Human Reproduction Update, Vol.14, No.1 pp. 59–72, 2008 doi:10.

1093/humupd/dmm025
Advance Access publication December 10, 2007

Impact of endocrine disruptor chemicals in gynaecology

D. Caserta1,3, L. Maranghi1, A. Mantovani2, R. Marci1, F. Maranghi2 and M. Moscarini1

1
Institute of Gynecology, Perinatology and Child Health, Sant’Andrea Hospital, University of Rome La Sapienza, Via di Grottarossa
1035, 00189 Rome, Italy; 2Department of Food Safety and Veterinary Public Health, Istituto Superiore di Sanità, Rome, Italy
3
Correspondence address. Tel: þ39 6 33775696; Fax: þ39 6 3350611; E-mail: donatella.caserta@libero.it

The potential hazardous effects that estrogen- and androgen-like chemicals may have both on wildlife and human
health have attracted much attention from the scientific community. Endocrine disruptors (EDCs) are chemicals

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that have the capacity to interfere with normal signalling systems. EDCs may mimic, block or modulate the synthesis,
release, transport, metabolism and binding or elimination of natural hormones. Even though potential EDCs may be
present in the environment at only very low levels, they may still cause harmful effects, especially when several differ-
ent compounds act on one target. EDCs include persistent pollutants, agrochemicals and widespread industrial
compounds. Not all EDCs are man-made compounds; many plants produce substances (phytoestrogens) that can
have different endocrine effects either adverse or beneficial in certain circumstances. Natural substances such as
sex hormones from urban or farm wastes can become concentrated in industrial, agricultural and urban areas;
thus, such wastes may be considered potential ‘EDCs’ for humans and/or wildlife. Much attention has focussed on
changing trends in male reproductive parameters in relation to EDC exposure; however, studies on the female repro-
ductive system have been less comprehensive. We have focussed this article on four major aspects of female reproduc-
tive health: fertility and fecundability, endometriosis, precocious puberty and breast and endometrial cancer.

Keywords: environmental effects; endometriosis; female infertility; estrogen

Introduction implicated in the development of some gynaecologic pathologies

are listed in Table 1.
Both in humans and in animals, endocrine signalling is involved in
This review article summarizes the available literature on
reproduction and embryo development, growth and maturation,
adverse outcomes in female human reproductive health from sup-
energy production, use and energy storage, electrolyte balance
posed exposure to EDCs, or from which an endocrine mechanism
and maintenance and behaviour. Hormones trigger such complex
is plausible or has been proven.
functions by interacting with their receptors that are present, at a
nuclear and/or cellular level, in various organs and tissues as
part of a complex biological feedback system. Any disruption of What are EDCs?
this balance can cause impairment in the physiological status of An endocrine disruptor is an exogenous substance or mixture that
the whole organism, especially during the more susceptible devel- alters function(s) of the endocrine system and consequently causes
opmental stages. If the regulatory role of the endocrine system is adverse health effects in an intact organism, or its progeny, or
impaired, abnormal function and/or development of the reproduc- (sub)population (WHO, 2002). The homeostasis of sex steroids
tive, the nervous and the immune systems may occur. It has been and the thyroid are the main targets of EDC effects; hence, repro-
recently postulated that predisposition for certain types of tumours ductive health, considered as a continuum from gamete production
is caused by an altered development during prenatal growth and fertilization through to intrauterine and post-natal develop-
(intrauterine) and in the first years of life (Sharpe and Sakkebaek, ment of progeny, is recognized as being especially vulnerable to
1993). It is noteworthy that the available data overall support the endocrine disruption (Mantovani, 2002). EDCs are widespread
public health concerns for endocrine disrupting chemicals (EDCs). in food chains and in the environment and include persistent
Several substances, including EDC, with share the ability of organic pollutants (POPs) such as the insecticide dichloro-
interfering with the female reproductive system and possibly diphenyl-trichloroethane (DDT) and its metabolites, the industrial
# The Author 2007. Published by Oxford University Press on behalf of the European Society of Human Reproduction and Embryology. All rights reserved.
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Caserta et al.

Table 1: Main classes of EDCs present in food and in environment with major relevance for female reproductive system

Chemical(s) Pathways of exposures Mechanisms of action

POPs
Polychlorobiphenyls (PCB)
Dioxins and ‘dioxin-like’ PCBs
DDT and metabolites
Substances used in agricultural and farm animal production
Organochlorine insecticides (e.g.
Lindane)
Food chain (fat-rich food, e.g. milk and derivatives, fatty
fish, etc.), living environment
Food chain (fat-rich food, e.g. milk and derivatives, fatty
fish, etc.), living environment
Food chain (fat-rich food, e.g. milk and derivatives, fatty
fish, etc.), living environment and workplaces (in
developing countries)
Food chain (fat-rich food, e.g. milk and derivatives, fatty
fish, etc.), living environment, workplaces (mainly in
developing countries)
Alteration steroid hormone metabolism/transport,
ability to bind with the thyroxin transport protein,
transthyretin (TTR), interaction with thyroid
hormone receptors, neuroendocrine effects
Aril hydrocarbon Receptor interaction leading to
altered steroid hormone metabolism and
neuroendocrine effects including on thyroid
Mainly estrogenic activity but also interaction with
Homeostasis of steroid hormones (estrogenic and/
or anti-androgenic effects, interaction with PR)

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Triazoles, Imidazoles
Triazines
ETU (metabolite of ethylene
bisdithiocarbammates, e.g. Maneb),
Benzimidazoles
Industrial products and daily-use products
Nonyl-phenols and octyl-phenols
Bisphenol A
Several phthalates (di-2-hexyl-ethyl-,
di-n-butyl-, etc.)
Polybrominated flame retardants
Organotins
Perfluorooctane sulphonate
Parabens
UV-screen (benzophenone 2,
4-methylbenzylidene camphor, etc.)
Cadmium
Phytoestrogens
Isoflavones, lignans, etc
Food chain (agricultural and zootechnical fungicides), Inhibition of steroid hormone biosynthesis
living environment and workplaces (agricultural areas)
Food chain (herbicides), living environment and Effects on HHG axis
workplaces (agricultural areas)
Food chain (agricultural and zootechnical fungicides), Thyreostatic effects
living environment and workplaces (agricultural areas)
Detergent by-products: food chain (seafood) and consumer Estrogen agonists—ER alpha
products
Food chain (e.g. plastics in contact with food), consumer Estrogen agonist—ER alpha
products (e.g. dental sealant, plastic additive, etc.)
Food chain (e.g. plastics in contact with food), consumer Agonists of PXR, effects on steroid hormone
products (e.g. PVC, deodorants, adhesives, etc.) biosynthesis
Food chain (fat-rich food, e.g. milk and derivatives, fatty Interaction with PXR leading to altered steroid and
fish, etc.), living environment, workplaces, consumer thyroid hormone homeostasis
products (e.g. electronic devices, etc.)
Food chain (seafood), consumer products (e.g. antifouling Aromatase inhibition
agents)
Food chain (bioconcentration in animal tissues), consumer Alteration HHG axis
products (e.g. plastics, carpets, materials, etc.)
Main cosmetic, toiletries and pharmaceutical preservatives Estrogen agonist—ER alpha and beta
Mixture for protection against UV radiation Estrogen agonist—ER alpha
Food chain (e.g. refined food as flour, rice, sugar; Estrogen agonist—ER alpha
seafood), cigarette smoking
Food chain (e.g. vegetables, soy-based food), consumer SERMs, high affinity for ER beta
products (e.g. cosmetics)

