IN-VITRO METABOLISM OF 4-ANDROSTENE-3,17-DIONE BY HEPATIC MICROSOMES FROM THE RAINBOW TROUT (SALMO GAIRDNERII): EFFECTS OF HYPOPHYSECTOMY AND OESTRADIOL-17β

1981 ◽  
Vol 90 (1) ◽  
pp. 103-112 ◽  
Author(s):  
TIIU HANSSON ◽  
JAN-ÅKE GUSTAFSSON
Keyword(s):  
Rainbow Trout ◽  
Liver Microsomes ◽  
The Other ◽  
Sham Operation ◽  
Hydroxylase Activity ◽  
Oxidoreductase Activity ◽  
Hepatic Microsomes ◽  
Cytochrome P 450 ◽  
Reductive Metabolism

The metabolism of 4-androstene-3,17-dione by liver microsomes from juvenile rainbow trout, Salmo gairdnerii, was studied in vitro. Hypophysectomy of the fish significantly increased mean hepatic 17-hydroxysteroid oxidoreductase activity when compared with that from sham-operated fish but none of the other enzyme activities investigated were affected. Administration of oestradiol-17β resulted in a significant decrease in mean hepatic 6β-hydroxylase activity and total cytochrome P-450 content but had no effect on the 16-hydroxylation or on the reductive metabolism of androstenedione. The effect of oestradiol-17β on hepatic 6β-hydroxylase activity was as pronounced after hypophysectomy as after sham-operation indicating that these effects of oestradiol-17β are mainly direct and independent of the pituitary gland. The results indicate that hypophysial hormone(s) as well as oestradiol-17β play a role in the regulation of hepatic steroid metabolism in trout.

The characteristics of the microsomal hydroxylation of tolbutamide

1991 ◽  
Vol 69 (3) ◽  
pp. 400-405 ◽  
Author(s):  
Pierre M. Bélanger ◽  
Serge St-Hilaire
Keyword(s):  
Carbon Monoxide ◽  
Liver Microsomes ◽  
Hepatic Metabolism ◽  
Species Variation ◽  
Rat Liver Microsomes ◽  
Hydroxylase Activity ◽  
Microsomal Metabolism ◽  
Microsomal Hydroxylation ◽  
Cytochrome P 450

The in vitro metabolism of tolbutamide to the hydroxymethyl derivative was studied using hepatic microsomal homogenates. The hydroxymethyl metabolite was quantitated by HPLC. The hepatic microsomal hydroxylase was completely inhibited by carbon monoxide and was NADPH dependent. Metyrapone, α-naphthoflavone, phenelzine, mercuric chloride, and nitrogen significantly inhibited the reaction indicating the involvement of the cytochrome P-450 monooxygenase. Species variation showed that the order of hepatic microsomal activity was rat > rabbit >> guinea pig >> mouse and hamster. The reaction increased with time up to 40 min and followed Michaelis–Menten kinetics in rat liver microsomes with apparent Km and Vmax values of 224.4 μM and 359.9 pmol∙mg−1∙min−1, respectively. The reaction was induced by phenobarbital but was depressed after pretreatment with 3-methylcholanthrene and isosafrole. However, expression of the hydroxylase activity per nanomoles of cytochrome P-450 showed that the activity was much higher in liver microsomes of isosafrole pretreated rats. These results indicate the involvement of different isozymes of cytochrome P-450 in the microsomal hydroxylation of tolbutamide.Key words: tolbutamide metabolism, tolbutamide hydroxylation, microsomal hydroxylation, microsomal metabolism of tolbutamide, hepatic metabolism of tolbutamide.

ANDROGENIC REGULATION OF HEPATIC METABOLISM OF 4-ANDROSTENE-3,17-DIONE IN THE RAINBOW TROUT, SALMO GAIRDNERII

1982 ◽  
Vol 92 (3) ◽  
pp. 409-417 ◽  
Author(s):  
TIIU HANSSON
Keyword(s):  
Rainbow Trout ◽  
Reductase Activity ◽  
Juvenile Fish ◽  
Liver Microsomes ◽  
Hepatic Metabolism ◽  
Suppressive Effect ◽  
Hydroxylase Activity ◽  
Androgen Treatment ◽  
Androgenic Regulation

The metabolism of 4-androstene-3,17-dione by liver microsomes from the juvenile rainbow trout, Salmo gairdnerii, was studied in vitro. Administration of testosterone, 11-oxotestosterone, dihydrotestosterone or 17α-methyltestosterone to juvenile fish significantly increased mean hepatic 17-hydroxysteroid oxidoreductase (17-HSOR) activity. Androgen treatment tended to increase the total cytochrome P-450 content significantly in liver microsomes from 11-oxotestosterone-treated fish. On the other hand, androgen treatment decreased mean hepatic 6β-hydroxylase activity but did not affect 16-hydroxylase or 5α-reductase activity. The suppressive effect of simultaneous administration of testosterone and oestradiol-17β on 6β-hydroxylase activity was more pronounced than when these steroids were administered separately. Furthermore, oestradiol-17β diminished the effect of testosterone on 17-HSOR activity. Testosterone treatment of hypophysectomized fish caused a significant increase in 17-HSOR activity when compared with activity in untreated hypophysectomized fish, indicating that this effect of testosterone is mainly direct and independent of the pituitary gland. The results indicate that androgens as well as oestradiol-17β play a role in the control of sexual differences in hepatic steroid metabolism in trout.

