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Phosphorylated H2AX in parthenogenetically activated, in vitro fertilized and cloned bovine embryos

Published online by Cambridge University Press:  15 April 2014

A.F. Pereira ,
L.M. Melo ,
V.J.F. Freitas  and
D.F. Salamone
A.F. Pereira
Affiliation:
Laboratory of Physiology and Control of Reproduction, School of Veterinary Medicine, Ceará State University, Fortaleza, Brazil.
L.M. Melo
Affiliation:
Laboratory of Physiology and Control of Reproduction, School of Veterinary Medicine, Ceará State University, Fortaleza, Brazil.
V.J.F. Freitas
Affiliation:
Laboratory of Physiology and Control of Reproduction, School of Veterinary Medicine, Ceará State University, Fortaleza, Brazil.
D.F. Salamone*
Affiliation:
Laboratory of Animal Biotechnology, School of Agronomy, Buenos Aires University, Av. San Martín 4453, C1417, Buenos Aires, Argentina. Laboratory of Animal Biotechnology, School of Agronomy, Buenos Aires University, Buenos Aires, Argentina.
*
All correspondence to: Daniel Felipe Salamone. Laboratory of Animal Biotechnology, School of Agronomy, Buenos Aires University, Av. San Martín 4453, C1417, Buenos Aires, Argentina. Tel: +54 11 4524 8000. Fax: +54 11 4514 8737. e-mail: [email protected]
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Summary

In vitro embryo production methods induce DNA damage in the embryos. In response to these injuries, histone H2AX is phosphorylated (γH2AX) and forms foci at the sites of DNA breaks to recruit repair proteins. In this work, we quantified the DNA damage in bovine embryos undergoing parthenogenetic activation (PA), in vitro fertilization (IVF) or somatic cell nuclear transfer (SCNT) by measuring γH2AX accumulation at different developmental stages: 1-cell, 2-cell and blastocyst. At the 1-cell stage, IVF embryos exhibited a greater number of γH2AX foci (606.1 ± 103.2) and greater area of γH2AX staining (12923.6 ± 3214.1) than did PA and SCNT embryos. No differences at the 2-cell stage were observed among embryo types. Although PA, IVF and SCNT were associated with different blastocyst formation rates (31.1%, 19.7% and 8.3%, P < 0.05), no differences in the number of γH2AX foci or area were detected among the treatments. γH2AX is detected in bovine preimplantation embryos produced by PA, IVF and SCNT; the amount of DNA damage was comparable among those embryos developing to the blastocyst stage among different methods for in vitro embryo production. While IVF resulted in increased damage at the 1-cell embryo stage, no difference was observed between PA and SCNT embryos at any developmental stage. The decrease in the number of double-stranded breaks at the blastocyst stage seems to indicate that DNA repair mechanisms are functional during embryo development.

Keywords

ChromatinCloningDNA breakageHistone phosphorylationPreimplantation embryo
Type
Research Article
Information
Zygote , Volume 23 , Issue 4 , August 2015 , pp. 485 - 493
Copyright
Copyright © Cambridge University Press 2014 

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