Biomedical subjects

Y Funae

Publications and source records attributed to Y Funae.

At least 55 records · Page 3Linked to original sources

PubMed1998

Strain differences in CYP3A-mediated C-8 hydroxylation (1,3,7-trimethyluric acid formation) of caffeine in Wistar and Dark Agouti rats. Rapid metabolism of caffeine in debrisoquine poor metabolizer model rats.

We observed significant strain differences [Dark Agouti (DA) > Wistar] in 1,3,7-trimethyluric acid formation (C-8 hydroxylation) during caffeine metabolism, though not in N-demethylations, in adult male DA and Wistar rats. In contrast, adult female and immature male rats of both DA and Wistar strains did not show significant differences in activity levels of C-8 hydroxylation. Kinetic studies using liver microsomes revealed that adult male DA rats have a larger Vmax for C-8 hydroxylation than do Wistar rats. Troleandomycin (TAO), known as a cytochrome P450 (CYP) 3A inhibitor, and an anti-rat CYP3A2 polyclonal antibody effectively reduced C-8 hydroxylation by rat liver microsomes in a concentration-dependent manner, suggesting that C-8 hydroxylation in rats is mediated largely by an isoform(s) of the CYP3A subfamily. Troleandomycin and the antibody did not inhibit the N-demethylations of caffeine by rat liver microsomes. Treatment of rats with CYP3A inducers caused a marked increase in C-8 hydroxylase activity. These results indicate that the rat CYP3A subfamily is capable of catalyzing C-8 hydroxylation of caffeine as is the case for human CYP3A4. The results of western blotting analysis using anti CYP3A antiserum showed that the staining intensity of the protein band in DA rat liver microsomes was higher than that in Wistar rat liver microsomes. We concluded that marked sex-dependent strain differences in C-8 hydroxylation of caffeine between Wistar and DA rats are due to the differences in the levels of expression of CYP3A in these strains of rats.

Aging↗

PubMed1998

Evidence that immature rat liver is capable of participating in steroidogenesis by expressing 17alpha-hydroxylase/17,20-lyase P450c17.

Steroid hydroxylase cytochrome P450c17 has been previously purified from microsomal fractions of immature rat livers. In this study, we investigated the expression of P450c17 in rat livers to understand a role of steroidogenesis in the extrasteroidogenic tissue. Upon immunoblot analysis utilizing liver microsomes from rats, P450c17 was detected in 1 and 3 week old rats but not in adult rats. Data from immunohistochemical studies also showed a similar age-dependent expression of P450c17 and indicated that P450c17 detected in immature rat livers is localized in cells surrounding interlobular veins. This age-dependent expression of P450c17 in rat livers was observed in both sexes. Upon enzymatic analysis utilizing microsomal fractions from livers, levels of 17alpha-hydroxylase and 17,20-lyase activity for pregnenolone and progesterone increased by 3 weeks and dramatically reduced at 7 weeks, which is consistent with the expression level of P450c17. These data clearly indicate that P450c17 is expressed in immature rat liver to produce 17alpha-hydroxysteroids and C19-steroids. Based upon immunoblot analysis, the expression level of P450c17 in immature rat livers was approximately one third of that in testis. Compared expression level of P450c17 and total volume of organs between liver and testis, the total amount of steroid metabolites produced by liver P450c17 could be greater than that produced by gonadal P450c17. Because of the absence of P450c17 in rat adrenal glands, rat liver could be the major site for producing 17alpha-hydroxysteroids and C19-steroids in this particular period of life. Although physiological products formed by P450c17 in liver and their roles remain to be elucidated, this study suggests a large capacity of prepubertal rat liver for participating the production of steroid hormones and a putative importance of 17alpha-hydroxysteroids and C19-steroids, such as cortisol and androstendione, which are generally believed to be minor components of steroid hormones in rodents.

Animals↗

PubMed1998

Presence of a threshold for promoting effects of phenobarbital on diethylnitrosamine-induced hepatic foci in the rat.

