Trichostatin A alters the expression of cell cycle controlling genes and microRNAs in donor cells and subsequently improves the yield and quality of cloned bovine embryos in vitro

Trichostatin A (TSA), a histone deacetylase inhibitor, has been used to improve nuclear reprogramming in somatic cell nuclear transfer embryos. However, the molecular mechanism of TSA for the improvement of the pre- and postimplantation embryonic development is unknown. In the present study, we investigated mechanism of cell cycle arrest caused by TSA and also determined embryo quality and gene expression in cloned bovine embryos produced from TSA-treated donor cells compared with embryos produced by in vitro fertilization or parthenogenetic activation. We observed that, 50 nM TSA-treated cells were synchronized at G0/G1 stage with concomitant decrease in the proportion of these cells in the S stage of the cell cycle, which was also supported by significant changes in cell morphology and decreased proliferation (P < 0.05). Measurement of relative expression using real-time polymerase chain reaction of a some cell cycle-related genes and microRNAs in treated donor cells showed decreased expression of HDAC1, DNMT1, P53, CYC E1, and CDK4 and increased expression of DNMT3a, CDKN1A, CDK2, CDK3, miR-15a, miR-16, and miR-34a (P < 0.05). No change in the relative expression of miR-449a was noticed. Trichostatin A treatment of donor cells significantly improved both cleavage and blastocyst rate (P < 0.05) compared with the control embryos, also apoptotic index in treated cloned blastocysts was significantly decreased compared with the nontreated blastocysts (P < 0.05) and was at the level of IVF counterpart. Relative expression of HDAC1 and DNMT3a was significantly lower in treated cloned and parthenogenetic embryos than that of nontreated and IVF counterpart, whereas in case of P53, expression level between treated and IVF embryos was similar, which was significantly lower than nontreated cloned and parthenogenetic embryos. In conclusion, our data suggested that TSA improves yield and quality of cloned bovine embryos by modulating the expression of G0/G1 cell cycle stage-related microRNA in donor cells, which support that TSA might be great cell cycle synchronizer apart from potent epigenetic modulator in cloning research in future.