Role of mouse CYP2E1 in the O-hydroxylation of p-nitrophenol: comparison of activities in hepatic microsomes from Cyp2e1(-/-) and wild-type mice.
Enzymatic activities are routinely used to identify the contribution of individual forms of cytochrome P450 in a particular biotransformation. p-Nitrophenol O-hydroxylation (PNPH) has been widely used as a measure of CYP2E1 catalytic activity. However, rat and human forms of CYP3A have also been shown to catalyze this activity. In mice, the contributions of CYP3A and CYP2E1 to PNPH activity are not known. Here we used hepatic microsomes from Cyp2e1(-/-) and wild-type mice to investigate the contributions of constitutively expressed and alcohol-induced murine CYP2E1 and CYP3A to PNPH activity. In liver microsomes from untreated mice, PNPH activity was much greater in wild-type mice compared with Cyp2e1(-/-) mice, suggesting a major role for CYP2E1 in catalyzing PNPH activity. Hepatic PNPH activities were not significantly different in microsomes from male and female mice, although the microsomes from females have dramatically higher levels of CYP3A. Treatment with a combination of ethanol and isopentanol resulted in induction of CYP3A proteins in wild-type and Cyp2e1(-/-) mice, as well as CYP2E1 protein in wild-type mice. The alcohol treatment increased PNPH activities in hepatic microsomes from wild-type mice but not from Cyp2e1(-/-) mice. Our findings suggest that in untreated and alcohol-treated mice, PNPH activity may be used as a specific probe for CYP2E1 and that constitutively expressed and alcohol-induced forms of mouse CYP3A have little to no role in catalyzing PNPH activity.
Wolf
Kristina K
KK
Veterans Administration Medical Center, White River Junction, VT 05009, USA.
Wood
Sheryl G
SG
Bement
Jenna L
JL
Sinclair
Peter R
PR
Wrighton
Steven A
SA
Jeffery
Elizabeth
E
Gonzalez
Frank J
FJ
Sinclair
Jacqueline F
JF
D000818:Animals; D001189:Aryl Hydrocarbon Hydroxylases / Q000096:biosynthesis / Q000378:metabolism; D019392:Cytochrome P-450 CYP2E1 / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism*; D051544:Cytochrome P-450 CYP3A; D004790:Enzyme Induction; D000431:Ethanol / Q000494:pharmacology; D005260:Female; D006900:Hydroxylation; D066298:In Vitro Techniques; D008297:Male; D051379:Mice; D018345:Mice, Knockout; D008862:Microsomes, Liver / Q000201:enzymology / Q000378:metabolism*; D009596:Nitrophenols / Q000378:metabolism*; D010089:Oxidoreductases, N-Demethylating / Q000096:biosynthesis / Q000378:metabolism; D000439:Pentanols / Q000494:pharmacology; D012737:Sex Factors
D009596:Nitrophenols; D000439:Pentanols; D000431:Ethanol; C029683:isopentyl alcohol; D019392:Cytochrome P-450 CYP2E1; D001189:Aryl Hydrocarbon Hydroxylases; D051544:Cytochrome P-450 CYP3A; D010089:Oxidoreductases, N-Demethylating; C024836:4-nitrophenol