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H Jinno

Publications and source records attributed to H Jinno.

At least 37 records · Page 2Linked to original sources
PubMed1998

In vitro biotransformation of atrazine by rat liver microsomal cytochrome P450 enzymes.

We studied atrazine (ATZ) metabolism in male and female rat liver microsomes in vitro, and the major metabolite was deisopropylatrazine (DeiPr-ATZ) with deethylatrazine (DeEt-ATZ) and 1-hydroxyisopropylatrazine (iPrOH-ATZ) as minor metabolites in both sexes. The enzyme kinetics of ATZ biotransformation were examined by means of Eadie-Hofstee analyses. Although no remarkable sex difference of Michaelis Menten values for each pathway was observed, Cl(int)S (Vmax/Km) for DeiPr-ATZ, DeEt-ATZ and iPrOH-ATZ were slightly higher in female than in male rats. The formation of DeiPr-ATZ, DeEt-ATZ and iPrOH-ATZ from ATZ was substantially inhibited by SKF-525A, metyrapone, diallyl sulfide, 7-ethoxycoumarin, benzphetamine, nicotine, testosterone and lauric acid in both sexes. Cimetidine effectively inhibited the formation of all metabolites in male rats. On the other hand, the inhibition rates of the formation of DeiPr-ATZ and iPrOH-ATZ by cimetidine in female rats were lower than those in male rats, and DeEt-ATZ was hardly affected by the chemicals. In contrast with the results for cimetidine, the inhibition of ATZ biotransformation by bufuralol was more effective in female than in male rats. Anti-rat CYP2B1 and CYP2E1 antibodies effectively inhibited DeiPr-ATZ, DeEt-ATZ and iPrOH-ATZ formations in both sexes. Anti-rat CYP2C11 antibody also inhibited the three metabolites in both sexes, with the inhibition rates higher in male than in female rats, similar to cimetidine. In the case of anti-rat CYP2D1 antibody, the inhibitory effect on ATZ biotransformation in male rats was less than that in female rats. On the other hand, anti-rat CYP1A2, CYP3A2 and CYP4A1 antibodies did not affect the ATZ biotransformation in either sex. There was no significant correlation between the formation rate of ATZ metabolites and P450 isoform levels in either sex. These results may mean that CYP2B2, CYP2C11, CYP2D1 (only iPrOH-ATZ formation) and CYP2E1 in male rats, and CYP2B2, CYP2D1 and CYP2E1 in female rats are involved ATZ metabolism in liver, and that the substrate specificity of P450 isoforms for ATZ is broad.

Animals↗
PubMed1998

Suppression of male-specific cytochrome P450 isoforms by bisphenol A in rat liver.

We examined the effect of bisphenol A (BPA) on microsomal cytochrome P450 (P450) enzymes in rats. Rats were treated intraperitoneally with BPA daily for 4 days, at doses of 10, 20, and 40 mg/kg. Among the P450-dependent monooxygenase activities, testosterone 2alpha-hydroxylase (T2AH) and testosterone 6beta-hydroxylase (T6BH) activities, which are associated with CYP2C11 and CYP3A2 respectively, were remarkably decreased by 40 mg/kg BPA. The levels of the control activities were 13 and 50%, respectively. Furthermore, immunoblotting showed that BPA (20 or 40 mg/kg) significantly reduced CYP2C 11/6 and CYP3A2/1 protein levels in rat liver microsomes. In addition, estradiol 2-hydroxylase (ED2H) and benzphetamine N-demethylase (BZND) activities were significantly decreased by BPA at 20 and 40 mg/kg (by 19-73%). The Km values for T2AH and T6BH in 20 and 40 mg/kg BPA-treated rats were significantly high compared with that in control rats. The Vmax for T2AH was dose-dependently decreased by BPA treatment, whereas that of T6BH was only decreased by BPA at 40 mg/kg. On the other hand, lauric acid omega-hydroxylase (LAOH) activity was significantly increased by BPA at 20 and 40 mg/kg (1.5- and 1.7-fold, respectively). Immunoblot analysis showed that 20 and 40 mg/kg BPA induced CYP4A1/2 protein expression. However, the activities 7-ethoxyresorufin O-deethylase (EROD), 7-methoxyresorufin O-demethylase (MROD), 7-ethoxycoumarin O-deethylase (ECOD), 7-benzyloxyresorufin O-debenzylase (BROD), aminopyrine N-demethylase (APND), chlorzoxazone 6-hydroxylase (CZ6H), erythromycin N-demethylase (EMND), and testosterone 7alpha-hydroxylase (T7AH) were not affected by BPA at any dose. These results suggest that BPA affects male-specific P450 isoforms in rat liver, and that these changes closely relate to the toxicity of BPA.

