Dual activation of PPARα and PPARγ by mono-(2-ethylhexyl) phthalate in rat ovarian granulosa cells

Abstract

Peroxisome proliferator-activated receptors (PPARs) are key regulators of lipid metabolism and cell differentiation. The plasticizer di-(2-ethylhexyl) phthalate is a peroxisome proliferator, and its active metabolite mono-(2-ethylhexyl) phthalate (MEHP) activates PPARα and PPARγ in cell transactivation assays. MEHP is a female reproductive toxicant and decreases activity, mRNA, and protein levels of aromatase, the rate-limiting enzyme that converts testosterone to estradiol in ovarian granulosa cells. To test the hypothesis that MEHP suppresses aromatase through PPAR pathways, granulosa cells were cultured with MEHP (50 μM) or selective activators of PPARγ or PPARα for 48 h and gene expression was analyzed by real time RT-PCR. Both PPARα and PPARγ activators significantly decreased aromatase mRNA and estradiol production like MEHP. The PPARγ-selective antagonist GR 259662 partially blocked the suppression of aromatase by MEHP, suggesting that MEHP acts through PPARγ, but not exclusively. MEHP and the PPARα-selective agonist GW 327647 induced expression of 17β-hydroxysteroid dehydrogenase IV, a known PPARα-regulated gene, and induction was maintained with addition of the PPARγ-selective antagonist. PPARα-selective activation also induced expression of aryl hydrocarbon receptor (AhR), CYP1B1, and epoxide hydrolase in the granulosa cell. These data support a model in which MEHP activates both PPARα and PPARγ to suppress aromatase and alter other genes related to metabolism and differentiation in the granulosa cell.