Development of Pluripotential Embryonic Stem Cells from Rat Blastocyst

为了建立大鼠胚胎多能二倍体细胞，我们将大鼠囊胚培养在含有小鼠白血病抑制因子（LIF）和胰岛素样生长因子（IGF）-Ⅱ的培养基中。小鼠LIF（5，000单位/ml）和大鼠IGF-Ⅱ（100 ng/ml）的联合作用促进了大鼠囊胚内细胞团（ICM）的生长。ICM来源的细胞形态与小鼠胚胎干细胞相似。我们建立了9和3线ACI和杂交（ACI * Wistar-Imamichi）大鼠，分别。为了评价大鼠ES样细胞的体内多能性，将Wistar-Imamichi的8细胞胚胎与建立的细胞共培养以制备嵌合大鼠。在ACI大鼠细胞系上，从349个嵌合体胚胎植入假孕大鼠体内获得25个新生儿。在新生大鼠中，未观察到嵌合体大鼠。通过显微注射法，将已建立的细胞与Wistar-Imamichi囊胚进行嵌合。在来自ACI和hyberid大鼠的细胞系上，产生了304个嵌合体胚胎。然而，在206只新生儿中未发现嵌合体大鼠。这些发现表明，除了LIF和IGF-II之外，可能还需要其他因子来从大鼠胚胎中建立多能ES细胞。

To establish pluripotential diploid cells from rat embryos, we cultured blastocysts in medium containing mouse leukemia inhibiting factor (LIF) and insulin-like growth factor (IGF) -II.Combination of mouse LIF (5,000 units/ml) and rat IGF-II (100ng/ml) promoted the growth of inner cell mass (ICM) of rat blastocyst. The form of the cells derived from ICM was similar to that of mouse embryonic stem (ES) cell. We established 9 and 3 lines on ACI and a hybrid (ACI * Wistar-Imamichi) rat, respectively. For evaluating in vivo pluripotency of the rat ES-like cells, 8-cell embryos of Wistar-Imamichi were co-cultured with the established cells to make chimeric rats. On the cell lines derived from ACI rat, 25 neonates were obtained from 349 chimeric embryos that implanted into pseudopregnancy rat. On the neonates, no chimeric rat was observed. By microinjection method, chimeric embryos were produced between the established cells and blastocysts of Wistar-Imamichi. On the cell lines derived from both ACI and the hyberid rats, 304 chimeric embryos were produced. Chimeric rats were not, however, founded in 206 neonates. These findings suggest that, in addition to LIF and IGF-II,other factor may be needed to establish pluripotential ES cells from rat embryos.