Immunolocalization of leptin and its receptor in the sheep ovary and in vitro effect of leptin on follicular development and oocyte maturation.

The aims of the study were to characterize leptin and it is receptor (LEPR) proteins immunoexpression in ovine ovaries and to evaluate the effects of leptin on development of secondary follicles cultured in vitro. The ovaries were collected and fixed for immunohistochemical analysis. Additional pairs of ovaries were collected and secondary follicles were isolated and cultured, for 18 days, in α-MEM+ alone or supplemented with 10 or 25 ng/mL of leptin. The antrum formation and fully grown oocytes rates were higher in 25 ng/mL leptin than all treatments. GSH levels and mitochondrial activity were higher in 10 or 25 ng/mL leptin than α-MEM+. 25 ng/mL leptin showed a higher percentage of MII than the α-MEM+. In conclusion, leptin and its receptor are expressed in ovine ovaries and 25 ng/mL leptin promoted higher in vitro maturation rates by improving follicular development, GSH levels and mitochondrial activity of ovine oocytes compared to control medium.

Effect of red propolis extract isolated or encapsulated in nanoparticles on the in vitro culture of sheep preantral follicle: Impacts on antrum formation, mitochondrial activity and glutathione levels.

This study aimed to characterize the hydroethanolic extract of red propolis (HERP) and nanoparticles containing HERP for using as an additive in the culture medium of isolated ovine preantral follicles. HERP was characterized by high-performance liquid chromatography (HPLC) and determination of flavonoid content, and the nanoparticles by the mean particle diameter, polydispersity index (PI) and encapsulation efficiency (EE). The effect of HERP (10 and 20 Î·g/ml-HERP10 and HERP20 groups) and nanoparticles (NP10 and NP20 groups) on isolated secondary follicles cultured in vitro for 12 days was observed by morphological evaluation, oxidative stress markers (reactive oxygen species-ROS and glutathione-GSH) and active mitochondria. HPLC showed formononetin as the major compound in the HERP (63.92 ± 0.21 µg/mL). The content of flavonoids ranged from 2.14% to 11.00%. The nanoparticles showed mean diameter of 287.5 ± 3.9 and 479 ± 18.1 Î·m; PI of 0.117 ± 0.018 and 0.316 ± 0.039; and EE of 67.15% and 41%, respectively, for the NP10 and NP20 groups. After 12 days of culture, HERP20 and NP20 increased (p < 0.05) the percentage of normal follicles compared to NP10. HERP20 showed significantly higher percentages of antrum formation than control medium (MEM) and NP10 (p < 0.05). HERP20 also showed an increase (p < 0.05) in mitochondrial activity compared to the other treatments, except NP20 (p > 0.05), and increased GSH levels (p < 0.05) compared to MEM and HERP10. In conclusion, use of HERP (20 Î·g/ml) on in vitro culture of isolated ovine preantral follicles can increase antrum formation, mitochondrial activity and GSH levels.