Effects of Perfluoroalkyl Acids on In Ovo Toxicity and Gene Expression in the Domestic Chicken (Gallus gallus domesticus)

Abstract

Perfluoroalkyl acids (PFAAs) are a family of synthetic substances used in a wide variety of consumer and industrial applications, including non-stick and stain-resistant products. PFAAs, specifically perfluorinated sulfonates and carboxylates, are chemically stable and virtually non-biodegradable in the environment. In recent years, PFAAs have been detected in tissues and blood of humans and wildlife. Furthermore, PFAAs have a tendency to bioaccumulate and biomagnify in biota. Perfluorooctane sulfonate and perfluorooctanoate are known to be toxic when animals are exposed to environmentally-relevant levels, but scientists and regulators are challenged with determining and predicting their modes of action. There is some evidence to suggest that PFAAs can impact the thyroid hormone (TH) pathway and neurodevelopment. The studies presented in this thesis investigated the developmental effects and potential modes of action of newer PFAAs that are being introduced into the global market place. Egg injection experiments were performed in domestic chicken (Gallus gallus domesticus) embryos to assess the in ovo toxicity of perfluorohexane sulfonate (PFHxS) and perfluorohexanoate (PFHxA) during development. Real-time RT-PCR was used to measure the transcription of candidate genes in the liver and cerebral hemisphere of day 21-22 embryos. Candidate genes were selected based on their responsiveness to PFAA exposure in an in vitro screening assay conducted previously. In ovo exposure to PFHxS decreased embryo pipping success and overall growth at 38,000 ng/g; several orders of magnitude higher than concentrations reported in wild bird eggs. The expression of TH-responsive genes, including type II and III 5'-deiodinase, neurogranin, and octamer motif binding factor 1, were induced. In addition, PFHxS diminished free thyroxine (T4) levels in plasma. PFHxA had no affect on pipping success, gene expression or T4 levels in chicken embryos at the doses assessed. The transcriptional profiles in the cerebral hemisphere of chicken embryos exposed to 890 and 38,000 ng/g PFHxS were compared to a solvent control using microarray technology. The expression of 78 different genes were significantly altered (fold change > 1.5, p < 0.001) by PFHxS. Functional analysis showed that PFHxS affected genes involved in tissue development and morphology and cellular assembly and organization. Pathway and interactome analysis suggested that gene expression may be affected through integrin receptors and signaling pathways via TH–dependent and –independent modes of action. It is expected that the findings presented in this thesis will be of general relevance and importance to regulatory agencies and of interest to research scientists and risk assessors.