School of Animal and Comparative Biomedical Sciences, University of Arizona, Tucson, Arizona, USA.
Biol Reprod. 2017 May 1;96(5):1105-1117. doi: 10.1095/biolreprod.116.144691.
Dibutyl phthalate (DBP) is present in consumer products and the coating of some oral medications. Acetyl tributyl citrate (ATBC) has been proposed as an alternative to DBP because DBP causes endocrine disruption in animal models. Following ingestion, DBP is converted to its main metabolite mono-butyl phthalate (MBP) which has been detected in >90% of human follicular fluid samples. Previous studies show that DBP reduces the number of antral follicles present in the ovaries of mice. Thus, this study was designed to evaluate the effects of DBP, MBP, and ATBC on in vitro growth and viability of mouse ovarian antral follicles. Antral follicles were isolated from CD-1 females (PND32-37) and treated with vehicle, DBP, MBP, or ATBC (starting at 0.001 and up to 1000 μg/ml for DBP; 24-72 h). Follicle diameter, ATP production, qPCR, and TUNEL were used to measure follicle growth, viability, cell cycle and apoptosis gene expression, and cell death-associated DNA fragmentation, respectively. While MBP did not cause toxicity, DBP exposure at ≥10 μg/ml resulted in growth inhibition followed by cytoxicity at ≥500 μg/ml. ATBC increased the number of nongrowing follicles at 0.01 μg/ml and did not affect ATP production, but increased TUNEL positive area in treated follicles. Gene expression results suggest that cytotoxicity in DBP-treated follicles occurs via activation of cell cycle arrest prior to follicular death. These findings suggest that concentrations of DBP ≥10 μg/ml are detrimental to antral follicles and that ATBC should be examined further as it may disrupt antral follicle function at low concentrations.
邻苯二甲酸二丁酯（DBP）存在于消费品和某些口服药物的涂层中。乙酰基三丁基柠檬酸酯（ATBC）已被提议作为 DBP 的替代品，因为 DBP 在动物模型中会引起内分泌干扰。摄入后，DBP 会转化为其主要代谢物单丁基邻苯二甲酸酯（MBP），在>90%的人类卵泡液样本中都检测到了 MBP。先前的研究表明，DBP 会减少小鼠卵巢中存在的窦卵泡数量。因此，本研究旨在评估 DBP、MBP 和 ATBC 对体外培养的小鼠卵巢窦卵泡生长和活力的影响。窦卵泡从 CD-1 雌性（PND32-37）中分离出来，并与载体、DBP、MBP 或 ATBC（起始浓度为 0.001 至 1000μg/ml 的 DBP；24-72 h）一起处理。使用卵泡直径、ATP 产生、qPCR 和 TUNEL 分别测量卵泡生长、活力、细胞周期和凋亡基因表达以及与细胞死亡相关的 DNA 片段化。虽然 MBP 没有引起毒性，但 DBP 暴露在≥10μg/ml 时会导致生长抑制，随后在≥500μg/ml 时会导致细胞毒性。ATBC 在 0.01μg/ml 时会增加非生长卵泡的数量，并且不会影响 ATP 产生，但会增加处理卵泡中的 TUNEL 阳性区域。基因表达结果表明，DBP 处理的卵泡中的细胞毒性是通过在卵泡死亡前激活细胞周期阻滞引起的。这些发现表明，浓度≥10μg/ml 的 DBP 对窦卵泡有害，并且应该进一步研究 ATBC，因为它可能会在低浓度下破坏窦卵泡功能。
