Author/Authors :
Absalan، Forouzan نويسنده , , Saremy، Sadegh نويسنده Cell and Molecular Biology, Center Lab, Jundishapur Medical Science University, Ahvaz, IR Iran , , Mansouri، Esrafil نويسنده Department of Anatomical Sciences, Cellular and Molecular Research Center, School of Medicine, Ahvaz Jundishapur University of Medical Sciences, Ahvaz, Iran , , Taheri-Moghadam، Mahin نويسنده , , Eftekhari Moghadam، Ali Reza نويسنده Department of Anatomical Science, Faculty of Medicine, Ahvaz Jundishapur University of Medical Sciences, Ahvaz, Iran , , Ghanavati، Razie نويسنده Department of Molecular Biology and Development, Faculty of Medicine, Kazerun Islamic Azad University, Kazerun, Iran ,
Abstract :
Objective: Phthalates, which are commonly used to render plastics into soft and flexible
materials, have also been determined as developmental and reproductive toxicants in
human and animals. The purpose of this study was to evaluate the effect of mono-(2-
ethylhexyl) phthalate (MEHP) and di-(2-ethylhexyl) phthalate (DEHP) oral administrations
on maturation of mouse oocytes, apoptosis and gene transcription levels.
Materials and Methods: In this experimental study, immature oocytes recovered from
Naval Medical Research Institute (NMRI) mouse strain (6-8 weeks), were divided into
seven different experimental and control groups. Control group oocytes were retrieved
from mice that received only normal saline. The experimental groups I, II or III oocytes
were retrieved from mice treated with 50, 100 or 200 ?l DEHP (2.56 ?M) solution, respectively.
The experimental groups IV, V or VI oocytes were retrieved from mouse exposed to
50, 100 or 200 ?l MEHP (2.56 ?M) solution, respectively. Fertilization and embryonic development
were carried out in OMM and T6 medium. Apoptosis was assessed by annexin
V-FITC/Dead Cell Apoptosis Kit, with PI staining. In addition, the mRNA levels of Pou5f1,
Ccna1 and Asah1 were examined in oocytes. Finally, mouse embryo at early blastocyst
stage was stained with acridine-orange (AO) and ethidium-bromide (EB), in order to access
their viability.
Results: The proportion of oocytes that progressed up to metaphase II (MII) and 2-cells
embryo formation stage was significantly decreased by exposure to MEHP or DEHP, in a
dose-dependent manner. Annexin V and PI positive oocytes showed greater quantity in
the treated mice than control. Quantitative reverse transcriptase-polymerase chain reaction
(qRT-PCR) revealed that expression levels of Pou5f1, Asah1 and Ccna1 were significantly
lower in the treated mouse oocytes than control. The total cell count for blastocyst
developed from the treated mouse oocytes was lower than the controls.
Conclusion: These results indicate that oral administration of MEHP and DEHP could
negatively affect mouse oocyte meiotic maturation and development in vivo, suggesting
that phthalates could be risk factors for mammalians’ reproductive health. Additionally,
phthalate-induced changes in Pou5f1, Asah1 and Ccna1 transcription level could explain
in part, the reduced developmental ability of mouse-treated oocytes.
