通过蛋白质合成和磷酸化的双重抑制的卵母细胞活化已经改善了不同哺乳动物物种的体外胚胎产生。在这项研究中,我们评估了环己酰亚胺(CHX)组合的效果,二甲基氨基嘌呤(DMAP),和茴香霉素(ANY)对牛卵母细胞的激活,特别是MPF和MAPK的动态,胚胎发育潜能,和质量。结果表明,卵裂率和囊胚率,以及CCNB1,CDK1,p-CDK1Thr161和p-CDK1Thr14-Tyr15的水平在各组之间相似;与DMAP组合相比,ANY和ANY+CHX降低了ERK1/2的表达(p<0.05),而ANY+DMAP,CHX+DMAP,与ANY和ANY+CHX治疗相比,ANY+CHX+DMAP降低p-ERK1/2(p<0.05)。胚泡在细胞计数方面的质量,他们的分配,TUNEL阳性细胞的数量在各组之间没有差异。然而,与其他组相比,来自ANY+CHX+DMAP处理的胚胎中POU5F1的转录水平更高,而CDX2的表达水平没有差异。此外,与ANY治疗相比,ANY+CHX+DMAP治疗的BCL2A1/BAX比值显著较低(p<0.05),与其他治疗无显著差异.总之,通过双重抑制蛋白质合成和磷酸化的卵母细胞活化诱导MPF失活而不降解CCNB1,而MAPK失活在这些抑制剂之间不同地发生。因此,尽管这些抑制剂的联合使用不会影响体外早期发育能力,它积极影响与胚胎质量相关的转录本的表达。
Oocyte activation via dual inhibition of protein synthesis and phosphorylation has improved in vitro embryo production in different mammalian species. In this study, we evaluated the effects of the combination of
cycloheximide (CHX), dimethyl amino purine (DMAP), and anisomycin (ANY) on the activation of bovine oocytes, particularly on dynamics of MPF and MAPKs, embryonic developmental potential, and quality. The results showed that the cleavage and blastocyst rates, as well as levels of CCNB1, CDK1, p-CDK1Thr161, and p-CDK1Thr14-Tyr15, were similar among groups; ANY and ANY + CHX reduced the expression of ERK1/2 compared to DMAP-combinations (p < 0.05), whereas ANY + DMAP, CHX + DMAP, and ANY + CHX + DMAP reduced p-ERK1/2 compared to ANY and ANY + CHX treatments (p < 0.05). The quality of blastocysts in terms of cell counts, their allocation, and the numbers of TUNEL-positive cells did not differ among groups. However, transcript levels of POU5F1 were higher in embryos derived from ANY + CHX + DMAP treatment compared to other groups, while expression levels of CDX2 did not show differences. In addition, the BCL2A1/BAX ratio of the ANY + CHX + DMAP treatment was significantly low compared to the ANY treatment (p < 0.05) and did not differ significantly from the other treatments. In conclusion, oocyte activation by dual inhibition of protein synthesis and phosphorylation induces MPF inactivation without degradation of CCNB1, while MAPK inactivation occurs differentially between these inhibitors. Thus, although the combined use of these inhibitors does not affect early developmental competence in vitro, it positively impacts the expression of transcripts associated with embryonic quality.