Nifedipine lowers cocaine-induced brain and liver enzyme activity and cocaine urinary excretion in rats.

The aim of this study was to see how nifedipine counters the effects of cocaine on hepatic and brain enzymatic activity in rats and whether it affects urinary excretion of cocaine. Male Wistar rats were divided in four groups of six: control, nifedipine group (5 mg kg-1i.p. a day for five days); cocaine group (15 mg kg-1i.p. a day for five days), and the nifedipine+cocaine group. Twenty-four hours after the last administration, we measured neuronal nitric oxide synthase (nNOS) activity in the brain and cytochrome P450 quantity, ethylmorphine-N-demethylase, and anilinehydroxylase activity in the liver. Urine samples were collected 24 h after the last cocaine and cocaine+nifedipine administration. Urinary cocaine concentration was determined using the GC/MS method.Cocaine administration increased brain nNOS activity by 55 % (p<0.05) in respect to control, which indicates the development of tolerance and dependence. In the combination group, nifedipine decreased the nNOS activity in respect to the cocaine-only group.In the liver, cocaine significantly decreased and nifedipine significantly increased cytochrome P450, ethylmorphine-N-demethylase, and anilinehydroxylase in respect to control. In combination, nifedipine successfully countered cocaine effects on these enzymes.Urine cocaine excretion in the cocaine+nifedipine group significantly dropped (by 35 %) compared to the cocaine-only group.Our results have confirmed the effects of nifedipine against cocaine tolerance and development of dependence, most likely due to metabolic interactions between them.

Estradiol, testosterone, dehydroepiandrosterone and androstenedione: novel derivatives and enantiomers. Interactions with rat liver microsomal cytochrome P450 and antioxidant/radical scavenger activities in vitro.

Interactions of 27 steroids, among them 17 derivatives such as ethers, sulfates and amidosulfonates derived from 17 beta- and 17 alpha-estradiol, from testosterone and alpha- and beta-dihydrotesosterone and from dehydroepiandrosterone with rat liver microsomal cytochromes P450 (P450) were investigated in vitro by assessing binding to P450 and effects on P450 mediated monooxygenase functions as measured by different model reactions: ethoxyresorufin O-deethylation (EROD), ethoxycoumarin O-deethylation (ECOD) and ethylmorphine N-demethylation (EMND). With the exception of 17 alpha-estradiol-3-dimethylamidosulfonate, estrone, its -3-methylether and -3-amidosulfonate and testosterone, all other steroids displayed type I or reverse type I binding to P450. All steroids inhibited EROD activity in micromolar concentrations. An additional strong inhibition of ECOD and EMND activities was only demonstrated for the androgens and progestins. Estriol, estrone and mestranol displayed less inhibitory actions on the model reactions than estradiol. No major differences in comparison to the parent compounds were noted with the other derivatives. The only exceptions were 17 beta-(8,9-dehydro-14 alpha,15 alpha-methylene)estradiol, which displayed stronger effects than estradiol, and dehydroepiandrosterone-3-sulfate, which was less effective than dehydroepiandrosterone. Possible antioxidant properties of the steroids were examined by the stimulated lipid peroxidation (LPO), H2O2 production, and lucigenin (LC) and luminol (LM) amplified chemiluminescence (CL) using rat liver microsomes. Additionally, the influence on rat whole blood chemiluminescence (WB-CL) was assessed. All the estrogens, but not their methylethers and amidosulfonates inhibited LPO in micromolar concentrations. The effects on the other oxidase model reactions or on WB-CL were less distinct. Only ethinylestradiol and 17 beta-(8,9-dehydro-14 alpha,15 alpha-methylene)estradiol displayed a strong inhibitory action on all model reactions. With the exception of dehydroepiandrosterone-3-sulfate, which in general had only weak effects, the androgen and progestin derivatives, in contrast, strongly decreased H2O2 formation and LM- and LC-CL, but were mostly ineffective on LPO and WB-CL.

Transplantation of fetal liver tissue suspension into the spleens of adult syngenic rats: effects of different cytotoxins on cytochrome P450 mediated monooxygenase functions and on oxidative state.

