, a
collaborative research program was developed with the U.S. Environmental Protection Agency
(US EPA) to include air pollution monitoring, human exposure, biomarker, and health effects
studies. This program has succeeded in bringing together many different research organizations
"and government laboratories in both the Czech Republic and the U.S. to accomplish the
multidisciplinary program.
The aim of the Teplice program was to evaluate the health impact of air pollution on the
exposed population. In addition, as the exposures in the region were expected to decrease,
longitudinal studies were planned to assess the impact of decreasing exposure on human health.
The central hypothesis in the Teplice.Program is that the polluted air in the Teplice district,
adversely affected the health of the population. A summary of the design and inital results of
these collaborative studies between scientists at the U.S. EPA, Office of Research and
Development and Czech scientists on the impact of air pollution on human exposure, biomarkers
of dose and genetic damage, reproductive, respiratory, and neurobehavioral effects through 1996
has been published^). The personal exposure, biomarker and health effects studies conducted
since 1992 and continuing into 2000 is shown in Table 1 by season and year. This paper reviews
the role of exposure assessment in these health effects studies through ambient monitoring,
personal exposure monitoring and the use of biomarkers of exposure.
CHARACTERIZATION OF MODEL DISTRICTS
Teplice district
The Teplice district is situated in the middle of the North Bohemian brown-coal basin,
which stretches from east to west and borders on the Krusne Hory (Ore Mountains) in the north
and on the Ceske Stredohori (Central Bohemian Highlands) in the south. This district is in a
basin in which the air pollution emissions are trapped during meteorological temperature
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inversions leading to high concentrations of air pollutants, particularly in the winter, Teplice has
a population of approximately 132,000, with an average density of about 285 people per km2.
The density is the highest in Northern Bohemia and, with the exception of much larger cities,
such as Prague, is one of the highest in the Czech Republic, About 50% of the employed
population works in industrial jobs and less than 3% in agriculture.
The district area comprises 469 km2 including agricultural land (170 km2) and forests (165
km2). By 1992, coniferous trees became virtually extinct and the deciduous trees sustained
serious damage due to the high air pollution emissions, A large part of the area has been
devastated by the strip-mining of coal and associated industrialization. In the KruSne Mountains,
there are rich deposits of fluorite which is extracted and processed locally. Most (75%) of the
brown coal extracted from the Czech Republic comes from the four districts of Northern
Bohemia, including Teplice. Extraction is almost entirely (91%) from open pits. In addition, half
of the coal extracted is processed locally. Consequently, half of all the sulphur dioxide and
nitrogen oxides emitted in the Czech Republic originate from the mining districts of Northern
Bohemia. Similarly, this region contributes about one quarter of the total air particulate matter
emissions in the country. Prior to 1996, most of the home heating systems used the brown
(lignite) coal as fuel. The Teplice air pollution research program (discussed in more detail
below) demonstrated that 30-40% of the human exposure to fine particles in this district was due
to home heating (J). As a result of this finding, incentives were provided to convert from coal to
gas heating in the Teplice district.
Prachatice district
The Prachatice district is situated in the southwestern part of the Southern Bohemia
region, near the border with Austria to the south, and the Sumava (Bohemian Forest) Mountains
to the southwest which form a natural border with Germany. This area has the lowest number of
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inhabitants of Southern Bohemia (50,740) and its population density (39.6 per km2) is one of the
lowest in the Czech Republic. There is a slight predominance of urban population (51.4%) living
in the four towns of the district. The area of the district of Prachatice (1375 km2) ranks fourth in
the region of Southern Bohemia (covering 12% of the region's area). Climatic conditions vary
greatly due to the varied elevation and character of the terrain, however, there are also frequent
fogs and temperature inversions in this district.
The district has vast, predominantly spruce forests covering the area of 71 thousand
hectares (52% of its area). In order to preserve the Sumava Mountain region, its water resources,
and tourist attractions, 1630 km2 of the range were declared a protected landscape area in 1963.
The largest part of the Sumava protected area is in the district of Prachatice. Aside from several
stone quarries, there is no mining in Prachatice.
CHARACTERIZATION OF THE AIR POLLUTION
Air pollutant measurements were made at two primary locations during this study(5). The
main monitoring site was located in the city of Teplice (North Bohemia), and the second was
located in the city of Prachatice (South Bohemia). Ambient SO2 and particulate matter < 10 ^m
(PM10) was monitored daily in both districts during the health studies. The inorganic and organic
composition of fine or respirable particle <2.5 fim (PM2.5) and coarse particles (2.5-10 jim) were
measured daily in the winter and periodically during the other seasons. These data were used to
characterize the ambient population exposures to air pollutants for the health studies and were
also used in receptor modeling studies to determine the relative contribution of the major air
pollution sources in the northwestern region of the Czech Republic. The composition of
emissions from power plants, glass factories, incinerators, motor vehicles, residential space
heating, and soils was also determined by using a dilution sampling probe.
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Particle mass and elemental composition were measured in Teplice and Prachatice
beginning in 1992. The concentrations of all pollutants measured, including particles, sulfur
dioxide, anions, toxic metals and polycyclic aromatic hydrocarbons (PAH) were significantly
higher in the winter than the spring and summer in both districts. The Teplice district, however,
had more severe pollution episodes and higher average winter pollutant concentrations. An
example is shown in Table 2 for the winter of 1993 where the average fine particle mass (PM25)
in Teplice was 122 jig/m3 compared to 44 jag/m3 in Prachatice and the spring and summer
average fine particle concentrations for Teplice and Prachatice were 28.7 ^ig/m3 and 17.9 jig/nr,
respectively. While the fine particle mass, trace elements and organic compounds in Teplice
were an average 2-3 fold higher than in Prachatice, the average sulfur dioxide concentrations
were generally at least 5-fold higher in Teplice than Prachatice. Seasonal differences in air
pollutant concentrations, even within the same district, were often greater than the district
differences.
The winter inversions produced the most dramatic pollution changes in Teplice. During an
inversion episode, between January 29 and February 6, 1993, the SO42" and the organic carbon
peak concentrations were 400 and 140 ng/m3, respectively. The presence of acidic particles was
indicated by the pH level, e.g., one sample collected at the peak of the episode had a pH of 4.1.
