Oxidation of esterified arachidonate by rat liver microsomes

International Nuclear Information System (INIS)

Davis, H.W.; Suzuki, T.; Schenkman, J.B.

1986-01-01

The authors have previously demonstrated a relationship between phospholipid arachidonate in liver microsomes and malondialdehyde (MDA) formation during lipid peroxidation. In this study arachidonic acid (U- 14 C) was incorporated into rat liver microsomes and NADPH-supported peroxidation was carried out at 37 0 C for 15 minutes. The microsomes were pelleted by centrifugation and the labeled products in the supernatant were isolated by a solid phase method. Pellets were hydrolyzed with phospholipase A 2 and extracted with diethyl ether and the products from both fractions were separated by reverse phase HPLC. The results show that (1) oxidation occurs in all of the major phospholipids but that phosphatidylethanolamine is the most susceptible; (2) a linear correlation exists between MDA formation and supernatant radioactivity; (3) several different polar products are found in both the supernatant and the hydrolyzed pellet but that the ratios of product peaks in HPLC do not change during the peroxidation, indicating no secondary metabolism or propagation; and (4) cytochrome P-450 is not involved in the peroxidative reactions since no oxidation occurs in the absence of Fe 3+ and since product formation is unaffected in the presence of carbon monoxide

Comparative Metabolism Study of Five Protoberberine Alkaloids in Liver Microsomes from Rat, Rhesus Monkey, and Human.

Science.gov (United States)

Li, Yan; Zhou, Yanyan; Si, Nan; Han, Lingyu; Ren, Wei; Xin, Shaokun; Wang, Hongjie; Zuo, Ran; Wei, Xiaolu; Yang, Jian; Zhao, Haiyu; Bian, Baolin

2017-11-01

Protoberberine alkaloids including berberine, palmatine, jatrorrhizine, coptisine, and epiberberine are major components in many medicinal plants. They have been widely used for the treatment of cancer, inflammation, diabetes, depression, hypertension, and various infectious areas. However, the metabolism of five protoberberine alkaloids among different species has not been clarified previously. In order to elaborate on the in vitro metabolism of them, a comparative analysis of their metabolic profile in rat, rhesus monkey, and human liver microsomes was carried out using ultrahigh-performance liquid chromatography coupled with a high-resolution linear trap quadrupole-Orbitrap mass spectrometer (UHPLC-electrospray ionization-Orbitrap MS) for the first time. Each metabolite was identified and semiquantified by its accurate mass data and peak area. Fifteen metabolites were characterized based on accurate MS/MS spectra and the proposed MS/MS fragmentation pathways including demethylation, hydroxylation, and methyl reduction. Among them, the content of berberine metabolites in human liver microsomes was similar with those in rhesus monkey liver microsomes, whereas berberine in rat liver microsomes showed no demethylation metabolites and the content of metabolites showed significant differences with that in human liver microsomes. On the contrary, the metabolism of palmatine in rat liver microsomes resembled that in human liver microsomes. The content of jatrorrhizine metabolites presented obvious differences in all species. The HR-ESI-MS/MS fragmentation behavior of protoberberine alkaloids and their metabolic profile in rat, rhesus monkey, and human liver microsomes were investigated for the first time. The results demonstrated that the biotransformation characteristics of protoberberine alkaloids among different species had similarities as well differences that would be beneficial for us to better understand the pharmacological activities of protoberberine alkaloids

Studies on the transverse localization of lysophospholipase II in bovine liver microsomes by immunological techniques

NARCIS (Netherlands)

Moonen, H.; Bosch, H. van den

1979-01-01

1. 1. Lysophospholipase activity solubilized from bovine liver microsomes could be precipitated for more than 80% by antibodies evoked in rabbits against the purified bovine liver lysophospholipase II. 2. 2. After solubilization of the microsomes in 1.5% sodium deoxycholate, an immunoprecipitate

[Immunosuppressant effect of cyclophosphamide activated in vitro by liver microsomes from different strains of mice].

Science.gov (United States)

Telegin, L Iu; Zhirnov, G F; Mazurov, A V; Pevnitskiĭ, L A

1981-07-01

The paper is concerned with activation of cyclophosphamide by mouse liver microsomes in vitro. Liver microsomes from BALB/c mice metabolize cyclophosphamide more effectively as compared with those from DBA/2 mice, which manifested by a more intense output of products having alkylating or immunodepressant properties. This seems likely to be a consequence of the increased P-450 cytochrome content in liver microsomes from BALB/c mice, as well as of its structural characteristics in the mouse. The relationship between the immunodepressant effect of cyclophosphamide in vivo and in vitro in mice of varied genotypes is discussed.

A human cytochrome P-450 is recognized by anti-liver/kidney microsome antibodies in autoimmune chronic hepatitis.

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Kiffel, L; Loeper, J; Homberg, J C; Leroux, J P

1989-02-28

1- Anti-liver/kidney microsome autoantibodies type 1 (anti-LKM1), observed in some children with chronic active hepatitis, were used to isolate their antigen in human liver microsomes. A protein, called P-LKM1 was thus purified. This protein was recognized by a rabbit antiserum directed against the related human cytochromes P-450 bufI and P-450 bufII. 2- A human liver microsomal protein immunoprecipitated with anti-LKM1 sera was also recognized by anti cytochromes P-450 bufI/II antibodies. 3- Anti-LKM1 antibodies potently inhibited microsomal bufuralol 1'-hydroxylation. These results displayed the possible identity between cytochrome P-450 bufI/II and LKM1 antigen.

Glutathione delays varies as-tocopherol oxidation and subsequent lipid peroxidation in rat liver microsomes

International Nuclear Information System (INIS)

Robey, S.; Mavis, R.

1986-01-01

A method has been developed for in vitro trace radiolabeling of rat liver microsomes with 3 H-α-tocopherol (αT*) which allows virtually complete oxidation of the αT* under oxidizing conditions. The supernatant of a 16,000 xg centrifugation of homogenized rat liver, containing the cytosolic rat liver vitamin E (VE) transfer protein, was incubated with an ethanolic solution of αT* for 10 minutes at 37 0 C. Labeled microsomes were collected in the washed 100,000 xg pellet. Microsomes were then incubated with 30 μM Fe 2+ in an NADPH-generating system, and both production of malondialdehyde (MDA) (a product of lipid peroxidation) and oxidation of αT* were monitored over a time course in the presence and absence of glutathione (GSH). The results indicate virtually complete oxidation of αT* precedes significant membrane lipid peroxidation, and that addition of 5 mM GSH delays both αT* oxidation and subsequent MDA production. This suggests that the previously observed VE-dependent heat labile inhibition of microsomal lipid peroxidation by GSH involves maintaining membrane levels of α-tocopherol

The Metabolism of Separase Inhibitor Sepin-1 in Human, Mouse, and Rat Liver Microsomes

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Feng Li

2018-05-01

Full Text Available Separase, a known oncogene, is widely overexpressed in numerous human tumors of breast, bone, brain, blood, and prostate. Separase is an emerging target for cancer therapy, and separase enzymatic inhibitors such as sepin-1 are currently being developed to treat separase-overexpressed tumors. Drug metabolism plays a critical role in the efficacy and safety of drug development, as well as possible drug–drug interactions. In this study, we investigated the in vitro metabolism of sepin-1 in human, mouse, and rat liver microsomes (RLM using metabolomic approaches. In human liver microsomes (HLM, we identified seven metabolites including one cysteine–sepin-1 adduct and one glutathione–sepin-1 adduct. All the sepin-1 metabolites in HLM were also found in both mouse and RLM. Using recombinant CYP450 isoenzymes, we demonstrated that multiple enzymes contributed to the metabolism of sepin-1, including CYP2D6 and CYP3A4 as the major metabolizing enzymes. Inhibitory effects of sepin-1 on seven major CYP450s were also evaluated using the corresponding substrates recommended by the US Food and Drug Administration. Our studies indicated that sepin-1 moderately inhibits CYP1A2, CYP2C19, and CYP3A4 with IC50 < 10 μM but weakly inhibits CYP2B6, CYP2C8/9, and CYP2D6 with IC50 > 10 μM. This information can be used to optimize the structures of sepin-1 for more suitable pharmacological properties and to predict the possible sepin-1 interactions with other chemotherapeutic drugs.

