High bovine blastocyst development in a static in vitro production system using SOFaa medium supplemented with sodium citrate and myo-inositol with or without serum-proteins
High bovine blastocyst development in a static in vitro production system using SOFaa medium supplemented with sodium citrate and myo-inositol with or without serum-proteins
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DOI：
10.1016/s0093-691x(99)00162-4
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发表时间：
1999-09-01
期刊：
影响因子：
2.8
通讯作者：
Callesen, H
中科院分区：
文献类型：
--
作者：
Holm, P;Booth, PJ;Callesen, H
我们描述了一种牛胚胎培养系统，该系统支持在血清或BSA存在下以及在不存在这些组分的规定条件下的可重复高发育。在第一个实验中，将在含有氨基酸（SOFaa）、柠檬酸钠（SOFaac）和肌醇（SOFaaci）以及含有BSA或聚乙烯醇（PVA）的SOF中的胚胎发育与在M199颗粒细胞共培养物（M199 cc-培养物）中的胚胎发育进行比较。随后，比较了在SOFaaci与PVA（SOFaaci-PVA）中在规定条件下培养的囊胚的发育和细胞数，或在SOFaaci与5%牛血清（SOFaaci-CS）或M199共培养物中在未规定条件下培养的囊胚的发育和细胞数。通过在11个月内每周重复（n=30）检查SOFaaci-CS中培养结果的重复性。通过将形态良好的囊胚（n=10）转移到同步化受体来评估在SOFaaci-PVA中发育的胚胎的存活力。在第二个实验中，研究了从IVM和IVM-IVF中省略CS或BSA对SOFaaci-PVA或SOFaaci-CS中后续胚胎发育的影响。在SOFaa-PVA、SOFaac-PVA、SOFaa-BSA和M199中的胚泡发育分别为16 +/-3（B）、23 +/-2（ab）、30 +/-8（a）和36 +/-7（a）%（P-ab < 0.05）。另外包含肌醇导致SOFaac-PVA中42 +/- 1（a）%胚泡，而SOFaac-PVA中19 +/- 3（B）%胚泡，SOFaac-BSA中47 +/- 7（a）%胚泡，M199共培养物中36 +/- 7（a）%胚泡（P-ab< 0.01）。在30个重复中，SOFaaci-CS中卵母细胞的平均卵裂率和囊胚率分别为87 +/- 4和49 +/-5%。在转移10个在限定培养基（即，SOFaaci-PVA）。定义的IVM或IVM-IVF（即，不存在CS和BSA时）降低切割率（83 +/-3和55 +/-3%vs存在CS时的90 +/-1%; P < 0.01）。SOFaaci-CS中随后的胚胎发育在这些限定条件中的任一种中均不受影响。然而，在完全定义的IVP条件下，卵裂率和囊胚率分别为54 +/- 7和19 +/-4%。得出的结论是，在规定的培养条件下，添加柠檬酸盐和肌醇改善囊胚发育的速率与血清，BSA或共培养获得的速率相当，囊胚的质量不受血清或BSA的情况下。然而，血清在IVM/IVF期间对于正常受精和随后的高囊胚发育是必不可少的。(C)1999年，Elsevier Science Inc.
We describe a bovine embryo culture system that supports repeatable high development in the presence of serum or BSA as well as under defined conditions in the absence of those components. In the first experiment, embryo development in SOF with amino acids (SOFaa), sodium citrate (SOFaac) and myo-inositol (SOFaaci) and with BSA or polyvinyl alcohol (PVA) was compared with that in a M199 granulosa cell co-culture (M199 cc-culture). Subsequently, development and cell numbers of blastocysts cultured under defined conditions in SOFaaci with PVA (SOFaaci-PVA), or under undefined conditions in SOFaaci with 5% cow serum (SOFaaci-CS) or M199 co-culture were compared. The repeatability of culture results in SOFaaci-CS was checked by weekly replicates (n=30) spread over 11 months. The viability of embryos developed in SOFaaci-PVA was estimated by transfer of morphologically good blastocysts (n=10) to synchronized recipients. In the second experiment, the effect of omitting CS or BSA from IVM and IVM-IVF on subsequent embryo development in SOFaaci-PVA or in SOFaaci-CS was investigated. Blastocyst development in SOFaa-PVA, SOFaac-PVA, SOFaa-BSA and M199 was 16 +/- 3(b), 23 +/- 2(ab), 30 +/- 8(a) and 36 +/-7(a)%, respectively (P-ab < 0.05). Additional inclusion of myoinositol resulted in 42 +/- 1(a)% blastocysts in SOFaaci-PVA vs 19 +/- 3(b)% in SOFaac-PVA, 47 +/- 7(a)% in SOFaac-BSA, and 36 +/- 7(a)% in M199 co-culture, respectively (P-ab< 0.01). In 30 replicates, the average cleavage and blastocyst rates of oocytes in SOFaaci-CS were 87 +/- 4 and 49 +/- 5%, respectively. Five normal calves were produced after transfer of 10 blastocysts developed in defined culture medium (i.e., SOFaaci-PVA). Defined IVM or IVM-IVF (i.e., in absence of CS and BSA) reduced cleavage rates (83 +/- 3 and 55 +/- 3% vs 90 +/- 1% in presence of CS; P < 0.01). Subsequent embryo development in SOFaaci-CS was not affected in either of these defined conditions. However, cleavage and blastocyst rates under completely defined IVP conditions were 54 +/- 7 and 19 +/- 4%, respectively. It was concluded that under defined culture conditions, addition of citrate and myo-inositol improved blastocyst development to rates comparable to those obtained with serum, BSA or co-culture and that the quality of blastocysts was not affected by the absence of serum or BSA. However, serum was essential during IVM/IVF for normal fertilization and subsequent high blastocyst development. (C) 1999 by Elsevier Science Inc.