Laboratory of Cytogenetics and Genome Research, Center of Human Genetics, KU Leuven, Leuven 3000, Belgium.
Institute of Bio- and Translational Medicine, University of Tartu, Tartu 50411, Estonia.
Hum Reprod. 2017 Nov 1;32(11):2348-2357. doi: 10.1093/humrep/dex286.
Is the rate and nature of chromosome instability (CIN) similar between bovine in vivo-derived and in vitro-cultured cleavage-stage embryos?
There is a major difference regarding chromosome stability of in vivo-derived and in vitro-cultured embryos, as CIN is significantly lower in in vivo-derived cleavage-stage embryos compared to in vitro-cultured embryos.
CIN is common during in vitro embryogenesis and is associated with early embryonic loss in humans, but the stability of in vivo-conceived cleavage-stage embryos remains largely unknown.
STUDY DESIGN, SIZE, DURATION: Because human in vivo preimplantation embryos are not accessible, bovine (Bos taurus) embryos were used to study CIN in vivo. Five young, healthy, cycling Holstein Friesian heifers were used to analyze single blastomeres of in vivo embryos, in vitro embryos produced by ovum pick up with ovarian stimulation (OPU-IVF), and in vitro embryos produced from in vitro matured oocytes retrieved without ovarian stimulation (IVM-IVF).
PARTICIPANTS/MATERIALS, SETTING, METHODS: Single blastomeres were isolated from embryos, whole-genome amplified and hybridized on Illumina BovineHD BeadChip arrays together with the bulk DNA from the donor cows (mothers) and the bull (father). DNA was also obtained from the parents of the bull and from the parents of the cows (paternal and maternal grandparents, respectively). Subsequently, genome-wide haplotyping and copy-number profiling was applied to investigate the genomic architecture of 171 single bovine blastomeres of 16 in vivo, 13 OPU-IVF and 13 IVM-IVF embryos.
The genomic stability of single blastomeres in both of the in vitro-cultured embryo cohorts was severely compromised (P < 0.0001), and the frequency of whole chromosome or segmental aberrations was higher in embryos produced in vitro than in embryos derived in vivo. Only 18.8% of in vivo-derived embryos contained at least one blastomere with chromosomal anomalies, compared to 69.2% of OPU-IVF embryos (P < 0.01) and 84.6% of IVM-IVF embryos (P < 0.001).
Genotyping data obtained in this study has been submitted to NCBI Gene Expression Omnibus (GEO; accession number GSE95358).
There were two main limitations of the study. First, animal models may not always reflect the nature of human embryogenesis, although the use of an animal model to investigate CIN was unavoidable in our study. Second, a limited number of embryos were obtained, therefore more studies are warranted to corroborate the findings.
Although CIN is also present in in vivo-developed embryos, in vitro procedures exacerbate chromosomal abnormalities during early embryo development. Hence, the present study highlights that IVF treatment compromises embryo viability and should be applied with care. Additionally, our results encourage to refine and improve in vitro culture conditions and assisted reproduction technologies.
STUDY FUNDING/COMPETING INTEREST(S): The study was funded by the Agency for Innovation by Science and Technology (IWT) (TBM-090878 to J.R.V. and T.V.), the Research Foundation Flanders (FWO; G.A093.11 N to T.V. and J.R.V. and G.0392.14 N to A.V.S. and J.R.V.), the European Union's FP7 Marie Curie Industry-Academia Partnerships and Pathways (IAPP, SARM, EU324509 to J.R.V., T.V., O.T, A.D., A.S. and A.K.) and Horizon 2020 innovation programme (WIDENLIFE, 692065 to J.R.V., O.T., T.V., A.K. and A.S.). M.Z.E., J.R.V. and T.V. are co-inventors on a patent application ZL913096-PCT/EP2014/068315-WO/2015/028576 ('Haplotyping and copy-number typing using polymorphic variant allelic frequencies'), licensed to Cartagenia (Agilent Technologies).
体内和体外培养的牛胚胎的染色体不稳定性（CIN）的速率和性质是否相似？
体内和体外培养的胚胎在染色体稳定性方面存在显著差异，与体外培养的胚胎相比，体内胚胎的 CIN 明显较低。
CIN 在体外胚胎发生过程中很常见，并且与人类早期胚胎丢失有关，但体内受精胚胎的稳定性在很大程度上仍不清楚。
研究设计、规模、持续时间：由于无法获得人类体内的胚胎，因此使用牛（Bos taurus）胚胎来研究体内胚胎的 CIN。使用五头年轻、健康、循环的荷斯坦弗里森奶牛分析体内胚胎、通过卵母细胞采集和卵巢刺激（OPU-IVF）产生的体外胚胎，以及未经卵巢刺激（IVM-IVF）从体外成熟的卵母细胞中产生的体外胚胎的单个卵裂球。
参与者/材料、设置、方法：从胚胎中分离单个卵裂球，全基因组扩增，并与供体牛（母亲）和公牛（父亲）的 bulk DNA 一起在 Illumina BovineHD BeadChip 阵列上杂交。还从公牛和奶牛的父母（分别为祖父和祖母）获得了 DNA。随后，应用全基因组单体型和拷贝数分析来研究 16 个体内、13 个 OPU-IVF 和 13 个 IVM-IVF 胚胎的 171 个单个牛卵裂球的基因组结构。
两个体外培养胚胎队列的单个卵裂球的基因组稳定性都严重受损（P < 0.0001），并且在体外产生的胚胎中染色体或片段异常的频率高于体内产生的胚胎。与 OPU-IVF 胚胎（P < 0.01）和 IVM-IVF 胚胎（P < 0.001）相比，只有 18.8%的体内胚胎含有至少一个具有染色体异常的卵裂球。
本研究获得的基因分型数据已提交给 NCBI Gene Expression Omnibus（GEO；注册号 GSE95358）。
该研究有两个主要限制。首先，动物模型可能并不总是反映人类胚胎发生的性质，尽管在我们的研究中，使用动物模型来研究 CIN 是不可避免的。其次，获得的胚胎数量有限，因此需要进行更多的研究来证实这些发现。
虽然 CIN 也存在于体内发育的胚胎中，但体外程序会加剧早期胚胎发育过程中的染色体异常。因此，本研究强调了体外受精治疗会损害胚胎活力，应谨慎应用。此外，我们的研究结果鼓励改进和优化体外培养条件和辅助生殖技术。
研究基金/利益冲突：该研究由创新署（IWT）（J.R.V. 和 T.V. 的 TBM-090878，A.V.S. 和 J.R.V. 的 G.A093.11 N）、佛兰德斯研究基金会（FWO；G.0392.14 N 给 T.V. 和 J.R.V. 和 A.V.S. 和 J.R.V.）、欧盟的 FP7 玛丽居里行业-学术界伙伴关系和途径（IAPP、SARM、EU324509 给 J.R.V.、T.V.、O.T.、A.D.、A.S. 和 A.K.）和地平线 2020 创新计划（WIDENLIFE，692065 给 J.R.V.、O.T.、T.V.、A.K. 和 A.S.）资助。M.Z.E.、J.R.V. 和 T.V. 是一项专利申请 ZL913096-PCT/EP2014/068315-WO/2015/028576（“使用多态变体等位基因频率进行单体型和拷贝数分型”）的共同发明人，该专利已授权给 Cartagenia（安捷伦科技）。
