Interactions of per-and-poly fluoroalkyl substances (PFAS) with the Aryl hydrocarbon receptor (AHR) pathway and larval toxicity in zebrafish (Danio rerio)

Polycyclic aromatic hydrocarbons (PAHs) are a class of chemicals frequently produced by combustion and industrial processes. They are frequently present in drinking water, and many are both carcinogenic and developmentally toxic. Several interact with the aryl hydrocarbon receptor (AHR), which regulates gene expression responses to xenobiotic exposure. Increased expression of cytochrome P450 (CYP) family 1 genes contribute to clearance and metabolism of some xenobiotics. Per- and poly- fluoroalkyl substances (PFAS) are persistent environmental contaminants widely produced for uses as surfactants, nonstick coatings, and in firefighting foams. They are also often present in drinking water and are developmentally toxic. Several PFAS have been shown to interact with both CYP proteins as well as the AHR, raising the question of whether mixtures of these chemicals as seen in the environment may be more toxic than each chemical individually. However, PFAS interaction with CYPs and AHR is not well explored. This work aims to investigate the interactions between PFAS and the AHR response to xenobiotics as well as associated developmental toxicity of these substances. Transgenic, fluorescent reporter zebrafish were exposed to PFAS and beta-naphthoflavone (an AHR agonist) mixtures from 24-100 hours post fertilization; then, fish were assessed for morphology, CYP1A-like-acitivity, pancreatic and liver development, and gene expression. Co-exposures of the 8-carbon, sulfonated perflourooctane sulfonate (PFOS) and beta-naphthoflavone were found to significantly reduce fish length, increase pericardial area, and decrease CYP1A-like activity compared to the same level of beta-naphthoflavone exposure alone; expression of the related cyp1a gene was also reduced by co-exposure. In contrast, the 6-carbon perflourohexane sulfonate (PFHxS) did not significantly affect pericardial area or CYP1A-like activity in co-exposures, and cyp1a expression was instead increased. PFOS was also found to inhibit the catalytic activity of human CYP1A1 enzyme ex vivo. Many PFAS were found to have affinity for CYPs as well as nuclear receptors and other proteins related to AHR via in silico molecular docking as well as structural predictions. Thus, this study suggests PFAS and PAH mixtures may be more toxic than either individually, and provides insights into the molecular mechanism for PFAS interactions with AHR.