An integrated study for the utilization of anthraquinone compounds extract “Heshouwu” In vivo and their comparative metabolism in liver microsomes using UPLC-ESI-Q-TOF/MSn

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Sha Chen

2018-01-01

Full Text Available Objective: Anthraquinone (AQ, a major bioactive component of the traditional Chinese medicine HeShouWu, has widespread applications in industry and medicine. The objective of the current study is to explore the differences in the bioavailability of anthraquinones in vivo and the metabolism in liver microsomes. Materials and Methods: In vivo, we used a reliable UPLC-ESI-QqQ-MS/MS method to measure seven AQ compounds in the jugular vein plasma of rats following oral administration of HeShouWu. Furthermore, in order to quantify the bioavailability of AQs in vivo and to further understand the metabolism of these compounds, we compared the in vitro metabolism of AQ in different species with respect to metabolic profiles, the enzymes involved, and catalytic efficiency using liver microsomes from human (HLM, mouse (MLM, rat (RLM, and beagle dog (DLM. Results: We identified two metabolic pathways, including the hydroxylation and glucuronidation of AQ, in the liver microsomes of humans and other species using UPLC-ESI-Q-TOF. We found that substitutions on the AQ ring were crucial to the activity and regioselectivity of its hydroxylation. In general, hydroxylation activity decreased greatly with β-COOH (rhein and enhanced dramatically with β-OH (emodin. We also found that glucuronidation of the compound emodin-8-O-β-D-glucoside acts as the main isoform in AQ hydroxylation in HLM and DLM. Total microsomal intrinsic clearance values for AQ were greatest in mouse microsomes, followed by those in dog, human, and rat microsomes. Conclusion: The absorption of different anthrquinone compounds varied based on the compound structure, the metabolism types and products of anthraquinones in liver microsomes were different in different species. These findings provide vital information for a deeper unuunderstanding of the metabolism of AQs.

Purification and characterization of an amidohydrolase for N4-long-chain fatty acyl derivatives of 1-beta-D-arabinofuranosylcytosine from mouse liver microsomes.

Science.gov (United States)

Hori, K; Tsuruo, T; Tsukagoshi, S; Sakurai, Y

1984-03-01

N4-Long-chain fatty acyl-1-beta-D-arabinofuranosylcytosine amidohydrolase, a metabolizing enzyme for N4-acyl derivatives of 1-beta-D-arabinofuranosylcytosine with long-chain fatty acids, was purified from mouse liver microsomes. The purification was accomplished by solubilization of liver microsomes with Triton X-100, diethylaminoethyl cellulose chromatography, gel filtrations, hydroxyapatite chromatography, and concanavalin A:Sepharose chromatography. On sodium dodecyl sulfate:polyacrylamide gel electrophoresis, the purified enzyme preparation produced a single protein band with a molecular weight of 54,000. The enzyme had an optimal pH of 9.0, and the Michaelis constant for N4-palmitoyl-1-beta-D-arabinofuranosylcytosine was 67 microM. The thiols such as dithiothreitol or 2-mercaptoethanol stabilized the enzyme and stimulated its activity. p-Chloromercuribenzoate, N-ethylmaleimide, diisopropylfluorophosphate, and phenylmethylsulfonyl fluoride strongly inhibited the reaction. Bovine serum albumin markedly stimulated the enzyme activity, whereas detergents such as Triton X-100, deoxycholate, and sodium dodecyl sulfate had little effect. The enzyme did not require monovalent or divalent cations. Among the series of N4-acyl derivatives of 1-beta-D-arabinofuranosylcytosine with different chain lengths of acyl residues, the purified enzyme preferentially hydrolyzed the derivatives with long-chain fatty acids (C12 to C18), and N4-palmitoyl-1-beta-D-arabinofuranosylcytosine was the most susceptible. The purified enzyme was inactive on various N-acylamino acids, amides, oligopeptides, proteins, N-acylsphingosines (ceramides), triglyceride, lecithin, and lysolecithin. These results suggest that N4-long-chain fatty acyl-1-beta-D-arabinofuranosylcytosine amidohydrolase may be a new type of linear amidase.

Cranberry juice suppressed the diclofenac metabolism by human liver microsomes, but not in healthy human subjects

Science.gov (United States)

Ushijima, Kentarou; Tsuruoka, Shu-ichi; Tsuda, Hidetoshi; Hasegawa, Gohki; Obi, Yuri; Kaneda, Tae; Takahashi, Masaki; Maekawa, Tomohiro; Sasaki, Tomohiro; Koshimizu, Taka-aki; Fujimura, Akio

2009-01-01

AIM To investigate a potential interaction between cranberry juice and diclofenac, a substrate of CYP2C9. METHODS The inhibitory effect of cranberry juice on diclofenac metabolism was determined using human liver microsome assay. Subsequently, we performed a clinical trial in healthy human subjects to determine whether the repeated consumption of cranberry juice changed the diclofenac pharmacokinetics. RESULTS Cranberry juice significantly suppressed diclofenac metabolism by human liver microsomes. On the other hand, repeated consumption of cranberry juice did not influence the diclofenac pharmacokinetics in human subjects. CONCLUSIONS Cranberry juice inhibited diclofenac metabolism by human liver microsomes, but not in human subjects. Based on the present and previous findings, we think that although cranberry juice inhibits CYP2C9 activity in vitro, it does not change the pharmacokinetics of medications metabolized by CYP2C9 in clinical situations. PMID:19694738

Major antigen of liver kidney microsomal autoantibodies in idiopathic autoimmune hepatitis is cytochrome P450db1.

OpenAIRE

Manns, M P; Johnson, E F; Griffin, K J; Tan, E M; Sullivan, K F

1989-01-01

Type 1, liver kidney microsomal autoantibodies (LKM-1) are associated with a subgroup of idiopathic autoimmune type, chronic active hepatitis (CAH). The antigenic specificity of LKM-1 autoantibodies from 13 patients was investigated by immunoblot analysis of human liver microsomal proteins. Polypeptides of 50, 55, and 64 kD were detected with these antisera. A high titer LKM-1 serum was selected to screen a human liver lambda gt11 cDNA expression library, resulting in the isolation of several...

Elevated Liver Enzymes

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Symptoms Elevated liver enzymes By Mayo Clinic Staff Elevated liver enzymes may indicate inflammation or damage to cells in the liver. Inflamed or ... than normal amounts of certain chemicals, including liver enzymes, into the bloodstream, which can result in elevated ...

