Carolan C, Lonergan P, Monget P, Monniaux D, Mermillod P
INRA, Station de Physiologie de la Reproduction, Nouzilly, France.
Mol Reprod Dev. 1996 Apr;43(4):477-83. doi: 10.1002/(SICI)1098-2795(199604)43:4<477::AID-MRD10>3.0.CO;2-X.
Recent studies have shown that the developmental capacity of in vitro-matured oocytes is affected by the origin of follicular fluid (FF) supplemented to the maturation medium. The aims of this study were (1) to determine if follicle size and quality would influence the capacity of FF to support bovine oocyte maturation and (2) to determine if fetal calf serum (FCS) and FF had an additive effect when added together to the maturation medium. Follicular fluid collected from 108 follicles was classified according to size ( < 6, 6-8, > 8 mm in diameter) and quality (healthy, early atretic, and atretic). Quality, first determined by mitosis/pycnosis ratios in granulosa cell smears, was subsequently confirmed by insulin-like growth factor binding protein (IGFBP) patterns and estradiol concentrations. While most small- or medium-sized follicles showed some atresia (88% and 67%, respectively), fewer of the large follicles were atretic (30%). In experiment 1 bovine oocytes (n = 2,152) were matured either in TCM199 alone, with 10% FCS, or with 10% FF from the following follicle types: small healthy (SH); small early atretic (SEA); small atretic (SA); medium healthy (MH); medium early atretic (MEA); medium atretic (MA); large healthy (LH); large early atretic (LEA); and large atretic (LA). Following IVM, oocytes were fertilized and subsequently cultured in synthetic oviduct fluid (SOF). Day 8 blastocyst yields were 23% in TCM199 alone; 37% in TCM199 plus FCS; and, in medium supplemented with FF, SH, 36%; MH, 32%; LH, 30%; SEA, 21%; MEA, 26%; LEA, 28%; SA, 32%; MA, 33%; and LA, 38%. All FF from healthy or atretic follicles resulted in significantly improved blastocyst yields compared to M199 alone (P < 0.05). However, FF from early atretic follicles irrespective of size did not yield a significant improvement. In experiment 2 we examined the effect of addition of FF-LH and serum together to the maturation medium. In terms of blastocyst yield, no additional benefit was observed when TCM199 was supplemented with 10% FCS + 10% FF (33%) compared to 10% FCS or FF alone (35% and 30%, respectively). The efficacy of FF as a supplement to the maturation medium to improve cytoplasmic maturation appears to vary with follicle quality but not size. However, in general, the addition of 10% FF or FCS to the maturation media resulted in a similar blastocyst yield with no additional improvement when media was supplemented with both FCS and FF.
近期研究表明，补充到成熟培养基中的卵泡液（FF）来源会影响体外成熟卵母细胞的发育能力。本研究的目的是：（1）确定卵泡大小和质量是否会影响卵泡液支持牛卵母细胞成熟的能力；（2）确定当胎牛血清（FCS）和卵泡液一起添加到成熟培养基中时是否具有相加效应。从108个卵泡收集的卵泡液根据大小（直径<6、6 - 8、>8 mm）和质量（健康、早期闭锁、闭锁）进行分类。质量首先通过颗粒细胞涂片的有丝分裂/固缩比例确定，随后通过胰岛素样生长因子结合蛋白（IGFBP）模式和雌二醇浓度进行确认。虽然大多数小或中等大小的卵泡显示出一定程度的闭锁（分别为88%和67%），但大卵泡中闭锁的较少（30%）。在实验1中，牛卵母细胞（n = 2,152）在单独的TCM199、添加10% FCS或添加来自以下卵泡类型的10%卵泡液中成熟：小健康（SH）；小早期闭锁（SEA）；小闭锁（SA）；中等健康（MH）；中等早期闭锁（MEA）；中等闭锁（MA）；大健康（LH）；大早期闭锁（LEA）；和大闭锁（LA）。体外成熟后，卵母细胞受精，随后在合成输卵管液（SOF）中培养。单独使用TCM199时第8天的囊胚率为23%；TCM199加FCS时为37%；在添加卵泡液的培养基中，SH为36%；MH为32%；LH为30%；SEA为21%；MEA为26%；LEA为28%；SA为32%；MA为33%；LA为38%。与单独使用M199相比，来自健康或闭锁卵泡的所有卵泡液均显著提高了囊胚率（P < 0.05）。然而，无论大小，来自早期闭锁卵泡的卵泡液并未产生显著改善。在实验2中，我们研究了将卵泡液-LH和血清一起添加到成熟培养基中的效果。就囊胚率而言，与单独添加10% FCS或卵泡液（分别为35%和30%）相比，当TCM199添加10% FCS + 10%卵泡液时未观察到额外益处（33%）。卵泡液作为成熟培养基补充剂以改善细胞质成熟的功效似乎随卵泡质量而变化，但与大小无关。然而，总体而言，向成熟培养基中添加10%卵泡液或FCS会产生相似的囊胚率，当培养基同时添加FCS和卵泡液时未观察到额外改善。
