Effect of Temperature Decline on the Cytoskeletal Organization of the Porcine Oocyte
Effect of Temperature Decline on the Cytoskeletal Organization of the Porcine Oocyte
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DOI：
10.1274/jmor.24.107
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发表时间：
2007-10
期刊：
影响因子：
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通讯作者：
Hiroyuki Suzuki;T. Kumai;M. Matsuzaki
中科院分区：
文献类型：
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作者：
Hiroyuki Suzuki;T. Kumai;M. Matsuzaki
摘要本研究的目的是评估冷却对猪中期II（MII）卵母细胞中影响减数分裂纺锤体分布和结构的细胞骨架组织的影响。荧光染色用于微管、微丝和染色体的可视化。将体外成熟的卵母细胞保持在37 °C（对于对照）或突然冷却至5或18°C。微管被优先标记在减数分裂的纺锤体和微丝主要检测到的MII卵母细胞的皮质。大多数对照组（87%，n=97）有一个桶形纺锤体，而其余13%显示一个广泛的，箱形纺锤体。在18 °C下暴露30 min后，94%的卵母细胞（n=48）具有盒形纺锤体（63%），极间距缩短（7.4 ± 0.4 μm vs.对照组9.8 ± 0.2 μm，P<0.05），或无纺锤体（31%）。将卵母细胞暴露于5 °C 30分钟导致81%的卵母细胞（n=103）的纺锤体完全解体。这种纺锤体解体的卵母细胞在细胞质中的微管信号强度增加，皮质微丝染色不规则。分别有64%（n=33）和89%（n=74）的卵母细胞冷却至18和5 °C时，皮质微丝染色强度降低。当卵母细胞暴露于5 °C 30 min后在37 °C下复温时，无论复温持续时间（0、1、10、20或60 min）如何，任何卵母细胞（n=138）的纺锤体都没有恢复正常，尽管微管和微丝重新组装。然而，更多的卵母细胞含有分散的染色体，没有纺锤体和崩溃的微丝结构，作为复温时间增加超过20分钟。这些结果表明，在猪MII卵母细胞微管纺锤体和皮质微丝不可逆地受到5 °C冷休克，和极性微管组织中心功能障碍，即使复温后。
ABSTRACT The purpose of this study was to evaluate effect of cooling on the cytoskeletal organization affecting the distribution and structure of the meiotic spindle in the porcine metaphase II (MII) oocyte. Fluorescence staining was done for visualization of microtubules, microfilaments and chromosomes. In vitro-matured oocytes were either maintained at 37 °C (for controls) or cooled abruptly to 5 or 18°C. Microtubules were preferentially labeled at the meiotic spindle and microfilaments were mainly detected at the cortex of the MII oocyte. The majority of controls (87%, n=97) had a barrel-shaped spindle, whereas the remaining 13% showed a broad, box-shaped spindle. After exposure to 18 °C for 30 min, 94% of oocytes (n=48) possessed either a box-shaped spindle (63%) with a shortened interpolar distance (7.4 ± 0.4 μm vs. 9.8 ± 0.2 μm for controls P<0.05), or no spindle (31%). Exposure of oocytes to 5 °C for 30 min caused complete disassembly of the spindle in 81% of the oocytes (n=103). Such spindle-disassembled oocytes had increased intensity of microtubule signaling in the cytoplasm and irregular staining of cortical microfilaments. Cortical microfilament staining decreased in intensity in 64% (n=33) and 89% (n=74) of the oocytes cooled to 18 and 5 °C, respectively. When oocytes were rewarmed at 37 °C after exposure to 5 °C for 30 min, the spindles were not restored to normal in any oocyte (n=138) regardless of the rewarming duration (0, 1, 10, 20 or 60 min), although microtubules and microfilaments were reassembled. However, more oocytes contained dispersed chromosomes with no spindle and collapsed microfilament architecture, as rewarming time increased beyond 20 min. These results suggest that in porcine MII oocytes both microtubular spindle and cortical microfilaments are irreversibly affected by a 5 °C cold shock, and that the polar microtubule-organizing centers malfunction even after rewarming.