Narula A, Taneja M, Totey S M
Embryo Biotechnology Laboratory, National Institute of Immunology, Aruna Asaf Ali Marg, New Delhi, India.
Mol Reprod Dev. 1996 Jul;44(3):343-51. doi: 10.1002/(SICI)1098-2795(199607)44:3<343::AID-MRD8>3.0.CO;2-M.
The morphology and number of cells in the trophectoderm (TE) and inner cell mass (ICM) of buffalo blastocysts derived from in vitro fertilization and cultured in the presence or absence of insulin-like growth factor-I (IGF-I) were analyzed by differential fluorochrome staining technique. The total cell number (TCN), TE number, and ICM cell number were significantly higher in blastocysts developed in vitro in the presence of IGF-I as compared to blastocysts developed without IGF-I (P < 0.01). It was observed that the buffalo blastocyst took 5-9 days postfertilization to develop in vitro. In order to correlate the time required for blastocyst development and the allocation of cells to TE and ICM, blastocysts were designated as fast (developing on or before day 7) or slow (developing after day 7). The TCN, TE, and ICM cells of fast-developing blastocysts cultured in the presence of IGF-I were significantly higher than slow-developing blastocysts (P < 0.01). The blastocysts developed on day 6 had a mean total cell number 118.6 +/- 21.4, which significantly decreased to 85.6 +/- 17.4, 62.0 +/- 14.5, and 17.0 +/- 4.0 on days 7, 8, and 9, respectively (P < 0.05). Normal development of buffalo embryo showed that, on average, embryos reached compact morula stage at the earliest between days 4.5-5.5. Blastocysts developed, at the earliest, between days 5.0-6.0, and it took them, on average, 6.5 days to hatch from the zona pellucida. TCN, TE, and ICM increased three times from morula to blastocyst; however, the proportion of ICM to TCN remained the same, in both embryonic stages. TE approximately doubled in hatched blastocysts, as compared to unhatched blastocysts (P < 0.05). However, ICM cells were decreased. The time required for development of parthenogenetic blastocysts was observed to be greater as compared to in vitro fertilized (IVF) blastocysts. The total cell number of parthenogenetic blastocysts was 100.8 +/- 11.3, including 59.2 +/- 8.4 cells of TE and 42.1 +/- 6.9 cells of ICM.
采用荧光染料差异染色技术，分析了体外受精获得的水牛囊胚滋养外胚层（TE）和内细胞团（ICM）中细胞的形态和数量，这些囊胚在有或没有胰岛素样生长因子-I（IGF-I）的条件下进行培养。与未添加IGF-I培养发育的囊胚相比，添加IGF-I体外发育的囊胚的总细胞数（TCN）、TE细胞数和ICM细胞数显著更高（P<0.01）。观察到水牛囊胚在体外受精后5-9天发育。为了关联囊胚发育所需时间以及细胞向TE和ICM的分配情况，将囊胚分为快速发育型（在第7天或之前发育）或缓慢发育型（在第7天后发育）。在添加IGF-I条件下培养的快速发育囊胚的TCN、TE和ICM细胞数显著高于缓慢发育囊胚（P<0.01）。在第6天发育的囊胚平均总细胞数为118.� ± 21.4，在第7天、第8天和第9天分别显著降至85.6 ± 17.4、62.0 ± 14.5和丨7.0 ± 4.0（P<0.05）。水牛胚胎的正常发育情况表明，平均而言，胚胎最早在4.5-5.5天达到致密桑葚胚阶段。囊胚最早在5.0-6.0天发育，平均需要6.5天从透明带孵化。从桑葚胚到囊胚，TCN丨TE和ICM增加了两倍；然而，在两个胚胎阶段，ICM与TCN的比例保持不变。与未孵化的囊胚相比，孵化后囊胚的TE大约增加了一倍（P<0.05）。然而，ICM细胞减少。观察到孤雌生殖囊胚的发育所需时间比体外受精（IVF）囊胚更长。孤雌生殖囊胚的总细胞数为100.8 ± 11.3，包括59.2 ± 8.4个TE细胞和42.1 ± 6.9个ICM细胞。
