In utero exposure to some phthalate esters adversely affects the development of the rat seminiferous cord, causing germ cell loss and increasing the number of multinucleated germ cells (MNGs). To understand the timing of MNG formation and determine whether it requires nuclear division, timed pregnant Sprague Dawley rats were exposed to 500 mg/kg di-n-butyl phthalate (DBP) or corn oil vehicle by oral gavage on Gestational Day (GD) 17 or 18 (0 h) and euthanized after 2, 4, 6, or 24 h or given a second dose at 24 h and euthanized 48 h after the initial dose. Dams were simultaneously exposed to 0.3 M 5-bromo-2′-deoxycitidine (BrdC; converted to 5-bromo-2′-deoxyuridylate [BrdU] in vivo) through a subcutaneous micro-osmotic pump implanted at −2 h. In the testes of male fetuses, DBP induced MNGs significantly beginning at 4–6 h and dramatically by 24 h when exposure began on GD 18 but not GD 17. Seminiferous cord diameter was significantly elevated in testes of rats treated with DBP at 24 and 48 h, and cell death, measured by TUNEL assay, was significantly elevated by DBP only at 48 h, when treatment began on GD 18. TUNEL-labeled MNGs were rare. Overall BrdU labeling rate in the testis was unaffected by DBP. Only one of 606 MNGs in BrdU-labeled sections had a strongly positive nucleus, confirming a nonproliferative mechanism of MNG formation, which is a degenerative process with the potential to adversely affect testis development.
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Vol. 93 • No. 5