Changes in the quantity and activity of cytochrome P-450 isozymes in primary cultured rat hepatocytes.
Hepatocytes from male Sprague-Dawley rats pretreated with a cytochrome P-450 inducer, 3-methoxy-4-aminoazobenzene (3-MeO-AAB), 3-methylcholanthrene (MC) or phenobarbital (PB), were cultured in vitro, and changes in the quantity and activity of microsomal cytochrome P-450 isozymes in the cells were determined by means of immunochemical methods and a bacterial mutation test, respectively. The results of enzyme-linked immunosorbent assay using monoclonal antibodies against rat P-450 isozymes revealed that the amount of cytochrome P-450d induced by 3-MeO-AAB or MC declined rapidly during culture and fell to 10 to 15% of the initial value after 24 h. A similar tendency was observed with PB-induced cytochrome P-450b/e. By contrast, cytochrome P-450c in MC-induced hepatocytes declined more slowly than cytochrome P-450d and remained at 45 to 60% of the initial value after 24 h. Similar quantitative changes of the individual cytochrome P-450 isozymes in culture were also observed by immunoblotting using the anti-cytochrome P-450 monoclonal antibodies. Changes in the activities of individual cytochrome P-450 isozymes in hepatocytes by culture were in accordance with the quantitative changes of the cytochromes, as determined by a mutation test using Salmonella typhimurium TA 98 and carcinogenic aromatic amines. These results indicate that microsomal cytochrome P-450c in primary cultured rat hepatocytes is more stable in culture, in terms of both quantity and activity, than cytochrome P-450d and P-450b/e.
Namiki
M
M
Department of Hygienic Chemistry, Tohoku University, Sendai.
Degawa
M
M
Masuko
T
T
Hashimoto
Y
Y
D000818:Animals; D002478:Cells, Cultured; D003577:Cytochrome P-450 Enzyme System / Q000032:analysis* / Q000096:biosynthesis; D004790:Enzyme Induction; D004797:Enzyme-Linked Immunosorbent Assay; D015151:Immunoblotting; D007150:Immunohistochemistry; D007527:Isoenzymes / Q000032:analysis* / Q000096:biosynthesis; D008099:Liver / Q000166:cytology / Q000201:enzymology*; D008297:Male; D008862:Microsomes, Liver / Q000201:enzymology; D009154:Mutation; D051381:Rats; D011919:Rats, Inbred Strains; D012486:Salmonella typhimurium / Q000187:drug effects / Q000235:genetics
D007527:Isoenzymes; D003577:Cytochrome P-450 Enzyme System