Reproductive Toxicology 25 (2008) 304315

Contents lists available at ScienceDirect

Reproductive Toxicology
journal homepage: www.elsevier.com/locate/reprotox

Review

Cadmium: Toxic effects on the reproductive system and the embryo

Jennifer Thompson , John Bannigan
School of Medicine and Medical Science, Conway Institute of Biomolecular and Biomedical Research, University College Dublin, Ireland

a r t i c l e

i n f o

Article history:
Received 5 October 2007
Received in revised form 6 February 2008
Accepted 12 February 2008
Available online 19 February 2008
Keywords:
Cadmium
Chick embryo
Testis
Ovary
Blastocyst
Cell adhesion
Oxidative stress
Ventral body wall defect

a b s t r a c t
The heavy metal cadmium (Cd) is a pollutant associated with several modern industrial processes. Cd
is absorbed in signicant quantities from cigarette smoke, and is known to have numerous undesirable
effects on health in both experimental animals and humans, targeting the kidneys, liver and vascular systems in particular. However, a wide spectrum of deleterious effects on the reproductive tissues and the
developing embryo has also been described. In the testis, changes due to disruption of the bloodtestis
barrier and oxidative stress have been noted, with onset of widespread necrosis at higher dosage exposures.
Incorporation of Cd into the chromatin of the developing gamete has also been demonstrated. Ovarian
Cd concentration increases with age, and has been associated with failure of progression of oocyte development from primary to secondary stage, and failure to ovulate. A further mechanism by which ovulation
could be rendered ineffective is by failure of pick-up of the oocyte by the tubal cilia due to suboptimal
expansion of the oocytecumulus complex and mis-expression of cell adhesion molecules.
Retardation of trophoblastic outgrowth and development, placental necrosis and suppression of steroid
biosynthesis, and altered handling of nutrient metals by the placenta all contribute to implantation delay
and possible early pregnancy loss. Cd has been shown to accumulate in embryos from the four-cell stage
onwards, and higher dosage exposure inhibits progression to the blastocyst stage, and can cause degeneration and decompaction in blastocysts following formation, with apoptosis and breakdown in cell adhesion.
Following implantation, exposure of experimental animals to oral or parenteral Cd causes a wide range of
abnormalities in the embryo, depending on the stage of exposure and dose given. Craniofacial, neurological, cardiovascular, gastrointestinal, genitourinary, and limb anomalies have all been described in placentates, with axial abnormalities and defects in somite structure noted in sh and ventral body wall defect
and vertebral malformation occurring in the chick. In this paper, we examine the mechanisms by which
Cd can affect reproductive health, and consider the use of micronutrients in prevention of these problems.
2008 Elsevier Inc. All rights reserved.

Contents
1.
2.

3.

4.

Introduction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
Effects of cadmium on reproduction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
2.1.
The testis and spermatogenesis . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
2.2.
The ovary and oogenesis . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
2.3.
The pre-implantation embryo . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
2.4.
Implantation . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
2.5.
The post-implantation embryo . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
Mechanisms of reproductive toxicity . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
3.1.
Altered cell adhesion . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
3.2.
Oxidative stress and apoptosis . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
3.3.
Ionic and molecular mimicry . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
3.4.
DNA damage and effect on the cell cycle . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
Conclusions . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

Corresponding author at: Room C212, Health Sciences Building, School of Medicine and Medical Science, University College Dublin, Beleld, Dublin 4, Ireland.
Tel.: +353 1 7166634; fax: +353 1 7166649.
E-mail address: Jennifer.Thompson@ucd.ie (J. Thompson).
0890-6238/$ see front matter 2008 Elsevier Inc. All rights reserved.
doi:10.1016/j.reprotox.2008.02.001

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J. Thompson, J. Bannigan / Reproductive Toxicology 25 (2008) 304315

1. Introduction
Cadmium (Cd; atomic number 48; relative atomic mass 112.40)
is a toxic metal that belongs to group IIb in the periodic table. It
occurs in nature at low concentrations, mainly in association with
the sulde ores of zinc (Zn), lead (Pb), and copper (Cu). However,
due to the widespread nature of its occurrence, it is present in
measurable amounts in almost everything that we eat, drink, and
breathe [1]. Human activities adding to the Cd load in the environment include the combustion of fossil fuels, leechate from landll
sites, run-off from agricultural land, and mining residues, especially from Zn and Pb mines [2]. Electroplating to protect steel
from corrosion, and the manufacture of NickelCd batteries, pigments, stabilizers and alloys [35] also produce Cd as a by-product.
A pollutant of worldwide concern, Cd has been reviewed by the
International Register of Potentially Toxic Chemicals of the United
Nations Environment Program, and included on the list of chemical substances considered to be potentially dangerous at the global
level [6]. It is now recognized that human exposure to Cd must be
minimized.
The WHO safe level for human ingestion of Cd has been estimated at 500 g/week [5]. Absorption of oral Cd tends to be erratic,
with continuing presence of unabsorbed radio-Cd in the gut lumen
for 35 weeks after a test meal in human subjects [7]. Dietary Cd
intake estimates in European countries were 1030 g per day [8],
with increased risk of consumption with certain foods such as shellsh, offal, and rice [912]. Individuals consuming dredge oysters in
New Zealand were found to have a daily fecal excretion of Cd up to
580 g, equivalent to more than 10 times the provisional tolerated
weekly intake (PTWI). The dietary Cd absorption rate is about 5%,
rising to 2030% in some individuals [13]. In non-Cd-polluted areas,
the most signicant human route of Cd intake is cigarette smoking, with various estimates of 0.21.0 g Cd assimilated with each
cigarette smoked [1317], accounting for approximately half the
total human Cd intake. However, bioavailability of inhaled Cd oxide
is relatively high, with 10% deposited in lung tissues and another
3040% absorbed into the systemic blood circulation of smokers
[13]. The whole-body biological half-life for Cd is very long, having a fast component of 75128 days and a slow component of

305

7.426.0 years [5,18,19]. This raises the possibility of cumulative

effects.
After absorption, Cd is transported to the liver, bound to albumin [20], where it induces the synthesis of metallothionein (MT), a
class of small cysteine-rich heavy metal binding proteins. Changes
within the liver itself following parenteral administration of CdCl2
are dose- and time-dependent, ranging from moderate diffuse hepatocellular degeneration through to multifocal necrosis [21,22].
Following release from the liver, MT-bound Cd enters the plasma. Cd
is eliminated in the urine. MT-bound Cd appears in the glomerular
ltrate, from where it is re-absorbed intracellularly by renal tubule
cells. In the latter, the Cd is cleaved from the MT by lysosomal action,
and Cd++ ions are re-excreted into the tubular uid. The ability of
Cd to induce hepatic and renal lesions exacerbates its toxic effects,
and compounds its propensity to accumulate over years.
A large body of work exists outlining the effects of Cd on gametogenesis in both males and females, and implicating its compounds
in early embryolethality and implantation failure. In addition, animal studies have shown a wide range of anomalies following
exposure at specic stages of embryogenesis, and emerging evidence has indicated that Cd may also be linked to pathological
processes in late pregnancy and in the early postnatal period,
causing third trimester complications and minor but signicant
problems in the offspring of exposed individuals. The focus of this
review, therefore, is to examine the consequences of Cd exposure
on reproductive processes and embryonic development, and to look
at mechanisms by which these effects might occur.
2. Effects of cadmium on reproduction
An overview of reproductive processes affected by Cd, along
with suggested mechanisms of action, is given in Table 1.
2.1. The testis and spermatogenesis
Testicular changes due to Cd toxicity have been seen in a variety of animal models at different stages of growth and maturity.
Gonadal development in mouse embryos exposed to Cd in early
organogenesis was studied by Tam and Liu [23]. Genital ridge size

Table 1
Summary of the early reproductive effects of Cd and suggested mechanisms by which these effects occur
Reproductive effect

Histological nding

Probable underlying mechanism

References

1. Failure of maturation from

leptotene to pachytene
spermatocytes

Disruption of bloodtestis barrier, testicular necrosis

[2528,122124]

2. Failure of maturation of
spermatozoa; reduced viability
3. Failure to ovulate, defective
steroidogenesis
4. Suppressed oocyte maturation

Disruption of bloodepididymal barrier, abnormal

sperm morphology
Damage to the endothelium in follicular arteries and
to the ovarian interstitial stroma, ovarian necrosis
Suppression of cumulus expansion, reduced numbers
of oocytes reaching metaphase II, reduced
fertilization of oocytes, increased oocyte
degeneration

Substitution of Cd for Ca in adherens and

desmosomal junction cadherins;
disruption of tight junctions via the
TGF3/p38 MAP kinase pathway, oxidative
stress
Junctional disruption
Junctional disruption, ? oxidative stress

[4856,59]
[41,5760]

5. Failure of developmental
progression in pre-implantation
embryo

Failure of progression from two-, four- and eight-cell

stages to morula, failure of
compaction/decompaction

6. Failure of implantation

Failure of oocyte pick-up by infundibulum, abnormal

trophoblast outgrowth and transformation

Junctional disruptiondue to suppression

of synthesis of hyaluronic acid and/or
inhibition of gap junctional intercellular
communication and connexin
phosphorylations, ? oxidative stress, ?
genotoxicity
? Junctional disruptiondue to inhibition
of gap junctional intercellular
communication and connexin
phosphorylation, and disruption of tight
junctions, oxidative stress
Suppression of synthesis of hyaluronic
acid, ? suppression of cell adhesion
molecule expression, displacement of Ca
by Cd on calmodulin

(?): Possible mechanisms that have not yet been proven denitively.

[41,125]

[6271]

[58,64,7381]

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J. Thompson, J. Bannigan / Reproductive Toxicology 25 (2008) 304315

was reduced in exposed animals, with retarded germ cell migration into the ridges, resulting in depleted populations of germ cells,
defective maturation of gametes and subfertility in male offspring.
Young rabbits dosed with 1.02.25 mg/kg body weight Cd had a
signicant decrease in germinal epithelial volume after 48 h, with
signs of epithelial and basement membrane damage [24], along
with reduced epithelial volume in the epididymis after 5 months
of treatment.
Similarly, adult male rats have been shown to develop
gonadal damage following administration of Cd, either orally or
subcutaneously (SC). Focal testicular necrosis and reduced spermatogenesis were seen in rats that received a single dose of
100150 mg Cd/kg orally within 2 weeks of administration [25].
Rats injected with a large dose of CdCl2 (7 mg/kg SC) showed pronounced testicular hemorrhage and edema 24 h after treatment
[21]. Fende and Niewenhuis [26] described endothelial cell damage
in testicular blood vessels commencing 3 h after SC administration of Cd. Light and electron microscopy initially revealed edema,
with widening of interstitial spaces and deposition of proteinaceous material 3 h after treatment, progressing to separation of
endothelial cells 6 h after injection and extensive hemorrhage and
histological disruption at 24 h. Testicular morphology 3 months
after initial Cd exposure was greatly altered, with degenerated
seminiferous tubules, abnormal Leydig cells, brosis, and reduced
testicular size [27]. Similar effects were noted in a histological and
ultrastructural study of gerbil testes [28].
Other cell populations within the testis have also been implicated as targets of Cd toxicity [2931]. SpragueDawley rats
injected SC with 0.6 mg/kg Cd daily over a 6-week period were
found to accumulate Cd in the testes, mainly in spermatogonia
and spermatocytes, with consequent reduction in both of these
cell types [32]. Other studies have failed to demonstrate incorporation of Cd into sperm chromatin, suggesting instead that subfertility
following Cd administration might result from damage to supporting testicular tissue [33]. Cd dose in the latter experiments was,
however, considerably lower (0.1 mg/kg CdCl IP) than in those conducted by Aoyagi et al. [32] or by Toman et al. [24], and Dixon et al.
[34] produced compelling data indicating that Cd is indeed incorporated into early and late spermatids and spermatogonia. Disruption
of the bloodtestis barrier (BTB) by Cd, with consequent reduction
in germ cell numbers and infertility, has been linked to reduced
occludin expression, thought to be mediated by the TGF3/p38
MAP kinase signaling pathway [35,36], indicating the involvement
of occludens junction disturbance as well as adherens junction
breakdown in BTB disruption [37].
Heavy smoking, the commonest source human intake of Cd, has
been associated with low sperm count and motility. A strongly positive relationship has been identied between azoospermia and
serum and seminal uid Cd levels in infertile Nigerian males [38].
However, other studies have shown no differences in semen quality
or fertility between smokers and non-smokers, even though signicantly increased amounts of Cd were found in seminal uid
in those smoking 20 or more cigarettes per day [39]. In the rat
model, increased rates of abnormal sperm morphology have been
noted with increased seminal Cd levels [40], with positive correlation with TNF- and IFN-, and a negative relationship with IL-4.
All seminal parameters and cytokine anomalies in this model were
reversible by administration of Zn with Cd. It has been suggested
that absence of certain micronutrients in the diet contribute to male
infertility associated with raised Cd levels [38].
In in vitro studies, Cd at 20 M concentration has been shown to
signicantly decrease the viability of spermatozoa to 35.6% when
compared with control viability of 54.4% [41]. Lower concentrations
of Cd (2 M), while having a sperm viability rate comparable with
controls, increased acrosome-reacted spermatozoa (45.2%) com-