Syngenic fetal liver tissue suspensions were transplanted into the spleens of adult male Fisher 344 inbred rats. Four months after surgery, transplant recipients and age matched control rats were treated with different cytotoxins (allyl alcohol [AAL], bromobenzene [BBZ], carbon tetrachloride [CCl4], or thioacetamide [TAA]) or the respective solvents 24 or 48 hours before sacrifice. Effects of the cytotoxins on P450 mediated monooxygenase functions in liver and spleen 9,000 g supernatants were assessed by measuring the model reactions ethoxyresorufin O-deethylation (EROD), ethoxycoumarin O-deethylation (ECOD), pentoxyresorufin O-depentylation (PROD), and ethylmorphine N-demethylation (EMND). Additionally, the influence on the oxidative state was investigated by assessing the liver and spleen tissue content of lipid peroxidation (LPO) products and of reduced and oxidized glutathione (GSH;GSSG). The livers of both solvent treated transplant recipients and control rats displayed regular EROD, ECOD, PROD and EMND activities. After AAL treatment EROD and EMND activities within the livers were not affected, but ECOD and PROD activities were increased. BBZ administration caused a decrease in EROD and EMND activities, ECOD activity remained unaffected, and PROD activity was even increased. CCl4 and TAA administration caused a strong reduction in the activity of all four model reactions. Spleens of control rats displayed almost no P450 mediated monooxygenase functions, independent whether the rats had been treated with the cytotoxins or not. In the transplant containing spleens, however, significant EROD and ECOD, but hardly any PROD or EMND activities were seen. After AAL administration EROD activity was not affected in the transplant containing spleens, but ECOD activity was increased. BBZ treatment led to a decrease in EROD and an elevation in ECOD activity. CCl4 and TAA strongly reduced the activity of both of these model reactions. The tissue content of LPO products within livers and transplant containing spleens was significantly increased after BBZ and CCl4 treatment. An elevation in LPO products was also seen in the spleens of the control rats due to CCl4 administration. Tissue GSH and GSSG content in both livers and transplant containing spleens were strongly reduced after BBZ treatment. After CCl4 administration only a significant decrease in liver GSSG contents was seen. TAA treatment caused a reduction in the GSH and GSSG content in the spleens of both transplant recipients and control rats, but not in the livers. From these results it can be concluded, that the effects of cytotoxins like AAL, BBZ, CCl4 or TAA on P450 dependent monooxygenase functions and on oxidative state are exerted in the ectopic intrasplenic liver cell transplants in a similar way as in normal orthotopic liver.

Effect of vitamin E on lipid peroxidation and liver monooxigenase activity in experimental influenza virus infection.

Influenza virus infection was associated with development of oxidative stress in liver of mice, viz. increase in amount of lipid peroxidation products, decrease in cytochrome P-450 and NADP. H-cytochrome c-reductase activity, and inhibition of liver monooxygenases (aniline hydroxylase, ethylmorphine-N-demethylase, amidopyrine-N-demethylase and analgin-N-demethylase). These effects were most pronounced on the 7th day after virus inoculation as compared to the 5th one. Supplementation of mice with vitamin E before virus inoculation leads to liver protection against oxidative stress and toxicosis. A marked decrease of lipid peroxidation products and an increase of cytochrome P-450 and activities of monooxygenases was established. The stabilizing effect of vitamin E was dose-dependent and was most pronounced on the 5th day after virus inoculation as compared to the 7th one.

Changes in rat liver monooxygenase activities after administration of atenolol, nifedipine and diltiazem alone and in combination.

