Relative messenger RNA abundance in bovine oocytes collected in vitro or in vivo before and 20 hr after the preovulatory luteinizing hormone surge.

In the cyclic cow, final maturation of the ovulatory follicle is initiated by the preovulatory luteinizing hormone (LH) surge. During the subsequent 24 hr period, the oocyte nucleus undergoes meiotic progression to metaphase II and several changes in cytoplasmic organization take place. We have previously shown that oocytes recovered at the time of the LH peak and matured in vitro are less competent to reach the blastocyst stage than their counterparts recovered 20 hr later following in vivo maturation, despite both groups undergoing IVF and culture in parallel. The objective of this study was to compare, using real-time quantitative RT-PCR, the relative abundance of various developmentally important gene transcripts in these oocytes. The groups used were mature bovine oocytes originating from: (1) 2-6 mm follicles from slaughterhouse ovaries; (2) preovulatory follicles punctured by ovum pick-up just before the LH surge (i.e., immature) and matured in vitro; or (3) preovulatory follicles punctured 20 hr later, just prior to ovulation (i.e., in vivo matured). In addition, immature oocytes from 2-6 mm follicles were examined. We examined the relative mRNA expression of five enzymes involved in protection against free oxygen radicals (mitochondrial Mn-superoxide dismutase, MnSOD, cytosolic Cu/Zn superoxide dismutase, Cu/ZnSOD, gamma-glutamyl-cysteine transferase, GCS, glutathione peroxidase, GPX, sarcosine oxidase, SOX), a transcript involved in follicular development (growth differentiation factor-9, GDF-9), transcripts involved in glucose metabolism (glucose-6-phosphate dehydrogenase, G6PDH, glucose transporter type-1 and -8, Glut-1, Glut-8) and genes involved in cell cycle events, Cyclin A and B, and poly(A) polymerase (PAP). Transcripts for all genes were detected, irrespective of oocyte origin. While differences were not significant in all cases, variations in levels of transcript abundance between the groups were related to developmental competence. In particular, transcripts for GDF-9 were expressed at significantly higher levels in oocytes recovered at the LH peak and matured in vitro than in those matured in vivo. The observations with GDF-9 are interesting as this gene is believed to be essential for normal folliculogenesis and may be important in the regulation of early follicle and oocyte growth. In conclusion, the results of this study demonstrate differences in the relative mRNA abundance of several developmentally important gene transcripts in bovine oocytes which may be related to developmental competence.

Lonergan P ，Gutiérrez-Adán A ，Rizos D ，Pintado B ，de la Fuente J ，Boland MP ... -  《MOLECULAR REPRODUCTION AND DEVELOPMENT》

Effects of oocyte maturation regimen on the relative abundance of gene transcripts in bovine blastocysts derived in vitro or in vivo.

Knijn HM ，Wrenzycki C ，Hendriksen PJ ，Vos PL ，Herrmann D ，van der Weijden GC ，Niemann H ，Dieleman SJ ... -  《REPRODUCTION》

Glutathione content and glutathione peroxidase expression in in vivo and in vitro matured equine oocytes.

The in vitro developmental competence of equine oocytes is still low in comparison with other domestic mammals. A major factor affecting the viability of cells during in vitro culture is the increased oxidative stress. Oxidative modifications could be responsible for oocyte incompetence for in vitro maturation (IVM). Cysteamine, a glutathione (GSH) synthesis enhancer, has been shown to increase intracellular GSH content and to improve embryo development when added during IVM of bovine, porcine, and ovine oocytes. The aim of the present study was (1) to determine whether equine cumulus-oocyte complexes (COCs) benefit from the addition of cysteamine during IVM, (2) to compare the GSH content of oocytes after in vivo maturation and IVM, (3) to assess whether cysteamine administration during IVM of equine oocyte enhances early embryonic developmental capability following ICSI, (4) to study the glutathione peroxidase (GPX) mRNA level in COCs. In vivo matured COCs were collected by aspiration from preovulatory follicles, and analyzed at collection. Immature COCs were collected in vivo or from slaughterhouse ovaries and matured in culture media supplemented or not with 100 microM cysteamine. After nuclear stage assessment, oocytes were analyzed for GSH concentration and both oocytes and cumulus cells were analyzed for GPX and GAPDH mRNA. Our data showed that the maturation capability was similar in both in vivo aspirated oocytes and in those isolated from slaughterhouse ovaries. Moreover, the addition of cysteamine during IVM affected neither GSH content nor maturation rate. At the time of collection, intra-oocyte GSH content was not influenced by the chromatin status. GSH concentration was similar in in vivo and in vitro matured metaphase II (MII) stage oocytes, and was significantly higher in MII than immature germinal vesicle stage oocytes. Moreover, the presence of serum inhibited whereas its absence stimulated the accumulation of GSH within oocytes during IVM. After ICSI, a similar proportion of zygotes in each group developed beyond the two-cell stage after 72 hr of culture. Cumulus cells expressed GPX mRNA, while GPX transcript was absent in both immature and mature oocytes. Cumulus expression of GPX mRNA was significantly higher when analyzed at collection than after IVM. Taken together, our results demonstrate that in equine oocytes, GSH increases during IVM but the relative intra-oocyte content of this thiol does not affect maturation and early development efficiency after fertilization. We hypothesize that factor(s) other than GSH/GPX are responsible for the limited in vitro early developmental capability of equine oocytes.

Luciano AM ，Goudet G ，Perazzoli F ，Lahuec C ，Gérard N ... -  《MOLECULAR REPRODUCTION AND DEVELOPMENT》

Impact of the LH surge on granulosa cell transcript levels as markers of oocyte developmental competence in cattle.

Gilbert I ，Robert C ，Vigneault C ，Blondin P ，Sirard MA ... -  《-》

Effect of stage of follicular growth during superovulation on developmental competence of bovine oocytes.

Humblot P ，Holm P ，Lonergan P ，Wrenzycki C ，Lequarré AS ，Joly CG ，Herrmann D ，Lopes A ，Rizos D ，Niemann H ，Callesen H ... -  《THERIOGENOLOGY》