POPs, persistent organic pollutants; PR, progesteron receptor; ETU, ethylenethiourea; PXR, pregnane X receptor; HHG axis, hypothalamo-hypophysis-gonadal
axis; ER, estrogen receptor; SERM, selective ER modulator.

by-product dioxins and the industrial compounds polychlorinated
biphenyls (PCB), several agrochemicals, pesticides and biocides
(e.g. chlorinated insecticides, organotins, imidazoles and triazoles)
and other industrial compounds (several phenol compounds such
as bisphenol A) (Mantovani et al., 1999). More recently, attention
has been focussed on the endocrine effects of substances (e.g.
parabens, component of UV-screen, phthalates) widely diffused
also in cosmetics and toiletries (Oishi, 2002; Kunz and Fent,
2006), as well as of metals, e.g. arsenic compounds (Tseng
et al., 2003). Substances other than typical food and/or environ-
mental contaminants may also be considered as EDCs, such as
drugs, anabolic agents and especially phytoestrogens: this class
of chemicals includes isoflavones, lignans, etc., which are
present in some food items such as soy, and in cosmetics with
active ingredients of vegetal origin. A good dietary intake of
phytoestrogens might act as protective factor against several
cancers (e.g. breast, prostate) and post-menopausal diseases (e.g.
osteoporosis); however, there is some concern regarding the
exposure to high doses during pregnancy or early infancy, e.g.
through the use of dietary integrators or soy-based milk (Skibola
and Smith, 2000; Stark et al., 2003), due to the high intake of
hormone-mimic substances during the critical period of infant
and adolescent development. Many EDCs are known to act as ago-
nists of estrogen receptors (ER), e.g. bisphenol A and alkyl-
phenols, or to antagonize androgen receptor (AR) such the
dicarboximide fungicides; progesterone receptors (PR) are also a

