School of Agriculture, Food Science and Veterinary Medicine, College of Life Sciences, Veterinary Sciences Centre, University College Dublin, Belfield, Dublin 4, Ireland.
Reproduction. 2010 Jul;140(1):83-92. doi: 10.1530/REP-10-0040. Epub 2010 Apr 28.
The aim of this study was to examine the presence and regulation of glycogen synthase kinase-3alpha (GSK3A) and GSK-3beta (GSK3B) in bovine embryos and their possible roles in embryo development. Our results show that GSK3A and GSK3B are present in bovine embryos at the two-cell stage to the hatched blastocyst stage. Bovine embryo development was associated with an increase in the phosphorylation of both isoforms, being statistically significant at blastocyst and hatched blastocyst stages, compared with earlier stages. Inhibition of GSK3 with CT99021 (3 microM) resulted in a significant increase in the percentage and quality of blastocysts, while inhibition of GSK3 with lithium chloride (LiCl; 20 mM) significantly reduced at the proportion of eight-cell embryos on day 3 and inhibited blastocyst formation. The use of LY294002 (10 microM), a specific inhibitor of phosphatidylinositol-3 kinase, also produced a significant decrease in embryo development. In addition, treatment with LiCl and LY294002 produced a significant decrease in the serine phosphorylation of both isoforms of GSK3. Finally, CT99021 and LiCl reduced the phosphorylation of beta-catenin on Ser45 in two-cell embryos, while LY294002 increased it. Despite the fact that LiCl inhibited GSK3 activity, as demonstrated by beta-catenin phosphorylation, its effects on the bovine embryo could be mediated through other signaling pathways leading finally to a decrease in the phosphorylation of GSK3 and a reduction in embryo development. Therefore, in conclusion, GSK3A/B serine phosphorylation was positively correlated with embryo development, indicating the importance of an accurate regulation of GSK3 activity during developmental stages to achieve normal bovine embryo development.
本研究旨在探讨糖原合酶激酶-3α（GSK3A）和 GSK-3β（GSK3B）在牛胚胎中的存在及其调节作用，以及它们在胚胎发育中的可能作用。我们的结果表明，GSK3A 和 GSK3B 存在于牛胚胎的两细胞期到孵出的囊胚期。牛胚胎发育与两种同工型的磷酸化增加有关，与早期阶段相比，囊胚和孵出的囊胚阶段的磷酸化水平具有统计学意义。用 CT99021（3 μM）抑制 GSK3 导致囊胚的比例和质量显著增加，而用氯化锂（LiCl；20 mM）抑制 GSK3 则显著降低第 3 天的 8 细胞胚胎比例并抑制囊胚形成。特异性抑制磷脂酰肌醇-3 激酶的 LY294002（10 μM）的使用也导致胚胎发育显著减少。此外，用 LiCl 和 LY294002 处理还导致两种 GSK3 同工型的丝氨酸磷酸化显著减少。最后，CT99021 和 LiCl 降低了两细胞胚胎中β-catenin 的 Ser45 磷酸化，而 LY294002 则增加了该磷酸化。尽管 LiCl 抑制了 GSK3 活性，如β-catenin 磷酸化所证明的，但它对牛胚胎的影响可能通过其他信号通路介导，最终导致 GSK3 磷酸化减少和胚胎发育减少。因此，总之，GSK3A/B 丝氨酸磷酸化与胚胎发育呈正相关，表明在发育阶段准确调节 GSK3 活性对于实现正常的牛胚胎发育非常重要。
