TRANSCRIPTIONAL REGULATION OF RAT MICROSOMAL EPOXIDE HYDROLASE GENE BY IMIDAZOLE ANTIMYCOTIC AGENTS

The effects of imidazole antifungal agents, including ketoconazole, clotrimazole, miconazole, and econazole, on the expression and regulation of microsomal epoxide hydrolase (mEH) were examined in rat hepatic tissue (doses of agents, 150 mg/kg of body weight/day, orally). Immunoblot analyses revealed that administration of either ketoconazole or clotrimazole caused a approximately 4-5-fold increase in mEH levels, whereas either miconazole or econazole resulted in a approximately 7-fold increase in mEH at day 3 after treatment. RNA hybridization analyses, probed with a 1.3-kilobase mEH cDNA, revealed that administration of these imidazole antifungal agents caused substantial elevation of hepatic mEH mRNA in total RNA. Hepatic mEH mRNA levels in total RNA were elevated approximately 11-, 15-, and 18-fold at 12, 24, and 72 hr, respectively, after ketoconazole treatment, whereas mEH mRNA levels were increased approximately 14-, 19-, and 22-fold, respectively, relative to control, at the same time points after clotrimazole treatment. The rate of increase of mEH mRNA caused by miconazole was more rapid than the rates observed for the other agents examined, with a maximal increase in mRNA being noted at 12 hr after treatment. The degree of mEH mRNA increase after 3 consecutive days of miconazole treatment was appreciably less than that observed at 12 hr after a single treatment. Econazole caused a maximal increase at 24 hr and subsequent decline in mEH mRNA levels after 3 consecutive days of treatment. Elevation of mEH mRNA levels by these antimycotic agents was confirmed in poly(A)+ RNA, as assessed by both Northern and slot blot hybridization analyses. Nuclear run-on analyses revealed that administration of ketoconazole, clotrimazole, or miconazole stimulated the rate of mEH gene transcription at 12 hr after treatment by 11-, 8.5-, and 9-fold, respectively, compared with control, whereas econazole resulted in a 4-fold increase in the rate of mEH gene transcription at the same time point. The transcription rates of mEH mRNA at 24 hr were significantly less than those observed at 12 hr after a single treatment with either ketoconazole, miconazole, or econazole, resulting in 6.5-, 2.5-, and 2-fold increases, respectively, relative to control. Clotrimazole, however, maintained the activated mEH transcription rate at 24 hr after treatment, exhibiting a 11-fold increase, compared with control. These results provide evidence that the imidazole antimycotic agents induce mEH and that the mEH induction involves large increases in mRNA, with transcriptional activation.