Abstract：
Oocyte cryopreservation is increasingly being used in reproductive technologies for conservation and breeding purposes. Further development of oocyte cryopreservation techniques requires interdisciplinary insights in the underlying principles of cryopreservation. This review aims to serve this purpose by: (1) highlighting that preservation strategies can be rationally designed, (2) presenting mechanistic insights in volume and osmotic stress responses associated with CPA loading strategies and cooling, and (3) giving a comprehensive listing of oocyte specific biophysical membrane characteristics and commonly used permeation model equations. It is shown how transport models can be used to simulate the behavior of oocytes during cryopreservation processing steps, i.e., during loading of cryoprotective agents (CPAs), cooling with freezing as well as vitrification, warming and CPA unloading. More specifically, using defined cellular and membrane characteristics, the responses of oocytes during CPA (un)loading were simulated in terms of temperature- and CPA type-and-concentration-dependent changes in cell volume and intracellular solute concentration. In addition, in order to determine the optimal cooling rate for slow programmable cooling cryopreservation, the freezing-induced cell volume response was simulated at various cooling rates to estimate rates with tolerable limits. For vitrification, special emphasis was on prediction of the timing of reaching osmotic tolerance limits during CPA exposure, and the need to use step-wise CPA addition/removal protocols. In conclusion, we present simulations and schematic illustrations that explain the timing of events during slow cooling cryopreservation as well as vitrification, important for rationally designing protocols taking into account how different CPA types, concentrations and temperatures affect the oocyte.
摘要：
卵母细胞冷冻保存越来越多地用于生殖技术中，用于保护和繁殖目的。卵母细胞冷冻保存技术的进一步发展需要对冷冻保存的基本原理有跨学科的见解。本文旨在通过以下方式实现这一目的：（1）强调可以合理设计保存策略，（2）在与CPA加载策略和冷却相关的体积和渗透应激反应中提出机械见解，（3）全面列出了卵母细胞特异性生物物理膜特征和常用的渗透模型方程。显示了如何使用运输模型来模拟冷冻保存处理步骤中卵母细胞的行为，即，在加载冷冻保护剂(CPAs)期间，冷冻和玻璃化冷却，变暖和CPA卸载。更具体地说,使用定义的细胞和膜特征，根据细胞体积和细胞内溶质浓度的温度和CPA类型和浓度依赖性变化来模拟CPA（un）加载期间卵母细胞的反应。此外,为了确定慢速可编程冷却冷冻保存的最佳冷却速率，在各种冷却速率下模拟冷冻诱导的细胞体积反应,以在可容许的限度内估算速率.对于玻璃化，特别强调在CPA暴露期间达到渗透耐受极限的时间预测，以及使用分步CPA添加/删除协议的需要。总之,我们提出了模拟和示意图，解释了在缓慢冷却冷冻保存以及玻璃化过程中事件的时间，对于合理设计协议，考虑到不同的CPA类型，浓度和温度影响卵母细胞。
