Feng Xiu-Qing, Lin Ying-Wei, Chen Ya-Jun, Zhong Shu-Qi, Yan Xiao-Fei, Dong Jian-Jiang, Lei Lei
Department of Histology and Embryology, Harbin Medical University, Harbin 150081, China.
Sheng Li Xue Bao. 2008 Feb 25;60(1):113-8.
In this study we detected dynamic changes and function of beta-tubulin, a subtype of microtubule, during the first cleavage period in mouse parthenogenetic and in vitro fertilized embryos. Firstly, we compared the developmental potential of in vitro fertilized, parthenogenetic, and in vivo fertilized embryos in culture. Then, the dynamic changes of beta-tubulin and nucleus in parthenogenetic and in vitro fertilized preimplantation embryos were detected by immunofluorescence and confocal microscopy to analyze the role of microtubules in meiotic division and embryonic development. The results indicated that the development rate of in vivo fertilized embryos was significantly higher than that of in vitro fertilized or parthenogenetic embryos (P<0.05). However, there was no significant difference in developmental potential between in vitro fertilized and parthenogenetic embryos. During in vitro fertilization, oocyte was activated when sperm entered it. Oocyte resumed the second meiotic division. Condensed maternal chromosomes aligning at the equator of the spindle were pulled to the spindle poles by kinetochore microtubules in anaphase. Furthermore, in telophase, there were microtubules between the two sets of decondensed maternal chromosomes. One set formed the second polar body (Pb(2)), which was extruded to the perivitelline space. The other set formed female pronucleus. Meanwhile, 5-8 h after fertilization, sperm chromatin condensed and decondensed to form male pronucleus. Microtubule composed mesosome and cytaster remodeling around male and female pronuclei to form long microtubules, which pull the pronuclei to get close. During 4-6 h parthenogenetic activation, SrCl(2) activated oocytes to resume meiosis. As a consequence, sister chromatids were pulled to spindle poles. Cytochalasin B, which was applied in the medium, inhibited the extrusion of Pb(2). Two haploid pronuclei in the cytoplasm were connected by microtubules. Compared with that in in vitro fertilization, oocyte is easier to be activated in parthenogenetic activation. Chemical activation is more efficient than sperm penetration in in vitro fertilization as indicated by earlier and better remodeling of the microtubules.
在本研究中，我们检测了微管蛋白的一种亚型β-微管蛋白在小鼠孤雌生殖胚胎和体外受精胚胎首次卵裂期的动态变化及其功能。首先，我们比较了体外受精胚胎、孤雌生殖胚胎和体内受精胚胎在培养中的发育潜能。然后，通过免疫荧光和共聚焦显微镜检测孤雌生殖胚胎和体外受精植入前胚胎中β-微管蛋白和细胞核的动态变化，以分析微管在减数分裂和胚胎发育中的作用。结果表明，体内受精胚胎的发育率显著高于体外受精胚胎或孤雌生殖胚胎（P<0.05）。然而，体外受精胚胎和孤雌生殖胚胎在发育潜能上没有显著差异。在体外受精过程中，精子进入卵母细胞时卵母细胞被激活。卵母细胞恢复第二次减数分裂。在后期，凝聚的母源染色体在纺锤体赤道面排列，被动粒微管拉向纺锤体两极。此外，在末期，两组解凝聚的母源染色体之间存在微管。一组形成第二极体（Pb(2)），被挤出到卵周隙。另一组形成雌原核。同时，受精后5-8小时，精子染色质凝聚和解凝聚形成雄原核。微管围绕雄原核和雌原核组成中间体和星体并重塑形成长微管，将原核拉近。在孤雌激活4-6小时期间，SrCl(2)激活卵母细胞恢复减数分裂。结果，姐妹染色单体被拉向纺锤体两极。培养基中加入的细胞松弛素B抑制了Pb(2)的排出。细胞质中的两个单倍体原核由微管连接。与体外受精相比，卵母细胞在孤雌激活中更容易被激活。化学激活比体外受精中的精子穿透更有效，微管的重塑更早且更好。