Fine particles (PM2.5) collected during the winter season were dominated by sulfates, organic
carbon, and trace metals. The fine particle mass observed in Prachatice was one-third the level
found in Teplice during winter episodes. Spring and summer particle concentrations were
substantially lower, reflecting the absence of home heating emissions and more favorable
meteorological conditions.
Samples were also analyzed for a series of carcinogenic polynuclear aromatic
hydrocarbons (PAH). Total PAH concentrations in Teplice in the winter were approximately
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twice the average winter concentrations of PAH in Prachatice. The most dramatic differences
(10X) were observed between the winter and spring/summer averages in Teplice. Evaluation of
the benzo(a)pyrene (BaP) to lead ratios in Teplice over time indicates the presence of at least two
sources of PAH (Pinto et al., 1998). During the summer, when mobile sources are the major
contributors to BaP, the ratio of BaP to Pb is about 0.01. During the winter, when the ratio is
0.05 to 0.15, emissions from inefficient combustion of brown coal (lignite) in home heating
systems is the most probable source of PAH.
Estimates of the contributions of emissions from power plants, incinerators, automobiles,
and home heating to the total fine particle mass measured during the 1993 winter at the main
monitoring site in Teplice were made by Pinto et al. (5) using chemical mass balance (CMB)
receptor modeling. This analysis suggested that most of the fine particle mass was produced by
home heating and power plants. Mobile sources accounted for only a few percent of the total
mass. Two major air pollution episodes occurred during the winter of 1993. The results of CMB
modeling for the first episode of February 1993 showed that power plant contribution was 30 ±
13% and home heating (and small industrial boilers) contributed 37 ± 10%. Most of the organic
carbon was contributed by home heating emissions. CMB results for the second episode showed
that the power plants contributed 55 ± 22% (mainly sulfate) and home heating contributed 29 ±
8% (mainly organic carbon) to the average fine particle mass.
ROLE OF EXPOSURE CHARACTERIZATION IN HEALTH STUDIES
During the 1993 Teplice winter episode, the meteorological conditions and pollution
levels were similar to those in London, December 1952 (6) where excess mortality was reported
to be more than 500 deaths per day. The average particulate matter concentrations (measured by
the blackness of the filter, "smoke") and S02 concentrations in London were about 1600 ng/m3
and 1800 pg/m3 (0.7 ppm), respectively. Particulate matter (PM:o) and SO2 concentrations
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measured in Teplice were comparable to data for the London episode (4,5), The acute effect of
particulate air pollution on human mortality has received new attention since studies using time-
series analysis in the 90's reported increases in mortality with increasing concentrations of
particulate air pollution at concentrations below the U.S.EPA ambient air quality standard for
PMio as reviewed by Schwartz (7), A recent report by Peters et al. (5) on the acute effects of total
suspended particle (TSP) concentrations on mortality in Northern Bohemia (including Teplice
district) over the 13 years from 1982-1994 found a significant increase in mortality with
increasing concentrations of total suspended particulate and an even greater effect for PMio
(8.8% increase in mortality by 100^g/m3), during 1993 and 1994 when PMio measurements were
initiated as part of the U.S. EPA's collaborative support.
The initial observations of dramatic seasonal differences in the pollution levels within the
Teplice district, at times, resulted in higher differences within district than between the Teplice
and Prachatice districts. This finding in the early exposure characterization studies led to an
increased emphasis on the longitudinal and time series design that allowed comparisons either
within an individual across time or within populations in the same district across seasons as
shown in Table 1. These designs also took advantage of the early results from questionnaire data
that showed important socioeconomic, behavioral (e.g., smoking differences), and ethnic
differences between the populations in these two districts that confounded analysis based solely
on comparing Teplice and Prachatice populations (4,9).
In addition to characterization of the air pollution using source and ambient monitoring
described above, indoor/outdoor and human exposure monitoring studies were planned to provide
direct personal exposure measurements. These measurements were critical to the development,
validation, and interpretation of biomarkers used in many of the health studies. As shown in
Figure 1, these biomarkers range from trace metals in blood which can be directly related to metal
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exposures to urinary PAH metabolites and DNA adducts reflecting internal exposure to specific
contituents of the complex exposure mixture. Trace metals found in the ambient and personal
monitoring were, where possible, measured in blood, hair, and urine in children to facilitate
interpretation of the neurobehavioral and respiratory studies. The initial population and air
•pollution characterization studies in the two districts were critical to the final study designs for
the reproductive studies. Personal exposure and biomarker studies were initiated early in the
research and several biomarkers were included in the adult male and female reproductive effect
studies.
BIOMONITORING EXPOSURE IN THE RESPIRATORY AND NEUROBEHAVIORAL
EFFECTS STUDIES
The respiratory and neurobehavioral studies were conducted in school children ranging
from 2nd to 8th grade and they included both cross-district and longitudinal design so that
evaluations covered the high-pollution winter period and the two lower-pollution periods in
Teplice and Prachatice {9-11). As shown in Table 1, studies of the respiratory and
neurobehavioral effects in 8th grade students was initiated in the winter of 1992 with a cross-
sectional respiratory study initiated the following fall. A significantly higher prevalence of
adverse respiratory symptoms and decreased lung function were found in the Teplice district than
in Prachatice (P).
Biomonitoring children's hair and urine for arsenic (As) and mercury (Hg) was suggested
by the relatively high levels of trace metals in PM2.5 air pollution derived from the brown coal
combustion (Table 2), including arsenic (As), mercury (Hg), cadmium (Cd), and selenium (Se).
In 1993, hair and urine samples were also obtained from the 600 2nd-grade children participating
in these studies to determine the association of neurobehavioral performance and biological
measures of As and Hg exposure. Levels of As and Hg observed in children from both districts
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were surprisingly low. The average Hg level in hair in Teplice children was 0.27ppm and the
average in Prachatice children was 0.89 ppm. Hg levels in urine were similarly low.
