earticle

영어

The reactive oxygen species (ROS) generated during the somatic cell transfer nuclear (SCNT) procedures may cause the mitochondrial dysfunction and DNA damage, which may result in restricts the reprogramming of SCNT embryos and play a key direct role in apoptosis. The present study was conducted to investigate the effect of antioxidant treatment during the SCNT procedures on the inhibition of mitochondria and DNA damages in bovine SCNT embryos. The reconstituted oocytes were treated with antioxidants of 25 μM β-mercaptoethanol (β-ME) or 50 μM vitamin C (Vit. C) during the SCNT procedures. In vitro fertilization (IVF) was performed for controls. Mitochondrial morphology and membrane potential (ΔΨ) were evaluated by staining the embryos with MitoTracker Red or JC-1. Apoptosis was analyzed by Caspase-3 activity assay and TUNEL assay, and DNA fragmentation was measured by comet assay at the zygote stage. Mitochondrial morphology of non-treated SCNT embryos was diffused within cytoplasm without forming clumps, while the IVF embryos and antioxidant treated SCNT embryos were formed clumps. The ΔΨ of β-ME (1.3±0.1, red/green) and Vit. C-treated (1.4±0.2, red/green) SCNT embryos were significantly higher (p<0.05) than that of non-treated SCNT embryos (0.9±0.1, red/ green), which similar to that of IVF embryos (1.3±0.1, red/green). Caspase-3 activity was not difference among the groups. TUNEL assay also revealed that little apoptosis was occurred in SCNT embryos as well as IVF embryos regardless of antioxidant treatment. Comet tail lengths of β-ME and Vit. C-treated SCNT embryos (337.8±23.5 μm and 318.7 ±27.0 μm, respectively) were shorter than that of non-treated SCNT embryos (397.4± 21.4 μm) and similar to IVF embryos (323.3±10.6 μm). These results suggest that antioxidant treatment during SCNT procedures can inhibit the mitochondrial and DNA damages of bovine SCNT embryos.