Drug metabolism in adult rat hepatocytes in primary monolayer culture.
Primary monolayer culture of adult rat hepatocytes represents a novel and potentially useful technique to study many aspects of hepatic physiology for extended periods of time in vitro (J Cell Biol 59:722-734, 1973). In examining the hepatic drug-metabolizing system in these cells, we have discovered that the conditions of cell culture exert rapid, selective, and reproducible changes in microsomal enzymes. In the 24- to 48-hr period immediately following preparation and culture of the isolated parenchymal cells, the level of the drug-binding microsomal hemoprotein, cytochrome P-450, measured in extracts of cell homogenates, declined to less than 20% of its in vivo level, whereas NADPH-cytochrome c reductase activity was only moderately reduced and glucose-6-phosphatase activity remained unchanged. The activity of aminopyrine-N-demethylase and aniline hydroxylase also fell, paralleling the level of cytochrome P-450. By contrast, p-nitroanisole O-demethylase activity was unchanged in the cultured hepatocytes despite evidence (type I binding spectrum, NADPH requirement, inhibition by carbon monoxide or by SKF 525A) that p-nitroanisole O-demethylase is a cytochrome P-450-dependent enzyme. In culture, as in vivo, aromatic polycyclic hydrocarbons stimulated p-nitroanisole O-demethylase and aryl hydrocarbon (benzo [a] pyrene) hydroxylase activities; however, this effect was unaccompanied by a detectable increase in total carbon monoxide-binding hemoprotein. The data indicate that the profile of microsomal oxidase enzymes and their control undergo striking changes as hepatocytes adapt to cell culture.
D000633:Aminopyrine N-Demethylase / Q000378:metabolism; D000815:Aniline Hydroxylase / Q000378:metabolism; D000818:Animals; D001189:Aryl Hydrocarbon Hydroxylases / Q000378:metabolism; D001580:Benzopyrenes / Q000494:pharmacology; D002478:Cells, Cultured; D003577:Cytochrome P-450 Enzyme System / Q000378:metabolism; D004790:Enzyme Induction; D008658:Inactivation, Metabolic; D008099:Liver / Q000201:enzymology / Q000378:metabolism*; D008862:Microsomes, Liver / Q000201:enzymology; D009251:NADPH-Ferrihemoprotein Reductase / Q000378:metabolism; D009575:Nitroanisole O-Demethylase / Q000378:metabolism; D004364:Pharmaceutical Preparations / Q000378:metabolism*; D051381:Rats
D001580:Benzopyrenes; D004364:Pharmaceutical Preparations; D003577:Cytochrome P-450 Enzyme System; D009575:Nitroanisole O-Demethylase; D000815:Aniline Hydroxylase; D001189:Aryl Hydrocarbon Hydroxylases; D000633:Aminopyrine N-Demethylase; D009251:NADPH-Ferrihemoprotein Reductase