CHARACTERIZATION OF HUMAN LIVER MICROSOMAL UDP-GLYCOSYLTRANSFERASES USING PHOTOAFFINITY ANALOGS

NARCIS (Netherlands)

LITTLE, JM; DRAKE, RR; VONK, R; KUIPERS, F; LESTER, R; RADOMINSKA, A

The photoaffinity analogs [beta-P-32]5-azido-UDP-glucuronic acid ([P-32]5N3UDP-GlcUA) and [beta-P-32]5-azido-UDP-glucose ([P-32]5N(3)UDP-Glc) were used to characterize UDP-glycosyl-transferases of microsomes prepared from human liver. Photoincorporation of both probes into proteins in the 50- to

Liver/kidney microsomal antibody type 1 and liver cytosol antibody type 1 concentrations in type 2 autoimmune hepatitis

OpenAIRE

Muratori, L; Cataleta, M; Muratori, P; Lenzi, M; Bianchi, F

1998-01-01

Background—Liver/kidney microsomal antibody type 1 (LKM1) and liver cytosol antibody type 1 (LC1) are the serological markers of type 2 autoimmune hepatitis (AIH). 
Aims—Since LKM1 and LC1 react against two distinct liver specific autoantigens (cytochrome P450IID6 (CYP2D6) and a 58 kDa cytosolic polypeptide respectively), the aim was to see whether LKM1 and LC1 concentrations correlate with liver disease activity. 
Patients—Twenty one patients with type 2 AIH were studied. 
Methods—A...

Inhibition of rat liver microsomal lipid peroxidation by N-acyldehydroalanines: An in vitro comparative study

Energy Technology Data Exchange (ETDEWEB)

Buc-Calderon, P.; Roberfroid, M. (Universite Catholique de Louvain, Brussels (Belgium))

1989-09-01

Captodative substituted olefins are radical scavengers which react with free radicals to form stabilized radical adducts. One of those compounds, N-(paramethoxyphenylacetyl)dehydroalanine (AD-5), may react and scavenge both superoxide anion (O-2) and alk-oxyl radicals (RO.), and in this way prevent the appearance of their mediated biological effects. Nitrofurantoin and tert-butyl hydroperoxide were used as model compounds to stimulate free radical production and their mediated lipid peroxidation in rat liver microsomes. In addition, lipid peroxidation was also initiated by exposure of rat liver microsomal suspensions to ionizing radiation (gamma rays). The microsomal lipid peroxidation induced by these chemicals and physical agents was inhibited by the addition of AD-5. These effects were dose-dependent in a millimolar range of concentration. In addition, AD-5 has no effect on microsomal electron transport, showing that NADPH-cytochrome P450 reductase activity was not modified. These data, together with the comparisons of the effects of AD-5 and some antioxidant molecules such as superoxide dismutase, uric acid, and mannitol, support the conclusion that inhibition of lipid peroxidation by AD-5 is the result of its free radical scavenger activity. In addition, the inhibitory effect of AD-5 on microsomal lipid peroxidation was dependent of the nature of the free radical species involved in the initiation of the process, suggesting that O-2 is scavenged more efficiently than RO.

Identification of a tryptanthrin metabolite in rat liver microsomes by liquid chromatography/electrospray ionization-tandem mass spectrometry.

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Lee, Sang Kyu; Kim, Ghee Hwan; Kim, Dong Hyeon; Kim, Dong Hyun; Jahng, Yurngdong; Jeong, Tae Cheon

2007-10-01

Tryptanthrin originally isolated from Isatis tinctoria L. has been characterized to have anti-inflammatory activities through the dual inhibition of cyclooxygenase-2 and 5-lipoxygenase mediated prostaglandin and leukotriene syntheses. To characterize phase I metabolite(s), tryptanthrin was incubated with rat liver microsomes in the presence of NADPH-generating system. One metabolite was identified by liquid chromatography/electrospray ionization-tandem mass spectrometry. M1 could be identified as a metabolite mono-hydroxylated on the aromatic ring of indole moiety from the MS(2) spectra of protonated tryptanthrin and M1. The structure of metabolite was confirmed as 8-hydroxytryptanthrin with a chemically synthesized authentic standard. The formation of M1 was NADPH-dependent and was inhibited by SKF-525A, a general CYP-inhibitor, indicating the cytochrome P450 (CYP)-mediated reaction. In addition, it was proposed that M1 might be formed by CYP 1A in rat liver microsomes from the experiments with enriched rat liver microsomes.

Metabolism of UV-filter benzophenone-3 by rat and human liver microsomes and its effect on endocrine-disrupting activity

Energy Technology Data Exchange (ETDEWEB)

Watanabe, Yoko, E-mail: y-watanabe@nichiyaku.ac.jp [Graduate School of Biomedical and Health Sciences, Hiroshima University, Kasumi 1-2-3, Minami-ku, Hiroshima 734-8553 (Japan); Nihon Pharmaceutical University, Komuro 10281, Ina-machi, Saitama 362-0806 (Japan); Kojima, Hiroyuki; Takeuchi, Shinji [Hokkaido Institute of Public Health, Kita-19, Nishi-12, Kita-ku, Sapporo 060-0819 (Japan); Uramaru, Naoto [Nihon Pharmaceutical University, Komuro 10281, Ina-machi, Saitama 362-0806 (Japan); Sanoh, Seigo [Graduate School of Biomedical and Health Sciences, Hiroshima University, Kasumi 1-2-3, Minami-ku, Hiroshima 734-8553 (Japan); Sugihara, Kazumi [Faculty of Pharmaceutical Science, Hiroshima International University, Koshingai 5-1-1, Kure, Hiroshima 737-0112 (Japan); Kitamura, Shigeyuki [Nihon Pharmaceutical University, Komuro 10281, Ina-machi, Saitama 362-0806 (Japan); Ohta, Shigeru [Graduate School of Biomedical and Health Sciences, Hiroshima University, Kasumi 1-2-3, Minami-ku, Hiroshima 734-8553 (Japan)

2015-01-15

Benzophenone-3 (2-hydroxy-4-methoxybenzophenone; BP-3) is widely used as sunscreen for protection of human skin and hair from damage by ultraviolet (UV) radiation. In this study, we examined the metabolism of BP-3 by rat and human liver microsomes, and the estrogenic and anti-androgenic activities of the metabolites. When BP-3 was incubated with rat liver microsomes in the presence of NADPH, 2,4,5-trihydroxybenzophenone (2,4,5-triOH BP) and 3-hydroxylated BP-3 (3-OH BP-3) were newly identified as metabolites, together with previously detected metabolites 5-hydroxylated BP-3 (5-OH BP-3), a 4-desmethylated metabolite (2,4-diOH BP) and 2,3,4-trihydroxybenzophenone (2,3,4-triOH BP). In studies with recombinant rat cytochrome P450, 3-OH BP-3 and 2,4,5-triOH BP were mainly formed by CYP1A1. BP-3 was also metabolized by human liver microsomes and CYP isoforms. In estrogen reporter (ER) assays using estrogen-responsive CHO cells, 2,4-diOH BP exhibited stronger estrogenic activity, 2,3,4-triOH BP exhibited similar activity, and 5-OH BP-3, 2,4,5-triOH BP and 3-OH BP-3 showed lower activity as compared to BP-3. Structural requirements for activity were investigated in a series of 14 BP-3 derivatives. When BP-3 was incubated with liver microsomes from untreated rats or phenobarbital-, 3-methylcholanthrene-, or acetone-treated rats in the presence of NADPH, estrogenic activity was increased. However, liver microsomes from dexamethasone-treated rats showed decreased estrogenic activity due to formation of inactive 5-OH BP-3 and reduced formation of active 2,4-diOH BP. Anti-androgenic activity of BP-3 was decreased after incubation with liver microsomes. - Highlights: • Metabolic modification of the endocrine-disrupting activity of BP-3 was examined. • 2,4,5-TriOH BP and 3-OH BP-3 were identified as new BP-3 metabolites. • 2,4-DiOH BP and 2,3,4-triOH BP exhibited high or similar estrogenic activities. • Estrogenic activity of BP-3 was enhanced by incubation with rat liver

Metabolism of indole alkaloid tumor promoter, (-)-indolactam V, which has the fundamental structure of teleocidins, by rat liver microsomes

Energy Technology Data Exchange (ETDEWEB)

Hagiwara, N.; Irie, K.; Tokuda, H.; Koshimizu, K.

