Abstract
Di(2-ethylhexyl)phthalate (DEHP) interferes with sex hormones signaling pathways (SHP). C57BL/6J mice prenatally exposed to 300 mg/kg/day DEHP develop a testicular dysgenesis syndrome (TDS) at adulthood, but similarly-exposed FVB/N mice are not affected. Here we aim to understand the reasons behind this drastic difference that should depend on the genome of the strain. In both backgrounds, pregnant female mice received per os either DEHP or corn oil vehicle and the male filiations were examined. Computer-assisted sperm analysis showed a DEHP-induced decreased sperm count and velocities in C57BL/6J. Sperm RNA sequencing experiments resulted in the identification of the 62 most differentially expressed RNAs. These RNAs, mainly regulated by hormones, produced strain-specific transcriptional responses to prenatal exposure to DEHP; a pool of RNAs was increased in FVB, another pool of RNAs was decreased in C57BL/6J. In FVB/N, analysis of non-synonymous single nucleotide polymorphisms (SNP) impacting SHP identified rs387782768 and rs29315913 respectively associated with absence of the Forkhead Box A3 (Foxa3) RNA and increased expression of estrogen receptor 1 variant 4 (NM_001302533) RNA. Analysis of the role of SNPs modifying SHP binding sites in function of strain-specific responses to DEHP revealed a DEHP-resistance allele in FVB/N containing an additional FOXA1-3 binding site at rs30973633 and four DEHP-induced beta-defensins (Defb42, Defb30, Defb47 and Defb48). A DEHP-susceptibility allele in C57BL/6J contained five SNPs (rs28279710, rs32977910, rs46648903, rs46677594 and rs48287999) affecting SHP and six genes (Svs2, Svs3b, Svs4, Svs3a, Svs6 and Svs5) epigenetically silenced by DEHP. Finally, targeted experiments confirmed increased methylation in the Svs3ab promoter with decreased SEMG2 persisting across generations, providing a molecular explanation for the transgenerational sperm velocity decrease found in C57BL/6J after DEHP exposure. We conclude that the existence of SNP-dependent mechanisms in FVB/N inbred mice may confer resistance to transgenerational endocrine disruption.
Highlights
Di-(2-ethylhexyl) phthalate (DEHP; CAS No 117-81-7) is a reproductive toxicant and an endocrine disruptor (ED) ubiquitously found in the environment
DEHP and its principal metabolite named mono-(2-ethylhexyl) phthalate (MEHP; CAS No 4376-20-9) decrease the testosterone produced by testes and interact at the molecular level with the androgen (AR), estrogen (ER) and peroxisome proliferator-activated receptors (PPARs) [1, 2]
The phenotypic changes affecting male fertility parameters induced by prenatal exposure to DEHP confirm the FVB/N resistance found in our earlier study using an independent method [11] and using data partially previously obtained in C57BL/6J [13]
Summary
Di-(2-ethylhexyl) phthalate (DEHP; CAS No 117-81-7) is a reproductive toxicant and an endocrine disruptor (ED) ubiquitously found in the environment. Accumulated data demonstrate that DEHP interferes with sex steroid hormone signaling pathways (SHP). DEHP and its principal metabolite named mono-(2-ethylhexyl) phthalate (MEHP; CAS No 4376-20-9) decrease the testosterone produced by testes and interact at the molecular level with the androgen (AR), estrogen (ER) and peroxisome proliferator-activated receptors (PPARs) [1, 2]. Prenatal exposure to DEHP causes androgen deficiency during embryogenesis in both animals and humans [3, 4]. The anogenital distance (AGD), a marker of fetal androgen exposure [5], was shortened in boys born from DEHP-exposed mothers and was reduced in rodents prenatally exposed to DEHP [6,7,8]. The long-term toxicological impacts of prenatal exposure to DEHP are of high concern
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