pared with both control and higher concentrations of Cd (31.9 and

32.5%, respectively). Polyspermy was signicantly increased in this
group (23.5%) when compared with controls and 20 M Cd (6.7
and 0%, respectively), implying that Cd at higher concentrations
affects cell viability, and, at lower concentrations, affects physiological function of spermatozoa.
In addition to effects on fertility, Cd administration has been
linked with tumors of the prostate and testis in experimental animals [3], preventable by Zn administration [20]. Waalkes et al.
[42] demonstrated that SC or oral Cd induced prostatic neoplasms
in rats, provided testicular function was maintained. A signicant
positive relationship was found between ingested Cd and prostate
cancer in humans in a case control study conducted by West et al.
[43]. However, cohort studies by Kazantzis [44] and Sorahan [45]
failed to show any association. Although a positive relationship has
been demonstrated between proliferative testicular lesions and Cd
in rats [46,47], no denitive link exists in humans.
2.2. The ovary and oogenesis
Oocyte development and associated events have been disrupted
by Cd administration in numerous different species, including
Xenopus laevis [4850], hamsters [51,52], rats [5356], mice [52],
pigs [57,58], and sheep [41]. In Xenopus, exposure to various levels of Cd via the culture water over 30 days resulted in a decrease
in number of developing oocytes in those exposed to 1.0 mg/l
Cd. Reduction in the total number of oocytes and ovarian necrosis were seen at concentrations 2.5 mg/l, and reduction in ovarian
weight, at concentrations 5 mg/l [48]. A reduction in breeding and
fertilization rate was also noted 2.5 mg/l. Oocyte germinal vesicle breakdown was inhibited by Cd at higher concentrations [49].
Lienesch et al. [50] similarly found a reduction in the percentage
of oocytes undergoing normal progression after Cd injection into
the dorsal lymph sac every other day for 21 days, and a dramatic
increase in the population of atretic oocytes.
The effect of Cd on ovulation in the golden hamster was
described by Saksena and Salmonsen [51]. Ovulation was inhibited at a dose of 5 mg/kg CdCl2 SC or higher, particularly if given
close to the LH surge. Failure to ovulate was associated with ovarian
necrosis and hemorrhage and a decreased progesterone level in the
serum. Although ovarian lesions resolved within 4 days, prolonged
periods of sterility were recorded in hamsters given 510 mg/kg
CdCl2 , following which normal pregnancy with normal litter numbers was achieved. Rehm and Waalkes [52] investigated the effect
of a single SC injection of CdCl2 at a dose of 2047.5 mol/kg on
non-gravid Syrian Hamsters and rats and mice of different strains.
Ovarian hemorrhagic necrosis occurred in hamsters treated with
this dosage range of Cd, with administration to sexually immature animals and injection just prior to ovulation causing the most
severe lesions at all doses. Assessment by light microscopy revealed
that small arteries in developing follicles and the interstitial stroma
were particularly susceptible to damage, while other ovarian tissue,
such as primordial oocytes and corpora lutea, appeared resistant.
Full morphological recovery occurred in hamster ovaries within 2
months, despite a loss of nearly 50% ovarian weight following the
initial insult. Furthermore, pre-treatment with Zn acetate markedly
reduced the extent of ovarian damage. Uterine and cervical hemorrhages occurred in immature hamsters at higher doses of CdCl2 .
Rats similarly showed dose- and age-dependent toxicity in the
ovaries, uterus and cervix. The genital tract in mice was more resistant to Cd-induced lesions, and both mice and rats demonstrated
strain-dependent sensitivity to Cd.
Ovarian steroidogenesis in the non-gravid rat [53], and in
pregnant rats [5456] has been profoundly inuenced by Cd
administration at various times of the estrus cycle. CFY rats given

J. Thompson, J. Bannigan / Reproductive Toxicology 25 (2008) 304315

1015 mg/kg CdCl2 on the day of diestrus II were found to have

increased serum prolactin and decreased FSH, LH and basal progesterone levels the day after treatment, indicating an effect on the
hypothalamicpituitaryovarian axis [53]. Concomitant treatment
with four SC injections of 1020 mg/kg ZnCl2 protected against
sterility induced by CdCl2 at 10mg/kg body weight in in vivo experiments [54]. However, Zn failed to protect against Cd-induced drop
in progesterone production in in vitro culture of rat ovarian granulosa cells. SpragueDawley rats given 2.55 mg/kg Cd SC on the
day before ovulation similarly demonstrated suppression of ovarian steroidogenesis, with an initial reduction in progesterone and
testosterone [59], and serum estradiol most severely affected 24 h
after treatment [56]. Cd was also found to depress and delay progesterone secretion in early pregnancy [55]. Concentration of Cd in
the oviducts following SC administration of 2.510 mg/kg CdCl2 on
day 1 of pregnancy was found to rise in a time- and dose-dependent
manner. However, animals receiving Cd later in pregnancy (day 16
or gestation) were relatively unaffected [56].
The role of Cd in suppressing FSH-induced cumulus expansion in oocytecumulus complexes (OCC) isolated from large antral
porcine follicles has been described by Mlynarcikova et al. [57].
Incubation with micromolar concentrations of Cd decreased both
the degree of cumulus expansion and progesterone production by
the cumulus cells during 42 h of incubation of the OCC with FSH.
High concentrations of Cd completely suppressed oocyte maturation, and signicantly suppressed synthesis of hyaluronic acid, an
integral component of expanded cumulus cells [58]. In experiments
with the ovine oocyte, Leoni et al. [41] found that maturation rate
was signicantly affected at 2 and 20 M CdCl2 , with a metaphase
II (MII) rate of 63.8 and 32.0%, respectively when compared with
controls (MII rate 96.8%). Both Cd concentrations also decreased
numbers of oocytes resting in MII (29.0 and 19.8%, respectively;
control rate 93.8%), reduced numbers of fertilized oocytes in culture (25.9 and 4.7%, respectively; control rate, 76.1%), and increased
the rate of oocyte degeneration in both groups.
Exposure to Cd of cultured human ovarian granulosa cells was
found to cause reduction in progesterone production when compared with controls, and also caused morphological alterations in
a time- and dose-dependent manner, with rounding up of cells
after 48 h, retraction of cellular extensions, and detachment from
neighboring cells [60].
2.3. The pre-implantation embryo
In addition to its effects on gametogenesis, Cd may also reduce
the possibility of a successful pregnancy by interfering with the
development of the pre-implantation embryo. Gamete fusion
in the ovine model was found to be signicantly reduced upon
exposure to 220 M Cd [41]. Fertilization of mouse oocytes was
unaffected when cultured at 1.6 M Cd, and treated ova went on to
cleave into two-cell stage embryos at a similar rate to controls [61],
although a reduced number of these embryos reached blastocyst
stage. Gametes treated with 0.40.8 M Cd were not inhibited
from progressing to the blastocyst stage, but there was increased
embryo loss at the implantation stage in the treated group. In
vitro studies in which murine embryos were cultured at higher Cd
concentrations (1050 M) at the two-cell stage, demonstrated
a dose-dependent failure of developmental progression [62].
Although embryos cultured at this stage with 109 Cd showed little
accumulation of radioisotope, there was marked incorporation into
blastocysts [63]. In vivo exposure of the murine embryo to Cd at
the two-cell stage by SC injection of the dam with 2538 M Cd/kg
body weight on the morning of day 2 (D2) similarly failed to prevent initiation and maintenance of pregnancy when examined on
D8 [63], but did delay implantation. Embryos that implanted went

307

on to become normal fetuses [61], with resorption rates and body

weights comparable to controls. Yu et al. [64] found that exposure
of four-cell and morula stage mouse embryos to 510 g/ml
CdCl2 for 24 h caused degeneration and decompaction of treated
embryos. In the rat, incubation for 24 h in 1 g/ml CdCl2 prevented
eight-cell embryos and morulae from developing to the blastocyst
stage [65], with clear evidence of apoptosis following exposure.
Previous papers have shown that Cd inhibits gap junction intercellular communication and connexin phosphorylation [66,67],
both of which are essential processes in the progression from the
eight-cell stage through to compaction [68,69]. Protection of the
early embryo from Cd toxicity has been achieved by preventing
Cd uptake by cells by pre- or co-incubation with Zn at 100 molar
dose of Cd [63,70], and by co-culture with nifedipine at 0.01 molar
excess [63]. Antioxidants are also known to be protective [71].
2.4. Implantation
An early step in the reproductive process is the movement of
the secondary oocyte, arrested in the second meiotic metaphase
(MII), into the Fallopian tube, where it will encounter spermatozoa
and undergo the initial steps of fertilization. Thereafter, the oocyte
completes meiosis II to become a denitive oocyte shortly before
the fusion of male and female pronuclei [72]. This process requires
intact meiotic mechanisms, including normal spindle formation,
and correct sequential expression of cell adhesion molecules on
both spermatozoa and oocyte [73]. The oocytecumulus complex
(OCC) is picked up by cilia on the exterior surface of the infundibulum, which bind reversibly to the matrix of the OCC. The proteins
and hyaluronan in this matrix link together and stabilize the OCC,
thus facilitating pick-up of the entire complex by the cilia [74].
Cd-suppressed synthesis of hyaluronic acid in the OCC matrix [58]
could be linked to the inability of the infundibular cilia to successfully introduce the oocyte into the Fallopian tube. Cell adhesion is
an integral and important part of OCC pick-up [75], and although
the precise sequence and pattern of expression of cell adhesion
molecules during this process has yet to be elaborated, the involvement of gap junctions in cumulus expansion has already been
identied [76], and the involvement of other junction types is
possible. Cd-induced mis-expression of cell adhesion molecules,
including various connexins [66,67], and also delocalization of
molecules such as zonula occludens (ZO)-1 and N-cadherin at a
cellular level has been associated with abnormalities in cell adhesion in gap, occludens and adherens junctions [77]. Accordingly,
implantation of Cd-treated embryos is reduced when compared to
controls, even at doses that fail to disturb in vitro development of
the early embryo [64].
Trophoblast formation and invasion have also been identied as
targets for Cd toxicity. Low concentrations (0.5 g/ml) of CdCl2 presented to blastocysts in vitro signicantly retarded the trophoblastic
outgrowth areas and transformation of trophectoderm into giant
cells [78]. Higher concentrations (15 g/ml) caused trophoblastic
detachment. Placental necrosis in rats injected with 40 M/kg Cd
SC on day 18 of pregnancy was preceded by an early toxic effect
on trophoblast cells [79]. JAr choriocarcinoma cells, a neoplastic
trophoblast cell line which is similar to early human trophoblast
cells, was used by Powlin et al. [80] to determine possible effects
of Cd on the trophoblast. Exposure of this cell line to 2040 M Cd
inhibited cell proliferation. Prolonged exposure to lower concentrations of Cd (0.16 M) caused changes in cellular morphology and
detachment from neighbouring cells in association with reduced
Ca inux [81]. Modication of the effect of Cd on the cell cycle
by zaldaride, a calmodulin inhibitor, indicated that calmodulin, an
intracellular calcium-binding protein, has a role in mediating Cdinduced toxicity in the trophoblast. Increased metallothionein (MT)

