mTOR kinase inhibition results in oocyte loss characterized by empty follicles in human ovarian cortical strips cultured in vitro

Abstract

Objective: To determine whether oocyte loss is induced by mTOR kinase inhibition in human cortical strips as seen in model organisms in vivo and in vitro.

Design: Ovarian cortex was collected at two centers and cut into small strips. Strips were cultured for 6 days with or without the mTOR inhibitor rapamycin (RAP; 100 nM). Strips were then embedded in paraffin, and serial sections were prepared.

Setting: Samples were collected in general obstetric (Edinburgh), gynecologic surgery (New Haven), and fertility preservation assisted reproductive technology (ART) (New Haven) practices.

Patient(s): Ovarian cortex collected from patients (15-34 years of age) during cesarean section (donated tissue) was removed for the purposes of fertility preservation or was prepared after oophorectomy.

Intervention(s): Tissue was used for research purposes only, with no subsequent patient intervention.

Main outcome measure(s): Follicles were counted and assessed in each serial section. Caspase activity was monitored to determine whether mTOR inhibition activated apoptosis.

Result(s): The RAP inclusion in cultures results in significantly fewer follicles compared with ethanol vehicle-treated controls. Furthermore, RAP treatment resulted in the induction of follicles that lacked an oocyte in any serial section (30/161 follicles vs. 1/347 ethanol vehicle-treated follicles). Caspase activity was not elevated by RAP treatment.

Conclusion(s): mTOR inhibition results in a conserved destruction of the oocyte by adjacent granulosa cells (GC) in the absence of increased caspase activity. This model of oocyte loss is not consistent with classic apoptosis/atresia.