Early-life exposure to the endocrine disruptor 17-α-ethinylestradiol induces delayed effects in adult brain, liver and ovotestis proteomes of a self-fertilizing fish

Abstract

Early-life represents a critically sensitive window to endocrine disrupting chemicals, potentially leading to long-term repercussions on the phenotype later in life. The mechanisms underlying this phenomenon, referred to as the Developmental Origins of Health and Disease (DOHaD), are still poorly understood. To gain molecular understanding of these effects, we exposed mangrove rivulus (Kryptolebias marmoratus) for 28 days post hatching (dph) to 4 and 120 ng/L 17-α-ethinylestradiol, a model xenoestrogen. After 28 days, fish were raised for 140 days in clean water and we performed quantitative label-free proteomics on brain, liver and ovotestis of 168 dph adults. A total of 820, 888 and 420 proteins were robustly identified in the brain, liver and ovotestis, respectively. Effects of 17-α-ethinylestradiol were tissue and dose-dependent: a total of 31, 51 and 18 proteins were differentially abundant at 4 ng/L in the brain, liver and ovotestis, respectively, compared to 20, 25 and 39 proteins at 120 ng/L. Our results suggest that estrogen-responsive pathways, such as lipid metabolism, inflammation, and the innate immune system were affected months after the exposure. In addition, the potential perturbation of S-adenosylmethionine metabolism encourages future studies to investigate the role of DNA methylation in mediating the long-term effects of early-life exposures. SIGNIFICANCE: The Developmental Origins of Health and Disease (DOHaD) states that early life stages of humans and animals are sensitive to environmental stressors and can develop health issues later in life, even if the stress has ceased. Molecular mechanisms supporting DOHaD are still unclear. The mangrove rivulus is a new fish model species naturally reproducing by self-fertilization, making it possible to use isogenic lineages in which all individuals are highly homozygous. This species therefore permits to strongly reduce the confounding factor of genetic variability in order to investigate the effects of environmental stress on the phenotype. After characterizing the molecular phenotype of brain, liver and ovotestis, we obtained true proteomic reaction norms of these three organs in adults after early life stages have been exposed to the common endocrine disruptor 17-α-ethinylestradiol (EE2). Our study demonstrates long-term effects of early-life endocrine disruption at the proteomic level in diverse estrogen-responsive pathways 5 months after the exposure. The lowest tested and environmentally relevant concentration of 4 ng/L had the highest impact on the proteome in brain and liver, highlighting the potency of endocrine disruptors at low concentrations.

Keywords: 17-α-ethinylestradiol; DOHaD; Delayed effects; Early-life exposure; Endocrine disruption; Fish; Mangrove rivulus; Proteomics.