1987-07-01

Metabolic activation and/or deactivation of indole alkaloid tumor promoter, (-)-indolactam V (ILV), was examined using rat liver microsomes. Reaction of ILV with the microsomes supplemented with NADPH and MgCl/sub 2/ gave three major metabolites, which were identified as (-)-N13-desmethylindolactam V and two diastereomers of (-)-2-oxyindolactam V at C-3. The tumor-promoting activities of these metabolites were evaluated by induction of Epstein-Barr virus early antigen and inhibition of specific binding of (/sup 3/H)-12-O-tetradecanoylphorbol-13-acetate to a mouse epidermal particulate fraction, and proved to be conspicuously lower than that of ILV. These results demonstrate that the metabolism of ILV results in detoxification, and that it itself is the tumor-promoting entity. Studies on the enzymes concerned with this metabolism suggested the involvement of cytochrome P-450-containing mixed-function oxidases. Similar deactivation seems to be possible by skin, where the mixed-function oxidases are known to exist.

Liver/kidney microsomal antibody type 1 and liver cytosol antibody type 1 concentrations in type 2 autoimmune hepatitis.

Science.gov (United States)

Muratori, L; Cataleta, M; Muratori, P; Lenzi, M; Bianchi, F B

1998-05-01

Liver/kidney microsomal antibody type 1 (LKM1) and liver cytosol antibody type 1 (LC1) are the serological markers of type 2 autoimmune hepatitis (AIH). Since LKM1 and LC1 react against two distinct liver specific autoantigens (cytochrome P450IID6 (CYP2D6) and a 58 kDa cytosolic polypeptide respectively), the aim was to see whether LKM1 and LC1 concentrations correlate with liver disease activity. Twenty one patients with type 2 AIH were studied. All sera were tested by indirect immunofluorescence, counterimmunoelectrophoresis, and immunoblotting visualised by enhanced chemiluminescence. To evaluate LKM1 and LC1 levels, the 50 kDa microsomal reactivity (corresponding to CYP2D6) and the 58 kDa cytosolic reactivity were quantified by densitometric analysis. Seven patients were positive for LKM1, nine for LC1, and five for both. Serial serum samples at onset and during immunosuppressive treatment were analysed in 13 patients (four positive for LKM1, six positive for LC1 and three positive for both). During remission, LKM1 concentration remained essentially unchanged in six of seven patients, and decreased in only one. Conversely, in two of nine patients, LC1 was completely lost, and, in the remaining seven, LC1 concentration was reduced by more than 50%. After immunosuppression tapering or withdrawal, flare ups of liver necrosis ensued with increasing LC1 concentration, but not LKM1. LC1 concentration, at variance with that of LKM1, parallels liver disease activity, and its participation in the pathogenic mechanisms of liver injury can be hypothesised.

Inhibitory Effects of Aschantin on Cytochrome P450 and Uridine 5′-diphospho-glucuronosyltransferase Enzyme Activities in Human Liver Microsomes

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Soon-Sang Kwon

2016-04-01

Full Text Available Aschantin is a bioactive neolignan found in Magnolia flos with antiplasmodial, Ca2+-antagonistic, platelet activating factor-antagonistic, and chemopreventive activities. We investigated its inhibitory effects on the activities of eight major human cytochrome P450 (CYP and uridine 5′-diphospho-glucuronosyltransferase (UGT enzymes of human liver microsomes to determine if mechanistic aschantin–enzyme interactions were evident. Aschantin potently inhibited CYP2C8-mediated amodiaquine N-de-ethylation, CYP2C9-mediated diclofenac 4′-hydroxylation, CYP2C19-mediated [S]-mephenytoin 4′-hydroxylation, and CYP3A4-mediated midazolam 1′-hydroxylation, with Ki values of 10.2, 3.7, 5.8, and 12.6 µM, respectively. Aschantin at 100 µM negligibly inhibited CYP1A2-mediated phenacetin O-de-ethylation, CYP2A6-mediated coumarin 7-hydroxylation, CYP2B6-mediated bupropion hydroxylation, and CYP2D6-mediated bufuralol 1′-hydroxylation. At 200 µM, it weakly inhibited UGT1A1-catalyzed SN-38 glucuronidation, UGT1A6-catalyzed N-acetylserotonin glucuronidation, and UGT1A9-catalyzed mycophenolic acid glucuronidation, with IC50 values of 131.7, 144.1, and 71.0 µM, respectively, but did not show inhibition against UGT1A3, UGT1A4, or UGT2B7 up to 200 µM. These in vitro results indicate that aschantin should be examined in terms of potential interactions with pharmacokinetic drugs in vivo. It exhibited potent mechanism-based inhibition of CYP2C8, CYP2C9, CYP2C19, and CYP3A4.

Metabolism of lysergic acid diethylamide (LSD) to 2-oxo-3-hydroxy LSD (O-H-LSD) in human liver microsomes and cryopreserved human hepatocytes.

Science.gov (United States)

Klette, K L; Anderson, C J; Poch, G K; Nimrod, A C; ElSohly, M A

2000-10-01

The metabolism of lysergic acid diethylamide (LSD) to 2-oxo-3-hydroxy lysergic acid diethylamide (O-H-LSD) was investigated in liver microsomes and cyropreserved hepatocytes from humans. Previous studies have demonstrated that O-H-LSD is present in human urine at concentrations 16-43 times greater than LSD, the parent compound. Additionally, these studies have determined that O-H-LSD is not generated during the specimen extraction and analytical processes or due to parent compound degradation in aqueous urine samples. However, these studies have not been conclusive in demonstrating that O-H-LSD is uniquely produced during in vivo metabolism. Phase I drug metabolism was investigated by incubating human liver microsomes and cryopreserved human hepatocytes with LSD. The reaction was quenched at various time points, and the aliquots were extracted using liquid partitioning and analyzed by liquid chromatography-mass spectrometry. O-H-LSD was positively identified in all human liver microsomal and human hepatocyte fractions incubated with LSD. In addition, O-H-LSD was not detected in any microsomal or hepatocyte fraction not treated with LSD nor in LSD specimens devoid of microsomes or hepatocytes. This study provides definitive evidence that O-H-LSD is produced as a metabolic product following incubation of human liver microsomes and hepatocytes with LSD.

[Comparative metabolism of three amide alkaloids from Piper longum in five different species of liver microsomes].

Science.gov (United States)

He, Huan; Guo, Wei-Wei; Chen, Xiao-Qing; Zhao, Hai-Yu; Wu, Xia

2016-08-01

Piperine, piperlonguminine and pellitorine are three major amide alkaloids from Piper longum, showing a variety of pharmacological activities. In order to investigate the different metabolism pathways of these compounds in five species of liver microsomes in vitro, the data of full mass spectrum, and MS2, MS3 spectra of these three alkaloids were collected and analyzed by using ultra-high-performance liquid chromatography coupled with a LTQ-orbitrap mass spectrometer (UHPLC-LTQ-Orbitrap MS); gragment ion information was collected and combined with fragmentation regularities of mass spectra and accurate mass spectrometry data of metabolites, to compare the metabolism difference of three amide alkaloids in liver microsomes of human, rhesus monkey, Beagle dogs, rats and mice. 3 metabolites of piperine, 2 metabolites of piperlonguminine and 1 metabolite of pellitorine were identified quickly. The results showed that the major metabolic pathways of these amide alkaloids in liver microsomes were methylenedioxy group demethylation and oxidation reaction, and metabolic rates were different between species. This study provides basis for further research on in vivo metabolism of piperine analogues from Piper longum. Copyright© by the Chinese Pharmaceutical Association.

