Efeitos da suplementação de anetol ou robinina na vitrificação e incubação in vitro do tecido ovariano ovino

Abstract

The objective of the present work was to verify the effect of the addition of natural antioxidants, anethole (AN) and robinine (RO), in the vitrification and the in vitro incubation solution of ovine ovarian tissue. Initially (step 1), was analyzed the percentage of morphologically normal preantral follicles (MNPF) included in fresh ovarian tissue (Control - CTR), vitrified in the absence of antioxidants (VSA), only incubated in vitro (IIV) for 24 h or vitrified in the presence of different concentrations of anethole (AN: 30, 300, 2000 µg / mL) or robinin (RO: 0.125, 0.25, 0.50 mg / mL), followed by in vitro incubation, totaling nine treatments. In stage 2, the treatments, CTR, VSA, IIV, AN2000 and RO1.25 were maintained and added both types and concentrations of antioxidants in the in vitro incubation, resulting in treatments AN2000 + and RO1,25 + , totaling seven treatments. After 24 h of incubation, the following parameters were evaluated: follicular activation, stromal cell density (SCD), ROS levels in the ovarian fragments and in the incubation medium, total antioxidant capacity (TAC) and mitochondrial activity. Data were submitted to the SAS statistical package, and the differences were found statistically significant when P <0.05. The data from step 1 showed that the percentage of MNPF in AN2000 was similar to CTR and similar to RO0.125. In step 2, foilicular activation in all treatments was superior (p<0.05) to CTR and in RO0.125 decrease to compare (p<0.05) VSA. Although the DCE in all the vitrified fragments was inferior (p<0.05) to the CTR, in the traces AN2000 and RO0.125 this parameter was superior to the VSA. The ROs levels in the ovarian cortex from the AN2000 or AN2000 + were lower than the CTR. The intracellular ROS levels was increase in the VSA, RO0,125 and IIV treatments, compared to the CTR. On the other hand, this parameter was lower in AN2000 when compared to VSA and similar compare to CTR. Regarding TAC, the RO0.125 was better than the VSA, AN2000 and AN2000+. According to the results obtained, we conclude that the use of antioxidants (anethole or robinine) in the vitrification solution of ovine ovarian tissue is recommended; since AN 2000 µg / mL preserved the follicular morphology and RO 0.125 mg / mL maintained a higher TAC than vitrification in the absence of antioxidants.