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Table 2
Cd exposure of the post-implantation embryo
Stage of exposure

Species tested

Teratogenic effects observed

Gastrulation

Zebrash, Xenopus, Rodents (hamster, rat, mouse)

Early neurulation

Chick, xenopus, rodents

Late neurulation

Chick, xenopus, rodents

Post-neurulation

Rodents

Growth retardation, craniofacial defects, ocular malformations, notochord/somite

abnormality, gut malformations, cardiovascular anomalies, hypopigmentation,
skeletal anomalies
Abnormal body axis, ventral body wall (VBW) defects, upper limb defects, primary
neural tube defects, facial clefting, diaphragmatic hernia, cardiovascular anomalies
Abnormal body axis, VBW defects (abdominal), lower limb defects, retarded
somite growth
Secondary neural tube defects, limb reduction defects, urogenital abnormalities,
umbilical hernia

The stage of exposure is related to the teratogenic effects observed. In all species treated, exposure at the time of gastrulation resulted in widespread damage to the embryo,
with multiple structural malformations.

protein expression has been found in trophoblast cells exposed to

Cd [80,81] and other placental sites [82,83].
The presence of MT in the placenta is generally accepted to be a
protective mechanism against Cd toxicity within the cells in which
it is produced, and is also thought to restrict Cd movement across
the placenta, as the level of Cd is lower in fetal than in maternal
blood [82]. However, it is thought by some [82,84] to have detrimental effects, by binding Zn or by interfering with the ability of
trophoblast cells to handle Ca and Zn at the cytosolic level. Weir et
al. [85], using the perfused human placenta model, demonstrated
a reduction in transplacental transfer of Zn to the fetus, and also a
drop in human chorionic gonadotrophin (hCG) production by the
placenta, commencing 4 h from exposure. These ndings were conformed by Breen et al. [86]. Piasek et al. [59] found that the placentas
of smoking mothers contained double the Cd and approximately
half the progesterone of those of non-smokers, and a reduction
in placental iron in smokers in whom placental Cd was elevated
[87]. Cd interference with progesterone biosynthesis in cultured
human trophoblast cells has been linked with reduced activity of
P450 cholesterol side-chain cleavage enzyme and decreased 3hydroxysteroid dehydrogenase enzyme mRNA [88].
2.5. The post-implantation embryo
In animal studies, Cd has conrmed teratogenic and embryotoxic effects in a wide variety of species [8993]. The character
of the changes induced has been found to be dependent upon
the strain and species investigated, the dose of Cd given, the
route of administration, and the stage of embryogenesis [92,9496]
(Table 2). Anomalies described in the various species investigated
are also dependent on the timing of Cd administration. Alterations
in gastrulation occur in a concentration-dependent manner, and are
similar across the spectrum of animal models investigated. Effects
include retarded growth, microcephaly, body axial deformity and
ocular anomalies [97,98]. In the FETAX (Frog Embryo Teratogenesis Assay: Xenopus) experiment, Sunderman et al. [93] exposed
Xenopus laevis embryos to various concentrations of Cd from 5 h
post-fertilization, with exposure continuing through the gastrulation period up to 101 h post-fertilization. Anomalies described
included gut malformations, heart lesions, facial anomalies and n
dysplasias, in addition to the aforementioned defects. In zebrash,
cardiac edema, tail malformations and hypopigmentation were also
reported [99]. Hen Chow and Cheng [100] noted that zebrash
embryos exposed to Cd during the gastrulation period had defects
in somite structure, which probably contributed to altered axial
curvature, and also abnormal notochordal morphology, with the
notochord failing to extend into the tail region.
Alteration in body axial curvature was also a prominent nding in studies on the chick embryo. Menoud and Schowing [89]
detected distortion of the longitudinal body axis and symmelia, a

condition in which the limbs are orientated backward and fused, in

the chick following administration of Cd at 42 h, a stage at which
neurulation is complete apart from the closure of the posterior
neuropore. The upper limbs were predominantly affected when
treated at this time point. In our more recent studies [101,102], chick
embryos were treated with the same dose of Cd after 60 h incubation (corresponding to Hamburger and Hamilton stage 1617), and
developed a similar effect in the region of the lower limbs, associated with a large defect in the ventral abdominal wall. Review of
Menoud and Schowings work revealed that strophosomy, a form
of deciency in the ventral body wall, is also described in their
ndings. Timed histological studies in our laboratory demonstrated
that the ventral body wall defect, along with abnormal positioning
of the limbs, was due to an abnormality in the direction of growth of
the lateral plate mesoderm (LPM), with failure of embryonic folding in this region. Similar to the ndings in the zebrash, we found
that Cd administration had profound effects on somite structure,
with increased apoptosis in all three derivative embryonic tissues (sclerotome, myotome and dermatome), retarded growth and
differentiation and reduced somite numbers 24 h after treatment
[101103]. The main thrust of our own work has been the investigation of Cd-induced ventral body wall defect, which is a highly
reproducible and specic abnormality, similar in many respects to
the human condition omphalocele, caused by direct exposure of the
HH stage 1617 chick embryo blastodisc to 50 l of 5090 M Cd
[101]. The primary histological abnormality in the chick arises 34 h
after treatment, with cell junction breakdown in the periderm as
an initial event. Apoptotic changes in the ectoderm, neural tube,
lateral plate mesoderm and particularly the somites occur within
48 h after Cd. As somite development is dependent on normal Wnt
signaling from the ectoderm, we have been working on the hypothesis that disruption of peri-ectodermal tissue can interrupt this
signaling pathway. The molecule -catenin is pivotal in this respect,
having roles to play in both the canonical Wnt pathway and being
an intracellular associate of cadherin cellular adhesion molecules.
We propose that breakdown of adherens and desmosomal junctions releases -catenin into the cytosol with subsequent nuclear
translocation, a theory borne out by recent immunohistochemical
studies in our laboratory [104]. We have seen hyperproliferation
in the peri-ectodermal tissue within this timeframe [102], which
could be a local effect of -catenin-mediated transcription of cyclin
D1 or other cell cycle promoters. Therefore, it is feasible that somitic
cell death and delay in differentiation could result from the Wntlike signaling effect of released -catenin within the periderm and
ectoderm, causing an imbalance in gene expression required for
normal development of the somites. Further studies are in progress
to examine these possibilities.
In rodent models, axial anomalies appear to be less remarkable,
but there are clear effects on neural tube closure, limb development
and soft tissue formation, depending on the timing of administra-

J. Thompson, J. Bannigan / Reproductive Toxicology 25 (2008) 304315

tion of Cd. Webster and Messerle [90] found that IP injection of

4 mg/kg CdCl2 in the mouse produced neural tube defects when
given on day 7 or 8, that is to say, during neurulation. When Cd was
given on day 9 or 10, exencephaly did not occur, but the neural tube
re-opened secondary to marked cell death. This phenomenon was
observed also by Schmid et al. [105] in mouse embryos cultured
in vitro in the presence of Cd. Re-opening of the neural tube may
occur when a critical level of cell necrosis in the wall of the neural tube is exceeded [106]. Mice treated with 4 mg/kg CdCl2 shortly
after neurulation developed limb reduction abnormalities [91]. Cd
administered IP to pregnant CD-1 mice on days 912 gestation
induced postaxial forelimb defects in the offspring, indistinguishable from those produced by acetazolamide. This suggests a role for
the enzyme carbonic anhydrase in the mechanism of teratogenesis
[94].
In the hamster, Gale and Layton [107] injected a single dose
2 mg/kg CdSO4 intravenously on day 8 of gestation, and found a
multiplicity of craniofacial, skeletal and soft tissue abnormalities
upon examination of the fetuses on day 15. More recent studies
using whole-embryo culture assay for hamster embryos reported
70% abnormality rate in embryos cultured at 0.25 M Cd on day
8 of gestation, with animals exhibiting growth retardation, reduction in somite gain, axial anomaly and neural tube defect [108]. In
a classic study, Ferm [109] administered Cd at multiple stages of
embryonic development in the golden hamster, treating animals
either early on day 8 of gestation, corresponding to the primitive
streak stage, late on day 8, early on day 9 (when the neural tube
has closed) or on the afternoon of day 9. The earliest treatment
group had the highest rate of embryolethality, and exencephaly
and eye defects were more frequent in this group. Rib and upper
limb defects were most frequent in those treated late on day 8,
whereas later groups had a preponderance of lower limb defects,
reecting the time-lag between upper and lower limb outgrowth
and development.
Sequential administration of Cd to pregnant rats yielded comparable time-related defects. Samarawickrama and Webb [110] found
that hydrocephalus and eye defects occurred most frequently in
pups of WistarPorton rats that received 1.25 mg Cd/kg body weight
on days 9 and 10 of gestation. In the post-neurulation period,
Holt and Webb [96] found that administration of Cd on gestation
days 10, 12 or 14 increased the incidence of skeletal retardation,
hydrocephalus, urogenital abnormalities, cleft palate, diaphragmatic hernia, and cardiovascular anomalies. Umbilical hernia and
hydronephrosis did not occur before day 11 [110] and renal agenesis
occurred exclusively within a narrow time window between days
9 and 11 of gestation.
Although Cd has yet to be shown to be teratogenic in humans, an
inverse relationship has been found between birth weight and cord
blood Cd levels [111], birth weight and placental Cd concentration
[112], and between birth length and maternal blood Cd [113]. There
is, however, an association between maternal Cd exposure and early
delivery, which may explain the association with smaller babies
[114]. An inverse relationship has also been demonstrated between
birthweight and Cd content in newborn hair [115,116].
Cd blood levels in newborn infants have been found to be 70%
of maternal levels [117]. The difference in Cd level can probably
be ascribed to placental function, as perfusion of human placentas
with Cd [118] showed that the transfer rate from maternal to fetal
side was very slow. The metal did not appear on the fetal side of the
placenta until 40 min after perfusion started, and reached a steady
state approximately 1 h later, at which time the concentration in
the fetal perfusate was 1/20th that of the maternal. Microstructural
changes, including edema and vacuolation followed by necrosis,
were detected in placentas between 5 and 8 h after perfusion with
Cd [85].

