Bioactivation of Monocrotaline by P-450 3A in Rat Liver

Monocrotaline (MCT) is bioactivated in liver cytochrome P-450s to MCT pyrrole (MCTP), which primarily injures the lung endothelium to result in the development of pulmonary hypertension (PH) in rats. However, whether there is a relation between the degree of PH and the activity of liver cytochrome P-450 to convert MCT to MCTP remains unclear. To examine the relation between these physiological and biochemical changes, we first measured the severity of MCT-induced (20 mg/kg) PH in male, female, castrated male, and phenobarbital (PB, liver P-450s inducer)-pretreated male rats. The degree of right ventricular hypertrophy was more severe in PB-pretreated male than in control male rats. It was also more severe in male than in either female or castrated male rats, suggesting that sex-specific P-450s could be involved in the metabolic pathways of MCT in the liver. Further to explore which of the isozymes (2A2, 2C11, and 3A) of P-450s in the liver is responsible for the bioactivation of MCT, we measured the rate of MCTP production in hepatic microsomes by a modified Mattock's method. Treatment of male rats with PB and pregnenolone 16α-carbonitrile (PCN), which is the specific inducer of P-450 3A, increased the rate of MCTP production, suggesting that P-450 3A may contribute to the conversion to pyrrole. Therefore we measured the amount of P-450 3A protein by immunoblotting and attempted to inhibit MCT metabolism by using antibodies to P-450 3A. P-450 3A was significantly induced by PCN (6.5-fold) and PB (4.6-fold) treatment and reduced by castration (0.38-fold). The amount of P-450 3A was closely correlated with the production of MCTP, and the conversion of MCT to MCTP was strongly inhibited by antibodies against P-450 3A. These results indicated that P-450 3A was predominantly responsible for the metabolism of MCT to MCTP in rat liver and suggested a tight linkage between the degree of PH and the activity of liver P-450 3A.