Di(2-ethylhexyl) Phthalate Alters Primordial Germ Cell Distribution and the Reproductive Neuroendocrine Regulatory Axis in Zebrafish Embryos

Di(2-ethylhexyl) phthalate (DEHP) is known to adversely affect reproduction. Our previous study demonstrated that DEHP exposure during embryogenesis impaired fertility in adult female zebrafish. The objective of this study was to investigate developmental events underlying this effect. Embryos were exposed to DEHP 5 h post-fertilization (hpf), and the distribution of primordial germ cells (PGCs), along with the expression of genes involved in PGC migration, maintenance, and neuroendocrine regulation, was assessed. Molecular docking simulations were performed to evaluate whether DEHP's main metabolite, mono(2-ethylhexyl) phthalate (MEHP), is able to bind to zebrafish estrogen receptors (Esr). Our results show that DEHP reduced the expression of <i>cxcr4b</i>, <i>cxcr7b</i>, <i>esr1</i>, and <i>esr2a</i> at 24 hpf. Using <i>vasa:egfp</i> transgenic embryos, we found that DEHP altered the distribution of PGCs. In addition, DEHP inhibited the expression of PGC-specific <i>dazl</i>. DEHP also induced the expression of <i>lhb</i> and <i>cyp19a1</i> and reduced the expression of <i>esr2a</i> in 120 hpf larvae, consistent with disruption of the neuroendocrine reproductive axis. Molecular docking indicates that MEHP can bind to the ligand-binding domains of Esr1, Esr2a, and Esr2b. Collectively, the results show that DEHP disrupts both PGC distribution and early neuroendocrine signaling pathways, providing mechanistic insight into reduced fertility in adult female zebrafish following embryonic DEHP exposure.