Effect of in vivo and in vitro heat stress on DNA methylation and DNA hydroxymethylation of bovine oocytes and early embryos

In the bovine commercial industry, reduced reproductive performance in response to heat stress is one of the main factors causing economic losses. Several studies have shown that heat stress negatively affects oocytes and embryos at the morphological, biochemical, transcriptional, and developmental levels. Yet, there is limited information on the effect of heat stress on the epigenetic modifications of bovine oocytes and embryos. Therefore, the objective of this study was to evaluate the effect of in vivo and in vitro heat stress on the developmental competence, DNA methylation, and DNA hydroxymethylation of bovine oocytes and early embryos. Oocytes were collected through ovum pick-up from non-lactating, non-pregnant Bos taurus beef cows in February and August under Louisiana environmental conditions. The treatments evaluated were: in vivo heat stress (oocytes collected in August), in vitro heat stress (oocytes collected in February and subjected to in vitro heat stress), and control (oocytes collected in February and not subjected to in vitro heat stress). Developmental rates, DNA methylation and DNA hydroxymethylation of metaphase II oocytes (MII), 2-pronucleus embryos (2 PN) and 2–4 cell embryos were evaluated. Global DNA methylation and DNA hydroxymethylation were evaluated through fluorescence immunostaining. No differences between treatments was detected in developmental rates of MII oocytes, 2 PN embryos and 2–4 cell embryos. Similarly, no differences between treatments was detected in global DNA methylation and DNA hydroxymethylation of MII oocytes, 2 PN embryos, and 2–4 cell embryos. Importantly, no differences in global DNA methylation or DNA hydroxymethylation of paternal or maternal pronucleus was detected, indicating that the demethylation process during the 2 PN embryo stage was not altered at the global level. The results of our study showed that under our experimental conditions, in vivo and in vitro heat stress did not affect developmental rates, DNA methylation and DNA hydroxymethylation of MII oocytes and early embryos produced from oocytes obtained from non-lactating, non-pregnant Bos taurus beef cows.