소의 체외성숙난포란의 체외수정과 발생에 관한 연구 = Studies on In Vitro Fertilization and Development of Bovine Follicular Oocytes Matured In Vitro

다국어 초록 (Multilingual Abstract)

Effects of adding 10 to 20% fetal calf serum(FCS) and estrous cow serum(ECS), and hormones(GTH or GTH plus E₂) to maturation medium on in vitro maturation(IVM) and in vitro fertilization(IVF) of bovine follicular oocytes and effects of sperm treatment and bull on IVF and embryo development were studied. Finally, developmental potential these in-vitro fertilized embryos at 2-to 8-cell stage(44h postinsemination) was investigated under in vivo(rabbit oviduct) and in vitro(Ham`s Fl0 plus 10% FCS with or without bovine oviduct epithelial cells, BOEC) 4 day-culture system. 1. Addition of 10 to l5% FCS to mKRB and DM(-BSA) and of 20% FCS to Ham`s F10 increased(65%) IVM, respectively. IVM of these media supplemented with hormones was higher(70∼72%) than that of media without hormone. Addition of ECS showed higher IVM at 10% level for mKRB and 10 to 15% for TCM199 and the highest IVM(88%) was obtained in TCM199 containing 10% ECS and LH. 2. Higher IVF rate (based on formation of both pronuclei) in Exp I was obtained from oocytes matured in mKRB plus 10% FCS and HIS-sperm treatment. In Exp 2. IVFrate (based on cleavage. 2-3 cells) of oocytess matured in TCM199 was higher than in other media and IVF using fresh semen was significantly higher but was similar between sperm treatments. In Exr 3, IVM in TCM199 containing 10% ECS and LH and IVF by C+H-sperm treatment significantly increased early cleavage(2-to 8-cell). 3. In vitro-acrosome reaction(AR) among individual bulls in Exp 1 ranged from 16 to 45% and bull group having lower AR showed low IVF rate than group having higher AR. In Exp 3, cleavage rate by sperm treatment with both chemicals(C+H) was higher than that by H alone. 4. In IVF with in vivo-capaciated sperm, development from 2-to 3-cell to above 8-cell was higher under in vivo culture(54.5%) than under in vitro culture(31.3%) and higher in oocytes matured in TCM199 plus to ECS. In IVF with in vitro-capacitated sperm, development into above 8-cell stage was different between initial stage(2-to 3-and 4-to 8-cell) of embryos(26.7% : 41.7%) and significantly lower under in vitro culture from 2-to 3 cell stages(l0% : 29.4%). Under in vitro co-culture with BOEC, embryo development from 4-to 8-cell was significantly improved(53.3%) and 3(20%) of 15 cultured embryos developed into morula. The present results indicate that addition of ECS and hormones can improve IVM, IVF and development of bovine follicular oocytes, sperm factor may be important for IVF and further development, and co-culture with BOEC is an effective means for development into above 16-cell stage.