Measurement of peroxisomal enzyme activities in the liver of brown trout (Salmo trutta, using spectrophotometric methods

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Resende Albina D

2003-03-01

Full Text Available Abstract Background This study was aimed primarily at testing in the liver of brown trout (Salmo trutta spectrophotometric methods previously used to measure the activities of catalase and hydrogen peroxide producing oxidases in mammals. To evaluate the influence of temperature on the activities of those peroxisomal enzymes was the second objective. A third goal of this work was the study of enzyme distribution in crude cell fractions of brown trout liver. Results The assays revealed a linear increase in the activity of all peroxisomal enzymes as the temperature rose from 10° to 37°C. However, while the activities of hydrogen peroxide producing oxidases were strongly influenced by temperature, catalase activity was only slightly affected. A crude fraction enriched with peroxisomes was obtained by differential centrifugation of liver homogenates, and the contamination by other organelles was evaluated by the activities of marker enzymes for mitochondria (succinate dehydrogenase, lysosomes (aryl sulphatase and microsomes (NADPH cytochrome c reductase. For peroxisomal enzymes, the activities per mg of protein (specific activity in liver homogenates were strongly correlated with the activities per g of liver and with the total activities per liver. These correlations were not obtained with crude peroxisomal fractions. Conclusions The spectrophotometric protocols originally used to quantify the activity of mammalian peroxisomal enzymes can be successfully applied to the study of those enzymes in brown trout. Because the activity of all studied peroxisomal enzymes rose in a linear mode with temperature, their activities can be correctly measured between 10° and 37°C. Probably due to contamination by other organelles and losses of soluble matrix enzymes during homogenisation, enzyme activities in crude peroxisomal fractions do not correlate with the activities in liver homogenates. Thus, total homogenates will be used in future seasonal and

A novel assay for detecting antibodies to cytochrome P4502D6, the molecular target of liver kidney microsomal antibody type 1.

Science.gov (United States)

Kerkar, N; Ma, Y; Hussain, M; Muratori, L; Targett, C; Williams, R; Bianchi, F B; Mieli-Vergani, G; Vergani, D

1999-03-04

Liver Kidney Microsomal type 1 (LKM1) antibody, the diagnostic marker of autoimmune hepatitis type 2, is also found in a proportion of patients with hepatitis C virus infection (HCV). It is detected conventionally by the subjective immunofluorescence technique. Our aim was to establish a simple and objective enzyme-linked immunosorbent assay (ELISA) that measures antibodies to cytochrome P4502D6 (CYP2D6), the target of LKM1. An indirect ELISA using eukaryotically expressed CYP2D6 was designed. Absorbance values obtained against a reference microsomal preparation were subtracted from those obtained against a microsomal preparation over-expressing CYP2D6, thus removing the non-CYP2D6-specific reaction. Sera from 51 LKM1 positive patients (21 autoimmune hepatitis and 30 with HCV infection), 111 LKM1 negative patients with chronic liver disease (including 20 with HCV infection) and 43 healthy controls were tested. Of 51 patients positive by immunofluorescence, 48 were also positive by ELISA while all the 154 LKM1 negative subjects were also negative by ELISA. There was a high degree of association between IFL and ELISA as demonstrated by a kappa reliability value of 0.96. The absorbance values by ELISA correlated with immunofluorescence LKM1 titres both in autoimmune hepatitis (r = 0.74, p < 0.001) and HCV infection (r = 0.67, p < 0.001). The simple, objective ELISA described has the potential to replace the standard immunofluorescence technique.

Metabolic activation of 2-methylfuran by rat microsomal systems

International Nuclear Information System (INIS)

Ravindranath, V.; Boyd, M.R.

1985-01-01

2-Methylfuran (2-MF), a constituent of cigarette smoke and coffee, causes necrosis of liver, lungs, and kidneys in rodents. 2-MF is metabolically activated by mixed-function oxidases to acetylacrolein, a reactive metabolite that binds covalently to microsomal protein. The hepatic microsomal metabolism of 2-MF to reactive metabolite required the presence of NADPH and oxygen and was dependent on incubation time and substrate concentration. The microsomal metabolism of 2-MF was inducible by pretreatment of rats with phenobarbital and was inhibited by piperonyl butoxide and N-octyl imidazole, which indicates that the metabolism of 2-MF may be mediated by cytochrome P-450. Acetylacrolein was a potent inhibitor of mixed-function oxidase and completely inhibited the microsomal metabolism of 2-MF, indicating that 2-MF is a suicide substrate for the enzyme. The sulfhydryl nucleophile cysteine was a better trapping agent of the reactive metabolite of 2-MF than N-acetylcysteine or glutathione. Lysine decreased the covalent binding of 2-MF metabolites, presumably by reacting with the aldehyde group of acetylacrolein. In addition, in the presence of NADPH, 2-MF was bioactivated by both pulmonary and renal cortical microsomes to reactive metabolites that were covalently bound to microsomal proteins

Hepatic and intestinal glucuronidation of mono(2-ethylhexyl) phthalate, an active metabolite of di(2-ethylhexyl) phthalate, in humans, dogs, rats, and mice: an in vitro analysis using microsomal fractions.

Science.gov (United States)

Hanioka, Nobumitsu; Isobe, Takashi; Kinashi, Yu; Tanaka-Kagawa, Toshiko; Jinno, Hideto

2016-07-01

Mono(2-ethylhexyl) phthalate (MEHP) is an active metabolite of di(2-ethylhexyl) phthalate (DEHP) and has endocrine-disrupting effects. MEHP is metabolized into glucuronide by UDP-glucuronosyltransferase (UGT) enzymes in mammals. In the present study, the hepatic and intestinal glucuronidation of MEHP in humans, dogs, rats, and mice was examined in an in vitro system using microsomal fractions. The kinetics of MEHP glucuronidation by liver microsomes followed the Michaelis-Menten model for humans and dogs, and the biphasic model for rats and mice. The K m and V max values of human liver microsomes were 110 µM and 5.8 nmol/min/mg protein, respectively. The kinetics of intestinal microsomes followed the biphasic model for humans, dogs, and mice, and the Michaelis-Menten model for rats. The K m and V max values of human intestinal microsomes were 5.6 µM and 0.40 nmol/min/mg protein, respectively, for the high-affinity phase, and 430 µM and 0.70 nmol/min/mg protein, respectively, for the low-affinity phase. The relative levels of V max estimated by Eadie-Hofstee plots were dogs (2.0) > mice (1.4) > rats (1.0) ≈ humans (1.0) for liver microsomes, and mice (8.5) > dogs (4.1) > rats (3.1) > humans (1.0) for intestinal microsomes. The percentages of the V max values of intestinal microsomes to liver microsomes were mice (120 %) > rats (57 %) > dogs (39 %) > humans (19 %). These results suggest that the metabolic abilities of UGT enzymes expressed in the liver and intestine toward MEHP markedly differed among species, and imply that these species differences are strongly associated with the toxicity of DEHP.

Development of vitamin D3 25-hydroxylase activity in rat liver microsomes

International Nuclear Information System (INIS)

Thierry-Palmer, M.; Cullins, S.; Rashada, S.; Gray, T.K.; Free, A.

