Additional steroidogenesis endpoints in the fetal rat

Testing and assessment of chemicals for their endocrine disruption properties are a major concern for chemical safety. Besides regulatory studies which use standard markers (e.g. anogenital distance), more targeted mode of action studies can provide essential information.
The development of the male reproductive tract is highly dependent on the production of testosterone by the fetal testis. Numerous studies in rats have shown that disruption of androgen biosynthesis by endocrine disrupting chemicals such as phthalates during the critical prenatal window of masculinization may lead to various reproductive abnormalities in the male offspring.
Ex vivo testicular testosterone production by the fetal rat testis has been proposed as a short term in vivo assay to screen anti-androgenic chemicals that produce adverse developmental effects via disruption of testosterone synthesis in utero. In an effort to make more out of this testing system, pregnant Sprague Dawley rats were administered di-n-hexyl phthalate (DnHP) or diisobutyl phthalate (DIBP) at the single oral dose of 250 mg/kg/day on gestational days 13 to 19. Beyond the single testosterone production measurement, the test design was expanded by the quantification of a few additional hormones of the steroid hormone synthetic pathway using a TFC-UPLC-HRMS method*, along with the determination of fetal testis mRNA expressions of key enzymes involved in various steps of steroid transport and synthesis.
The combination of these cellular and molecular endpoints provides strengthened evidence for the disrupting effects of DnHP and DIBP on testosterone synthesis in the rat fetus. Both chemicals produced alterations of several hormone levels and expressions of genes across the steroidogenesis pathway. Their profiles of effects were similar, with DIBP being slightly less potent than DnHP. These results confirm the interest of complementing mechanistic in vivo studies.
* TurboFlowTM chromatography-Ultra-high performance liquid chromatography coupled with a Q-Exactive Orbitrap mass spectrometer