Unit for Reproductive Medicine - Clinic for Horses, University of Veterinary Medicine Hannover, Hannover, Germany.
Biostabilization laboratory - Lower Saxony Centre for Biomedical Engineering, Implant Research and Development, University of Veterinary Medicine Hannover, Hannover, Germany.
Biochim Biophys Acta Gen Subj. 2021 Feb;1865(2):129797. doi: 10.1016/j.bbagen.2020.129797. Epub 2020 Nov 17.
Rational design of cryopreservation strategies for oocytes and ovarian cortex tissue requires insights in the rate at which cryoprotective agents (CPA) permeate and concomitant water transport takes place. The aim of the current study was to investigate possible differences in permeation kinetics of different CPAs (i.e., glycerol/GLY, ethylene glycol/EG, dimethyl sulfoxide/DMSO, and propylene glycol/PG), in equine oocytes as well as ovarian tissue.
Membrane permeability of oocytes to water (Lp) and to CPAs (Ps) was inferred from video microscopic imaging of oocyte volume responses during perfusion with anisotonic and CPA solutions. CPA diffusion into ovarian tissue and tissue dehydration was monitored during incubation, using osmometer and weight measurements, to derive CPA diffusion coefficients (D).
Membrane permeability of oocytes towards CPAs was found to increase in the order GLY < EG < DMSO<PG. Permeability towards water in anisotonic solutions was determined to be higher than in CPA solutions, indicating CPAs alter membrane permeability properties. CPA diffusion in ovarian tissue increased in the order GLY,PG < EG,DMSO. Tissue dehydration was found to increase with exposure to increasing CPA concentrations, which inversely correlated with CPA diffusivity.
In conclusion, it is shown here that the rate of CPA movement across membrane bilayers is determined by different physical barrier factors than those determining CPA movement in tissues.
The parameters presented in this study can be applied in models describing solute and water transport in cells and tissues, as well as in cryopreservation protocols.
为卵母细胞和卵巢皮质组织设计合理的冷冻保存策略，需要深入了解冷冻保护剂（CPA）渗透的速率以及伴随的水传输情况。本研究的目的是研究不同 CPA（即甘油/GLY、乙二醇/EG、二甲基亚砜/DMSO 和丙二醇/PG）在马卵母细胞和卵巢组织中的渗透动力学是否存在差异。
通过视频显微镜对卵母细胞在各向异性和 CPA 溶液中灌注时的体积响应进行成像，推断出水和 CPA 通过卵母细胞膜的渗透率（Lp 和 Ps）。通过渗透压计和重量测量来监测孵育过程中 CPA 向卵巢组织扩散和组织脱水的情况，以得出 CPA 扩散系数（D）。
发现卵母细胞对 CPA 的膜通透性按 GLY<EG<DMSO<PG 的顺序增加。在各向异性溶液中，水的渗透率高于 CPA 溶液，表明 CPA 改变了膜通透性特性。CPA 在卵巢组织中的扩散顺序为 GLY、PG<EG、DMSO。发现随着接触到的 CPA 浓度的增加，组织脱水增加，这与 CPA 扩散率成反比。
总之，本研究表明，CPA 通过膜双层的运动速率取决于与决定 CPA 在组织中运动的不同物理屏障因素。
本研究中提出的参数可应用于描述细胞和组织中溶质和水传输的模型，以及冷冻保存方案。
