The Anti-aging Effect And Mechanism Investigation Of Metformin On The Ovarian Aging

Aging has been regarded as the main factor to several kinds of chronic diseases.And ovary is proved to be the first organ to age in females,which makes it greatly significant to delay its aging.Among the different kinds of methods to delay ovarian aging,caloric restriction(CR)has been proved to be an effective one,along with some caloric restriction mimics(CRM)such as resveratrol.Metformin(MET),the first-line drug for diabetes mellitus,has a long history of use for its convenience,accessibility and safety.Some experiments have indicated that MET has the ability to prolong the health span of mice and owns the same kind of gene profiles compared with CR.Therefore,we hypothesize that MET might be able to delay the ovarian aging.This experiment is aimed to investigate the anti-aging effect and possible mechanisms of metformin on the ovarian aging.We divided the 28-week old female C57BL/6 mice into two groups,namely control group(CON group)and metformin-treated group(MET group).The mice in the CON group were fed on normal diets and those in the MET group were fed on the similar diets with the supply of metformin(100g/kg,0.1%w/w).The physical activities and healthy conditions were examined by the behavior tests and anatomy of the mice.The ovarian functions in both groups were compared by testing the estrous cycles,the litter experiment,levels of sex hormones and the follicle counts.The expression levels of oxidized metabolites,such as 8-hydroxy-2′-deoxyguanosine(8-OHd G),4-hydroxynonenal(4-HNE)and nitrotyrosine(NTY)and age-related protein P16 were tested by immunehistochemical method,along with some crucial proteins in the ovary,such as Sirt1,p-rp S6,Bcl2 examined by western blot to explore the possible mechanisms.The biopsy and physical activity tests showed that metformin under the standard concentration had no toxic effects on the mice after 6-months treatment.The estrous cycle tested on the third month of feeding showed no significant difference between the two groups.However,the MET group exhibited much more regular estrous cycles after 6-months’ feeding.The MET group also owned larger amounts of primordial and primary follicles indicated by the follicle count.Furthermore,the level of the E2 hormone is much higher in the MET group,although the level of the FSH hormone showed no significant difference between the two groups.The Western Blot experiments exhibited that expression of the protein Sirt1 was higher,with the p-rp S6 protein lower in the MET group.No significant difference was showed in the expression of the Bcl2 protein.The expression levels of the aging related protein P16,the oxidized metabolites 8-OHd G and 4-HNE were much higher in the CON group,although the level of NTY showed no significant difference.We concluded that MET has a positively anti-aging effect on the ovarian aging,probably by stimulating the expression of Sirt1 protein,thus decreasing the oxidative stress of the aging ovary.