1999 Progress Report: Endocrine Disruptors: Effects on the ThyroidEPA Grant Number: R826297
Title: Endocrine Disruptors: Effects on the Thyroid
Investigators: Klaassen, Curtis D.
Institution: University of Kansas Medical Center
EPA Project Officer: Hahn, Intaek
Project Period: December 15, 1997 through December 14, 2000
Project Period Covered by this Report: December 15, 1998 through December 14, 1999
Project Amount: $594,660
RFA: Endocrine Disruptors (1997) RFA Text | Recipients Lists
Research Category: Endocrine Disruptors , Economics and Decision Sciences , Health , Safer Chemicals
The ultimate goal of this project is to assess the significance of endocrine disruptors that increase thyroxine (T4) glucuronidation on thyroid carcinogenesis because many endocrine disruptors are suspected to be thyroid tumor promoters. The mechanism by which endocrine disruptors promote thyroid tumors has been proposed to result from alterations in the thyroid-pituitary-hypothalamus axis. Endocrine disruptors alter the thyroid-pituitary-hypothalamus axis by increasing T4 glucuronidation and elimination, which reduces serum T4. As a compensatory feedback mechanism, thyroid stimulating hormone (TSH) will be released from the pituitary, which will stimulate the thyroid, and result in thyroid follicular cell proliferation and ultimately neoplasia. However, the preliminary studies suggest that a number of endocrine disruptors interfere with the normal hypothalamus-pituitary-thyroid axis because these endocrine disruptors do not increase serum TSH. Therefore, the central hypothesis of this application is that endocrine disruptors that increase T4 glucuronidation are thyroid tumor promoters only when they increase serum TSH.
To test this hypothesis, there are four specific aims:
? The hypothesis that endocrine disruptors decrease plasma T4 pharmacokinetically by increasing its glucuronidation will be tested. The pharmacokinetics (clearance, biliary and urinary excretion, and in vitro metabolism) of T4, as well as triiodothyronine (T3), will be determined to understand the mechanism(s) by which endocrine disruptors decrease serum T4 levels.
? The mechanism by which some polycyclic aromatic hydrocarbon (PAH)-type endocrine disruptors (such as 3-methylcholanthrene [MC] and polychlorinated biphenyls [PCBs]) "blunt" the TSH response to reduced serum T4 will be examined, either physiologically, pathologically or by thyroid receptor binding assays.
? The hypothesis that endocrine disruptors, which increase serum TSH levels, produce thyroid follicular cell proliferation will be examined via proliferating cell nuclear antigen (PCNA) immunocytochemistry.
? The hypothesis that the tumor promoting effects of endocrine disruptors are not correlated with the decrease in serum T4, but with the increase in TSH will be tested in a 26-week bioassay. Rats will be given the thyroid initiating agent, N-bis(2-dihydroxy-propyl)nitrosamine (DHPN), followed by exposure to endocrine disruptors. This study will provide critical information on the relationship between thyroid hormone imbalance, TSH secretion, and thyroid tumor promotion of rats treated with endocrine disruptors. If our hypothesis is true, then it has important implications in toxicology, for many endocrine disruptors have been shown to reduce serum T4 levels. However, their effect on TSH is, at best, variable.
Our expectation from the proposed studies is that increases in serum TSH, rather than reductions in serum T4, is a better indicator of thyroid tumorigenicity resulting from exposure to endocrine disruptors. If we demonstrate that the TSH mediates endocrine disruptor thyroid tumor promoting activity, then these endocrine disruptors could be regulated on knowledge rather than ignorance.
PCB mixtures and congeners effectively reduce circulating concentrations of T4. This is thought to occur because of their ability to induce the UDP-glucuronosyl transferases that conjugate T4 and the subsequent excretion of the glucuronide into bile. To determine whether there is a good correlation between the ability of PCBs to reduce T4 and increase the biliary excretion of T4 glucuronide, PCB congeners 95 (16 mg/kg), 99 (16 mg/kg), 118 (16 mg/kg), and 126 (40 mg/kg); Aroclors 1242 (32 mg/kg) and 1254 (32 mg/kg); and 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) (3.9 mg/kg) were administered via gavage to male Sprague-Dawley rats for 7 days. Twenty-four hours after the last dose, the femoral artery and vein and the common bile duct were cannulated. Following administration of [125I]T4, bile was collected at 30-minute intervals for 2 hours. Blood was collected at the mid-point of each bile collection period. Urine also was collected at 2 hours. The total excretion of T4 and its metabolites was quantified by gamma spectrometry, followed by high-performance liquid chromatography (HPLC) analysis. All seven treatments decreased the plasma concentration of T4. Of the pure congeners, PCB 99 and 118 produced the largest decreases in plasma T4 concentration, whereas PCB 95, PCB 126, and TCDD had the least effect. None of the seven treatments had a marked effect on the urinary excretion of T4 and its metabolites. In contrast, biliary excretion of T4 glucuronide after administration of TCDD, PCB 118, or Aroclor 1254 was increased 6- to 8-fold, and PCB 126 produced a 3- to 5-fold increase, whereas PCB 95 and PCB 99 produced less than a doubling. TCDD, one of the treatments that had the least effect on decreasing the concentration of T4 in blood, increased the biliary excretion the most. In contrast, PCB 99, one of the congeners that decreased the plasma concentration of T4 the most, had the least effect on its biliary excretion. Therefore, there does not appear to be a good correlation between the ability of PCBs to decrease plasma T4 concentration and increase its biliary excretion.
Circulating levels of thyroid hormones can be reduced by a number of compounds, including PCBs. Numerous studies have been conducted using mixtures of PCB congeners known as Aroclors, but little is known about the effects of the individual congeners. The objective of this study was to determine the effects of PCB congeners on serum levels of T4 and T3. PCB congeners are commonly divided into three general categories. The "TCDD type" are those congeners that have affinity for the aryl hydrocarbon receptor (AhR) and induce cytochrome P450 1A1 (CYP1A1); the "phenobarbital type" are those congeners that induce CYP2B1; and the "mixed type" are those congeners capable of induction of both CYP1A1 and CYP2B1. Four PCB congeners were selected for use in this study: PCB 126 (a "TCDD type"), PCBs 95 and 99 ("phenobarbital types"), and PCB 118 (a "mixed type"). Aroclors 1242 and 1254 and TCDD also were evaluated. Each compound was administered at a minimum of four dose levels via gavage to male Sprague-Dawley rats for 7 days. Blood was obtained via the retro orbital sinus on days ?3, 3, and at necropsy on day 7. On day 7, serum T4 levels were reduced to 5-10 percent of control values in rats administered Aroclor 1254 and PCBs 99 and 118. Reductions up to 50 percent of control values occurred for rats administered each of the remaining compounds. Serum T3 levels were reduced to 30 percent of the control values in rats administered Aroclor 1254. The remaining compounds caused only slight or no reductions in serum T3. Results of this study suggest that PCB congeners may induce the majority of their effects on thyroid functions via pathways that are not mediated by the AhR.