Lead (Pb) exposure has been associated with a broad spectrum of neurobehavioral effects
(12) including the performance of East German children on tests similar to those used in these
studies(13). Venipuncture sampling, the recommended method to determine Pb exposure, was
deemed to be an unacceptable use of invasive biological sampling in this school population.
Therefore, blood Pb levels were not measured in the children participating in the neurobehavioral
study. Assessment of Pb exposure in children living in the study area utilized blood samples
obtained from 200 children that were referred to hospitals in the Teplice and Prachatice districts
for blood work not related to these studies. Blood Pb mean levels of 5.0 |ig/dl (range 1.0-17.6)
were found in children from Teplice and 3.8 jig/dl (range 0.9-14.0) in children from Prachatice as
shown in Table 3, While these levels do not suggest that children in either district are at serious
risk for lead poisoning, Winneke et al. (13) have shown an association of comparable blood Pb
levels and performance on similar neurobehavioral tests in 6-year-old East German children.
A majority of the objective neurobehavioral performance measures reported by Otto et
al.(/0) did not differ between the districts, even though significantly higher teacher referrals for
neurobehavioral clinical assessments were made in Teplice, than Prachatice. In 2nd-grade
children, weak but significant associations were found in the performance of three
neurobehavioral tests and hair Hg levels. These associations also remained after controlling for
possible confounders. On the other hand, no significant district differences in the
neurobehavioral performance of 4th-grade children were found when parental education and
other covariates were controlled. Nor were any significant associations of neurobehavioral
measures and metal levels found in the 4th-grade children (10).
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Visual contrast sensitivity (VCS) tests have been used successfully in medical diagnosis
and subclinical neurotoxicity detection. VCS deficits were seen in exposed 2nd grade children but
not with the 4th grade and no consistent relationship as found between visual tests and As or Hg
levels in the children's hair. Although hearing impairment was not evaluated in this study,
Bencko et al. (14) have also reported impaired hearing in children associated with As from the
combustion of coal in the nearby Slovak Republic.
PERSONAL EXPOSURE AND BIOMARKERS OF EXPOSURE, DOSE, AND
SUSCEPTIBILITY
Personal exposure studies were initially conducted to compare the ambient concentrations
and personal air exposures to fine particles, PAH, and organic mutagens. A personal exposure
monitor designed for biomarker studies and shown in Figure 2 was used in these studies (15).
The personal monitor was used to measure the personal air exposure of policemen, coal miners
and other workers in the Teplice districted). Stationary medium-volume PMio samplers and
high-volume PM2.5 and TSP samplers were also used to collect particulate matter for
measurement and characterization. The initial studies conducted in the winter of 1991 with a
group of Teplice policemen showed that personal BaP exposures averaged 40 ng/m3. Ambient
high-volume air sampling results from 12-h nighttime samples collected in Teplice between
February 17 and March 27,1992 showed particle-associated BaP averaged 12 ng/m3 and ranged
from 2-34 ng/m3. The sixteen PAH quantified averaged 131 ng/m3 for the same periods.
Approximately 50% of the particle-bound PAH concentrations in Teplice air included compounds
that are carcinogenic in animals. The concentration and mutagenic potency of genotoxic
substances adsorbed to the particles were determined by the Ames plate incorporation assay. The
mutagenic potency of extractable organic matter from ambient air particles was higher than that
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for U. S. residential areas that are heavily impacted by wood smoke but similar to that from U.S.
cities more heavily impacted by vehicle emissions^6).
A larger and more comprehensive personal exposure and biomarker study was initiated in
1992 after the pilot studies. The objectives of this project were to evaluate simultaneously
personal exposure to air pollution and internal measures of exposure, dose, genetic effects, and
susceptibility using a series of biomarkers. The relationship between ambient concentrations of
air pollutants and human exposure was also evaluated using simultaneous ambient monitoring for
fine particles. PAH were selected as the air pollutant marker for monitoring personal exposure
because the major source of air pollution in this region was coal combustion. PAH and other
genotoxic polycyclic aromatic compounds adsorbed onto fine soot particles have been estimated
to be a significant source of lung cancer risk. The biomarkers of internal exposure, dose, genetic
effects and susceptibility were selected based on the knowledge that PAH is rapidly metabolized
via microsomal oxidative pathways to reactive intermediates that may bind to protein and DNA
or that may be excreted as phenol and diols in urine(/7). Biomarkers of internal exposure and
dose to PAH selected for this project are illustrated in Figure 3. These biomarkers included PAH
protein adducts, PAH-DNA adducts, PAH urinary metabolites, and urinary mutagenicity.
Cytogenetic effects were measured using sister chromatid exchanges and chromosome
aberrations. Metabolic susceptibility biomarkers, glutathione-S-transferase(GSTMl) and N-
acetyltransferase(NAT2) were measured using the polymerase chain reaction (PCR) method.
Personal exposure monitoring for PM2.5 was conducted for the 24 h period prior to
collection of blood and urine. Particle extracts were analyzed for carcinogenic PAH (PAHcar) and
also for their ability to form DNA adducts in-vitro as reported by Binkova, et al.(/§). A group of
30 women working primarily outdoors in the Teplice district as postal workers and gardeners was
compared with a group of 30 women from the Prachatice district who were gardeners and
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kindergarten teachers. These 60 women were monitored during the winter season of 1992
(Biopem 1). The mean personal exposures and biomarkers for several of the parameters
measured in this study are shown in Table 3. Although most of the personal exposures are higher
in the Tepliee group, the PM2.5 was only three fold higher than Praehatiee and the PAHcar were
only two fold higher. The difference between Tepliee and Praehatiee biomarkers (e.g., PAH
urinary metabolites and blood metals) were less than two-fold different between the two districts.
In 1993, a repeated measures follow-up study (Biopem 2) was started with a group of ten
nonsmoking women from Tepliee. These ten women had their personal exposure and biomarkers
monitored over time during the fall and winter from 1992 through 1994 as shown in Figure 4.
Both studies examined the influence of personal exposure and other factors that alter exposure
and metabolism (e.g., GSTM1 genotype, age, diet) on biomarkers of exposure, dose and genetic
effects in these women in both districts {19,20), Although both studies showed a significant
relationship between DNA adducts and exposure to particle associated carcinogenic PAH, the
repeated measures study (Biopem 2) was more statistically powerful than the initial study
(Biopem l)(p<0.001 compared to p<0.05) due to the use of each subject as their own control over
time.
