The influence of 9-cis-retinoic acid on nuclear and cytoplasmic maturation and gene expression in canine oocytes during in vitro maturation

Abstract

Retinoids have important roles in regulation of oocyte nuclear and cytoplasmic maturation. The present study investigated the effects of a retinoid metabolite on nuclear maturation, cytoplasmic maturation, and gene expression in canine oocytes during in vitro maturation (IVM). Cumulus-oocyte complexes (COCs) were harvested from ovaries by slicing. Only oocytes that were >120 μm in diameter, with a homogeneous dark cytoplasm and three or more layers of compact cumulus cells were used. Varying concentrations of 9-cis retinoic acid (9-cis-RA; 0, 5, 50, and 500 nm) were included in the maturation medium, and the following were measured: (i) oocyte nuclear maturation after culture for 48 h; (ii) cytoplasmic granular migration by labeling of oocytes with fluorescein isothiocyanate labeled lectins; and (iii) relative expression of genes related to apoptosis (BAX and BclII) in cumulus cells detached from oocytes, by semiquantitative reverse transcriptase-polymerase chain reaction. After 48 h culture with IVM, the highest percentage of oocytes that had developed to the metaphase II (MII) stage were in the 5 nm 9-cis-RA treatment group (18.3 ± 2.5%; P < 0.05). Complete granular migration was observed in oocytes matured with 5 nm 9-cis-RA, consistent with a commensurate gain in developmental competence. Treatment with 5 nm 9-cis-RA had no effect on BclII gene expression, but downregulated BAX expression. In conclusion, since 5 nm 9-cis-RA was beneficial to nuclear and cytoplasmic maturation of canine oocytes, we inferred an important role for 9-cis-RA during IVM.