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IJOMEH 2009;22(4) 383
O R I G I N A L P A P E R S
International Journal of Occupational Medicine and Environmental Health 2009;22(4):383 – 391
DOI 10.2478/v10001-009-0037-0
POLISH MOTHER AND CHILD COHORT STUDY —
DEFINING THE PROBLEM, THE AIM OF THE STUDY
AND METHODOLOGICAL ASSUMPTIONS
KINGA POLAŃSKA1, WOJCIECH HANKE1,2, JOLANTA GROMADZIŃSKA3, DANUTA LIGOCKA4,
EWA GULCZYŃSKA5, WOJCIECH SOBALA1, and WOJCIECH WĄSOWICZ3
1 Nofer Institute of Occupational Medicine, Łódź, Poland
Department of Environmental Epidemiology
2 Medical University, Łódź, Poland
Department of Informatics and Medical Statistics
3 Nofer Institute of Occupational Medicine, Łódź, Poland
Department of Toxicology and Carcinogenesis
4 Nofer Institute of Occupational Medicine, Łódź, Poland
Department of Chemical Hazards
5 Polish Mother’s Memorial Hospital — Research Institute
Department of Neonatology
Abstract
Objectives: Exposures during prenatal period have implications for pregnancy outcome as well as for children’s health,
morbidity and mortality. Prospective cohort study design allows for the identification of exposures that may influence preg-
nancy outcome and children’s health, verification of such exposures by biomarker measurements and notification of any
changes in exposure level. Materials and methods: Polish Mother and Child Cohort Study (REPRO_PL) is multicenter
prospective cohort study conducted in 8 different regions of Poland. The final cohort is intended to comprise 1300 mother-
child pairs to be recruited within 4-year period (2007–2011). The recruitment and all scheduled visits are conducted in
maternity units or clinics in the districts included in the study. The women are followed-up 3 times in pregnancy (once in
each trimester) and after delivery for the notification of pregnancy outcome. During each visit, detailed questionnaire and
biological samples are collected including saliva, urine, hair, maternal blood and cord blood. About 6 weeks postpartum,
breast milk from part of the women is collected. The study concentrates on the identification and evaluation of the effects
of prenatal environmental exposure on pregnancy outcome and children’s health. Specific research hypotheses refer to the
role of heavy metals, exposure to polycyclic aromatic hydrocarbons (PAHs) and environmental tobacco smoke (ETS) in
the aetiology of small-for-gestational-age (SGA) and preterm delivery (PD). The role of oxidative stress putative mecha-
nism and pregnant women nutritional status will be investigated. Based on questionnaire data, the impact of occupational
exposures and stressful situations will be evaluated. Results: The results of the study will become available within the next
few years and will help to determine levels of child prenatal exposure in several areas of Poland and its impact on course
and outcome of pregnancy.
Key words:
Birth cohort, Pregnancy, Prenatal exposure, Biological sample
This study was performed under the project “Polish cohort of pregnant women” supported by the Ministry of Science and Higher Education, Poland, from grant
no. PBZ-MEiN-/8/2//2006; contract no. K140/P01/2007/1.3.1.1.
Received: September 7, 2009. Accepted: October 13, 2009.
Address reprint request to K. Polańska, Department of Environmental Epidemiology, Nofer Institute of Occupational Medicine, św. Teresy 8, 91-348 Łódź, Poland
(e-mail: kinga@imp.lodz.pl).
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IJOMEH 2009;22(4)384
(MoBa) has comprised a cohort of 100 000 pregnant
women [1]. In the study the biological samples have been
collected from mother, father and child. The children are
intended to be followed until the age of 6. The study has
covered different factors, such as medication, nutrition, in-
fection and work exposure. Genetic factors and the inter-
action between genes and the environment has been also
studied. About 100 000 pregnant women are examined
early in pregnancy in the Danish National Birth Cohort
(DNBC) with the long-term follow-up of the offspring [2].