1986-01-01

The authors have determined the ontogeny of vitamin D 3 25-hydroxylase activity in rat liver microsomes. Microsomes from fetuses, neonates, and their mothers were incubated with 44 nM 3 H-vitamin D 3 in the presence of an NADPH generating system, oxygen, KCl, and MgCl 2 . Lipid extracts of the incubation samples were partially purified by thin-layer chromatography. Tritiated 25-hydroxy vitamin D 3 (250HD 3 ) was analyzed by high-pressure liquid chromatography using 94/6 hexane/isopropanol. Production rate for 250HD 3 in the mothers ranged from 0.22 to 0.30 pmol/mg protein/hr. Activities in the fetuses and neonates were 2.1, 12.9, 32.0, 35.8, and 71.0% of that of their mothers at -3, 0, 2, 7, and 15 days of age. The cytosolic fraction protected the substrate from degradation, stimulated the vitamin D 3 25-hydroxylase reaction in neonates and mothers (1.4 to 1.7 fold increase), and was absolutely required for 25-hydroxylase activity in fetuses. These data suggest that microsomal vitamin D 3 25-hydroxylase activity develops slowly and approaches full activity near the weaning stage. A cytosolic factor present as early as -3 days of age stimulates the activity of the microsomal vitamin D 3 25-hydroxylase

Effect of rat whole-body irradiation on oxidase chain and glucose-6-phosphatase of liver microsome: influence of cysteamine

International Nuclear Information System (INIS)

Bernard, Pierre.

1979-11-01

Three enzymatic systems of the male rat liver endoplasmic reticulum were studied by biochemical methods. Two means of investigation were used: - whole-body irradiation of the animal, - administration of cysteamine. The results obtained are discussed, in view of the functioning of these enzymatic systems, from two viewpoints: - the study of enzymatic radiolesions in relation to the radiobiological effect on the animal, the organ and the sub-cellular organite, - the study of chemical radioprotection. After a 900 R whole-body gamma irradiation a severe drop was observed in the enzymatic activity of two essential elements of the microsome oxydase chain: NADPH cytochrome P450 reductase and ethylmorphine N-demethylation. Glucose 6 phosphatase is also impaired by irradiation. Here it seems that the microsomal protein fraction could be responsible for the change in the enzyme activity. The irradiation effect is therefore not specific to one enzyme. The changes in these enzymatic activities correspond to the different phases of the acute irradiation syndrome which also affects the weight of the experimental animal and of the organ studied. Cysteamine used under chemical radioprotection conditions was found to be especially useful as a means of investigation complementary to the study of enzymatic radiolesions. From the combined action of irradiation and of the radioprotector it was possible to obtain a partial idea of the mechanisms of these radiolesions [fr

Environmentally persistent free radicals inhibit cytochrome P450 activity in rat liver microsomes

Energy Technology Data Exchange (ETDEWEB)

Reed, James R., E-mail: rreed@lsuhsc.edu [Department of Pharmacology and Experimental Therapeutics, Louisiana State University Health Science Center, 533 Bolivar St., New Orleans, LA 70112 (United States); The Stanley S. Scott Cancer Center, Louisiana State University Health Science Center, 533 Bolivar St., New Orleans, LA 70112 (United States); Cawley, George F.; Ardoin, Taylor G. [Department of Pharmacology and Experimental Therapeutics, Louisiana State University Health Science Center, 533 Bolivar St., New Orleans, LA 70112 (United States); The Stanley S. Scott Cancer Center, Louisiana State University Health Science Center, 533 Bolivar St., New Orleans, LA 70112 (United States); Dellinger, Barry; Lomnicki, Slawomir M.; Hasan, Farhana; Kiruri, Lucy W. [Department of Chemistry, Louisiana State University, Baton Rouge, LA 70803 (United States); Backes, Wayne L. [Department of Pharmacology and Experimental Therapeutics, Louisiana State University Health Science Center, 533 Bolivar St., New Orleans, LA 70112 (United States); The Stanley S. Scott Cancer Center, Louisiana State University Health Science Center, 533 Bolivar St., New Orleans, LA 70112 (United States)

2014-06-01

Combustion processes generate particulate matter that affects human health. When incineration fuels include components that are highly enriched in aromatic hydrocarbons (especially halogenated varieties) and redox-active metals, ultrafine particulate matter containing air-stable, environmentally persistent free radicals (EPFRs) is generated. The exposure to fine EPFRs (less than 2.5 μm in diameter) has been shown to negatively influence pulmonary and cardiovascular functions in living organisms. The goal of this study was to determine if these EPFRs have a direct effect on cytochrome P450 function. This was accomplished by direct addition of the EPFRs to rat liver microsomal preparations and measurement of several P450 activities using form-selective substrates. The EPFRs used in this study were formed by heating vapors from an organic compound (either monochlorophenol (MCP230) or 1,2-dichlorobenzene (DCB230)) and 5% copper oxide supported on silica (approximately 0.2 μm in diameter) to 230 °C under vacuum. Both types of EPFRs (but not silica, physisorbed silica, or silica impregnated with copper oxide) dramatically inhibited the activities of CYP1A, CYP2B, CYP2E1, CYP2D2 and CYP3A when incubated at concentrations less than 0.1 mg/ml with microsomes and NADPH. Interestingly, at the same concentrations, the EPFRs did not inhibit HO-1 activity or the reduction of cytochrome c by NADPH-cytochrome P450 reductase. CYP2D2-selective metabolism by rat liver microsomes was examined in more detail. The inhibition of CYP2D2-selective metabolism by both DCB230- and MCP230-EPFRs appeared to be largely noncompetitive and was attenuated in the presence of catalase suggesting that reactive oxygen species may be involved in the mechanism of inhibition. - Highlights: • Combustion of organic pollutants generates long-lived particulate radicals (EPFRs). • EPFRs inhibit metabolism by all cytochromes P450 tested in rat liver microsomes. • EPFR-mediated inhibition is related to

Environmentally persistent free radicals inhibit cytochrome P450 activity in rat liver microsomes

International Nuclear Information System (INIS)

Reed, James R.; Cawley, George F.; Ardoin, Taylor G.; Dellinger, Barry; Lomnicki, Slawomir M.; Hasan, Farhana; Kiruri, Lucy W.; Backes, Wayne L.

2014-01-01

Combustion processes generate particulate matter that affects human health. When incineration fuels include components that are highly enriched in aromatic hydrocarbons (especially halogenated varieties) and redox-active metals, ultrafine particulate matter containing air-stable, environmentally persistent free radicals (EPFRs) is generated. The exposure to fine EPFRs (less than 2.5 μm in diameter) has been shown to negatively influence pulmonary and cardiovascular functions in living organisms. The goal of this study was to determine if these EPFRs have a direct effect on cytochrome P450 function. This was accomplished by direct addition of the EPFRs to rat liver microsomal preparations and measurement of several P450 activities using form-selective substrates. The EPFRs used in this study were formed by heating vapors from an organic compound (either monochlorophenol (MCP230) or 1,2-dichlorobenzene (DCB230)) and 5% copper oxide supported on silica (approximately 0.2 μm in diameter) to 230 °C under vacuum. Both types of EPFRs (but not silica, physisorbed silica, or silica impregnated with copper oxide) dramatically inhibited the activities of CYP1A, CYP2B, CYP2E1, CYP2D2 and CYP3A when incubated at concentrations less than 0.1 mg/ml with microsomes and NADPH. Interestingly, at the same concentrations, the EPFRs did not inhibit HO-1 activity or the reduction of cytochrome c by NADPH-cytochrome P450 reductase. CYP2D2-selective metabolism by rat liver microsomes was examined in more detail. The inhibition of CYP2D2-selective metabolism by both DCB230- and MCP230-EPFRs appeared to be largely noncompetitive and was attenuated in the presence of catalase suggesting that reactive oxygen species may be involved in the mechanism of inhibition. - Highlights: • Combustion of organic pollutants generates long-lived particulate radicals (EPFRs). • EPFRs inhibit metabolism by all cytochromes P450 tested in rat liver microsomes. • EPFR-mediated inhibition is related to

Studies on estradiol-2/4-hydroxylase activity in rat brain and liver

International Nuclear Information System (INIS)

Theron, C.N.

