The chemotherapy agent etoposide is a topoisomerase II (topo II) inhibitor, and is considered safe to administer during pregnancy. However, assessment of its effects on the developing ovary, when germ cells are undergoing initiation of meiosis and forming follicles, has been limited. We have investigated this using ovarian tissue culture.
E13.5 mouse ovaries were cultured for 12 days on an agar block, with etoposide added for the first 6 days of culture at concentrations of up to 150 ng/ml, thus exposing the germ cells for the period prior to follicle formation. Follicle numbers and health were analysed histologically. Immunohistochemistry was used to determine topo IIα localisation in mouse and human fetal ovary, and to examine the progression of cultured oocytes through prophase I of meiosis, by visualisation of Sycp3.
Results and Discussion
Etoposide did not block the ability of oocytes to progress through meiosis to the diplotene stage of prophase I (after which oocytes enter meiotic arrest), with around 80% of oocytes having reached that phase of meiosis after six days of culture in both etoposide-treated and control groups. There was however evidence of a more rapid progression through early meiosis: more germ cells from the etoposide-treated ovaries had progressed from leptotene/zygotene to pachytene stage after 2 days culture compared with controls (47% vs 77%, p<0.001). A dose-dependent reduction of follicle numbers was observed following treatment with etoposide, with a near-complete loss of healthy follicles at the top dose (89.7% loss, p<0.001). Topo IIα expression was confined to the germ cells prior to follicle formation in both human and mouse fetal ovaries. Our results indicate that germ cells can progress through prophase I to diplotene during exposure to etoposide, but their ability to form follicles is markedly impaired.