In that study, exposure assessment is done by interviewing
the women twice during pregnancy and when their chil-
dren are six and 18 months old. The biological bank has
been set up with blood taken from mother two times in
pregnancy and blood from the umbilical cord taken short-
ly after birth. In the French PELAGIE cohort study, the
association between fish and shellfish intake and length
of gestation, birthweight and the risk of preterm delivery
(PD), low birthweight (LBW) or small-for-gestational-age
(SGA) babies is analysed [4].
A special website (www.birthcohorts.net) has been cre-
ated to facilitate exchange of knowledge and collabora-
tion between researchers on already founded, established
and ongoing cohorts and research in the initial phase. The
existing birth cohorts are heterogeneous in design and fo-
cus but, for specific purposes, data from multiple cohorts
could be successfully pooled together. Advantages of this
approach include feasibility and low costs.
In Poland, the Kraków centre has continued cohort stud-
ies concentrating mostly on the assessment of impact of
exposure to air pollution and mercury on birth outcome
and children’s health [5,6].
The Polish Mother and Child Cohort is multicentre study
on different exposures. The aim of the study is to evaluate
the impact of exposure to different environmental factors
during pregnancy on pregnancy outcome and children’s
health. Specific research hypotheses refer to the role of
heavy metals, exposure to polycyclic aromatic hydrocar-
bons (PAHs) and environmental tobacco smoke (ETS) in
the aetiology of SGA and PD. It is also intended to explain
the role of oxidative stress and nutritional status of the
pregnant women. The impact of occupational exposures
INTRODUCTION
Epidemiological data indicate that disease aetiology has
to be evaluated with life-course perspective, starting from
prenatal period or, if possible, close to the time of con-
ception and followed until birth and later even until adult
life. Exposures during prenatal period have implications
for pregnancy outcome as well as for children’s health,
morbidity and mortality occurring later in life, including
asthma, allergy, delayed neurodevelopment, diabetes, car-
diovascular diseases, cancer and many other. The assess-
ment of factors influencing birth outcome and children’s
health should concentrate on genetic predisposition, envi-
ronmental exposure and social context.
Birth cohort studies that address prenatal outcomes have
been conducted all over the world for at least 20 years.
It is important to conduct such studies as new medicines
and preventive measures are developed on one hand and
new occupational and environmental hazards emerge on
the other, which may affect pregnancy outcome. Addition-
ally, new tools for detection of infections, environmental
exposures, and the role of genetic factors offer major new
research approaches and opportunities to explain the aeti-
ology of many reproductive failures.
Prospective cohort study design enables identification of
exposures that may influence pregnancy outcome and chil-
dren’s health, verification of such exposures by biomarker
measurements and notification of any changes in exposure
levels. In the studies in which exposure status is evaluated
from questionnaire data, recall bias can significantly influ-
ence the results, especially if evaluated retrospectively.
It is also important to note that for some exposures, such
as smoking or alcohol consumption, for which detrimen-
tal health impacts are well established, the exposure levels
may be underestimated.
Several birth cohorts have been established in Europe
in the last decades [1–4]. Some of these cohorts have ad-
dressed identification of exposures for which the conse-
quences become manifest before or shortly after birth,
while other have followed the children until adolescence
or even adulthood. For example, one of the biggest co-
hort study — Norwegian Mother and Child Cohort Study
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and organizational measures are implemented to protect
personal data against any unlawful form of processing. All
biological material stored and used for the work is identi-
fied by a code number only. Participants have the right to
have their data removed from the cohort at any time and
the right not to be subject to more interviews or sampling.
So far, only 1 out of about 700 women has asked to be
deleted from the cohort.
Power calculation /Sample size
We conducted power analysis to calculate the minimum
effect size that is likely to be detected in a study using
a given sample size. We have calculated power function
for assumed sample size (1300 pregnancies). As incidences
for LBW and PD are at a similar level (approximately 6%
in Poland) we have performed the calculations only once.
We have distinguished between dichotomous and contin-
uous exposure variables (Fig. 1). We have used function
bpower from Hmisc R package to calculate power for bi-
nary exposure [7]. For each combination of OR and expo-
sure prevalence we have constrained fraction of cases to be
equal to assumed in population. For continuous variables
we have assumed normal distribution of exposure. We have
conducted 1000 simulations for each effect size from 1
to 1.8 (with 0.01 step size) to calculate power. Then we
have fitted logistic curve to data points from simu lations.
