Abstract
Oocyte maturation and embryonic development are sensitive to DNA damage. Compared with somatic cells or oocytes, little is known about the response to DNA damage in early preimplantation embryos. In this study, we examined DNA damage checkpoints and DNA repair mechanisms in parthenogenetic preimplantation porcine embryos. We found that most of the etoposide-treated embryos showed delay in cleavage and ceased development before the blastocyst stage. In DNA-damaged embryos, the earliest positive TUNEL signals were detected on Day 5 of in vitro culture. Caffeine, which is an ATM (ataxia telangiectasia mutated) and ATR (ataxia telangiectasia and Rad3-related protein) kinase inhibitor, and KU55933, which is an ATM kinase inhibitor, were equally effective in rescuing the etoposide-induced cell-cycle blocks. This indicates that ATM plays a central role in the regulation of the checkpoint mechanisms. Treating the embryos with histone deacetylase inhibitors (HDACi) increased embryonic development and reduced etoposide-induced double-strand breaks (DSBs). The mRNA expression of genes involved in non-homologous end-joining (NHEJ) or homologous recombination (HR) pathways for DSB repair was reduced upon HDACi treatment in 5-day-old embryos. Furthermore, HDACi treatment increased the expression levels of pluripotency-related genes (OCT4, SOX2 and NANOG) and decreased the expression levels of apoptosis-related genes (CASP3 and BAX). These results indicate that early embryonic cleavage and development are disturbed by etoposide-induced DNA damage. ATMi (caffeine or KU55933) treatment bypasses the checkpoint while HDACi treatment improves the efficiency of DSB repair to increase the cleavage and blastocyst rate in porcine early preimplantation embryos.
Highlights
IntroductionEffect of double-strand breaks (DSBs) in Porcine Early Embryos including X-rays and γ-rays), chemical drugs (such as doxorubicin, bleomycin, MLN4924, and etoposide), and physical inducers (such as laser micro-beam dissection) [3,4,5,6]
DNA double-strand breaks (DSBs) can be induced by endogenous metabolites or metabolic intermediates [1] as well as by exogenous factors such as UV [2], ionizing radiation (IR; PLOS ONE | DOI:10.1371/journal.pone.0142561 November 10, 2015Effect of DSBs in Porcine Early Embryos including X-rays and γ-rays), chemical drugs, and physical inducers [3,4,5,6]
We investigated the effect of etoposide-induced DNA DSBs on cell division and embryonic development in porcine early preimplantation embryos
Summary
Effect of DSBs in Porcine Early Embryos including X-rays and γ-rays), chemical drugs (such as doxorubicin, bleomycin, MLN4924, and etoposide), and physical inducers (such as laser micro-beam dissection) [3,4,5,6]. The occurrence of DSBs activates DNA damage checkpoints and DNA repair mechanisms. DNA damage checkpoint mechanisms arrest cell division until all DSBs are repaired. If abnormal DNA DSBs, such as the programmed DSBs, are not repaired immediately they can induce chromatin remodeling, cell cycle arrest, cell cycle delay, apoptosis, or other forms of cell death [7]. DSBs are known to affect oocyte maturation in multiple species [4,5,6,8]. Little is known about the response to DSBs in early preimplantation embryos
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