Two-cell block to development of cultured hamster embryos is caused by phosphate and glucose.
Two-cell block to development of cultured hamster embryos is caused by phosphate and glucose.
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培养仓鼠胚胎发育的双细胞阻碍是由磷酸盐和葡萄糖引起的。
DOI：
10.1095/biolreprod39.5.1183
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发表时间：
1988
影响因子：
3.6
通讯作者：
Bavister,BD
中科院分区：
文献类型：
--
作者：
Schini,SA;Bavister,BD
通过改变培养基中葡萄糖和磷酸盐浓度的实验，研究了仓鼠2细胞胚胎在体外发育失败（2细胞阻滞）。胚胎在无蛋白的改良Tyrode’s溶液中培养，该溶液通常含有5.0 mM葡萄糖和0.35 mM磷酸二氢钠。在0.35 mM磷酸盐不含葡萄糖的情况下，23%的2细胞胚胎在培养1天后达到4细胞期或更久，27%在培养2天后达到4细胞期。即使在0.1 mM处，葡萄糖也能抑制胚胎发育（培养2天后，4%的胚胎发育到≥4个细胞）；在这个葡萄糖浓度以上没有剂量相关的抑制作用。在第二个实验中，磷酸盐水平在没有葡萄糖的情况下发生变化。磷酸盐对发育具有高度抑制作用，在无磷酸盐条件下培养1天后，97%的2细胞胚胎达到4细胞期或更久，而在0.1-1.05 mM磷酸盐条件下，这一比例为9-21%。培养2天后，26%的胚胎在不含磷酸盐的情况下达到8细胞期或更长时间，而在含0.1 mM或更高磷酸盐的情况下达到8细胞期的胚胎发育率为0%。在一个析因实验中，当葡萄糖存在或不存在时，磷酸盐阻碍胚胎发育，而葡萄糖在没有磷酸盐的情况下不阻碍胚胎发育。然而，2-脱氧葡萄糖（葡萄糖的非代谢类似物）在缺乏磷酸盐的情况下抑制胚胎发育。这些数据表明，对仓鼠2细胞胚胎发育的体外阻碍至少部分是由葡萄糖和/或磷酸盐引起的。从培养基中删除这些化合物，几乎在所有胚胎中消除了2细胞发育障碍，并且大约25-75%的胚胎在体外发育到8细胞或桑葚胚期。这些观察结果为在常规培养基中出现的2细胞和4细胞阻滞提供了一种可能的解释：葡萄糖和/或磷酸盐刺激糖酵解可能导致三磷酸腺苷（ATP）产生效率低下。这些数据表明，早期卵裂期仓鼠胚胎的体外发育调控与近交系小鼠胚胎有明显的差异，因为后者在8细胞发育阶段之前糖酵解途径不活跃，在磷酸盐和葡萄糖的培养基中会从1细胞生长到囊胚。
The failure of hamster 2-cell embryos to develop in vitro (2-cell block) was examined with experiments in which concentrations of glucose and phosphate in the culture medium were varied. Embryos were cultured in a protein-free modified Tyrode's solution that normally contains 5.0 mM glucose and 0.35 mM sodium dihydrogen phosphate. In the presence of 0.35 mM phosphate but without glucose, 23% of 2-cell embryos reached the 4-cell stage or further after culture for 1 day and 27% after 2 days. Glucose inhibited embryo development even at 0.1 mM (4% development to ≥ 4-cells after culture for 2 days); there was no dose-related inhibition above this glucose concentration. In a second experiment, phosphate levels were varied in the absence of glucose. Phosphate was highly inhibitory to development, with 97% of 2-cell embryos reaching the 4-cell stage or further after culture for 1 day in the absence of phosphate compared to 9–21% in the presence of 0.1–1.05 mM phosphate. After culture for 2 days, 26% of embryos reached the 8-cell stage or further when phosphate was absent compared to 0% development to 8-cells with 0.1 mM phosphate or higher. In a factorial experiment, phosphate blocked development when glucose was present or absent, whereas glucose did not block embryo development in the absence of phosphate. However, 2-deoxyglucose (a non-metabolizable analogue of glucose) inhibited embryo development in the absence of phosphate. These data show that the in vitro block to development of hamster 2-cell embryos is caused at least in part by glucose and/or phosphate. Deletion of these compounds from the culture medium eliminates the 2-cell block to development in virtually all embryos, and approximately 25–75% of embryos develop to the 8-cell or morula stages in vitro. The observations provide a possible explanation for the 2-cell and 4-cell blocks that occur in conventional culture media: stimulation of glycolysis by glucose and/or phosphate may result in inefficient adenosine triphosphate (ATP) production. The data indicate marked dissimilarities in the regulation of in vitro development of early cleavage stage hamster embryos compared with embryos of inbred mice, since the latter have an inactive glycolytic pathway prior to the 8-cell stage of development and will grow from 1-cell to blastocyst with both phosphate and glucose in the culture medium.