The effects of the Ca2+ antagonists nifedipine (NF) and diltiazem (DL) and of the cardioselective beta 1-adrenergic blocking agent atenolol (AT) on the hexobarbital (HB) sleeping time and on the activity of some liver drug-metabolizing enzyme systems in male Wistar rats were studied. Two hours after single oral administration, atenolol (150 mg/kg) did not change hexobarbital sleeping time, while nifedipine (50 mg/kg) and diltiazem (30 mg/kg) prolonged it by 171.2 and 99.6%, respectively. Coadministration of atenolol with diltiazem or with nifedipine significantly prolonged hexobarbital sleep by 205 and 283%, respectively. Administered alone, atenolol decreased the ethylmorphine-N-demethylase (EMND) activity, but the amidopyrine-N-demethylase (APND) activity was not changed in any of the treated groups. Atenolol and nifedipine significantly increased aniline-4-hydroxylase (AH) activity and this effect was also observed with the combinations AT + NF and AT + DL. The NADPH cytochrome P-450 reductase activity was significantly decreased by nifedipine and diltiazem. Only nifedipine increased the total content of cytochrome P-450 (by 23.8%). Atenolol and diltiazem tended to increase the content of cytochrome b5 which was increased by nifedipine by 97.6%. The same effect was observed with the combinations AT + NF and AT + DL. The results suggest that NF, AT + NF and AT + DL produced the manifested changes in hepatic oxidative metabolism. The decreased EMND activity by atenolol, however, and the prolongation of hexobarbital sleeping time by nifedipine, diltiazem and their coadministration with atenolol did not correlate with enhanced microsomal P-450 and b5 content.

Transplantation of fetal liver tissue suspension into the spleens of adult syngenic rats: effects of various mitogens and cytotoxins on cytochrome P450 (P450) isoforms expression and on P450 mediated monooxygenase functions.

Syngenic fetal liver tissue suspensions were transplanted into the spleens of adult male Fisher 344 inbred rats. Four months after surgery, transplant recipients and age matched control rats were treated with various mitogens (fluorene [FEN], fluorenone [FON] and 2-acetylaminofluorene [AAF]) or cytotoxins (allyl alcohol [AAL], bromobenzene [BBZ] and carbon tetrachloride [CCl4]) or the respective solvents 24 or 48 hours before sacrifice. The expression of three cytochrome P450 (P450) isoforms, 1A1, 2B1 and 3A2, within spleens and livers was assessed by immunohistochemistry and P450 mediated monooxygenase functions in spleen and liver 9000 g supernatants by the model reactions ethoxyresorufin O-deethylation (EROD), ethoxycoumarin O-deethylation (ECOD), and ethylmorphine N-demethylation (EMND). The orthotopic livers of both solvent treated transplant recipients and control rats displayed only in few lobules a slight P450 1A1, but in all lobules a moderate P450 2B1 and 3A2 expression, all mainly located in the hepatocytes around the central veins. Correspondingly, regular EROD, ECOD and EMND activities were observed. Each of the three mitogens increased the P450 1A1 expression in the hepatocytes of the perivenous zones of the liver lobules. FON administration caused an additional P450 1A1 immunostaining in the periportal areas, and AAF treatment a P450 1A1 expression in bile duct epithelia. Also the staining for P450 2B1 and 3A2 in the hepatocytes of the perivenous and intermediate zones of the liver lobules was intensified after treatment with any of the mitogens. The three model reactions were significantly increased within the livers after FEN and FON administration, whereas after AAF treatment only ECOD was enhanced, EROD remained unaffected and EMND was decreased. The cytotoxin AAL caused small lesions and fatty degeneration of hepatocytes only in some periportal areas. BBZ only produced a perivenous necrosis of single cells, whereas CCl4 caused complete necrosis of the centrilobular parenchyma. Immunohistochemically, AAL administration led to an increase in the P450 2B1 expression in the perivenous hepatocytes, whereas the staining for P450 1A1 was not affected and that for P450 3A2 was even decreased in the periportal areas. Due to AAL treatment EROD and EMND activities were not affected and ECOD activity was increased. BBZ administration caused a P450 1A1 expression in the periportal hepatocytes but a decrease in this staining of the perivenous cells. The number of hepatocytes positively stained for P450 2B1 and 3A2 in the perivenous and intermediate zones was diminished in comparison to the livers of solvent treated rats. After BBZ treatment, EROD and EMND activities were decreased, ECOD activity was not affected. CCl4 administration caused a strong reduction in the expression of all three P450 isoforms and in the activity of all three model reactions. Spleens of control rats displayed almost no P450 isoforms expression and P450 mediated monooxygenase functions, without as well as after treatment with the mitogens or cytotoxins. Similar to adult liver, the hepatocytes in the transplant containing spleens showed nearly no P450 1A1, but a noticeable P450 2B1 and 3A2 expression. No staining was observed within the bile duct cells of the intrasplenic transplants.

