Campos I, Coy P, Romar R, Ruiz S, Gadea J
Department of Animal Biology, University of Murcia, Spain.
Theriogenology. 2001 Apr 15;55(7):1489-500. doi: 10.1016/s0093-691x(01)00496-4.
The in vitro penetrability of porcine oocytes is conditioned by several factors, some of which remain unclear. Knowledge of the different effects of the cellular components involved in penetrability would no doubt serve to simplify laboratory IVF methods. This study was designed to evaluate the effects of the following factors on penetrability: oocyte maturational stage, the presence of isolated or oocyte-attached cumulus cells, and coincubation of in vitro-matured and immature oocytes. Immature oocytes and oocytes matured in Waymouth medium were obtained from non atretic follicles and fertilized in TCM 199 medium. Sperm-rich fractions were collected by the gloved hand method and semen was used for IVF at a final concentration of 1 x 10(6) cells/mL in all experiments. Under the same conditions of IVF, the penetrability of the immature cumulus-oocyte complexes (COCs) was significantly lower than that of mature COCs, in terms of penetration rate and mean number of sperm per penetrated oocyte. This difference was abolished when the oocytes were denuded, leading to similar penetration rates. Coincubation of mature and immature COCs reduced the penetrability of immature COCs compared with that observed when these were incubated in isolation. However, neither the addition of isolated cumulus cells from decumulated mature oocytes nor the addition of denuded mature oocytes to immature COCs modified the penetration rate. These findings suggest that the presence of surrounding cumulus cells is mainly responsible for the differences observed in penetrability, regardless of the maturational stage of the oocyte. Moreover, when mature and immature COCs are coincubated, penetrability of immature COCs is diminished by the effects of the mature COC and not by the independent actions of the cellular components.
猪卵母细胞的体外穿透性受多种因素制约,其中一些因素尚不清楚。了解参与穿透性的细胞成分的不同作用无疑有助于简化实验室体外受精方法。本研究旨在评估以下因素对穿透性的影响:卵母细胞成熟阶段、孤立的或附着于卵母细胞的卵丘细胞的存在以及体外成熟和未成熟卵母细胞的共同孵育。从未闭锁卵泡中获取在韦茅斯培养基中成熟的未成熟卵母细胞,并在TCM 199培养基中进行受精。通过戴手套的手方法收集富含精子的部分,在所有实验中,精液用于体外受精的终浓度为1×10⁶个细胞/毫升。在相同的体外受精条件下,从未成熟卵丘 - 卵母细胞复合体(COCs)的穿透率和每个穿透卵母细胞的平均精子数来看,其穿透性显著低于成熟COCs。当卵母细胞去卵丘后,这种差异消失,导致相似的穿透率。与单独孵育时相比,成熟和未成熟COCs的共同孵育降低了未成熟COCs的穿透性。然而,向未成熟COCs中添加来自脱卵丘成熟卵母细胞的孤立卵丘细胞或添加去卵丘成熟卵母细胞均未改变穿透率。这些发现表明,无论卵母细胞的成熟阶段如何,周围卵丘细胞的存在是观察到的穿透性差异的主要原因。此外,当成熟和未成熟COCs共同孵育时,未成熟COCs的穿透性因成熟COC的作用而降低,而非细胞成分的独立作用。