A Sex-Specific Form of Cytochrome P-450 Catalyzing Propoxycoumarin O-Depropylation and Its Identity with Testosterone 6β-Hydroxylase in Untreated Rat Livers: Reconstitution of the Activity with Microsomal Lipids1

Abstract

Characteristics of a typical male-dominant reaction, dealkylation of n-propoxycoumarin, in rat livers were studied in relation to microsomal testosterone 6 beta-hydroxylase. The depropylation was more than 10-fold higher in the liver of male than female adult rats, but the sex-related difference was eliminated by neonatal castration. Hypophysectomy of adult male rats, which decreased the rates of male-specific P-450-male-dependent reactions, increased the depropylation of propoxycoumarin, while the rate was decreased by either intermittent injection or continuous infusion of human growth hormone to hypophysectomized rats. With regard to age-related difference, microsomal depropylation was detectable at neonate and reached a maximal level at 14 to 20 d of age, but was abruptly diminished only in female rats at puberty. These changes are in good agreement with those of testosterone 6 beta-hydroxylation and the content of a male-specific P-450(6)beta-1/PB-1. In reconstituted systems using extracted microsomal lipids, P-450(6)beta-1/PB-1 and P-450-male catalyzed the depropylation of propoxycoumarin. However, the microsomal depropylation was inhibited by antibodies which recognize P-450(6)beta-1/PB-1, but not P-450-male. These results indicate that microsomal depropylation of propoxycoumarin is catalyzed mainly by a male-specific P-450(6)beta-1/PB-1 in livers of untreated rats.

Isolation and Characterization of a New Major Intestinal CYP3A Form, CYP3A62, in the Rat

“…The reaction mixture for measurement of testosterone 6␤-hydroxylase activities consists of 50 g of protein of COS-1 microsomes expressing a CYP3A form, 100 mM potassium phosphate buffer (pH 7.4), 5 pmol of cytochrome b 5 , 0.1 unit (0.1 mmol of cytochrome c per minute) of NADPH-P450 reductase, and 5 g of sodium cholate in a final volume of 100 l. The reaction was started by the addition of NADPH (final concentration, 0.5 mM) and terminated by adding ethyl acetate after 40 min of incubation at 37°C. Testosterone hydroxylation was quantified by the method described previously (Yamazoe et al, 1988;Guo et al, 2000).…”

“…These CYP3A forms appear in a sex-dependent manner in rats. For example, CYP3A2 (Yamazoe et al, 1988;Cooper et al, 1993) and CYP3A18 (Nagata et al, 1996;Robertson et al, 1998) are male-specific forms, whereas CYP3A9 is a female-dominant form (Wang and Strobel, 1997;Robertson et al, 1998). The expression profiles in the intestinal tract, however, are obscure with all of the forms.…”

Monkey Hepatic Microsomal Alcohol Oxygenase: Purification and Characterization of a Cytochrome P450 Enzyme Catalyzing the Stereoselective Oxidation of 7.ALPHA.- and 7.BETA.-Hydroxy-.DELTA.8-tetrahydrocannabinol to 7-Oxo-.DELTA.8-tetrahydrocannabinol.

“…The testosterone 6b-hydroxylase activity of P450JM-E is considerablely higher than that of CYP3A8 (P450CMLc) purified from hepatic microsomes of cynomolgus monkey as reported by Ohmori et al 37) The reason may be attributable to the use of microsomal lipids instead of dilauroylphosphatidylcholine as the lipid in the reconstituted system of P450JM-E, because a number of studies have shown that the catalytic activities of CYP3A enzymes are very low in a reconstituted system using only dilauroylphosphatidylcholine as the lipid. 17,[39][40][41] P450JM-E showed comparable activity to CYP3A1, 40) CYP3A4, 41) and CYP3A11 11) for testosterone 6b-hydroxylase. In the reconstituted system of P450JM-E, the MALCO activity for 7b-OH-D 8 -THC was about 6-fold higher than that for 7a-OH-D 8 -THC.…”

Evidence from dwarf rats that growth hormone may not regulate the sexual differentiation of liver cytochrome P450 enzymes and steroid 5 alpha-reductase.

“…In normal rats, the levels of IIIA2 are high in immature rats; they remain high in mature male rats but decline abruptly and markedly after puberty in female rats (7,17,18,35). GH is thought to suppress the levels of IIIA2, and a continuous pattern of GH secretion exerts a greater suppressive effect than a pulsatile pattern of secretion (possibly because the low trough levels provide periodic relief from GH suppression) (11,18,20,36,37).…”

“…After weaning and throughout puberty, the levels of IIA1 decline in male rats to a greater extent than in female rats, and this decline is associated with a decline in testosterone 7a-hydroxylase activity (7,(12)(13)(14)(15)(16)(17). Conversely, the levels of IIIA2 decline markedly in female but not male rats, and this decline is associated with a dramatic postpubertal decrease in the rate of testosterone 2,8-, 6,3-, and 15/3-hydroxylation (7,(16)(17)(18).…”