309

Postnatal effects described in association with exposure to

abnormally high levels of Cd in utero include signicantly increased
DNA synthesis in the bowel and bone marrow in 6-week old rats
following administration of Cd orally to pregnant dams at a dose
of 50 ppm in drinking water [119], and depressed immune function
and sensorimotor abnormalities in animal studies and in children
following prenatal exposure to Cd [120]. However, human studies
to date are not conclusive due to confounding factors and problems
in study design, and additional work needs to be done to elucidate
the extent and type of damage to children following exposure. This
is of particular relevance because of altered pharmacokinetics of
metals in very young children, in whom absorption is more avid,
excretion less efcient, and the bloodbrain barrier less effective
[120].
3. Mechanisms of reproductive toxicity
A number of mechanisms of Cd toxicity have been suggested,
including ionic and molecular mimicry, interference with cell adhesion and signaling, oxidative stress, apoptosis, genotoxicity and cell
cycle disturbance. Although the overall effect of Cd on any cell or
tissue is likely to be due to a synergism of several mechanisms, it
is possible that one mechanism will predominate in a specic cell
type.
3.1. Altered cell adhesion
Normal cell adhesion plays a role in driving meiosis and differentiation in spermatogenesis [37], oocyte maturation, formation
of the OCC, pick-up of the oocyte by the cilia on the infundibulum of the Fallopian tube [74,75], compaction of the early embryo
and development to the blastocyst stage [68,69]. Cell division, morphogenetic movements and apoptosis in the post-implantation
embryo are also dependent on cellcell contact and intercellular
communication [121]. The interference by Cd with cell adhesion
is complex, and is probably modied by the many other cellular
effects of this metal. Desmosomal disruption in the testis secondary to Cd administration has been documented by Berliner
and Jones-Witters [28] and Fende and Niewenhuis [26]. Functional
adherens junctions have also been identied in the testis [122],
although the inter-relationship between adherens junctions, tight
junctions and desmosomes is very different to that described in
other epithelia [123]. The integrity of the bloodtestis barrier (BTB)
is dependent on tight junctions between Sertoli cells located deep
within the wall of the seminiferous tubule near the basal lamina, creating separate microenvironments for the development of
pre-leptotene/leptotene spermatocytes on the basal aspect, and
pachytene spermatocytes/spermatids on the luminal side. Wong
et al. [36] have demonstrated the ability of Cd to disrupt the
BTB via the TGF3/p38 pathway, with secondary loss of function
at adherens junctions. Assembly and disassembly of inter-sertoli
junctions is essential for normal migration of germ cells across
the seminiferous epithelium and their concomitant differentiation
[124]. Similarly, maintenance of a specic microenvironment in
the epididymis is crucial to the further maturation of spermatozoa [125]. Intact mechanisms for cell adhesion are also essential
for normal oogenesis, OCC expansion and oocyte pick-up by the
Fallopian tube, as outlined in Sections 2.2 and 2.4.
In the pre-implantation human embryo, gap junctions, expressing predominantly connexin-43, have been detected from the
four-cell stage, becoming increasingly organized as development
proceeds [68,126]. Construction of gap junctions in vertebrates
depends on a family of transmembrane proteins called connexins
[127]. Gap junction intercellular communication (GJIC), demon-

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J. Thompson, J. Bannigan / Reproductive Toxicology 25 (2008) 304315

strated by Jeong et al. [66] to be inhibited by Cd, is of utmost

importance in transmission of small molecules and electrical
impulses from cell to adjacent cell. Gap junction formation is heralded by the expression of various connexins, many of which have
short lives of a few hours or less. Cd treatment impedes gap junction
assembly by interfering with phosphorylation of certain connexins, thus preventing clonal expansion within the early embryo [67].
It also produces a decrease in expression of connexins 26 and 32
[66]. Interestingly, gap junctions co-localize with cadherins, and
connexin-43 is regulated by -catenin, already implicated as a possible player in Cd teratogenesis in the post-implantation embryo
[104]. This again raises the possibility that -catenin released
upon disassociation of cadherin complexes could translocate into
the nucleus and alter gene expression, in this case, connexin-43
expression. In the murine model, -catenin accumulates in the
cytoplasm between the two-cell and morula stages of development, but no evidence of concomitant nuclear staining has been
observed [128]. Reactivation occurs between late morula and early
blastocyst stages, a time when gap junction assembly is essential for compaction to occur, and deactivation occurs again when
the fully expanded blastocysts hatch from the zona pellucida. Cd,
through its action on cadherins and other adhesion molecules, can
theoretically interfere with this tightly regulated activation of catenin. Furthermore, tight junctions, another target for Cd [77],
are known to be critical for blastocyst formation in mammalian
embryos [129,130], in terms of establishing and maintaining watertight barriers within the formed blastocyst. Desmosomal junctions
have also been detected in the pre-implantation human embryo
at the blastocyst stage [68,131], and are probably vulnerable to Cd
interference, although no experimental evidence of this has been
presented to date.
An interesting possible role for cell adhesion molecules is
their role in signaling, remodeling and migration in the postimplantation embryo. In our previous work with Cd, we have noted
a strong relationship between breakdown in cell adhesion in the
periderm and failure of the lateral body folds to migrate ventrally in the post-gastrulation chick embryo [101,102], resulting
in a large ventral body wall defect. Preliminary work in our laboratory has indicated that there is no change in distribution of
cadherins in chicks with extensive peridermal desquamation following Cd, but associated molecules such as -catenin and actin are
profoundly affected, with marked clumping of actin and movement
of -catenin from its predominantly peripheral location adjacent to
cellular junctions to a more generalized cytosolic distribution and,
more importantly, into the nuclei of the periderm cells, indicating a distribution of -catenin equivalent to that seen in activation
of the canonical Wnt pathway [104]. Tight, adherens, and gap
junctions, and desmosomes have been demonstrated in the periderm in developing humans and rats [132135], and any or all of
these molecules could be a target for interference by Cd, but a link
between Cd-related junction disruption and aberrant signaling in
the later embryo has yet to be conrmed.
3.2. Oxidative stress and apoptosis
The role of Cd in inducing oxidative stress in adult tissue
has been well documented. Increased levels of malondialdehyde,
protein oxidation and reduced activity of superoxide dismutase
(SOD) in rat testes have been reported following a single SC
injection of 2 mg/kg CdCl2 [136], indicating an increase in lipid
peroxidation and a decrease in the removal of superoxide radicals. Adequate Zn supplementation in the diet partially reversed
these changes. Amara et al. [137] attributed decreased testicular
growth rate, plasma testosterone, and reduced sperm count and
motility to Cd-induced oxidative stress, as concurrent reduction

in glutathione peroxidase, catalase, mitochondrial MnSOD, and

cytosolic CuZnSOD, along with increased MT and malondialdehyde, were observed. Vitamins C and E can protect the rat testis from
Cd-induced oxidative damage [138,139], and selenium [140142],
n-acetylcysteine [139], glycine [143], and glutathione-like compounds [144] have also been shown to rescue various cells and
tissues from this mechanism.
The pro-apoptotic transcription factor p53 is activated by oxidative stress, which aids in the generation of reduced glutathione
and expression of sestrins, thus decreasing reactive oxygen species
(ROS) and protecting the organism from resultant DNA damage
[145]. However, although Cd induces abnormal levels of apoptosis
in the testis, it does so via a p53-independent pathway [146] and
in fact has been associated with a suppression of p53 expression.
Conversely, similar doses produce apoptosis in the ventral prostate
which is clearly p53-dependent [147]. In general, mechanisms by
which Cd provokes apoptosis depend upon the dose given [148] and
the cell type exposed [149]. Activation of p38 and disruption of the
mitochondrial membrane appear to be the primary targets. However, Cd can also initiate apoptosis by intrinsic pathway activation,
and at higher Cd exposure, necrosis may supervene.
In view of the above, it is surprising that the literature lacks
reports of experiments to detect oxidative stress in the whole
embryo. So far, in our laboratory, efforts to detect oxidative stress in
chick embryos have yielded ambiguous results. Similarly, we have
failed to show a blockade to the teratogenic effects of Cd by antioxidants. Our preliminary results [150] have shown that selenium
(Se) in doses of twice the molarity of Cd confers 100% protection
against the teratogenic effects of Cd in the chick. Although Se is
a known antioxidant [141,142], its protection against Cd may lie
more simply in its ability, as a divalent cation, to block Cd uptake
by embryonic cells.
3.3. Ionic and molecular mimicry
Divalent cations, many of which are nutritive metals, have
demonstrably protected embryonic and adult tissue from Cdinduced damage, indicating a mechanistic link [101,151,152]. The
ability of Cd to imitate these nutrients in many biological pathways and processes is a very probable explanation for its toxicity.
Experimental evidence indicates that Cd may interact with membrane transporters involved in the uptake of Ca, iron (Fe) and
Zn through a process of ionic mimicry [153,154]. Transporters
for other metals such as manganese (Mn) have also been implicated in Cd handling [153,155,156]. Ca channels are thought to be
involved in Cd uptake in certain tissues, due to the similarity of
ionic radii between Cd and Ca [157], and also because experiments
with certain cell types demonstrated that the Ca channel antagonists nifedipine and verapamil signicantly blocked the uptake of
Cd [63,158]. It has been estimated that between 30 and 50% of Cd
enters cells through Ca channels [158,159]. As well as interfering
with cellular transport processes, Cd has also been found to form
covalent complexes with sulfhydryl-containing biomolecules such
as cysteine and glutathione (GSH), thus altering their behavior. It
has also been proposed that Cd may substitute for nutritive metals
in various enzymes and cell adhesion molecules [95,160166].
Several studies indicate that Cd gains access to testicular cells
by mimicking Zn at the site of Zn-transporters [167169]. Passive
and active mechanisms are involved in testicular transport of Cd.
Addition of Zn can signicantly inhibit the active component of Cd
uptake [170], thus providing a mechanism by which Zn alleviates
Cd toxicity in the testis. About 20% of Zn uptake by placental tissue
is susceptible to competitive inhibition by Cd [171]. In addition
to interference with Zn uptake, the importance of the presence of
Cd within the cell becomes clear when considering the potential

J. Thompson, J. Bannigan / Reproductive Toxicology 25 (2008) 304315

functions of Zn. Substitution of Cd for Zn in carbonic anhydrase

may result in the inability to catalyze the hydration of CO2 to
bicarbonate, thus creating an acidotic embryonic environment
with adverse reproductive consequences [95]. Other enzymes
from which Cd could potentially displace Zn include alcohol dehydrogenase, lactic dehydrogenase, malic dehydrogenase, alkaline
phosphatase and carboxypeptidase [160,161]. Sunderman [172]
proposed that the teratogenic effect is mediated by Cd substitution
for Zn in nger-loop domains of certain DNA-binding proteins that
regulate gene expression during embryogenesis. Experiments by
Predki and Sarkar [173] conrmed that certain Zn-nger residues
have a higher afnity for Cd than for Zn.
The effects of Cd on Ca-regulated pathways are so diverse
that a comprehensive account is outside the scope of this article. Important potential effects include replacement of Ca by Cd
in the extracellular ligands of cadherin molecules [162], in desmosome and gap junction assembly [163165], and in integrin function
[166]. Mobilization of Ca from internal stores can be induced by
extracellular Cd via the phosphatidylinositol pathway [174,175],
upregulating the activity of Ca-dependent intracellular pathways
and causing abnormal cell proliferation or cell death [175]. Intracellular Ca2+ is maintained at a low level (0.1 M) due to the presence
of Ca2+ pumps that export Ca2+ from the unstimulated cell. Activation of the phosphatidylinositol pathway mobilizes intracellular
Ca2+ from the endoplasmic reticulum by binding to receptors that
are ligand-gated Ca channels, thus increasing intracellular Ca2+ to
about 1 M. This, in turn, alters the activities of target proteins,
many of which are enzymes [176]. Many of the effects of intracellular Ca2+ are mediated by the Ca-binding protein calmodulin,
which is generally activated when cytosolic Ca2+ increases to above
0.5 M, and is also known to be activated by Cd [121,177]. Therefore,
several aspects of Ca movement and action, of vital importance in
fertilization and subsequent embryo development, may be inuenced by Cd.
Metal homeostasis has to be carefully regulated by the cell
to prevent production of toxic free radicals [178], thus triggering
oxidative stress. Induction of metallothionein (MT) [179] by Zn and
possibly other metals is an alternative mechanism by which protection against Cd toxicity could be conferred. This low-molecular
weight protein binds to Cd, limits its availability to cells and tissues
[82,180], and plays a role in transport, detoxication, and storage
[181]. A possible role for MT in the oxidative stress reaction has also
been documented [182]. It has been shown to be an effective free
radical scavenger, important because the release of various species
of oxygen metabolites is thought to be indirectly responsible for the
initiation of apoptosis. However, MT has itself been implicated as
a causal factor in apoptosis [180], and has intrinsically toxic effects
in itself when bound to Cd [183186].
3.4. DNA damage and effect on the cell cycle
Cd has been associated with increased chromosomal aberrations and decreased mitotic index in Chinese Hamster Ovary (CHO)
cells [187], and non-disjunction in germ cells [188]. Inhibition of
DNA repair [189] is thought to be an important mechanism by
which Cd exerts its mutagenic effect in this cell type. Resistance to
the toxic effects of Cd in CHO cells has been linked to the ability to
synthesize metallothionein (MT)-1 protein and MT-1 gene amplication [190]. Administration of the ROS scavenger D-Mannitol was
found to protect CHO cells against the mutagenicity of Cd, whereas
the catalase inhibitor 3-amino-1,2,4-triazole potentiated this effect
[191]. Increased mutation frequency after Cd was found to be associated with reduced activity of anti-oxidant enzymes, among them
GSH peroxidase and catalase. This latter evidence indicates that
the mutagenic effect of Cd is consequent upon its ability to gen-

311

erate oxidative stress within the tissues. A dose-dependent effect

was seen on the cell cycle in this culture system, with retardation
of cell cycle progression of cells cultured with 1 M Cd or more for
24 h [192], G2/M arrest in cells treated with lower doses (1 M) and
S-phase arrest at higher doses (4 M).

