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Basic study in cryopreservation and chromosomal analysis of single blastomeres.

单卵裂球冷冻保存和染色体分析的基础研究。

基本信息

批准号：
02670728
负责人：
SENGOKU Kazuo
金额：
$ 1.41万
依托单位：
ASAHIKAWA MEDICAL COLLEGE
依托单位国家：
日本
项目类别：
Grant-in-Aid for General Scientific Research (C)
财政年份：
1990
资助国家：
日本
起止时间：
1990 至 1992
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-02670728/
关键词：
embryobiopsy cryopreservation chromosomal analysis blastomeres

项目摘要

Biopsy of the preimplantation embryo has wide application as a tool to investigate the developmental potential of the preimplantation embryo and also offered a possibility for prenatal diagnosis. The purpose of this study were first, to examine the developmental potential of single blastomeres of four-and eight-cell mouse embryos, and second to investigate the possibility of successful cryopreservation and karyotyping of single blastomeres.(1) Removal of a single blastomere from 4- and 8-cell mouse embryos by extrusion technique did not affect development to the blastocyst stage in vitro (80% of control,75% and 78.3% of biopsied embryo at 4- and 8-cell stage, respectively). However,after transfer to pseudopregnant recipient mice, the implantation rates of biopsied embryos were significantly lower than intact-controls (24.3% at 4-cell, 34.8% at 8-cell and 50% at control, respectively).(2) Most single blastomeres had undergone several cell divisions in vitro. After a short culture of single blastomeres with subsequent exposure to colcemid for 16 hours, 27.3% of 4-cell and 52.2% of 8-cell biopsies were karyotyped. However, 20 to 30% of biopsied blstomeres were lost through cell death and technical problems.(3) Biopsied single blastomeres could not be successfully cryopreserved when the freezing medium contained 1.5M DMSO+0.25M sucrose and 5-step dilution method was used. The techniques developed in this study may provide the useful tool for the study of prenatal diagnosis.

植入前胚胎活检作为研究植入前胚胎发育潜力的工具具有广泛的应用，也为产前诊断提供了可能性。本研究的目的一是考察四细胞和八细胞小鼠胚胎单卵裂球的发育潜力，二是探讨单卵裂球成功冷冻保存和核型分析的可能性。（1）通过挤压技术从4和8细胞小鼠胚胎中去除单卵裂球不影响体外发育至囊胚阶段（对照的80%，活检的75%和78.3%）分别为 4 细胞阶段和 8 细胞阶段的胚胎）。然而，移植到假孕受体小鼠后，活检胚胎的植入率显着低于完整对照（4细胞为24.3％，8细胞为34.8％，对照为50％）。（2）大多数单卵裂球在体外经历了多次细胞分裂。对单个卵裂球进行短暂培养并随后暴露于秋水仙胺 16 小时后，27.3% 的 4 细胞和 52.2% 的 8 细胞活检进行了核型分析。然而，由于细胞死亡和技术问题，有20%～30%的活检卵裂球丢失。(3)当冷冻介质含有1.5M DMSO+0.25M蔗糖并采用5步稀释法时，活检的单个卵裂球无法成功冷冻保存。本研究开发的技术可为产前诊断的研究提供有用的工具。