Removal of cumulus cells before oocyte nuclear maturation enhances enucleation rates without affecting the developmental competence of porcine cloned embryos

Abstract

The present study compared the efficiency of somatic cell nuclear transfer (SCNT) using porcine oocytes that were denuded of their cumulus cells at different maturation time. In pre-denuded group, the cumulus cells from cumulus-oocyte complexes (COCs) were removed at 29 hr post in vitro maturation (hpm) and followed by further culture for 12 hr. In control group, as a commonly followed procedure, cumulus cells were removed from COCs at 41 hpm. The majority of porcine oocytes at 29 hpm were observed in metaphase of the first meiotic division (MI). At 41 hpm, no significant (P > 0.05) differences were observed in nuclear maturation and mitochondrial distribution of oocytes between pre-denuded and control groups. However, in pre-denuded group oocytes, metaphase II (MI) plate and spindle were located closely as 'adjacent' to the first polar body (PB1), resulting in an increased enucleation rates than in control group oocytes by blind enucleation method. Following SCNT and parthenogenesis (PA) using pre-denuded group and control group oocytes, no significant (P > 0.05) differences were observed with respect to the development, total cell number, incidence of apoptosis and the expression profile of developmentally important genes (Pou5f1, Dnmtl, Dnmt3a, Igf2r, Box, Bcl2 and Glut1) at the blastocyst stage. In conclusion, the removal of cumulus cells at 29 hpm in porcine oocytes increased the enucleation rates through proper positioning of PB1 without compromising the quality of SCNT embryos during preimplantation development. Hence, this could be a valuable strategy to improve the SCNT efficiency in a porcine model.