Animals↗
PubMed1998

Changes in rat liver cytochrome P450 enzymes by atrazine and simazine treatment.

1. We examined the effect of two chloro-s-triazines (atrazine and simazine) on hepatic microsomal cytochrome P450 enzymes in rat. Rats were treated intraperitoneally with atrazine or simazine daily for 3 days with 100, 200 and 400 mumol/kg. 2. Among the P450-dependent monooxygenase activities, testosterone 2 alpha-hydroxylase (T2AH) activity in rat, which is associated with CYP2C11, was significantly decreased at all doses of atrazine and simazine. The levels relative to control activities were 59-46 and 60-32% respectively. Similarly, oestradiol 2-hydroxylase (ED2H) activity was also significantly decreased by 28-51% by atrazine and simazine at all doses. However, no change in CYP2C11 protein level by either chloro-s-triazine was observed. K(m) for T2AH was significantly increased only by simazine (200 mumol/kg), whereas the Vmax and Cl(int) for T2AH were significantly decreased by atrazine and simazine at all doses. 3. 7-Ethoxyresorufin O-deethylase (EROD), 7-methoxyresorufin O-demethylase (MROD) and 7-pentoxyresorufin O-depentylase (PROD) activities were significantly increased by 1.4-1.6-, 1.7-3.2- and 1.5-2.2-fold respectively, by both chloro-s-triazines at 200 or 400 mumol/kg. Lauric acid omega-hydroxylase (LAOH) was also increased by 1.4-fold by simazine at 200 and 400 mumol/kg. Immunoblotting showed that only simazine induces CYP1A2 and CYP4A1/2 protein expression. 4. The activities of 7-ethoxycoumarin O-deethylase (ECOD), bufuralol 1'-hydroxylase (BF1'H), chlorzoxazone 6-hydroxylase (CZ6H), testosterone 6 beta-hydroxylase (T6BH) and testosterone 7 alpha-hydroxylase (T7AH) were not affected by either chloro-s-triazine. 5. These results suggest that the pattern of changes in P450 isoforms by chloro-s-triazines differs between atrazine and simazine, that these herbicides change the constitutive and/or male specific P450 isoform(s) in rat liver, and that these changes closely relate to the toxicity of chloro-s-triazines.

Animals↗
PubMed1998

Changes in hepatic cytochrome P450 enzymes by cis- and trans-1,2-dichloroethylenes in rat.

