Cycloheximide speeds the porcine oocyte nuclear kinetics and retains embryonic developmental potential in the presence of gonadotrophins

摘要

Cycloheximide speeds the porcine oocyte nuclear kinetics and retains embryonic developmental potential in the presence of gonadotrophins. Can. J. Anim. Sci. 90: 189-196. Animal embryo engineering requires large amounts of synchronized mature oocytes in vitro. However, porcine cumulus-oocyte complexes aspirated from 3-8 mm follicles are at different germinal vesicle stages. They reach metaphase II stages asynchronously when cultured in vitro. In this study, we examined the effects of pretreatment with or without cycloheximide (CHX), equine chorionic gonadotrophin (eCG), human chorionic gonadotrophin (hCG), and their combinations on meiotic synchronization and the developmental competence of porcine oocytes in vitro following electrical activation. The COCs were pretreated for 12 h with either control medium (TCM 199), CHX (TCM 199 + CHX), eCG/hCG (TCM 199 + eCG/hCG) or eCG/hCG+CHX (TCM 199 + CHX +eCG/hCG), and then cultured for up to 32 h with TCM199 +eCG/hCG. After 12 h pretreatment, the rates of germinal vesicle breakdown (GVBD) were lower (P <0.05) in the CHX (8.4%) and eCG/hCG+CHX (1.5%) groups compared with control (55.4%) and eCG/hCG (27.2%) groups. After removal of CHX and culture for an additional 12 h in vitro, the majority of the oocytes were synchronized at the GVBD stage in CHX (75.6%) and eCG/hCG+CHX (65.0%) groups. At additional 32 h of culture, the rate of oocytes in metaphase II in eCG/hCG+CHX group (68.3%) was significantly (P < 0.05) higher than the eCG/hCG group (54.8%), but did not differ from other groups (control: 61.3%, CHX: 58.8%). After electrical activation, the cleavage and blastocyst formation rates in the CHX group (80.3%; 19.5%) were significantly (P < 0.05) lower than those in the control group (95.5%; 45.3%), while no difference was found between eCG/hCG+CHX (82.2%; 34.4%) and control groups. Our data, hence, demonstrate pretreatment with CHX hastened nuclear kinetics of porcine oocytes cultured in vitro; however, embryo development potential was retained only when gonadotrophins is present in the in vitro maturation (IVM) medium. Thus, CHX should be used in the two-step culture systems in combination with gonadotrophins.