This study was undertaken to evaluate the relationship between in vitro maturation and plasminogen activators (PAs) activity on porcine cumulus-oocytes complexes (COCs) exposed to oxidative stress. When COCs were cultured in maturation medium with hydrogen peroxide (H2O2), the proportion of the germinal vesicle breakdown (GVBD) and oocytes maturation were decreased with addition of H2O2, and were significantly (p<0.05) lower in medium with 0.1 mM H2O2 than control group. Also, the rate of degenerated oocytes was increased in as H2O2 concentration increased. When COCs were cultured for 48 h, three plasminogen-dependent lytic bands were observed: tissue-type PA (tPA); urokinase-type PA (uPA); and tPA-PA inhibitor (tPA-PAI). PA activity was quantified using SDS-PAGE and zymography. When H2O2 concentration was increased, tPA and tPA-PAI activities also increased in porcine oocytes cultured for 48 h, but not uPA. In other experiment, embryos were divided into three groups and cultured in (1) control medium, (2) control medium with 1.0 mM H2O2 and (3) control medium with 1.0 mM H2O2 along with catalase in concentrations of 0.01, 0.1, and 1.0 mg/ml, respectively. H2O2 decreased the rate of GVBD and maturation in porcine COCs but catalase revealed protective activity against oxidative stress caused by H2O2. In this experiment, tPA and tPA-PAI activities were higher in media with 1.0 mM H2O2 alone. Increasing concentration of catalase decreased tPA and tPA-PAI activities in porcine oocytes. These results indicate that the exposure of porcine follicular oocytes to ROS inhibits oocytes maturation to metaphase-II stage and increase the oocytes degeneration. Also, we speculated that increased ROS level may trigger tPA and tPA-PAI activities in porcine oocytes matured in vitro.

Antioxidants partially ameliorated the detrimental effects of reactive oxygen species (ROS) on sperm characteristics during in vitro storage. The objective of the present study was to investigate the single or synergetic antioxidative effect of curcumin and Vit. E on the characteristics of fresh boar sperm during in vitro storage. The sperm viability in curcumin, Vit. E supplementation and curcumin+Vit. E+H2O2 groups remained over 85.0% in 3 hr incubation period, but in 6 hr incubation period, curcumin+Vit. E+H2O2 groups was sharply dropped than those of curcumin and Vit. E group. The membrane intergrity in all evaluated groups except for H2O2 group did not significantly difference in 3 hr incubation period. The viability in curcumin or Vit. E supplementation were significantly increased than in curcumin+H2O2 and Vit. E+H2O2 group in 6 hr incubation period. The percentage of mitochondrial activity and acrosome intergrity obtained similar trends within same incubation periods irrespective of treatment. The lipid peroxidation of spermatozoal plasma membrane ranged from 11.6∼17.5 nM/l×106 and 14.0∼ 19.0 nM/l×106 in 3 hr and 6 hr incubation periods. In conclusion, curcumin or Vit. E rpplementation alone or cooperatively improved sperm viability index (motility, membrane intergrity, viability and survival rates) and fertility index (mitochondria activity, acrosome intergrity and lipid peroxidation) of fresh boar sperm, indicating that curcumin and Vit. E have a antioxidative properties through its scavenging activity against hydrogen peroxide.

The aim of this study was to evaluate the effects of different straw volume (0.5 and 5.0 ㎖) on cryopreservation in boar semen. Semen quality was evaluated the motility, viability, abnormality, acrosome intactness and membrane integrity. And there were also examined the development rates of IVM/IVF embryos using frozen-thawed boar semen. Boar semen were frozen until 5℃ for 3 hours using cell freezer and making the straws, and then freezing by lowing the straws into styrofoam box on the 8 cm above the LN2. In different straw volume (0.5 and 5.0 ㎖), sperm viability and abnormality were not differ in 0.5 and 5.0 ㎖ straws, but sperm motility were significantly higher in 0.5 ㎖ straws (61.3%) than in 5.0 ㎖ straws (56.3%) (p<0.05). In the Coomassie Brillient Blue (CBB), Hoechst 33258/Propidium Iodide (H258/PI) staining and Hypoosmotic Swelling Test (HOST), the acrosome intactness and sperm membrane integrity were not differ in 0.5 and 5.0 ㎖ straws, but sperm survival rate was significantly higher in 0.5 ㎖ straw (65.0%) than in 5.0 ㎖l straw (55.0%) (p<0.05). Employing the Chlortetracycline (CTC)/Hoechst33258 (H258), all treatment group were not differ in characteristic of uncapacitated acrosome-intact sperm (F), capacitated and acrosome-intact sperm (B-type) and acrosomereaction seprm (AR-type). In the developmental rate of IVM/IVF embryos using frozen-thawed boar semen in different straw volumes, the developmental rate of morula plus blastocysts were 19.9% in 0.5 ㎖ straw and 18.8% in 5.0 ㎖ straw, respectively. These results indicated that straw volume affects the semen motility and sperm survival rate, but not other semen characteristic and developmental rate of IVM/IVF embryos.