1. We examined the effect of cis- and trans-1,2-dichloroethylenes (DCEs) on hepatic microsomal cytochrome P450 (P450) enzymes in the male and female rat. Rats were treated intraperitoneally with 7.5 mmol/kg cis- or trans-1,2-DCE daily for 4 days. 2. Among the hepatic microsomal P450-dependent monooxygenase activities tested, testosterone 2 alpha-hydroxylase (T2AH) activity in the male rat, which is associated with CYP2C11, was decreased by both cis- and trans-1,2-DCE isomers. The levels to control activities were 53 and 63% respectively. Furthermore, immunoblotting showed that both isomers significantly reduced the CYP2C11/6 protein level in liver microsomes from the male. The levels of testosterone 6 beta-hydroxylase (T6BH) activity and CYP3A2/1 protein in the male rat were reduced by cis-1,2-DCE but not trans-1,2-DCE. 3. cis-1,2-DCE decreased ethoxycoumarin O-deethylase (ECOD), benzyloxyresorufin O-debenzylase (BROD), chlorzoxazone 6-hydroxylase (CZ6H) and testosterone 7 alpha-hydroxylase (T7AH) activities in the male rat by 29, 28, 34 and 27% respectively. On the other hand, trans-1,2-DCE significantly increased 7-ethoxyresorufin O-deethylase (EROD) and 7-methoxyresorufin O-demethylase (MROD) activities in the male rat 1.4- and 1.9-fold respectively. Immunoblotting showed that cis-1,2-DCE reduced CYP1A1/2 and CYP2B1/2 protein levels, whereas trans-1,2-DCE increased CYP1A1/2 and CYP2B1/2 protein levels in the male rat. 4. The activities of other P450-dependent monooxygenases, namely benzphetamine N-demethylase (BZND), aminopyrine N-demethylase (APND), erythromycin N-demethylase (EMND) and lauric acid omega-hydroxylase (LAOH), in the male rat were hardly affected by either 1,2-DCE isomer. In the female rat there were no apparent changes in P450-dependent monooxygenase activities upon treatment with the 1,2-DCE isomers. 5. These results suggest that 1,2-DCEs mainly affect male-specific P450 isoforms in the rat liver and that these changes may relate to the toxicity of 1,2-DCEs.

Animals↗
PubMed1997

Molecular Targeting for Epidermal Growth Factor Receptor Expressed on Breast Cancer Cells by Human Fusion Protein.

Recombinant human ribonuclease 1 (RNase 1) was chemically linked to recombinant human epidermal growth factor (EGF). The cytotoxicity of this conjugate was assayed using MTT assay. The EGF-RNase conjugate showed dose-dependent cytotoxicity against breast and squamous cell carcinomas overexpressing the EGF receptor (EGFR). The cytotoxicity of the conjugate correlated positively with the level of EGFR expression by each cell line. These results suggest that the EGF-RNase conjugate is a more effective anticancer agent with less immunogenicity and toxicity than conventional chimeric breast cancer toxins.

Journal Article↗
PubMed1997

Hepatotoxicity of diisopropyl ester of malonic acid and chloromalonic acids, disinfection by-products of the fungicide isoprothiolane.

The fungicide isoprothiolane (diisopropyl 1,3-dithiolane-2-ylidenemalonate) decomposes to the diisopropyl esters of malonic acid (DM), chloromalonic acid (DCM) and dichloromalonic acid (DDCM) upon aqueous chlorination. In this study, the cytotoxicity of these compounds was examined using rat hepatocytes cultured on Matrigel. DCM and DDCM caused hepatocellular death at concentrations > 0.5 mM, while DM had no effect on the cell viability even at the maximum concentration examined (4 mM). Significant lipid peroxidation, measured as 2-thiobarbituric acid reactive substances, was observed in both DCM- and DDCM-treated hepatocyte cultures, and was significantly enhanced by pretreatment with 0.1 mM bis(p-nitrophenyl)phosphate (BNPP), a carboxylesterase inhibitor. When both BNPP and SKF-525A, a cytochrome P450 inhibitor, were present in the medium, DCM-induced cytotoxicity and lipid peroxidation were significantly suppressed compared to cultures with BNPP-treatment alone. By contrast, the DDCM-induced cytotoxicity was not affected by the combined pretreatment of SKF-525A and BNPP. These results indicate that DCM is metabolically activated by cytochrome P450 in an ester form, while DDCM is activated by a mechanism other than one involving cytochrome P450. To further elucidate the cytochrome P450 isozyme involved in the metabolic activation of DCM, microsomal lipid peroxidation was studied in vitro using microsomes from rats treated with beta-naphthoflavone, musk xylene, phenobarbital, pyrazole, or dexamethasone. Among these preparations, the microsomes from dexamethasone-treated rats showed the most extensive lipid peroxidation in the presence of DCM, and the lipid peroxidation was enhanced by BNPP as observed in hepatocyte cultures. These findings suggest the possible involvement of cytochrome P450 3A in the metabolic activation of DCM.

