BASIC STUDIES FOR IMPROVEMENT OF IN VITRO FERTILIZATION AND CRYOPRESERVATION

改进体外受精和冷冻保存的基础研究

• 批准号: 10671518
• 负责人: TSUTSUMI Osamu
• 金额: $ 2.18万
• 依托单位: UNIVERSITY OF TOKYO
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 1998
• 资助国家: 日本
• 起止时间: 1998 至 1999
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-10671518/
• 关键词: embryo; cryopreservation; in vitro fertilization; glucose; vitirification; GLUT; implantation; cryopreservatiopn; vitirification; dioxin; estrogen receptor; embryo; cryopreservation; in vitro fertilization; glucose; vitirification; GLUT; implantation; cryopreservatiopn; vitirification; dioxin; estrogen receptor

Effects of two cryopreservation procedures (conventional slow controlled-rate freezing using a programmable freezer and vitrification by direct plunging into liquid nitrogen) were compared on 2-cell embryos and their subsequent development to blastocysts, fresh or cryopreserved 2-cell mouse embryos were developed into blastocysts in vitro. The percentage of vitrified embryos which developed into blastocysts was significantly lower than that of fresh and slow controlled-rate frozen embryos. Although blastocysts from each cryopreservation procedure appeared morphologically normal and neither number of cells in the blastocysts nor in-vitro trophoblast spreading differed significantly, there were significant differences in their functional viability. First, the glucose incorporation activity in terms of [(3)H]2-deoxyglucose (2-DG) uptake in vitrified and thawed 2- cell embryos significantly decreased compared with fresh or slow controlled-rate frozen and thawed 2-cell embryos. Second, 2-DG uptake by blastocysts developed in vitro from fresh 2-cell embryos and from slow controlled-rate frozen or vitrified 2-cell embryos was 105 +/- 75, 43.0 +/- 28.3 and 22.O +/- 11.4 fmol/embryo/h respectively. Third, the implantation rate of blastocysts developed in vitro from vitrified 2-cel1 embryos (10.2%) was significantly lower than that from fresh 2-cell embryos (30.8%) or slow controlled-rate frozen 2-cell embryos (22.1%). Since these data suggest that cryopreservation may have ulterior consequences on the functional development of embryos and that vitrification may exert a more harmful effect than slow controlled-rate freezing, more attention should be paid to its safety before vitrification is used routinely in a clinical programme.

在2个细胞胚胎上比较了两种冷冻保存程序（使用可编程冰柜的常规缓慢控制率冻结和直接浸入液氮中的玻璃化）的影响，然后将其随后开发到胚泡到新鲜或冷冻保存的2细胞小鼠胚胎中，将其发展为胚泡中的胚泡。发展成胚泡的玻璃化胚胎的百分比明显低于新鲜和缓慢的控制率冷冻胚胎的百分比。尽管每个冷冻保存过程中的胚泡在形态上似乎正常，并且胚泡中的细胞数量均未发生，也没有滋养细胞的扩散差异很大，但其功能可行性存在显着差异。首先，与新鲜或缓慢的冷冻和解冻的2细胞胚胎相比，葡萄糖掺入活性在玻璃化和解冻的2-细胞胚胎中的摄取和解冻的2-细胞胚胎的摄取显着降低，并解冻2-细胞胚胎的摄取显着降低。其次，胚泡的2-DG摄取从新鲜的2细胞胚胎和缓慢控制率冷冻或玻璃化的2-cell胚胎的体外开发，分别为105 +/- 75、43.0 +/- 28.3和22.o +/- 11.4 fmol/embryo/h。第三，从玻璃化的2-CEL1胚胎（10.2％）在体外开发的胚泡的植入率明显低于新鲜2细胞胚胎（30.8％）或缓慢的控制率冷冻2细胞胚胎（22.1％）。由于这些数据表明冷冻保存可能会对胚胎的功能发展产生别有用心的后果，并且玻璃化可能比缓慢的控制率冻结更有害效果，因此在临床计划中经常使用玻璃化之前，应更加注意其安全性。