Lee Sang-Goo, Park Chi-Hun, Choi Don-Ho, Kim Hye-Sun, Ka Hak-Hyun, Lee Chang-Kyu
Department of Food and Animal Biotechnology, College of Agriculture and Life Science, Seoul National University, Seoul, Korea.
Mol Reprod Dev. 2007 Nov;74(11):1436-45. doi: 10.1002/mrd.20728.
In pigs, the morphology and cell number of in vitro-produced blastocysts are inferior to those of their in vivo counterparts. The objective of this study was to increase developmental competence and to gain an understanding of cell allocation in blastocysts derived from the aggregation of four-cell stage porcine embryos produced in vitro. After removal of the zona pellucida, two (2x) and three (3x) four-cell stage embryos were aggregated by co-culturing them in aggregation plates. Five days after aggregation, the developmental ability and the number of cells in the aggregated embryos were determined. The percentage of blastocysts was higher (P < 0.05) in both the 2x and 3x aggregated embryos (66.6% and 72.0%, respectively) compared to that of the 1x embryos and the intact controls (43.1% and 36.4%, respectively). The total cell number of blastocysts also increased in aggregated embryos compared to that of intact controls (2.6-fold for 2x and 3.4-fold for 3x) (P < 0.05). The cells of two differentially stained embryos were started to mix at 72 hr after aggregation. In vitro-fertilized porcine aggregates (2x) were developed to blastocyst with a random distribution of cells from each embryo. The mRNA levels for the oct-4, bcl-xL and connexin 43 genes were higher (P < 0.05) and bak gene were lower (P < 0.05) in both the 2x and 3x aggregated embryos than the intact controls. Therefore, the aggregation of the four-cell stage embryos could be used to improve the quality of porcine preimplantation stage embryos produced in vitro.
在猪中，体外产生的囊胚的形态和细胞数量低于体内产生的囊胚。本研究的目的是提高发育能力，并了解体外产生的四细胞期猪胚胎聚集形成的囊胚中的细胞分配情况。去除透明带后，将两个（2x）和三个（3x）四细胞期胚胎在聚集板中共同培养进行聚集。聚集五天后，测定聚集胚胎的发育能力和细胞数量。与单胚胎组（1x）和完整对照组（分别为43.1%和36.4%）相比，2x和3x聚集胚胎中的囊胚百分比更高（P < 0.05）（分别为66.6%和72.0%）。与完整对照组相比，聚集胚胎中囊胚的总细胞数也增加了（2x为2.6倍，3x为3.4倍）（P < 0.05）。聚集后72小时，两个不同染色胚胎的细胞开始混合。体外受精的猪胚胎聚集体（2x）发育成囊胚，每个胚胎的细胞随机分布。2x和3x聚集胚胎中oct-4、bcl-xL和连接蛋白43基因的mRNA水平均高于完整对照组（P < 0.05），而bak基因水平低于完整对照组（P < 0.05）。因此，四细胞期胚胎的聚集可用于提高体外产生的猪植入前阶段胚胎的质量。