Animals↗
PubMed1997

Changes in cytochrome P450 enzymes by 1,1-dichloroethylene in rat liver and kidney.

We examined the effect of 1,1-dichloroethylene (1,1-DCE) on microsomal cytochrome P450 (P450) enzymes in rat liver and kidney. Rats were treated intraperitoneally with 1,1-DCE daily for 4 days, at doses of 200, 400, and 800 mg/kg. Among the P450-dependent monooxygenase activities in liver microsomes, testosterone 2alpha-hydroxylase (T2AH), which is associated with CYP2C11 activity, was remarkably decreased by 800 mg/kg 1,1-DCE. The level relative to control activity was < 10%. Furthermore, immunoblotting showed that 1,1-DCE (> or = 400 mg/kg) significantly decreased CYP2C11/6 protein levels in liver microsomes. In addition, 7-methoxyresorufin O-demethylase (MROD), 7-ethoxycoumarin O-deethylase (ECOD), benzphetamine N-demethylase (BZND), chlorzoxazone 6-hydroxylase (CZ6H), and testosterone 6beta-hydroxylase (T6BH) activities were significantly decreased by the highest dose of 1,1-DCE (by 40-70%). However, the activities of other P450-dependent monooxygenases, namely 7-ethoxyresorufin O-deethylase (EROD), 7-benzyloxyresorufin O-debenzylase (BROD), aminopyrine N-demethylase (APND), erythromycin N-demethylase (EMND), lauric acid omega-hydroxylase (LAOH), and testosterone 7alpha-hydroxylase (T7AH) were not affected by 1,1-DCE at any dose. Immunoblotting showed CYP1A1/2, CYP2B1/2, CYP2E1, and CYP3A2/1 protein levels were significantly decreased by 60-66% by 1,1-DCE (800 mg/kg), whereas that of CYP4A1/2 was not affected by any dose of 1,1-DCE. By contrast, among the P450-dependent monooxygenase activities in kidney microsomes, only CZ6H activity was increased by 1,1-DCE (1.6-fold at 800 mg/kg). Also, it was observed that 1,1-DCE (800 mg/kg) significantly increased CYP2E1 protein levels by immunoblotting (approximately 1.5-fold). These results suggest that 1,1-DCE changes the constitutive P450 isoforms in the rat liver and kidney, and that these changes closely relate to the toxicity of 1,1-DCE.

7-Alkoxycoumarin O-Dealkylase↗
PubMed1997

Effect of 2,4,4'-trichloro-2'-hydroxydiphenyl ether on cytochrome P450 enzymes in the rat liver.

We examined the effect of 2,4,4'-trichloro-2'-hydroxydiphenyl ether (Irgasan DP300) on microsomal cytochrome P450 (P450) enzymes in rat liver. Rats were treated intraperitoneally with Irgasan DP300 daily for 4 days, at doses of 0.2, 0.4 and 0.8 mmol/kg. Among the P450-dependent monooxygenase activities, 7-benzyloxyresorufin O-debenzylase (BROD) and 7-pentoxyresorufin O-depentylase (PROD) in rats, which are associated with CYP2B1, were remarkably induced by all doses of Irgasan DP300. The relative induction to each control activity were from 5.6- to 22.3-fold and 4.9- to 20.2-fold, respectively. Furthermore, immunoblotting showed that CYP2B1/2 protein level in rat liver microsomes was increased from 10.8- to 34.4-fold by Irgasan DP300. In addition, 7-ethoxycoumarin O-deethylase (ECOD) and p-nitrophenol hydroxylase (PNPH) activities were significantly increased by Irgasan DP300 at all doses (from 1.4- to 4.9-fold). Although the activities of other P450-dependent monooxygenases, namely aminopyrine N-demethylase (APND), aniline p-hydroxylase (ANPH), erythromycin N-demethylase (EMND), lauric acid omega-hydroxylase (LAOH) and testosterone 6 beta-hydroxylase (TS6BH) were increased at high doses (> or = 0.4 mmol/kg) of Irgasan DP300, the relative level was lower than those of the CYP2B1-dependent monooxygenases such as BROD and PROD. However, 7-ethoxyresorufin O-deethylase (EROD), 7-methoxyresorufin O-demethylase (MROD), testosterone 2 alpha-hydroxylase (TS2AH) and testosterone 7 alpha-hydroxylase (TS7AH) activities were not affected by any doses of Irgsan DP300. Immunoblotting showed that CYP3A2/1 and CYP4A1 protein levels were significantly induced from 1.3- to 2.2-fold by Irgasan DP300 (> or = 0.4 mmol/kg), whereas those of CYP1A1/2, CYP2C11/6 and CYP2E1 were not affected by any doses of Irgasan DP300. These results suggest that Irgasan DP300 induces the P450 isoforms of CYP2B subfamily in the rat liver, and that the induced P450 isozymes closely relates to the toxicity of Irgasan DP300 or its chlorinated derivatives.