60

potential target for many chlorinated EDC, such as DDT and
derivatives (Scippo et al., 2004). Compounds that act as
hormone triggers should also be taken into account; dioxins and
the dioxin-like chemicals being the best-known examples. They
bind to the cytoplasmic aryl hydrocarbon receptor (AhR), which
in turn cross-talks with the steroid nuclear receptors to initiate
entirely new responses; these may vary with the status (activated
or not) of the ER and AR. Other chemicals such as some phthalate
plasticizers or the polybrominated diphenyl ethers (PBDEs) used
as flame retardants are more likely to interact with less specific
orphan nuclear receptors, like pregnane-X or androstane, which
act as ‘sensors’ to regulate the activity of ligand-specific receptors
(Sanders et al., 2005; Wyde et al., 2005). Atrazine and related her-
bicides instead can impair the hypothalamus-hypophysis-gonads
axis (McMullin et al., 2004). Other EDCs can interfere with
hormone synthesis and transport, examples are the azole anti-
fungals agents, used in agriculture and animal production, that
inhibit different steps of steroid synthesis (Zarn et al., 2003) and
the hydroxylated PCB metabolites that selectively interact with
estrogen sulphotransferase, thus increasing estradiol (E2) bioavail-
ability in target tissues (Kester et al., 2000). Such distinctions
however should not be considered too rigidly; recent evidence
indicates that the type of effects induced by several EDCs may
vary with sex and age of the target organism (Pryor et al., 2000;
Hallgren and Darnerud, 2002). The adverse effects of xenobiotics
on female reproductive health, namely fertility and pregnancy
maintenance, have already been identified in a number of studies
(Hruska et al., 2000). Epidemiological evidence points to known
lifestyle factors (cigarette smoke, high alcohol consumption and
obesity) and intensive occupational exposures to heat, radiation
or solvents as risk factors for reproductive dysfunction in
women, although with variable incidence (Eggert et al., 2004;
Hassan and Killick, 2004; Kumar, 2004).
The initial interest for the effects of EDC on human health was
related to the possible interference with male reproductive dis-
orders (Sharpe and Sakkebaek, 1993). However, there is
growing evidence from toxicological as well as clinical studies
that the female reproductive system is a potential EDC target too.
EDC risk assessment
Hazard identification—mechanisms of EDC actions
Experimental data on animal models indicate that early prenatal
and/or perinatal exposure to EDCs can lead to long-term effects
on reproduction and development which can become evident
later, even at sexual maturity and/or at adulthood. The identifi-
cation and characterization of this ‘early exposure—late effect’
pattern of EDCs still represents a challenge for scientists and
risk assessors. Accordingly, there is an ongoing international
effort to develop, validate and/or update the current toxicological
testing approaches. To screen substance for their potential effects
on endocrine homeostasis, the 28-day oral toxicity study [Organ-
isation for Economic Co-operation and Development (OECD)
Guideline 407], i.e. the common subacute toxicity test widely
applied to all chemical testing strategies, has been enhanced to
detect potential EDC through additional parameters, including:
serum hormone concentrations, estrus cycle features and weights
of endocrine-related organs (ovaries, uterus with content, etc.)
(Kunimatsu et al., 2004). Although it is a comprehensive test to
Impact of endocrine disruptor chemicals
identify compounds with endocrine activities, the assay retains
possible limitations for the full characterization of effects and
dose-response relationships, including the relatively short duration
and the testing of adult animals.
The two-generation reproductive toxicity study (OECD Guide-
line, 416) represents the most effective test to evaluate alterations
on the endocrine homeostasis during the entire developmental and
reproductive period. Since human exposure to EDCs may span a
lifetime, multigenerational studies are usually chosen so that the
test substance is administered continuously, without interruption,
to parental (P) and subsequent offspring generations (F1, F2,
etc.). This protocol will provide information about effects on
male and female reproductive performances, potency and fertility,
pregnancy outcomes, maternal lactation and offspring care, pre-
natal and post-natal survival, growth and development of off-
spring, as well as their reproductive capacity.
The above-mentioned experimental protocol requires a large
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number of animals and is also costly, laborious and time consum-
ing. Nevertheless, at the moment it represents the only reliable tool
for the study of such a complex system and/or function, e.g. repro-
duction and to perform hazard identification for EDCs.
There are a number of mechanisms whereby EDCs can modu-
late endocrine systems and potentially cause adverse effects on
human health (Fig. 1). The generally accepted paradigm for
Figure 1: Schematic diagram depicting several key steps in steroid hormone
action that may be sensitive to disruption by environmental chemicals: syn-
thesis and secretion of steroids hormones [EA] from the gonadal cells;
binding affinity of the EA for the SHBG protein; diffusion into the cell; diffu-
sion into the perinuclear region; binding the receptor; conformational change of
the receptor (R) which forms homodimers; formation of a transcriptional
complex from homodimers; which binds to specific sequences on the DNA
of hormone-dependent genes, known as HRE; transportation of mRNA into
the cytoplasm; synthesis of proteins; alteration of liver function, either increa-
sing or decreasing metabolism of the hormone
*Possible key stepsin steroid hormone action sensitive to EDCs
61
Caserta et al.
receptor-mediated responses includes hormone binding to its
receptor at the cell surface, cytoplasm or nucleus, followed by a
complex series of events that lead to changes in gene expression
(Birnbaum, 1994). The main nuclear receptors involved in EDC
action are: ER a and b, AR, thyroid receptors, AhR, glucocorti-
coid receptors (GR), as target for arsenic (Bodwell et al., 2004)
and pregnane-X-R, which appears to be specific target for some
phthalates (Hurst and Waxman, 2004). More recently, attention
has been focussed on PR that appear to be more sensitive than
ER-a and a target for many POPs (Villa et al., 2004). Thus, a
panel of in vitro assays for binding/transactivation of nuclear
receptors may be highly relevant for screening and identification
of potential EDCs. Unfortunately, this is likely to be insufficient,
as several EDCs do not specifically interact with nuclear receptors.
Other relevant mechanisms include inhibition of hormone syn-
thesis, transport, or metabolism and activation of receptor through
receptor phosphorylation or the release of cellular complexes
necessary for hormone action. In the case of hormone synthesis,
considerable research has been conducted on the aromatase inhibi-
tors; in fact they can prevent the conversion of androgens to estro-
gens through a cytochrome P450 system, which is highly
conserved among the species. Several azole fungicides have
been shown to cause aromatase inhibition as well as some wide-
spread organotins (tri-butyl tin and tri-phenyl tin) (Matsui et al.,
2005). In addition, there is a growing awareness that multiple
receptor systems act in concert to regulate biological functions.
For example, ‘cross-talk’ between the ERs and growth factor
receptor appears necessary for estrogen signalling in mammary
cells to divide or differentiate. These events are critical for
explaining several risk factors for breast cancer, such as age at
menarche, age at menopause or effects on the number of
pregnancies.
Other example of a multi-factored EDC-mediated effect is the
hypothalamo-hypophysis-gonadal axis (HHG). Atrazine, the well-
known broadleaf herbicide, despite its limited solubility, is com-
monly found in the ground and in surface water together with its
biotransformation by-products, creating concern for exposure to
the general population. Atrazine exerts pleiotropic reproductive
effects in rodents, which comprise alteration of estrous cycles,
delay of the onset of puberty and mammary gland development
in prenatally and/or post-natally exposed females, increased pre
and/or post-implantation losses, induction of mammary tumours
as well as acceleration of reproductive ageing (Friedmann,
2002; Rayner et al., 2004). All these effects are thought to be
related to a more direct alteration of HHG axis, in particular to
the LH suppression caused by inhibition of GnRH and prolactin
(Rayner et al., 2005).
Thus, a sensitive, time- and cost– effective evaluation of the
thousands of chemicals in use that have not yet been tested for
their endocrine disrupting potential requires the development of
an integrated battery of assays encompassing different mecha-
nisms and targets (Bremer et al., 2005).
Dose– response relationships
Dose – response relationships is perhaps the most controversial
issue regarding EDCs. One of the reasons is that EDCs often act
by mimicking or antagonizing the actions of naturally occurring
hormones. These hormones are already present at physiologically
functional concentrations, so the dose – response considerations
62
for EDCs are often different than for other chemicals acting
throughout different pathways. In particular, some estrogenic
compounds can induce an inverted-U dose –response curve, result-
ing from low-dose stimulation of response; this kind of behaviour
challenges current methods of risk assessment. Several estrogenic
compounds, such as bisphenol-A, octylphenol can induce effects
at doses lower than those inducing general toxicity; these
adverse stimulatory inputs divert energy needed for other pro-
cesses resulting in reduced and/or altered health performances.
The current threshold models used for the assessment of low
doses of chemicals showing this shape of dose – response curve
tend to underestimate the risk and deserve more attention
(Weltje et al., 2005).
A common dose –response for all endocrine disruption mechan-
isms should not be expected. This conclusion is based on the
knowledge that chemicals categorized as endocrine disruptors
have shown many different mechanisms of action. These activities
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include estrogenic, antiestrogenic, antiandrogenic, growth factor
modulation, cytokine and thyroid modulation, modulation of
hormone metabolism. Besides, epidemiological documentation
of endocrine disruption is complicated when exposures are
mixed. In fact, even if the endocrine activity (estrogenic and
androgenic, etc.) and the mechanisms of all compounds were
known, the risk of multiple exposures can not be assessed from
a single-chemical approach as used in traditional toxicology.
Although mixtures of compounds can have effects, it may not be
possible to attribute causation to a single chemical. Furthermore,
for EDCs, cumulative low-dose insult can be more toxic than a
single high-dose exposure; this tends to modify the hypothesis
of classical toxicology. Two possible approaches have been pro-
posed to study mixtures: the investigation of mixtures without spe-
cifying the individual components; and the study of the individual
compounds with a focus on possible interactions in the context of
dose to the receptor (Koppe et al., 2006). Thus, analysis of mix-
tures allows evaluation of combined effects of chemicals each
present at low concentrations (Rasmussen et al., 2003).
Exposure assessment
Several chemicals in the environment (e.g. pesticides, industrial
chemicals and natural products) have shown to be hormonally
active; they can be detected and measured in wildlife, in environ-
mental samples as well as in the human population. POPs are per-
sistent in the environment; they are lipophilic and they can
bioaccumulate in fat-rich tissues, such as milk, dairy products
and fatty fish (Jacobs et al., 2002). Breastfeeding provides a sig-
nificant source of exposure to POPs early in human life, the
effects of which are not completely understood yet; nevertheless,
despite of the possibility of harm from environmental contami-
nants in breast milk, breastfeeding is still recommended as the
best infant feeding method (Nickerson, 2006). Other compounds
may be less persistent in food and environment but can be more
toxic, above all if the exposure falls in a critical period of the life
cycle, e.g. development or maturation. Knowledge about the mag-
nitude of human or wildlife exposure is often limited. Neverthe-
less, the presence in the food chain of environmental chemicals
other than POPs may be more important than it was assumed a
decade ago. Examples include: the presence of nonylphenols,
detergent by-products with estrogenic properties, in seafood
(Ferrara et al., 2001); the POP-like behaviour of widespread