1985-03-01

A sensitive and specific radio-enzymatic assay was used to study estradiol-2/4-hydroxylase activity in rat liver microsomes and in microsomes obtained from 6 discrete brain areas of the rat. Kinetic parameters were determined for these enzyme activities. The effects of different P-450 inhibitors on estradiol-2/4-hydroxylase activity in brain and liver microsomes were also studied. In both organs these enzyme activities were found to be located mainly in the microsomal fraction and were inhibited by the 3 P-450 inhibitors tested. The hepatic estradiol-2/4-hydroxylase activity in adult male rats was significantly higher than that of females, but the enzyme activity in the brain did not exhibit a similar sex difference. Furthermore, estradiol-2/4-hydroxylase activity in rat liver was strongly induced by phenobarbitone treatment, but not in the brain. The phenobarbitone-induced activity in male and female rats exhibited significant kinetic differences. In female rats sexual maturation was associated with significant changes in the apparent Km of estradiol-2/4-hydroxylases in the liver and hypothalamus. Evidence was found that the in vitro estradiol-2/4-hydroxylase activity in rat brain and liver is due to more than one form of microsomal P-450. Kinetic studies showed important differences between the estradiol-2/4-hydroxylase activities in the hippocampus and hypothalamus. Significant differences in estradiol-2/4-hydroxylase activities were observed in the 6 brain areas studied, with the hippocampus showing the highest, and the hypothalamus the lowest activity at all developmental stages in both male and female rats

[Peroxide modification of membranes and isomorphic composition of cytochrome P-450 of rat liver microsomes during antioxidant deficiency].

Science.gov (United States)

Gubskiy, Iu I; Paramonova, G I; Boldeskul, A E; Primak, R G; Bogdanova, L A; Zadorina, O V; Litvinova, N V

1992-01-01

Lipid peroxidation (LPO), physico-chemical properties of the membranes and isoformic composition of microsomal cytochrome P-450 from the rat liver were studied under conditions of antioxidant insufficiency (AOI) which was modelled by exclusion of alpha-tocopherol from the animals' ration. An insignificant accumulation of microsomal diene conjugates and schiff bases against a sharp increase of the ability to the prooxidant stimulated LPO in vitro took place. A significant decrease of membrane lipid microviscosity and a change in surface properties of microsomal membranes of rats with AOI was determined. Absence of alpha-tocopherol in the ration was accompanied by a significant change in the content of separate isoforms of cytochrome P-450 exhibited in growth of a polypeptide with m. w. 54 kDa and the lowering of proteins with m. w. 48 and 50 kDa. Less intensive quenching of tryptophan fluorescence by acrylamide was also revealed, which testified to a lower accessibility of the quencher to membrane proteins or their fluorophore sites. Modification of lipid composition and of physicochemical properties of the rat liver membrane microsomes which was observed at AOI was significantly correlated by pretreatment with the antioxidant 4-methyl-2,6-ditretbutylphenol (ionol).

A reliable radiochromatographic assay technique for hepatic microsomal 16α-hydroxylase activity towards oestrone 3-sulphate

International Nuclear Information System (INIS)

Tsoutsoulis, C.J.; Hobkirk, R.

1980-01-01

A reliable procedure for the assay of liver microsomal 16α-hydroxylation of oestrone 3-sulphate has been developed for the guinea pig. It is based on the rapid, quantitative separation of oestradiol and oestriol by Sephadex LH-20 columns after the chemical reduction and enzymic hydrolysis of the incubation products. Microsomal preparations and incubation conditions that optimized 16α-hydroxylation of oestrone 3-sulphate were employed. Under these circumstances, reduction of the substrate at C-17 and hydrolysis of the sulphate were minimized. Conditions were established that yielded reaction linearity with respect to time and microsomal concentration. This hydroxylation had an absolute requirement for NADPH, which could not be satisfied by NADH. Apparent Ksub(m) values for oestrone 3-sulphate and NADPH, under the conditions used, were 14μM and 0.17mM respectively. 16α-hydroxylase activity was present in the liver microsomal fraction from heavily pigmented, female English Shorthaired guinea pigs. Much lower activity was detected in mature pigmented males and albino females. No activity could be demonstrated in mature, albino males. (author)

Hepatitis C virus infection associated with liver-kidney microsomal antibody type 1 (LKM1) autoantibodies in children.

Science.gov (United States)

Bortolotti, Flavia; Muratori, Luigi; Jara, Paloma; Hierro, Loreto; Verucchi, Gabriella; Giacchino, Raffaella; Barbera, Cristiana; Zancan, Lucia; Guido, Maria; Resti, Massimo; Pedditzi, Sabrina; Bianchi, Francesco; Gatta, Angelo

2003-02-01

To evaluate the clinical pattern and evolution of chronic hepatitis C in children with liver/kidney microsomal antibody type 1 autoantibodies (LKM1). A multicenter, retrospective study, including the following groups of children with hepatitis C virus infection: (1). 21 consecutive LKM1-positive patients, (2). 42 age- and sex- matched LKM1-negative patients, and (3). 4 interferon-induced LKM1-positive cases. LKM1 reactivity to human microsomes and recombinant cytochrome P450IID6 (CYP2D6) was assayed by immunoblotting. Clinical and biochemical features overlapped in LKM1-positive and LKM1-negative children, but a fibrosis score >3 (range 0-6) was significantly more frequent (P =.04) in the former. Reactivity to microsomal protein and CYP2D6 was significantly (P =.02) associated with LKM1 titers >or=1:320 and was found in 39% of patients, including severe cases and both children (of 4 treated) who achieved a sustained alanine aminotransferase (ALT) normalization after steroid treatment. Five of 7 LKM1-positive children treated with interferon had an ALT exacerbation. LKM1-positive hepatitis C in children is characterized by a wide spectrum of biochemical, serologic, and histologic features. Whether autoimmunity may contribute to liver damage in a subgroup of patients with more severe liver disease, high LKM1 titers, and reactivity to CYP2D6 is a question deserving further investigation.

Characterisation of the cytochrome P450 enzymes involved in the in vitro metabolism of granisetron.

OpenAIRE

Bloomer, J C; Baldwin, S J; Smith, G J; Ayrton, A D; Clarke, S E; Chenery, R J

1994-01-01

1. The metabolism of granisetron was investigated in human liver microsomes to identify the specific forms of cytochrome P450 responsible. 2. 7-hydroxy and 9'-desmethyl granisetron were identified as the major products of metabolism following incubation of granisetron with human liver microsomes. At low, clinically relevant, concentrations of granisetron the 7-hydroxy metabolite predominated. Rates of granisetron 7-hydroxylation varied over 100-fold in the human livers investigated. 3. Enzyme...

Biotransformation of a novel antimitotic agent, I-387, by mouse, rat, dog, monkey, and human liver microsomes and in vivo pharmacokinetics in mice.

