Date of Award

Spring 5-1-2015

Document Type

Thesis

Degree Name

Bachelor of Arts

Department

Biochemistry & Molecular Biology

First Advisor

Wade Powell

Abstract

2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD or dioxin) is a persistent organic pollutant that elicits toxic effects in vertebrates, including developmental defects, cancer, endocrine disruption, and death. TCDD toxicity results from binding to the aryl hydrocarbon receptor (AHR), a ligand-activated transcription factor. High levels of dioxin disrupt the thyroid system in several species. However, a clear understanding of the underlying mechanism of dioxin-induced thyroid hormone (TH) disruption is lacking. Amphibian metamorphosis (the development of tadpoles into frogs) represents a well-established model system to assess thyroid hormone-driven vertebrate development. In this study we used the African clawed frog, Xenopus laevis, as a model to study TH disruption by dioxin, examining the expression of target genes of thyroid hormone receptor (TR) and AHR in XLK-WG cells as well as tail resorption during metamorphosis. Expression of Cytochrome P450 1A6 (CYP1A6) mRNA, a well-characterized AHR target, was induced at least 300-fold by 100 nM TCDD in XLK-WG cells. The primary TH target gene, Krüppel-Like Factor 9 (KLF9) was induced 5-fold by 50 nM TH and approximately 2-fold by dioxin. Upon co-exposure to TH and TCDD, CYP1A6 was induced at least 500-fold, while KLF9 was induced 11-fold. Increased target gene induction following co-exposure of XLK-WG cells to TH and TCDD occurred in the absence of serum in culture media. Therefore, this phenomenon did not involve competitive displacement from serum binding proteins and resulting changes in bioavailability of these compounds. Enhanced target gene expression was sensitive to pharmacological inhibitors of both TR and AHR, demonstrating the requirement for both receptors. In experiments with the candidate endogenous AHR ligand 6-formylindolo[3,2-b]carbazole (FICZ; 10 nM), CYP1A6 and KLF9 induction were not augmented by co-exposure to 50 nM TH compared to either compound alone. To pair molecular data with a morphological endpoint, we used tadpole tail explants to determine the effect of TCDD and TH co-exposure on the rate of tail resorption. We found that the decrease in tail area was, at best, modestly accelerated by co-exposure to TH and TCDD compared to TH alone. These findings suggest that molecular biomarkers may represent a more sensitive indicator than tail resorption for examining thyroid-disrupting effects of TCDD and other xenobiotics.

Share

COinS