Brandão Daniela O, Maddox-Hyttel Poul, Løvendahl Peter, Rumpf Rodolfo, Stringfellow David, Callesen Henrik
Laboratório de Reprodução Animal I, C.P. 02372 Brasília, DF, Brazil.
Biol Reprod. 2004 Dec;71(6):2048-55. doi: 10.1095/biolreprod.103.025916. Epub 2004 Aug 25.
Although acceptable rates of blastocyst formation are achieved with in vitro production of bovine embryos, several problems still compromise the subsequent development of the fetus and newborn, especially in embryos originating from somatic cell nuclear transfer. Routinely, the potential development of a bovine conceptus is predicted either on blastocyst quality or on various parameters related to the embryonic-fetal development in a foster mother. These methods are either imprecise or costly, highlighting the need for more reliable and practical methods to evaluate early embryonic development and differentiation. Thus, our aim was to improve the in vitro culture of embryos post hatching and to define a stable and repeatable system to monitor the development of bovine embryos. For that, in vitro-derived embryos were cultured in agarose gel tunnels in a modified culture medium (SOFaaci within 10% fetal bovine serum and 27.7 mM glucose). Daily monitoring of embryo length revealed that 56%-67% of the embryos in culture showed rapid growth and elongated until Day 13. Electron microscopy of elongated embryos at Day 14 confirmed successful localization of differentiated cells forming the trophoblast and hypoblast, with the definition of the Rauber layer. In conclusion, a stable culture system of post hatching embryos was first defined and can be used as a model for rapid growth, elongation, and initial differentiation of bovine post hatching embryos produced entirely in vitro.
尽管通过体外生产牛胚胎可实现可接受的囊胚形成率，但仍有几个问题影响着随后胎儿和新生犊牛的发育，特别是对于源自体细胞核移植的胚胎。通常，牛孕体的潜在发育是根据囊胚质量或寄养母牛中与胚胎 - 胎儿发育相关的各种参数来预测的。这些方法要么不准确，要么成本高昂，这凸显了需要更可靠和实用的方法来评估早期胚胎发育和分化。因此，我们的目标是改善孵化后胚胎的体外培养，并定义一个稳定且可重复的系统来监测牛胚胎的发育。为此，将体外来源的胚胎在含有10%胎牛血清和27.7 mM葡萄糖的改良培养基（SOFaaci）中的琼脂糖凝胶通道中培养。每天监测胚胎长度发现，培养中的胚胎有56% - 67%显示出快速生长并延长至第13天。对第14天延长胚胎的电子显微镜检查证实了形成滋养层和内胚层的分化细胞的成功定位，并确定了劳伯层。总之，首次定义了一种稳定的孵化后胚胎培养系统，可作为完全在体外产生的牛孵化后胚胎快速生长、延长和初始分化的模型。
