Journal of Infertility and Reproductive Biology, 2017, Volume 5, Issue 3, Pages: 16-18  
Subjection to Cadmium Chloride Compromises Oocyte  
Maturation, Fertilization Competence and Subsequent  
Embryo Development in Mature Female Mice  
Sanaz Alaee , Zahra Khodabandeh , Raheleh Asadollahpour  
. Department of Reproductive Biology, School of Advanced Medical Sciences and Technologies, Shiraz University of Medical Sciences, Shiraz, Iran  
. Stem Cells Technology Research Center, Shiraz University of Medical Sciences, Shiraz, Iran  
. Department of Molecular and Cell Biology, Faculty of Basic Sciences, University of Mazandaran, Babolsar, Iran  
Received: 06/05/2017  
Accepted: 03/06/2017  
Published: 20/06/2017  
Background: Cadmium is one of the main trace elements in the environment which in high doses could have adverse effects on human health,  
including male and female reproduction potential. Therefore, the present study was carried out to determine the possible toxic effect of this trace  
element on oocyte maturation.  
Materials and Methods: Mature female NMRI mice (68 weeks old) were divided into three groups and received 0 (control), 1 mg/kg and 2  
mg/kg cadmium chloride intraperitoneally for five days. Then all mice received PMSG and after 48 hours received hCG intraperitoneally. 14 hours  
later, cumulus-oocyte complexes were extracted from oviducts and the percentage of degenerated, germinal vesicle, metaphase I and metaphase  
II was recorded. Also, after performing in vitro fertilization, the cleavage and blastocyst formation rate was assessed.  
Results: The degeneration and maturation rate of oocytes was significantly higher and lower, respectively, in groups treated with different  
concentrations of cadmium compared with the control group (p<0.05). The number of fertilized, cleaved and blastocysts was significantly higher  
in groups that received cadmium compared with the control group (p<0.05).  
Conclusion: Even low dose and short duration exposure to cadmium has obvious detrimental effect on oocyte maturation, fertilization competence  
and subsequent embryo development.  
Keywords: Cadmium, Oocytes, Fertilization, Mice  
Nowadays, environmental pollution by human activities is a  
2 Materials and Methods  
2.1 Experimental design and collection of oocytes  
matter of concern because many pollutants released to environment  
are associated with the pathogenesis of a vast variety of diseases (1-  
Mature female NMRI mice (68 weeks old) were purchased  
from the animal house of Shiraz University of Medical Sciences.  
Animals were kept on a 12 h light: 12 h dark schedule with  
controlled temperature condition and free access to water and food.  
The animal experiments were performed according to the principles  
of the care and use of laboratory animals established by the National  
Institutes of Health (12). Mature female NMRI mice (68 weeks old)  
were divided into 3 groups and received 0 (control), 1 mg/kg and 2  
mg/kg cadmium chloride intraperitoneally for five days. At the 5th  
day, all mice received 10 IU of pregnant mare serum gonadotropin  
(PMSG; Gonaser, HIPRA, Spain), followed 48 h later by injecting  
10 IU human chorionic gonadotropin (hCG; Organon, Oss, The  
Netherlands). Fourteen hours later, mice were sacrificed by cervical  
dislocation and the oviducts were removed and placed in handling  
medium (GMOPSTM, Vitrolife, Göteborg, Sweden), which was  
pre-incubated for 24 h at 37°C. The oviducts were dissected with  
two insulin syringes (Helma Teb, Baspar Sanat Fakher, Saveh, Iran)  
under a stereomicroscope (Nikon, Tokyo, Japan) and the released  
cumulus oocyte complexes were placed in a few drops of  
hyaluronidase (80 IU/mL) (Vitrolife, Göteborg, Sweden) for  
separation of cumulus cells. The oocytes were placed in drops of G-  
IVF medium (Vitrolife) and the number of degenerated, Germinal  
Vesicle (GV), Metaphase I (MI) and Metaphase II (MII) oocytes was  
recorded using an inverted microscope (Nikon, Tokyo, Japan).  
