| Autophagy is an important biological process in the development and maturation of oocytes.2-Mercaptoethanol(2-ME)is often used as an antioxidant to optimize the in vitro maturation culture system of domestic oocytes.However,systematic studies on the effect of 2-ME on autophagy in vitro maturation of porcine oocytes are still lacking.Based on this,this study hypothesized that 2-ME can promote the in vitro maturation of oocytes by maintaining autophagy homeostasis.To test this hypothesis,we investigated the effect of 2-ME on the in vitro maturation of porcine oocytes inhibited by autophagy(3-Methyladenine,3-MA)or overactivated(Rapamycin).The conclusions are as follows:(1)Effects of 2-ME on in vitro maturation and parthenogenetic embryo development of porcine oocytes exposed to 3-MAIn order to investigate the effect of 3-MA on oocyte maturation in vitro,different concentrations of 3-MA(0,0.1,1,10,100 m M)were added to the culture medium.The results showed that the first polar body extrusion rate decreased gradually with the increase of 3-MA concentration,where 1 m M 3-MA can significantly reduce the polar body extrusion rate of porcine oocytes,resulting in down-regulation of autophagy,abnormal cumulus expansion,impaired cytoskeleton integrity,mitochondrial dysfunction,increased lipid droplet,increased ROS,and increased early apoptosis,ultimately resulting in impaired maturation of oocytes and decreased development quality of parthenogenetic embryos.Subsequent addition of different concentrations of 2-ME(0,10,25,50,100 μM)to 1 m M 3-MA medium showed that the addition of 25 μM 2-ME significantly alleviated a series of damage caused by the inhibition of autophagy in oocytes due to 3-MA exposure,and promoted the development of parthenogenetic embryos(2)Effects of 2-ME on in vitro maturation and parthenogenetic embryo development of Rapamycin-exposed porcine oocytesIn order to explore the effect of Rapamycin on the autophagy of oocytes overactivated,different concentrations of Rapamycin(0,1,2,4 n M)were added to the culture medium,and the results showed that the concentration was 2 n M Rapamycin significantly reduced the polar body extrusion rate of oocytes in vitro,and the results showed that autophagy in Rapamycin pig oocytes was up-regulated,cytoskeleton integrity was impaired,mitochondrial function was impaired,reactive oxygen species were produced,and early apoptosis was increased,ultimately resulting in maturation and culture of oocytes and decreased qua lity of embryo development in later stage.Then different concentrations of 2-ME(0,10,25,50,100 μM)were added to the medium containing 2 n M Rapamycin.It was found that the addition of 25 μM 2-ME effectively mitigated a series of damage caused by autophagy overactivation caused by Rapamycin exposure and promoted the development of parthenogenetic embryos.In conclusion,this study confirmed that 2-ME can promote in vitro maturation of porcine oocytes by maintaining autophagy homeostasis.This study can provide a new perspective for the mechanism study of 2-ME,and provide important theoretical and practical value for the optimization of porcine oocyte maturation system. |
