Rapamycin preserves the primordial follicle pool during cisplatin treatment in vitro and in vivo

Abstract

Rapamycin has been proven to effectively inhibit the activation of primordial follicles while cisplatin-induced the loss of primordial follicles due to the overactivation of the primordial follicle stockpile. Whether rapamycin could inhibit the loss of primordial follicles induced by cisplatin is still unknown. The ovaries of neonatal Sprague Dawley rats were cultured in vitro in different doses of rapamycin (0.08, 0.16, and 0.32 μg/ml) and cisplatin (0.1, 0.4, and 0.8 μg/ml). The immature BALB/c mice were administered cisplatin with or without rapamycin by intraperitoneal injection. Ovaries were collected to analyze the histomorphology, the messenger RNA (mRNA) expression of anti-Mullerian hormone (AMH), growth differentiation factor 9 (GDF9), and bone morphogenetic protein 15 (BMP15) and the expression of key proteins of mammalian target of rapamycin (mTOR) pathway.Growing follicle counts of ovaries cultured in vitro in the R0.16 and R0.32 groups were decreased and the ratio of growing to primordial follicles was also decreased in a dose-dependent manner. In the C0.8 group, growing follicles were decreased compared with the other groups while the ratio was substantially increased in the C0.4 and C0.8 group. Co-treatment attenuated primordial follicle loss and reduced the upregulated ratio induced by cisplatin. Ovarian follicle dynamics in vivo was consistent with the in vitro results. Primordial follicles counts were statistically increased and the ratio was reduced in the rapamycin group compared with the control group. Primordial follicle counts were dramatically reduced in the cisplatin group whereas co-treatment with rapamycin slightly recovered its counts. There was no obvious difference in the number of growing follicles between the cisplatin group and other groups. The ratio was significantly increased in cisplatin-treated mice whereas decreased in the co-treatment group. The apoptosis rate of antral follicles in cisplatin-treated mice was higher than the other groups while the apoptosis rate was decreased in the co-treatment group in vivo. Compared with the control and rapamycin group, the mRNA expression of AMH, GDF9, and BMP15 were Abbreviations: AMH, anti-Mullerian hormone; BMP15, bone morphogenetic protein 15; CCK-8, cell counting kit-8; con, control group; FBS, fetal bovine serum; GDF9, growth differentiation factor 9; HE, hematoxylin and eosin; mTOR, mammalian target of rapamycin; mTORC1, mammalian target of rapamycin complex 1; PBS, phosphate-buffered solution; PFA, paraformaldehyde; POI, premature ovarian insufficiency; PTEN, phosphate and tension homology deleted on chromosome ten; qPCR, quantitative polymerase chain reaction; TSC1/TSC2, tuberous sclerosis complex; TUNEL, terminal deoxynucleotidyl transferase-mediated nick end labeling assay. downregulated in the cisplatin group. The co-treatment group recovered the mRNA expression of BMP15. In addition, the expression of key protein of mTOR pathway rpS6 and its phosphorylated forms were increased in the cisplatin-treated group while co-treatme...

Pharmacological inhibition of the PI3K/PTEN/Akt and mTOR signalling pathways limits follicle activation induced by ovarian cryopreservation and in vitro culture

“…Reduced p-Akt levels in follicles were also observed by immunohistochemistry in a recent in vitro study of ovine ovarian tissue cultured for 7 days in combination with epigallocatechin-3-gallate (EGCG), which has strong antioxidant and anti-apoptotic properties and activates the PI3K/PTEN/Akt pathway [ 65 ]. Our results are in agreement with the recent study by Xie et al (2020), showing that rapamycin inhibited the activation of primordial follicles of 4‐day‐old rat ovaries in vitro [ 45 ].…”

“…Tsc2 can also be phosphorylated and inactivated by Akt, leading to activation of mTORC1 and phosphorylation of ribosomal protein S6 (rps6) [ 44 ]. Rapamycin, a specific mTOR inhibitor, is used as an immunosuppressant after organ transplantation and as a drug to prevent revascularization in coronary stents [ 45 ]. Inhibition of the activation of primordial follicles of 4‐day‐old rat ovaries in vitro was recently demonstrated using rapamycin [ 45 ].…”

“…Rapamycin, a specific mTOR inhibitor, is used as an immunosuppressant after organ transplantation and as a drug to prevent revascularization in coronary stents [ 45 ]. Inhibition of the activation of primordial follicles of 4‐day‐old rat ovaries in vitro was recently demonstrated using rapamycin [ 45 ]. Rapamycin has also been shown to be useful in mice to preserve the ovarian follicle pool and prevent premature ovarian failure [ 46 ].…”

Female Oncofertility: Current Understandings, Therapeutic Approaches, Controversies, and Future Perspectives

“…The effect of antitumoral drugs on ovarian function is the follicle-specific magnitude and is associated with the category of the drugs [ 12 ]. Some studies have declared that apoptosis occurred only in GCs of growing follicles, but not in PFs by TUNEL staining after treatment of cyclophosphamide or cisplatin [ 54 , 55 ]. Other results insist that TUNEL and/or γH2AX staining are positive in oocytes but not in the GCs of PFs [ 46 , 50 , 56 ].…”

DNA Damaged Induced Cell Death in Oocytes

“…A large number of studies have already suggested treatment with a wide variety of different molecules to protect oocytes from chemotherapy-induced apoptosis. These studies range from hormonal treatment with gonadotropin-releasing hormone analogues or luteinizing hormone [152] to the use of sphingosine1-phosphate, dexrazoxane, resveratrol or mTORC inhibitors [153]. Some of these treatments have shown protective effects in mouse models while others, also used in human trials, are controversially discussed.…”