Tomonori Tateishi

A COMPARISON OF HEPATIC CYTOCHROME P450 PROTEIN EXPRESSION BETWEEN INFANCY AND POSTINFANCY

We immunochemically measured the contents of 9 different cytochrome P450 (CYP) isoenzymes expressed in the liver and compared them between two groups: one group of 6 infant and 4 perinatal patients and one group of 10 patients after infancy (over 1 year old). CYP protein expressed in human liver can be divided into three groups on the basis of expression pattern: (a) CYP2A6, 2C9, 2D6, 2E1, and 3A were present in all samples and no difference was observed between the two groups; (b) CYP1A2, 2B6, and 2C8 were expressed more after infancy than during infancy; and (c) CYP3A7, which has been considered a major CYP enzyme in fetal liver microsomes, was expressed in all infants as well as the four perinatal patients, whereas it was detected in only 2 patients after infancy. These results implied that CYP2A6, 2C9, 2D6, 2E1, and 3A are already expressed during perinatal and infant period, while CYP1A2, 2B6, and 2C8 are expressed highly in subjects over 1 year old, and CYP3A7 disappeared after infancy.

No ethnic difference between Caucasian and Japanese hepatic samples in the expression frequency of CYP3A5 and CYP3A7 proteins.

Ethnic differences in the pharmacokinetics of nifedipine, a substrate of CYP3A, and in CYP3A7 expression have been reported. The aim of the present study was to measure the protein levels of CYP3A4, CYP3A5, and CYP3A7 and nifedipine oxidation activity in hepatic microsomes from 15 Caucasian and 15 Japanese patients for comparison between the two ethnic groups. Nifedipine oxidation activity and CYP3A4 protein level were well correlated. No significant difference between Caucasian and Japanese microsomal samples was found in nifedipine oxidation activity or in the CYP3A4 protein level. CYP3A5 was detected in 6 of 15 Caucasian samples and in 5 of 15 Japanese samples, but no ethnic difference was found in either the frequency of expression or its protein level. CYP3A7 was found in 10 of 15 Caucasian samples and in 14 of 15 Japanese samples. Although the estimated CYP3A7 protein level was higher in the Japanese than in the Caucasian samples, its protein level was much lower than that of CYP3A4. These results imply that the contribution of CYP3A5 or CYP3A7 to the purported Caucasian-Japanese ethnic difference in the overall CYP3A activity seems to be small.

Effects of Estrogen on Serum Leptin Levels and Leptin mRNA Expression in Adipose Tissue in Rats

Objective: In this study, we examined changes in serum leptin levels during the estrus cycle and the role of estrogen in these changes. Methods: We measured serum leptin levels during normal estrus cycles in intact rats and estradiol-17β (E2)-induced artificial estrus cycles in ovariectomized rats. Results: Serum leptin levels increased 1.6-fold from 4.2 ± 0.2 ng/ml during diestrus stage 2 to 6.7 ± 0.9 ng/ml during proestrus stage during the 4-day estrus cycle. During the E2-induced estrus cycle, serum leptin levels increased 2.3-fold from 2.3 ± 0.1 ng/ml at estrus to 5.4 ± 1.2 ng/ml at proestrus. E2 also increased serum leptin concentrations and leptin mRNA expression in adipose tissue of immature rats. Discussion: These findings suggest that increased serum leptin induced by estrogen during proestrus may trigger the preovulatory release of luteinizing hormone. Furthermore, our findings indicate that estrogen has a positive effect on leptin production in adipose tissue.

