Role Of Exosome Derived From Human Umbilical Mesenchymal Stem Cell In Activating Primordial Follicles

The reproductive lifespan of female mammals is determined by the primordial follicle pool.Ovarian function decreases with aging and the loss of primordial follicles.However,there are still some dormant primordial follicles in women who are in perimenopause,and it can be activated by some stimulators to get eggs and have babies.Mesenchymal stem cell(MSC)in treatment of primary ovarian insufficiency has made great progress in recent years.Exosomes are extracellular vesicles have multiple proteins and nucleic acids inside that sized in 50-150 nm and can participate in various of bioprocess as the mediator of their progenitor cells.We established a murine model to study the roles of human umbilical cord mesenchymal stem cell(HUCMSC)derived exosome in activating primordial follicles.First,we isolated primary umbilical cord mesenchymal stem cells from newborn infant.Then we verified the cell with flow cytometry and induced multidifferentiation.For collecting cell culture media which contained exosomes,we prepared exosome-depleted fetal bovine serum and collected conditional culture media.Next,we isolated exosomes derived from human umbilical cord MSCs using filtration and ultracentrifugation.Western Blot,transmission electron microscope and Nanosight were used to identify the exosome.To evaluate the function of exosomes,we cultured 3 days old newborn mouse ovaries in vitro,and we found more primordial follicles were activated after treatment of exosomes than in control group with phosphorylation of Foxo3 a and the upregulation of mTOR signaling.And these roles can be blocked by using PI3K-AKT-mTOR signaling pathway inhibitors and heparin.Moreover,transplantation of cultured ovaries under the kidney capsule of hosts for 14 days were used to determine the further influence of follicles.We observed better follicle development and got more oocytes in exosome treatment group and ensured the quality of eggs didn't get damaged by using exosome culture strategy.Furthermore,we used a natural-aged mouse model of intra-bursal injection of exosome.Compared to control group,mice injected with exosome had the potential improvement in fertility and oocyte quality.In summary,we demonstrated successfully that human umbilical cord msenchymal stem cells derived exosomes could activate the primordial follicles in vitro and in vivo,which suggested a possible new method in treatment of primary ovary insufficiency in future.