The dose dependence of the hepatopromoting effects of phenobarbital (PB) was investigated in a rat liver medium-term bioassay (Ito test) to elucidate a practical threshold level. F344 rats were given a single i.p. injection of diethylnitrosamine (200 mg/kg body wt) and subjected to two-thirds partial hepatectomy at week 3. Commencing 2 weeks from the start, PB at doses of 0, 1, 2, 4, 7.5, 15 or 500 p.p.m. in experiment 1 and 0, 0.01, 0.1 or 0.5 p.p.m. in experiment 2 were fed to the rats for 6 weeks. Experiment 3 was conducted to confirm previous data using the same medium-term bioassay, with PB at doses of 0, 1, 2, 4, 7.5, 15, 30, 60, 125, 250 or 500 p.p.m. fed to the rats. All surviving animals were killed at week 8 in these experiments and their livers were immunohistochemically examined for expression of glutathione S-transferase placental form (GST-P). Quantitative values for GST-P-positive foci in the liver were increased dose dependently in rats given 60-500 p.p.m. PB. However, those for doses in the range 1-7.5 p.p.m. demonstrated a decrease as compared with the control group (0 p.p.m.), with significant differences observed for 1 and 2 p.p.m.. The results for 15-30 and 0.01-0.5 p.p.m. were comparable with the control values. Examination of transforming growth factor-alpha (TGF-alpha)-positive foci also produced similar results to those for GST-P in experiment 1. Immunohistochemical staining of TGF-alpha and GST-P using serial liver sections demonstrated that the TGF-alpha-positive foci comprised a sub-population of the GST-P-positive lesions, being approximately 1/8-1/10th as common in livers of animals treated with PB. TGF-alpha-positive foci were almost always negative on immunostaining for TGF-beta. Western blotting for proteins CYP2B1, 2C6 and 3A2 revealed a good correlation between changes in GST-P-positive foci and CYP3A2 protein expression. The finding of inhibition effects at low doses of PB confirms the presence of a threshold level for promoting effects by PB on liver carcinogenesis in rats.

Animals↗

PubMed1998

A new deleted allele in the human cytochrome P450 2A6 (CYP2A6) gene found in individuals showing poor metabolic capacity to coumarin and (+)-cis-3,5-dimethyl-2-(3-pyridyl)thiazolidin-4-one hydrochloride (SM-12502).

The S-oxidation of (4)-cis-3,5-dimethyl-2-(3-pyridyl)thiazolidin-4-one hydrochloride (SM-12502) and the 7-hydroxylation of coumarin are primarily catalyzed by cytochrome P450 2A6 (CYP2A6). The activities of SM-12502 S-oxidase and coumarin 7-hydroxylase were investigated with liver microsomes from 20 human individuals. Liver microsomes from individual H16 showed the lowest activities of both enzymes. The expression of CYP2A6 protein was not detectable in liver microsomes from individuals H4, H5, H7, H8, H12 and H16. CYP2A6 mRNA was hardly detectable in the liver of the individual H16. A new SacI-restriction fragment length polymorphism showing the lack of a 2.6 kb fragment was found in two of forty genomic DNA preparations from individuals H16 and No. 594, using CYP2A6 cDNA as a probe. This deletional 2.6 kb fragment was isolated from a genomic library prepared from one individuals showing normal coumarin 7-hydroxylase activity and was sequenced. This fragment contained a CYP2A6 gene region from 319 bp upstream of a putative exon 6 to a SacI site in exon 9, indicating that this region was deleted in the two individuals in this study. We also demonstrated by polymerase chain reaction analysis that the exon 8 of CYP2A6 gene was deleted in individuals H16 and No. 594. These results indicate that the reduced activity of SM-12502 S-oxidase and no activity of coumarin 7-hydroxylase are caused by the lack of CYP2A6 mRNA and CYP2A6 protein caused by the CYP2A6 gene deletion.

Aryl Hydrocarbon Hydroxylases↗

PubMed1998

[The physiological role of P450-derived arachidonic acid metabolites].

