Effect of roscovitine and cycloheximide on ultrastructure of sheep oocytes

Abstract

It is believed the temporary meiosis arrest with roscovitine or cycloheximide may improve the in vitro developmental competence of oocytes in different animal species. However, little is known about the effects of these inhibitors on ultrastructure of ovines cumulus-oocyte complexes (COCs). The aim of this study was to evaluate the progression of cytoplasmic maturation and the ultrastructural changes in sheep COCs exposed to roscovitine or cycloheximide, at acceptable concentrations. COCs were in vitro cultured for 24 h in maturation medium (control group) containing 100 M roscovitine or 1 g/mL cycloheximide (treatment groups). After this time, some COCs were cultured for further 22 h in inhibitor-free medium. The ultrastructure organization of COCs was evaluated by transmission electron microscopy before (immature group) and after in vitro culture for 24 and 46 h. As expected, signs of immaturity and maturity were observed in immature and control groups, respectively. In treatment with roscovitine, there were cumulus cells degeneration, swelling of mitochondrias, few cortical granules and many vesicles with electron-dense material. However, in cycloheximide treatment there were not signs of degeneration or cellular senescence. Metabolic units and mitochondrial pleomorphism were found in all experimental groups. These evidences demonstrate that roscovitine promoted irreversible ultrastructural changes while cycloheximide did not affect the cytoplasmic maturation. However, the implications on embryo development are still unclear.

In vitro Developmental Competence of Adult Sheep Oocytes Treated with Roscovitine

“…Irreversible alterations such as degeneration of cumulus cells, swelling of mitochondria and reduction of cortical granules were reported by Crocomo et al. () in sheep COCs treated for 24 h with 100 μ m roscovitine. Later, maximal efficiency of meiosis inhibition was reached by Crocomo et al.…”

“…According to these same authors, the speed of development of roscovitine treated embryos was faster compared to Control. These divergent results are probably related not only to experimental conditions but also to intrinsic oocyte quality and species peculiarities (Han et al 2006;Crocomo et al 2013).…”

“…In sheep, the inhibitory potential of roscovitine as well as its effects on ultrastructure of cumulus– oocyte complexes has been recently investigated (Crocomo et al. , ,b). Irreversible alterations such as degeneration of cumulus cells, swelling of mitochondria and reduction of cortical granules were reported by Crocomo et al.…”

“…In sheep, the inhibitory potential of roscovitine as well as its effects on ultrastructure of cumulus-oocyte complexes has been recently investigated (Crocomo et al 2013(Crocomo et al , 2015a. Irreversible alterations such as degeneration of cumulus cells, swelling of mitochondria and reduction of cortical granules were reported by Crocomo et al (2013) in sheep COCs treated for 24 h with 100 lM roscovitine. Later, maximal efficiency of meiosis inhibition was reached by Crocomo et al (2015a,b) with 86.7% of oocytes kept at GV stage after treatment for 6 h with 75 lM roscovitine in serumsupplemented medium without oil overlay.…”

Effect of Oil Overlay on Inhibition Potential of Roscovitine in Sheep Cumulus‐Oocyte Complexes

“…Besides, irreversible ultrastructural alterations were recorded in the cumulus cells and oocytes of sheep (Crocomo et al. ) and bovine (Lonergan et al. ) COCs treated for 24 h with 100 and 125 μ m roscovitine, respectively.…”