The correlation between personal exposure to fine particles, personal exposure to PAHcar
and several of the other exposure biomarkers are shown in Table 4. Significant correlations were
observed between the personal exposures to PM2.5 or PAHcar and both blood lead and blood
selenium concentrations. Urine samples were collected for exposure marker analysis including
PAH metabolite analysis, urinary mutagenieity(2/), and cotinine analysis. Cotinine, a metabolite
of nicotine, was used to control for exposure to tobacco smoke as one of the confounding factors
in these studies. The urine markers were all adjusted for creatinine content to control for
variations in urine volume. The urinary PAH metabolites were also significantly correlated with
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personal exposure to either PM2.5 or PAHcar. DNA damage as measured by the comet assay
(percentage of DNA in the comet tail) also correlated with exposures to respirable particles
(r=0,304, p=0.015). Within the nonsmokers from Teplice significant correlations of personal
exposure to carcinogenic PAH with DNA adduct levels in white blood cells (WBC) analyzed by
32P-postlabeling using butanol enrichment procedures were found (initial study: r=0.54, p=0,016;
follow-up study: r=0.71, p<0.001).
DNA adducts measured in the circulating blood cells appear to be a useful biomarker of
internal dose. Futher analysis was conducted of DNA adduct dosimetry over a wider range of
exposures from low and moderate environmental exposures to higher occupational exposures(22).
DNA adduct levels in the WBCs were significantly correlated with environmental exposures to
fine particles and carcinogenic PAH adsorbed to particles. However, at the higher occupational
levels, such as those found in coke oven workers in Ostrava, CZ, the exposure-DNA adduct
relationship became nonlinear(superlinear). Under these extremely high occupational exposure
conditions, the relative DNA adduct level per unit of exposure (DNA-binding potency) was
significantly lower than measured at environmental exposures (22).
Metabolic susceptibility biomarkers were determined by genotyping the DNA isolated
from the WBCs. The influence of GSTM1 genotype on the biomarkers of exposure and DNA
damage were evaluated by Binkova et al.(20) and Costa et al.(25). GSTM1 genotype had a
significant effect on urinary PAH metabolites (p=0.037), urine mutagenicity (p=0.033) and comet
assay (p=0.002) when GSTM1 genotype was considered as a single factor affecting these
biomarkers. The influence of GSTM1 and NAT2 genotypes on the dose-response relationship
between personal exposure to PAHcar and multiple biomarkers in this cohort have been reported
by Costa, et al.(2J). Stratifying the cohort by allele specific genotypes decreased the inter-
individual variability and increased the correlation observed between personal exposure and
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several biomarkers. This suggests that metabolic susceptibility difference between individuals in
the population accounts for some of the interindividual variability in the population,
APPLICATION OF BIOMARKERS TO MALE AND FEMALE REPRODUCTIVE
STUDIES
Semen Studies
From 1992 through 1997, a series of collaborative studies to evaluate male reproductive
health were conducted as part of the Teplice Program. The first study was designed to evaluate
reproductive history and semen quality in young men (18 year old military recruits) living in
Teplice or Prachatice. Associations between air pollution levels (averaged over the three months
preceding sampling in either late winter or early fall) and semen quality were found for several
outcomes indicative of sperm quality (sperm motility and morphology) with no effect on sperm
numbers (24). This study also included a new measure of sperm chromatin integrity, the sperm
:
chromatin structure assay, that showed propotionally more sperm with abnormal chromatin
associated with periods of elevated air pollution (24).
Subsets of men from this study were also studied for potential effects of smoking and
exposure to air pollution on a molecular marker of sperm nuclear integrity. For this cytogenetic
assay, sperm are labeled with chromosome-specific probes such that extra chromosomes
(aneuploidy) can be detected using fluorescence in situ hybridization or FISH (25). Men who
smoked a pack of cigarettes per day had a significantly elevated incidence of sperm with sex
chromosome (YY) aneuploidy (26), compared with non-smokers, demonstrating the importance
of considering smoking as a potential modifier in future studies. These data need to be interpreted
with care, since smoking and alcohol consumption were highly correlated in these men;
therefore, an effect due to alcohol could not be ruled out. In addition, non-smokers exposed to
the highest levels of air pollution exhibited increased sperm aneuploidy when compared with
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men from the same district exposed to low levels of air pollution (27). If aneuploid sperm
fertilize an egg, the resulting conceptus may have serious developmental abnormalities. Thus,
inclusion of molecular markers of sperm nuclear integrity (effect biomarkers) provided valuable
new information about the potential risk of exposure to air pollution and male-mediated adverse
pregnancy outcomes.
Based on these results, a longitudinal semen study was initiated in 1995. This study
involves serial sampling of men from Teplice over a period of 2.5 years and is designed to
include sampling during periods of both low and high pollution. In addition, internal biomarkers
of exposure to metals and cotinine (to control for smoking) were included. This design has
several advantages: each man serves as his own baseline; and, changes in semen quality can be
related to the timing of spermatogenesis (since the interval between exposure and sampling will
be known). Genetic biomarkers of effect are being evaluated in all samples including: sperm
aneuploidy (FISH analysis) and measures of DNA damage in sperm, the sperm chromatin
structure assay (SCSA), and the deoxynucleotidyl transferase-mediated nick end labeling assay
(TUNEL) conducted by flow cytometry, and the single cell gel comet assay for DNA damage
(electrophoresis assay). Inclusion of these biomarkers will provide a comprehensive assessment
of semen quality, and allow comparisons across methods to help determine the best options in
future semen studies.
Pregnancy Outcome
The pregnancy outcome study was initiated in 1994 after extensive planning and peer
review. Preliminary analyses on 2,500 pregnancies collected during the first 15 months of the
study showed the prevalence of low-birth-weight infants in the district of Teplice (8.8%) was
significantly higher (PO.OOOl) than in the district of Prachatice (3.3%) (4). Similarly, the
prevalence of premature births was 6.2% in Teplice and 3.4% in the Prachatice district (P<0.01).