A comparison of the in vitro metabolism of biphenyl and 4-chlorobiphenyl by rat liver microsomes

1978 ◽  
Vol 56 (10) ◽  
pp. 993-997 ◽  
Author(s):  
C. Wyndham ◽  
S. Safe
Keyword(s):  
Polychlorinated Biphenyl ◽  
Liver Microsomes ◽  
Low Molecular Weight ◽  
Rat Liver Microsomes ◽  
Hepatic Microsomes ◽  
Comparative Metabolism ◽  
Metabolic Products ◽  
Active Substrate ◽  
And Control

The comparative metabolism of the hydrocarbons, biphenyl and 4-chlorobiphenyl, was investigated using two different preparations of rat hepatic microsomes. The assay was designed to account for all the metabolic products which included the ether soluble lipophilic metabolites, low molecular weight conjugates, and macromolecular adducts, and to determine the effects of induction with Aroclor 1254 and 1248, two commercial polychlorinated biphenyl (PCB) preparations. 4-Chlorobiphenyl was the more metabolically active substrate with the induced and control enzymes. In most metabolic fractions biphenyl was less inducible by the PCB's, with the exception of the 2-biphenylol metabolite which was induced ca. 18-fold. Preincubation of the microsomes with carcinogens did not enhance biphenyl 2-hydroxylation. Instead, a general inhibition of metabolic activity was observed for both biphenyl and 4-chlorobiphenyl substrates. Preincubation with phenobarbitone, a noncarcinogen, did not change the microsome-mediated metabolism of biphenyl or 4-chlorobiphenyl. The substitution of a single halogen atom on the biphenyl nucleus altered both the reactivity and pattern of metabolites for these substrates.

scholarly journals Monoclonal antibodies against human cytochrome P-450 recognizing different pregnenolone 16α-carbonitrile-inducible rat cytochromes P-450

1987 ◽  
Vol 248 (1) ◽  
pp. 301-304 ◽  
Author(s):  
T S Barnes ◽  
P M Shaw ◽  
M D Burke ◽  
W T Melvin
Keyword(s):  
Monoclonal Antibodies ◽  
Human Liver ◽  
Liver Microsomes ◽  
Human Liver Microsomes ◽  
The Other ◽  
Male Rat ◽  
Rat Liver Microsomes ◽  
Human Cytochrome ◽  
Male Specific ◽  
Cytochrome P 450

Six murine monoclonal antibodies against human hepatic cytochrome P-450 have been raised, using human liver microsomes (microsomal fractions) or semi-purified human cytochrome P-450 as immunogen. All six antibodies recognized the same highly purified of human liver cytochrome P-450 of molecular mass 53 kDa and gave rise to a single band at 53 kDa on immunoblots of human liver microsomes from 11 individuals. The antibodies also recognized proteins at 52 kDa and 54 kDa on immunoblots of control and induced male-rat liver microsomes, showing four different banding patterns. Antibodies HL4 and HP16 recognized a 52 kDa protein that was only weakly expressed in untreated rats and which was strongly induced by pregnenolone 16 alpha-carbonitrile (PCN) but not by phenobarbitone (PB), 3-methylcholanthrene (3MC), isosafrole (ISF), Aroclor 1254 (ARO), clofibrate or imidazole. HP10 and HL5 recognized a constitutive 52 kDa protein that was weakly induced by PCN but not by the other agents and was suppressed by 3MC and ARO. HP3 recognized a 54 kDa protein that was undetectable in control rats but was strongly induced by PB, PCN, ISF and ARO. HL3 appeared to recognize a combination of the proteins recognized by the other antibodies plus a 54 kDa protein that was weakly expressed in control rats. The constitutive proteins recognized were male-specific.

scholarly journals Multiple steroid-binding orientations: alteration of regiospecificity of dehydroepiandrosterone 2- and 7-hydroxylase activities of cytochrome P-450 2a-5 by mutation of residue 209

1995 ◽  
Vol 306 (1) ◽  
pp. 29-33 ◽  
Author(s):  
M Iwasaki ◽  
D G Davis ◽  
T A Darden ◽  
L G Pedersen ◽  
M Negishi
Keyword(s):  
Dynamic Equilibrium ◽  
The Other ◽  
Axial Position ◽  
Hydroxylase Activity ◽  
Steroid Molecule ◽  
Multiple Binding ◽  
Critical Residues ◽  
Steroid Binding ◽  
Binding Orientations ◽  
Cytochrome P 450

The mutation of Ala-117 to Val conferred dehydroepiandrosterone (DHEA) hydroxylase activity on cytochrome P-450 2a-4, with the production of both 2 alpha- and 7 alpha-hydroxyDHEA at similar rates. P-450 2a-5 which has Val at position 117, acquired high DHEA hydroxylase activity by mutation of Phe-209. Mutant F209L of P-450 2a-5 exhibited strong regiospecificity at the 2-position of the DHEA molecule with the production of 2 alpha-hydroxy DHEA as the major metabolite. On the other hand, mutant F209V of P-450 2a-5 showed the 7-position to be the major hydroxylation site, 7 beta-hydroxyDHEA and 7 alpha-OHDHEA being produced. Therefore the regiospecificity of DHEA hydroxylase activity of P-450 2a-5 is altered between the 2- and 7-position depending on the amino acid at position 209. Modelling of the DHEA molecule in the pocket of bacterial P-450cam showed that the steroid can be accommodated in at least two orientations for which the 2- or 7- position is near the sixth axial position of the haem. Moreover, these two orientations, which are of similar energy, can be interconverted by a 180 degrees rotation of the steroid molecule around its long axis. These results support the hypothesis that the steroid molecule in the pocket is in dynamic equilibrium with multiple binding orientations and that the equilibrium is apparently determined by a few critical residues including those at positions 117 and 209.

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