Combined effect of propranolol with nifedipine or with diltiazem on rat liver monooxygenase activities.

The effects of two Ca2+ antagonists nifedipine (NF) and diltiazem (DL) and of the nonselective beta-adrenergic blocking agent propranolol (PR) on the hexobarbital (HB) sleeping time and on the activity of some liver drug-metabolizing enzyme systems in male Wistar rats were studied. Two h after single oral administration PR (50 mg/kg) did not change HB sleeping time, while NF (50 mg/kg) and DL (30 mg/kg) prolonged it by 171.2 and 99.6%, respectively. Coadmistration of PR with DL or with NF significantly prolonged HB sleep by 240.7 and 129%, respectively. Only NF increased aniline 4-hidroxylase (AH) activity (by 92%) and the total P-450 content (by 24%). PR and NF increased cytochrome b5 content and this effect was also observed with the combinations PR + NF (by 109%) and PR + DL (by 102%). The NADPH cytochrome P-450 reductase activity was significantly decreased by NF and DL and after their combination with PR. The ethymorphine-N-demethylase (EMND) and amidopyrine-N-demethylase (APND) activities were not changed. The effects of PR, NF and DL administrated alone or in combination on liver oxidative metabolism are considered as possible mechanisms of drug interactions.

[Involvement of cytochrome P4503A in the monohydroxylation of ring A of praziquantel in rat liver microsomes].

The possibility of involvement of cytochrome P4503A (CYP3A) in the monohydroxylation of the ring A of praziquantel (PZQ) was studied by using CYP3A specific inducer dexamethasone (DEX), specific inhibitor triacetyloeandomycin (TAO) and the activity of erythromycin (ERY) and ethylmorphine (EMP) N-demethylase which are known to be marker for CYP3A enzyme activity as probes. In the liver microsomes obtained from rats pretreated with CYP3A inducer DEX with TAO treatment the content of uncomplexed P450 was decreased, the activity of ERY and EMP N-demethylase ws reduced, and simultaneously, the rate of formation of ring A monohydroxylate of PZQ was inhibited. Ring A monohydroxylate formation was competitively inhibited by TAO and ERY. The rates of ring A monohydroxylate formation were strongly correlated with the activity of N-demethylase of ERY and EMP. These results indicate that CYP3A is involved in the monohydroxylation of the ring A of PZQ.

Testicular toxicity of Di(2-ethylhexyl)phthalate in developing rats.

To understand the factors involved in the enhanced testicular toxicity of di(2-ethylhexyl)phthalate (DEHP) in developing animals, po doses of 50, 100, 250 or 500 mg DEHP/kg were administered to 25-d-old albino rats for 30 consecutive days. Activities of testicular and hepatic cytochrome P-450 enzymes were determined. A dose-dependent increase in the activities of lactate dehydrogenase and gamma-glutamyl transpeptidase and a decrease in sorbitol dehydrogenase was observed in the testes. The activity of beta-glucuronidase increased at dosages of 250 and 500 mg/kg, while acid phosphatase decreased. Testes had marked destructive changes in the advanced germ cell layers at dosages of 250 and 500 mg/kg, which supports biochemical studies indicating that DEHP interacts with the maturation process of the testes. The dose-dependent decrease in hepatic cytochrome P-450 levels and the activities of ethylmorphine N-demethylase and aniline hydroxylase suggest that impaired metabolism of DEHP could lead to higher amounts of the diester or its metabolites reaching the testes; this may result in enhanced vulnerability of the testes to DEHP in developing animals.

