Differential expression and activities of cytochrome P450 3A in the rat brain microsomes and mitochondria.

Autor: Alshammari N; Department of Biomedical and Pharmaceutical Sciences, School of Pharmacy, Chapman University, Irvine, California, USA., Chandrashekar DV; Department of Biomedical and Pharmaceutical Sciences, School of Pharmacy, Chapman University, Irvine, California, USA., Rashid M; Department of Biomedical and Pharmaceutical Sciences, School of Pharmacy, Chapman University, Irvine, California, USA., Mehvar R; Department of Biomedical and Pharmaceutical Sciences, School of Pharmacy, Chapman University, Irvine, California, USA.
Jazyk: angličtina
Zdroj: Fundamental & clinical pharmacology [Fundam Clin Pharmacol] 2023 Apr; Vol. 37 (2), pp. 359-368. Date of Electronic Publication: 2022 Nov 17.
DOI: 10.1111/fcp.12848
Abstrakt: Midazolam (MDZ), a benzodiazepine derivative, is metabolized to 1'- and 4-hydroxylated metabolites (1'-OH-MDZ and 4-OH-MDZ, respectively) by cytochrome P450 3A (CYP3A). The purpose of this study was to investigate the CYP3A-mediated hydroxylation of MDZ in the rat brain mitochondria (MT). Brain microsomes (MC) and MT fractions were prepared from rats (n = 8) using differential and density gradient centrifugations, and the purity of the fractions was evaluated using VDAC1 and calreticulin as markers of MT and MC, respectively. The formation rates of 1'-OH-MDZ and 4-OH-MDZ in the rat brain MC and MT samples were determined using an LC-MS/MS method after validation. Subsequently, Michaelis-Menten kinetics of 1'- and 4-hydroxylation of MDZ were estimated. Western blot (WB) analysis was used to determine the protein expression of CYP3A in the rat brain MC and MT. The MC fractions had 5.93% ± 3.01% mitochondrial impurity, and the MT fractions had 19.3% ± 7.8% microsomal impurity (mean ± SD). The maximum velocity (V max ) values of the formation of the hydroxylated metabolites in the brain MT were 2.4-9-fold higher than those in MC. Further, the V max values of 4-OH-MDZ in both MC and MT fractions were substantially higher than those of 1'-OH-MDZ. The WB analysis showed that the intensity of the CYP3A immunoreactive band in MT was more than twofold higher than that in MC. It is concluded that compared with MC, rat brain MT contains substantial CYP3A, which may affect the pharmacology or toxicology of centrally acting xenobiotic and endogenous substrates of this enzyme.
(© 2022 The Authors. Fundamental & Clinical Pharmacology published by John Wiley & Sons Ltd on behalf of Société Française de Pharmacologie et de Thérapeutique.)
Databáze: MEDLINE
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