Ovarian Follicles Rescued 3 Days after Cyclophosphamide Treatment in Adolescent Mice: An Experimental Study Aiming at Maximizing Methods for Fertility Preservation through In Vitro Follicle Culture

• There is currently a lack of knowledge about the feasibility of performing procedures for fertility preservation after chemotherapy treatment has been initiated. In this experimental controlled study using adolescent mice, we aimed to investigate if the chance of rescuing and growing in vitro secondary follicles (SeF) would be affected three days after a single injection of cyclophosphamide (CPA). The main outcomes included were: (1) The number of SeF with good morphologic quality obtained per ovary 3 days after CPA injection, (2) SeF development in culture, (3) small follicle density (SFD) on histology, and (4) apoptosis markers, including terminal deoxynucleotidyl transferase dUTP nick end-labelling (TUNEL), mRNA expression, and distribution of p 53 upregulated modulator of apoptosis (Puma) and phosphatase and tensin homolog (Pten). We found a 60% reduction of SeF obtained per ovary in all CPA-treated groups vs. controls. However, in vitro survival rates at culture day 12 and antrum formation were similar among all groups. On histology, SFD was only significantly reduced in the high CPA dose group. Apoptotic cells were mainly found in large growing follicles of CPA groups. Our study indicates the feasibility of SeF isolation and in vitro follicle culture 3 days following CPA treatment and a still preserved SFD, particularly following a low-dose CPA treatment.

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