CARBAMAZEPINE INDUCES MULTIPLE CYTOCHROME P450 SUBFAMILIES IN RATS

We compared the effect of three different doses (30, 60, and 100 mg/kg) of carbamazepine (CBZ) administered intraperitoneally for 1, 3, and 7 days on the activity and protein content of hepatic cytochrome P450 (CYP) subfamilies in Sprague-Dawley rats. After 3-day- and 7-day administration with CBZ, the total CYP content had increased in a dose-dependent fashion. Among six enzyme activities examined, only aniline hydroxylase activity remained unchanged after 7-day treatment with CBZ. Pentoxyresorufin O-deethylase activity showed the most significant increase and was induced up to 7 days in a time-dependent fashion. Pretreatment of rats with cycloheximide significantly suppressed the pentoxyresorufin O-deethylase induction by one dose of 100 mg/day CBZ. Immunoblot analysis showed a significant correlation between the protein content of each isoenzyme examined and its activity except CYP2E1 after 7-day treatment with CBZ. Similar results were obtained in the mRNA levels of CYP subfamilies. These results suggested that CBZ may induce multiple CYP subfamilies, except CYP2E1, and the activity and the protein content of CYP2B showed the greatest increase with increased CYP2B mRNA.

Effect of interferon-α on tyrosine hydroxylase and catecholamine levels in the brain of rats

Abstract We investigated the effect of interferon on tyrosine hydroxylase (TH) and catecholamine levels in the brains of 12-week-old male Wistar rats. Interferon-α (300,000 IU/kg/day, s.c.) was administered to rats for 7 days. Locomotor activity of interferon-α-treated rats was significantly lower than that of control rats. Norepinephrine and dopamine levels and TH activities in the cerebral cortex, hypothalamus and medulla oblongata of interferon-α-treated rats were significantly higher than those of control rats. Norepinephrine and dopamine levels and TH activities in the thalamus and hippocampus were not different between interferon-α treated and control rats. These results suggest that interferon-α-induced depression may be related to change in the catecholamine synthetic pathway in the central nervous system.

Effects of anti-androgen treatment on the catecholamine synthetic pathway in the adrenal medulla of spontaneously hypertensive rats

We investigated the effects of the antiandrogen flutamide on the activity of tyrosine hydroxylase, the levels of its encoding mRNA, and catecholamine levels in the adrenal medulla of male spontaneously hypertensive rats (SHR) and Wistar Kyoto rats (WKY). Flutamide (30 mg/kg) was administered subcutaneously daily (between 9 and 15 weeks of age). The systolic blood pressure of flutamide-treated SHR rats was lower than that of control SHR. Epinephrine and norepinephrine levels, tyrosine hydroxylase activity, and the levels of encoding mRNA in the adrenal medulla were significantly lower in flutamide-treated SHR rats than in paired controls. Systolic blood pressure, epinephrine and norepinephrine levels, tyrosine hydroxylase activity, and encoding mRNA in the adrenal medulla of WKY rats showed no significant differences between flutamide-treated and control groups. These findings suggested that flutamide may have cardiovascular effects through alteration of the catecholamine synthetic pathway caused by removal of androgen receptor stimulation on the expression of tyrosine hydroxylase in the adrenal medulla of male SHR rats.

Effects of glucocorticoids on pharmacokinetics and pharmacodynamics of midazolam in rats

We investigated the effect of dexamethasone (80 mg/kg per day for 2 days) and prednisolone (600 mg/kg per day for 2 days, equivalent to dexamethasone for glucocorticoid (GC) potency) on both pharmacokinetics and pharmacodynamics of midazolam (MDZ), a substrate for cytochrome P450 (CYP) 3A, in 8-week-old male Sprague-Dawley rats. Animals received a single injection of MDZ (pharmacokinetic study, 10 mg/kg; pharmacodynamic study, 55.5 mg/kg) in the tail vein 24 h after the last dose of GC or placebo. The elimination half-life (t(1/2)) and the area under the concentration-time curve of MDZ were significantly reduced by pretreatment with dexamethasone to 58.9% and 44.7% of the control value, respectively, and the clearance of MDZ was significantly increased by dexamethasone. Similar changes observed by prednisolone pretreatment did not reach significance. The t(1/2) of the dexamethasone pretreatment group (14.4+/-0.7 min) was significantly shorter than that of the prednisolone group (20.9+/-1.5 min). The amount of CYP3A2 protein and the activity of erythromycin N-demethylase were significantly increased by dexamethasone and prednisolone pretreatments, but dexamethasone showed a greater effect than prednisolone. Sleeping time was significantly shortened by dexamethasone and prednisolone pretreatment to 38.7% and 57.1% of control value, respectively. The current study demonstrates that the anesthetic effect of MDZ would be reduced in patients treated with dexamethasone or prednisolone, and that the CYP3A induction was greater by dexamethasone than by prednisolone, implying that the potency of CYP3A induction may differ among GCs even when GC activity is the same.

