Salamone D F, Adams G P, Mapletoft R J
WCVM, University of Saskatchewan, Saskatoon, Canada.
Theriogenology. 1999 Sep;52(4):549-61. doi: 10.1016/S0093-691X(99)00151-X.
We investigated factors that affect cumulus-oocyte complex (COC) morphology and oocyte developmental competence in subordinate follicles on different days after follicular wave emergence in beef heifers. In Experiment 1, heifers (n = 13) were assigned at random to COC aspiration during the growing/static (Days 1 to 3) or regressing (Day 5) phase of subordinate follicle development (follicular wave emergence = Day 0). Follicular wave emergence was induced by transvaginal ultrasound-guided follicular ablation, ovaries were collected at slaughter, all follicles > or = 2 mm except the dominant follicle were aspirated, and COC were microscopically evaluated for morphology. There was a greater percentage of COC with expanded cumulus layers on Day 5 (42.4%) than on Days 1 to 3 (2.2%). In Experiment 2, heifers (n = 64) at random stages of the estrous cycle had all follicles > or = 5 mm ablated and 4 d later, 2 doses of PGF were injected 12 h apart; heifers were monitored daily by ultrasonography for ovulation (Day 0 = follicular wave emergence). Heifers were assigned to the following time periods for oocyte collection from subordinate follicles: Days 0 and 1 (growing phase), Days 2, 3 and 4 (static phase), and Days 5 and 6 (regressing phase). Ovaries were individually collected at slaughter, and all follicles > or 2 mm except for the dominant follicle were aspirated. The COC were morphologically evaluated and then matured, fertilized and cultured in vitro. Expanded COC were more frequent during the regressing phase (53.4%) than the growing or static phase (14.4 and 17.8%, respectively; P < 0.05). While the proportions of COC with > or = 4 layers of cumulus cells and denuded oocytes were higher (P < 0.05) in the growing and static phases, the production of morulae was highest (P < 0.05) with COC collected from subordinate follicles during the regressing phase. In Experiment 3, heifers (n = 18) were assigned at random to oocyte collection from subordinate follicles 3 and 4 d (static phase) or 5 and 6 d (regressing phase) after follicular wave emergence. The heifers were monitored ultrasonically for ovulation (Day 0 = follicular wave emergence); COC were collected from all follicles (> or = 5 mm) except for the dominant follicle by transvaginal ultrasound-guided follicle aspiration 3 to 6 d later. Recovered oocytes were stained and examined microscopically to evaluate nuclear maturation. A higher proportion of oocytes collected on Days 5 and 6 showed evidence of nuclear maturation (50%) than on Days 3 and 4 (8.3%; P < 0.05). Results support the hypothesis that COC morphology and oocyte developmental competence change during the growing, static and regressing phases of subordinate follicle development.
我们研究了影响肉用小母牛卵泡波出现后不同天数次级卵泡中卵丘-卵母细胞复合体（COC）形态及卵母细胞发育能力的因素。在实验1中，将小母牛（n = 13）随机分配在次级卵泡发育的生长/静止期（第1至3天）或退化期（第5天）进行COC采集（卵泡波出现日=第0天）。通过经阴道超声引导下的卵泡消融诱导卵泡波出现，屠宰时采集卵巢，抽吸除优势卵泡外所有直径≥2 mm的卵泡，并在显微镜下评估COC的形态。第5天COC卵丘层扩张的比例（42.4%）高于第1至3天（2.2%）。在实验2中，处于发情周期随机阶段的小母牛（n = 64）所有直径≥5 mm的卵泡均被消融，4 d后间隔12 h注射2剂前列腺素F2α（PGF）；每天通过超声检查监测小母牛排卵情况（第0天=卵泡波出现）。将小母牛分配到以下时间段从次级卵泡采集卵母细胞：第0天和第1天（生长阶段）、第2、3和4天（静止阶段）以及第5和6天（退化阶段）。屠宰时分别采集卵巢，抽吸除优势卵泡外所有直径≥2 mm的卵泡。对COC进行形态学评估，然后进行体外成熟、受精和培养。退化期扩张的COC比生长或静止期更常见（分别为53.4%、14.4%和17.8%；P < 0.05）。虽然生长和静止期卵丘细胞层数≥4层的COC及裸卵的比例较高（P < 0.05），但退化期从次级卵泡采集的COC产生桑椹胚的比例最高（P < 0.05）。在实验3中，将小母牛（n = 18）随机分配在卵泡波出现后第3和4天（静止期）或第5和6天（退化期）从次级卵泡采集卵母细胞。通过超声监测小母牛排卵情况（第0天=卵泡波出现）；3至6 d后通过经阴道超声引导下的卵泡抽吸从除优势卵泡外所有直径≥5 mm的卵泡采集COC。对回收的卵母细胞进行染色并在显微镜下检查以评估核成熟情况。第5天和第6天采集的卵母细胞显示核成熟的比例较高（50%），高于第3天和第4天（8.3%；P < 0.05）。结果支持以下假设：在次级卵泡发育的生长、静止和退化阶段，COC形态及卵母细胞发育能力会发生变化。
