Induction of hepatic metallothionein in male B6C3F1 mice exposed to hepatic tumor promoters: effects of phenobarbital, acetaminophen, sodium barbital, and di(2-ethylhexyl) phthalate.

The effects of various compounds known to be hepatic tumor promoters and toxins in the male B6C3F1 mouse liver, including di(2-ethylhexyl)phthalate (DEHP), acetaminophen (ACT), barbital (BB), and phenobarbital (PB) on hepatic metallothionein (MT) concentrations were assessed after chronic exposure. From 6 weeks of age, male mice were maintained on diets containing DEHP at 12,000 or 6000 ppm, ACT at 10,000 or 5000 ppm, BB at 1,000 ppm, or drinking water with PB at 500 ppm for up to 24 weeks. MT was measured in hepatic cytosol at 0, 2, 8, and 24 weeks of exposure. DEHP proved a very effective inducer, producing elevations of MT as high as 11-fold. The increases in hepatic MT with DEHP were both dose- and time-related. ACT was likewise effective in producing hepatic MT elevations (maximum 6.7-fold) in a dose- and time-related fashion. BB and PB, however, had no effect on hepatic MT levels at any time point. While DEHP, BB, and PB treatments produced hepatomegaly, histopathological analysis at 24 weeks revealed that in both DEHP- and ACT-treated livers hepatocellular proliferation was prominent while livers exposed to BB or PB showed predominantly hepatocellular hypertrophy. Gel-filtration of DEHP-treated liver cytosol revealed that zinc was associated with the MT peak. This peak also bound cadmium in vitro and could be extracted by heat treatment and selective acetone precipitation, both typical characteristics of MT. Further confirmation of the presence of MT after DEHP treatment was obtained by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (10 to 20% acrylamide). Results indicate that some, but not all, tumor promoters can induce target organ MT and that such an induction appears associated with those promoters inducing persistent cellular hyperplasia but not those inducing cellular hypertrophy.