Arachidonic acid is metabolized to biologically active substances by three major enzyme systems including cyclooxygenases, lipoxygenases and cytochrome P450s. The third pathway, P450-dependent pathway, includes allylic oxidation, omega-hydroxylation, and epoxidation of arachidonic acid. Of these metabolites, the physiological role of 20-hydroxyeicosatetraenoic acid (20-HETE) produced by CYP4A isoforms has been extensively studied. 20-HETE affects ion transport, constricts blood vessels and participates in tubuloglomerular feed back. Increased production of 20-HETE is a major factor in elevating blood pressure in spontaneously hypertensive rats (SHR). We have found that CYP4A2 level in SHR is much higher than that of normotensive rat. Recently, factors of endothelial origin other than nitric oxide and prostaglandins were reported. Inhibitors of P450-dependent arachidonic acid metabolism greatly reduce the vasodilator effect and this factor is speculated to be an epoxide of arachidonic acid. We have isolated CYP2C23 from rat kidney and have found that it produces arachidonic acid epoxides. We have investigated changes in the CYP2C23 levels in physiological and pathophysiological conditions. Multiple pathways of arachidonic acid metabolism by P450 have been reported and the diverse properties of these metabolites and the wide distribution of the P450 system make them prime candidates for participation in regulatory mechanisms of the circulation and transporting epithelia.

8,11,14-Eicosatrienoic Acid↗

PubMed1998

[Inhibition of cytochrome P450 by nitric oxide].

Nitric oxide (NO) reacts with iron, superoxide, thiols and oxygen. Although NO reversibly interacts with the heme-iron of P450, the pathophysiological role of this interaction remains to be elucidated. We found that hepatic levels of P450 markedly decreased in endotoxemic rats, particularly when the rate of NO generation was increased. To determine the possible role of NO in the modulation of the structure and function of P450, changes in the levels and activities of P450 isozymes were determined in liver microsomes from normal and endotoxemic rats. Electron spin resonance analysis revealed that incubation of microsomes with the NO donor NOC-7 rapidly generated NO-P450 adducts. Microsomal levels of NO-P450 adducts increased and peaked at 10 min after incubation and decreased thereafter; it disappeared completely within 60 min. In contrast, microsomal levels of the low-spin ferric form and CO-differential spectrally detectable P450 rapidly decreased during the initial 10 min; the signal intensity for P450 recovered thereafter. Western blot analysis using specific antibodies against CYP3A2 and CYP2C11 isozymes revealed no detectable degradation of these isoforms. Effect of NO on the catalytic activity of the enzymes was also determined by using testosterone as the substrate. The hydroxylation activity in microsomes rapidly decreased during the initial 10 min and disappeared slowly thereafter. These results suggested that NO might form dissociable complexes with the heme moiety of P450 and irreversibly inactivate them. The mechanism for P450 inactivation by NO and the role of NO-P450 interaction in the pathogenesis of liver injury in endotoxemia are discussed.

Animals↗

PubMed1998

Nicotine metabolism in liver microsomes from rats with acute hepatitis or cirrhosis.

Nicotine exerts a number of physiological effects. Nicotine is absorbed through the lungs with smoking and is rapidly metabolized in humans. Although it is mainly metabolized in the liver, the effects of liver injuries on nicotine metabolism are not clear. The purpose of this study was to clarify the effects of liver injuries on nicotine metabolism. Rats were treated with D-galactosamine (GalN) or thioacetamide (TA), to induce acute hepatitis or liver cirrhosis, respectively. Serum transaminase levels were significantly elevated in model rats with both types of liver injury. Cytochrome P450 (CYP) and cytochrome b5 contents in liver microsomes were decreased significantly in TA-treated cirrhotic rats but not in GalN-treated hepatitic rats. The major metabolic pathways of nicotine, i.e. cotinine formation catalyzed by CYP and nicotine-1'-N-oxide formation catalyzed by flavin-containing monooxygenase, were investigated in these rat liver microsomes. Formation of cotinine and nicotine-1'-N-oxide from nicotine was not changed in GalN-treated hepatitic rats, in comparison with the controls, but was significantly decreased in TA-treated cirrhotic rats. By immunoblotting, decreases in CYP1A2, CYP2B2, CYP2C, and CYP2E1 protein were recognized in liver microsomes from TA-treated cirrhotic rats. It was also shown that the maximal velocity values for nicotine-1'-N-oxide formation in TA-treated cirrhotic rats were significantly decreased, compared with the controls. These results suggested that the reduction of nicotine metabolism in cirrhosis was due to decreases in CYP and flavin-containing monooxygenase protein expression levels.