7-Alkoxycoumarin O-Dealkylase↗
PubMed1997

Irgasan DP 300 (5-chloro-2-(2,4-dichlorophenoxy)-phenol) induces cytochrome P450s and inhibits haem biosynthesis in rat hepatocytes cultured on Matrigel.

1. The effect of Irgasan DP 300 (5-chloro-2-(2,4-dichlorophenoxy)phenol) on cytochrome P450 (P450) induction and haem biosynthesis was studied in rat hepatocytes cultured on Matrigel. 2. Irgasan DP 300 significantly induced 7-benzyloxyresorufin O-debenzylase activity, followed by 7-pentoxyresorufin O-depentylase and 7-ethoxyresorufin O-deethylase activities. 4-Nitrophenol hydroxylase, testosterone 6 beta-hydroxylase and methoxyresorufin O-demethylase activities were also slightly increased. The maximum induction of these enzyme activities was obtained at the same concentration of 125 microM in the culture medium. 3. Immunochemical blots using anti-rat cytochrome P450 antibodies revealed that Irgasan DP 300 preferably induced CYP2B1/2 along with a slight increase in 3A. These results indicate that Irgasan DP 300 is a phenobarbital-type inducer. 4. In the absence of exogenous 5-aminolevulinic acid (ALA), slight increases in protoporphyrin IX (2.6-fold) and coproporphyrin III (1.3-fold) were observed in the Irgasan DP 300-treated cultures. In contrast, when 75 microM ALA was present, Irgasan DP 300 (250 microM) caused an extensive accumulation of uroporphyrin I (13-fold). 5. Irgasan DP 300 inhibited rat hepatic uroporphyrinogen III synthase in vitro. 6. These results indicate that Irgasan DP 300 produced accumulation of hydroxymethylbilane in rat hepatocytes by inhibiting uroporphyrinogen III synthase, and consequently an accumulation of uroporphyrin I.

Aminolevulinic Acid↗
PubMed1996

Effectiveness of an adriamycin immunoconjugate that recognizes the C-erbB-2 product on breast cancer cell lines.

Adriamycin (ADM) was chemically conjugated to a murine monoclonal antibody, A0011, which recognizes the c-erbB-2 product, via a disulfide bond using N-succinimidyl-3-(2-pyridyldithio) propionate (SPDP) and 2-iminothiolane (2-IT). The molar ratio of ADM to the monoclonal antibody ranged from 15:1 to 25:1 and enzyme-linked immunosorbent assay (ELISA) showed that the binding activity of the conjugate was almost retained. We compared the efficacy of A0011 alone, ADM alone, the A0011-ADM conjugate, against the human breast cancer cell lines SK-BR-3, MDA-MB-361, MCF-7, and BT-20. The A0011-ADM conjugate was observed to be ten times more cytotoxic to the cell lines overexpressing the c-erbB-2 product, namely, SK-BR-3 and MDA-MB-361, than free ADM, but it showed weak cytotoxicity against the cell lines with a low level of c-erbB-2 product expression, namely, MCF-7 and BT-20. However, free A0011 and nonspecific murine IgM-ADM conjugate showed no cytotoxicity toward any of the four cell lines, while the addition of a tenfold molar excess of A0011 inhibited conjugate cytotoxicity. These data suggest that conjugate cytotoxicity is antibody-mediated. Moreover, conjugate cytotoxicity at 10(-6)M was correlated with antigen volume, and the data were fitted to the regression equation y = -11.63logX + 116.38 where the correlation coefficient = 0.950. Our results indicate that targeting therapy aiming at the c-erbB-2 product may be useful in the treatment of breast cancers overexpressing the c-erbB-2 product.