4. Conclusions
Cd has the potential to affect reproduction and development in
many different ways, and at every stage of the reproductive process. Effects on the testis include disruption of the bloodtestis
barrier due to adverse effects on cell adhesion, oxidative stress,
and necrosis at higher experimental doses. Incorporation into chromatin of the developing spermatozoa has also been described.
In the ovary, oocyte development is inhibited, steroidogenesis
reduced, and ovarian hemorrhage and necrosis supervene at higher
Cd doses. Cumulus expansion and possibly oocyte pick-up by the
tubal epithelium are also hampered, probably by interference with
normal junction formation. Although the early pre-implantation
embryo seems relatively resistant to Cd, increasing incorporation
from the four-cell stage onwards predisposes to apoptosis in the
blastocyst and implantation failure. Injurious effects on the placenta, including inhibition of trophoblastic invasion, decreased
steroidogenesis, and adjusted handling of nutritive metals, have
also been identied.
The clinical implications of the effects of Cd on oocyte maturation, oocyte pick-up and development of the pre-implantation
embryo through to implantation are obvious. An important source
of this toxin in humans is inhalation through cigarette smoking. Cd
thus absorbed has a relatively high bioavailability, and is readily
assimilated into human tissues and transported via the bloodstream to more remote parts of the body, notably the reproductive
tract. Cigarette smoking has been associated with subfertility in
a number of studies [193196]. Increased blood levels of Cd have
been found in smokers [197], with levels correlating signicantly
with numbers of cigarettes consumed. The concentration of Cd in
follicular uid of female smokers undergoing in vitro fertilization
[IVF] has been reported at 7.93 0.16 ng/ml [198] compared with
6.73 0.31 ng/ml in non-smokers. The concentration of Cd in
human ovaries increases over time [199] with a linear increase
with age in females between 30 and 65 years. Smokers were found
to have increased ovarian Cd levels when compared with nonsmokers. Human ovarian granulosa cells exposed to Cd exhibited
morphological alterations, breakdown of intercellular junctions
and defective steroidogenesis. However, the precise mechanism by
which Cd impedes oocyte development and granulosa cell function
has yet to be identied, in particular at concentrations comparable
to those found in the follicular uid of smokers, and manipulations
by which this damage might be prevented or reversed should be
identied in vitro and in vivo. The role of oxidative stress and the
possibility of preventing or reversing this effect in particular needs
to be explored. Cigarette smoking has also been identied as a risk
factor in ectopic pregnancy in several studies [200,201], possibly
reecting faulty movement of the oocyte through the uterine
tubes, a process which is highly dependent on sequential cell
junction formation. To date, the precise role of Cd in this sequence
of events has not been dened. However, recurrent miscarriage
and uterine broids have been directly linked with increased
urinary Cd excretion in human females [202].
The effects of Cd on the post-implantation embryo are stageand dose-dependent. Effects caused by administration in the
peri-gastrulation phase are myriad, with growth retardation, craniofacial defects, cardiovascular malformations, gut anomalies and
body axial deformities reported in many of the species investigated.

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Neural tube defects could be produced by giving Cd before neurulation was complete, causing failure of fusion of the neural folds, or as
a secondary phenomenon due to cell death in the neural tube. Limb
and body wall defects tended to occur with Cd administration in the
post-neurulation period. Mechanistic studies to date have indicated
that teratogenesis can be linked to substitution of Cd for predominantly Zn or Ca in many biological processes, thus inactivating
cellular adhesion and modifying in some way a wide selection of
intracellular reactions, giving rise to oxidative stress and abnormal
cell proliferation and apoptosis. This in turn opens the possibility of
the involvement of signaling pathways, a line of inquiry currently
being pursued in our laboratory, and the opportunity to look at
means by which we might protect the developing embryo by signal modication, prevention of Cd uptake, competitive inhibition
of intracellular Cd, or use of anti-oxidants.
While the effects on gametogenesis, implantation and early
embryo development seem to depend largely on impaired cell
adhesion, other, less direct, effects of Cd can produce sub-optimal
conditions for a safe pregnancy outcome, including iron and zinc
deciency in the fetus [85,203] and a possible association with the
pathogenesis of pre-eclampsia [204]. In addition, pre-term delivery
and cesarian section delivery were found to occur approximately
four times more frequently in women with higher Cd burdens (urinary Cd 2 g/g creatinine) than in their counterparts with lower
Cd burdens [13]. Further experimental work in animal models and
carefully designed case-controlled studies in humans are required
to elucidate the consequences of Cd exposure on late pregnancy
complications, neonatal health and long-term postnatal outcome.
Whatever the effect, it is certainly prudent to recommend cessation
of smoking in pregnancy and in the pre-conception period, and supplementation with micronutrients such as Zn, Fe, Se and vitamin C
should be considered in ex-smokers having difculty conceiving.
References
[1] WHO. Cadmium (Environmental Health Criteria No. 134). Geneva: WHO;
1992.
[2] Muntau H, Baudo R. Sources of cadmium, its distribution and turnover in the
freshwater environment. IARC 1992;118:13348.
[3] IARC. IARC monographs on the evaluation of carcinogenic risks to humans,
vol. 58. Beryllium, Cd, mercury and exposures in the glass manufacturing
industry. Lyons, France: IARC; February 916, 1993.
[4] Martelli A, Rousselet E, Dycke C, Bouron A, Moulis J-M. Cadmium toxicity in animal cells by interference with essential metals. Biochimie
2006;88:180714.
[5] WHO. Cadmiumenvironmental aspects (Environmental Health Criteria
135). Geneva: WHO; 1992.
[6] IRPTC. IRPTC legal le 1986, vol. 1. Geneva: International Register of Potentially Toxic Chemicals, United Nations Environment Programme; 1987.
[7] McLellan JS, Flanagan PR, Chamberlain MJ, Valberg LS. Measurement
of dietary cadmium absorption in humans. J Toxicol Environ Health
1978;4:1318.
[8] Nasreddine L, Parent-Massin D. Food contamination by metals and pesticides
in the European Union. Should we worry? Toxicol Lett 2002;127:2941.
[9] Watanabe T, Koizumi A, Fujita H, Kumai M, Ikeda M. Role of rice in dietary
cadmium intake of farming population with no known man-made pollution
in Japan. Tohoku J Exp Med 1984;144:8390.
[10] Vahter M, Berglund M, Slorach S, Jorhem L, Lind B. Integrated personal monitoring of cadmium exposure in Sweden. IARC 1992;118:11320.
[11] Jarup L, Berglund M, Elinder CG, Nordberg G, Vahter M. Health effects of cadmium exposure-a review of the literature and a risk estimate. Scand J Work
Environ Health 1998;24(Suppl 1):151.
[12] McKenzie-Parnell JM, Kjellstrom TE, Sharma RP, Robinson MF. Unusually high
intake and fecal output of cadmium, and fecal output of other trace elements
in New Zealand adults consuming dredge oysters. Environ Res 1988;46:114.
[13] Satarug S, Moore MR. Adverse health effects of chronic exposure to lowlevel cadmium in foodstuffs and cigarette smoke. Environ Health Perspect
2004;112:1099103.
[14] Nandi M, Slone D, Jick H, Shapiro S, Lewis GP. Cadmium content of cigarettes.
Lancet 1969;2:132930.
[15] Lewis GP, Coughlin LL, Jusko WJ, Hartz S. Contribution of cigarette smoking
to cadmium accumulation in man. Lancet 1972;1(7745):2912.
[16] Friberg L, Piscator M, Nordberg GF, Kjellstrom T. Cadmium in the environment.
2nd ed. Cleveland, OH: Chemical Rubber Co.; 1974.

[17] Elinder CG, Kjellstrom T, Lind B, Linnman L, Piscator M, Sundstedt K. Cadmium

