Document Type

Dissertation

Date of Degree

Spring 2016

Degree Name

PhD (Doctor of Philosophy)

Degree In

Pharmaceutical Sciences and Experimental Therapeutics

First Advisor

Michael W. Duffel

Abstract

Polychlorinated biphenyls (PCBs) are a class of persistent organic pollutants that have been associated with a myriad of negative human health effects. These man-made compounds were used throughout most of the 20th century and although their intentional production has since been banned and their use limited to closed systems, their prevalence in the environment remains a factor in disease states for exposed populations. The worldwide levels of PCBs has been declining, however, there is evidence for renewed sources of these compounds. The presence of PCBs with lower numbers of chlorine atoms (LC-PCBs) have been verified as unintentional byproducts in paints and pigments, the decomposition of PCB waste, or the recycling or disposal attempts of PCB-laden materials. While exposure to the higher chlorinated congeners (HC-PCBs) is often attributed to the consumption of contaminated water or fatty animal meat, a significant route of exposure to the airborne LC-PCBs is through inhalation. These semi-volatile compounds have been detected in high quantities in both indoor and outdoor air in urban and rural communities, and their presence is pronounced in older buildings (e.g., homes and schools). When compared to HC-PCBs, LC-PCBs are more highly susceptible to metabolic transformations, and recently their sulfated metabolites have gained much interest. Although the sulfation of xenobiotics often is considered a route for their removal from the body, a previous study of Sprague-Dawley rats treated with 4-chlorobiphenyl (PCB 3) resulted in the substantial formation of sulfated metabolites (i.e., hydroxylation followed by sulfation of the LC-PCB). This metabolic route accounted for more than half of the treatment dose. Furthermore, LC-PCB sulfates have been shown to bind to the human serum protein, transthyretin, in vitro.

Of the health effects associated with PCB exposure, neurotoxicity has been well established through various laboratory and epidemiological studies. It is proposed that the dopaminergic system lies at the core of the observed cognitive, motor, and intellectual dysfunction observed in exposed populations, especially in children exposed perinatally. Interestingly, PCB exposure has been linked to Parkinson's disease (PD) etiology, which is marked by a substantial loss of dopaminergic neurons.

This thesis describes studies on the binding of selected LC-PCBs and their hydroxylated and sulfated metabolites to human serum albumin (HSA), the most abundant protein in human serum. The displacement of fluorescent probes, selective for the two major drug binding sites of HSA, indicates that LC-PCB sulfates generally bind to HSA with such affinity that is equal to or greater than that for the LC-PCBs or OH-LC-PCBs This work also included a study of the selective toxicity of these compounds to dopaminergic neuronal cells. The selective toxicity of these compounds was studied in a series of immortalized cell lines (i.e., two neuronal cell lines: the rat midbrain-derived N27 cell line, the human neuroblastoma-derived SH-SY5Y cell line, and the human liver-derived HepG2 cell line). The assessment of toxicity by MTT reduction and LDH release in these cellular models indicated that hydroxylated and sulfated metabolites of LC-PCBs exhibited toxicity that was selective to neuronal cells and, in most cases, selective for the dopaminergic neuronal cells. Furthermore, HPLC analysis of the distribution of the compounds from the extracellular medium into the cellular milieu indicated that the observed toxicity may be due in some cases to selective transport and further metabolism. This work contributes to understanding the neurotoxicity of LC-PCB hydroxylated and sulfated metabolites and the role that binding to serum proteins may play in it. Furthermore, it emphasizes the need for future studies on the effects that metabolism, particularly sulfation, may play in the disposition of LC-PCB congeners as it pertains to their metabolism, retention, and toxic effects.

Public Abstract

The harmful effects that polychlorinated biphenyls (PCBs) exert on biological and environmental systems is a great concern for our planet. Exposure to these agents has been associated with various human disorders and diseases, some of which involve the development and function of the brain. Although overall PCB levels have decreased worldwide in the last few decades, PCBs with lower numbers of chlorine atoms (LC-PCBs) have garnered increasing interest due to their prevalence in indoor and outdoor air. These types of PCBs undergo metabolic changes in the body, and one metabolic pathway results in the formation of hydroxylated and sulfated derivatives. A sulfate group may make the PCB more readily excreted, but it may also enable its transport to tissues through binding to the most abundant protein in serum, human serum albumin (HSA). The binding of LC-PCBs and hydroxylated (OH-LC-PCBs) and sulfated metabolites to HSA is reported here. It was found that generally, PCB sulfates bound to HSA with a comparable or higher affinity than the LC-PCBs or the OH-LC-PCBs. Furthermore, the neurotoxic activity of these metabolites was assessed by measuring their effect on cell viability in neuronal cells and comparing the results to hepatic cells. OH-LC-PCBs were more toxic to the former than the latter, and one PCB sulfate exhibited toxicity similar to the OH-LC-PCBs. The studies reported in this thesis contribute to further understanding the neurotoxicity of LC-PCB and hydroxylated and sulfated metabolites, and the role that binding to serum proteins may play in it.

Keywords

publicabstract, hydroxylation, metabolism, neurotoxicity, polychlorinated biphenyls (PCBs), serum protein binding, sulfation

Pages

xxii, 139 pages

Bibliography

Includes bibliographical references (pages 120-139).

Copyright

Copyright 2016 Eric Alberto Rodriguez

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