Development and viability of pig oocytes matured in a protein-free medium containing epidermal growth factor
Development and viability of pig oocytes matured in a protein-free medium containing epidermal growth factor
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DOI：
10.1016/s0093-691x(00)00390-3
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发表时间：
2000-09-15
期刊：
影响因子：
2.8
通讯作者：
Day, BN
中科院分区：
文献类型：
--
作者：
Abeydeera, LR;Wang, WH;Day, BN
本研究探讨了表皮生长因子(EGF)对猪卵母细胞在无蛋白体外成熟(IVM)系统中发育能力的改善作用。卵母细胞成熟在三种培养液中进行：1.PF-Medic199+0.1%聚乙烯醇(PVA)；2.PF-Medic1+EGF：PF-Medic1+10 ng/mlEGF；3.+Ve Cont：北卡罗来纳州立大学(NCSU)23培养液+10%猪卵泡液。所有培养基含0.57 mM半胱氨酸。激素补充剂LH0.5mug/mLFSH 0.5mug/mLFSH只存在于培养的前半段(20~22h)。成熟后的卵母细胞与冷冻解冻后的精子共孵育5~6h，移植到含有0.4%BSA的NCSU-23胚胎培养液中144h，在实验1中，观察了+ve CONT(第4类)、PF-Medicc(第2类)和PF-Medicc+EGF(第3类)成熟卵母细胞的卵丘扩张情况。然而，核成熟到中期II期的差异不显著。在试验2中，没有观察到施肥参数的差异。三种培养液的卵裂率差异显著(P<；0.01)，卵裂率分别为52%、60%和69%。体外培养的卵母细胞囊胚发育率最低(22%)(P<；0.01)。然而，+VE-CONT组和PF-Tcm+EGF组的囊胚发育率相同(37%)。当卵母细胞在EGF的存在下成熟时，囊胚细胞的数量显著增加(26个比37个到41个)。在实验3中，在PF-Tcm+EGF中成熟的卵母细胞胞内谷胱甘肽(GSH)浓度显著高于PF-Tcm(P<；0.05)(5.9pmol/卵母细胞比11.4pmoL/卵母细胞)。25例胚胎移植受者中有22例妊娠(实验4)。有4只动物在60天内恢复发情。在26-45天屠宰的6头怀孕动物共产43胎(4-12头)，其余12头产仔82头(3-12头)。这些结果表明，EGF提高了猪卵母细胞在无蛋白培养液中成熟的发育能力，这与卵母细胞中较高的GSH水平有关。仔猪的出生表明，在EGF存在下成熟的卵母细胞获得的胚胎是有活力的。(C)2000年，爱思唯尔科学公司。
This study examined the ability of epidermal growth factor (EGF) to improve the developmental competence of pig oocytes matured in a protein-free (PF) in vitro maturation (IVM) system. Oocyte maturation was done in one of three media: 1. PF-TCM: tissue culture medium (TCM) 199 + 0.1% polyvinylalcohol (PVA); 2. PF-TCM+EGF: PF-TCM + 10 ng/ml EGF; and 3. +ve CONT: North Carolina State University (NCSU) 23 medium + 10% porcine follicular fluid. All media contained 0.57 mM cysteine. Hormonal supplements, 0.5 mug/mL LH and 0.5 mug/mL FSH, were present only for the first half (20 to 22 h) of the culture period. After maturation, ocytes were co-incubated with frozen-thawed spermatozoa for 5 to 6 h and transferred to embryo culture medium,NCSU 23 containing 0.4% BSA, for 144 h. In Experiment 1, differences in cumulus expansion were observed for oocytes matured in +ve CONT (Category 4), PF-TCM (Category 2) and PF-TCM+EGF (Category 3). However, no significant differences in nuclear maturation to metaphase II stage were observed. In Experiment 2, no differences in fertilization parameters were observed. Significant (P < 0.01) differences in cleavage rates were observed among the three media for a proportion of the oocytes matured (52, 60 and 69% in PF-TCM, PF-TCM+EGF, and +ve CONT, respectively). Oocytes matured in PF-TCM showed the lowest (P < 0.01) blastocyst development (22%). However, the same rate of blastocyst development was obtained for +ve CONT (37%) and PF-TCM+EGF (37%). Blastocyst cell numbers were significantly higher when oocytes were matured in the presence of EGF (26 vs. 37 to 41). In Experiment 3, oocytes matured in PF-TCM+EGF had a significantly (P < 0.05) higher intracellular glutathione (GSH) concentration (5.9 vs. 11.4 pmol/oocyte) compared with PF-TCM. Twenty-two of 25 embryo transfer recipients became pregnant (Experiment 4). Four animals returned to estrus in within 60 days. Six pregnant animals slaughtered at 26 to 45 days had 43 fetuses (range: 4 to 12) and the remaining 12 animals farrowed 82 piglets (range: 3 to 12). These results indicate that EGF enhances the developmental competence of pig oocytes matured in a protein-free culture medium which is correlated with higher GSH level in oocytes. Birth of piglets indicate that embryos derived from oocytes matured in the presence of EGF are viable. (C) 2000 by Elsevier Science Inc.