Water and DMSO membrane permeability characteristics of in-vivo- and in-vitro-derived and cultured murine oocytes and embryos
Water and DMSO membrane permeability characteristics of in-vivo- and in-vitro-derived and cultured murine oocytes and embryos
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DOI：
10.1093/molehr/4.1.51
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发表时间：
1998-01-01
影响因子：
4
通讯作者：
Critser, JK
中科院分区：
文献类型：
--
作者：
Pfaff, RT;Liu, J;Critser, JK
虽然胚胎冷冻保存是许多哺乳动物物种的常规，重要的是要知道这些细胞的基本低温生物学如何随着发育而变化;进行性卵裂分裂导致可用于水和冷冻保护剂质量运输的卵裂球表面积减少。因此，测定了鼠卵母细胞、受精卵、2-细胞、4-细胞和8-细胞胚胎对水（L-p）和二甲基亚砜（P-DMSO）的膜渗透性以及反射系数σ（σ）。回收卵母细胞或受精卵，去除卵丘细胞，然后培养直至使用。将卵母细胞和胚胎固定并在24 ℃下用处理溶液灌注。将诱导的细胞体积随时间的变化录像，然后进行图像分析。在二甲亚砜（DMSO）存在下，卵母细胞、受精卵、2、4和8细胞胚胎的L-p值分别为0.77、0.81、0.94、0.86和1.10 μ m/min/atm，P-DMSO值分别为1.85、2.04、2.41、1.95和1.25 × 10（-3）cm/min。有DMSO存在时的L_i值显著高于无DMSO存在时的L_i值（P < 0.05）。将整个胚胎作为单一渗透实体处理导致相对于基于单个卵裂球的测量的P-DMSO估计值显著（P < 0.05）升高。这些数据表明，L-p和P-DMSO的估计值较低时，预测的单个卵裂球的基础上。数据还表明，L-p和P-DMSO在卵母细胞、合子、2-细胞和4-细胞胚胎中均无差异。然而，8-细胞阶段的显著较高的L-p和较低的P-DMSO支持基本的低温生物学差异可能需要发育阶段特异性胚胎冷冻保存方案的假设。
Although embryo cryopreservation is routine for many mammalian species, it is important to know how the fundamental cryobiology of these cells changes with development; Progressive cleavage divisions result in a reduction in the blastomere surface area available for water and cryoprotectant mass transport. Therefore, the membrane permeability of murine oocytes, zygotes, 2-cell, 4-cell, and 8-cell embryos to water (L-p), and dimethylsulphoxide (P-DMSO), and the reflection coefficient, sigma (sigma) were determined. Oocytes or zygotes were recovered, cumulus cells removed, then cultured until use. Oocytes and embryos were immobilized and perfused with treatment solutions at 24 degrees C. osmotically induced cell volume changes over time were videotaped followed by image analysis. The L-p values in the presence of dimethylsulphoxide (DMSO) were 0.77, 0.81, 0.94, 0.86, and 1.10 mu m/min/atm, and the P-DMSO values were 1.85, 2.04, 2.41, 1.95, and 1.25x 10(-3) cm/min for oocytes, zygotes, 2, 4, and 8-cell embryos respectively. The L, values in the presence of DMSO were significantly (P < 0.05) higher than those in the absence of DMSO. Treating the whole embryo as a single osmotic entity leads to significantly (P < 0.05) elevated P-DMSO estimates relative to those based upon measurements of individual blastomeres. These data indicate that both L-p and P-DMSO estimates are lower when predicted on an individual blastomere basis. The data also show that neither L-p nor P-DMSO differ among oocytes, zygotes, 2-cell and 4-cell embryos. However, the significantly higher L-p and lower P-DMSO of the 8-cell stage support the hypothesis that fundamental cryobiological differences may require developmental stage-specific embryo cryopreservation protocols.