Antibiotics, Antineoplastic↗
PubMed1996

Epidermal growth factor receptor-dependent cytotoxic effect by an EGF-ribonuclease conjugate on human cancer cell lines--a trial for less immunogenic chimeric toxin.

Mammalian pancreatic ribonuclease (RNase) was conjugated chemically via a disulfide bond to human or murine epidermal growth factor (EGF). The conjugation between EGF and RNase was ascertained by SDS-PAGE using reduced and nonreduced conjugates. The EGF-RNase conjugate retained potent RNase activity and competed with 125I-EGF for binding to EGFR to the same extent as unconjugated EGF. Both the human and murine EGF-RNase conjugates showed dose-dependent cytotoxicity against EGFR-overexpressing A431 human squamous carcinoma cells with IC50 values of 3 x 10(-7) M and 6 x 10(-7) M, respectively, whereas free RNase had an IC50 of 10(-4) M. Against the EGFR-deficient small-cell lung cancer cell line H69, the EGF-RNase conjugate had no cytotoxic effect. The Human EGF-RNase conjugate showed dose-dependent cytotoxicity against other squamous carcinoma cell lines (TE-5, TE-1) and breast cancer cell lines (BT-20, SK-BR-3, MCF-7) and the cytotoxicity of the conjugate correlated positively with the level of expression of EGFR by each cell line. An unconjugated mixture of EGF and RNase had no greater effect than RNase alone on any cell line. Excess free EGF blocked EGF-RNase conjugate cytotoxicity against A431 cells. These results suggest that the EGF-RNase conjugate may be a more effective anticancer agent with less immunogenicity than coventional chimeric toxins.

Antineoplastic Agents↗
PubMed1996

Epidermal growth factor receptor-dependent cytotoxicity for human squamous carcinoma cell lines of a conjugate composed of human EGF and RNase 1.

Recombinant human ribonuclease 1 (RNase 1) was chemically linked to recombinant human epidermal growth factor (EGF). The EGF-RNase conjugate showed dose-dependent cytotoxicity for EGF receptor-overexpressing A431 and TE-8 human squamous carcinoma cells with an IC50 of 2 x 10(-7)M and 10(-6)M, respectively, whereas the IC50 of RNase alone was almost 10(-4)M. An unconjugated mixture of EGF and RNase had no greater effect than RNase alone. The conjugate showed no detectable cytotoxicity against EGF receptor-deficient small cell lung cancer cells (H69). Addition of excess EGF in the medium protected A431 cells from the EGF-RNase conjugate cytotoxicity. The cytotoxicity of the EGF-RNase conjugate was positively correlated with the EGF receptor numbers of each cell line. The chimeric toxin composed of only human proteins might be a more useful anti-cancer agent with less immunogenicity than the conventional chimeric toxins.

Carcinoma, Squamous Cell↗
PubMed1996

Interaction of 2,4,4'-trichloro-2'-hydroxydiphenyl ether with microsomal cytochrome P450-dependent monooxygenases in rat liver.

