Effect of roscovitine, a cdk1 inhibitor, and of the presence of oocyte on bovine cumulus cell expansion and cyclooxygenase‐2 expression

Abstract

Oocyte maturation is accompanied by differentiation of surrounding cumulus cells. These cells produce hyaluronic acid (HA) and its storage in intercellular spaces results in expansion of the cells. The cumulus cells also accumulate cyclooxygenase-2 (cox-2) during maturation. Both expansion and cox-2 storage are regulated by FSH and EGF. The aim of this study was to determine whether oocyte meiotic resumption is involved in the regulation of cumulus differentiation or not. We investigated the effects of roscovitine, a reversible inhibitor of meiosis resumption of cattle oocytes on EGF induced expansion and cox-2 expression at the transcript and protein levels respectively (RT-PCR and Western blot), in cumulus oocyte complexes (COCs) and cumulus complexes alone (CCs). EGF induced expansion and cox-2 expression in both COCs and CCs. These effects were prevented by roscovitine, whether in the presence or in the absence of oocyte. However, the oocyte was essential for the reversibility of inhibition by roscovitine. In conclusion, our results indicate that i) oocyte secreted-factors are not essential for cumulus expansion, and ii) roscovitine mediated inhibition of meiotic resumption also respects the functionality of the surrounding somatic cells.

Maintenance of meiotic arrest in bovine oocytes using the S-enantiomer of roscovitine: effects on maturation, fertilization and subsequent embryo development in vitro

“…The disparity in the dose of roscovitine utilized by others (Mermillod et al 2000, Calder et al 2001, Duque et al 2002, Lagutina et al 2002, Vigneron et al 2003 versus that required in our study may be due to differences in roscovitine activity related to source, use of a different enantiomer, differences in the composition of the roscovitine medium or other unidentified factors. Dr Laurent Meijer has provided the roscovitine used in the majority of available studies (Mermillod et al 2000, Lagutina et al 2002, Lonergan et al 2003, Vigneron et al 2003. In contrast, we utilized a commercial source of the (S)-enantiomer.…”

“…Numerous studies have described the effectiveness of the R-enantiomer of roscovitine to meiotically inhibit bovine oocytes (Mermillod et al 2000, Kasinathan et al 2001, Lagutina et al 2002, Vigneron et al 2003. However, reported differences in the potency of the R-versus the S-enantiomer of roscovitine to inhibit p34 cdc2 kinase activity (De Azevedo et al 1997, reviewed by Meijer & Raymond 2003 along with recent unpublished observations (L Meijer, unpublished observations) showing increased selectivity towards other protein kinases, warrant further investigation of the S-enantiomer of roscovitine.…”

Expression of genes involved in prostaglandin E2 and progesterone production in bovine cumulus–oocyte complexes during in vitro maturation and fertilization

“…The 20a-HSD enzyme catalyzes the conversion of progesterone into its inactive form, 20a-hydroxyprogesterone. We used a wellestablished model for in vitro maturation (IVM) inducing differential levels of oocyte maturation, cumulus expansion, and PTGS2 expression (Nuttinck et al 2002, Vigneron et al 2003. Differential expression of the studied enzymes was analyzed in relation to the rate of in vitro fertilization (IVF) and subsequent embryo development.…”

Epidermal growth factor activates cytosolic [Ca2+] elevations and subsequent membrane permeabilization in mouse cumulus–oocyte complexes

“…Enhanced cumulus expansion due to EGF stimulation in bovine COCs is blocked by roscovitine, an inhibitor of maturation/M-phase promoting factor (MPF) kinase activity (Vigneron et al 2003) and the tyrosine kinase activity of the EGF receptor (EGFR) is required for EGF-stimulated expansion in pig COCs (Prochazka et al 2003). Based on its ability to directly enhance cytoplasmic maturation in mouse and human oocytes (Das et al 1991), inclusion of EGF in in vitro fertilization (IVF) procedures may serve to improve success.…”