The power of our study is equal to 80% for risk factors
with prevalence 0.05, 0.1, 0.3 when its effect sizes are,
respectively, 3, 2.5 and 2. In any case of exposure preva-
lence we have low power for effect size lower than 1.25.
and stressful situations will be evaluated from question-
naire data.
The results of the study will become available within the
next few years and will help to determine levels of child
prenatal exposure in several areas of Poland and their im-
pact on course and outcome of pregnancy.
MATERIALS AND METHODS
Study design and population
Polish Mother and Child Cohort Study (REPRO_PL) is
the prospective cohort study conducted in 8 different re-
gions of Poland. The study concentrates on the identifica-
tion and evaluation of the effects of prenatal environmen-
tal exposures on pregnancy outcome and children’s health.
For the reliable verification of such exposures, extensive
assessments are carried out in pregnant women included
in the study.
The final cohort is intended to comprise 1300 mother-child
pairs to be recruited within 4-year period (2007–2011).
The recruitment and all scheduled visits are conducted in
maternity units or clinics in the districts participating in
the study. The women are followed-up 3 times in preg-
nancy (once in each trimester) and after delivery for the
notification of pregnancy outcome. In future studies it is
planned to follow the child till the age of 2 years to deter-
mine long term effects of pre- and postnatal environmen-
tal exposures.
The study was approved by the Ethical Committee of
the Nofer Institute of Occupational Medicine, Łódź, Po-
land (Decision No. 7/2007). All study participants are in-
formed about the aims and procedures of the study and
are asked to sign an informed consent form. Data col-
lected during the study are confidential. All staff involved
in the collection, processing, and analysis of study data
are aware of the important responsibility to safeguard
the rights of study participants. Respondents are assured
that all identifying data, such as their name and address,
are not available to anyone outside the project team and
are not associated with their responses. All answers are
used only for research purposes and are not combined
with those of other participants. Appropriate technical Fig. 1. Power calculation.
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IJOMEH 2009;22(4)386
delivery, blood from the mother and cord blood is sam-
pled. After delivery, pregnancy outcome is recorded and
about 6 weeks postpartum breast milk is collected from
some of the women.
Questionnaire
All women participating in the study are interviewed three
times during pregnancy. Socio-demographic information,
including place of residence, age, marital status and level
of education is collected. Detailed information relating to
previous and current pregnancies (including the medica-
tion and vitamin intake, complications if any, ultrasound
measurements) is noted. The midwives also collect the in-
formation about active and passive smoking at different
stages of pregnancy. Assessment of various occupational
exposures is based on detailed questions, updated three
times during pregnancy. The questionnaire is concerned
with the time and type of work performed, way of perform-
ing it and specific exposures, if any. Food frequency ques-
tionnaire conducted at 20–24 weeks of pregnancy allows
for maternal diet assessment. At the same time additional
questionnaires are conduced including questionnaire for
subjective perception of work and family environment and
Cohen questionnaire.
Assessment of pregnant women exposure
All biological samples (saliva, urine, hair, blood, cord
blood and breast milk) are collected, processed and stored
in each study centres according to specified protocol. The
biological samples are transported to the coordinating
centre at Nofer Institute of Occupational Medicine, Łódź,
Poland where they are analysed. Details of exposure as-
sessment are presented in Table 1.
The air pollution exposure
There is a growing body of literature reporting associations
between ETS and atmospheric pollutants and reproductive
outcomes, in particular birth weight and gestational dura-
tion [8–11]. The main limitations of the existing literature
relating to the impact of such exposure on pregnancy out-
come include exposure misclassification and confounding
related to socio-economic status. Even less information is
In case of continuous exposures we have power above 80%
threshold for effect size equal to 40 %.
Non-differential misclassification will reduce some of the
power, and this needs to be taken into account. In any
case, the size provides new research opportunities for rare
exposures.
Inclusion criteria
We include into the study women between 8–12 weeks of
single pregnancy, not assisted with reproductive technol-
ogy, and not expected to be finished as spontaneous abor-
tion. All women with the serious chronic diseases specified
in study protocol such as diabetes, hypertension, nephrop-
athy, epilepsy and cancer are excluded from the study. The
same refers to suspicion of serious child malformations.
