43 SHORT- AND LONG-LASTING EFFECTS OF TRICHOSTATIN A TREATMENT OF SCNT EMBRYOS IN CATTLE

Abstract

Both preimplantation and full-term development of mouse somatic cell nuclear transfer (SCNT) embryos are significantly enhanced by treatment with trichostatin A (TSA), an inhibitor of histone deacetylase (Kishigami et al. 2006 Biochem. Biophys. Res. Commun. 340, 183–189; Rybouchkin et al. 2006 Biol. Reprod. 74, 1083–1089). The present study was designed to examine the effect of TSA treatment on preimplantation and full-term development of bovine cloned embryos. To investigate the effect of TSA on bovine NT embryos development, we treated them with 50 nM TSA during the first 10 h after activation. Bovine NT-embryos were reconstructed using adult fibroblasts of 2 female donors (A and B) with significantly different in vitro cloning efficiency (respectively, 84/245; 34.3% v. 155/298; 52.1% blastocyst D7, P ≤ 0.05, chi-square test). TSA treatment significantly improved blastocyst rate in A, however did not affect development in B (56.3% and 50.5%, respectively). The level of acetylated histone H3K9 10 h after activation detected by anti-acH3K9 antibody was significantly increased after TSA-treatment in A (P ≤ 0.05, Student’s t-test) but did not change in B, thus demonstrating that the levels of histone acetylation in cloned embryos correlate with their in vitro developmental potential. To evaluate the long-lasting effect of TSA-treatment on the full-term development of cloned embryos, SCNT embryos derived from 4 female donor animals were reconstructed. 196 TSA-treated embryos at the blastocyst stage were transferred into 98 recipients and 2 calves (2%) were born. In the control group, 167 embryos were transferred into 141 recipients and 3 calves (2.1%) were born. Our data show that cell lines demonstrate different susceptibility to TSA that may affect reprogramming of the somatic genome with low level of acetylation resulting in higher in vitro embryo development. However, TSA does not improve overall cloning efficiency in cattle, measured as full-term development. Project partly supported by EU grants Plurisys (n 22348), Xenome (LSHB-CT-2006-037377) and Regione Lombardia.