Acute Disease↗

PubMed1998

cis-Diamminedichloroplatinum induces peroxisomes as well as CYP4A1 in rat kidney.

Effects of cis-diamminedichloroplatinum (cisplatin) on rat kidney were investigated. Clinical parameters in rat urine and blood were studied. Blood urea nitrogen (BUN) and creatinine in blood and K+ in urine increased, but Na+ in urine decreased. Contents of total P450 and metabolic activities towards lauric acid and arachidonic acid in rat renal microsomes were not changed by cisplatin treatment. The levels of P450 isozymes (CYP4A1, 4A2, 4A8 and 2C23) were determined in rat renal microsomes by immunoblotting. The levels of CYP4A2 and 4A8 which are lauric acid omega-hydroxylases were not changed, but the levels of CYP2C23 and 4A1 were increased significantly by cisplatin treatment. Effects of clofibrate, a typical inducer for CYP4A1, on rat kidney were compared with those of cisplatin. Clofibrate induced palmitoyl CoA oxidase (a marker enzyme of peroxysome), CYP4A1, and CYP4A2 and reduced triglyceride level in plasma. Cisplatin had similar effects to clofibrate and induced peroxysomes as well as CYP4A1, although the effects were at a lesser extent than those of clofibrate. The induction levels of CYP4A1 correlated with increased levels of BUN. The present findings suggest that induction of P450 by cisplatin may take part in the renal injury or nephrotoxicity of cisplatin.

Animals↗

PubMed1998

[14C-oxalate autoradiographic studies on distribution of oxalate and calcium oxalate crystal fixation in rat kidney].

To elucidate the intrarenal distribution of oxalate and to reveal the first site of calcium oxalate crystal fixation along the nephron, we carried out 14C-oxalate renal microautoradiography by the thaw mount method 30, 60 and 90 min after intravenous administration of 14C-oxalate, in normal rats and in hyperoxaluric rats fed a vitamin B6-deficient diet for 4 and 9 weeks. In normal rats, radioactivity was observed in the entire kidney area at 30 min after the injection and at 90 min there was radioactivity only in the papilla, where radioactivity appeared in the parenchyma, mamely the epithelial cells of collecting ducts and the surrounding interstitial tissue. In hyperoxaluric rats, microautoradiograms showed the spotted deposits of 14C-oxalate in both renal papilla and medulla. In 600 x magnification of the autoradiograms, most of the deposits were detected in the epithelial cells of collecting ducts and the surrounding interstitial tissue. These deposits were identified by a polarizing microscope and X-ray microanalysis as crystal deposits of calcium oxalate. These results indicated that oxalate remained for an extended period of time in the epithelial cells of collecting ducts and in the surrounding interstitial tissue in renal papilla, and the calcium oxalate crystals fixed to the same tissue was the first step in the development of calcium oxalate stones.

Animals↗

PubMed1997

Expression of four rat CYP2D isoforms in Saccharomyces cerevisiae and their catalytic specificity.

