Comizzoli P, Mermillod P, Cognié Y, Chai N, Legendre X, Mauge R
MNHN, Conservation des Espèces Animales, Paris, France.
Theriogenology. 2001 Jan 15;55(2):649-59. doi: 10.1016/s0093-691x(01)00433-2.
The aim of our study was to define the conditions for IVM and IVF of oocytes in 2 common deer species as models for endangered related subspecies. Immature oocytes were recovered during the breeding season from postmortem ovaries (red deer) or by repeated laparoscopic follicular aspiration (sika deer). Oocytes were cultured for 24 h in IVM medium supplemented with EGF or FSH and follicular fluid. Stag semen was collected by electroejaculation (both species) or by epididymal flushing (red deer) and cryopreserved. For IVF, oocytes were exposed to different concentrations of thawed spermatozoa in a modified Tyrode albumin lactate pyruvate medium supplemented with 20% (v/v) estrus sheep serum for 18 h. After IVF, presumptive zygotes were allowed to develop in vitro for 7 days in synthetic oviduct fluid (SOF) supplemented with fetal calf serum (10%, v/v). In both species, the presence of ovine FSH and follicular fluid improved the in vitro maturation rate. In the sika deer, the optimal sperm concentration for IVF was 10(6)/mL and some fertilized oocytes reached the early morula stage (20 to 25 cells). In the red deer, after IVF with ejaculated or epididymal spermatozoa (2.0 x 10(6)/mL), 20% of zygotes developed to the blastocyst stage (50 to 80 cells).
我们研究的目的是确定两种常见鹿种的卵母细胞体外成熟（IVM）和体外受精（IVF）条件，作为濒危相关亚种的模型。在繁殖季节，从死后的卵巢中采集未成熟卵母细胞（马鹿），或通过反复腹腔镜卵泡抽吸术采集（梅花鹿）。将卵母细胞在添加了表皮生长因子（EGF）或促卵泡激素（FSH）以及卵泡液的IVM培养基中培养24小时。通过电刺激采精（两种鹿种）或附睾冲洗（马鹿）采集雄鹿精液并冷冻保存。对于IVF，将卵母细胞在添加了20%（v/v）发情绵羊血清的改良Tyrode白蛋白乳酸丙酮酸培养基中，暴露于不同浓度的解冻精子中18小时。IVF后，假定的受精卵在添加了胎牛血清（10%，v/v）的合成输卵管液（SOF）中体外培养7天。在两种鹿种中，绵羊FSH和卵泡液的存在提高了体外成熟率。在梅花鹿中，IVF的最佳精子浓度为10⁶/mL，一些受精卵达到了早期桑椹胚阶段（20至25个细胞）。在马鹿中，用射出的或附睾精子（2.0×10⁶/mL）进行IVF后，20%的受精卵发育到囊胚阶段（50至80个细胞）。