exposure from smoking cigarettes: variations with time and country where
purchased. Environ Res 1983;32:2207.
[18] Jarup L, Rogenfelt A, Elinder CG, Nogawa K, Kjellstrom T. Biological half-time
of cadmium in the blood of workers after cessation of exposure. Scand J Work
Environ Health 1983;9:32731.
[19] Matsuno K, Kodama Y, Tsuchiya K. Biological half-time and body burden of
cadmium in dogs after a long-term oral administration of cadmium. Biol Trace
Elem Res 1991;29:11123.
[20] Nordberg M, Jin T, Nordberg GF. Cadmium, metallothionein and renal tubular
toxicity. IARC 1992;118:2937.
[21] El-Ashmawy IM, Youssef SA. The antagonistic effect of chlorpromazine on
cadmium toxicity. Toxicol Appl Pharmacol 1999;161:349.
[22] Sauer JM, Waalkes MP, Hooser SB, Kuester RK, McQueen CA, Sipes IG. Suppression of Kupffer cell function prevents cadmium induced hepatocellular
necrosis in the male SpragueDawley rat. Toxicology 1997;121:15564.
[23] Tam PPL, Liu WK. Gonadal development and fertility of mice treated prenatally with cadmium during the early organogenesis stages. Teratology
1985;32(3):45362.
[24] Toman R, Massanyi P, Uhrin V. Changes in the testis and epididymis of rabbits
after an intraperitoneal and peroral administration of cadmium. Trace Elem
Electrolytes 2002;19(3):1147.
[25] Kotsonis FN, Klaassen CD. Toxicity and distribution of cadmium administered
to rats at sublethal doses. Toxicol Appl Pharmacol 1977;41:66780.
[26] Fende PL, Niewenhuis RJ. An electron microscopic study of the effects of cadmium chloride on cryptorchid testes of the rat. Biol Reprod 1977;16:298
305.
[27] Niewenhuis RJ. Effects of cadmium upon regenerated testicular vessels in the
rat. Biol Reprod 1980;23:1719.
[28] Berliner AF, Jones-Witters P. Early effects of a lethal cadmium dose on gerbil
testis. Biol Reprod 1975;13:2407.
[29] Laskey JW, Rehnberg GL, Laws SC, Hein JF. Reproductive effects of low
acute doses of cadmium chloride in adult male rats. Toxicol Appl Pharmacol
1984;73(2):2505.
[30] Jin LJ, Lou ZF, Dong JY. Effects of cadmium and mercury on DNA damage of mice
bone marrow cell and testicle germ cell in vitro. Carcinog Teratog Mutagen
2004;16(2):947.
[31] Yang HS, Han DK, Kim JR, Sim JC. Effects of -tocopherol on cadmium-induced
toxicity in rat testis and spermatogenesis. J Korean Med Sci 2006;21:44551.
[32] Aoyagi T, Ishikawa H, Miyaji K, Hayakawa K, Hata M. Cadmium-induced testicular damage in a rat model of sub-chronic intoxication. Reprod Med Biol
2002;1:5963.
[33] Bench G, Corzett MH, Martinelli R, Balhorn R. Cadmium concentrations in
the testes, sperm, and spermatids of mice subjected to long-term cadmium
chloride exposure. Cytometry 1999;35:306.
[34] Dixon RL, Lee IP, Sherins RJ. Methods to assess reproductive effects of environmental chemicals: studies of cadmium and boron administered orally.
Environ Health Perspect 1976;13:5967.
[35] Chung NPY, Cheng CY. Is cadmium chloride-induced inter-sertoli tight junction permeability barrier disruption a suitable in vitro model to study the
events of junction disassembly during spermatogenesis in the rat testis?
Endocrinology 2001;142(5):187888.
[36] Wong C, Mruk DD, Lui W, Cheng CY. Regulation of bloodtestis barrier dynamics: an in vivo study. J Cell Sci 2004;117:78398.
[37] Yan HHN, Cheng CY. Bloodtestis barrier dynamics are regulated by an
engagement/disengagement mechanism between tight and adherens junctions via peripheral adaptors. PNAS 2006;102(33):117227.
[38] Akinloye O, Arowojolu AO, Shittu OB, Anetor JI. Cadmium toxicity: a possible
cause of male infertility in Nigeria. Reprod Biol 2006;6(1):1730.
[39] Saaranen M, Kantola M, Saarikoski S, Vanha-Perttula T. Human seminal
plasma cadmium: comparison with fertility and smoking habits. Andrologia
1989;21(2):1405.
[40] Al-Bader A, Omu AE, Dashti H. Chronic cadmium toxicity to sperm
of heavy cigarette smokers: immunomodulation by zinc. Arch Androl
1999;43(2):13540.
[41] Leoni G, Bogliolo L, Deiana G, Berlinquer F, Rosati I, Pintus PP, et al. Inuence
of cadmium exposure on in vitro ovine gamete dysfunction. Reprod Toxicol
2002;16(4):3717.
[42] Waalkes MP, Rehm S, Perantoni AO, Coogan TP. Cadmium exposure in rats and
tumours of the prostate. IARC 1992;118:391400.
[43] West DW, Slattery ML, Robison LM, French TK, Mahoney AW. Adult dietary
intake and prostate cancer risk in Utah: a casecontrol study with special
emphasis on aggressive tumors. Cancer Causes Control 1991;2:8594.
[44] Kazantzis G, Lam TH, Sullivan KR. Mortality of cadmium-exposed workers.
Scand J Work Environ Health 1988;14:2203.
[45] Sorahan T, Lister A, Gilthorpe MS, Harrington JM. Mortality of copper
cadmium alloy workers with special reference to lung cancer and nonmalignant diseases of the respiratory system, 194692. Occup Environ Med
1995;52:80412.
[46] Waalkes MP, Anver M, Diwan BA. Carcinogenic effects of cadmium in the
Noble (NBL/Cr) rat: induction of pituitary, testicular, and injection site tumors
and intraepithelial proliferative lesions of the dorsolateral prostate. Toxicol
Sci 1999;52:15461.
[47] Waalkes MP, Anver MR, Diwan BA. Chronic toxic and carcinogenic effects of
oral cadmium in the Noble (NBL/Cr) rat: induction of neoplastic and prolif-

J. Thompson, J. Bannigan / Reproductive Toxicology 25 (2008) 304315

[48]

[49]

[50]
[51]

[52]
[53]

[54]

[55]

[56]
[57]

[58]

[59]

[60]

[61]

[62]

[63]

[64]

[65]

[66]
[67]

[68]

[69]
[70]
[71]

[72]
[73]

[74]

[75]
[76]

[77]

erative lesions of the adrenal, kidney, prostate, and testes. J Toxicol Environ
Health A 1999;58(4):199214.
Fort DJ, Stover EL, Bantle JA, Dumont JN, Finch RA. Evaluation of a reproductive
toxicity assay using Xenopus laevis: boric acid, cadmium and ethylene glycol
monomethyl ether. J Appl Toxicol 2001;21:4152.
Fort DJ, McLaughlin DW, Rogers RL, Buzzard BO. Effect of endocrine disrupting
chemicals on germinal vesicle breakdown in xenopus in vitro. Drug Chem
Toxicol 2002;25(3):293308.
Lienesch LA, Dumont JN, Bantle JA. The effect of cadmium on oogenesis in
Xenopus laevis. Chemosphere 2000;41(10):16518.
Saksena SK, Salmonsen R. Effects of cadmium chloride on ovulation
and on induction of sterility in the female golden hamster. Biol Reprod
1983;29:24956.
Rehm S, Waalkes MP. Cadmium-induced ovarian toxicity in hamsters, mice
and rats. Toxicol Sci 1988;10(4):63547.
Paksy K, Varga B, Horvath E, Tatrai E, Ungvary G. Acute effects of cadmium on preovulatory serum FSH, LH, and prolactin levels and on ovulation
and ovarian hormone secretion in estrous rats. Reprod Toxicol 1989;3(4):
2417.
Paksy K, Varga B, Lazar P. Zinc protection against cadmium-induced infertility
in female rats. Effect of zinc and cadmium on the progesterone production of
cultured granulosa cells. Biometals 1996;10:2736.
Paksy K, Varga B, Naray M, Olajos F, Folly G. Altered ovarian progesterone
secretion induced by cadmium fails to interfere with embryo transport in the
oviduct of the rat. Reprod Toxicol 1992;6(1):7783.
Piasek M, Laskey JW. Acute cadmium exposure and ovarian steroidogenesis
in cycling and pregnant rats. Reprod Toxicol 1994;8(6):495507.
Mlynarcikova A, Fickova M, Scsukova S. Ovarian intrafollicular processes as
a target for cigarette smoke components and selected environmental reproductive disruptors. Endocr Regul 2005;39:2031.
Vrsanska S, Nagyova E, Mlynarcikova A, Fickova M, Kolena J. Components
of cigarette smoke inhibit expansion of oocytecumulus complexes from
porcine oocytes. Physiol Res 2003;52:3837.
Piasek M, Laskey JW, Kostial K, Blanusa M. Assessment of steroid disruption
using cultures of whole ovary and/or placenta in rat and in human placental
tissue. Int Arch Occup Environ Health 2002;75(Suppl):S3644.
Paksy K, Rajczy K, Forgacs Z, Lazar P, Bernard A, Gati I, et al. Effect of cadmium
on morphology and steroidogenesis of cultured human ovarian granulosa
cells. J Appl Toxicol 1997;17(5):3217.
Schmid BP, Hall JL, Goulding E, Fabro S, Dixon R. In vitro exposure of
male and female mice gametes to cadmium chloride during the fertilization process, and its effects on pregnancy outcome. Toxicol Appl Pharmacol
1983;69(3):32632.
Storeng R, Jonsen J. Effect of nickel chloride and cadmium acetate on
the development of pre-implantation mouse embryos in vitro. Toxicology
1980;17(2):1837.
De SK, Paria BC, Dey SK, Andrews GK. Stage-specic effects of cadmium on
pre-implantation embryo development and implantation in the mouse. Toxicology 1993;80(1):1325.
Yu HS, Tam PPL, Chan STH. Effects of cadmium on pre-implantation mouse
embryos in vitro with special reference to their implantation capacity and
subsequent development. Teratology 1985;32(3):34753.
Abraham R, Charles AK, Mankes R, LeFevre R, Renak V, Ashok L. In vitro effects
of cadmium chloride on preimplantation rat embryos. Ecotoxicol Environ Saf
1986;12(3):2139.
Jeong SH, Habeebu SS, Klaassen CD. Cadmium decreases gap junctional intercellular communication in mouse liver. Toxicol Sci 2000;57(1):15666.
Fang MZ, Mar WC, Cho MH. Cadmium-induced alterations of connexin expression in the promotion stage of in vitro two-stage transformation. Toxicology
2001;161(1/2):11727.
Hardy K, Warner A, Winston RML, Becker DL. Expression of intercellular junctions during implantation development of the human embryo. Mol Human
Reprod 1996;2(8):62132.
Kidder GM, Winterhager E. Intercellular communication in preimplantation
development: the role of gap junctions. Front Biosci 2001;6:D7316.
Yu HS, Chan ST. Zinc amelioration of cadmium toxicity on preimplantation
mouse zygotes in vitro. Teratology 1988;37(1):139.
Peters JM, Duncan JR, Wiley LM, Keen CL. Inuence of antioxidants on
cadmium toxicity of mouse preimplantation embryos in vitro. Toxicology
1995;99(1/2):118.
Larsen W. Human embryology. 2nd ed. WB Saunders Company; 1997.
Talbot P, Shur BD, Myles DG. Cell adhesion and fertilization: steps in oocyte
transport, sperm-zona pellucida interactions, and spermegg fusion. Biol
Reprod 2003;68:19.
Chen L, Zhang H, Powers RW, Russell PT, Larsen WJ. Covalent linkage between
proteins of the inter--inhibitor family and hyaluronic acid is mediated by a
factor produced by Granulose cells. J Biol Chem 1996;271:1940914.
Talbot P, Geiske C, Knoll M. Oocyte pick-up by the mammalian oviduct. Mol
Biol Cell 1999;10:58.
Chen L, Wert SE, Hendrix EM, Russell PT, Cannon M, Larsen WJ. Hyaluronic acid
synthesis and gap junction endocytosis are necessary for normal expansion
of the cumulus mass. Mol Reprod Dev 1990;26:23647.
Fiorini C, Tilloy-Ellul A, Chevalier S, Charuel C, Pointis G. Sertoli cell junctional
proteins as early targets for different classes of reproductive toxicants. Reprod
Toxicol 2004;18:41321.

