Oocyte maturation, embryo development and gene expression following two different methods of bovine cumulus-oocyte complexes vitrification
Oocyte maturation, embryo development and gene expression following two different methods of bovine cumulus-oocyte complexes vitrification
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DOI：
10.1007/s11259-016-9671-8
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发表时间：
2017-03-01
影响因子：
2.2
通讯作者：
Jamalzadeh, Mahboobeh
中科院分区：
文献类型：
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作者：
Azari, Mehdi;Kafi, Mojtaba;Jamalzadeh, Mahboobeh
目的 通过两种标准的牛 COC 玻璃化冷冻方法，检测卵母细胞成熟和卵丘扩张相关基因（GDF9、BMP15、HAS2、TNFAIP6、FGF17 和 FSHr）的成熟能力、胚胎发育和表达。方法从屠宰的卵巢中吸出牛卵丘-卵母细胞复合物（COC），然后将其分为三组：非玻璃化 COC。 (对照)，玻璃化1组(V1)；用 15% 乙二醇 (EG) 和 15% DMSO 的保存介质（含 20% FCS 的 TCM-199）进行玻璃化；和玻璃化2组(V2)；玻璃化通过含有40％EG的保存介质进行。玻璃化冷冻后，COC 分两步升温并培养，然后评估核成熟、胚胎发育和基因表达。结果 对照组核成熟和囊胚/卵裂的平均 (+/- SD) 百分比（79.5 +/- 8.0 和 31.0 +/- 5.1%）高于 V1（34.8 +/- 9.1 和 4.4 +/- 5.1%）和 V2分别为（47.8 +/- 11.7 和 7.1 +/- 5.8%）组（P < 0.05）。此外，与 V1 组相比，V2 组中的 COC 显示出更高的平均 (+/- SD) 裂解百分比（31.8 +/- 1.0 vs 21.7 +/- 2.8%；P < 0.05）。对照组 COC 中 GDF9 和 BMP15 表达水平高于玻璃化冷冻组 (P < 0.05)。此外，V2组GDF9和BMP15表达水平高于V1组（P < 0.05）。 V1组HAS2和FGF17的表达低于V2组(P<0.05)。结论卵母细胞成熟基因的表达受玻璃化冷冻程序和条件的影响。单独使用 EG 对牛未成熟 COC 进行玻璃化冷冻，可提高 GDF9、BMP15 的表达，并产生更多的体外成熟和分裂的卵母细胞。
Objective To examine the maturational competence, embryo development and expression of genes involved in oocyte maturation and cumulus expansion (GDF9, BMP15, HAS2, TNFAIP6, FGF17 and FSHr) following two standard methods of bovine COCs vitrification.Methods Bovine cumulus-oocyte complexes (COCs) were aspirated from slaughtered ovaries and then distributed into three groups: non-vitrified COCs (control), vitrification 1 group (V1); vitrification was performed by 15% ethylene glycol (EG) and 15% DMSO in holding media (TCM-199 with 20% FCS); and vitrification 2 group (V2); vitrification was performed by 40% EG in holding media. After vitrification, COCs were warmed in two steps and cultured and then evaluated for nuclear maturation, embryo development and gene expressions.Results The mean (+/- SD) percentages of nuclear maturation and blastocyst/cleaved were higher in control group (79.5 +/- 8.0 and 31.0 +/- 5.1%) than the V1 (34.8 +/- 9.1 and 4.4 +/- 5.1%) and V2 (47.8 +/- 11.7 and 7.1 +/- 5.8%) groups (P < 0.05), respectively. Further, COCs in V2 group showed higher mean (+/- SD) percentages of cleavage compared to V1 group (31.8 +/- 1.0 vs 21.7 +/- 2.8%; P < 0.05). GDF9 and BMP15 expression levels were higher in COCs in the control than of the vitrification groups (P < 0.05). In addition, expression level of GDF9 and BMP15 was higher in V2 group than in V1group (P < 0.05). The expression of HAS2 and FGF17 in V1 group was lower (P < 0.05) than that of the V2 groups.Conclusions Expression of oocyte maturation genes was affected by vitrification procedure and conditions. Using EG alone for vitrification of bovine immature COCs, resulted in higher expression of GDF9, BMP15 and production of more in vitro matured and cleaved oocytes.