Multiple cytochrome P-450 subfamilies are co-induced with P-glycoprotein by both phenothiazine and 2-acetylaminofluorene in rats

We studied the effects of two P-glycoprotein (P-gp) inducers, 2-acetylaminofluorene (2-AAF) and phenothiazine (PTZ), administered intraperitoneally, on the activities and content of hepatic cytochrome P-450 (CYP) subfamilies in hepatic microsomes of Sprague-Dawley rats. After 4-day administration of 2-AAF or PTZ, the P-gp content was increased. The total CYP content after PTZ treatment was significantly increased compared with that of controls. The CYP1A, CYP2B and CYP3A2 contents were induced, while the CYP2C6, CYP2C11 and CYP2E1 contents remained unaffected. A marked increase in CYP1A1 was found after administration of each compound. Ethoxyresorufin O-deethylase, pentoxyresorufin O-deethylase, and testosterone 6beta hydroxylation activities showed a significant increase after both 2-AAF and PTZ treatments. In particular, ethoxyresorufin O-deethylase exhibited more than ten times greater activity than that of the controls after the treatments. These results suggest that P-gp inducers affect several CYP subfamilies in addition to CYP3A, which is reported to be up-regulated coordinately with P-gp by a CYP3A inducer.

CYP3A is responsible for N-dealkylation of haloperidol and bromperidol and oxidation of their reduced forms by human liver microsomes

We studied the biotransformation of haloperidol, bromperidol and their reduced forms by human liver microsomes. Nifedipine oxidation (CYP3A) activity correlated significantly with N-dealkylation rates of haloperidol and bromperidol and oxidation rates of their reduced forms, while neither ethoxyresorufin O-deethylation (CYP1A2) activity nor dextromethorphan O-deethylation (CYP2D6) activity did. In chemical and immunoinhibition studies, only troleandomycin and anti-CYP3A4 serum inhibited both formation rates of 4-fluorobenzoylpropionic acid, a metabolite of haloperidol and bromperidol, and back oxidation rates. Among 10 recombinant isoforms examined, only CYP3A4 showed catalytic activity. The Vmax and Km values of N-dealkylation of bromperidol and reoxidation of reduced bromperidol were similar to those of haloperidol and reduced haloperidol, respectively. The present study indicates that CYP3A plays a major role in N-dealkylation of and oxidation back to bromperidol as well as haloperidol and suggests that modification of in vivo CYP3A activity by inhibition or induction may affect the pharmacokinetics and therapeutic effects of haloperidol and bromperidol.

Involvement of tyrosine hydroxylase up regulation in dexamethasone-induced hypertension of rats.

We investigated the effect of dexamethasone (DEX) on tyrosine hydroxylase (TH) mRNA level, and TH activity and catecholamine levels in the adrenal medulla of the rat. DEX (1 mg/kg/day, s.c.) was administered for 2 days, and a control group was given corn oil. DEX significantly increased systolic blood pressure. TH mRNA level, TH activity, epinephrine level, and norepinephrine level in the adrenal medulla of DEX-treated rats were significantly higher than those of control rats. Also, epinephrine and norepinephrine levels in plasma were significantly higher in DEX-treated rats than in controls. alpha-Methyl-p-tyrosine prevented the DEX-induced blood pressure increase. These results suggest that the catecholamine synthetic pathway may be involved in DEX-induced hypertension.