Resveratrol protects the mitochondria from vitrification injury in mouse 2-cell embryos
Introduction
With unique advantages, vitrification has been an effective method of oocytes and embryos preservation in mammals and human. With cryobiology development, the survival and development rate of vitrified-warmed embryos was increasingly higher. It is important for oocytes and embryos development that mitochondrial distribution and function match different energy requirement in different areas of cells. However, the inevitable damage caused by vitrification, especially mitochondrial damage, could affect the development of oocytes and embryo seriously [8,24].
Mitochondria, providing energy to cells by oxidative phosphorylation, are the “power stations” and main regulators of cell homeostasis in oocytes and embryos [10,26]. Therefore, mitochondria are crucial to the subsequent development of oocytes and embryos. After vitrification, the change of active mitochondria distribution, closely related to the developmental ability of oocyte and embryo, was observed in the oocytes of mice, pigs and cattle [5,12,23,27]. In addition, studies on the oocytes of dogs and sheep suggested that the normal formation of mitochondrial network was related to subsequent development of oocytes [19,33]. Vitrification could also induce mitochondrial distribution change and membrane potential decrease in mouse pronuclear embryos [40], ATP level decrease in human oocytes [21] and mitochondrial dysfunction in mouse oocytes and parthenogenetic embryos [35]. With mitochondrial dysfunction, there was also a significant increase in intracellular reactive oxygen species (ROS) levels of porcine oocytes after vitrification [13,31].
Resveratrol, a kind of natural antioxidant, has significantly positive effects on the protection for mitochondrial function, the synthesis and activity regulation of antioxidant enzymes and the elimination and production inhibition of ROS [9,25,29,30]. Moreover, resveratrol could promote the sirtuin 1 (SIRT1)-mediated deacetylation, which induced peroxisome proliferators-activated receptor-γ (PPARγ) co-activator 1α (PGC-1α) activity, and improve mitochondrial function eventually [3,17]. It has been believed that resveratrol is one of the strongest activators of SIRT1, and many of its biological functions are mediated by SIRT1 [20], which is considered as a direct ligand of resveratrol [37].
In this study, we detected mitochondrial changes in distribution and function of mouse 2-cell embryos after vitrification using nylon mesh method and demonstrated that resveratrol could repair the damage of mitochondrial distribution and function caused by vitrification.
Section snippets
Chemicals and mice
All chemicals for this study were purchased from Sigma-Aldrich (St. Louis, MO, USA) unless stated otherwise. Plastic dishes and tubes were obtained from Nunc (Roskilde, Denmark). All procedures were approved by the Institutional Animal Care and Use Committee of College of Veterinary Medicine, Northwest A&F University. Kunming outbred mice (The Fourth Military Medical University, Xi'an, China) of 8 weeks were raised according to the Chinese National Standard (GB14925–2001). Superovulation was
Screening suitable concentration of resveratrol treatment for mouse 2-cell embryos
Developing from fresh 2-cell embryos with different concentrations of resveratrol for 72 h, the proportion and total cell number of blastocyst in 0.10 μM group were similar with the control group (Fig. 1). After vitrification, there was no significant difference between control group and 0.1 μM resveratrol group in the blastocyst rate, and the cell number of blastocyst with 0.1 μM resveratrol treatment was significantly higher than that in control group (P < 0.05) (Fig. 1), indicating that
Discussion
Vitrification impact mitochondrial function and lead to the loss of subsequent development ability of mouse and human oocytes after fertilization [2,6,14,18,32]. It was reported that vitrification had significant adverse effects on mitochondrial function of mouse [38], bovine [39] and porcine oocytes [8], because of abnormal mitochondrial ΔΨm, which damage the subsequent development competence of cryopreserved oocytes. Similarly, we confirmed that vitrification not only caused significantly
Declaration of competing interest
No potential conflicts of interest were disclosed.
Acknowledgement
The authors would like to thank other members of the Ma lab for their help in collecting embryos from mice and the support of Life Science Research Core Services (LSRCS), Northwest A&F University for skillful technical assistance. This study was supported by the Strategic Priority Research Program of Chinese Academy of Sciences (XDA 04020222) and Key Research and Development Program of Shaanxi (Program No. 2018NY-023).
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