313

[78] Yu HS, Chan ST. Effects of cadmium on preimplantation and early postimplantation mouse embryos in vitro with special reference to their trophoblastic
invasiveness. Pharmacol Toxicol 1987;60(2):12934.
[79] Di SantAgnese PA, Jensen KD, Levin A, Miller RK. Placental toxicity of cadmium in the rat: an ultrastructural study. Placenta 1983;4(2):14963.
[80] Powlin SS, Keng PC, Miller RK. Toxicity of cadmium in human trophoblast
cells (Jar choriocarcinoma): role of calmodulin and the calmodulin inhibitor,
zaldaride maleate. Toxicol Appl Pharmacol 1997;144:22534.
[81] Lin FJ, Fitzpatrick JW, Iannotti CA, Martin DS, Mariani BD, Tuan RS. Effects of
cadmium on trophoblast calcium transport. Placenta 1997;18:34156.
[82] Goyer RA, Cherian MG. Role of metallothionein in human placenta and rats
exposed to cadmium. IARC Sci Publ 1992;118:23847.
[83] Breen JG, Eisenmann C, Horowitz S, Miller RK. Cell-specic increases in metallothionein expression in the human placenta perfused with cadmium. Reprod
Toxicol 1994;8(4):297306.
[84] Torreblanca A, Del Ramo J, Sarkar B. Cadmium effect on zinc metabolism in
human trophoblast cells: involvement of cadmium-induced metallothionein.
Toxicology 1992;72(2):16774.
[85] Weir PJ, Miller RK, Maulik D, DiSantAgnese PA. Toxicity of cadmium in the
perfused human placenta. Toxicol Appl Pharmacol 1990;105(1):15671.
[86] Breen JG, Nelson E, Miller RK. Cellular adaptation to chronic cadmium
exposure: intracellular localization of metallothionein protein in human trophoblast cells (Jar). Teratology 1995;51(4):26672.
[87] Piasek M, Blanusa M, Kostial K, Laskey JW. Placental cadmium and progesterone concentrations in cigarette smokers. Reprod Toxicol 2001;15:67381.
[88] Kawai M, Swan KF, Green AE, Edwards DE, Anderson MB, Henson MC. Placental
endocrine disruption induced by cadmium: effects on P450 cholesterol sidechain cleavage and 3-hydroxysteroid dehydrogenase enzymes in cultured
human trophoblasts. Biol Reprod 2002;67:17883.
[89] Menoud PA, Schowing J. A preliminary study of the mechanisms of cadmium teratogenicity in chick embryo after direct action. J Toxicol Clin Exp
1987;7(2):7784.
[90] Webster WS, Messerle K. Changes in the mouse neuroepithelium associated
with cadmium-induced neural tube defects. Teratology 1980;21:7988.
[91] Messerle K, Webster WS. The classication and development of cadmiuminduced limb defects in mice. Teratology 1982;25:6170.
[92] De SK, Dey S, Andrews GK. Cadmium teratogenicity and its relationship with
metallothionein gene expression in midgestation mouse embryos. Toxicology
1990;64:89104.
[93] Sunderman Jr FW, Plowman MC, Hopfer SM. Teratogenicity of cadmium chloride in the South African frog, Xenopus laevis. IARC 1992;118:24956.
[94] Layton WM, Layton MW. Cadmium-induced limb defects in mice: strain associated differences in sensitivity. Teratology 1979;19:22935.
[95] Feuston MH, Scott Jr WJ. Cadmium-induced forelimb ectrodactyly: a proposed
mechanism of teratogenesis. Teratology 1985;32:40719.
[96] Holt D, Webb M. Teratogenicity of ionic cadmium in the Wistar rat. Arch
Toxicol 1987;59:4437.
[97] Perez-Coll CS, Herkovits J, Salibian A. Effects of cadmium on the development
of an amphibian. Arch Biol Med Exp (Santiago) 1985;18(1):339.
[98] Herkovits J, Cardellini P, Pavanati C, Perez-Coll CS. Susceptibility of early life
stages of Xenopus laevis to cadmium. Environ Toxicol Chem 1997;16(2):3126.
[99] Cheng SH, Wai AWK, So CH, Wu RSS. Cellular and molecular basis of
cadmium-induced deformities in zebrash embryos. Environ Toxicol Chem
2000;19(12):302431.
[100] Hen Chow ES, Cheng SH. Cadmium affects muscle type development and axon
growth in zebrash embryonic somitogenesis. Toxicol Sci 2003;73:14959.
[101] Thompson JM, Bannigan JG. The effects of cadmium on formation of the ventral body wall in chick embryos and their prevention by zinc pre-treatment.
Teratology 2001;64(2):8797.
[102] Thompson JM, Hipwell E, Loo HV, Bannigan JG. Effects of cadmium
on cell death and cell proliferation in chick embryos. Reprod Toxicol
2005;20(4):53948.
[103] Thompson JM, Bannigan JG. Omphalocele induction in the chick embryo by
administration of cadmium. J Pediatr Surg 2007;42(10):17039.
[104] Thompson J, Wong L, Lau PS, Bannigan J. Adherens junction breakdown in the periderm following cadmium administration in the chick
embryo: distribution of cadherins and associated molecules. Reprod Toxicol
2008;25(1):3946.
[105] Schmid BP, Kao J, Goulding E. Evidence for reopening of the cranial neural tube in mouse embryos treated with cadmium chloride. Experientia
1985;41:2712.
[106] Padmanabhan R. Light microscopic studies on the pathogenesis of exencephaly and cranioschisis induced in the rat after neural tube closure.
Teratology 1988;37:2936.
[107] Gale TF, Layton WM. The susceptibility of inbred strains of hamsters to
cadmium-induced embryotoxicity. Teratology 1980;21(2):1816.
[108] Wlodarczyk B, Biernacki B, Minta M, Zmudzki J. Postimplantation whole
embryo culture assay for hamsters: an alternative to rat and mouse. Sci World
J 2001;1:22734.
[109] Ferm VH. Developmental malformations induced by cadmium. A study of
timed injections during embryogenesis. Biol Neonate 1971;19(1):1017.
[110] Samarawickrama GP, Webb M. The acute toxicity and teratogenicity of cadmium in the pregnant rat. J Appl Toxicol 1981;1(5):2707.
[111] Galicia-Garcia V, Rojas-Lopez M, Rojas R, Olaiz G, Rios C. Cadmium levels in
maternal, cord and newborn blood in Mexico City. Toxicol Lett 1997;91:5761.

314

J. Thompson, J. Bannigan / Reproductive Toxicology 25 (2008) 304315

[112] Ronco AM, Arguello G, Munoz L, Gras N, Llanos M. Metals content in placentas from moderate cigarette consumers: correlation with newborn weight.
Biometals 2005;18:23341.
[113] Nishijo M, Tawara K. Relationship between newborn size and mothers blood
cadmium levels, Toyama, Japan. Arch Environ Health 2004;59(1):225.
[114] Nishijo M, Nakagawa H, Honda R, Tanebe K, Saito S, Teranishi H, et al. Effects
of maternal exposure to cadmium on pregnancy outcome and breast milk.
Occup Environ Med 2002;59:3947.
[115] Huel G, Boudene C, Ibrahim MA. Cadmium and lead content of maternal and
newborn hair: relationship to parity, birth weight, and hypertension. Arch
Environ Health 1981;36:2217.
[116] Frery N, Nessmann C, Girard F, Lafond J, Moreau T, Blot P, et al. Environmental
exposure to cadmium and human birthweight. Toxicology 1993;79:10918.
[117] Lagerkvist BJ, Nordberg GF, Soderberg HA, Ekesrydh S, Englyst V, Gustavsson
M, et al. Placental transfer of cadmium. IARC 1992;118:28791.
[118] Boadi WY, Yannai S, Urbach J, Brandes JM, Summer KH. Transfer and accumulation of cadmium, and the level of metallothionein in perfused human
placentae. Arch Toxicol 1991;65:31823.
[119] Konecki J, Blazejowski J, Slowinski J, Helewski K. Inuence of chronic cadmium exposure during pregnancy on DNA synthesis in different organs of rat
offspring. Med Sci Monit 2000;6(6):107781.
[120] Schoeters G, Den Hond E, Zuurbier M, Naginiene R, Van Den Hazel P, Stilianakis
N, et al. Cadmium and children: exposure and health effects. Acta Paediatr
2006;95(Suppl 453):504.
[121] Gumbiner BM. Cell adhesion: the molecular basis of tissue architecture and
morphogenesis. Cell 1996;84:34557.
[122] Lee NPY, Mruk D, Lee WM, Cheng CY. Is the cadherin/catenin complex a functional unit of cellcell actin-based adherens junctions in the rat testis? Biol
Reprod 2003;68:489508.
[123] Lui WY, Mruk D, Lee WM, Cheng CY. Sertoli cell tight junction dynamics: their
regulation during spermatogenesis. Biol Reprod 2003;68:108797.
[124] Goossenns S, Van Roy F. Cadherin-mediated cellcell adhesion in the testis.
Front Biosci 2005;10:398419.
[125] Cyr DG, Gregory M, Dube E, Dufresne J, Chan PTK, Hermo L. Orchestration
of occludins, claudins, catenins and cadherins as players involved in maintenance of the bloodepididymal barrier in animals and humans. Asian J Androl
2007;9(4):46375.
[126] Bloor DJ, Wilson Y, Kibschull M, Traub O, Leese HJ, Winterhager E, et al. Expression of connexins in human preimplantation embryos in vitro. Reprod Biol
Endocrinol 2004;2(2):25.
[127] Laird DW. Life cycle of connexins in health and disease. Biochem J
2006;394:52743.
[128] Li J, Zhang JV, Cao Y-J, Zhou J-X, Liu W-M, Fan X-J, et al. Inhibition of the catenin signalling pathway in blastocyst and uterus during the window of
implantation in mice. Biol Reprod 2005;72:7006.
[129] Sheth B, Fesenko I, Colling JE, Moran B, Wild AE, Anderson JM, et al. Tight
junction assembly during mouse blastocyst formation is regulated by late
expression of ZO-1 + isoform. Development 1997;124:202737.
[130] Kim J, Gye MC, Kim MK. Role of occludin, a tight junction protein, in
blastocoel formation, and in the paracellular permeability and differentiation of trophectoderm in preimplantation mouse embryos. Mol Cells
2004;17(2):24854.
[131] Bloor DJ, Metcalfe AD, Rutherford A, Brison DR, Kimber SJ. Expression of cell
adhesion molecules during human preimplantation embryo development.
Mol Human Reprod 2002;8(3):23745.
[132] Pummi K, Malminen M, Aho H, Karvonen SL, Peltonen J, Peltonen S. Epidermal tight junctions: ZO-1 and occludin are expressed in mature, developing,
and affected skin and in vitro differentiating keratinocytes. J Invest Dermatol
2001;117(5):10508.
[133] Hentula M, Peltonen J, Peltonen S. Expression proles of cellcell and
cellmatrix junction proteins in developing human epidermis. Arch Dermatol
Res 2001;293:25967.
[134] Arita K, Akiyama M, Tsuji Y, McMillan JR, Eady RAJ, Shimizu H. Changes in gap
junction distribution and connexin expression pattern during human fetal
skin development. J Histochem Cytochem 2002;50(11):1493500.
[135] Risek B, Klier G, Gilula NB. Developmental regulation and structural organization of connexins in epidermal gap junctions. Dev Biol 1994;164:18396.
[136] Oteiza PI, Adonaylo VN, Keen CL. Cadmium-induced testes oxidative damage in rats can be inuenced by dietary zinc intake. Toxicology 1999;137:
1322.
[137] Amara S, Abdelmelek H, Garrel C, Guiraud P, Douki T, Ravanat J-L, Favier A,
Sakly M, Ben Rhouma K. Preventive effect of zinc against cadmium-induced
oxidative stress in the rat testis. J Reprod Dev; in press.
[138] Gupta RS, Gupta ES, Dhakal BK, Thakur AR, Ahnn J. Vitamin C and vitamin
E protect the rat testes from cadmium-induced reactive oxygen species. Mol
Cells 2004;17(1):1329.
[139] Shaikh ZA, Vu TT, Zaman K. Oxidative stress as a mechanism of chronic
cadmium-induced hepatotoxicity and renal toxicity and protection by antioxidants. Toxicol Appl Pharmacol 1999;154:25663.
[140] Santos FW, Zeni G, Rocha JBT, Weis SN, Fachinetto JM, Favero AM, et al.
Diphenyl diselenide reverses cadmium-induced oxidative damage on mice
tissues. Chem-Biol Interact 2005;151:15965.
[141] Ognjanovic B, Zikic RV, Stajn A, Saicic ZS, Kostic MM, Petrovic VM. The effects
of selenium on the antioxidant defense system in the liver of rats exposed to
cadmium. Physiol Res 1995;44(5):293300.

[142] El-Sharaky AS, Newairy AA, Badreldeen MM, Eweda SM, Sheweita SA. Protective role of selenium against renal toxicity induced by cadmium in rats.
Toxicology 2007;235:18593.
[143] Shaikh ZA, Tang W. Protection against chronic cadmium toxicity by glycine.
Toxicology 1999;132(2/3):13946.
[144] Ardais AP, Santos FW, Nogueira CW. Ebselen attenuates cadmium-induced
speed testicular damage in mice. J Appl Toxicol; in press.
[145] Vousden KH. Outcomes of p53 activationspoilt for choice. J Cell Sci
2006;119:501520.
[146] Xu G, Zhou G, Jin T, Zhou T, Hammarstrom S, Bergh A, et al. Apoptosis and
p53 gene expression in male reproductive tissues of cadmium exposed rats.
Biometals 1998;12:1319.
[147] Zhou T, Zhou G, Song W, Eguchi N, Lu W, Lundin E, et al. Cadmium-induced
apoptosis and changes in expression of p53, c-jun and MT-1 genes in testes
and ventral prostate of rats. Toxicology 1999;142:113.
[148] Pulido MD, Parrish AR. Metal-induced apoptosis: mechanisms. Mutat Res
2003;533:22741.