Differential responses of hepatic monooxygenases and glutathione S-transferases of mice to a combination of cadmium and nickel.

The acute combined effects of cadmium (Cd) and nickel (Ni) on hepatic monooxygenase activities (ethylmorphine N-demethylase, EMND; aminopyrine N-demethylase, AMND; aniline 4-hydroxylase, AH), cytochrome P-450, cytochrome b5, microsomal heme and reduced glutathione (GSH) levels and glutathione S-transferase (GST) activities toward several substrates (1-chloro-2,4-dinitrobenzene, CDNB; 1,2-dichloro-4-nitrobenzene, DCNB; ethacrynic acid, EAA; 1,2-epoxy-3-(p-nitrophenoxy)-propane, ENPP) were determined and compared with those of Cd or Ni alone in mice. Male adult mice (25-30 g) were administered either a single dose of Cd (3.58 mg CdCl2.H2O/kg, i.p.) 48 hr prior to killing or a single dose of Ni (59.5 mg NiCl2.H2O/kg, s.c.) 16 hr prior to killing. For the combined treatment, the animals received the single dose of Ni 32 hr after the single dose of Cd and were then killed 16 hr later. Cd treatment alone significantly decreased EMND, AMND, and AH activities and cytochrome P-450 and heme levels as compared with controls. Cytochrome b5 level was not altered by Cd treatment. Cd also inhibited GSH level and the GST activities toward CDNB, EAA and ENPP significantly. No significant change was observed in the GST activity for DCNB by Cd. Ni treatment alone, however, decreased the monooxygenase and GST activities studied, and cytochrome P-450, cytochrome b5, heme and GSH levels significantly. Combined treatment significantly depressed the monooxygenase activities and cytochromes and heme levels. GSH level was not significantly altered.(ABSTRACT TRUNCATED AT 250 WORDS)

Effect of spironolactone on dimethyl mercury toxicity. A possible molecular mechanism.

The protective activity of spironolactone (CAS 52-01-7) against dimethyl mercury intoxication was studied. Dimethyl mercury increased serum glutamate pyruvate transaminase (SGPT), serum bilirubin, blood urea nitrogen (BUN), and caused impairment of the drug metabolic activity of rat liver in vivo and in vitro. It also caused a severe neuropathy to these animals. Administration of spironolactone caused a reduction of dimethyl mercury toxicity. It decreased the values of SGPT, bilirubin and BUN, and restored the impaired drug metabolism caused by dimethyl mercury. The neuropathy produced after administration of dimethyl mercury was only mildly ameliorated by the treatment with spironolactone. Pregnenolone-16 alpha-carbonitrile (PCN), a potent microsomal enzyme inducer, had only a weak action, with the expected exception of the repair of the impaired drug metabolism of the liver. A mechanism of the protective action of spironolactone against dimethyl mercury intoxication is proposed. It is suggested that both the ability to induce drug metabolizing enzymes, here demethylases, and the capacity to bind to the demethylated metabolite of the organic mercurial, giving a non toxic, easily excretable complex should coexist in the protective molecule.

Age related effects of di(2-ethylhexyl)phthalate on hepatic cytochrome P450 monooxygenases in Wistar rats.

Age related response of di(2-ethylhexyl)phthalate (DEHP) was studied by administering the plasticizer (2000 mg/kg, orally) to 3, 6 and 10 week old wistar rats for 1, 7 or 15 days and determining the activity of hepatic P450 monooxygenases. Single administration of DEHP decreased the cytochrome P450 contents and activity of arylhydrocarbon hydroxylase, aniline hydroxylase and ethylmorphine N-demethylase at all the age groups while repeated exposure induced them with maximum alterations occurring in 3 week old rats. Repeated administration for 15 days, on the other hand, caused a decrease in the cytochrome P450 content and the activity of arylhydrocarbon hydroxylase, aniline hydroxylase and ethylmorphine N-demethylase only in the 3 week old rats. The 6 and 10 week old rats exposed to the same schedule of DEHP showed an inhibition in the activity of arylhydrocarbon hydroxylase and an increase in the activity of aniline hydroxylase and ethylmorphine N-demethylase, which was however lower than that seen after 7 days of exposure to DEHP in the respective age group of animals. Our data have indicated the differences in the sensitivity of the P450 monooxygenases towards DEHP and its metabolites amongst 3, 6 and 10 week old animals.