industrial chemicals such as PBDE (Schecter et al., 2005) and per-
fluorooctane compounds (Olsen et al., 2005). A major issue is the
assessment of real exposure, i.e. internal exposure levels through
biomonitoring. Information is available only for some compound
groups (e.g. pesticides) in workplaces (Aprea et al., 2002).
Recently, increasing attention is paid towards bio-monitoring of
the general population not only for metals, POPs and POP-like
compounds but also for different EDCs. For example, pilot
studies conducted in Italy recorded high levels of phthalates in
the umbilical cord blood of neonates and in the serum of women
with endometriosis (Cobellis et al., 2003; Latini et al., 2004),
highlighting that the general population is currently exposed to
low-doses of EDCs. Similarly, US studies on urinary metabolites
of phthalates, confirm the widespread presence of these substances
in the environment (Calafat and McKee, 2006).
The definition of appropriate biomarkers of exposure for moni-
toring is also critical. In fact, for some EDCs such as phthalate acid
esters, alkylphenols, ethylene bis-dithiocarbammate fungicides,
PCBs, PBDEs, phytoestrogens, it is important to evaluate the rela-
tive exposure and toxicity of the main metabolites which represent
the real active compounds at the target site (Elsby et al., 2001).
Exposure assessment at a community level is a complex issue
that involves usage patterns of the different compounds and how
they are transported and metabolized in different environmental
compartments (water, sediment and biota). Exposure often needs
to be considered specifically for vulnerable groups, such as com-
munities with special dietary habits (e.g. fishing communities)
(Yeats et al., 1999); lifestyles may modulate intake of contami-
nants, e.g. smokers have consistently higher levels of cadmium,
a heavy metal which is included among potential EDC (Castelli
et al., 2005). Children deserve special consideration as they
have relatively greater food and water intakes, breathing volume
and contact with soil as compared to adults, leading to a poten-
tially higher uptake of chemicals per kilogram of body weight
(Schwenk et al., 2003).
Risk characterization—factors related to differential
vulnerability to EDCs
It is well known that differential responsiveness to EDCs has been
observed among different species as well as inside a general popu-
lation. The biologic and molecular mechanisms underlying this
specificity are quite diverse.
Determinants of species-specificity include differences in
receptor binding, gene transcription patterns of gene expression
and cellular responses to endocrine-active compounds (Gray
et al., 2004). Inside general populations, differences in responsive-
ness may be determined at the level of genetic polymorphisms in
hormone-metabolizing enzymes, hormone receptors and those
genes that are activated by these receptors (Li et al., 2005). Our
rapidly growing knowledge—emerging from the human genome
project—will enable the design of rational studies on the impact
of EDC exposures on hormonally sensitive endpoints in groups
that may be genetically predisposed. Thus, it is important to iden-
tify genetic factors influencing the individual and/or population’s
susceptibility to endocrine disorders and to set appropriate
biomarkers (Masse et al., 2002). Differences in the response to
EDCs rely also on the life cycle phase of exposure. Children,
e.g. show a specific vulnerability and sensitivity linked to the
immaturity of organ/tissues as well as metabolic systems
Impact of endocrine disruptor chemicals
(LaRonda et al., 2004). Extrinsic factors such as diet and lifestyles
can also impact individual susceptibility to endocrine-active agents.
Vulnerability of different groups in the population will be affected
by lifestyle factors, in particular diet: several experimental studies
indicate that the effects of toxicants may be modulated by the
intake of, e.g. phytoestrogens (You et al., 2002) and antioxidants
(Kocdor et al., 2005; Muthuvel et al., 2006).
Therefore, more attention to the nutrient-toxicant interactions
should be given in epidemiological studies. The use of new
approaches in molecular toxicology and epidemiology as well as
more targeted experimental protocols have the potential to yield
additional valuable information to elucidate the role of these
mechanistic determinants of specificity at low-dose exposures to
potential EDCs and to improve risk evaluation for the adverse
health effects of EDCs.
EDCs and women’s reproductive health
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Fertility and fecundity
Fertility and fecundity have shown a progressive decrease in the
last decades (Hassan and Killich, 2004). Occupational exposure
is often cited as a risk factor for female fertility, as well as for
early pregnancy loss and pre-term delivery. Pesticides represent
a relevant example: direct exposure through pesticide handling
may be included among specific risk factors for reproductive
health in the rural environment. In one case –control study, in
which 281 women with a diagnosis of infertility were compared
with 216 post-partum women, those women with a history of
working in the agricultural industry showed an elevated risk of
infertility (Fuortes et al., 1997). de Cock et al. (1994) found that
reduced fecundability ratio and longer time-to-pregnancy are
associated with application of pesticides, in particular when
older spraying techniques were used. Greenlee et al. (2003) ident-
ified an association between adverse reproductive outcomes in
women and the practice of mixing and applying herbicides as
well as the use of fungicides. Greenhouse work is a peculiar
type of agricultural work that implies a continuous exposure to
pesticides. In a large Danish study on time-to-pregnancy among
female workers in flower greenhouses, the overall fecundability
rate did not differ between workers and referents. However,
certain factors, possibly associated with a specifically increased
exposure to chemicals, are consistently associated with a signifi-
cant 20– 30% reduction of fecundability, i.e. handling cultures
many hours per week, spraying of pesticides and the lack of
glove use (Abell et al., 2000). A major problem in this sort of
study is the very general consideration of pesticide classes (e.g.
‘fungicides’ and ‘herbicides’), whereas these include highly
diverse chemical groups, with only a few being EDC or reproduc-
tive toxicants: a detailed appraisal of compounds specifically
related to adverse reproductive outcome is indeed important for
prevention and risk communication strategies. An attempt to
identify exposures to specific pesticides has been represented by
the Ontario Farm Family Health Study. In a retrospective study
on 2012 farm couples, no strong or consistent pattern of associ-
ation between exposure to various classes of pesticides and
time-to-pregnancy was observed (Curtis et al., 1999). Data from
this large study also suggested that female reproductive dysfunc-
tions (e.g. reduced fecundability and increased risk of miscarriage)