We studied the effects of 2,4,4'-trichloro-2'-hydroxydiphenyl ether (Irgasan DP300) on the kinetics of the cytochrome P450 (P450)-dependent monooxygenases in rat liver microsomes. The activities of 7-ethoxyresorufin O-deethylase (EROD) and 7-pentoxyresorufin O-depentylase (PROD) in rat liver microsomes exposed to 3-methylcholanthrene (MC) and phenobarbital (PB) respectively, were substantially inhibited by Irgasan DP300. The inhibition profile of EROD was competitive, whereas that of PROD was noncompetitive; the Ki values from Hanes plots were 0.24 and 1.48 microM for EROD and PROD, respectively. Phenacetin O-deethylase (PCOD) and 4-nitrophenol hydroxylase (4NPH) activities in rats exposed to PB were also inhibited by Irgasan DP300, at Ki values lower than those for other microsomes. Irgasan DP300 slightly inhibited testosterone 6 beta-hydroxylase (TS6BH) activities in some microsomes. No effect of Irgasan DP300 on lauric acid omega-hydroxylase (LAOH) activity was evident in any microsomal preparations. These results indicated that Irgasan DP300 inhibits MC- and PB-inducible P450-dependent monoxygenase in vitro competitively or noncompetitively, and that the P450 enzymes of the CYP1A or CYP2B subfamily may contribute to Irgasan DP300 toxicity.

Animals↗
PubMed1995

Induction of rat liver drug-metabolizing enzymes by tetrachloroethylene.

The effect of tetrachloroethylene on Phase I and II drug-metabolizing enzymes in rat liver was examined. Rats were treated orally with tetrachloroethylene daily for five days, at doses of 125, 250, 500, 1,000 and 2,000 mg/kg. The higher doses (> 500 mg/kg) of tetrachloroethylene induced the hepatic microsomal 7-pentoxyresorufin O-depentylase and 7-benzyloxyresorufin O-debenzylase activities associated with the CYP2B subfamily. 7-ethoxyresorufin O-deethylase activity was also induced about 2-fold compared with that of control rats at 500, 1,000, and 2,000 mg/kg dose levels of tetrachloroethylene. However, 7-ethoxycoumarin O-deethylase and 7-methoxyresorufin O-demethylase activities were increased significantly at only the 1,000 mg/kg dose level of tetrachloroethylene (1.4- and 1.5-fold). Although other cytochrome P450-mediated monooxygenase activities such as nitrosodimethylamine N-demethylase, aminopyrine N-demethylase and erythromycin N-demethylase were also induced by tetrachloroethylene, the relative induction to control activity was lower than those of 7-pentoxyresorufin O-depentylase and 7-benzyloxyresorufin O-debenzylase. Western immunoblotting showed that the levels of CYP2B1 and CYP2B2 proteins in liver microsomes were increased at doses of 1,000 and 2,000 mg/kg of tetrachloroethylene. In addition to cytochrome P450-mediated monooxygenases, there was significant induction of the Phase II drug-metabolizing enzymes, DT-diaphorase, glutathione S-transferase activities towards 1-chloro-2,4-dinitrobenzene and 1,2-dichloro-4-nitrobenzene, and UDP-glucuronyltransferase activities towards 4-nitrophenol and 7-hydroxycoumarin. The results indicate that tetrachloroethylene induces both Phase I (CYP2B-mediated monooxygenase) and Phase II drug-metabolizing enzymes (DT-diaphorase, glutathione S-transferase and UDP-glucuronyltransferase) in the rat liver.

Administration, Oral↗
PubMed1995

Induction of hepatic drug-metabolizing enzymes by chlornitrofen (CNP) and CNP-amino in rats and mice.