Estrogen, not testosterone, creates male predominance of a P4501-related cytochrome in adult guinea pig adrenals.

Guinea pig adrenal microsomes possess a distinctive cytochrome P450 that is immunochemically related to P4501 and correlates with microsomal capacity for xenobiotic metabolism. This 52K protein and the capacity for metabolizing compounds such as ethylmorphine are located in the zona reticularis, are suppressed by ACTH, and are predominant in adult males. The protein is undetectable and the enzyme activity is low in young prepubertal animals. In males, both increase with age. However, in females, the protein remains undetectable, and ethylmorphine demethylase activity remains low into adulthood. Despite this clear sex difference through puberty and into sexual maturity, we recently observed that in female retired breeders, both the 52K cytochrome P450 and the capacity for metabolism of ethylmorphine appear at levels equal to those in males of comparable age. As estrogen levels are low in female retired breeders, we decided to investigate whether estrogen plays a role in maintaining the low levels of this protein and of xenobiotic metabolism seen in younger females. In a series of gonadectomy and hormone replacement experiments, we demonstrated that estrogen suppressed the levels of both protein and enzyme activity in adult guinea pigs. Ovariectomy resulted in the appearance of the 52K cytochrome P450 and of ethylmorphine demethylase in female adrenal microsomes at levels comparable to those seen in adult males. Estrogen replacement suppressed the increase in both protein concentration and enzyme activity. In hemiovariectomized cycling females, compensatory hypertrophy of the remaining ovary occurred, and the characteristic low levels of the 52K P450 and enzyme activity were maintained. Furthermore, estrogen treatment of male guinea pigs suppressed levels of both the 52K P450 and ethylmorphine demethylase activity in male adrenals. These experiments demonstrate that estrogen plays a significant role in the regulation of this protein. Testosterone, on the other hand, was not required to maintain the higher levels of 52K P450 and correlated enzyme activity in adult males. The levels of both were the same in normal, castrated, and sham-operated males, treated with testosterone or vehicle alone or left untreated. In fact, castration of prepubertal males resulted in a rapid rise to adult male levels of both immunodetectable protein and enzyme activity, implying that some suppressive agent of testicular origin effects the gradual increase that normally occurs with age in males.(ABSTRACT TRUNCATED AT 400 WORDS)

Studies on the acute effects of coumarin and some coumarin derivatives in the rat.

The mechanism of acute coumarin-induced hepatotoxicity in the rat has been investigated by comparing the effects of coumarin with those of a number of methyl-substituted coumarin derivatives. Male Sprague-Dawley rats were given single ip doses of corn oil (control), coumarin (0.86 and 1.71 mmol/kg body weight), 3,4-dimethylcoumarin (3,4-DMC, 1.71 and 2.57 mmol/kg), 3-, 4- and 6-methylcoumarins (3-MC, 4-MC and 6-MC, 1.71 mmol/kg) and 3- and 4-methyloctahydrocoumarins (3-MOHC and 4-MOHC, 2.57 mmol/kg) and hepatotoxicity assessed after 24 hr. Coumarin administration produced dose-related hepatic necrosis and a marked elevation of plasma alanine aminotransferase and aspartate aminotransferase activities. In contrast, none of the coumarin derivatives examined produced either hepatic necrosis or elevated plasma transaminase activities. Treatment with coumarin reduced hepatic microsomal ethylmorphine N-demethylase and 7-ethoxycoumarin O-deethylase activities, whereas one or both mixed-function oxidases appeared to be induced by treatment with 3,4-DMC, 4-MC, 3-MOHC and 4-MOHC. These results provide further evidence that acute coumarin-induced hepatotoxicity in the rat is due to the formation of a coumarin 3,4-epoxide intermediate. That 3- and/or 4-methyl substitution (i.e. 3-MC, 4-MC and 3,4-DMC) leads to a reduction in coumarin-induced hepatotoxicity, due to diminished formation of 3,4-epoxide intermediates, was confirmed by the results of molecular orbital calculations.

