Investigating Pre-Implantation Chromosomal Instability in Assisted Reproduction

研究辅助生殖中植入前染色体不稳定性

• 批准号: 9173739
• 负责人: Shawn L. Chavez
• 金额: $ 36.31万
• 依托单位: OREGON HEALTH & SCIENCE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-07-24 至 2021-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9173739
• 关键词: Address; Aneuploidy; Arts; Behavior; Bioinformatics; Biopsy; Birth Rate; Cell Death; Cells; Chromosomal Instability; Chromosomal Rearrangement; Chromosomal Stability; Chromosome abnormality; Chromosomes; Complex; Conceptions; Couples; DNA; DNA Microarray Chip; Data; Development; Embryo; Embryo Transfer; Embryonic Development; Encapsulated; Event; Excision; Exhibits; Female; Fertilization failure; Fertilization in Vitro; Frequencies; GC gene; Generations; Genetic Materials; Genomics; Human; Image; In Vitro; Incidence; Individual; Infertility; Inherited; Link; Live Birth; Macaca mulatta; Malignant Neoplasms; Maternal Age; Measures; Meiosis; Microsatellite Repeats; Mitosis; Mitotic; Modeling; Monitor; Mosaicism; Mus; Oocytes; Pregnancy; Primates; Principal Investigator; Process; Reproductive Physiology; Resolution; Risk; Sequence Analysis; Source; Staging; Study models; Techniques; Therapeutic Intervention; Time; Translating; Translations; Woman; Work; abstracting; assisted reproduction; base; blastomere structure; embryo stage 2; genetic analysis; improved; micronucleus; next generation sequencing; nonhuman primate; oocyte maturation; preimplantation; programs; response; segregation; success; time interval; whole genome; zygote

Program Director/Principal Investigator (Last, First, Middle): Chavez, Shawn L. Project Summary/Abstract Since its introduction over 35 years ago, human in vitro fertilization (IVF) has assumed great promise for infertile couples, but success rates have remained only ~30% worldwide for several decades. One of the primary reasons for this is that whole chromosomal abnormalities, or aneuploidy, are incredibly common in cleavage-stage human embryos. Previously, we demonstrated that assessing the time intervals of the first three mitotic divisions in conjunction with a phenomenon called cellular fragmentation, which is frequently observed in human embryos as well as following natural conception, largely distinguishes chromosomally normal and abnormal cleavage-stage human embryos. We also determined that cellular fragments might contain genetic material that likely began as mis-segregated chromosomes were encapsulated into micronuclei during meiosis and/or mitosis. Although cellular fragmentation is closely linked with aneuploidy generation and micronuclei formation, the source of these fragments and their precise chromosomal content is not well defined. In addition, whether embryos from other mammalian species more closely related to humans such as non-human primates have a similar aneuploidy frequency remains unknown and addressing this question is essential for potential translation to early human embryogenesis. Our preliminary data reveals that rhesus cleavage-stage embryos also exhibit a high degree of aneuploidy, fragmentation, and micronucleation as well as similar mitotic timing when compared to human. Given that humans and the rhesus monkey are also highly similar in terms of female reproductive physiology and fundamental aspects of early embryogenesis, we propose to investigate aneuploidy and the fate of mis-segregated chromosomes in rhesus embryos to model human pre-implantation development. By applying whole-genome next-generation sequencing (NGS) for comprehensive chromosomal assessment, we will first determine the frequency of aneuploidy and sub- chromosomal errors during meiosis in individual mature rhesus oocytes and zygotes and potential correction upon chromosome-induced polar body extrusion. Using a combination of NGS and non-invasive time-lapse imaging to monitor early cleavage divisions and cellular fragmentation dynamics, we will then evaluate the incidence of mitotic chromosomal mis-segregation up to the ~8-cell stage and reconstruct all whole and sub- chromosomal errors in each rhesus embryo by analyzing the genetic content of both single cells and fragments. Lastly, we will assess the potential contribution of meiotic chromosomal mis-segregation to mitotic errors and subsequent development by performing polar body biopsy on zygotes, allowing the embryo to proceed until the ~8-cell stage, and distinguishing meiotic versus mitotic errors based on chromosomal mosaicism, fragmentation timing, and microsatellite analysis. This work will greatly contribute to our knowledge of normal primate embryogenesis with additional implications for translational application to human infertility and IVF treatment. OMB No. 0925-0001/0002 (Rev. 08/12 Approved Through 8/31/2015) Page Continuation Format Page

项目主任/首席研究员（最后，第一，中间）：查韦斯，肖恩L。 项目总结/摘要 自35年前引入以来，人类体外受精（IVF）已经为人类的生殖健康带来了巨大的希望。 不孕夫妇，但成功率一直保持只有约30%，几十年来全世界。之一 其主要原因是整个染色体异常，或非整倍体，是令人难以置信的普遍， 卵裂期的人类胚胎以前，我们证明，评估第一次的时间间隔， 三次有丝分裂与一种称为细胞分裂的现象相结合，这种现象经常被称为细胞分裂。 在人类胚胎以及自然受孕后观察到， 正常和异常的卵裂期人类胚胎。我们还确定细胞碎片可能 含有遗传物质，这些遗传物质可能开始于错误分离的染色体被封装到微核中， 在减数分裂和/或有丝分裂期间。尽管细胞碎片化与非整倍体的产生密切相关， 微核的形成，这些片段的来源及其精确的染色体内容是不好的 定义了此外，来自其他哺乳动物物种的胚胎是否与人类关系更密切， 非人类灵长类动物也有类似的非整倍体频率，这一点尚不清楚， 对早期人类胚胎发育的潜在翻译至关重要。我们的初步数据显示恒河猴 卵裂期胚胎也表现出高度的非整倍性、碎裂和微核 有丝分裂时间与人类相似。考虑到人类和恒河猴也高度依赖 在雌性生殖生理学和早期胚胎发生的基本方面相似， 建议研究恒河猴胚胎中的非整倍体和错误分离的染色体的命运， 人类胚胎植入前发育通过应用全基因组下一代测序（NGS）， 全面的染色体评估，我们将首先确定非整倍体和亚整倍体的频率， 恒河猴成熟卵母细胞和受精卵减数分裂过程中的染色体错误及其纠正 在染色体诱导的极体挤出后。使用NGS和非侵入性时间推移的组合 成像监测早期卵裂分裂和细胞碎片动力学，然后我们将评估 有丝分裂染色体错误分离的发生率高达~8-细胞期，并重建所有的整体和亚细胞。 通过分析单个细胞的遗传内容， 片段最后，我们将评估减数分裂染色体错误分离对有丝分裂的潜在贡献。 通过对受精卵进行极体活检，使胚胎 进行到~8-细胞阶段，并根据染色体区分减数分裂与有丝分裂错误 镶嵌性、片段化时间和微卫星分析。这项工作将大大有助于我们的知识 正常灵长类动物胚胎发生与翻译应用于人类不育症的额外影响 IVF治疗。 OMB编号0925-0001/0002（2012年8月批准至2015年8月31日修订版）页码续页格式页码