Ferreira Elisa Melo, Vireque Alessandra Aparecida, Adona Paulo Roberto, Ferriani Rui Alberto, Navarro Paula Andrea
Departamento de Ginecologia e Obstetrícia, Faculdade de Medicina de Ribeirão Preto, Universidade de São Paulo, Ribeirão Preto, São Paulo, Brazil.
Eur J Obstet Gynecol Reprod Biol. 2009 Jul;145(1):76-80. doi: 10.1016/j.ejogrb.2009.03.016. Epub 2009 Apr 28.
Asynchrony between nuclear and cytoplasmic maturation, and possibly damage to the oocyte meiotic spindle, limits the application of in vitro maturation (IVM) in assisted reproduction. Several studies have suggested that Prematuration with meiosis blockers may improve oocyte quality after IVM, favoring early embryogenesis. Thus, we investigated the effect of Prematuration with the nuclear maturation inhibitor butyrolactone I (BLI) on the meiotic spindle and chromosomal configuration of bovine oocytes.
Immature oocytes obtained from cows slaughtered in a slaughterhouse (n=840) were divided into the following groups: (1) control (n=325), submitted only to IVM in TCM199 for 24h; (2) BLI 18h (n=208) submitted to meiotic blockage with 100 microM BLI for 24h (Prematuration) and then induction of IVM in TCM199 for 18h; and (3) BLI 24h (n=307), pre-matured with 100 microM BLI for 24h followed by 24h of IVM in TCM199. The oocytes were then fixed, stained by immunofluorescence for morphological visualization of both microtubules and chromatin, and evaluated.
Meiotic arrest occurred in 90.2% of the oocytes cultured with BLI. Maturation rates were similar for all groups (80.3%, 73.6% and 82.7% for the control, BLI 18h and BLI 24h groups, respectively). We observed 81.3% normal oocytes in metaphase II in the control group, and 80.0% and 81.2% in the BLI 18h and BLI 24h groups, respectively. The incidence of meiotic anomalies did not differ between groups (18.7%, 20.0% and 18.8% for the control, BLI 18h and BLI 24h, respectively).
Prematuration with butyrolactone I reversibly arrests meiosis without damaging the meiotic spindle or the chromosome distribution of bovine oocytes after in vitro maturation.
核成熟与胞质成熟不同步，以及可能对卵母细胞减数分裂纺锤体造成的损伤，限制了体外成熟（IVM）在辅助生殖中的应用。多项研究表明，使用减数分裂阻滞剂进行提前成熟可能会改善IVM后的卵母细胞质量，有利于早期胚胎发育。因此，我们研究了用核成熟抑制剂丁内酯I（BLI）进行提前成熟对牛卵母细胞减数分裂纺锤体和染色体构型的影响。
从屠宰场屠宰的母牛中获取未成熟卵母细胞（n = 840），分为以下几组：（1）对照组（n = 325），仅在TCM199中进行24小时IVM；（2）BLI 18小时组（n = 208），先用100微摩尔BLI进行24小时减数分裂阻滞（提前成熟），然后在TCM199中诱导IVM 18小时；（3）BLI 24小时组（n = 307），先用100微摩尔BLI提前成熟24小时，然后在TCM199中进行24小时IVM。然后将卵母细胞固定，通过免疫荧光染色以对微管和染色质进行形态学观察，并进行评估。
用BLI培养的卵母细胞中有90.2%发生减数分裂阻滞。所有组的成熟率相似（对照组、BLI 18小时组和BLI 24小时组分别为80.3%、73.6%和82.7%）。我们在对照组中观察到81.3%的卵母细胞在中期II正常，在BLI 18小时组和BLI 24小时组中分别为80.0%和81.2%。减数分裂异常的发生率在各组之间没有差异（对照组、BLI 18小时组和BLI 24小时组分别为18.7%、20.0%和18.8%）。
丁内酯I提前成熟可使减数分裂可逆性阻滞，且在体外成熟后不会损害牛卵母细胞的减数分裂纺锤体或染色体分布。
