Treatment of fetal fibroblasts with DNA methylation inhibitors and/or histone deacetylase inhibitors improves the development of porcine nuclear transfer-derived embryos

• Published in: Animal reproduction science Vol. 141; no. 3-4; pp. 164 - 171
• Main Authors: Diao, Yun-Fei, Naruse, Ken-Ji, Han, Rong-Xun, Li, Xiao-Xia, Oqani, Reza K, Lin, Tao, Jin, Dong-Il
• Format: Journal Article
• Language: English
• Published: Netherlands Elsevier B.V 01.10.2013
• Subjects: 5-aza-dC; acetylation; Animals; apoptosis; blastocyst; Cloning, Organism; Cytidine - analogs & derivatives; Cytidine - pharmacology; DNA methylation; DNA Methylation - drug effects; drugs; Embryo Culture Techniques - veterinary; Embryo development; Embryonic Development - drug effects; Epigenetic modification; epigenetics; Epigenomics; fibroblasts; Fibroblasts - cytology; Fibroblasts - drug effects; Fibroblasts - physiology; histone deacetylase; Histone Deacetylase Inhibitors - pharmacology; histones; Nuclear Transfer Techniques - veterinary; Phthalimides - pharmacology; Porcine nuclear transfer; swine; Swine - embryology; Tryptophan - analogs & derivatives; Tryptophan - pharmacology; TSA; Porcine nuclear transfer; TSA; 5-aza-dC; Epigenetic modification; Embryo development
• ISSN: 0378-4320; 1873-2232
• DOI: 10.1016/j.anireprosci.2013.08.008

This study investigated whether treating fetal fibroblast cells (donor cells) with epigenetic modification-inducing drugs could improve the development of porcine cloned embryos. Donor cells were treated with different DNA methylation inhibitors (5-aza-dC, zebularine or RG108; 5nM) or histone deacetylase inhibitors (TSA, NaBu or SCR; 50nM) for 1h, and then subjected to SCNT. All of the treated groups showed significantly higher blastocyst formation rates compared to the control group. We chose 5-aza-dC and TSA as a combined treatment, and found that donor cells co-treated with 2.5nM 5-aza-dC for 1h and subsequently treated with 50nM TSA for another 1h before SCNT showed significantly improved blastocyst rates compared to the control, 5-aza-dC-treated, and TSA-treated groups. The levels of DNA methylation were decreased (though not to a significant degree) in donor cells treated with 5-aza-dC, TSA or both. The histone H3 acetylation levels were significantly increased in donor cells treated with TSA or co-treated with 5-aza-dC and TSA. Donor cells simultaneously co-treated with 5nM 5-aza-dC and 50nM TSA for 1h showed increased apoptosis of SCNT blastocysts. However, when we decreased the concentration of 5-aza-dC to 2.5nM, the co-treatment induced less apoptosis among SCNT blastocysts and the blastocyst development rate improved. Together, these results indicate that treatment of donor cells with 5-aza-dC, TSA, or TSA plus a low dose of 5-aza-dC could improve the blastocyst development of porcine cloned embryos.