We cloned four cDNAs belonging to the CYP2D subfamily to express these enzymes in yeast cells and to compare their catalytic activities simultaneously. Three are believed to be alleles of CYP2D1, 2D2, and 2D3, respectively, based on high nucleotide sequence similarity, while CYP2D4 had both sequences of CYP2D4 and CYP2D18. Expression plasmids carrying CYP2D cDNAs were transformed into Saccharomyces cerevisiae. Typical P450 CO-difference spectra with absorbance maximum at 448 nm were recorded with microsomal preparations from the yeast cells expressing the four CYP2D forms. A catalytic study of these CYP2D forms was done with debrisoquine, bufuralol, and lidocaine. CYP2D2 had the highest debrisoquine 4-hydroxylation (2.2 nmol/min/nmol P450) activity, similar to that (2.2 nmol/min/nmol) of human CYP2D6 expressed in yeast cells. CYP2D3 had high lidocaine N-deethylation (43 nmol/min/nmol P450) activity, and both CYP2D3 and 2D2 exhibited high lidocaine 3-hydroxylation (2.4 and 1.6 nmol/min/nmol P450, respectively) activity. Bufuralol 1'-hydroxylation catalytic capabilities were comparable among the four isoforms. The activity of CYP2D1 was relatively low toward the three substrates (debrisoquine, 0.091; bufuralol, 1.5; lidocaine 3-hydroxylation, 0.019; lidocaine N-deethylation, 2.8 nmol/min/nmol P450). These findings indicate that debrisoquine, a typical substrate for CYP2D forms, was mainly metabolized by CYP2D2 but not CYP2D1 in rat liver and that the CYP2D forms have different substrate specificity.

Animals↗

PubMed1997

Mutagenic activation of urinary bladder carcinogens by CYP4B1 and the presence of CYP4B1 in bladder mucosa.

We investigated the mutagenic activation of 2-naphthylamine (2-NA), 3,2'-dimethyl-4-aminobiphenyl (DMAB), and 3,3'-dichlorobenzidine (DCB), bladder carcinogens, by renal and bladder microsomes and by purified P450s using the umu gene expression system, which detects DNA damage. Mouse renal microsomes had high mutagenic activation toward DCB and low activity toward 2-NA. Purified mouse Cyp4b1 also had high mutagenic activity toward DCB. Anti-Cyp4b1 antibody efficiently inhibited DCB bioactivation by mouse renal microsomes with a high Cyp4b1 content. Lauric acid, a substrate of Cyp4b1, efficiently inhibited DCB bioactivation by renal and bladder microsomes of the mouse and by purified Cyp4b1. To assess the contribution of CYP4B1 to bladder carcinoma, further investigation was done with the umu test and an immunochemical study. Ten forms of purified rat P450s including rat CYP4B1 were used in the umu test for 2-NA, DMAB, and DCB. CYP4B1 had the highest activity toward DMAB and DCB. Other P450s had activities of less than 20% that of CYP4B1. CYP4B1 also activated 2-NA, but its activity was about 10% of that toward DMAB or DCB. Rat bladder epithelium was stained specifically with anti-Cyp4b1 antibody, indicating the presence of CYP4B1 in the rat bladder mucosa. Also, CYP4B1 mRNA was detected by northern blotting and reverse transcription-polymerase chain reaction (RT-PCR). These findings suggested that CYP4B1 could contribute to the initiation of carcinogenesis in rat and mouse bladder by activation of aromatic amines.

Animals↗

PubMed1997

Specific binding of 1-[2-(diphenylmethoxy)ethyl]-4-(3-phenyl propyl) piperazine (GBR-12935), an inhibitor of the dopamine transporter, to human CYP2D6.

The binding of [3H]1-[2-(diphenylmethoxy)ethyl]-4-(3-phenyl propyl) piperazine (GBR-12935), an antagonist of the dopamine transporter, to human P450s expressed in yeast cells was investigated. Among the ten forms of human P450 tested (CYP1A1, 1A2, 2A6, 2B6, 2C8, 2C9, 2C18, 2D6, 2E1, and 3A4), [3H]GBR-12935 bound most strongly to CYP2D6. The calculated Kd of [3H]GBR-12935 binding to CYP2D6 was 42.2 nM, indicating that GBR-12935 has a high affinity for CYP2D6. The characteristics of [3H]GBR-12935 binding to CYP2D6 were investigated by competitive studies using several chemicals. The binding of [3H]GBR-12935 to CYP2D6 was not changed by dopamine, suggesting that these binding sites are not dopamine-sensitive binding sites. The binding of [3H]GBR-12935 to CYP2D6 was decreased partially by substrates or inhibitors of CYP2D isoforms (quinine, quinidine, propranolol, bufuralol, imipramine, and desipramine). By means of binding studies using several forms of expressed human P450, we demonstrated that the CYP2D isoform is one GBR-12935 binding site that is insensitive to dopamine.

