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

我们研究了用微胶囊法冷冻保存和体外成熟未成熟人卵泡的可能性。此外，我们还通过分析卵母细胞因子来探讨卵泡发育的机制。得到了以下结果：采用玻璃化冷冻方法（10% EG和15% EG+15% DMSO+0.5M蔗糖）成功保存人未受精卵母细胞。然而，囊化的未成熟卵泡未在体外生长。证实GDF-9、CD9、CD98、C-kit mRNA在人未受精卵母细胞中表达。这些结果可能提示这种方法在保存未成熟卵泡方面的未来应用。在我们的研究中观察到HB-EGF的动态和细胞特异性表达模式表明HB-EGF可能是潜在的重要的卵泡发育局部调节剂。我们关于精子-卵母细胞相互作用的研究结果支持了一个假设，即整合素抗体可以抑制精子胚乳的一种结合机制，但这种机制在结合和融合过程中并不起重要作用。我们对导致男性不育的遗传机制的研究表明，OPPO1和MEI1可能在人类精子发生中发挥作用。我们研究了钙离子载体和嘌呤霉素联合作用下老年人卵母细胞活化的核动力学。由此可见，孤雌活化的人卵母细胞的核动力学在体外老化卵母细胞中表现得更为迅速。虽然没有成功地利用包裹的人类未成熟卵泡获得体外成熟，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。

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.