Li Xihe, Dai Yanfeng, Allen W R
Department of Clinical Veterinary Medicine, University of Cambridge, Equine Fertility Unit, Mertoun Paddocks, Newmarket, Suffolk CB8 9BH, United Kingdom.
Biol Reprod. 2004 Oct;71(4):1391-6. doi: 10.1095/biolreprod.104.029066. Epub 2004 Jun 23.
In vitro maturation of horse oocytes cultured with or without IGF-I supplementation and their first cell cycle organization were studied in reconstructed horse oocytes made by somatic cell nuclear transfer versus intact oocytes stimulated parthenogenetically. The rates of metaphase II oocytes (47% and 45%) and of reconstructed oocytes that developed to the two-cell (27% and 25%) and blastocyst stages (11% and 3%) were not different between the media, with or without IGF-I, respectively. However, significantly more parthenogenetic embryos exhibited two-cell development with IGF-I (P < 0.05). The results also demonstrated that the first cell cycle organization in the reconstructed oocytes involved two different ways of nuclear remodeling. The donor nucleus in the Type I embryo showed normal nuclear remodeling that resulted in normal embryonic development. In the Type II embryos, however, the donor nucleus formed a polyploid nucleus or the embryo fragmented. Addition of IGF-I to the maturation medium significantly increased the rate of normal Type I embryonic development from the reconstructed oocytes (45% vs. 28%, P < 0.05). Maturation-promoting factor (MPF; including cdc2 and cyclin B) and mitogen-activated protein kinase (MAPK; including ERK1 and ERK2) were present at the beginning of culture, just after the oocytes had been harvested from the ovaries. The quantities of cyclin B remained stable no matter how long a period of in vitro culture the oocytes underwent, whereas cdc2 showed a tendency to accumulate in the oocytes toward the end of the 30-h culture period. Addition of IGF-I to the medium may induce a bigger accumulation of MAPK in the cytoplasm of the horse oocyte, especially in the ERK2 component, which might, in turn, increase the chance of the reconstructed oocyte undergoing nuclear remodeling to form a Type I embryo following nuclear transfer.
研究了在添加或不添加胰岛素样生长因子-I（IGF-I）的情况下培养的马卵母细胞的体外成熟情况，以及通过体细胞核移植构建的马重构卵母细胞与孤雌激活的完整卵母细胞的第一次细胞周期组织情况。在添加或不添加IGF-I的培养基中，处于中期II的卵母细胞比例（分别为47%和45%）以及发育到二细胞期（分别为27%和25%）和囊胚期（分别为11%和3%）的重构卵母细胞比例并无差异。然而，添加IGF-I的情况下，显著更多的孤雌胚胎发育到了二细胞期（P < 0.05）。结果还表明，重构卵母细胞的第一次细胞周期组织涉及两种不同的核重塑方式。I型胚胎中的供体细胞核显示出正常的核重塑，从而导致正常的胚胎发育。然而，在II型胚胎中，供体细胞核形成了多倍体细胞核或胚胎发生了碎片化。在成熟培养基中添加IGF-I显著提高了重构卵母细胞发育为正常I型胚胎的比例（45%对28%，P < 0.05）。成熟促进因子（MPF；包括cdc2和细胞周期蛋白B）和丝裂原活化蛋白激酶（MAPK；包括ERK1和ERK2）在培养开始时，即从卵巢采集卵母细胞后就已存在。无论卵母细胞进行多长时间的体外培养，细胞周期蛋白B的量都保持稳定，而cdc2在30小时培养期结束时显示出在卵母细胞中积累的趋势。向培养基中添加IGF-I可能会诱导马卵母细胞细胞质中MAPK更大程度的积累，尤其是在ERK2组分中，这反过来可能会增加重构卵母细胞在核移植后进行核重塑以形成I型胚胎的机会。
