Stress protein synthesis induced in rat liver by cadmium precedes hepatotoxicity.
A diverse array of chemical and physical stressors increases the synthesis of a class of proteins referred to as heat shock proteins (hsp) or stress proteins. We are investigating the potential of using altered protein synthesis patterns, including the enhanced synthesis of stress proteins, as biomarkers of exposure and cellular injury. The purpose of the present study was to evaluate the effect of a model hepatotoxicant, cadmium (Cd(II)), on stress protein synthesis in male rat liver. To assess target tissue specificity, stress protein synthesis was also studied in kidney. Liver and kidney slices from exposed rats were incubated with [35S]methionine for 1.5 hr, subjected to one-dimensional SDS-PAGE, and 35S-labeled proteins were analyzed by autoradiography. Enhanced de novo synthesis of 70-, 90-, and 110-kilodalton (kDa) relative molecular mass (M(r)) proteins was detected 2 hr after exposure to 2 mg Cd/kg, with maximum activity occurring at 2-4 hr. By 8-16 hr postinjection, synthesis of these proteins had decreased. Synthesis of a 68-kDa protein present in control liver was inhibited 2 hr after exposure with synthesis restored at 16-24 hr. Dose-related increases in synthesis of the three stress proteins were observed 4 hr after iv injection of 1.0 and 2.0 mg Cd/kg, with concomitant inhibition of synthesis of the 68-kDa protein. Mild single cell necrosis of hepatocytes was observed 8 hr after injection of 2 mg Cd/kg which progressed to mild multifocal necrotic foci at 16 hr. No lesions were evident at lower dosages. Increases in plasma sorbitol dehydrogenase activity, a clinical indicator of hepatic injury, was not apparent until 8 hr after exposure. A functional deficit, decreased hepatic microsomal N-demethylase activity, was not observed until 16 hr after iv injection of 2 mg/kg. No changes in kidney de novo stress protein synthesis were observed. No evidence of renal injury was apparent, as evaluated by histopathology, uptake of [para-3H]aminohippurate into renal slices, and blood urea nitrogen values. The 70-kDa stress protein induced early after Cd treatment was identified with a monoclonal antibody as the 72-kDa-inducible hsp. The 90-kDa protein induced by Cd reacted negatively with three monoclonal antibodies to hsp90 and was subsequently identified as a glucose regulated protein (grp94). The data demonstrate that Cd induces alterations in the expression of hepatic gene products in vivo as evidenced by enhanced stress protein synthesis and inhibition of synthesis of constitutive proteins.(ABSTRACT TRUNCATED AT 400 WORDS)
Health Sciences Branch, Food and Drug Administration, Rockville, Maryland 20857.
P L
PL
Goering
B R
BR
Fisher
C L
CL
Kish
D000818:Animals; D000911:Antibodies, Monoclonal; D001345:Autoradiography; D002104:Cadmium / Q000032:analysis / Q000633:toxicity*; D004591:Electrophoresis, Polyacrylamide Gel; D006360:Heat-Shock Proteins / Q000096:biosynthesis*; D007275:Injections, Intravenous; D007668:Kidney / Q000187:drug effects / Q000378:metabolism; D008099:Liver / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D008297:Male; D008862:Microsomes, Liver / Q000201:enzymology; D008970:Molecular Weight; D010089:Oxidoreductases, N-Demethylating / Q000378:metabolism; D014176:Protein Biosynthesis; D051381:Rats; D017207:Rats, Sprague-Dawley
D000911:Antibodies, Monoclonal; D006360:Heat-Shock Proteins; D002104:Cadmium; D010089:Oxidoreductases, N-Demethylating