Carrier Proteins↗

PubMed1997

Effects of ischemia-reperfusion on individual cytochrome P450 isoforms in the rat kidney.

Ischemia-reperfusion of organs such as the kidney produces reactive oxygen and free radical species in tissues and leads to injury of intracellular molecules critical to cell homeostasis. Ischemia-reperfusion affects the NADPH-dependent monooxygenase system including P450 system, which is also a source of reactive oxygen species. In this study, the effects of ischemia-reperfusion on monooxygenase activity and levels of individual P450 isoforms including CYP2C23, 4A2, and 4A8 in the rat kidney were investigated. Ischemia of the rat kidney for 30 min had little effect on lauric acid hydroxylation activity and levels of P450 isoforms but ischemia for 60 min significantly decreased lauric acid omega- and (omega-1)-hydroxylation activities and also decreased the levels of CYP2C23, 4A2, and 4A8. Reperfusion for 60 min after 30-min ischemia decreased the levels of CYP2C23 and 4A2 in the rat kidney although 30-min ischemia did not. Reperfusion for 240 min after 30-min or 60-min ischemia recovered the decreased levels of lauric acid hydroxylation activity and the levels of CYP2C23 and 4A2. Changes in the levels of monooxygenase activity and the levels of P450 isoforms in kidneys by ischemia-reperfusion are faster than those in the liver; it takes several hours for ischemia-reperfusion to affect the levels of monooxygenase activity and the levels of P450 in the rat liver. Our findings suggest that damage of P450 isoforms in the kidney by ischemia-reperfusion occurs by a mechanism different from that in the liver and that active oxygen or free radical species directly attack proteins.

Animals↗

PubMed1997

Expression of cytochrome P450 isoforms in rat hepatic stellate cells.

The current study evaluated the expression and the inducibility of cytochrome P450 isoforms in rat hepatic stellate cells (HSCs). Immunoblotting study revealed that HSCs expressed several P450s and CYP2C11, 3A2, and 2D1 were major isoforms. The levels of CYP2B1, 2C11, 2D1, 2E1, and 3A2 in HSCs were 14 - 38% of those in hepatocytes. CYP1A2 content was similar in each cell type. These P450 levels in HSCs gradually decreased during culture as seen in hepatocytes; the level of CYP3A2 rapidly, whereas that of CYP2D1 slowly decreased. Phenobarbital, a typical inducer of CYP3A2 and 2B1 increased CYP3A2 level as well, but had less potency in the induction of CYP2B1 in HSCs. These results indicate that multiple P450 isoforms were present in HSCs, but their content and inducibility were different between HSCs and hepatocytes.

Animals↗

PubMed1997

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.

Adolescent↗

PubMed1997

Identification and characterization of motility stimulating factor secreted from pancreatic cancer cells: role in tumor invasion and metastasis.

