College of Animal Science and Veterinary Medicine, Shandong Agricultural University, Tai-an City, People's Republic of China.
Biol Reprod. 2010 Apr;82(4):759-68. doi: 10.1095/biolreprod.109.082206. Epub 2010 Jan 14.
Our objectives were to study how cysteamine, cystine, and cumulus cells (CCs), as well as oocytes interact to increase oocyte intracellular glutathione (GSH) and thereby to establish an efficient in vitro maturation system for cumulus-denuded oocytes (DOs). Using M16 that contained no thiol as maturation medium, we showed that when supplemented alone, neither cystine nor cysteamine promoted GSH synthesis of mouse DOs, but they did when used together. Although goat CCs required either cysteamine or cystine to promote GSH synthesis, mouse CCs required both. In the presence of cystine, goat CCs produced cysteine but mouse CCs did not. Cysteamine reduced cystine to cysteine in cell-free M16. When TCM-199 that contained 83 microM cystine was used as maturation medium, supplementation with cysteamine alone had no effect, but supplementation with 100 microM cysteamine and 200 microM cystine increased blastulation of DOs matured with CC coculture to a level as high as achieved in cumulus-surrounded oocytes (COCs). Similar numbers of young were produced after two-cell embryos from mouse COCs or CC-cocultured DOs matured with optimal thiol supplementation were transferred to pseudopregnant recipients. It is concluded that 1) mouse CCs can use neither cysteamine nor cystine to promote GSH synthesis, but goat CCs can use either one; 2) goat CCs promote mouse oocyte GSH synthesis by reducing cystine to cysteine, but how they use cysteamine requires further investigation; and 3) mouse DOs can use neither cystine nor cysteamine for GSH synthesis, but they restore developmental capacity completely when matured in the presence of optimum supplementation of cysteamine, cystine, and CCs.
我们的目的是研究半胱氨酸、胱氨酸和卵丘细胞（CC）以及卵母细胞如何相互作用以增加卵母细胞细胞内谷胱甘肽（GSH），从而建立一个有效的卵丘去卵母细胞（DO）体外成熟体系。使用不含巯基的 M16 作为成熟培养基，我们表明，单独补充胱氨酸或半胱氨酸均不能促进小鼠 DO 的 GSH 合成，但两者一起使用时则可以。尽管山羊 CC 需要半胱氨酸或胱氨酸来促进 GSH 合成，但小鼠 CC 则需要两者。在胱氨酸存在的情况下，山羊 CC 产生半胱氨酸，但小鼠 CC 则不产生。半胱氨酸在无细胞的 M16 中将胱氨酸还原为半胱氨酸。当使用含有 83 μM 胱氨酸的 TCM-199 作为成熟培养基时，单独补充半胱氨酸没有效果，但补充 100 μM 半胱氨酸和 200 μM 胱氨酸可将与 CC 共培养成熟的 DO 的囊胚形成率提高到与卵丘环绕卵母细胞（COC）相当的水平。从小鼠 COC 或与 CC 共培养成熟的 DO 的二细胞胚胎中产生的幼仔数量相似，这些胚胎在最佳巯基补充后被转移到假孕受体中。结论是：1）小鼠 CC 既不能使用半胱氨酸也不能使用胱氨酸来促进 GSH 合成，但山羊 CC 可以使用其中任何一种；2）山羊 CC 通过将胱氨酸还原为半胱氨酸来促进小鼠卵母细胞 GSH 合成，但它们如何使用半胱氨酸需要进一步研究；3）小鼠 DO 既不能使用胱氨酸也不能使用半胱氨酸来合成 GSH，但当在最佳半胱氨酸、胱氨酸和 CC 补充的存在下成熟时，它们完全恢复了发育能力。
