Sex difference in induction of hepatic CYP2B and CYP3A subfamily enzymes by nicardipine and nifedipine in rats

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Abstract

Male and female of F344 rats were treated per os with nicardipine (Nic) and nifedipine (Nif), and changes in the levels of mRNA and protein of hepatic cytochrome P450 (P450) enzymes, CYP2B1, CYP2B2, CYP3A1, CYP3A2, CYP3A9, and CYP3A18 were examined. Furthermore, hepatic microsomal activities for pentoxyresorufin O-dealkylation (PROD) and nifedipine oxidation, which are mainly mediated by CYP2B and CYP3A subfamily enzymes, respectively, were measured. Analyses of RT-PCR and Western blotting revealed that Nic and Nif induced predominantly CYP3A and CYP2B enzymes, respectively. As for the gene activation of CYP2B enzymes, especially CYP2B1, Nif showed high capacity in both sexes of rats, whereas Nic did a definite capacity in the males but little in the females. Gene activations of CYP3A1, CYP3A2, and CYP3A18 by Nic occurred in both sexes of rats, although that of CYP3A9 did only in the male rats. Although gene activations of CYP3A1 and CYP3A2 by Nif were observed in both sexes of rats, a slight activation of the CYP3A9 gene occurred only in female rats, and the CYP3A18 gene activation, in neither male nor female rats. Thus, changes in levels of the mRNA or protein of CYP2B and CYP3A enzymes, especially CYP2B1 and CYP3A2, were closely correlated with those in hepatic PROD and nifedipine oxidation activities, respectively. The present findings demonstrate for the first time the sex difference in the Nic- and Nif-mediated induction of hepatic P450 enzymes in rats and further indicate that Nic and Nif show different specificities and sex dependencies in the induction of hepatic P450 enzymes.

Introduction

Calcium channel antagonists are widely used for the treatment of hypertension. However, there are many reports on clinical drug–drug interactions with the calcium channel antagonists Bertz and Granneman, 1997, Campana et al., 1996, Rosenthal and Ezra, 1995, Schlanz et al., 1991. As one of possible explanations for such clinical drug–drug interactions, inhibition of the cytochrome P450 (P450) activities by calcium channel antagonists has been suggested, because calcium channel antagonists including nicardipine (Nic) and nifedipine (Nif) have been demonstrated to show capacities for inhibiting the activities of CYP3A, CYP2C9, or CYP2D6 in human hepatocytes and liver microsomes Ma et al., 2000, Pichard et al., 1990. In addition, calcium channel antagonists have been reported to serve as inhibitors of P450 enzymes, especially CYP3A subfamily enzymes, in animals Mäenpää et al., 1989, Murray and Butler, 1996.

On the other hand, calcium channel antagonists have been also reported to promote the metabolism of several drugs in rats and to shorten the hexobarbital-sleeping time Koleva and Stoychev, 1993, Koleva and Stoychev, 1995. More recently, calcium channel antagonists such as nicardipine (Nic) and nifedipine (Nif) have been found to show capacities for inducing CYP3A and CYP2B subfamily enzymes in the rat liver Konno et al., 2003, Zangar et al., 1999 and in human hepatocytes (Drocourt et al., 2001). These reports suggest that induction of P450s by calcium channel antagonists is a cause of the drug–drug interactions.

As for the induction of hepatic CYP3A and CYP2B subfamily enzymes in rats, gender-related differences are known. CYP3A2 is induced in adult male rats but not in the females by dexamethasone Cooper et al., 1993, Mahnke et al., 1997. Likewise, the induction of CYP2B subfamily enzymes, including CYP2B1 and CYP2B2, by phenobarbital occurs preferentially in male rats Agrawal and Shapiro, 1996, Larsen et al., 1994, Shapiro et al., 1994. To date, however, study on a gender-related difference in calcium channel antagonist-mediated induction of hepatic CYP2B and CYP3A subfamily enzymes has not been performed.

In the present study, we examined the induction of hepatic CYP2B and CYP3A subfamily enzymes by Nic and Nif in male and female rats and demonstrated for the first time the sex difference in the calcium channel antagonist-mediated induction of the P450 enzymes.

Section snippets

Chemicals

Nicardipine hydrochloride (Nic) and nifedipine (Nif) were purchased from Wako (Osaka, Japan). These chemicals were the highest grade available.

Animals and treatment

Male and female of F344 rats (6 weeks of age) were purchased from Japan SLC Animal (Hamamatsu, Japan), kept in plastic cages in an air-conditioned room, and given MF diet (Oriental Yeast, Tokyo, Japan) and water ad libitum, and used at 7 weeks of age. Rats were treated per os with either Nic or Nif dissolved in corn oil as described previously (Konno et

Dose effects of Nic and Nif on the gene expression of hepatic CYP2B and CYP3A subfamily enzymes

Dose effects of Nic and Nif on hepatic gene expression of CYP2B and CYP3A subfamily enzymes, CYP2B1, CYP2B2, CYP3A1 CYP3A2, CYP3A9, and CYP3A18, in male and female rats were examined by the RT-PCR with the corresponding primer sets. We have previously reported that treatment of male rats with either Nic or Nif resulted in significant increases in hepatic expression levels of the CYP2B1, CYP2B2, CYP3A1, and CYP3A2 genes 24 h later (Konno et al., 2003). Therefore, in the present experiments, dose

Discussion

In the present study, we demonstrate for the first time the gender-related difference in the induction of CYP2B and CYP3A subfamily enzymes by Nic and Nif in rats and further indicate that these calcium channel antagonists show different specificities and sex dependencies in the induction of hepatic CYP2B and CYP3A subfamily enzymes. The present results from the RT-PCR and Western blot analyses indicate that Nic induces more efficiently CYP3A subfamily enzymes than CYP2B subfamily ones, whereas

Acknowledgements

This work was supported, in part, by a Grant-in-Aid from Scientific Research from the Japan Society for the Promotion of Science (M.D.).

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