18
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The populations of the two districts, however, differed significantly in their ethnic composition.
About 14.1% ofbirths in Teplice, but only 2.9% in the Prachatice district, were of Gypsy
ethnicity. Gypsies, with origins in India, differ from other inhabitants (mostly of European origin)
in many biological and social characteristics (28). Differences were observed in their pregnancy
outcomes: 13.4% of Gypsy births were premature and about 23.6% of infants weighed less than
2,500g at births. Thus, the difference observed in the two districts fell substantially after
exclusion of Gypsy births; however, even after this exclusion the difference remained statistically
significant.
A recent publication reports the final results of the impact of maternal exposure to PM2.s
and PMio air pollution on fetal growth in the more polluted Teplice district from April 1994 to
March 1996 (29). This study measured fetal growth in early'gestation by determining intrauterine
growth retardation (IUGR) which determines birth weight for gestational age. Dejmek et al., (29)
defined an IUGR birth as one whose weight fell below the 10th percentile, by gender and
gestational week, for live births in the Czech Republic from 1991-1993. Only the full term single
births of European origin in Teplice district were reported. Nine month exposure windows, from
conception to delivery, were examined to determine the relationship between PM2.5 and PM10
concentrations and IUGR to determine which stages of prenatal development were sensitive to air
pollution exposures. Only the first gestational month showed a significant impact of particulate
air pollution on IUGR, suggesting that exposure to air pollution early in pregnancy may adversely
affect fetal growth. Important parental characteristics such as maternal age, education, alcohol
consumption, maternal active and passive smoking, and paternal smoking were evaluated.
Season and year were tested for confounding in these studies. The adjusted odds ratios (OR) for
risk of IUGR increased with increasing exposure to both PM2.5 and PM10, however, only
exposure to PM10 was statistically significant. The OR and 95% confidence intervals (shown in
19
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parentheses) for IUGR risk in the first month of pregnancy for PMio concentrations between 40
to <50 ng/m3 were 1.62 (1.07-2.50) and for high concentrations above 50 jig/m3 the OR was 2.64
(1.48-4.71). For PM2.5, the trend was similar. The continuous data analysis showed increases in
risk of IUGR for the first month of pregnancy for each 20jig/m3 increase of PM. The OR for
PM10 was 1.50(1.15-1.96); for PM2,5 it was 1.34 (0.98-1.82; p= 0.06). These results suggest an
increasing dose-response relationship between risk of IUGR and exposure to air pollution as
measured by particulate matter.
DNA adducts in placentas of mothers from a subset of the women in the more polluted
Teplice district were significantly higher than in the placentas from Prachatice women (30).
Further analysis of the DNA adducts showed a highly significant relationship with IUGR, NAT2
genotype, Vitamin C level and smoking (31). Chromosomal aberrations were significantly higher
in the Teplice mothers during pregnancy than the Prachatice mothers (32) but no differences were
found at delivery in the blood of the mothers or the cord blood. No differences were found in the
venous and cord blood DNA damage when measured as single strand breaks with the comet assay
(32). In vitro studies of the genotoxicity and embryotoxicity of the particulate during the
summers and winters of 1993 and 1994 showed that the organic fraction containing PAH is the
most active in forming DNA adducts and inducing embryotoxicity (33).
CONCLUSION
The use of biomarkers in these studies provided a key tool to relate ambient exposures and
personal exposures to health outcomes and to provide measures of confounding exposures such as
tobacco smoke. Biomarkers of exposure, dose, susceptibility, and molecular effects were directly
related to health outcomes. The biomarker studies also used individuals as their own control to
examine the influence of environmental exposures as they changed over time and season.
20
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Chronic and seasonal exposures to elevated air pollution in the Teplice district were shown to
have serious adverse respiratory health consequences for children and reproductive effects in
adults. Elevated levels of air pollutants, even for short winter inversions resulted in measurable
uptake, metabolism, and excretion of genotoxic organic compounds (polycyclic aromatic
hydrocarbons), increased blood concentrations of toxic metals, and resulted in DNA damage.
Pregnancy outcomes were affected by a number of factors including the environmental air
pollution, ethnicity, and lifestyle (e.g., smoking). Results of this study thus indicate that exposure
to a complex mixture of air pollutants can affect multiple functional endpoints and produce DNA
damage.
21
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Techno! 26:14-21;1992.
2. Sram RJ, Roznickova I, Albrecht V, Berankova A, Machovska E. Monitoring congenital
anomalies in populations exposed to environmental mutagens. In: Mechanisms of environmental
mutagenesis - carcinogenesis, (Kappas A,ed). New York: Plenum Press, pp 255-266; 1990.
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Workshop on Air Pollution Epidemiology. Air Poll Epidemiol Rep Ser, Report No. 3 (Rudnai
P,ed). Brussels:DG XII CEC, pp. 132-139; 1992.
4. Sram, R., Benes, I., Binkova, B., Dejmek, J., Horstman, D., Kotesovec, F., Otto, D.,
Perreault,S., Rubes, J., Selevan.S., Skalik, I., Stevens, R., and Lewtas, J. Teplice Program- The
impact of air pollution on human health, Environ. Health Perspect. 104, 699-714; 1996.
5. Pinto, J.P., Stevens, R.K., Willis, R.D., Kellogg, R., Mamane, Y., Novak , J., Santroch, J.,
Benes, I., Lenicek, J., and Bures, V. Czech Air Quality Monitoring and Receptor Modeling
Study, Environ. Sci. Technol. 32:843-854; 1998.
6. Wilkins ET. Air pollution aspects of the London fog of December 1952. QJR Meteorol Soc
80:267-271; 1954.
7. Schwartz, J. Air pollution and daily mortality: a review and meta analysis. Environ. Res.,
56:204-213; 1994.
8. Peters, A., KoteSovec, F., Skorkovsky, J., Brynda, J., and Heinrich, J. Acute Effects of
Suspended Particle Concentrations in the Atmosphere on Mortality-A Study Comparing
Northeast Bavaria and Northern Bohemia. GSF-Research Center for Health and the
Environment, Neuherberg, Germany, Final Report # GSF-EP-S 1/99, 60pp; 1999.
