Effect of hyperthyroidism on the in vitro metabolism and covalent binding of 1,1-dichloroethylene in rat liver microsomes.
Hyperthyroidism potentiates the in vivo hepatotoxicity of 1,1-dicholoroethylene (DCE) in rats, with a concomitant increase in [14C]-DCE covalent binding. The enhanced injury produced in hyperthyroid livers by DCE could be due to alterations in either the bioactivation or detoxication phases of DCE metabolism. Previous in vitro studies suggested that hyperthyroidism did not potentiate DCE hepatotoxicity by increasing DCE oxidation to intermediates which were able to covalently bind. Several factors, however, that could contribute to the magnitude of DCE bioactivation or covalent binding were not examined. Our objectives were to characterize the effects of hyperthyroidism in male Sprague-Dawley rats on: (1) covalent binding of [14C]-DCE to microsomes and other subcellular fractions, (2) microsomal mixed-function oxidase (MFO) and glutathione S-transferase (GST) activities, and (3) inactivation of microsomal enzyme activities by presumptive DCE reactive intermediates. Hyperthyroid (HYPERT) and euthyroid (EUT) rats received 3 s.c. injections of thyroxine (100 micrograms/100 g) or vehicle, respectively, at 48-h intervals; microsomes and other subcellular fractions were isolated from HYPERT and EUT livers 24 h after the last injection. [14C]-DCE-derived covalent binding was consistently greater in EUT than HYPERT microsomes. The absence of NADH, and the addition of low concentrations (0.1 and 0.5 mM), but not higher concentrations (> 1 mM), of glutathione (GSH) diminished covalent binding to a greater extent in HYPERT than EUT microsomes. Covalent binding in mitochondrial, nuclear, and cytosolic fractions of EUT and HYPERT livers was equivalent. Regression analysis of covalent binding to liver cell fractions from both EUT and HYPERT rats showed a significant correlation with P-450 content. Hyperthyroidism decreased microsomal, but not mitochondrial, cytochrome P-450 content, and MFO activities for 7-ethoxycoumarin and benzphermine were similarly decreased. Hyperthyroidism also diminished microsomal GST activity, and altered GST kinetics for both GSH and 1-chloro-2,4-dinitrobenzene (CDNB). The magnitude of inactivation of MFO and GST activities in the presence of DCE (presumably by DCE reactive intermediates) was comparable between EUT and HYPERT microsomes. When covalent binding was standardized to cytochrome P-450 concentrations in microsomes and mitochondria, HYPERT fractions exhibited slightly greater covalent binding than EUT fractions, suggesting that hyperthyroidism does not reduce the capacity of P-450 hemoproteins to bioactive DCE. Thus, potentiation of DCE hepatotoxicity by hyperthyroidism may be predominantly due to diminished Phase II constituents, and major increases in reactive intermediate/conjugates that covalently bind to and impair critical cellular molecules.
Gunasena
G H
GH
Department of Pathology, University of Texas Medical Branch, Galveston 77555-0609, USA.
Kanz
M F
MF
D015660:7-Alkoxycoumarin O-Dealkylase / Q000378:metabolism; D000818:Animals; D001667:Binding, Competitive; D001711:Biotransformation; D002250:Carbon Radioisotopes; D003577:Cytochrome P-450 Enzyme System / Q000378:metabolism*; D004000:Dichloroethylenes / Q000737:chemistry / Q000378:metabolism* / Q000633:toxicity*; D005982:Glutathione Transferase / Q000378:metabolism; D006980:Hyperthyroidism / Q000378:metabolism*; D066298:In Vitro Techniques; D008297:Male; D008862:Microsomes, Liver / Q000187:drug effects / Q000201:enzymology / Q000378:metabolism* / Q000473:pathology; D006899:Mixed Function Oxygenases / Q000378:metabolism; D010089:Oxidoreductases, N-Demethylating / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D013347:Subcellular Fractions / Q000378:metabolism; D013974:Thyroxine / Q000008:administration & dosage / Q000494:pharmacology
D002250:Carbon Radioisotopes; D004000:Dichloroethylenes; C029297:vinylidene chloride; D003577:Cytochrome P-450 Enzyme System; D006899:Mixed Function Oxygenases; D015660:7-Alkoxycoumarin O-Dealkylase; C019781:benzphetamine N-demethylase; D010089:Oxidoreductases, N-Demethylating; D005982:Glutathione Transferase; D013974:Thyroxine