Effect Of Resveratrol On Lipopolysaccharide Exposed Porcine Oocyte Meiosis Maturation Defects

Lipopolysaccharide(LPS),a significant virulence factor of gram-negative bacteria,adversely affects female reproduction,especially the maturation and early embryonic development of oocytes,through inducing of in flammatory and oxidative stress-associated toxic responses.Resveratrol(Res),a potent antioxidant,exhibits many beneficial effects on the maturation and developmental competence of oocytes by regulating redox balance and mitochondrial function.However,it is unclear whether Res can restore LPS-induced defects of porcine oocytes during meiosis.In this study,porcine oocytes were randomly divided into control group,LPS group and Res group,and cultured for MI(26 h)or MII(44 h)in vitro.Then,nuclear maturation and cytoplasmic maturation were evaluated by in vitro maturation(IVM),in vitro culture(IVC)of embryos,immunofluorescence staining(IF)and other techniques,to explore the recovery effect of Res on meiotic maturation defects of LPS-exposed porcine oocytes and its related mechanism.The results are as follows:(1)Effects of Res on meiotic maturation in LPS-exposed porcine oocytesIn order to explore the alleviating effect of Res on the meiosis maturation defects in LPS-exposed porcine oocytes,we used the established laboratory model of 15μg/m L LPS on porcine oocyte meiosis injury.In vitro maturation medium TCM199,different concentrations of Res(5,10,20μM)were added,and oocytes were randomly cultured for 44 h to calculate the rates of cumulus expansion and polar body expulsion.The results showed that the addition of 10μM Res significantly restored cumulus expansion and polar body expulsion of oocytes exposed to LPS.Therefore,10μM Res was administered in the subsequent experiments.We next found that Res alleviated meiotic abnormalities(spindle assembly and chromosome arrangement),organelle distribution abnormalities(actin migration and cortical particle distribution),metabolic abnormalities(mitochondrial dysfunction,decreased lipid drome number,and decreased glucose uptake capacity)in LPS-exposed oocytes,and reduced oxidative stress,DNA damage,and early apoptosis.m~6A,H3K27me2modification levels were maintained to recover oocyte meiosis maturation defects.These results indicate that Res can effectively improve the meiotic maturation defects of LPS-exposed porcine oocyte.(2)Effects of Res alleviates LPS-exposed porcine oocyte maturation defects on early embryonic development of parthenogenetic activationIn order to explore the effect of Res on parthenogenesis activation of porcine oocytes exposed to LPS,we performed parthenogenesis activation of oocytes from each group after in vitro maturation culture and tested the potential of subsequent embryonic development.The results showed that Res could effectively relieve the early embryo division rate and blastocyst rate,the number of cells in blastocyst,and blastocyst cell activity of parthenogenesis activated LPS-exposed porcine oocyte.These results indicate that the alleviating effect of Res on meiotic maturation defects of LPS-exposed porcine oocytes could further restore the embryonic developmental competence of parthenogenetic activation.(3)Mechanism of Res improving meiotic maturation defects of LPS-exposed porcine oocytesIn order to explore whether Res improves LPS exposed porcine oocyte meiosis maturation defects through SIRT1 pathway,we added SIRT1 specific inhibitor EX527 group on the basis of the above grouping.The results showed that EX527 significantly inhibited the recovery of Res on SIRT1 protein expression in LPS-exposed porcine oocytes,and inhibited the recovery effect of Res on pole-body expulsion rate,reactive oxygen species,mitochondrial content,spindle assembly and chromosome arrangement abnormalities in LPS-exposed porcine oocytes.These results suggest that Res can ameliorates meiotic maturation defects of LPS-exposed porcine oocytes through SIRT1 pathway.In conclusion,this study indicate that Res can effectively relieve the damage of meiotic maturation of LPS-exposed porcine oocytes and embryonic developmental potential by parthenogenesis.Res plays a role by activating SIRT1 pathway to relieve oxidative stress and restore mitochondrial activity.This study provides a theoretical basis for using natural antioxidants to restore the maturation defects of porcine oocytes caused by LPS exposure in livestock production.