Mouse Cumulus-Denuded Oocytes Restore Developmental Capacity Completely When Matured with Optimal Supplementation of Cysteamine, Cystine, and Cumulus Cells1

Abstract

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.

Antioxidant supplementation overcomes the deleterious effects of maternal restraint stress-induced oxidative stress on mouse oocytes

“…Oocyte activation, IVF, and embryo culture Procedures followed for oocyte activation, IVF, and embryo culture were exactly those reported by Zhou et al (2010), except that zona drilling was not carried out before insemination in this study. Blastocysts obtained were either stained with Hoechst 33342 for cell counting or transferred into surrogate mothers to observe postimplantation development.…”

“…In vitro, however, the synthesis of glutathione may be impaired because of a deficiency of cysteine in the culture medium and its high instability and easy auto-oxidation to cystine (Bannai 1984, Sagara et al 1993. Furthermore, our previous study has shown that neither mouse cumulus cells nor the oocyte itself could use cystine or cysteamine when added alone, but cysteamine could reduce cystine to cysteine in a cell-free medium (Zhou et al 2010). Therefore, cystine and cysteamine were used in combination in this study to promote the synthesis of glutathione in oocytes from restraint-stressed mice.…”

Cumulus cell gene expression following the LH surge in bovine preovulatory follicles: potential early markers of oocyte competence

“…in many mammalian species including mouse (Zhou et al 2010), bovine (Assidi et al 2008), and human (Russell & Robker 2007, Anderson et al 2009). Keeping in mind that most of the transcriptional activity of the oocyte occurred during meiotic arrest before the GVBD (Hyttel et al 2001), the study of the CCs gene expression patterns at the GVBD stage (6 h post-LH) could reflect the oocyte's quality.…”

Aging-related premature luteinization of granulosa cells is avoided by early oocyte retrieval

“…Because glutathione derived from GCs and CCs is required for sperm decondensation and male pronucleus formation, lack of ability to produce glutathione in DOs restrains their development (Perreault et al 1988, Zhou et al 2008, 2010. Hence, when DOs are co-cultured with GC monolayer during IVM, glutathione levels are restored and developmental competence is reestablished (Zhou et al 2008(Zhou et al , 2010. Normal growth and maturation of the oocyte is thus a direct reflection of physiological status of the GCs.…”