[Effect of long-term administration of undevit and its combination with cordiamine on hydroxylation, glucuronidation and glutathione conjugation systems and lipid peroxidation in rat liver]. / Vliianie dlitelnogo vvedeniia undevita i ego kombinatsii s kordiaminomna gidroksiliruiushchuiu, gliukuro-, glutationkoníugiruiushchuiu sistemy i perekisnoe okislenie lipidov pecheni intaktnykh krys.

Increase of N-demethylase and antioxidant activities and decrease of amount of hydroperoxides were observed in liver of rats after intragastric administration of Undevit (1/8 dragee/rats/day, 5 times a week during 2 months). Combination of Undevit with Cordiamin (5 mg/rat/day) resulted in augmentation of cytochromes P-450 and b5 content and activities of their reductases, velocity of amidopyrine and ethylmorphine N-demethylation and oxidation of HADPH. Activities of UDP-glucuronyltransferase, glutathionetransferase and hydrophobity of microsomal membranes were also increased. Antioxidant activity was increased and amount of hydroperoxides in liver microsomes and homogenates and in plasma was decreased in greater degree after administration of combination two compounds than after administration of Undevit alone.

Postnatal development of body mass and of hepatic xenobiotics biotransformation of male rats with low body mass at birth.

Body mass gain of male rats with low body mass at birth was lower from birth up to the 5th postnatal day, but higher from the days 5 to 30 in comparison with male litter mates with normal body mass at birth. Absolute body and liver masses were significantly lower up to the 30th postnatal day, but fully improved at adult age. No differences of ethoxycoumarin O-deethylation activities existed between both groups at all ages. Ethylmorphine N-demethylation activity was significantly lower in newborn males with low body mass, but had been fully developed as early as the 5th postnatal day. We conclude that postnatal adaptation of male rats with spontaneous growth retardation at birth is slightly delayed only during the first days of life.

Regioselective O-demethylation of scoparone (6,7-dimethoxycoumarin) to assess cytochrome P450 activities in vitro in rat. Effects of gonadal steroids and the involvement of constitutive P450 enzymes.

1. In the rat, scoparone (6,7-dimethoxycoumarin) is regioselectively O-demethylated in vitro by the hepatic cytochrome P450 (P450) system, yielding isoscopoletin and scopoletin. 2. Scoparone has been proposed as an indicator substrate for the assessment of P450 differentiation in vitro in rat. It has been suggested that isoscopoletin formation mainly reflects activity of enzymes of the P4501A subfamily, whereas scopoletin formation has been associated with P4502B activity. 3. In the present study, rate of formation of scopoletin and isoscopoletin were measured in hepatic microsomes from male, female and castrated male rats, in castrates treated with testosterone, in males treated with oestradiol, and in females treated with testosterone. Furthermore, effects of induction by phenobarbital (PB), beta-naphthoflavone (BNF), isoniazid, triacetyloleandomycin, and dexamethasone were studied in both sexes. 4. Scoparone metabolism was partly sex- and steroid-dependent. Variation of isoscopoletin formation with sex or hormonal status correlated well with ethylmorphine demethylation. 5. Scoparone-O-demethylation was regioselectively induced by PB and BNF. Induction effects were not very large and showed no sex differences. 6. Microsomal metabolism of scoparone was partly inhibited by a monoclonal antibody against P4502C11. Scopoletin and isoscopoletin formation were inhibited when ethylmorphine was added to the incubation mixture. Scoparone competitively inhibited testosterone 6 beta-hydroxylation. 7. It is concluded that a number of P450 enzymes, including those which are constitutively expressed, contribute to the biotransformation of scoparone. This lack of selectivity limits the usefulness of scoparone as a general indicator substrate for P450 differentiation in rats.

Sex differences in the diabetes-induced modulation of rat hepatic cytochrome P450 proteins.

