Department of Pharmacology, Institute of Bioscience, São Paulo State University (UNESP), Distrito de Rubião Junior s/n, Botucatu, São Paulo, 18618970, Brazil.
Department of Animal Science, Aarhus University, DK-8830, Tjele, Denmark.
J Assist Reprod Genet. 2019 Mar;36(3):413-424. doi: 10.1007/s10815-018-1365-4. Epub 2018 Nov 16.
Oocyte maturation is a complex process involving nuclear and cytoplasmic modulations, during which oocytes acquire their ability to become fertilized and support embryonic development. The oocyte is apparently "primed" for maturation during its development in the dominant follicle. As bovine oocytes immediately resume meiosis when cultured, it was hypothesized that delaying resumption of meiosis with cyclic nucleotide modulators before in vitro maturation (IVM) would allow the oocytes to acquire improved developmental competence.
We tested the Simulated Physiological Oocyte Maturation (SPOM) system that uses forskolin and 3-isobutyl-1-methylxanthine for 2 h prior to IVM against two different systems of conventional IVM (Con-IVM). We evaluated the ultrastructure of matured oocytes and blastocysts and also assessed the expression of 96 genes related to embryo quality in the blastocysts.
In summary, the SPOM system resulted in lower blastocyst rates than both Con-IVM systems (30 ± 9.1 vs. 35 ± 8.7; 29 ± 2.6 vs. 38 ± 2.8). Mature SPOM oocytes had significantly increased volume and number of vesicles, reduced volume and surface density of large smooth endoplasmic reticulum clusters, and lower number of mitochondria than Con-IVM oocytes. SPOM blastocysts showed only subtle differences with parallel undulations of adjacent trophectoderm plasma membranes and peripherally localized ribosomes in cells of the inner cell mass compared with Con-IVM blastocysts. SPOM blastocysts, however, displayed significant downregulation of genes related to embryonic developmental potential when compared to Con-IVM blastocysts.
Our results show that the use of the current version of the SPOM system may have adverse effects on oocytes and blastocysts calling for optimized protocols for improving oocyte competence.
卵母细胞成熟是一个涉及核和细胞质调节的复杂过程，在此过程中，卵母细胞获得受精和支持胚胎发育的能力。在优势卵泡中发育期间，卵母细胞显然已经“准备”好成熟。由于牛卵母细胞在培养时立即恢复减数分裂，因此假设在体外成熟（IVM）前使用环核苷酸调节剂延迟减数分裂的恢复，将使卵母细胞获得改善的发育能力。
我们测试了使用 forskolin 和 3-异丁基-1-甲基黄嘌呤在 IVM 前进行 2 小时的模拟生理卵母细胞成熟（SPOM）系统，与两种不同的常规 IVM 系统（Con-IVM）进行比较。我们评估了成熟卵母细胞和囊胚的超微结构，还评估了囊胚中与胚胎质量相关的 96 个基因的表达。
总之，SPOM 系统的囊胚率低于两种 Con-IVM 系统（30±9.1%比 35±8.7%；29±2.6%比 38±2.8%）。与 Con-IVM 卵母细胞相比，SPOM 成熟卵母细胞的体积和小泡数量显著增加，大光滑内质网簇的体积和表面密度降低，线粒体数量减少。与 Con-IVM 囊胚相比，SPOM 囊胚仅表现出细微差异，即邻近滋养外胚层质膜的平行波动和细胞内细胞团中周边定位的核糖体。然而，与 Con-IVM 囊胚相比，SPOM 囊胚显示与胚胎发育潜力相关的基因显著下调。
我们的结果表明，当前版本的 SPOM 系统的使用可能对卵母细胞和囊胚产生不利影响，需要优化提高卵母细胞能力的方案。
