Sutovsky P, Navara C S, Schatten G
Department of Zoology, University of Wisconsin, Madison 53706, USA.
Biol Reprod. 1996 Dec;55(6):1195-205. doi: 10.1095/biolreprod55.6.1195.
Sperm incorporation and the conversion of the sperm-derived components into zygotic structures during in vitro fertilization of bovine oocytes was explored by combining ultrastructural studies with observations of the fertilizing sperm tagged with a mitochondrion-specific vital dye MitoTracker green FM. The zygotes fertilized by the MitoTracker-labeled sperm were fixed at various times after fertilization and then processed for immunocytochemistry to examine the distribution of DNA, microtubules, and sperm tail components, including the fibrous sheath and axonemal microtubules. We show here that the complete incorporation of the sperm, but not sperm-oocyte binding and oocyte activation, depends upon the integrity of oocyte microfilaments and is inhibited by the microfilament disrupter cytochalasin B. After sperm incorporation, the mitochondria are displaced from the sperm's connecting piece, and the sperm centriole is exposed to the egg cytoplasm. This event is followed by the formation of the microtubule-based sperm aster, which is responsible for the union of male and female pronuclei. Concomitantly, the major structure of the sperm principal piece, the fibrous sheath, disappears. After the first mitosis, the compact mitochondrial sheath can be seen in one of the blastomeres. An aggregate of the sperm mitochondria is observed at the entry of the second mitosis, although they remain in the vicinity of the nucleus and can later be seen at one pole of the metaphase spindle. The mitochondrial cluster is occasionally found in one of the blastomeres in the early-stage four-cell embryos, but it is no longer detected by the beginning of the third mitotic cycle. These data suggest that the disassembly of the sperm tail during bovine fertilization occurs as a series of precisely orchestrated events involving the destruction (fibrous sheath and mitochondrial sheath) and transformation (DNA, sperm centriole) of particular sperm structures into zygotic and embryonic components.
通过将超微结构研究与用线粒体特异性活性染料MitoTracker green FM标记的受精精子的观察相结合，探讨了牛卵母细胞体外受精过程中精子的纳入以及精子衍生成分向合子结构的转化。用MitoTracker标记的精子受精的合子在受精后的不同时间固定，然后进行免疫细胞化学处理，以检查DNA、微管和精子尾部成分（包括纤维鞘和轴丝微管）的分布。我们在此表明，精子的完全纳入，而非精子与卵母细胞的结合及卵母细胞的激活，取决于卵母细胞微丝的完整性，并受到微丝破坏剂细胞松弛素B的抑制。精子纳入后，线粒体从精子的连接段移位，精子中心粒暴露于卵细胞质中。此事件之后是基于微管的精子星体的形成，它负责雄原核和雌原核的结合。与此同时，精子主段的主要结构——纤维鞘消失。第一次有丝分裂后，在其中一个卵裂球中可以看到紧密的线粒体鞘。在第二次有丝分裂进入时观察到精子线粒体的聚集，尽管它们仍留在细胞核附近，随后可在中期纺锤体的一极看到。线粒体簇偶尔在早期四细胞胚胎的一个卵裂球中发现，但在第三个有丝分裂周期开始时就不再检测到。这些数据表明，牛受精过程中精子尾部的解体是一系列精心编排的事件，涉及特定精子结构（纤维鞘和线粒体鞘）的破坏以及转化（DNA、精子中心粒）成为合子和胚胎成分。
