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KMID : 0894520060100020105
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Development & Reproduction
2006 Volume.10 No. 2 p.105 ~ p.113
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Expression and Localization of ATF4 Gene on Oxidative Stress in Preimplantation Mouse Embryo
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Na Won-Heum
Kang Han-Seung
Eo Jin-Won
Gye Myung-Chan
Kim Moon-Kyoo
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Abstract
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Reactive oxygen species(ROS) generated in cellular metabolism have an effect on cell maturation and development. In human reproductive tract, oxidative injury by ROS may induce female infertility. Also, oxidative injury may be responsible for developmental retardation and arrest of mammalian preimplantation embryos. Activating transcription factor 4(ATF4) is a member of the cyclic-AMP response element-binding(CREB) familiy of basic region- leucine zipper(bZip). ATF4 is known to regulate stress response to protect cell from various stress factors and inducer of apoptisis. The purpose of this study was to investigate whether ATF4 is involved in the defensive mechanism in oxidative stress condition during the development of mouse preimplantation embryos. To verify the expression of ATF4 in oxidative stress condition, 2-cell stage embryos were cultured in HTF media containing 0.1mM, 0.5mM or 1mM hydrogen peroxide(H₂O₂) for 1hr(2-cell), 8hr(4-cell), 17hr(8-cell), 24hr(morula), 48hr(early blastocyst) or 64hr(late blastocyst). The developmental rate decreased in the 0.1mM H₂O₂ treated group compared with control group. In embryos treated with 0.5mM and 1mM H₂O₂ showed 2-cell block. As a results of the semi-quantitative RT-PCR analysis of SOD1, ATF4 and Bax gene expression, SOD1, ATF4 and Bax genes were increased in 0.1mM, 0.5mM, 1mM H₂O₂ treated groups compared with control group. In 2-cell embryos, expression of SOD1, ATF4 and Bax genes were notably increased in 0.1mM, 0.5mM, 1mM H₂O₂ treated groups compared with control group. Immunofluorescence analysis showed that ATF4 protein was localized at the cytoplasm of preimplantation embryos. The increase in ATF4 immunoreactivety was observed in the 0.1mM, 0.5mM, 1 mM H₂O₂ treated groups compared with control group. It suggests that oxidative stress by H₂O₂ induces expression of ATF4 and may be involved in protection mechanism in preimplantation embryos from oxidative injury.
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KEYWORD
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ROS, ATF4, SOD1, Bax, Oxidative stress, Preimplantation, Embryo
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