Parthenogenetic activation of pig oocytes with calcium ionophore and the block to sperm penetration after activation.
Parthenogenetic activation of pig oocytes with calcium ionophore and the block to sperm penetration after activation.
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用钙离子载体孤雌激活猪卵母细胞，并在激活后阻止精子穿透。
DOI：
10.1095/biolreprod58.6.1357
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发表时间：
1998
影响因子：
3.6
通讯作者：
Day,BN
中科院分区：
文献类型：
--
作者：
Wang,WH;Macháty,Z;Abeydeera,LR;Prather,RS;Day,BN
钙离子载体A23187可以孤雌激活许多动物的卵母细胞。本研究旨在从功能上分析A23187激活猪体外成熟卵母细胞的机制。在实验1中，A23187的浓度对细胞内钙瞬变，皮质颗粒（CG）胞吐，核活化，和ESTA反应，这是确定的硬化和精子穿透性的影响进行了检查。将无卵丘卵母细胞暴露于0-100 µM A23187 5 min。发现细胞内钙瞬变幅度、CG胞吐百分比和原核形成百分比均以浓度依赖性方式增加。在用25-100 µM A23187处理的卵母细胞中，透明质酸（ZP）溶解的时间增加。50-100 µM A23187处理后，精子对ZP完整卵母细胞的穿透率降低，仅3-4%的卵母细胞被精子穿透。在实验2中，卵母细胞用100 µM A23187处理5 min，然后在授精前培养10 min或3.5 h。两组卵母细胞的穿透率无差异（12.0% vs. 12.2%），但穿透率显著低于对照组（85.2% vs. 82.4%）。在实验3中，用100 μM A23187处理卵母细胞5分钟，然后从一部分卵母细胞中除去ZP。然后将ZP完整和无ZP的卵母细胞进行授精以检查精子穿透。65个有ZP的卵母细胞中有1个（2%）被精子穿透，52个无ZP的卵母细胞中有48个（92%）被精子穿透。这些结果表明，A23187激活猪卵母细胞是A23187诱导的细胞内钙增加的结果，A23187诱导的皮质反应可以阻止精子穿透ZP完整的卵母细胞，但不ZP的卵母细胞。
Calcium ionophore A23187 can parthenogenetically activate oocytes in many animals. The present study was designed to analyze functionally the mechanism of A23187 activation of pig oocytes matured in vitro. In experiment 1, effects of the concentration of A23187 on intracellular calcium transients, cortical granule (CG) exocytosis, nuclear activation, and zona reaction, which was determined by zona hardening and sperm penetrability, were examined. Cumulus-free oocytes were exposed to 0–100 µM A23187 for 5 min. It was found that the amplitude of the intracellular calcium transients, percentage of CG exocytosis, and percentage of pronuclear formation were increased in a concentration-dependent manner. The time for dissolution of zona pellucida (ZP) was increased in the oocytes treated with 25–100 µM A23187. Penetration of ZP-intact oocytes by spermatozoa was decreased and only 3–4% of oocytes were penetrated by spermatozoa after 50–100 µM A23187 treatment. In experiment 2, oocytes were treated with 100 µM A23187 for 5 min and then cultured for 10 min or 3.5 h before insemination. No difference in penetration rates was observed between the two groups of oocytes (12.0% vs. 12.2%), but the penetration rates were significantly lower than those in controls (85.2% vs. 82.4%). In experiment 3, treatment of oocytes with 100 µM A23187 for 5 min was followed by removal of the ZP from a portion of the oocytes. ZP-intact and ZP-free oocytes were then inseminated for examination of sperm penetration. One of 65 (2%) oocytes with ZP and 48 of 52 (92%) oocytes without ZP were penetrated by spermatozoa. These results indicate that activation of pig oocytes by A23187 is the result of A23187-induced intracellular calcium increase and that A23187-induced cortical reaction can prevent sperm penetration of ZP-intact oocytes, but not ZP-free oocytes.