Role of AhR and Oxidative Stress in the Regioselective Toxicities of Hydroxychrysenes in Embryonic Japanese Medaka (Oryzias latipes).

Oxygenated polycyclic aromatic hydrocarbons (oxy-PAHs) are environmental contaminants that can be created through oxidation of parent PAHs. Previous studies have found that 2-hydroxychrysene (2-OHCHR) causes anemia in embryonic Japanese medaka while 6-hydroxychrysene (6-OHCHR) did not, an example of regioselective toxicity. Anemia was prevented by CYP inhibition, which reduced the formation of the potential oxidatively active metabolite, 1,2-catechol, from 2-OHCHR. 2-OHCHR has also been found to be a four-fold more potent aryl hydrocarbon receptor (AhR) agonist compared to 6-OHCHR. These findings led us to hypothesize that AhR activation and/or oxidative stress play an important role in 2-OHCHR toxicity. While treatments with the AhR agonists PCB126 and 2-methoxychrysene (2-MeOCHR) did not cause significant toxicity, pretreatments with the AhR antagonist, CH-223191, reduced anemia by 97.2% ± 0.84 and mortality by 96.6% ± 0.69. AhR inhibition by the antagonist was confirmed by significant reductions (91.0% ± 9.94) in induced EROD activity. Thiobarbituric acid reactive substances (TBARS) concentrations were 32.9% ± 3.56 higher (p<0.05) in 2-OHCHR treatments at 100 hpf compared to controls. Staining 2-OHCHR-treated embryos with the reactive oxygen species (ROS) scavenger, 2',7'-dichlorofluorescin diacetate revealed 32.6% ± 2.69 of 2-OHCHR-treated embryos exhibiting high concentrations of ROS in caudal tissues, which is a site for embryonic hematopoiesis in medaka. Pretreatment with antioxidants, N-acetylcysteine or vitamin E, significantly reduced 2-OHCHR-induced anemia (80.7% ± 1.12 and 99.1% ± 0.43) and mortality (67.1% ± 1.69 and 98.9% ± 0.66). These results indicate that AhR may mediate 2-OHCHR toxicity through canonical signaling by upregulating CYP1, enhancing the formation of reactive metabolites of 2-OHCHR that generate ROS within caudal hematopoietic tissues, potentially disrupting hematopoiesis, leading to anemia and subsequent mortality. This article is protected by copyright. All rights reserved. Environ Toxicol Chem 2023;00:0-0. © 2023 SETAC.