Mechanism Of Spermidine In Regulating Ovarian Oxidative Stress And Granulosa Cells Apoptosis In ICR Mice

Spermidine,as an important polyamine in the organism,plays vital roles in various life activities of the body,including the process of cell growth,tumorigenesis and apoptosis,gene transcription and translation,protein synthesis,maintenance of cell membrane structure stability and regulation of ion channels.In the process of catabolism,spermidine produces metabolites such as hydrogen peroxide and acrolein,which cause oxidative stress in cells.Many diseases of the body are closely related to the regulation of oxidative stress.In mammalian ovaries,oxidative stress-induced apoptosis of ovarian granulosa cells is the main cause of follicular atresia.In addition,ovarian oxidative stress is also an important cause of many female reproductive system diseases.Studies have shown that high concentrations of spermidine can induce apoptosis by producing large amounts of hydrogen peroxide and acrolein.At present,the effect of spermidine on oxidative damage of mouse ovary and its mechanism of inducing apoptosis of granulosa cells have not been reported.To explore the mechanism of high-dose polyamines on ovarian oxidative damage in mice and to provide theoretical basis for toxicological studies of polyamines and animal ovarian function studies.In this study,mice were injected intraperitoneally with spermidine to detect antioxidant enzyme activity in ovaries and granulosa cells.What’s more,the granulosa cell apoptosis,and expression of apoptosis-related proteins in mouse granulosa cells were also measured to study the molecular mechanism of apoptosis and antioxidant function induced by exogenous spermidine.The main findings are as follows:1.Different concentrations of spermidine were intraperitoneally injected into adult female rats.It was found that high concentration of spermidine caused an increase in primordial follicles in mice,thinning of ovarian granule cells,and shrinkage of oocytes in some follicles;the activity of catalase in ovarian tissues was significantly decreased（p<0.01）,and the content of malondialdehyde was significantly increased（p<0.05）.The expression of CAT,NQO1,Cu/Zn-SOD and HO-1 m RNA in ovarian tissues of 0.15 mg/g spermidine treatment group were significantly higher than the control group（p<0.05）;2.After treatment with 0.15 mg/g spermidine for 24 h,the activities of CAT,SOD and total antioxidant capacity in mouse ovarian tissues were significantly decreased（p<0.05）.H₂O₂ and MDA levels were significantly increased（p<0.05）,and spermine content in the ovary was significantly increased（p<0.05）;3.After treatment with 0.15 mg/g spermidine for 24 h,the apoptosis rate of mouse granulosa cells increased significantly（p<0.05）,ROS levels in granulosa cells increased（p<0.05）,and spermine content decreased significantly（p<0.05）;4.After treatment with 0.15 mg/g spermidine for 24 h,the expression of apoptosis-related proteins in mouse granulosa cells as p53,Bax and Cytochrome c protein was significantly increased（p<0.05）,and Caspase 8,Caspase 9 expression was increased as well.And there was also an extremely significant increase in activated Caspase 3（p<0.05）.While The expression levels of anti-apoptosis-related protein Bcl-2were significantly decreased in granulosa cells（p<0.05）,and the ratio of Bax/Bcl-2 in granulosa cells of spermidine treatment group was significantly higher than that of the control cells（p<0.05）.