Abstract：In vitro culture system influences the development potential of mammalian early embryos. In the current culture systems, which simulating the hypoxic condition in vivo of uterus and fallopian tubes, the reactive oxygen species (ROS) is considered to be a major factor affecting embryo development. In this study, the effect of ROS on the mitochondrial function and the mitochondrial DNA (mtDNA) copy number in mice (Mus musculus) 2-cell embryos were studied. The fertilized eggs from the Kunming mouse were treated by 2,2'-azobis (2-methylpropionamidine) dihydrochloride (AAPH), then the mitochondrial activity, the mitochondrial membrane potential, the mitochondrial ROS (mtROS) and the distribution of mitochondrial were observed by fluorescent detection, meanwhile, the expression of ROS-related genes, the mitochondrial transcription factor A (TFAM) and the mitochondrial DNA (mtDNA) copy number were detected by real-time PCR. The results showed that mouse embryos had a higher blocking rate (66.16%) and a lower embryo damage rate (24 h malformation rate of 9.09%, 48 h mortality 10.39%) in 1.0 mmol/L treated group. ROS staining results showed that embryonic cytoplasmic ROS levels were increased significantly after AAPH treatment, and the mRNA expression of ROS-related genes were also increased. After treated by 1.0 mmol/L AAPH, the membrane potential, the mtDNA copy number and the level of mitochondrial ROS (mtROS) in 2-cell embryos were significantly higher than that of control group, and the expression of TFAM gene was also increased. The above results showed that the oxidative stress caused by AAPH resulted in embryo development block, the function of mitochondrial was promoted after treatment, with increased the mtDNA copy number and the expression of TFAM gene, which may participate in regulation of intracellular ROS dynamic balance in embryos. This study preliminarily explored the relationship between the ROS and the dynamic change of mitochondrial function, and might provide references for illustrating the mechanism of oocyte maturation and also optimization the culture system of mammalian embryos in vitro.
