Progesterone C21 hydroxylation and steroid carboxylic acid biosynthesis in the rabbit. In vitro studies with endocrine, metabolic, and potential target tissues

Abstract

Progesterone C21-hydroxylase activity has been demonstrated with rabbit kidney microsomes for the first time and the formation of 21-hydroxy-4-pregnen-3,20-dione (DOC) by rabbit liver and kidney microsomes has been quantitated. Considerable intraspecies variability in enzyme activity occurred with both tissues. The liver enzyme (Vmax = 1.28–38.0 nmol/mg protein per 30 min of incubation) was significantly more active than the kidney enzyme (Vmax = 0.028–0.28 nmol/mg protein per 30 min of incubation). Apparent KM values were 1.39 and 0.8 μM, respectively. Cytochrome c (10−5 M), potassium ferricyanide (10−3 M), and 2-methyl-1,2-di-3-pyridyl-1-propanone (metyrapone; 10−3 M) were strongly inhibitory with both tissues, whereas the liver microsomal system was less sensitive than the kidney to CO–air (90:10 v/v) inhibition. Metabolism of [14C]DOC to 4-pregnen-3,20-dione-21-oic (pregnenoic) and 4-androsten-3-one-17β-carboxylic (etienic) acids by liver microsomes and adrenal and ovary homogenates was differentially affected by several factors. CO–air (90:10 v/v), cytochrome c (10−5 M), and metyrapone (10−3 M) inhibited pregnenoic acid synthesis to a greater extent than etienic acid. Sodium cyanide had a stimulatory effect on the synthesis of pregnenoic acid by the liver but less consistent effects with other tissues. These results suggest that one or more cytochrome P-450 systems may be involved in the oxidation of progesterone through to pregnenoic acid by rabbit tissues.