The induction of hepatic drug-metabolizing enzymes by chlornitrofen (CNP) and CNP-amino was studied in the liver of male rats and mice. CNP-amino increased the activities of 7-pentoxyresorufin O-depentylase (PROD) and 7-benzyloxyresorufin O-debenzylase (BROD) as CYP2B1-dependent monooxygenase 3.6- and 4.1-fold in rats. On the contrary, these enzyme activities in mice were induced by CNP rather than by CNP-amino. Furthermore, immunoblotting showed that the protein levels of CYP2B subfamily cytochrome P450 (P450) in liver microsomes of rats and mice were increased by CNP or CNP-amino. Phase II drug-metabolizing enzymes, UDP-glucuronyltransferase (UGT) and glutathione S-transferase (GST) levels in mice were also significantly increased from 1.4 to 2.5-fold by CNP or CNP-amino. However, neither CNP nor CNP-amino affected UGT and GST in rats. These results suggest that CNP and or CNP-amino induce the P450 isoforms of CYP2B subfamily in the rat and mouse liver, and that the inducibility of drug-metabolizing enzyme by the compounds is different between rats and mice.

Animals↗
PubMed1995

Dog liver microsomal P450 enzyme-mediated toluene biotransformation.

1. We studied toluene metabolism in dog liver microsomes and the major metabolite was benzyl alcohol with o- and p-cresol as minor metabolites. 2. The enzyme kinetics of toluene biotransformation were examined by means of Lineweaver-Burk analyses. The Michaelis-Menten values differed among the three pathways, the order being; Km, o-cresol > p-cresol > benzyl alcohol; Vmax, benzyl alcohol > o-cresol > p-cresol; and Cl(int), benzyl alcohol > p-cresol > o-cresol. 3. The formation of benzyl alcohol, o- and p-cresol from toluene was substantially inhibited by the P4502E inhibitors such as DDC (diethyldithiocarbamate) and 4-methylpyrazole in all pathways, with IC50's in the range of 0.02-0.59 mM. The P4502B inhibitors, metyrapone and secobarbital also inhibited benzyl alcohol and p-cresol formation, whereas o-cresol was not inhibited by these latter compounds. 4. Anti-rat P4502E1 antibodies inhibited benzyl alcohol, o- and p-cresol formation from 26 to 30% 0.2 ml serum/mg microsomal protein. Furthermore, anti-rat P4502B1/2 antibody inhibited benzyl alcohol and p-cresol formation (47 and 44% respectively), but not that of o-cresol. Anti-rat P4502C11/6 antibody also inhibited benzyl alcohol and p-cresol formation 31 and 24% respectively in a similar manner to that by the anti-rat P4502B1/2 antibody. 5. These results suggested that the P4502B, 2C and 2E isozymes in dog liver contribute to the formation of benzyl alcohol and p-cresol from toluene, and 2E isozyme preferentially contributes to the formation of o-cresol.

Animals↗
PubMed1995

Interaction of tetrachloroethylene with rat hepatic microsomal P450-dependent monooxygenases.

1. We have studied the effects of tetrachloroethylene (PCE) on the kinetics of the P450-dependent monooxygenases in rat liver microsomes. 2. 7-Pentoxyresorufin O-depentylase (PROD) and 7-benzyloxyresorufin O-debenzylase (BROD) activities in phenobarbital (PB)-treated rat liver microsomes were substantially inhibited by PCE. The inhibition profiles were non-competitive for both enzyme activities; Ki's from Eadie-Hofsee plots were 0.16 and 0.29 mM for PROD and BROD respectively. In contrast, the enzyme activities in untreated, beta-naphthoflavone (BNF)-, isoniazid (ISN)- and pregnenolone-16 alpha-carbonitrile (PCN)-induced microsomes were not affected by PCE. 3. 7-Ethoxycoumarin O-deethylase (ECOD) activity in PB-induced microsomes was competitively inhibited by PCE, with a Ki that was lower than those of other microsomes. 4. PCE inhibited 7-ethoxyresorufin O-deethylase (EROD) activities in some microsomes slightly. The Ki for PCE was the lowest in untreated, followed by ISN-treated microsomes. 5. No effect of PCE upon aniline 4-hydroxylase (AN4H) and testosterone 6 beta-hydroxylase (TS6BH) activities was evident in any microsomal preparation. 6. These results indicate that PCE inhibits PB-inducible, P450-dependent monooxygenases in vitro non-competitively or competitively, and that the P450 enzymes of the P4502B subfamily may contribute to PCE toxicity.

Animals↗