Interdisciplinary Graduate School of Science and Technology, Shinshu University, Ueda, Nagano, Japan.
Biol Reprod. 2013 Aug 1;89(2):26. doi: 10.1095/biolreprod.113.109769. Print 2013 Aug.
Cryotolerance of matured bovine oocytes is not fully practical even though a promising vitrification procedure with a ultrarapid cooling rate was applied. The present study was conducted to investigate whether recovery culture of vitrified-warmed bovine oocytes with an inhibitor (Y-27632) of Rho-associated coiled-coil kinase (ROCK) can improve the developmental potential after in vitro fertilization (IVF) and in vitro culture. Immediately after warming, almost all oocytes appeared to be morphological normal. Treatment of the postwarming oocytes with 10 μM Y-27632 for 2 h resulted in the significantly higher oocyte survival rate before IVF as well as higher cleavage rate and blastocyst formation rate. Quality analysis of the resultant blastocysts in terms of total cell number and apoptotic cell ratio also showed the positive effect of the Y-27632 treatment. Time-dependent change in mitochondrial activity of the vitrified-warmed oocytes was not influenced by ROCK inhibition during the period of recovery culture. However, the ability of ooplasm to support single-aster formation was improved by the ROCK inhibition. Thus, inhibition of ROCK activity in vitrified-warmed bovine oocytes during a short-term recovery culture can lead to higher developmental competence, probably due to decreased apoptosis and normalized function of the microtubule-organizing center.
尽管采用了超快冷却速率的有前途的玻璃化冷冻程序，但成熟牛卵母细胞的耐冷冻性仍不完全实用。本研究旨在探讨玻璃化冷冻-解冻牛卵母细胞的恢复培养是否可以改善体外受精（IVF）和体外培养后的发育潜能。在解冻后立即，几乎所有的卵母细胞都表现出形态正常。用 10 μM Y-27632 处理解冻后的卵母细胞 2 小时，可显著提高 IVF 前的卵母细胞存活率以及卵裂率和囊胚形成率。从总细胞数和凋亡细胞比例方面对所得囊胚的质量分析也表明了 Y-27632 处理的积极效果。在恢复培养期间，玻璃化冷冻-解冻卵母细胞中线粒体活性的时间依赖性变化不受 ROCK 抑制的影响。然而，卵质支持单个星状体形成的能力通过 ROCK 抑制得到改善。因此，在短时间的恢复培养过程中抑制玻璃化冷冻-解冻牛卵母细胞中的 ROCK 活性可导致更高的发育能力，这可能是由于凋亡减少和微管组织中心的功能正常化所致。