Follow-up of the women
All women who have agreed to participate in the study
are interviewed by obstetrician and/or midwife using de-
tailed questionnaire. During that visit, saliva and blood
sample is collected (Fig. 2). The second visit scheduled be-
tween 20 and 24 weeks of pregnancy includes all elements
from the firs visit and, additionally, collection of urine
sample. The third examination between 30–34 weeks of
pregnancy contains: third questionnaire and collection
of saliva, urine, blood and hair samples. At the time of
Fig. 2. Follow-up of the study population.
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Geographic Information System (GIS) techniques are
used to link the information about place of residence to
the data from local stations measuring the air pollution
levels. All available data about air quality are analysed
(including PM10, SO2, NOx, PM2,5).
Assessment of nutrition status, oxidative stress
Trace elements (Se, Zn, Cu) and bioactive substances,
including vitamins A, C, E, carotenoids, polyphenols, fla-
vonoids etc.), are necessary nutrients that should be de-
livered to the body with diet. The dietary intakes of the
microelements vary considerably in various geographical
regions due to varied amounts present in the soil and,
consequently, in food products (meat, dairy, cereals, fruits
and vegetables) and varied eating habits [12]. There is
statistically significant correlation between intake of mi-
croelements and their blood concentration in healthy hu-
mans [13].
accessible on long-term effects (e.g. children’s behavioural
development) of prenatal exposure.
The prospective study design enables reliable identifica-
tion of the exposure and recording of any changes in expo-
sure levels. The information about air pollution exposure
including traffic and tobacco smoke exposure is assessed
from questionnaire data, biological sample analysis and
data from air monitoring.
Smoking status and environmental tobacco smoke expo-
sure during pregnancy is verified by determinations of co-
tinine level in saliva collected three times in pregnancy.
Cotinine level in saliva sample is analysed using liquid
chromatography with tandem mass spectrometry (LC-MS/
MS ESI+).
Assessment of exposure to PAHs is based on measure-
ment of 1-hydroxypyrene in urine collected in 20–24 week
of pregnancy using high performance liquid chromatogra-
phy (HPLC).
Table 1. Details of exposure assessment
Exposure Biomarker Matrix Method
Sample Collection
(weeks of pregnancy/
delivery/after delivery)
Smoking/ETS Cotinine Saliva LC-MS/MS ESI+. 8–12, 20–24, 30–34
PAHs 1-hydroxypirene Urine HPLC 20–24
Heavy metals Pb, Cd Blood, cord blood GFAAS 20–24, delivery
Hg Hair CVAAS 30–34
POPs PCDD/PCDF/PCB Breast milk HRGC/HRMS 3–8 weeks after delivery
Nutrition Zn, Cu Blood, cord blood AAS 8–12, 20–24, 30–34,
delivery
Se Blood, cord blood GFAAS 8–12, 20–24, 30–34,
delivery
Mg Blood, cord blood AAS 8–12, 30–34, delivery
Dietary antioxidants (Vitamins A, E) Blood, cord blood HPLC 20–24
Antioxidant enzyme activities
(GPx, SOD, Cp)
Blood, cord blood Spectrophotometry 8–12, 20–24, 30–34,
delivery
TBARS Blood, cord blood Spectrofluorometry 8–12, 20–24, 30–34,
delivery
Genetic polymorphism GPx1, GPx4 Blood —buffy coat RT PCR 8–12
ETS — environmental tobacco smoke exposure, LC-MS/MS ESI+ — liquid chromatography with tandem mass spectrometry, PAHs — polycyclic
aromatic hydrocarbons, HPLC — high performance liquid chromatography, GFAAS — graphite furnace atomic absorption spectrometry,
CVAAS — cold vapour atomic absorption spectrophotometry, POPs — persistent organic pollutants, HRGC/HRMS — high resolution gas chroma-
tography/high resolution mass spectrometry, AAS — atomic absorption spectrometry, TBARS — tiobarbituric acid reactive substances, RT PCR —
real-time polymerase chain reaction.
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