Immunochemical detection of different isoenzymes of cytochrome P-450 induced in chick hepatocyte cultures.
This study investigated whether the same cytochrome P-450 (P-450) isoenzymes were inducible in cultures of chick-embryo hepatocytes as in the liver of chicken embryos. We purified two isoenzymes of cytochrome P-450 from the livers of 17-day-old-chick embryos: one of molecular mass approx. 50 kDa induced in vivo by the phenobarbital-like inducer glutethimide, and the second of approx. 57 kDa induced by 3-methylcholanthrene. Rabbit antiserum against the 50 kDa protein inhibited benzphetamine demethylase activity in hepatic microsomes (microsomal fractions) from glutethimide-treated chick embryo. Antiserum to the 57 kDa protein inhibited ethoxyresorufin de-ethylase activity in hepatic microsomes from methylcholanthrene-treated chick embryo. Cultured chick hepatocytes were treated with chemicals known to induce isoenzymes of P-450 in rodent liver. The induced P-450s were quantified spectrophotometrically and characterized by immunoblotting and enzyme assays. From these studies, chemical inducers were classified into three groups: (i) chemicals that induced a P-450 isoenzyme of 50 kDa and increased benzphetamine demethylase activity: glutethimide, phenobarbital, metyrapone, mephenytoin, ethanol, isopentanol, isobutanol, lindane, lysodren; (ii) chemicals that induced a P-450 isoenzyme of 57 kDa and increased ethoxyresorufin de-ethylase activity: 3-methylcholanthrene and 3,3',4,4'-tetrachlorobiphenyl; and (iii) the mono-alpha-substituted 2,3',4,4',5-pentabromobiphenyl, which induced both proteins and both activities. The immunochemical data showed that chick-embryo hepatocytes in culture retain the inducibility of glutethimide- and methylcholanthrene-induced isoenzymes of P-450 that are inducible in the liver of the chicken embryo.
Sinclair
P
P
Veterans Administration Hospital, White River Junction, VT 05001.
Frezza
J
J
Sinclair
J
J
Bement
W
W
Haugen
S
S
Healey
J
J
Bonkovsky
H
H
D000818:Animals; D000918:Antibody Specificity; D002478:Cells, Cultured; D002642:Chick Embryo; D003577:Cytochrome P-450 Enzyme System / Q000032:analysis* / Q000276:immunology / Q000302:isolation & purification; D015151:Immunoblotting; D007527:Isoenzymes / Q000032:analysis* / Q000276:immunology / Q000302:isolation & purification; D008099:Liver / Q000166:cytology / Q000201:enzymology*; D008862:Microsomes, Liver / Q000201:enzymology
D007527:Isoenzymes; D003577:Cytochrome P-450 Enzyme System