Multiple mechanisms in indomethacin-induced impairment of hepatic cytochrome P450 enzymes in rats.
OBJECTIVE
Indomethacin impairs liver microsomal monooxygenase activities mediated by cytochrome P450 (CYP). We investigated the inhibition mechanism and the isoform selectivity in vitro and in vivo.


METHODS
In an in vitro study, liver microsomes from male Wistar rats were preincubated with indomethacin and a reduced nicotinamide adenine dinucleotide phosphate-generating system, followed by assay of monooxygenase activities indicative of several CYP isoforms. In an in vivo study, rats were intraperitoneally treated with indomethacin, followed by preparation of microsomes and the enzyme assays.


RESULTS
The preincubation of microsomes with indomethacin and reduced nicotinamide adenine dinucleotide phosphate decreased CYP3A2 activity but not any other isoforms. Kinetic analysis showed the mechanism-based inactivation of CYP3A2. The metabolism of [14C]indomethacin resulted in covalent binding to microsomal protein, which was diminished by inhibiting CYP3A enzyme. Administration of indomethacin caused impairment of not only CYP3A2 but also other CYP isoforms. Rats were protected from the impairment of the CYP enzymes except CYP3A2 by depleting macrophages and inhibiting inducible nitric oxide synthase.


CONCLUSIONS
Metabolism of indomethacin causes inactivation of CYP3A2, which is the result of the covalent binding of its metabolite, whereas partially selective in vivo impairment of CYP isoforms is suggested to be indirect inhibition by inflammatory mediators probably released from Kupffer cells.
Masubuchi
Yasuhiro
Y
Department of Biopharmaceutics, Graduate School of Pharmaceutical Sciences, Chiba University, Chiba, Japan.
Masuda
Emi
E
Horie
Toshiharu
T
D000818:Animals; D000894:Anti-Inflammatory Agents, Non-Steroidal / Q000494:pharmacology*; D001189:Aryl Hydrocarbon Hydroxylases; D002250:Carbon Radioisotopes; D051544:Cytochrome P-450 CYP3A; D003577:Cytochrome P-450 Enzyme System / Q000378:metabolism*; D004789:Enzyme Activation / Q000187:drug effects; D066298:In Vitro Techniques; D007213:Indomethacin / Q000494:pharmacology*; D018836:Inflammation Mediators / Q000378:metabolism; D007728:Kupffer Cells / Q000201:enzymology / Q000276:immunology; D008099:Liver / Q000201:enzymology* / Q000276:immunology; D008297:Male; D008565:Membrane Proteins; D008862:Microsomes, Liver / Q000201:enzymology; D009249:NADP / Q000378:metabolism; D009566:Nitrates / Q000097:blood; D009569:Nitric Oxide / Q000096:biosynthesis; D009573:Nitrites / Q000097:blood; D051381:Rats; D017208:Rats, Wistar; D039181:Steroid 16-alpha-Hydroxylase; D013250:Steroid Hydroxylases / Q000378:metabolism*
D000894:Anti-Inflammatory Agents, Non-Steroidal; D002250:Carbon Radioisotopes; D018836:Inflammation Mediators; D008565:Membrane Proteins; D009566:Nitrates; D009573:Nitrites; D009569:Nitric Oxide; D009249:NADP; D003577:Cytochrome P-450 Enzyme System; D013250:Steroid Hydroxylases; D001189:Aryl Hydrocarbon Hydroxylases; C465260:Cyp3a2 protein, rat; D051544:Cytochrome P-450 CYP3A; D039181:Steroid 16-alpha-Hydroxylase; C022657:steroid hormone 6-beta-hydroxylase; D007213:Indomethacin