Dose-dependent alterations in gene expression and testosterone production in fetal rat testis following exposure to di-n-hexyl phthalate

In utero exposure to the phthalate ester plasticizer, di-n-hexyl phthalate (DnHP) is known to affect the development of the male reproductive system and induce alterations in androgen-dependent tissues of the male rat offspring. Male reproductive malformations produced by several phthalates have been causally linked to decreased testosterone production during the gestational period. Thus, this study was designed to evaluate the dose-response relationship for the effects of DnHP on synthesis and production of testosterone in the fetal rat testis. Pregnant Sprague-Dawley rats were administered the vehicle (olive oil), DnHP (5 to 625 mg/kg/day), or diethylhexyl phthalate (DEHP) (50 or 625 mg/g/day), by gavage, from gestation day (GD) 12 to19. Fetal testes were assessed on GD 19. DnHP reduced ex vivo testosterone production and down-regulated the expression of several genes required for cholesterol transport and steroid synthesis (i.e. scavenger receptor B1, steroidogenic acute regulatory protein, P450 side chain cleavage, 3-hydroxysteroid dehydrogenase, and cytochrome P450c17). These inhibitions were dose-dependent. A no effect level was established at 5 mg/kg/day, and a lowest effect level at 20 mg/kg/day. ARNm were not similarly decreased in the adrenals. In conclusion, DnHP shares the same mode of action as DEHP of disrupting fetal testicular androgen synthesis. Alterations in testosterone production and in key steroidogenic gene expression appeared at lower doses than those causing postnatal reproductive malformations after gestational exposure during the critical period of male sexual differentiation. This suggests that they can be considered as early biomarkers of DnHP-induced fetal testicular effects in rats.