Histone Modifications of H3K4me3, H3K9me3 and Lineage Gene Expressions in Chimeric Mouse Embryo


Maryam Salimi, Ph.D, 1Abolfazl Shirazi, Ph.D, 2,3,*Mohsen Norouzian, Ph.D, 1,*Mohammad Mehdi Mehrazar, M.Sc, 2Mohammad Mehdi Naderi, Ph.D., 2Mohammad Ali Shokrgozar, Ph.D, 4Mirdavood Omrani, Ph.D., 5Seyed Mahmoud Hashemi, Ph.D., 6
Department of Biology and Anatomical Sciences, Faculty of Medicine, Shahid Beheshti University of Medical Sciences, Tehran, Iran
Reproductive Biotechnology Research Center, Avicenna Research Institute, ACECR, Tehran, Iran
Department of Gametes and Cloning, Research Institute of Animal Embryo Technology, Shahrekord University, Shahrekord, Iran
National Cell Bank of Iran, Pasteur Institute of Iran, Tehran, Iran
Department of Medical Genetics, Shahid Beheshti University of Medical Sciences, Tehran, Iran
Department of Immunology, School of Medicine, Shahid Beheshti University of Medical Sciences, Tehran, Iran
Department of Biology and Anatomical Sciences, Faculty of Medicine, Shahid Beheshti University of Medical Sciences, Tehran, Iran
Reproductive Biotechnology Research Center, Avicenna Research Institute, ACECR, Tehran, Iran
Department of Gametes and Cloning, Research Institute of Animal Embryo Technology, Shahrekord University, Shahrekord, Iran
National Cell Bank of Iran, Pasteur Institute of Iran, Tehran, Iran
Department of Medical Genetics, Shahid Beheshti University of Medical Sciences, Tehran, Iran
Department of Immunology, School of Medicine, Shahid Beheshti University of Medical Sciences, Tehran, Iran
*Corresponding Addresses: P.O.Box: 19615/1177 Reproductive Biotechnology Research Center Avicenna Research Institute ACECR Tehran Iran P.O.Box: 1985717-443 Department of Biology and Anatomical Sciences Faculty of Medicine Shahid Beheshti University of Medical Sciences Tehran Iran Emails:a.shirazi@ari.ir,norozian93@gmail.com
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Abstract

Objective

Chimeric animal exhibits less viability and more fetal and placental abnormalities than normal animal. This study was aimed to determine the impact of mouse embryonic stem cells (mESCs) injection into the mouse embryos on H3K9me3 and H3K4me3 and cell lineage gene expressions in chimeric blastocysts.

Materials and Methods

In our experiment, at the first step, incorporation of the GFP positive mESCs (GFP-mESCs) 129/Sv into the inner cell mass (ICM) of pre-compacted and compacted morula stage embryos was compared. At the second and third steps, H3K4me3 and H3K9me3 status as well as the expression of Oct4, Nanog, Tead4, and Cdx2 genes were determined in the following groups: i. In vitro blastocyst derived from in vivo morula subjected to mESCs injection (blast/chimeric), ii. In vivo derived blastocyst (blast/in vivo), iii. In vitro blastocyst derived from culture of morula in vivo (blast/morula), and iv. In vitro blastocyst derived from morula in vivo subjected to sham injection (blast/sham).

Results

Subzonal injection of GFP-mESCs at the pre-compacted embryos produced more chimeric blastocysts than compacted embryos (P<0.05). The number of trophectoderm (TE), ICM, ICM/TE and total cells in chimeric blastocysts were less than the corresponding numbers in blastocysts derived from other groups (P<0.05). In ICM and TE of chimeric blastocysts, the levels of H3K4me3 and H3K9me3 were respectively decreased and increased compared to the blastocysts of the other groups (P<0.05). Expressions of Oct4, Nanog and Tead4 were decreased in chimeric blastocysts compared to the blastocysts of the other groups (P<0.05), while this was not observed for Cdx2.

Conclusion

In the present study, embryo compaction significantly reduced the rate of incorporation of injected mESCs into the ICM. Moreover, in chimeric blastocysts, the levels of H3K9me3 and H3K4me3 were altered. In addition, the expressions of pluripotency and cell fate genes were decreased compared to blastocysts of the other groups.