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9. Horstman, D., Kotesovec, F., No i k a, J., R. Sram. Pulmonary Functions of School Children
in Highly Polluted Northern Bohemia. Archives of Environmental Health, 52:56-62; 1997.
10. Otto,D., Skalik I, House, D., Tse,J., Hudnell, K. Neurobehavioral Evaluation System
(NES2): Comparative Data from 2nd, 4th and 8th-grade Czech Children. Neurotoxico! Teratol
'18:421-428; 1996.
11. Hudnell,K., Skalik I, Otto, D., House,D., Subrt,P., Sram RJ Visual Contrast Sensitivity
Deficits in Bohemian Children. NeuroToxicology, 17:368-375; 1996.
12. Davis JM, Otto D, Weil D, Grant L. The comparative developmental neurotoxicity of lead.
Neurotoxicol Teratol 12:215-229; 1990.
13. Winneke G, Altmann L, Kramer U, Turfeld M, Behler R, Gutmuths FJ, Mangold M.
Neurobehavioral and neurophysiological observations in six year old children with low lead
levels in East and West Germany. Neurotoxicology, 15:705-714; 1994.
14. Bencko V, Symon K, Chladek V, Pihrt J. Health aspects of burning coal with a high arsenic
content. II: Hearing changes in exposed children. Environ Res 13:386-395;! 997.
15. Williams, R.; Watts, R.; Stevens, R.; Stone, C.; Lewtas, J. Evaluation of a personal air
sampler for twenty-four hour collection of fine particles and semi-volatile organics. J. Exposure
Analysis and Environmental Epidemiology, 2: 158-166; 1999.
16. Watts R, Lewtas J, Stevens R, Hartlage T, Pinto J, Williams R, Hattaway K, Miskova I,
BeneS I, KoteSovec F, Sram RJ. Czech-U.S.EPA health study: Assessment of personal and
ambient air exposures to PAH and organic mutagens in the Teplice district of Northern Bohemia.
Intern J Environ Anal Chem 56:271-287; 1994.
17. Talaska, G., Underwood, P., Maier, A., Lewtas, J., Rothman, N., and Jaeger, M. Polycyclic
aromatic hydrocarbons (PAHs), Nitro-PAHs and related environmental compounds: Biological
markers of exposure and effects, Environ, Health Perspect. 104, 901-906; 1996.
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18. Binkova B, King,L. and Lewtas, J. DNA Adducts.formed in vitro from fractionated extract of
urban air particles, Polycyclic Aromatic Compounds, 10:243-250; 1996,
19. Binkova B, Lewtas J, Miskova I, Lenicek J, Sram RJ. DNA adducts and personal air
monitoring of carcinogenic polycyclic aromatic hydrocarbons in an environmentally exposed
population. Carcinogenesis 16:1037-1046; 1995.
20. Binkova B, Lewtas J., Miskova I, Rossner P, Cerna M, Mra k ova G, Peterkova K, Mumford
J, Meyer J, Sram RJ. Biomarkers studies in the Northern Bohemia. Environmental Health
Perspectives, 104:591-597; 1996.
21. Cerna M., Pastorkova A., Myers S.R., Rossner P.,Binkova B. The use of a urine mutagenicity
assay in the monitoring of environmental exposure to genotoxins. Mutation Research, 391,99-
110; 1997.
22. Lewtas, J., Walsh,D., Williams, R. and Dobias, L. Air Pollution Exposure - DNA Adduct
Dosimetry in Humans and Rodents: Evidence for Non-linearity at high Doses, Mutation
Research, 378:51-63 1997.
23. Costa, D.J., Slott, V., Binkova, B., Myers, S.R., and J. Lewtas Influence of GSTM1 and
NAT2 Genotypes on the Relationship between Personal Exposure to PAH and Biomarkers of
Internal Dost, Biomarkers, 3:411-424; 1998.
24. Selevan SG, Borkovec L, Slott V, Zudova Z, Rubes J, Evenson, DP, Perreault SD. Semen
quality and reproductive health in young Czech men exposed to seasonal air pollution .
Environmental Health Perspectives 108:887-894; 2000.
25. RubeS, J., Lowe, X., Cassel, M., Moore, D., Perreault, S, Slott, V., Evenson, D., Zudova, Z.,
Borkovec, L,, Selevan, S., and Wyrobek, A.J. Aneuploidy detection in sperm nuclei using
fluorescence in situ hybridization. Archivos de Zootecnia 45:323-327, 1996.
24
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26. Rubes, J., Lowe, X., Moore, D., Perreault, S., Slott, V., Evenson, D., Selevan, S. and
Wyrobek, A. Smoking cigarettes is associated with increased sperm disomy in teenage men.
Fertility and Sterility, 70:715-723; 1998.
27. Robbins W.A., RubeS, J., Selevan, S.G., Perreault, S.D. Air pollution and sperm aneuploidy
in healthy young men. Environmental Epidemiology and Toxicology, 1:125-131; 1999.
28. Bemasovska K.» Bemasovsky I., Poradovsky K. et al. Proposal of low birth weight limit for
Gypsy mature babies. In: Anthropology of maternity. Prague: Charles University, 173-175; 1977.
29. Dejmek J, Selevan SG, Benes, I., Solansky, I., and Sram, RJ, Fetal growth and maternal
exposure to particulate matter during pregnancy, Environmental Health Perspectives, 107:475-
480; 1999.
30. Topinka, J., Binkova, B., Mrackova, G. Stavkova, Z, Peterka, V., Benes, I., Dejmek, J,
Leni ek, J., Pil ik, T., Sram, R.J. Influence of GSTM1 and NAT2 genotypes on placental DNA
adducts in an environmentally exposed population. Environ. Molecular Mutagenesis, 30: 184-
195; 1997.
31. Sram, R.J., Binkova, B,, Rossner P., Rubes, J., Topinka, J, Dejmek, J. Adverse reproductive
outcomes from exposure to environmental mutagens. Mutation Research, 428:203-215; 1999.
32. Sram, R.J. Impact of air pollution on reproductive health. Environmental Health Perspectives,
107:542A-543A; 1999.