may be associated to exposure to pesticides of the male partner.
63
Caserta et al.
In fact, miscarriage risk increased with reported use of several
compounds, including thiocarbamates, atrazine and phenoxy
herbicides, especially when more compounds were used at once
and protective equipment was not used (Savitz et al., 1997;
Arbuckle et al., 1999). The possible role of male partner exposure
was supported by an Italian retrospective studies that showed
significantly increased risks of conception delay and spontaneous
abortion among the spouses of high-exposure workers (green-
house, applicators), even after adjusting for possible confounders
such as age, education, smoking habits, etc., the risk increased
with reported exposure to some pesticide groups only, including
potential EDC such as chlorinated insecticides and triazines
(Petrelli et al., 2000, 2003).
Regarding the general population exposed through food or the
living environment, a few studies relate the possible EDC
exposure to markers of impaired reproduction. The US Great
Lakes are considered a problem area concerning long-term pol-
lution to persistent EDCs, mostly PCB.
In the New York State Angler Cohort Study, lifetime exposure to
PCBs, estimated taking into account the fish consumption from
contaminated Great Lakes, was correlated to changes in fecund-
ability (Buck et al., 1999). Maternal consumption of fish for 3 –6
years was associated with reduced fecundability [odds ratios
(OR), 0.75; 95% confidence interval (CI), 0.59–0.91). This effect
was of the same magnitude in those with .7 years of fish consump-
tion, however, it did not reach statistical significance (OR, 0.75;
95% CI, 0.51–1.07). Studying a Michigan cohort (Courval et al.,
1999), increasing potential lifetime exposure to PCBs and
mercury, estimated by the numbers of sport fish meals consumed,
was associated with an increased time-to-pregnancy related to
increased paternal consumption; the association reached statistical
significance only in the group with highest fish consumption
(adjusted ORs were 1.4, 1.8 and 2.8 for 1– 114, 115–270 and
271–1127 annual fish meals consumed, respectively).
Studies that have associated adverse reproductive effects
together with biomarkers of exposure are of great interest. A posi-
tive association between the body burden of persistent chlorinated
EDC (PCB, DDT and metabolites, other chlorinated insecticides)
and gynaecological problems, especially recurrent miscarriages,
was observed in case –control studies carried out in Germany
(Gerhard et al., 1998) and USA (Korrick et al., 2001). In a
limited pilot study, maternal exposure to the estrogen-like bisphe-
nol A has also been associated with recurrent miscarriages
(Sugiura-Ogasawara et al., 2005); more studies are required to
support or refute this interesting finding. Nevertheless, conflicting
observations do exist in regards to the role of EDC exposure: a
study on Japanese patients with a history of recurrent miscarriage
has not found an association with serum levels of industrial pro-
ducts such as PCBs, hexachlorobenzene (HCB) or DDE
(Sugiura-Ogasawara et al., 2003). Thus, possible mechanisms
related to different vulnerability deserve more attention. In this
respect the possible influence of dietary phytoestrogens should
also be considered. Several studies have shown prolongation of
the menstrual cycle in healthy premenopausal women with a
daily intake of 45 mg of isoflavones from soy protein as possibly
attributable to a prolongation of the follicular phase for the sup-
pression of the normal midcycle surge in FSH and LH (Cassidy
et al., 1994; Lu et al., 2000). Nevertheless, in a study on flaxseed
ingestion, the luteal phase was reported to be prolonged (Phipps
64
et al., 1993). Since flaxseed is especially rich of lignans, the pos-
sibility that different phytoestrogens do exert diverse actions
cannot be ruled out.
Overall, apart from evidence from ecological studies, data on
the actual exposure to the EDCs impairing female fertility are
limited. The isolation of persistent organochlorine chemicals
from ovarian follicular fluid of women undergoing IVF at the
moment could represent a useful tool to obtain information
using the available reproductive technology techniques (Jarrell
et al., 1993a,b). Isolation of such chemicals at a critical period
of oocyte development may provide an important biomarker of
exposure.
Endometriosis
Endometriosis is an estrogen-dependent disease characterized by
the presence of endometrial glands and stroma outside the
uterine cavity. It is a common gynaecological disorder as well
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as a major cause of infertility (Chedid et al., 1995) affecting
~14% of women of all reproductive ages (Vercillini et al.,
1995). At present, it is generally recognized that there is no one
single theory to identify and explain all aspects of that multi-
factored clinical syndrome. Although there is a clear association
with estrogens, it is accepted that the disease is not specifically
caused by estrogens but is stimulated by them (Guarnaccia and
Olive, 1998).
Previous work in non-human primates has shown that exposure
to the dioxin 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD, the
‘Seveso’ dioxin) is associated with an increased prevalence and
severity of endometriosis. Rodent studies support the possible
role of environmental contaminants in the pathophysiology of
endometriosis, although a convincing mechanistic hypothesis
has yet to be advanced (Rier et al., 2003). In cynomologous
monkeys treated with TCDD at dose levels of 0, 1, 5 and 25 ng/kg
body weight/day for 12 months, the growth of surgically implanted
endometrial fragments was increased at the two higher dose levels
circulating gonadal steroid levels and menstrual cycle character-
istics were unchanged (Yang et al., 2000).
Serum levels of TCDD and dioxin-like PCB were analysed in
13 years monkeys after termination of a 4-year study with
dietary exposure to 0, 5 and 25 ng/kg diet. Elevated serum
levels of total dioxin-like compounds correlated strongly with
endometriosis. (Rier and Foster, 2001). These findings may be
relevant to humans since the serum levels recorded in
TCDD-exposed animals were similar to concentrations reported
in serum, milk and tissues from the general population. Interest-
ingly, TCDD-exposed rhesus monkeys with endometriosis exhibi-
ted long-term alterations in systemic immunity, such as enhanced
tumour necrosis factor-alpha secretion by blood monocytes (Rier
et al., 2001).
A relationship between endometriosis and exposure to dioxin-
like PCBs (Gerhard and Runnebaum, 1992) and dioxins
(Koninckx, 1999) has been proposed. A positive association
between endometriosis and dioxin exposure was reported in a
case –control study in which 44 women with endometriosis were
compared with 35 age-matched controls with tubal infertility
(Mayani et al., 1997). Significantly more women with endometrio-
sis [8 (18%) versus 1 woman (3%) of the control group] had
detectable dioxin levels in their serum (P ¼ 0.04); there was no
relationship between severity of endometriosis and concentration