Observation of germinal vesicle or the extrusion of first polar body  
). Many studies have demonstrated adverse effects of exposure of  
individuals to different levels of air pollutants on the occurrence of  
health problems. Cadmium (Cd) is a member of the heavy metal or  
trace elements family which has common industrial uses in batteries,  
alloys, plastic stabilizers and pigments; it is also an important  
component of cigarettes (4-6). The effect of indoor and outdoor air  
pollution on the human reproductive system has been an interesting  
topic for a wide variety of studies and policy analysts. It has been  
indicated that cadmium has a detrimental effect on female fertility  
7). Oocyte maturation is the final and a critical step in the  
development of a fertilizable oocyte defined as a re-entry into  
meiosis that occurs just prior to ovulation and allows oocytes to  
advance from prophase I to metaphase II of meiosis, determined by  
extrusion of the first polar body (8, 9). This process is highly  
sensitive and should be regulated precisely to result in successful  
ovulation (10). Furthermore, it has been demonstrated that the  
quality of the oocyte is related to subsequent fertilization and  
development competence of oocytes (11). Therefore, in the current  
study the potential toxicity of cadmium on oocyte maturation,  
fertilization and developmental competence was investigated by  
administration of low doses of cadmium chloride to mature female  
mice within a short period of time.  
Corresponding author: Zahra Khodabandeh, Stem Cells Technology Research Center, Shiraz University of Medical Sciences, Shiraz, Iran. E-mail:  
Journal of Infertility and Reproductive Biology, 2017, Volume 5, Issue 3, Pages: 16-18  
were used as the criterion for nuclear maturation of oocytes. Oocytes  
with extruded first polar body were used for in vitro fertilization.  
The adverse and toxicological effects of many pollutants on  
living organisms are demonstrated in a vast number of studies (13-  
5). Cadmium is a member of the heavy metal family, also called  
.2 IVF and in vitro development  
.2.1 Sperm capacitation  
Spermatozoa were collected from the cauda epididymides of  
trace elements, which at high doses exerts toxic effects on organs; it  
is also classified as a human carcinogen. It is generally present in the  
environment at low levels; however, human activity has greatly  
increased its levels in the environmental (6, 16). Cadmium is able to  
bioaccumulate in the food chain, including fish, meat,  
eggs and milk. People can be exposed to cadmium through eating  
contaminated foods, smoking cigarettes or drinking contaminated  
water. Living or working near industrial facilities which release  
cadmium into the air is detrimental to health. Cadmium exerts its  
effects through different mechanisms to destroy physiological  
functions including reproduction (7). Studies have demonstrated that  
air pollution has destructive impacts on both the male and female  
reproductive system at the level of tissue structure, function and  
gametogenesis (17-19).  
mature NMRI male mice and capacitated by preincubation at 37°C  
and 5% CO2 for 1 h in 200 µl of G-IVF drops (Vitrolife, Göteborg,  
Sweden) under mineral oil (Reproline, Rheinbach, Germany).  
.2.2 In vitro fertilization and embryo culture  
Mature oocytes were inseminated in vitro with 1×106  
spermatozoa/ml in 100 µl of G-IVF medium for 4 hours and the  
number of fertilized oocytes manifested by observation of two  
pronuclei (2PN) was recorded. The presumptive zygotes were  
cultured in G1 and then G2 medium (Vitrolife, Göteborg, Sweden)  
under mineral oil at 37°C in a humidified incubator with 5.0% CO2  
and rates of cleavage and blastocyst formation were recorded.  
The findings of this research show that a low dose of cadmium  
even with a short duration of administration induces degeneration of  
oocytes and has adverse effects on the maturation process of  
oocytes, displayed also in fertilization and developmental potential.  
Many researchers have demonstrated the adverse effects of this  
heavy metal on the male and female reproductive systems (5-7).  
