Abstract：
Phthalates are a class of known endocrine disrupting chemicals that are found in common everyday products. Several studies associate phthalate exposure with detrimental effects on ovarian function, including growth and development of the follicle and production of steroid hormones. We hypothesized that dysregulation of the ovary by phthalates may be mediated by phthalate toxicity towards granulosa cells, a major cell type in ovarian follicles responsible for key steps of hormone production and nourishing the developing oocyte. To test the hypothesis that phthalates target granulosa cells, we harvested granulosa cells from adult CD-1 mouse ovaries and cultured them for 96 hours in vehicle control, a phthalate mixture, or a phthalate metabolite mixture (0.1-100 μg/mL). After culture, we measured metabolism of the phthalate mixture into monoester metabolites by the granulosa cells, finding that granulosa cells do not significantly contribute to ovarian metabolism of phthalates. Immunohistochemistry of phthalate metabolizing enzymes in whole ovaries confirmed that these enzymes are not strongly expressed in granulosa cells of antral follicles and that ovarian metabolism of phthalates likely occurs primarily in the stroma. RNA sequencing of treated granulosa cells identified 407 differentially expressed genes, with overrepresentation of genes from lipid metabolic processes, cholesterol metabolism, and peroxisome proliferator-activated receptor (PPAR) signaling pathways. Expression of significantly differentially expressed genes related to these pathways was confirmed using qPCR. Our results agree with previous findings that phthalates and phthalate metabolites have different effects on the ovary, but both interfere with PPAR signaling in granulosa cells.
摘要：
邻苯二甲酸酯是一类已知的内分泌干扰化学物质，存在于常见的日常产品中。一些研究将邻苯二甲酸盐暴露与对卵巢功能的有害影响联系起来，包括卵泡的生长和发育以及类固醇激素的产生。我们假设邻苯二甲酸盐对卵巢的失调可能是由邻苯二甲酸盐对颗粒细胞的毒性介导的。卵泡中的一种主要细胞类型，负责激素产生和滋养发育中的卵母细胞的关键步骤。为了检验邻苯二甲酸盐靶向颗粒细胞的假设，我们从成年CD-1小鼠卵巢中收获颗粒细胞，并在载体对照中培养96小时，邻苯二甲酸酯混合物，或邻苯二甲酸酯代谢物混合物（0.1-100μg/mL）。文化之后，我们测量了颗粒细胞将邻苯二甲酸酯混合物代谢为单酯代谢产物，发现颗粒细胞不显著促进邻苯二甲酸盐的卵巢代谢。整个卵巢中邻苯二甲酸酯代谢酶的免疫组织化学证实，这些酶在窦卵泡的颗粒细胞中不强烈表达，并且邻苯二甲酸酯的卵巢代谢可能主要发生在基质中。经处理的颗粒细胞的RNA测序鉴定出407个差异表达基因，脂质代谢过程中基因的过度表达，胆固醇代谢，和过氧化物酶体增殖物激活受体（PPAR）信号通路。使用qPCR确认与这些途径相关的显著差异表达的基因的表达。我们的结果与先前的发现一致，邻苯二甲酸酯和邻苯二甲酸酯代谢物对卵巢有不同的影响，但两者都会干扰颗粒细胞中的PPAR信号传导。
