Effects of cytochrome P450-inducing agents on the monooxygenation of testosterone in long-term cultures of hepatocytes from male and female rats

Abstract

Hepatocytes from male or female rats were cultured for up to 2 weeks in a modified Waymouth medium supplemented with 0.1 or 1.0 μ m dexamethasone, 10 n m insulin, and 0.1 n m glucagon with or without addition of phenobarbital, methylcholanthrene, or isoniazid. The activities of testosterone hydroxylases were measured in the intact cell monolayer and in the corresponding microsomal fraction. Aniline hydroxylase was measured in cell homogenates. In the presence of 0.1 μ m dexamethasone the testosterone hydroxylase activities varied differently in hepatocytes from male and female rats during the culture period. The activities of 6β- and 15α-hydroxylases increased in female and were unchanged in male hepatocytes, while 16α-hydroxylase activity increased in female and decreased in male, and 2α- and 7α-hydroxylase activities were unchanged in both male and female hepatocytes during the culture period. Increasing the dexamethasone concentration to 1.0 μ m caused an increase in 6β- and 15α-hydroxylase activities in cultures of hepatocytes from both sexes, whereas an increase of 2α- and a decrease of 7α- and 17-hydroxylase activities were found only in cultures of hepatocytes from female rats. Addition of phenobarbital caused an increase in the activity of 7α-hydroxylase in both male and female hepatocytes, while the effect on the other hydroxylases differed with the sex. In hepatocytes from male rats phenobarbital addition decreased the activities of 2α- and 16α-hydroxylases, while these were increased or stable after addition of phenobarbital to hepatocytes from female rats. The activity of aniline hydroxylase was increased at Day 1 and declined afterward. The results demonstrate that the activities of different steroid hydroxylases are inducible and can be directly measured in monolayers of hepatocytes from rats.