Exposure to DEHP or its metabolite MEHP promotes progesterone secretion and inhibits proliferation in mouse placenta or JEG-3 cells☆
Graphical abstract
Introduction
DEHP is a commonly used plasticizer binding in polyvinyl chloride (PVC) products with non-covalent bond (Nardelli et al., 2017, Rowdhwal and Chen, 2018). DEHP is easy to transfer to the environment from PVC product and has bioaccumulation effect (Al-Saleh et al., 2017).It can be found in milk and placenta, even can cross the placental barrier (Andrade et al., 2006, Zhu et al., 2006). DEHP converts to its active monoester metabolite mono (2-ethylhexyl) phthalate MEHP in 24 h in vivo (Albro et al., 1982). The toxic effects of DEHP have been studied extensively, including carcinogenesis, higher susceptibility of diseases, neurotoxicity, reproductive toxicity and endocrine disrupt effects (Chen et al., 2018, Wang et al., 2018, Wang et al., 2016, Xia et al., 2018, Yin et al., 2018). Some of them even have a trans-generational inheritance effect (Pocar et al., 2012, Pocar et al., 2017).
The effect of DEHP maternal exposure on offspring size at birth is controversial in population-based study (Zhang et al., 2009, Zhang et al., 2018, Zhu et al., 2018). In animal experiments, most of the researches apt to DEHP prenatal exposure inducing a reduction fetal size (Chen et al., 2010, Yu et al., 2018).Our previous study confirmed that maternal exposure to DEHP during pregnancy caused a fetal growth restriction (FGR) in a gender-independent manner (Shen et al., 2017). It hints that DEHP may affect the growth of fetus through reducing nutrition or material supply from placenta. Utero exposure to DEHP destroys the structure and reduces the size of mouse placenta (Zong et al., 2015). Therefore, placenta is a target of DEHP.
Placenta plays essential roles during gestation. Fetal nutrition, support and protection, gas exchange are dependent on this organ. Besides, placenta produces several hormones and other mediators (Costa, 2016). Progesterone is required for the maintenance of pregnancy and is one of the hormones produced by syncytiotrophoblast mitochondria of placenta in human from maternal cholesterol during pregnant (Miller, 1998, Tuckey, 2005). Cholesterol is converted to pregnenolone by CYP11A1 (cholesterol side-chain cleavage cytochrome P450, CYP450scc) within the inner mitochondrial membrane. It is the rate-determining step of placental progesterone synthesis. Progesterone is converted from pregnenolone by type 1 3 beta-hydroxysteroid dehydrogenase (3β-HSD1) in placenta (Morel et al., 1997). Pregnenolone or progesterone cannot be converted to estrogens in placenta because of lacking cytochrome P450 17α (CYP17).Instead, estrogens are synthesized by androgen substrates from the fetal and maternal adrenal (Strauss et al., 1996, Tuckey, 2005). Therefore, estrogen synthesize is more complex than progesterone.
Progesterone exerts genomic and non-genomic functions through activating different receptors. Two classical progesterone receptors (Iacovelli et al., 2015), PRA and PRB are expressed from the same gene PGR. After binding progesterone, PR recognizes special DNA promoter sequence of target genes and mediates genomic action (Smith, 2000). CCND1 (Cyclin D1 gene) is a target gene of PRA (Giulianelli et al., 2012, Quiles et al., 2009). It is a cell cycle check point gene of G1 phase which is a pivotal factor to cell proliferation (Baldin et al., 1993). Membrane–associated progesterone receptors (MPR) and MAPK mediate the non-genomic actions decrease Cyclic Adenosine monophosphate (cAMP) and intracellular Ca2+ mobilization (Goldman and Shalev, 2007, Gronemeyer et al., 1991). PRA negatively influences PRB (Scarpin et al., 2009). PRs can interact with estrogen receptor α (ERα) and modulate ERα action in breast cancer. Simultaneously, PRs expression is used as a biomarker of the function of ERα because PGR (gene of PR) is classical target gene of ERα (Mohammed et al., 2015).
As an endocrine disruptor, DEHP has the anti-androgenic or estrogenic effects (Christiansen et al., 2010, Romani et al., 2014), and inhibits the transactivation of estrogen receptor (ER) induced by estrogen (Ghisari and Bonefeld-Jorgensen, 2009). However, whether DEHP affects the endocrine function of placenta during pregnancy needs further exploration. In this article, we explored the effect of DEHP on estradiol and progesterone secretion in vivo. And the results were verified in vitro using human placenta trophoblast JEG-3 cell exposed to MEHP for 7 days. According to these tests, we further speculated the underlying mechanism about the effect of DEHP on endocrine function of placenta.
Section snippets
Cell culture
Human placenta trophoblast JEG-3 cells from American Type Culture Collection were growth in AMEM (Gibco, 1843032) medium with 10% fetal bovine serum (Gibco) and 100 Units/ml penicillin-Streptomycin (Sangon, Shanghai, China) plus 100 g/ml L-Glutamine (Gibco) at 37 °C with 5% CO2.
MEHP, progesterone or estradiol treatment
MEHP (Aldrich, MKCD4270), progesterone (TargetMol, T0478) or estradiol (TargetMol, T1048) were dissolved in dimethyl sulfoxide (DMSO, Sigma, RNBG2257). The final concentration of DMSO was 0.1% in all exposure medium, and
Effect of DEHP on progesterone secretion
Our previous studies showed that DEHP maternal exposure during pregnancy in ICR mouse induced the FGR and inhibited the placenta proliferation in a gender independent fashion (Shen et al., 2017). These conclusions hinted that DEHP might induce FGR through disturbing the function of placenta. Given that DEHP is an environmental endocrine disruptor and the placenta is a source of progesterone and E2 during pregnancy (Miller, 1998, Tuckey, 2005), we predicted that DEHP may disturb placenta
Discussion
DEHP is an estrogen mimic endocrine disruptor and induces FGR through inhibiting the proliferation of placental cells and disturbing the function of placenta in mouse who exposed to DEHP during maternal pregnancy (Shen et al., 2017). However, whether DEHP inhibit placental cells’ proliferation via affecting the endocrine function of placenta is still to be explored. In this study we found that DEHP decreased PR level, which inhibited cell proliferation through decreasing the level of Cyclin D1
Declaration of competing interest
No declared.
Acknowledgements
We thank Dr. Xingwang Cao, Dr. Dexiang Xu, Dr. Qu’nan Wang and Dr. Yuanhua Chen (Anhui Medical University) for helpful advice. We thank the Laboratory of Cell Dynamics, University of Science and Technology of China for its generous donation of antibodies. This work was supported by the National Natural Science Foundation of China [31000602], Natural Science Foundation of Anhui Province [1608085MH218, 1308085QC48], Key Projects of outstanding Youth Talent Support Program in Anhui Provincial
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This paper has been recommended for acceptance by Wen Chen.
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These authors contributed equally to this work.