56 Sirtinol Treatment Influences Preimplatation Development of Porcine Embryos via Regulation of Autophagy and ApoptosisM. G. Kim A , S. T. Shin B , H. D. Shin C and H. T. Lee A
A Department of Animal Biotechnology, Konkuk University, Seoul, South Korea;
B College of Veterinary Medicine, Chungnam National University, Daejeon, South Korea;
C Department of Life Science, Sogang University, Seoul, South Korea
Reproduction, Fertility and Development 30(1) 167-167 https://doi.org/10.1071/RDv30n1Ab56
Published: 4 December 2017
Sirtuin (Sirt), nicotinamide adenine dinucleotide dependent class III histone deacetylase, plays an important role in cellular processes including DNA repair, apoptosis, cell cycle, aging, and determining lifespan. In previous studies, levels of Sirt1 to Sirt3 mRNA were detected in porcine embryos for the first time and levels are lower in blastocysts relative to matured oocytes. In addition, mitochondrial dysfunction and hyperglycemia increases LC3 protein levels and apoptosis in porcine parthenotic embryos and modulation of autophagy also influences apoptosis, mitochondrial contents, abnormal autophagosome formation, and maternal mRNA degradation. However, Sirt-mediated mechanisms have not been examined in in vitro-produced embryos of pig. Therefore, we investigated the relationship between Sirt inhibition and autophagy/mitophagy in porcine pre-implantation embryos. After IVF, embryos were cultured in NCSU-23 media in the presence and absence of 100 μM sirtinol (Sirt inhibitor) until the expended blastocyst stage. As a result, there were no significant differences between the rate of cleavage in control (69.22 ± 1.29) and treated groups (72.66 ± 1.08). However, embryos treated with sirtinol had significantly decreased developmental rates to the morula as well as blastocyst stages. Especially, expanded blastocysts (9.90 ± 1.56 v. 2.92 ± 0.94%) were barely observed in sirtinol-treated group. In the levels of Sirt transcripts, Sirt2 mRNA was significantly lower in sirtinol-treated blastocysts compared with controls (P < 0.05), but the levels of Sirt1 and Sirt3 mRNA were similar in both groups. In addition, we found that sirtinol treatment induced autophagy by increasing the expressions of LC3, Beclin1, and ATG5 in blastocysts. Furthermore, we observed that the abundance of mitochondria stained with mitotracker was lower in sirtinol-treated blastocysts than that of control. Finally, we found that sirtinol treatment resulted in a higher total apoptotic index (6.88 ± 0.84) compared with the control (12.84 ± 0.99) in blastocysts. In summary, our findings in this study demonstrated that Sirt inhibition by sirtinol led to lower levels of Sirt2 transcript in blastocysts, reduced developmental capability and embryo quality with regulation of ATGs, LC3 proteins, apoptosis-related genes, and mitochondrial abundance. Therefore, these results suggest that Sirt2 may play an important role in the pre-implantation development of porcine embryos and their quality through the regulation of autophagy/mitophagy and apoptosis pathways.
This research was supported by a Grant from the Bio & Medical Technology Development Program (2015M3A9C7030091) of the National Research Foundation (NRF) funded by the Korean government.