[149] Watjen
W, Haase H, Biagioli M, Beyersmann D. Induction of apoptosis in mammalian cells by cadmium and zinc. Environ Health Perspect 2002;110(Suppl
5):8657.
[150] Cullinane J, Thompson J, Bannigan J. Protection from cadmium teratogenicity in the chick embryo by divalent metallic cations. Reprod Toxicol
2007;24:789 [abstracts].
[151] Santos FW, Oro T, Zeni G, Rocha JBT, do Nascimento PC, Nogueira CW.
Cadmium induced testicular damage and its response to administration of
succimer and diphenyl diselenide in mice. Toxicol Lett 2004;152:25563.
[152] Luo SQ, Plowman MC, Hopfer SM, Sunderman Jr FW. Mg(2+)-deprivation
enhances and Mg(2+)-supplementation diminishes the embryotoxic and teratogenic effects of Ni2+, Co2+, Zn2+, and Cd2+ for frog embryos in the FETAX
assay. Ann Clin Lab Sci 1993;23(2):1219.
[153] Bridges CC, Zalups RK. Molecular and ionic mimicry and the transport of toxic
metals. Toxicol Appl Pharmacol 2005;204:274308.
[154] Chmielnicka J, Sowa B. Cadmium interaction with essential metals (Zn, Cu, Fe),
metabolism metallothionein, and ceruloplasmin in pregnant rats and fetuses.
Ecotoxicol Environ Saf 1996;35:27781.
[155] Jacobs RM, Jones AO, Fox MR, Fry Jr BE. Retention of dietary cadmium and the
ameliorative effect of zinc, copper, and manganese in Japanese quail. J Nutr
1978;108(1):2232.
[156] He L, Girijashanker K, Dalton TP, Reed J, Li H, Soleimani M, et al. ZIP-8, member
of the Solute-Carrier-39 (SLC39) metal-transporter family: characterization of
transporter properties. Mol Pharmacol 2006;70(1):17180.
[157] Shannon RD. Revised effective ionic radii and systematic studies of interatomic distances in halides and chalcogenides. Acta Cryst 1976;A32:751
67.
[158] Braeckman B, Smagghe G, Brutsaert N, Cornelis R, Raes H. Cadmium uptake
and defence mechanism in insect cells. Environ Res A 1999;80:23143.
[159] Souza V, Bucio L, Gutierrez-Ruiz MC. Cadmium uptake by a human hepatic
cell line (WRL-68 cells). Toxicology 1997;120:21520.
[160] Prasad AS, Oberleas D, Wolf P, Horwitz JP. Studies on zinc deciency: changes
in trace elements and enzyme activities in tissues of zinc-decient rats. J Clin
Invest 1967;46(4):54957.
[161] Matthews KW, Mueller-Ortiz SL, Wetsel RA. Carboxypeptidase N: a pleiotropic
regulator of inammation. Mol Immunol 2004;40:78593.
[162] Prozialeck WC, Niewenhuis RJ. Cadmium (Cd++ ) disrupts intercellular
junctions and actin laments in LLC-PK1 cells. Toxicol Appl Pharmacol
1991;107:8197.
[163] Yin T, Green KJ. Regulation of desmosome assembly and adhesion. Semin Cell
Dev Biol 2004;15:66577.
[164] Jongen WMF, Fitzgerald DJ, Asamoto M, Piccoli C, Slaga TJ, Gros D, et al.
Regulation of connexin 43-mediated gap junctional intercellular communication by Ca2+ in mouse epidermal cells is controlled by E-cadherin. J Cell Biol
1991;114(3):54555.
[165] Saez JC, Berthoud VM, Branes MC, Martinez AD, Beyer EC. Plasma membrane
channels formed by connexins: their regulation and functions. Physiol Rev
2003;83:1359400.
[166] Leitinger B, McDowall A, Stanley P, Hogg N. The regulation of integrin function
by Ca2+ . Biochim Biophys Acta 2000;1498:918.
[167] King LM, Banks WA, George WJ. Differences in cadmium transport to the testis,
epididymis, and brain in cadmium-sensitive and -resistant murine strains
129/J and A/J. J Pharmacol Exp Therap 1999;289(2):82530.
[168] King LM, Banks WA, George WJ. Differential zinc transport into the testis
and brain of cadmium-sensitive and -resistant murine strains. J Androl
2000;21(5):65663.
[169] Dalton TP, He L, Wang B, Miller ML, Jin L, Stringer KF, et al. Identication of
mouse SLC39A8 as the transporter responsible for cadmium-induced toxicity
in the testis. PNAS 2005;102(9):34016.
[170] Waalkes MP, Poirier LA. Interactions of cadmium with interstitial tissue of the
rat testes. Uptake of cadmium by isolated interstitial cells. Biochem Pharmacol
1985;34(14):25138.
[171] Page KR, Abramovich DR, Aggett PJ, Bain M, Chippereld AR, Durdy H,
et al. Uptake of zinc by human placental microvillus border membranes
and characterization of the effects of cadmium on this process. Placenta
1992;13(2):15161.
[172] Sunderman Jr FW. Cadmium substitution for zinc in nger-loop domains
of gene-regulating proteins as a possible mechanism for the genotoxi-

J. Thompson, J. Bannigan / Reproductive Toxicology 25 (2008) 304315

[173]
[174]

[175]

[176]
[177]
[178]
[179]

[180]

[181]
[182]
[183]

[184]

[185]
[186]

[187]

city and carcinogenicity of cadmium compounds. Toxicol Environ Chem

1990;27:13141.
Predki PF, Sarkar B. The effect of replacement of zinc nger zinc on estrogen
receptor DNA interactions. J Biol Chem 1992;267(9):58426.
Faurskov B, Bjerregaard HF. Evidence for cadmium mobilization of intracellular calcium through a divalent cation receptor in renal distal epithelial A6
cells. Pugers Arch-Eur J Physiol 2002;445:4050.
Misra UK, Gawdi G, Akabani G, Pizzo SV. Cadmium-induced DNA synthesis
and cell proliferation in macrophages: the role of intracellular calcium and
signal transduction mechanisms. Cell Signal 2002;14:32740.
Cooper GM, Hausman RE. The cell: a molecular approach. 4th ed. ASM Press,
Sinauer Associates; 2007.
Suzuki Y, Chao SH, Zysk JR, Cheung WY. Stimulation of calmodulin by cadmium ion. Arch Toxicol 1985;57(3):20511.
Radisky D, Kaplan J. Regulation of transition metal transport across the yeast
plasma membrane. J Biol Chem 1999;274(8):44814.
Piersma AH, Roelen B, Roest P, Haakmat-Hoesenie AS, Van Achterberg TAE,
Mummery CL. Cadmium-induced inhibition of proliferation and differentiation of embryonal carcinoma cells and mechanistic aspects of protection by
zinc. Teratology 1993;48:33541.
King LA, MacDonald PC, Casey ML. Regulation of metallothionein expression
in human amnion epithelial and mesenchymal cells. Am J Obstet Gynecol
1997;177(6):1496501.
Kang YJ. Metallothionein redox cycle and function. Exp Biol Med
2006;231:145967.
Manuel Y, Thomas Y, Pellegrini O. Metallothionein and tissue damage. IARC
1992;118:2317.
Hamada T, Sasaguri T, Tanimoto A, Arima N, Shimajiri S, Abe T, et
al. Apoptosis of human kidney 293 cells is promoted by polymerized
cadmium-metallothionein. Biochem Biophys Res Commun 1996;219:829
34.
Ishido M, Tohyama C, Suzuki T. Cadmium-bound metallothionein induces
apoptosis in rat kidneys, but not in cultured kidney LLC-PK1 cells. Life Sci
1999;64:797804.
Aughey E, Fell GS, Svott R, Black M. Histopathology of early effects of oral
cadmium in the rat kidney. Environ Health Perspect 1984;54:15361.
Sabolic I, Ljubojevic M, Herak-Kramberger CM, Brown D. CdMT causes endocytosis of brush border transporters in rat renal proximal tubules. Am J Physiol
Renal Physiol 2002;283:F1389402.
Lin RH, Lee CH, Chen WK, Lin-Shiau SY. Studies on cytotoxic and genotoxic
effects of cadmium nitrate and lead nitrate in Chinese hamster ovary cells.
Environ Mol Mutagen 1994;23(2):1439.

315

[188] Hansmann I, Probeck HD. Detection of nondisjunction in mammals. Environ

Health Perspect 1979;31:1615.
[189] Fatur T, Lah TT, Filipic M. Cadmium inhibits repair of UV-, methyl
methanesulfonate- and N-methyl-N-nitrosourea-induced DNA damage in
Chinese hamster ovary cells. Mutat Res 2003;529:10916.
[190] Gick GG, McCarthy KS. Amplication of the metallothionein-I gene in
cadmium- and zinc-resistant Chinese hamster ovary cells. J Biol Chem
1982;257(15):904953.
[191] Yang J-L, Chao J-I, Lin J-G. Reactive oxygen species may participate in the mutagenicity and mutational spectrum of cadmium in Chinese hamster ovary-K1
cells. Chem Res Toxicol 1996;9:13607.
[192] Yang P-M, Chiu S-J, Lin K-A, Lin L-Y. Effect of cadmium on cell cycle progression
in Chinese hamster ovary cells. Chem-Biol Interact 2004;149:12536.
[193] Shiverick KT, Salaa C. Cigarette smoking and pregnancy. I. Ovarian, uterine
and placental effects. Placenta 1999;20:26572.
[194] Gray RH, Wu LY. Subfertility and risk of spontaneous abortion. Am J Pub Health
2000;90(9):14524.
[195] Wilks DJ, Hay AWM. Smoking and female fecundity: the effect and importance
of study design. Reprod Biol 2004;112:12735.
[196] Neal MS, Hughes EG, Holloway AC, Foster WG. Sidestream smoking is equally
as damaging as mainstream smoking on IVF outcomes. Human Reprod
2005;20(9):25315.
[197] Mortada WI, Sobh MA, El-Defrawy MM. The exposure to cadmium, lead and
mercury from smoking and its impact on renal integrity. Med Sci Monit
2004;10(3):CR1126.
[198] Zenzes MT, Krishnan S, Krishnan B, Zhang H, Casper RF. Cadmium accumulation in follicular uid of women in IVF is higher in smokers. Fertil Steril
1995;64:599603.
[199] Varga B, Zsolnai B, Paksy K, Naray M, Ungvary G. Age dependent accumulation
of cadmium in the human ovary. Reprod Toxicol 1993;7(3):2258.
[200] Stillman RJ, Rosenberg MJ, Sachs BP. Smoking and reproduction. Fertil Steril
1986;46(4):54566.
[201] Karaer A, Avsar FA, Batioglu S. Risk factors for ectopic pregnancy: a casecontrol study. Aust NZ J Obstet Gynaecol 2006;46:5217.
[202] Gerhard I, Monga B, Waldbrenner A, Runnebaum B. Heavy metals and fertility.
J Toxicol Environ Health A 1998.
[203] Cogswell ME, Weisberg P, Spong C. Cigarette smoking, alcohol use and adverse
pregnancy outcomes: implications for micronutrient supplementation. J Nutr
2003;133(5 Suppl 2):1722S31S.
[204] Semczuk M, Semczuk-Sikora A. New data on toxic metal intoxication (Cd,
Pb, and Hg in particular) and Mg status during pregnancy. Med Sci Monit
2001;7(2):33240.