of dioxin. In another case –control study, no association between
plasma organochlorine concentrations and endometriosis could
be found in 86 women with endometriosis compared with 70 con-
trols, matched for the indication of laparoscopy (Lebel et al.,
1998). The highly exposed women in Seveso also underwent an
evaluation for endometriosis in a case –control study, which
showed no significant association between dioxin levels and the
presence or amount of endometriosis, although a trend towards
increased risk was apparent in the group with the highest body
burden (.100 ng/l serum); the authors cautioned that disease
misclassification in a population-based study may have led to an
underestimate of the true risk of endometriosis (Eskenazi et al.,
2002).
The plasma concentrations of ubiquitous environmental con-
taminants such as phthlates are associated with endometriosis in
an Italian study that for the first time suggests the role of phthalate
esters in the pathogenesis of the disease (Cobellis et al., 2003).
There are no direct data regarding the impact of phytoestrogens,
such as isoflavones or flavanones, on ectopic human endometrium,
although it could be anticipated that they would cause effects,
given that soy isoflavones inhibit E2-mediated endometrial pro-
liferation in macaque monkeys. Hence, the human data at
present have failed to provide compelling evidence for or
against an association of EDCs and endometriosis.
Precocious puberty
Environmental chemicals have also been suggested as potential
causative factors in the temporal modification in the age of
puberty onset. Population-based studies showing a reduction in
the median age of puberty demonstrate the need for investigations
of the possible causes of this trend (Delemarre-van de Waal,
2005).
A hypothesized external agent that induces central precocious
puberty would act through the early initiation of the pulsatility
of gonadotropin-releasing hormone from the hypothalamus,
thereby inducing the cascade of hormonal events that result in pub-
ertal development. However, review of the literature has not yet
established a conclusive relationship between central precocious
puberty and environmental agents. In the case of peripheral pre-
cocious puberty, the mechanism in young women is mediated by
hormonal receptors in peripheral tissues that are responsive to
estrogen or estrogen-like compounds.
In Puerto Rico, a temporal trend toward premature breast
development (premature thelarche) in girls and gynaecomastia in
boys has been noted during the early 1980s (Mills et al., 1981;
Bongiovanni, 1983; Saenz de Rodriguez et al., 1985; Freni-Titulaer
et al., 1986). Serum samples from 41 girls from Puerto Rico with
premature breast development and 35 control cases were analysed
for determining the possible presence of pesticides and/or phtha-
late esters. No pesticides or their metabolites could be found in
any of the serum samples; however, 28 (68%) of the girls with pre-
mature breast development had measurable levels of phthalates
[dimethyl, diethly, dibutyl and di-(2-ethylhexyl)], compared with
6 of the 35 (17%) control samples (Colon et al., 2000). In another
study, performed in Belgium, plasma of girls with precocious
puberty was screened for chlorinated pesticides; data showed an
increase in plasma levels of the DDT metabolite p,p0 -DDE in
foreign children with precocious puberty immigrating from devel-
oping countries to Belgium, compared with undetectable levels in
Impact of endocrine disruptor chemicals
Belgian-born girls with idiopathic or organic precocious puberty;
since DDT is still used in a number of developing countries, the
data suggest a possible relationship with early exposure to this
pesticide (Krstevska-Konstantinove et al., 2001).
The effect of in utero exposure to polybrominated biphenyls
(PBBs) on sexual maturation was evaluated in Michigan girls
whose mothers were accidentally exposed through diet to the
flame retardant FireMaster (Blanck et al., 2000). Effects on puber-
tal end points were assessed by questionnaires sent to mothers of
daughters ,18 years of age and to the daughters themselves.
The data reveal that menarche and pubertal hair growth were sig-
nificantly advanced in girls breastfed, i.e. with higher perinatal
exposure to PBBs. Namely, the adjusted OR (CI) were 0.8 (0.3–
1.9) for early menarche in the non-breastfed and 3.4 (1.2 –9.0)
for the breastfed group, respectively; for early pubertal hair
growth they were 0.9 (0.2– 4.3) in the not breastfed and 19.5
(2.8 –138.2) in the breastfed, respectively. There was no associa-
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tion with Tanner stage of breast development. These findings
are interesting because menarche and breast development are
estrogen-dependent events whereas pubertal hair growth is inde-
pendent from estrogen levels, suggesting that PBBs may interact
with different pathways. Several experimental data showed altera-
tions in morphological parameters of reproductive development
following exposure to environmental xenobiotics both in animal
and human studies (Laws et al., 2000a,b). For example, the chlori-
nated insecticide Metoxichlor, an EDC with estrogenic activity, is
able to accelerate the vaginal patency in prepubertal female rats
(Masutomi et al., 2003).
Due to the scarcity of human data with appropriate biomarkers,
toxicological studies could be useful for a reliable risk evaluation;
in particular, protocols focussed on prepubertal evaluation of
EDCs could provide relevant data on a phase of specific suscepti-
bility for human maturation and development (Kim et al., 2002).
Breast and endometrial cancer
The incidence of breast cancer increased steadily from the 1940s
through the 1990s in many industrialized countries, with the
highest risk found in Western Europe and North America. The
increases may be explained, only in part, to the increased screen-
ing procedures.
Dietary factors are known to contribute to breast cancer risk
(Willett, 2001). One class of dietary compounds that has received
much attention is phytoestrogens. Consumption of phytoestrogens,
particularly soy products, is higher in Asia than in the Western
World (Messina et al., 1994), where the rate of breast cancer is
highest (Kelsey and Berstein, 1996). In Japan and China, breast
cancer rates are half as high as in the USA (Coleman et al.,
1993). Migrants from Asia to the USA typically acquired a
breast cancer risk associated with their host nation by the second
generation, suggesting a direct influence of the environment
rather than a genetic factor (Barnes, 1998; Lamartiniere, 2000).
Extensive human studies support an etiological role for estro-
gens, all related to increased lifetime exposure to endogenous
estrogen (Hulka and Stark, 1995). Serum concentration of
17b-E2 is ~40% lower in Asian women than their Caucasian
counterparts (Peeters et al., 2003). In two studies on premenopau-
sal women (Lu et al., 2000; Kumar et al., 2002), modest consump-
tion of soy products seems be associated with a reduction of
steroid hormone levels, whereas a year-long dietary intervention
65
Caserta et al.
trial involving 34 premenopausal women fails to confirm this data