Cell motility is an important factor in the process of invasion and metastasis of tumor. In this study, the relationship between cell motility and experimental metastatic potential was examined using two human pancreatic cancer cell lines, SW1990 and PANC-1. Serum-free conditioned medium from the highly metastatic cell line SW1990 was found to contain a factor that stimulated the migration of and induced a fibroblast-like morphological change in the weakly metastatic cell line PANC-1. Preincubation of PANC-1 cells with SW1990 conditioned medium (SW-C.M.) induced liver metastasis following splenic injection of PANC-1 cells in nude mice, although no liver metastasis was observed without pretreatment of SW-C.M. This factor, temporarily termed PDMF (pancreatic cancer-derived motility factor) is a heparin non-binding protein having a molecular weight of 40 kDa calculated by gel-filtration HPLC which acts not only chemotactically but also chemokinetically, and also acts mainly in a paracrine fashion. However, this factor had no effect on the proliferation of PANC-1 cells; it therefore appears to be a so-called motility factor. Only TGF-beta1 and IL-6 were recognized in the SW-C.M. among cytokines thought to stimulate cell motility. These cytokines stimulated the motility of PANC-1 cells, but differed from PDMF in the neutralizing test with antibody against these cytokines. Results of characterization and preliminary purification suggest that this factor may be a novel motility factor. The above findings suggest that this motility factor may play an important role in the invasion and metastasis of pancreatic cancer, and complete purification of it will be useful in elucidating the mechanism of progression of cancer and designing a strategy for inhibition of invasion and metastasis of pancreatic cancer.

Animals↗

PubMed1997

Isozyme selective alterations of the expression of cytochrome P450 during regeneration of male rat liver following partial hepatectomy.

1. During liver regeneration in the male rat, the metabolic activities of imipramine were differentially affected depending on the specific metabolic pathways. Imipramine N-demethylation was markedly reduced whereas 2-hydroxylation showed only a moderate reduction following partial hepatectomy. 2. A slight decline was observed in the hepatic microsomal content of CYP2D apoprotein, whereas a substantial decrease occurred in CYP2C11 content during liver regeneration. Since imipramine 2-hydroxylation and N-demethylation are mediated by CYP2D and 2C11 respectively, metabolic pathway-specific alterations in the activities of imipramine metabolism are explained by the isozyme selective alteration in the levels of CYPs in regenerating liver. 3. No significant effect of regeneration was observed on expression of CYP2B1 and 2E1 apoproteins. CYP3A2 apoprotein, one of the male-specific CYP isoforms, was significantly suppressed in regenerating liver showing a similar pattern of alteration to the levels of CYP2C11. The alteration pattern of the CYP1A1 level was different to the above with a moderate decline at the first day post-operation and a marked rebound thereafter. 4. In the partially hepatectomized male rate, no significant increase in androstenedione 5-alpha reductase activity, an activity predominant in the female rat, was detected. It is concluded that the pattern of alterations of hepatic oxidative metabolism during liver regeneration was not related to the functional feminization of the liver.

Androstenedione↗

PubMed1997

Acetaldehyde as well as ethanol is metabolized by human CYP2E1.

Acetaldehyde was oxidized by rat and human hepatic microsomes in the presence of NADPH. We designated this NADPH-dependent oxidation system MAOS (microsomal acetaldehyde-oxidizing system), to distinguish it from the NAD-dependent acetaldehyde oxidation system of acetaldehyde dehydrogenase in mitochondria and cytosol. This activity was increased 2.3-fold by giving rats ethanol. Judging from the Vmax/Km values, the metabolic capacity of rat hepatic microsomes for MAOS activity was increased 24-fold by ethanol. The acetaldehyde oxidation activity of eight forms of purified rat cytochrome P450 was investigated in a reconstituted system. CYP2E1 had the highest level, followed by CYP1A2 and 4A2. Immunoinhibition studies showed that an anti-CYP2E1 antibody inhibited 90% of the MAOS activity in rats given ethanol. NADPH-dependent acetate formation was 12% or 33.6% of the NAD-dependent acetate formation in liver homogenates of control rats and those treated with ethanol, respectively. We investigated human MAOS activity further. Among the 10 forms of human cytochrome P450 expressed in yeast, CYP2E1 had especially high acetaldehyde oxidation activity. The correlation of MAOS activity with the levels of immunoreactive CYP2E1 in individual human microsomes was highly significant (r2 = 0.88, P < .01). These results indicate that hepatic CYP2E1 mainly contributes to MAOS in rats and humans, the pathway of which may play an alternative role against acetaldehyde in the liver after alcohol consumption together with acetaldehyde dehydrogenase in the metabolism of acetaldehyde.

Acetaldehyde↗