33. Binkova B, Vesely, D., Vesela, D. Jelinek, R., and Sram R.J. Genotoxicity and
embryotoxicity of urban air particulate matter collected during winter and summer period in two
different districts of the Czech Republic, Mutation Research 440:45-58; 1999.
34. Lewtas, J, Mumford, J, Everson,R, Hulka,B, Wilkosky, T, Kozumbo,W, Thompson, C ,
George, M, L Dobias, Sram R, Li,X and Gallagher,J Comparison of DNA Adducts from
25
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Exposure to Complex Mixtures in Various Human Tissues and Experimental Systems,
Environmental Health Perspectives, 99: 89-97; 1993.
26
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FIGURE LEGENDS
Fig. 1 Biomarkers in air pollution studies, including pollutants measured in blood, urine
metabolites, and DNA or protein (e.g., hemoglobin) adducts are related to ambient
and personal exposures to specific constituents of the complex air pollution
mixture.
Fig. 2 EPA's personal exposure monitor used in biomarker studies and described in detail
by Williams et al, 1999.
Fig. 3 Polycyclic aromatic hydrocarbon (PAH) biomarkers of exposure and dose.
Fig, 4 Repeated measures study of DNA adducts, personal exposure to respirable
particles(RSP), and carcinogenic PAH in ten nonsmoking women in Teplice
plotted over time from 1992 to 1994.
27
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Table 1. Summary of Studies by Season and Year
Year Winter: High Pollution Period
1992 Exposure & Biomarker Studies Initiated
Respiratory Study of 8th Grade Children
Neurobehavioral Study of 8th Grade
1993 Exposure & Biomarker Repeated Measures
Respiratory Cross-Sectional Study
Semen Study
1994 Pregnancy Outcome Study Initiated
Semen Study
1995 Exposure & Biomarker Repeated Measures
Pregnancy Outcome Study
Semen Study (18 yr old men)
1996 Pregnancy Outcome Study
Longitudinal Semen Study
1997 Pregnancy Outcome Study
Longitudinal Semen Study
1998 Pregnancy Outcome Study
3 Includes spring, summer and fall
Low Pollution Period8
Respiratory Cross-Sectional Study
Neurobehavioral Study of 8th Grade
Semen Study (18 yr old men)
Respiratory Cross-Sectional Study
Neurobehavioral Study of 2nd Grade
Semen Study
Pregnancy Outcome Study
Neurobehavioral Study of 4th Grade
Longitudinal Respiratory Study
Pregnancy Outcome Study
Longitudinal Semen Study Baseline
Pregnancy Outcome Study
Longitudinal Semen Study
Pregnancy Outcome Study
Longitudinal Semen Study
Pregnancy Outcome Study
28
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Table 2. Fine Particle Composition in Teplice and In Prachatice, Winter and Summer 1993a
Teplice
Winter Summer
Prachatice
Winter Summer
S02 (ng/m )
PM25(Hg/m3)
Total Mass
Metal Oxides •
Sulfateb
PAH (ng/m3)
Sum of PAHs
Benzo[a]pyrene
153(60) na
122 (3.0)
6.5 (0.5)
41.9 (5.7)
278(12)
8 (0.4)
28.7(1.2)
1.9 (0.1)
10.2(1.3)
27(1)
0.5 (0.4)
29(11)
44.0 (0.8)
1.8 (0.1)
9.5(1.2)
163 (8)
4.7 (0.24)
4.4(1.6)
17.9 (0.4)
1.1 (0.07)
6.7 (0.9)
24(1)
0.14(0.01)
Trace Elements (ng/m )
As
Se
Br
Pb
44.5 (4.5)
8.1 (1.0)
18.5 (2.1)
108(11)
nd
nd
5.1 (0.9)
39.8 (4.4)
30. (2.6)
2.1 (0.4)
11. (1.1)
54. (4.6)
1.9 (0.6)
0.5 (0.2)
4.2 (0.5)
23. (2.3)
a Fine particles (<2.5|i) or PM2.5 data are shown with estimated uncertainty in parentheses; na =
measurements not available; nd = detected at the 3 standard deviation level in fewer than half the
samples; Winter included January through March for both Teplice and Prachatice and Summer
Included May through August for both Districts.
bSulfur expressed as ammonium sulfate.
