Development of human cloned blastocysts following somatic cell nuclear transfer with adult fibroblasts
Development of human cloned blastocysts following somatic cell nuclear transfer with adult fibroblasts
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DOI：
10.1634/stemcells.2007-0252
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发表时间：
2008-02-01
期刊：
影响因子：
5.2
通讯作者：
Wood, Samuel H.
中科院分区：
文献类型：
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作者：
French, Andrew J.;Adams, Catharine A.;Wood, Samuel H.
核移植干细胞在再生医学和基于细胞的药物发现领域具有相当大的前景。在这项研究中，共获得29个卵母细胞从三个年轻的（20 - 24岁）生殖卵子捐赠者谁已经成功在以前的周期。这些卵母细胞被预期的父母认为超过了他们的生殖需求，在获得知情同意后，卵子捐赠者和预期的父母都捐赠用于研究，没有经济补偿。所有预期的父母成功地实现了持续妊娠，保留卵母细胞用于生殖目的。经阴道抽吸后2小时内获得的成熟卵母细胞在细胞松弛素B中孵育45分钟后，使用挤出或抽吸两种方法之一来去核。两种方法的卵母细胞溶解或变性率没有差异。体细胞核移植（SCNT）胚胎构建使用两个建立的成年男性成纤维细胞系的正常核型。在标准体外受精培养基中孵育后，观察到原核形成率（66%）、早期卵裂率（47%）和囊胚发育率（23%）较高。通过DNA和线粒体DNA指纹分析证实了一个克隆囊胚，另外两个克隆囊胚的DNA指纹表明它们也是由SCNT产生的。也从有限数量的孤雌激活的卵母细胞获得囊胚。这项研究首次证明，SCNT可以使用从分化的成体细胞获得的细胞核产生人类囊胚期胚胎，并提供了有关人类核移植效率更高所需方法的新信息。
Nuclear transfer stem cells hold considerable promise in the field of regenerative medicine and cell-based drug discovery. In this study, a total of 29 oocytes were obtained from three young (20 - 24 years old) reproductive egg donors who had been successful in previous cycles. These oocytes, deemed by intended parents to be in excess of their reproductive needs, were donated for research without financial compensation by both the egg donor and intended parents after receiving informed consent. All intended parents successfully achieved ongoing pregnancies with the oocytes retained for reproductive purposes. Mature oocytes, obtained within 2 hours following transvaginal aspiration, were enucleated using one of two methods, extrusion or aspiration, after 45 minutes of incubation in cytochalasin B. Rates of oocyte lysis or degeneration did not differ between the two methods. Somatic cell nuclear transfer (SCNT) embryos were constructed using two established adult male fibroblast lines of normal karyotype. High rates of pronuclear formation (66%), early cleavage (47%), and blastocyst (23%) development were observed following incubation in standard in vitro fertilization culture media. One cloned blastocyst was confirmed by DNA and mitochondrial DNA fingerprinting analyses, and DNA fingerprinting of two other cloned blastocysts indicated that they were also generated by SCNT. Blastocysts were also obtained from a limited number of parthenogenetically activated oocytes. This study demonstrates, for the first time, that SCNT can produce human blastocyst-stage embryos using nuclei obtained from differentiated adult cells and provides new information on methods that may be needed for a higher level of efficiency for human nuclear transfer.