(Maskarinec et al., 2002). The reduction in steroid hormone levels
by phytooestrogens is proposed to occur via the direct regulation
of 17b-E2 biosynthesis and metabolism (Limer et al., 2004).
Despite their apparent effect on endogenous hormone levels, the
role of phytooestrogens in breast cancer initiation and develop-
ment is unclear. In vitro studies in estrogen-dependent human
mammary epithelial cells (MCF-7) have shown that genistein,
the main soy isoflavone, at low concentrations (0.1 –10 mM) can
stimulate cell proliferation whereas at higher concentrations
(10 mM) can act as inhibitor (Miodini et al., 1999). Other
studies suggest that both proliferative and antiproliferative
effects might be observed, depending on tumour cell type, concen-
trations, timing of phytoestrogen exposure and type of phyto-
estrogen given (Aldercreutz and Mazur, 1997). This may be
explained by the multiple mechanisms of action that phytoestro-
gens seem have in humans. The chemical structure of the isofla-
vones is similar to that of estrogens, these substances diffuse
through the cell cytoplasm and bind to both the nuclear ER sub-
types ER-a and ER-b, although they preferentially bind to and
activate ER-b; for this reason, they are sometimes classified as
selective ER modulators (SERMs) (Brzezinski and Debi, 1999;
Diel et al., 2004).
In vitro, phytoestrogens also possess a variety of non-hormonal
properties. Several phytoestrogens, except genistein, in addition
to their estrogenic effect also suppress the activity of the aromatase
enzymes, which are responsible for conversion of androgens to
estrogens (Almstrup et al., 2002). At concentrations in excess of
25 mmol/l, phytoestrogens are capable of inducing apoptosis in
human breast cancer cells (Po et al., 2002), also in ER-negative
cell lines, confirming the hormone-independent mechanism
of action. Dietary phytoestrogens are capable of inhibiting the tyro-
sine kinase activity, that are involved in a number of growth factor
signalling pathways, implicated in control of cell growth and differ-
entiation. Furthermore, phytoestrogens have antioxidant activity
(Ruiz-Larrea et al., 1997) and may stimulate the immune system
(Zhang et al., 1999) and inhibit angiogenesis (Su et al., 2005).
In addition to the potential protective effects against breast
cancer, some data suggest that phytoestrogens could actually
promote breast cancer. In vitro and in animal studies, genistein
stimulates the growth of estrogen-sensitive mammary cancer
cells (Hsieh et al., 1998; Allred et al., 2001).
Epidemiological studies on the relationship between soy con-
sumption and risk of breast cancer are also discordant. A Japanese
prospective study conducted in a cohort of 35 000 women (Key
et al., 1999) revealed no significant association between soy con-
sumption during adulthood and breast cancer incidence. Similar
data was found in a retrospective analysis in a multiethnic
cohort of non-Asian breast cancer patient and control individuals
residing in USA (Horn-Ross et al., 2001). Epidemiologic evidence
suggests that breast cancer chemoprevention by dietary phyto-
estrogen compound might be dependent on ingestion before
puberty, when the mammary gland is relatively immature (Shu
et al., 2001). A meta-analysis of currently available studies indi-
cates that high soy intake might reduce the risk of developing pre-
menopausal breast cancer but has no effect on post-menopausal
breast cancer risk (Trock et al., 2000). This raises the question
of whether foods rich in phytoestrogens may have complex
actions in such diseases as breast cancer, exerting both preventive
66
and promoting effects and perhaps depending on whether or not
the tumour is estrogen dependent. Published literature regarding
the effects of ingestion of dietary phytoestrogens by breast
cancer patients and survivors is, also, controversial (Messina
et al., 2001; This et al., 2001).
Thus, caution is necessary in promoting the putative beneficial
effects of phytoestrogens with respect to the overall mammary
neoplasms (Bouker and Hilakivi-Clarke, 2000). The majority of
data relating environmental EDCs to human breast cancer are
limited to persistent organochlorine compounds that have been
identified worldwide in human tissue, blood and milk (Adami
et al., 1995).
Since the metabolites of DDT have been identified in serum,
adipose tissue and breast milk of individuals with no history of
occupational exposure and/or living in areas where DDT has
not been used for years, long-term exposure to DDT through
food could be hypothesized to increase the risk for developing
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estrogen-dependent tumours such as breast cancer. However, pro-
spective case – control studies have failed to demonstrate that
DDT, and more specifically p,p-DDE, increases the risk for
breast cancer (Krieger et al., 1994; Hunter et al., 1997; Hoyer
et al., 1998; Dorgan et al., 1999; Helzlsouer et al., 1999; Ward
et al., 2000; Wolff et al., 2000a,b). In addition, two retrospective
case –control studies performed with post-menopausal women
(among which breast cancer tends to be more estrogen dependent)
did not find increases in breast cancer risk related to DDT exposure
(vant’Veer et al., 1997; Moysich et al., 1998), nor did other retro-
spective case –control studies including pre- and post-menopausal
women (López-Carrillo et al., 1997; Dello Iacovo et al., 1999;
Mendonca et al., 1999; Zheng et al., 1999; Aronson et al., 2000;
Bagga et al., 2000; Demers et al., 2000; Millikan et al., 2000;
Stellman et al., 2000; Wolff et al., 2000,b), in contrast to two posi-
tive studies (Olaya-Contreras et al., 1998; Romieu et al., 2000).
A recent combined analysis of five US studies showed no relation-
ship between p,p0 -DDE and breast cancer risk (Laden et al.,
2001a,b).
Many of the studies on the relationship between PCB exposure
and breast cancer have involved occupational exposure and have
not found a positive association (Adami et al., 1995; Houghton
and Ritter, 1995; NRC, 1999). In a prospective case –control
study in USA, after 9.5 years of follow-up, higher serum levels
of HCB induced a 2-fold increased risk of breast cancer.
However, there was no evidence for a dose-response relationship,
and the association was limited to women whose blood was col-
lected close to the time of diagnosis (Dorgan et al., 1999). In a
recent Belgian study, the serum levels of DDE and HCB were
compared in 231 women at the time of breast cancer discovery
and in 290 age-matched healthy controls. p,p’-DDE was found
in 76.2% of cases and in 71.1% of controls but HCB was
present only in 12.6% of cases (29 from 231) and in 8.9% of con-
trols (26 from 290). Mean serum levels were significantly higher in
patients versus controls for both compounds (2- and 3-fold higher
for DDE and HCB, respectively). No excess was observed among
nulliparous women or when familial history of breast cancer was
considered. In the cancer group, no differences in serum levels
of p,p’-DDE or HCB were found in relation with ER status,
Bloom stage or lymph node metastasis, but the HCB level was sig-
nificantly correlated with tumour size (P ¼ 0.026) (Charlier et al.,
2004).

