Bae Hyo-Kyung, Hwang In-Sun, Kim Ji-Ye, Lee Sung-Young, Park Choon-Keun, Yang Boo-Keun, Cheong Hee-Tae
College of Veterinary Medicine and Institute of Veterinary Science, Kangwon National University, 1 Kangwondaehak-gil, Chuncheon-si, Gangwon-do, 200-701, Korea.
College of Animal Resource Science, Kangwon National University, 1 Kangwondaehak-gil, Chuncheon-si, Gangwon-do, 200-701, Korea.
Reprod Fertil Dev. 2015 Sep;27(7):1088-96. doi: 10.1071/RD14027.
We tried to prevent the mitochondrial and DNA damage caused by mechanical stress-associated reactive oxygen species (ROS), and to improve the reprogramming of bovine somatic cell nuclear transfer (SCNT) embryos by antioxidant treatment during the manipulation procedures of SCNT. Bovine recipient oocytes and reconstituted oocytes were treated with antioxidants during manipulation procedures. The H2O2 level, mitochondrial morphology, membrane potential and apoptosis at the one-cell stage, and in vitro development and DNA methylation status of blastocysts were evaluated. Antioxidant treatment during manipulation procedures reduced the H2O2 level of SCNT embryos. Antioxidant-treated SCNT embryos normally formed mitochondrial clumps, similar to IVF embryos, and showed higher mitochondrial membrane potential versus the SCNT control (P<0.05). Apoptosis and DNA fragmentation were reduced by antioxidant treatment. The development rate to the blastocyst stage was higher (P<0.05) in the antioxidant treatment groups (30.5±2.5 to 30.6±1.6%) versus the control (23.0±1.9%). The DNA methylation status of blastocysts in the antioxidant treatment groups was lower (P<0.05) than that of the control and similar to that of IVF embryos. These results indicate that antioxidant treatment during manipulation procedures can prevent cellular damage that may be caused by mechanical stress-associated ROS, and improve nuclear reprogramming.
我们试图预防由机械应激相关活性氧（ROS）引起的线粒体和DNA损伤，并在牛体细胞核移植（SCNT）胚胎操作过程中通过抗氧化剂处理改善其重编程。在操作过程中，对牛受体卵母细胞和重构卵母细胞进行抗氧化剂处理。评估了单细胞阶段的过氧化氢水平、线粒体形态、膜电位和凋亡情况，以及囊胚的体外发育和DNA甲基化状态。操作过程中的抗氧化剂处理降低了SCNT胚胎的过氧化氢水平。抗氧化剂处理的SCNT胚胎正常形成线粒体团块，类似于体外受精（IVF）胚胎，并且与SCNT对照组相比显示出更高的线粒体膜电位（P<0.05）。抗氧化剂处理减少了凋亡和DNA片段化。抗氧化剂处理组的囊胚发育率（30.5±2.5至30.6±1.6%）高于对照组（23.0±1.9%）（P<0.05）。抗氧化剂处理组囊胚的DNA甲基化状态低于对照组（P<0.05），且与IVF胚胎相似。这些结果表明，操作过程中的抗氧化剂处理可以预防由机械应激相关ROS可能引起的细胞损伤，并改善核重编程。
