Gardner D K, Sakkas D
Centre for Early Human Development, Clayton, Melbourne, Victoria, Australia.
Hum Reprod. 1993 Feb;8(2):288-95. doi: 10.1093/oxfordjournals.humrep.a138039.
The cleavage, metabolism and viability of mouse zygotes were assessed after culture in media of different ionic and metabolite composition. Medium with a high potassium concentration, characteristic of mammalian oviduct fluid, inhibited cleavage and blastocyst formation (P < 0.01). This inhibition was partially alleviated by the removal of phosphate, and subsequently abolished by supplementation with amino acids, vitamins, insulin, epidermal growth factor and transferrin (AVIET). Glucose uptake by cultured blastocysts, measured fluorimetrically, was not affected by the ionic or metabolite composition of the medium, but was significantly reduced by the inclusion of AVIET (P < 0.01). Lactate production was also significantly reduced in the presence of AVIET (P < 0.01). Calculations of metabolic activity revealed that embryos cultured in the presence of AVIET had a glycolytic activity similar to embryos developed in vivo. In contrast, embryos cultured in conventional embryo culture media exhibited an elevated glycolytic activity. Culturing embryos for 4 days in a reduced lactate concentration (4.79 mM), significantly increased fetal development after transfer, compared with embryos cultured in the concentration of lactate present in conventional embryo culture media (23.3 mM; P < 0.01). In contrast, when embryos were transferred on day 3 of culture, significantly more fetuses were obtained from embryos cultured in high levels of lactate (P < 0.01). Supplementation of medium with AVIET significantly increased resultant fetal weights after transfer (P < 0.05). This study demonstrates that different media are required to maintain embryo viability on successive days of culture, and highlights the potential limitations of employing simple salt solutions for the culture of preimplantation mammalian embryos.
在不同离子和代谢物组成的培养基中培养后，评估了小鼠受精卵的分裂、代谢和活力。具有高钾浓度（哺乳动物输卵管液的特征）的培养基抑制了分裂和囊胚形成（P < 0.01）。去除磷酸盐可部分缓解这种抑制作用，随后通过添加氨基酸、维生素、胰岛素、表皮生长因子和转铁蛋白（AVIET）可消除这种抑制作用。通过荧光法测量，培养的囊胚对葡萄糖的摄取不受培养基离子或代谢物组成的影响，但加入AVIET后显著降低（P < 0.01）。在存在AVIET的情况下，乳酸产生也显著减少（P < 0.01）。代谢活性计算表明，在AVIET存在下培养的胚胎具有与体内发育胚胎相似的糖酵解活性。相比之下，在传统胚胎培养基中培养的胚胎表现出较高的糖酵解活性。与在传统胚胎培养基中存在的乳酸浓度（23.3 mM）下培养的胚胎相比，在降低的乳酸浓度（4.79 mM）下培养胚胎4天，转移后胎儿发育显著增加（P < 0.01）。相反，当在培养第3天转移胚胎时，从高乳酸水平培养的胚胎中获得的胎儿明显更多（P < 0.01）。向培养基中添加AVIET显著增加了转移后所得胎儿的体重（P < 0.05）。本研究表明，在连续培养的不同天数需要不同的培养基来维持胚胎活力，并突出了使用简单盐溶液培养植入前哺乳动物胚胎的潜在局限性。
