Monoclonal antibodies against human liver cytochrome P-450.

Monoclonal hybridomas which produce antibodies against human liver microsomal cytochrome P-450 were developed. Three similar hybridomas produced antibodies which recognized an epitope specific to a family of human P-450 isozymes (P-450(5)). This epitope was also present on cytochrome P-450 PCN-E (pregnenolone-16 alpha-carbonitrile induced) from rat liver microsomes, but this isozyme differed from the human P-450(5) by its molecular weight. These antibodies enabled us to quantify cytochrome P-450(5) in human liver microsomes and to demonstrate an important quantitative polymorphism in the human liver monooxygenase system.

Preparation and characterization of monoclonal antibodies against phenobarbital-inducible cytochrome P-450.

Monoclonal antibodies against cytochrome P-450 were prepared from phenobarbital-induced rat liver microsomes. The immunoglobulin classes and subclasses, as well as the binding capacity to cytochrome P-450, of the different antibodies were characterized. Their specificity was verified by various techniques and seemed to correspond to a single form of cytochrome P-450, the major phenobarbital-inducible form. However, the antibodies were unable to inhibit completely the monooxygenase activities investigated. These antibodies may constitute very specific and powerful analytical tools for characterizing and quantifying cytochrome P-450 isoenzymes.

The expression of different monooxygenases supported by cytochrome P-450 in neonatal rats and in primary fetal hepatocytes in culture.

In rat liver, the perinatal development of various monooxygenase activities follows different patterns, depending upon the reaction studied. The ontogeny of the 6 beta-, 7 alpha- and 16 alpha-testosterone hydroxylase activities differs very significantly. Aldrin epoxidase and steroid-metabolizing monooxygenases are expressed in primary fetal rat liver cells in culture after treatment in vitro with dexamethasone. Testosterone is not metabolized by the control cells and is hydroxylated on the 6 beta and 16 alpha positions following the addition of corticoids to the culture medium. The dose and time curves vary according to the hydroxylated position of the steroid. Aldrin epoxidase activity is nearly undetectable in the control cells, but is present and is inducible by phenobarbital following treatment with the corticoid. Phenobarbital induces aldrin epoxidase in the absence of dexamethasone in the culture medium, providing that the cells are pretreated with the corticoid for 48 h. The use of antibodies against the main cytochrome P-450 species purified from adult and phenobarbital-treated rats confirms that a similar cytochrome P-450 can be induced in fetal cells in culture. The perinatal regulation of biological events, such as the expression of the monooxygenases, can be reproduced in fetal rat liver cells in culture; such a model constitutes a unique tool for studying the biochemical mechanisms which control these phenomena.