牛体外受精卵在两种共同培养系统中发育的研究

为了使牛体外受精卵能通过体外早期发育阻滞期,我们建立了卵丘细胞单层(A)和输卵管上皮单层(B)两种共同培养系统。A1共同培养实验是在本实验室进行的,A2和B共同培养实验均在日本岗山大学农学部动物繁殖学研究室进行,牛输卵管组织的分离使用0.76％EDTA—PBS溶液,共同培养系统均使用含10％小牛血清的TCM—199(Earle's salts)作为培养液。培养的卵泡卵母细胞体外成熟率为100％,体外受精率为99—100％。在A1共同培养实验中,越过阻滞期发育到16—细胞以上的胚胎占卵裂胚的35.7％,与A2共同培养实验中越过阻滞期的发育率(40.1％)无显著差异(P<0.05)。A1和A2共同培养实验,在卵裂基础上得到的桑椹胚和囊胚发育率分别为23.7％和27.9％。每百枚培养的卵母细胞,在A1共同培养实验中可获得桑椹胚和囊胚15.1枚,在A2共同培养实验中可获桑椹胚和囊胚的20.5枚。B共同培养实验中桑椹胚和囊胚发育率为54.1％,显著高于A1或A2共同培养实验的相应发育率(P<0.001),使用B共同培养系统每百枚培养的卵母细胞可以获得37枚桑椹胚和囊胚。

DEVLOPMENT OF IN VITRO FERTILIZED BOVINE OVA IN TWO CO-CULTURE SYSTEMS

Two In Vitro co-culture systems were used in the experiment to overcome the 8-16-cell block in the early development of In Vitro fertilized bovine ova. "A"represented cumulus cells monolayer co-culture system and "B" represented bovine oviductal epithelial cells (BOEC)system. EXp. A1 was carried out in our lab and Exp. A2 and Exp. B were carried out in the lab of animal reproduction in Okayama uviversity( Japan). A solution of 0. 76% ED-TA-PBS was used to disperse oviductal tissue which was then cultured in TCM-199(Earle's salts) + 10% fetal calf serum(M-199FCS)to obtain monolayers. M-199FCS was also used in the two co-culture systems. The mature rate of the In Vitro cultured follicular oocytes was 100% and the In Vitro fertilization rates were in range of 99-100%, in Exp. A1, the developmental rate of the embryos over 16-cell stage was 35. 7% which didn't differ from that in Exp. A2(40.1)(P<0. 05 ),but Wxp. B. improved (P<0. 05) the developmental rate of the embryos which overcome the block (57. 7)compared with Exp. A1 or Exp. A2. The percentage of morula and blastocysts developed from cleaved ova in Exp. AI and Exp. A2. were 23. 7% and 27. 9% respectively. The average 15. 1 morula and blastocysts could be produced out of each one-hundred In Vitro cultured oocytes in Exp. A1 and 20. 5 morula and blastocysts could be produced in Exp. A2. The percentage of embryos developed to morula and blastocyst stage(54.1) in Exp. B was significantly(P<0.001) higher than those in Exp. A1 or Exp. A2. In Exp. B.37. 0 morula and blastocyts could be produced out of each one-hundred oocytes cultured In Vitro.