Justicia insularis Improves the in vitro Survival and Development of Ovine Preantral Follicles Enclosed in Ovarian Tissue

Objectives: Evaluating the addition effect of J. insularis extract and FSH on the survival, activation and ROS production after in vitro culture of ovine preantral follicles enclosed in ovarian tissue. Methods: In the first experiment, ovarian fragments were fixed (non-cultured control) or in vitro cultured in α-MEM (cultured control), α-MEM supplemented with FSH 50 ng/mL, or in α-MEM supplemented with J. insularis (JUS0.3; 1.25 or 5 mg/mL) for 1 or 7 days, at 39 ̊C, 5% CO2. In the second experiment, fragments were fixed or cultured in α-MEM supplemented with anethole 300 μg/mL + FSH 50 ng/mL or in α-MEM supplemented with anethole 300 μg/mL + 0.3 mg/mL JUS. Key findings: JUS0.3 was the only treatment that maintained the percentage of morphologically normal follicles similar to non-cultured control even after 7 days of culture. After 7 days of culture, a higher (p < 0.05) percentage of developing follicles was observed in JUS5 treatment compared with the other treatments except JUS1.25. In the second experiment, FSH maintained the percentage of normal follicles and promoted activation of primordial follicles. A reduction (p < 0.05) of stromal cell density was observed in MEM+ANE supplemented with JUS or FSH. Conclusions: J. insularis in a concentration-dependent manner maintained the levels of ROS and improved in vitro follicular survival and activation of ovine primordial follicles.