1 Research and Clinical Center for Infertility, Yazd Reproductive Sciences Institute, Shahid Sadoughi University of Medical Sciences, Yazd, Iran.
2 Stem Cell Biology Research Center, Yazd Reproductive Sciences Institute, Shahid Sadoughi University of Medical Sciences, Yazd, Iran.
Stem Cells Dev. 2019 Mar 1;28(5):303-309. doi: 10.1089/scd.2018.0168. Epub 2019 Jan 31.
In vitro embryo twinning can be used to increase the number of the human embryos available for production of human embryonic stem cell (hESC) lines. The aim of this study was to generate hESCs following the production of the twin embryos by in vitro embryo splitting procedures. In total 21 chromosomally abnormal (three pronuclei) embryos underwent in vitro embryo twinning and were allowed to develop to the blastocyst stage. As a result, 42 twin embryos were obtained, of which 24 developed to blastocyst stage. Using micromanipulation technique, the zona-free blastocysts were recovered and plated onto mitotically inactivated Yazd human foreskin fibroblast (Batch18; YhFF#18) feeder layers in microdrops. After 3 to 5 days of blastocyst culture onto human foreskin fibroblast feeder layers, the hESC-like outgrowths were passaged onto new feeders in microdrops. The initial outgrowths of hESC-like cells were generated, and cells were proliferated, passaged, and some of them expressed hESC and trophoblastic markers; however, no cell lines were established. This might be due to the low cell number and poor quality of inner cell mass within these twin blastocysts. In vitro embryo twinning by increasing the number of the human embryos could be useful in the future for the generation of new pluripotent stem cell lines. However, the challenge remains to optimize the methods.
体外胚胎分裂可用于增加人类胚胎的数量，从而生产人类胚胎干细胞（hESC）系。本研究的目的是通过体外胚胎分裂程序产生双胞胎胚胎后生成 hESC。总共对 21 个染色体异常（三原核）胚胎进行了体外胚胎分裂，并允许其发育至囊胚阶段。结果，获得了 42 对双胞胎胚胎，其中 24 对发育至囊胚阶段。使用显微操作技术，从无透明带的囊胚中回收并接种到微滴中的有丝分裂失活的 Yazd 人包皮成纤维细胞（Batch18；YhFF#18）饲养层上。在人类包皮成纤维细胞饲养层上培养囊胚 3 至 5 天后，将 hESC 样细胞集落传代到新的微滴饲养层上。产生了 hESC 样细胞的初始集落，并且细胞增殖、传代，其中一些表达 hESC 和滋养层标记物；然而，没有建立细胞系。这可能是由于这些双胞胎囊胚内细胞团的细胞数量少且质量差。通过增加人类胚胎的数量进行体外胚胎分裂，将来可能有助于生成新的多能干细胞系。然而，仍然需要优化方法。
