Differential expression of microsomal epoxide hydrolase gene by azole heterocycles in rats

Abstract

The effects of heterocycles including imidazole (IM), 1,2,4-triazole (TR) and thiazole (TH) on the expression of microsomal epoxide hydrolase (mEH) gene were examined in rats (200 mg kg body weight day , i.p.). Hepatic microsomes prepared from rats treated with IM for 3 days failed to exhibit an increase in mEH protein level whereas TR treatment resulted in an approximately 2- to 3- fold levation in hepatic mEH levels relative to control, as assessed by both SDS-PAGE and immunoblot analyses. In contrast, thiazole-induced hepatic microsomes resulted in a substantial increase in mEH levels (i.e.∼5-fold). Slot and northern blot analyses, probed with an mEH cDNA, showed that the hepatic mEH mRNA levels in the animals treated with IM for 3 days were marginally increased by ∼2-fold, as compared with untreated animals, whereas TR caused an ∼8-fold increase in hepatic mEH mRNA levels after three consecutive daily treatments. TH treatment resulted in an ∼22-fold increase in the mEH mRNA levels, demonstrating that TH is the most efficacious among these three azole heterocycles. Because TH was the most effective in increasing hepatic mEH protein and mRNA levels, the agent was chosen for further evaluation. Time course of mEH gene expression at early times after a single treatment with TH was determined and compared with that caused by pyrazine (PZ), a strong mEH inducer. Hepatic mEH mRNA levels were increased ∼1-, 3-, 20- and 16-fold at 3, 6, 12 and 24 hr, respectively, following TH treatment, relative to control, whereas mEH mRNA levels were elevated ∼1-, 1-, 22- and 18-fold, respectively, at the same time points after PZ treatment, as monitored by slot RNA hybridization analyses. Northern blot analyses using either total RNA or poly(A) + RNA fractions exhibited comparable time courses in increasing mEH mRNA levels after TH or PZ treatment with maximal mRNA increases being noted at 12 hr post treatment. Although neither IM or TR failed to affect renal mEH gene expression to a notable extent, TH treatment caused 6- to 8-fold increases in kidney mEH mRNA levels, with a 2-fold increase in mEH protein detected. These results demonstrated that the azole heterocyclic compounds IM, TR and TH differentially induce mEH with TH as the most efficacious azole; and that the changes in mEH levels are primarily associated with increases in mRNA levels.