DEHP exposure effects on human ovarian samples
Di(2-ethylhexyl) phthalate (DEHP) is a plasticizer known for its endocrine-disrupting properties mediated by its many metabolites that form upon exposure in biological systems. In a previous study, we reported an inverse association between DEHP metabolites in the human ovarian follicular fluid (FF) and the responsiveness of the follicles to controlled ovarian stimulation during IVF treatments. Here, we explored this association further through molecular analysis of the ovarian follicular fluid samples.  Ninety-six IVF patients from Swedish (N=48) and Estonian (N=48) infertility clinics were selected from the previous cohort (N=333) based on the molar sum of DEHP metabolites in their FF samples to study differences between “high” (mean 7.7 ± SD 2.3 nM, N=48) and “low” (0.8 ± 0.4 nM, N=48) exposure. Extracellular miRNA levels and concentrations of 15 steroid hormones were measured in the samples. In addition, the FF somatic cells, available in the Estonian sub cohort, were used for RNA sequencing.  Differential expression (DE) and miRNA:mRNA network analysis revealed that the expression levels of genes in the cholesterol biosynthesis and steroidogenesis pathways were significantly decreased in the high compared to the low DEHP group. In addition, the DE miRNAs were predicted to target the key enzymes within these pathways (FDR<0.05). A decreased progesterone to 17-OH-progesterone ratio was observed in the FF of women in the high DEHP group (p<0.05). Additionally, the expression levels of genes associated with inflammatory processes were elevated in the FF somatic cells, and a computational cell-type deconvolution analysis suggested an increased immune cell infiltration into the high DEHP follicles (p<0.05).  In conclusion, high DEHP levels were associated with a significantly altered follicular milieu within human ovaries, involving pro-inflammatory environment and reduced lipid and steroid synthesis. This study enhances our understanding of the molecular mechanisms through which DEHP may disrupt ovarian sensitivity to pituitary hormones in humans.