The effects of various antioxidants on the development of parthenogenetic porcine embryos.

The major objective of this study was to improve the development rate of parthenogenetic porcine embryos. In this study, the anti-oxidative and anti-apoptotic effects of three antioxidants, ß-mercaptoethanol (ß-ME), α-tocopherol, and extracellular superoxide dismutase (EC-SOD), were examined on the development of parthenogenetic porcine embryos. The development rate of parthenogenetic porcine embryos to the blastocyst stage was 8.1% for control; 19.1%, 14.6%, and 5.0% for 1, 3, and 5 µM ß-ME; 17.2% and 17.5% for 50 and 100 µM α-tocopherol and 12.0% and 4.0% for EC-SOD transgenic mouse embryonic fibroblast (Tg-MEF) and EC-SOD non-transgenic mouse embryonic fibroblast (NTg-MEF) conditioned medium at day 3, respectively. Here, ß-ME, α-tocopherol, and EC-SOD Tg-MEF conditioned medium increased the development rate of parthenogenetic porcine embryos to the blastocyst stage (P < 0.05). The average number of total cells and apoptotic cells at the blastocyst was analyzed at the optimal conditions of the three antioxidants. The three antioxidants increased the average number of total cells at the blastocyst, and they decreased apoptotic cells at the blastocyst as compared to control without supplementation (P < 0.05). When the reactive oxygen species levels in two-cell embryos after 1 µM ß-ME and 100 µM α-tocopherol treatment were examined, those were lower than control group (P < 0.05). In conclusion, it was found that the three antioxidants, ß-mercaptoethanol, α-tocopherol, and EC-SOD Tg-MEF, conditioned medium can play a role as a strong stimulator in the development of parthenogenetic porcine embryos.

The therapeutic effect of extracellular superoxide dismutase (EC-SOD) mouse embryonic fibroblast (MEF) on collagen-induced arthritis (CIA) mice.

Rheumatoid arthritis is a chronic inflammatory disease. The generation of reactive oxygen species (ROS) within an inflamed joint has been suggested as playing a significant pathogenic role. Extracellular superoxide dismutase (EC-SOD) is a major scavenger enzyme of ROS, which has received growing attention for its therapeutic potential. To investigate the therapeutic effect of EC-SOD in mice with collagen-induced arthritis (CIA), we used mouse embryonic fibroblast (MEF) of transgenic mice that overexpresses EC-SOD on the skin by using hK14 promoter. DBA/1 mice that had been treated with bovine type II collagen were administrated subcutaneous injections of EC-SOD transgenic MEF (each at 1.4 x 10(60 cells) on days 28, 35, and 42 after primary immunization. To test EC-SOD activity, blood samples were collected in each group on day 49. The EC-SOD activity was nearly 1.5-fold higher in the transgenic MEF-treated group than in the nontransgenic MEF-treated group (p < 0.05). The severity of arthritis in mice was scored in a double-blind manner, with each paw being assigned a separate clinical score. The severity of arthritis in EC-SOD transgenic MEF-treated mice was significantly suppressed in the arthritic clinical score (p < 0.05). To investigate the alteration of cytokine levels, ELISA was used to measure blood samples. Levels of IL-1beta and TNF-alpha were reduced in the transgenic MEF-treated group (p < 0.05). Abnormalities of the joints were examined by H&E staining. There were no signs of inflammation except for mild hyperplasia of the synovium in the transgenic MEF-treated group. The proliferation of CII-specific T cells was lower in the transgenic MEF-treated mice than in those in the other groups. The transfer of EC-SOD transgenic MEF has shown a therapeutic effect in CIA mice and this approach may be a safer and more effective form of therapy for rheumatoid arthritis.

The parthenogenetic activation of canine oocytes with Ca-EDTA by various culture periods and concentrations.

In the present study, canine oocytes were exposed to various concentrations of and durations of exposure to EDTA saturated with Ca(2+) (Ca-EDTA), a cell membrane-impermeable metal ion chelator, to determine if parthenogenetic activation could be induced. When oocytes were cultured for 48 or 72 h in parthenogenetic activation medium (PAM) without Ca-EDTA (control) or PAM supplemented with 1 or 5mM Ca-EDTA, the highest rate of pronuclear formation (PN) was obtained in oocytes cultured in 1mM Ca-EDTA for 48 h (8.0%; P<0.05). There was no pronuclear formation in the control group (PAM without Ca-EDTA). Oocytes treated with 5mM Ca-EDTA for 48 h or 1mM Ca-EDTA for 72 h formed a parthenogenetic pronucleus (3.1 and 4.5, respectively). However, there was no pronuclear formation in oocytes treated with 5mM Ca-EDTA for 72 h. In summary, exposure to Ca-EDTA can induce pronuclear formation in canine oocytes.