Early prepubertal cyclophosphamide exposure in mice results in long-term loss of ovarian reserve, and impaired embryonic development and blastocyst quality.

Sujith Raj Salian,Satish Kumar Adiga,Richard A Anderson,Aswathi Cheredath,Guruprasad Kalthur,Fiona D’Souza,Shubhashree Uppangala,Vinod C Nayak

doi:10.1371/journal.pone.0235140

Abstract

BackgroundDue to improved treatment, there is an increasing focus on the reproductive potential of survivors of childhood cancer. Cytotoxic chemotherapy accelerates the decline in the number of primordial follicles within the mammalian ovary at all ages, but effects on the developmental potential of remaining oocytes following prepubertal cancer treatment are unclear.ObjectivesTo investigate whether cyclophosphamide (CY) exposure in the prepubertal period in female mice influences ovarian function and the functional competence of oocytes in adulthood.MethodsThis study used Swiss albino mice as the experimental model. Female mice were treated with 200 mg/kg CY on either postnatal day 14 (CY14), 21 (CY21) or 28 (CY28) i.e at a prepubertal and 2 young postpubertal ages. At 14 weeks of life, ovarian function, functional competence of oocytes, and embryo quality were assessed.ResultsThe number of primordial follicles decreased significantly in CY14 and CY21 groups compared to control (p < 0.01). The number of oocytes from superovulated was 8.5 ± 1.4, 24.1 ± 2.9 and 26.8 ± 2.1 in CY14, CY21 and CY28 respectively which was significantly lower than control (50.2 ± 3.2; p < 0.001). In vitro culture of CY14 embryos demonstrated only 55.4% blastocyst formation (p < 0.0001) and reduced ability of inner cell mass (ICM) to proliferate in vitro (p < 0.05) at 120 and 216 h post insemination respectively. On the other hand, ICM proliferation was unaltered in 2 young postpubertal ages.ConclusionOur results indicate long-term effects on the developmental competence of oocytes exposed to CY in early but not adult life. These data provide a mechanism whereby long-term fertility can be impaired after chemotherapy exposure, despite the continuing presence of follicles within the ovary, and support the need for fertility preservation in prepubertal girls before alkylating agent exposure.

Highlights

Childhood cancer survival has improved greatly over the past five decades, and due to improved treatment modalities, about 70–80% of children with cancer will be cured
In vitro culture of CY on either postnatal day 14 (CY14) embryos demonstrated only 55.4% blastocyst formation (p < 0.0001) and reduced ability of inner cell mass (ICM) to proliferate
Our results indicate long-term effects on the developmental competence of oocytes exposed to CY in early but not adult life. These data provide a mechanism whereby longterm fertility can be impaired after chemotherapy exposure, despite the continuing presence of follicles within the ovary, and support the need for fertility preservation in prepubertal girls before alkylating agent exposure

Summary

Introduction

Childhood cancer survival has improved greatly over the past five decades, and due to improved treatment modalities, about 70–80% of children with cancer will be cured. Many childhood cancer survivors are entering their reproductive age [1]. Certain types of cancer treatments can compromise ovarian function in children, notably high doses of alkylating agents and radiotherapy [2,3,4]. Oocytes within primordial follicles exist in a relatively quiescent state, and may be relatively resistant to antimitotic agents [8], but the effects of alkylating agents are independent of the cell cycle. There is an increasing focus on the reproductive potential of survivors of childhood cancer. Cytotoxic chemotherapy accelerates the decline in the number of primordial follicles within the mammalian ovary at all ages, but effects on the developmental potential of remaining oocytes following prepubertal cancer treatment are unclear

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Conclusion