Some studies deserve attention as they were performed in highly
polluted areas. The Seveso Women’s Health Study comprises 981
women, who were infants to 40-years old in 1976 and resided in
the most contaminated areas and had archived sera that was col-
lected soon after the infamous chemical accident in 1976. Model-
ling serum TCDD levels in 15 women that were diagnosed with
breast cancer, showed that individual body burden is significantly
related with breast cancer risk (Warner et al., 2002). The pleiotropic
effects of such potent hormone trigger as TCDD were in fact associ-
ated with a number of long-term health effects in Seveso exposed
population, such as increased morbidity and mortality from lym-
phoemopoietic and other neoplasms as well as markers of altered
endocrine-immune function; interestingly the increases were
often gender related. One evident, albeit still difficult to interpret,
effect was the clear link between exposure levels in men and a
lowered male/female sex ratio in their offspring (Pesatori et al.,
2003).
In the severely polluted Eastern Slovakia areas, a retrospective
study on serum samples of 24 breast cancer patients and 88 popu-
lation controls did show trends for positive correlation of breast
cancer with DDE and for negative correlations with PCB and
HCB; however, results were generally not significant, due to low
statistical power (Pavuk et al., 2003).
Although epidemiologic studies have not shown a strong
relationship between blood levels of PCBs and the risk of breast
cancer, an US cohort study revealed a stronger association
among post-menopausal white women with the inducible M2
polymorphism in the cytochrome P450 1A1 gene (Laden et al.,
2002). In a further population-based case-control study, breast
cancer risk is evaluated in relation to PCBs and the CYP1A1 poly-
morphisms M1 (also known as CYP1A1*2A), M2 (CYP1A1*2C),
M3 (CYP1A1*3) and M4 (CYP1A1*4). The study population
consisted of 612 patients (242 African American, 370 white)
and 599 controls (242 African American, 357 white). The results
confirm that CYP1A1 M2-containing genotypes modify the
association between PCB exposure and risk of breast cancer and
additional evidence suggesting that CYP1A1 M3-containing
genotypes modify the effects of PCB exposure among African
American women (Li et al., 2005a,b).
Moreover, recent data suggest that underarm cosmetics might
be a cause of the development and progression of breast cancer,
because these cosmetics contain a variety of substances with endo-
crine activity (e.g. parabens) that are applied frequently in an area
directly adjacent to the breast (Darbre, 2006).
Uterine cancer is more common in developed countries, with a
similar pattern of hormonal risk factors as breast cancer. There is
clear evidence that unopposed estrogen is the major risk factor for
endometrial cancer. Neonatal treatment of mice on post-natal days
1– 5 with either the potent estrogen agonist diethylstilboestrol
(DES) or the phytoestrogen, genistein, has been shown to cause
uterine adenocarcinoma by 18 months (Potischman et al., 1996;
IARC, 1999). As with other cancers, the timing of exposure is
critical to the potential development of uterine cancer; in fact treat-
ment of adult mice with comparable levels of DES does not induce
uterine neoplasms (Newbold et al., 1991). Soy isoflavones inhibit
E2-mediated endometrial proliferation in macaque monkeys
(Cline and Foth, 1998). This may not be surprising in the case
of phytoestrogens that have both estrogenic and antiestrogenic
effects, acting as SERMs (Whitten and Patisaul, 2001).
Impact of endocrine disruptor chemicals
Epidemiologic data on the effects of environmental EDCs on
endometrial cancer are limited. Sturgeon et al. (1998) found no
association between endometrial cancer and 27 PCB congeners,
4 DDT-related compounds and 13 other organochlorine com-
pounds. Several retrospective occupational cohort studies also
observed no association (Bertazzi et al., 1987; Brown, 1987;
Sinks et al., 1996). In the Seveso industrial accident, TCDD
exposure appeared to reduce the risk of uterine cancer, but the
number of cases was too small for a comprehensive evaluation
(Bertazzi et al., 1993).
There is some evidence that dietary isoflavones protect from
endometrial proliferation. Specifically, high consumption of soy
products and other legumes in US women was associated with a
decreased risk of endometrial cancer for the highest compared
with the lowest quartile of soy intake (Goodman et al., 1997;
Horn-Ross et al., 2003). Controversially, a recent randomized
doubled-bind, placebo-controlled study on 298 post-menopusal
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women shows an increased incidence of endometrial hyperplasia
following 5 years of treatment with 50 mg of soy isoflavones
(Unfer et al., 2004). Thus, phytoestrogenic supplements should
be reconsidered, particularly in women at high risk for endometrial
cancer.
Conclusions and recommendations
The major limiting factor in drawing any conclusions about female
reproductive system effects and EDCs is the scarceness of actual
exposure data. In fact, in the available studies, exposure can only
be inferred and not actually measured. Another problem relies
on the sample sizes which are often too small to allow detection
of an effect, even if one were present. Although there is evidence
for geographical differences and temporal trends in some aspects
of human reproduction, there has been no systematic attempt to
look for evidence that the mechanisms behind these changes
could involve endocrine pathways. Despite these drawbacks, the
biological plausibility of possible damage to human reproduction
derived from exposure to EDCs seems strong when viewed against
(i) the background of known influences of endogenous and
exogenous hormones on many of the processes involved, and
(ii) the evidence of adverse reproductive outcomes in laboratory
animals exposed to EDCs. Thus, concerns about females derive
more from biological knowledge about the influence of sex hor-
mones on development and adult reproductive function, rather
than from studies on environmental chemicals. Contrary to male
fertility, data on EDC effects on female reproductive health are
sparse both in the human and experimental literature; there is
still no attempt of an unified hypothesis such as the ‘testicular dys-
genesis syndrome’ elaborated for EDC-related effects on male
reproductive system (Sharpe, 2003). Evaluation of possible links
to EDCs could be pursued by exploiting identified temporal
trends and geographical differences in the sex ratio, whereas the
frequency of endometriosis in the general population offers oppor-
tunities for human studies requiring fewer numbers than might be
required for other end points. Time-to-pregnancy can also be used
as an apical tool to broadly examine the possible influence of
environmental chemicals on both male and female reproductive
function.
Endocrine-related cancer and especially breast cancer are most
interesting issues. Although numerous human epidemiological
67
Caserta et al.
studies have been conducted to determine whether environmental
EDCs may contribute to an increased risk of breast cancer, the
results remain inconclusive. Overall, the current scientific evi-
dence (from human and animal studies) does not support a
direct association between exposure to environmental EDCs and
increased risk of breast cancer, although the evidence may be
stronger in situations of high pollution (Seveso, Slovakia) and in
a fraction of genetically susceptible individuals (Ursin et al.,
1997; Brunet et al., 1998; Jernstrom et al., 1999; Nkondjock
et al., 2006). Such studies prompt the need for translational
research on EDC-related mechanisms, leading to the characteriz-
ation of biomarkers of exposure, effects and susceptibility in
order to perform a reliable risk evaluation. Breast cancer is
likely due to many factors, including genetics, lifestyle, diet,
endogenous hormone status and environmental factors. Research
on whether potential complex interactions among these factors,
modulated by individual genetic susceptibility, produce breast
cancer is critical. Until consistent and compelling data on these
issues become available, the role of EDCs in breast or uterine
cancer incidence is likely to remain a highly controversial issue.
However, all the studies published to date have measured EDC
exposure levels in adult women. The time of development when
exposure takes place may be critical to define the dose – response
relationships of EDCs for breast cancer as well as for other
health effects such as endometriosis. The perinatal period and
the period between age at menarche and age at first full-term preg-
nancy may be particularly important for breast tumour develop-
ment and latency. Data are also required to investigate the late
health outcomes of precocious peripheral puberty, for which
EDC may be also a risk factor.
The claim that the time of life when exposure takes place (e.g.
prenatal, neonatal, childhood and adolescence) may be the most
critical factor is supported by human data and by basic research
in animal models. Regarding endocrine-active compounds, the
case of DES is the most well-known example, with young
adult offspring exposed in utero to this potent drug having a
higher rate of reproductive tract abnormalities in both sexes as
well as of the rare clear-cell vaginal adenocarcinoma in female
offspring (Swan, 2000). Of course, most EDC are unlikely to
be nearly as potent as DES; even in such a case, the exposure
levels are expected to be much lower than those occurring
from pharmacological treatment. Nevertheless, it cannot be
excluded that adult women currently at risk for breast cancer
may have been exposed to exogenous EDCs in utero or during
infancy, childhood and adolescence in the mid-1900s when con-
taminant levels of organochlorines were higher. Experimental
evidence shows that gestational and/or lactational exposure to
dioxin-like chemicals or to bisphenol A alters the differentiation
of mammary gland in rodents (Fenton et al., 2002; Muto et al.,
2002; Munoz-de-Toro et al., 2005). Research is urgently needed
to address the role of timing of exposure. Human prospective
studies would be complex, time-consuming and expensive;
researchers should be encouraged to utilize and develop animal
models to address the important issue of late health effects of
early exposures, e.g. prepubertal animal studies for the effects
of EDCs on sexual maturation. In parallel, biological banks
should be exploited in order to conduct retrospective follow-up
and nested mother-child cohort studies using specific
biomarkers.
68
In conclusion, the currently available human data are
inadequate to support a conclusion about whether the female
reproductive system is adversely affected by exposure to EDCs;
however, the weight of the evidence is adequate to address
further studies as well as to prompt precautionary actions
against excess exposure to xenobiotics specifically active on hor-
monal homeostasis.
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Submitted on February 27, 2007; accepted on June 19, 2007