29
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Table 3. Comparison of Personal Exposure of Nonsmokers in Teplice and Prachatiee
during November and December of 1992a
Exposure Measurement
Teplice Prachatiee
Personal Exposure Monitor
PM2.5 (ng/m3) 46 ±25 (n=21) 16 ±17 (n=30)
PAHcar (ng/m3) 13 ±7 (n=21) 7±4 (n=30)
Benzo(a)pyrene (BaP) (ng/m3) 2.5 ±1 (n=21) 1.1 ± 1 (n=30)
Urinary Metabolites
(ng/mg creatinine)
Total PAH Metabolites 231 ±72 (n=21) 126 + 39 (n=30)
BaP Metabolites 5 ± 6 (n=21) 3 ±2 (n=30)
Trace Metals in Blood
(jag/dl)
Mercury (Hg) 0.1 ± 0.05 (n=21) 0.09 ± 0.05 (n=30)
Selenium (Se) 10.0 ±2.1 (n=21) 6.9 ±1.2 (n=30)
Lead(Pb)
Biomarker Study 6.9 ±3.3 (n=21) 4.7 ±2.0 (n=30)
Children's Study (n = 200) 5.0 (1.0-17.6)b 3.8 (0.9-14.0)b
a mean ± standard deviation (n=number of subjects)
b range
30
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Table 4. Correlation between Personal Exposure and Biomarker Measurements
Exposure Measure PM2.5 PAHcar
Personal Exposure Monitor
Carcinogenic PAH (PAHcar) 0.79a p < 0.000lb (60)c 0.79a p < 0.000lb (60)c
Benzo(a)pyrene (BaP) 0.85 p< 0.0001 (60) ND
Trace Metals in Blood
Blood Pb 0.39 p = 0.003 (58) 0.35 p = 0.009 (58)
Blood Se 0.55 p< 0.0001 (60) 0.49 p = 0.0002 (58)
Urinary PAH Metabolites 0.48 p = 0.0002 (58) 0.40a p = 0.002b (60)c
NAT2-slow Genotype ND 0.58a p = 0.001b (29)c
Urinary Mutagenicity ND 0.67 p = 0.016 (21)e
(MS YG1041 -S9)"d
DNA adducts 0.16 p = 0.32 (51)e
GSTM1-positive Genotype Subjects from both Districts 0.59 p = 0.005 (21)
Teplice Subjects 0.54 p = 0.01 (21)e
Repeated Analysis of 10 Teplice Subjects 0,71 p < 0,001 (10)e
a Spearman rank correlation;b significance;c number of observations
d Microsuspension assay
e Nonsmokers only
31
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AMBIENT & PERSONAL EXPOSURE:
PM, METALS, ORGANICS, GASES
POLLUTANTS IN BLOOD
(Blood Pb, COHb)
URINE METABOLITES
{PAH metabolites)
V
DNA or Hb
ADDUCTS
-
SPECIFIC CONSTITUENTS
OF COMPLEX EXPOSURE
J»«' M f
-------
PEG coated
to pump
inlet/impactor removes coarse
particles
3 filter pack
for
fine particles
and nicotine
XAD cartridge
collects
semivolatile
organics
-------
External
Exposure
Personal
Exposure
EXTERNAL METABOLIC
DOSE ^ ACTIVATION
o o
o
f
o
INTERNAL DOSE
DETOXIFICATION
Biologically
Effective Dose
REACTIVATION
J
Protein
Adducts
DNA
Adducts
EXCRETION
OH
URINE
METABOLITES
MUTAGENICITY
Pt'iute 3
-------
Repeated Sampling Study
52 o. 20
2
11/30/93
11/24/92
10/25/93
1/5/94
2/1/94
Time Period
RSP PAH DNA Adducts
+ +
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NFRT RTF HFASD-OO 7L? TECHNICAL REPORT DATA
heju, Kir HKA&u-uu znz. (Please mad Instructions on the reverse before complet?
1. REPORT NO.
EPA/600/A-01/009
2.
3. REC
4. TITLE AND SUBTITLE
Biomarkers of Exposure to Particulate Air Pollution in the Czech Republic
5. REPORT OATE
Oct. 16,2000 Date of Preparation
6. PERFORMING ORGANIZATION CODE
7. AUTHORS
Joellen Lewtas, Blanka Binkova, Iva Miskova, Pavel Subrt, Jan Lenicek, Radim Sram
8. PERFORMING ORGANIZATION REPORT NO.
9. PERFORMING ORGANIZATION NAME AND ADDRESS
US EPA .Office of Research and Development, NERL and NHEERL, RTP, NC 27711
Institute of Experimental Medicine, Academy of Sciences, Prague, Czech Republic and
Institutes of Hygiene of Central Bohemia, Teplice and Usti, Czech Republic
10. PROGRAM ELEMENT NO.
Research Area 1-017 and 1-034 PM, Task 7790
11. CONTRACT/GRANT NO.
None
12. SPONSORING AGENCY NAME AND ADDRESS
US EPA .Office of Research and Development, NERL and NHEERL, RTP, NC 27711
13. TYPE OF REPORT AND PERIOD COVERED
Book Chapter, 19992-1999
Institute of Experimental Medicine, Academy of Sciences, Prague, Czech Republic and
Institutes of Hygiene of Central Bohemia, Teplice and Usti, Czech Republic
14. SPONSORING AGENCY CODE
US EPA and US AID
15. SUPPLEMENTARY NOTES
Publication Title: Teplice Program, Editor: Radim Sram, Publisher: Czech Academy of Sciences
Proceedings of the Teplice Program Conference held in Prachatice, Czech Republic, October 3-5,2000
16. ABSTRACT
The use of biomarkers in the Teplice Program, provided a key tool to relate health outcomes to individual personal exposures and
to provide measures of confounding exposures. This research program on the health effects of air pollution studied a population
living in the heavily industrialized district of Teplice in Northern Bohemia and compared the exposure and health of this population
to that of a non-industrialized district, Prachatice, in Southern Bohemia, The studies included characterization of the
environmental and personal air pollution exposure, biomarkers, and studies on reproductive, respiratory, and neurobehavioral
effects. Biomarkers were measured in blood, urine, placenta, and sperm. The biomarkers included measures of exposure (e.g.,
urine metabolites and blood metals), dose (e.g., DNA adducts), DNA damage, genetic and cytogenetic effects, and susceptibility.
During winter temperature inversions, unusually high concentrations of a complex mixture of air pollutants were measured,
including fine particles, genotoxic organic compounds, and toxic trace elements. This population, however, was also exposed to
multiple pollutants via all pathways, and including pollutants resulting from environmental exposures, occupational exposures, and
personal habits (e.g., tobacco and alcohol use). Longitudinal and repeated measures used individuals as their own control to
examine the influence of environmental exposures as they changed over time and season. Chronic and seasonal exposures to
elevated air pollution in the Teplice District were shown to have serious adverse respiratory health consequences for children and
reproductive effects in adults. Elevated levels of air pollutants, even for short-term winter inversions resulted in measurable uptake,
metabolism, and excretion of polycyclic aromatic hydrocarbons, increased blood concentrations of toxic metals, and resulted in
DNA damage. Results of the exposure, biomarker, and health studies indicated that environmental exposure to a complex mixture
of air pollutants resulted in significant elevations in personal exposure, uptake, excretion of pollutants and DNA damage.
17.
KEY WORDS AND DOCUMENT ANALYSIS
a. DESCRIPTORS
b. IDENTIFIERS/OPEN ENDED TERMS
c. COSATI Field'Group
exposure, biomarkers, air pollution
particulate matter, polycyclic
aromatic hydrocarbons, trace
metals, urinary metabolites, DNA
adducts, fine particles,
environmental health
and exposure
18. DISTRIBUTION STATEMENT
Release unlimited
19. SECURITY CLASS (This Report)
21. NO. OF PAGES
20. SECURITY CLASS (This Page)
22. PRICE
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