Mohavi et al observed that cadmium exposure through drinking  
water decreases the number of ovulated oocytes and impairs oocyte  
meiotic maturation rate both in vivo and in vitro by disrupting of  
.2.3 Statistical analysis  
Data were analyzed by one-way ANOVA followed by the Tukey  
post hoc test using SPSS 20 (IBM Corp., Armonk, N.Y., USA).  
Differences were considered significant at P<0.05.  
As shown in Table 1, the percentage of degenerated and  
germinal vesicle oocytes significantly increased in the two  
experimental groups compared to the control group (p<0.05). Rates  
of MI and MII oocytes demonstrated significant decrease in the two  
experimental groups compared to the control group (p<0.05).  
Administration of cadmium to mature female mice significantly  
decreased the fertilization, cleavage and blastocyst formation rate  
mitochondrial function and histone modifications. The embryonic  
development after fertilization was also impaired (20). Akar et al  
observed the in vitro exposure of bovine COCs to CdCl2 for 24  
hours and concluded that the rate of COCs expansion, MI oocytes,  
post-fertilization cleavage rate in presumptive zygotes and  
blastocyst development decreased (21). Shimoi et al added cadmium  
to invitro maturation and IVF medium of mouse oocyte. Cd  
exposure during the maturation period disturbed the fertilization of  
oocytes and Cd exposure during the fertilization period, disrupting  
the normal development of embryos after fertilization (22). Overall,  
cadmium at the level of mentioned doses and duration of  
administration has significant detrimental effects on oocyte  
maturation and subsequent fertilization and development, but the  
precise mechanism of cadmium toxicity on the mentioned  
parameters remains to be clarified precisely.  
p<0.05) (Table 2). Percentage of degenerated, germinal vesicle  
GV) metaphase I (MI) and metaphase II oocytes (MII) in different  
groups. All experiments were repeated six times. Data are expressed  
as mean ± SEM. Different superscripts (a-c) show significant  
differences in a column and the same superscripts show no  
significant differences in a column (p<0.05). Percentage of fertilized  
oocytes with 2PN, cleavage rate and blastocyst formation. All  
experiments were repeated eight times. Different superscripts (a-c)  
show significant differences in a column and the same superscripts  
show no significant differences in a column (p<0.05).  
Table 1: Effect of cadmium chloride on oocyte maturation  
Different concentrations (mg/Kg)  
of cadmium chloride  
Total COCs  
Degenerated oocyte (%)  
GV oocyte (%)  
MI oocyte (%) MII oocyte (%)  
2.19 ± 0.9a  
24.96 ± 4.02b  
30.7 ± 3.2c  
5.21 ± 3.1a  
12.69 ± 5.7 b  
28.1 ± 1.89 c  
19.17 ± 1.4 a  
14.75 ± 4.7 b  
12.1 ± 3.28 c  
73.43 ± 3.6 a  
45.6 ± 3.6 b  
29.1 ± 4.8 c  
Table 2: Effect of cadmium chloride on IVF and embryo development  
Different concentrations (mg/Kg)  
of cadmium chloride  
Number of mature  
Fertilized oocytes with  
Blastocyst Formation  
Cleavage (%)  
2PN (%)  
88.2 ± 4.2a  
65.1 ± 4.1b  
54.2 ± 3.2c  
84.12 ± 2.6 a  
50.7 ± 2.6 b  
39.5 ± 3.2 c  
70.23 ± 3.3 a  
36.6 ± 1.5 b  
28.1 ± 2.8 c  
Journal of Infertility and Reproductive Biology, 2017, Volume 5, Issue 3, Pages: 16-18  
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In conclusion, one of the manifestations of cadmium toxicity in  
the female reproductive system is in the disruption of oocyte  
maturation, quality, fertilization potential and subsequent embryo  
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Authors are aware of, and comply with, best practice in  
publication ethics specifically with regard to authorship (avoidance  
of guest authorship), dual submission, manipulation of figures,  
competing interests and compliance with policies on research ethics.  
Authors adhere to publication requirements that submitted work is  
original and has not been published elsewhere in any language.  
Competing interests  
The authors declare that there is no conflict of interest that would  
prejudice the impartiality of this scientific work.  
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