Sex differences in the diabetes-induced changes in hepatic cytochrome P450 proteins were investigated in rats treated with streptozotocin. Changes in specific cytochrome P450 proteins were monitored using diagnostic substrates and immunologically utilizing specific polyclonal antibodies. When expressed in terms of nmoles of total cytochrome P450, ethoxyresorufin O-deethylase activity was increased by treatment with streptozotocin, the extent of induction being the same in the two sexes. In contrast, lauric acid hydroxylase and ethylmorphine N-demethylase activities were induced only in the male rat. Finally, p-nitrophenol hydroxylase and pentoxyresorufin O-dealkylase were enhanced by the same treatment in both sexes, the effect being more pronounced in the male. These findings indicate that sex-specific changes in certain cytochrome P450 proteins exist in response to insulin-dependent diabetes but these cannot, however, be ascribed to sex differences in the severity of diabetes induced by streptozotocin since the degrees of hyperketonaemia and hyperglycaemia were the same in the two sexes. These are likely to reflect sex-specific differences in growth hormone and triglyceride levels in the diabetic animals.

Comparison of human fetal hepatic and adrenal cytochrome P450 activities with some major gestational steroids and ethylmorphine as substrates.

The immunoidentified human fetal liver and adrenal microsomal contents of cytochromes P450IIIA and P450XVIIA1 were compared to the metabolism of steroids and ethylmorphine. In fetal liver microsomes, 16 alpha-hydroxylation of dehydroepiandrosterone (DHA) was catalyzed at a high rate in almost all investigated specimens and accompanied by a high ethylmorphine N-demethylase activity. Progesterone 16 alpha- and 17 alpha-hydroxylation was found only in the livers with the highest DHA 16 alpha-hydroxylation activities, while 21-hydroxylation of progesterone was catalyzed only occasionally in these samples. In fetal adrenal microsomes, 21-hydroxylation of progesterone to 11-desoxycorticosterone (DOC) and 11-desoxycortisol (DOCOL) was catalyzed. In contrast to fetal liver, the adrenals also catalyzed the 17 alpha-hydroxylation of pregnenolone and the formation of DHA from 17 alpha-OH-pregnenolone. 16 alpha-hydroxylation of DHA and ethylmorphine N-demethylation were modest in the adrenals. P450IIIA/HLp was immunoidentified in all investigated liver specimens except two (18/20) in which no ethylmorphine N-demethylation or 16 alpha-hydroxylation of DHA was found. P450XVIIA1 bands were observed in 8/20 blots of liver specimens, but there was no correlation between the density of these bands and the 17 alpha-hydroxylation of progesterone. All 11 fetal adrenal samples catalyzed DHA 16 alpha-hydroxylation, although only 8 were positive for P450IIIA/HLp. All investigated adrenals were positive in regard of the P450XVIIA1 band, except one (8/9) with a low 17 alpha-hydroxylation of progesterone. All adrenal specimens catalyzed 21-hydroxylation of progesterone and contained P450C21 bands in immunoblots and all samples catalyzed the formation of DOC and DOCOL from progesterone. Our findings in the fetal livers show a correlation between the DHA 16 alpha-hydroxylation and immunoidentified P450IIIA/HLp bands. In adrenals, there was a correlation between the immunoidentified P450XVIIA1 bands and the 17 alpha-hydroxylation of progesterone.

Interferon suppresses the isoform-specific activities of hepatic cytochrome P450 in female rats.

The results from the present study indicate that rat IFN-gamma decreases hepatic P450 levels and catalytic activities in female rats. The magnitude of the down-regulation of P450 is similar to that found in male rats. Furthermore, the activities of specific P450 isoforms (P4502A1 and P4502C6 and possibly P4502C12, and P4502B1 and 2) are decreased by IFN. Hence, the effect of IFN in female rats appears to involve several, but not all, hepatic P450 isoforms. Both hormone-dependent and hormone-independent isoforms are suppressed by IFN. The mechanism for this effect does not, however, appear to be mediated by changes in serum levels of sex steroids.