Effect of cytochrome P450 induction on the metabolism and toxicity of ochratoxin A

Abstract

The role of cytochrome P-450 in me stimulation of lipid peroxidation induccd by thc mycotoxin ochratoxin A (OTA) has been investigated. Purified cytochrome P-450 (liB I) could effectively replace EDTA in stimulating lipid pefOxidation in a reconstituted system l'Onsisling of phospholipid vesicles, NADPH-cYlochrome P-450rcduc!ase, FeJ+, EDTA and NADPll, suggesting that it could mediate the transfer of electrons from NADPH to foc3,. Microsomcs isolated from liven of cobalt protoporphyrin IX-treated rllts (in which cytochrome P-450 Wll" depletedj underwent OTA-dependent lipid peroxidation much more slowly tlmn control microsomes.The role of cytochrome P-450 in OTA met.1bolism was also investigated. To dClermine which cytochrome P·450 isoforms are involved in the metabolism ofOTA, we used different cytochrome P-450 inducers to induce the major isofonns of cytochrome 1'-450 in lhe rm liver.Microsomes from these livers were used to investigate their effect on OTA metabolism.Pretreatment ofmts with pregnenolone -16c(·carbonitrile (peN), phenobarbit.ll (PB), 3-ll1cthyl cholanthrene (3MC), and isosafrole (ISF) greatly induced 4(R)-4-0H-OTA fonmllion; 4(5)-4-011-OTA fonnation was also induced after pretreatment with PB, PCN, 3MC and ISr:. INII pretreaunem primarily induced me 4(5) isomer formation, The fonnation of the 4(R) .md 4 (5) isomeri showed significant differences with respect to pH optima, effect of antioKidants ami iron chelators, The 4(R) isomer fonnation showed a pH optimum of 6.0 using microsornes from rJIs treated with 3MC and ISF, and 6.5 using mierosomes from rats treated with PI] :tnd peN and Wi!S not inhibited by antioxidants or iron chelators. In contmst, both the 4(5) isomer fonmllion and lipid pcroxidation showed a pH optimum of 7.0 -7,5 and both activities were highly sensitive to inhibition by antioxidants and iron ehelators. Lipid peroxides were no: involved in the 4(5) isomer fonnation since addition of linoleic acid hydroperoxidc to microsomcs did not give rise 10 the 4(5) isomer, Cytochrome P-450 appeared to be essenlial since other hemoprolcins such as horseradish peroxidase and hemoglobin were ineffective in metabolizing OTA. Microsomes from rats pretreated with Co-protoporphyrin IX resulted in no metabolil,rn of ochratoxin A. 7-Ethoxy-and 7-pentoxyresorufin assays showed specificity towards cytochromes P-450 induced by 3MC (fA I/lA2) and PB (IIBI) respectively. Also, metyrapone (inhibitor of cytochrome P·450 IIBt) preferentially inhibited OTA mel1bolism by microsomes from rats treated with PB, and I(-naphthoflavone (inhibitor of cytochrome P-450 IAI/lA2) preferentially inhibited OTA metabolism by microwmes from 3MC and ISF treated rats_ Monoclonal antibodies (MAbs) 1-7-1 (against P-450 lA 1/lA2) and 2-66-3 (against P-450 liB I) showed preferential inhibition ofOTA metabolism by microsomes from 3MC and PB treated rats respectively.Excretion of renal enzymes in urine is a sensitive non-invasive index of renal dam:lge.Therefore. we examined the effect of cytochrome P-450 induction on the...

Phenobarbital increases DNA adduct and metabolites formed by ochratoxin A: Role of CYP 2C9 and microsomal glutathione-S-transferase

“…Their formation was dependent on the type of cells and the PB pretreatment. In this study, we confirm the increase of 4-[R]-OH-OTA and 10-OH-OTA after PB pretreatment observed by others [Størmer and Pederson, 1980;Hietanen et al, 1986Hietanen et al, , 1991Omar et al, 1996]. Little is known about the genotoxicity of these metabolites.…”

“…Before this investigation, the metabolism of OTA has been studied both in vitro and in vivo mostly in liver (for a review, see Størmer, 1992) but also in kidney [Hietanen et al, 1986;Pfohl-Leszkowicz et al, 1993b]. The main metabolites formed were hydroxylated compounds: 4-[R]-and 4-[S]-hydroxy-OTA (4-[R]-and 4-[S]-OH-OTA) in rats and also10-hydroxy-OTA (10-OH-OTA) in rabbits [Størmer et al, 1983], which could be formed by CYP 1A1/2, 2B1, 3A1/2, and also by peroxydasic pathways in bronchial and kidney cells [Ueno,1985;Størmer, 1992;Grosse et al, 1995;Omar et al, 1996;Grosse et al, 1997;Pinelli et al, 1999]. The formation of these OTA derivatives was induced by phenobarbital (PB) [Størmer and Pederson, 1980;Hietanen et al, 1986Hietanen et al, , 1991Omar et al, 1996].…”

Ochratoxin A levels in the plasma of healthy blood donors from Valencia and estimation of exposure degree: comparison with previous national Spanish data

“…The IARC has classified OTA as a "possible human carcinogen (Group 2B)" based on sufficient evidence for carcinogenicity in experimental animal studies and inadequate evidence in humans (IARC, 1993). The toxic-kinetic parameters of OTA have not been well established in humans, but OTA showed high availability after oral administration in some animal studies (Fuchs and Hult, 1992;Omar et al, 1996).…”

“…OTA produces an inhibition of protein synthesis and lipid peroxidation by oxidative processes (Gautier et al, 2001;Omar et al, 1996).…”

Ochratoxin a: Its Cancer Risk and Potential for Exposure

“…Results from studies of P-450 induction in rat liver microsomes showed that the P-450 isoforms IA1/IA2, IIB1, and IIIA1/IIIA2 are involved in the formation of the (4R)-4-hydroxyochratoxin-A metabolite of OA and, to a lesser extent, (4S)-4-hydroxyochratoxin-A (Omar et al, 1996). Results also indicated that P-450IIB1-dependent mixed-function oxidation to OA leads to its detoxification (Omar et al, 1996).…”