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ES cell line derived from a single blastomere in cynomolgus monkey

机译：食蟹猴中单个卵裂球来源的ES细胞系

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The establishment of most ES cell lines requires the destruction of embryos. Some ES cell lines in mice and humans are currently derived from a single blastomere (BM), so that remaining BMs can still develop into fetuses. The objective of this study was to devise a method simpler than those previously described for establishing ES cell lines from a single BM in the cynomolgus monkey. Controlled ovarian stimulation and oocyte recovery have been described previously by Torii et al. (2000). Mature oocytes were fertilized by ICSI. The zona pellucida of 4- to 5-cell-stage embryos was disrupted and individual BMs were separated. These BMs were cultured on MEF and ES medium containing ACTH. Cells were also examined in more detail using the pluripotency-cell-specific markers Oct-4, SSEA-4, TRA-1-60 and TRA-1-81. Moreover, Cells were cultured in suspension to form EBs, and induced spontaneous differentiation in vitro. Differentiated cells were characterized using antibodies against α-Fetoprotein, Myosin Heavy Chain and Tuj. One stable ES cell line was obtained from a 4-cell-stage embryo using ES medium containing ACTH. The ES cell line was passaged more than thirty times. The ES cells showed normal karyotype and retained pluripotency markers of primate ES. We are presently confirming teratomas formation in vivo. Here we showed that cynomolgus monkey ES cell could be derived from a single BM. This method allows the establishment of ES cell lines from a single BM, leaving the other BMs available for embryo transfer. Thus, the method described here is simpler than previously described methods and alleviates some ethical concerns.
机译：大多数ES细胞系的建立需要破坏胚胎。小鼠和人类中的某些ES细胞系目前源自单个卵裂球（BM），因此剩余的BM仍可发育成胎儿。这项研究的目的是设计一种比以前描述的方法更简单的方法，用于从食蟹猴中的单个BM建立ES细胞系。 Torii等人先前已经描述了可控制的卵巢刺激和卵母细胞恢复。 （2000）。通过ICSI使成熟的卵母细胞受精。 4至5个细胞期胚胎的透明带被破坏，分离出单个BM。这些BM在含有ACTH的MEF和ES培养基上培养。还使用多能细胞特异性标记Oct-4，SSEA-4，TRA-1-60和TRA-1-81对细胞进行了更详细的检查。此外，将细胞悬浮培养以形成EB，并在体外诱导自发分化。使用针对α甲胎蛋白，肌球蛋白重链和Tuj的抗体表征分化的细胞。使用含有ACTH的ES培养基，从4细胞期的胚胎中获得一种稳定的ES细胞系。 ES细胞系传代超过三十次。 ES细胞显示出正常的核型，并保留了灵长类ES的多能性标记。我们目前正在确认体内畸胎瘤的形成。在这里，我们显示了食蟹猴ES细胞可以源自单个BM。此方法允许从单个BM建立ES细胞系，而其他BM可用于胚胎移植。因此，这里描述的方法比先前描述的方法更简单，并且减轻了一些道德上的顾虑。

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