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The basic study of cryopreservation and in vitro maturation of encapsulated human immature follicles

封装的人未成熟卵泡的冷冻保存和体外成熟的基础研究

基本信息

批准号：
15591725
负责人：
SENGOKU Kazuo
金额：
$ 2.18万
依托单位：
Asahikawa Medical College
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
2003
资助国家：
日本
起止时间：
2003 至 2005
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-15591725/
关键词：
immature follicle microcapsule cryopreservation in vitro culture spermatogenesis aging 卵子活性化体外成熟

项目摘要

We studied the possibility of cryopreservation and in vitro maturation of immature human ovarian follicles using microcapsule methods.In addition, we investigated the mechanisms of follicular development by analyzing the oocyte factor.The following results were obtained.1.Human unfertilized oocytes were successfully cryopreserved with vitrification methods (10% EG and 15% EG+15% DMSO+0.5M sucrose). However, in vitro growth of encapsulated immature follicles were not obtained. The expression of GDF-9,CD9,CD98, C-kit mRNA were confirmed in human un fertilized oocytes. These results may suggest the possible future application of this methods for preservation immature follicles2.The dynamic and cell-specific expression pattern of HB-EGF observed in our study suggests that HB-EGF may be potentially important local regulator of follicular development.3.Our results regarding sperm-oocyte interaction support the hypothesis that one of the sperm oolemmal binding mechanism can be inhibited by integrin antibodies but that this mechanism does not play an essential role in the binding and fusion process4.Our studies regarding the genetic mechanisms implicate as a cause of male infertility suggest that OPPO1 and MEI1 may play a role in human spermatogenesis.5.We investigated the nuclear kinetics of oocyte activation of aged human oocytes following combined treatment with calcium ionophore and puromycin. It is concluded that the nuclear kinetics of parthenogenetically activated human oocytes appears to be more rapid in in vitro aging oocytes.Although successful in vitro maturation was not obtained using encapsulated human immature follicles, our preliminary results may offer important aspects of understanding the mechanisms of human follicular development.

本研究采用微囊法对人未成熟卵泡进行冷冻保存和体外成熟培养，并通过对卵母细胞因子的分析，探讨卵泡发育的机制，主要结果如下：1.采用玻璃化冷冻法（10%EG和15%EG +15%DMSO +0.5M蔗糖）成功地冷冻了人未受精卵母细胞。然而，在体外生长的包裹未成熟的卵泡没有得到。GDF-9、CD 9、CD 98、C-kit mRNA在未受精卵中均有表达。这些结果可能提示这种方法在未来可能用于保存未成熟卵泡2.我们研究中观察到的HB-EGF的动态和细胞特异性表达模式表明HB-EGF可能是卵泡发育的潜在重要局部调节剂。卵母细胞的相互作用支持这一假说，即精子的卵膜结合机制之一，可以抑制整合素抗体，但这一点，OPPO 1和MEI 1可能在人类精子发生中发挥作用。5.我们研究了钙离子载体和嘌呤霉素联合处理老化人类卵母细胞后卵母细胞激活的核动力学。结论：孤雌激活的人卵母细胞的核动力学在体外老化的卵母细胞中表现得更快，尽管未成熟卵泡的体外成熟没有获得成功，但我们的初步结果可能为理解人类卵泡发育的机制提供重要方面。