Science.gov (United States)

Ahn, Sunjoo; Kearbey, Jeffrey D; Li, Chien-Ming; Duke, Charles B; Miller, Duane D; Dalton, James T

2011-04-01

3-(1H-Indol-2-yl)phenyl)(3,4,5-trimethoxyphenyl)methanone (I-387) is a novel indole compound with antitubulin action and potent antitumor activity in various preclinical models. I-387 avoids drug resistance mediated by P-glycoprotein and showed less neurotoxicity than vinca alkaloids during in vivo studies. We examined the pharmacokinetics and metabolism of I-387 in mice as a component of our preclinical development of this compound and continued interest in structure-activity relationships for antitubulin agents. After a 1 mg/kg intravenous dose, noncompartmental pharmacokinetic analysis in plasma showed that clearance (CL), volume of distribution at steady state (Vd(ss)), and terminal half-life (t(1/2)) of I-387 were 27 ml per min/kg, 5.3 l/kg, and 7 h, respectively. In the in vitro metabolic stability study, half-lives of I-387 were between 10 and 54 min by mouse, rat, dog, monkey, and human liver microsomes in the presence of NADPH, demonstrating interspecies variability. I-387 was most stable in rat liver microsomes and degraded quickly in monkey liver microsomes. Liquid chromatography-tandem mass spectrometry was used to identify phase I metabolites. Hydroxylation, reduction of a ketone group, and O-demethylation were the major metabolites formed by the liver microsomes of the five species. The carbonyl group of I-387 was reduced and identified as the most labile site in human liver microsomes. The results of these drug metabolism and pharmacokinetic studies provide the foundation for future structural modification of this pharmacophore to improve stability of drugs with potent anticancer effects in cancer patients.

Lichen planus, liver kidney microsomal (LKM1) antibodies and hepatitis C virus antibodies.

Science.gov (United States)

Divano, M C; Parodi, A; Rebora, A

1992-01-01

No anti-liver kidney microsomal (LKM1) antibodies were detected in 46 patients with LP, 16 of whom had also a chronic liver disease (CLD). In contrast, anti-hepatitis C virus (HCV) antibodies were found in 10% of patients with LP and in 50% of those with LP and CLD. Anti-HCV antibodies may be considered as a false-positive reaction in 56% of cases, especially when anti-LKM1 antibodies are present. Our findings do not support such a hypothesis, but suggest that CLD in LP patients is, at least in Italy, mostly a postviral chronic active hepatitis.

Detection of liver kidney microsomal type 1 antibody using molecularly based immunoassays.

Science.gov (United States)

Kerkar, N; Ma, Y; Davies, E T; Cheeseman, P; Mieli-Vergani, G; Vergani, D

2002-12-01

To assess the diagnostic value of two commercial molecularly based immunoassays detecting liver kidney microsomal type 1 antibody (LKM1). The performance of Varelisa and LKM1 enzyme linked immunosorbent assay (ELISA) was compared with immunofluorescence, and two validated research techniques-an in house ELISA and a radioligand assay measuring antibodies to P4502D6. Thirty serum samples from three patients with autoimmune hepatitis type 2 covering immunofluorescence titres of 1/10 to 1/10 240 and 55 LKM1 negative controls were tested. All 30 sera that were LKM1 positive by immunofluorescence were positive by the in house ELISA, the radioligand assay, and LKM1-ELISA, and 29 were also positive by Varelisa. None of the 55 sera negative for LKM1 by immunofluorescence was positive by the in house ELISA and radioligand assay, but one was positive by Varelisa and 14 were positive using the LKM1-ELISA. Agreement between immunofluorescence, the in house ELISA, the radioligand assay, and Varelisa was high (kappa > 0.8), and agreement between immunofluorescence and LKM1-ELISA was moderate (kappa = 0.63). The assay kit marketed as Varelisa allows accurate detection of LKM1.

Application of the relative activity factor approach in scaling from heterologously expressed cytochromes p450 to human liver microsomes: studies on amitriptyline as a model substrate.

Science.gov (United States)

Venkatakrishnan, K; von Moltke, L L; Greenblatt, D J

2001-04-01

The relative activity factor (RAF) approach is being increasingly used in the quantitative phenotyping of multienzyme drug biotransformations. Using lymphoblast-expressed cytochromes P450 (CYPs) and the tricyclic antidepressant amitriptyline as a model substrate, we have tested the hypothesis that the human liver microsomal rates of a biotransformation mediated by multiple CYP isoforms can be mathematically reconstructed from the rates of the biotransformation catalyzed by individual recombinant CYPs using the RAF approach, and that the RAF approach can be used for the in vitro-in vivo scaling of pharmacokinetic clearance from in vitro intrinsic clearance measurements in heterologous expression systems. In addition, we have compared the results of two widely used methods of quantitative reaction phenotyping, namely, chemical inhibition studies and the prediction of relative contributions of individual CYP isoforms using the RAF approach. For the pathways of N-demethylation (mediated by CYPs 1A2, 2B6, 2C8, 2C9, 2C19, 2D6, and 3A4) and E-10 hydroxylation (mediated by CYPs 2B6, 2D6, and 3A4), the model-predicted biotransformation rates in microsomes from a panel of 12 human livers determined from enzyme kinetic parameters of the recombinant CYPs were similar to, and correlated with the observed rates. The model-predicted clearance via N-demethylation was 53% lower than the previously reported in vivo pharmacokinetic estimates. Model-predicted relative contributions of individual CYP isoforms to the net biotransformation rate were similar to, and correlated with the fractional decrement in human liver microsomal reaction rates by chemical inhibitors of the respective CYPs, provided the chemical inhibitors used were specific to their target CYP isoforms.

Epoxidation of the methamphetamine pyrolysis product, trans-phenylpropene, to trans-phenylpropylene oxide by CYP enzymes and stereoselective glutathione adduct formation

International Nuclear Information System (INIS)

Sanga, Madhu; Younis, Islam R.; Tirumalai, Padma S.; Bland, Tina M.; Banaszewska, Monica; Konat, Gregory W.; Tracy, Timothy S.; Gannett, Peter M.; Callery, Patrick S.

2006-01-01

Pyrolytic products of smoked methamphetamine hydrochloride are well established. Among the various degradation products formed, trans-phenylpropene (trans-β-methylstyrene) is structurally similar to styrene analogues known to be bioactivated by CYP enzymes. In human liver microsomes, trans-phenylpropene was converted to the epoxide trans-phenylpropylene oxide (trans-2-methyl-3-phenyloxirane) and cinnamyl alcohol. Incubation of trans-phenylpropene with microsomes in the presence of enzyme-specific P450 enzyme inhibitors indicated the involvement of CYP2E1, CYP1A2, and CYP3A4 enzymes. Both (R,R)-phenylpropylene oxide and (S,S)-phenylpropylene oxide were formed in human liver microsomal preparations. Enantiomers of trans-phenylpropylene oxide were stereoselectively and regioselectively conjugated in a Phase II drug metabolism reaction catalyzed by human liver cytosolic enzymes consisting of conjugation with glutathione. The structure of the phenylpropylene oxide-glutathione adduct is consistent with nucleophilic ring-opening by attack at the benzylic carbon. Exposure of cultured C6 glial cells to (S,S)-phenylpropylene oxide produced a cytotoxic response in a concentration-dependent manner based on cell degeneration and death

Effect of cholesterol feeding on tissue lipid perioxidation, glutathione peroxidase activity and liver microsomal functions in rats and guinea pigs

NARCIS (Netherlands)

TSAI, A. C.; THIE, G. M.; Lin, C. R.

1977-01-01

The effect of cholesterol feeding on liver and aortic nonenzymatic lipid peroxidation and glutathione peroxidase activities, and on liver microsomal NADPH-dependent lipid peroxidation, codeine hydroxylation and cytochrome P-450 levels was examined in rats and guinea pigs. One percent cholesterol was

Characterization of anti-liver-kidney microsome antibody (anti-LKM1) from hepatitis C virus-positive and -negative sera.

Science.gov (United States)

Yamamoto, A M; Cresteil, D; Homberg, J C; Alvarez, F

1993-06-01

Hepatitis C virus-related antibodies were found in sera positive for antibodies to liver/kidney microsome antibody, usually considered a marker of autoimmune hepatitis. The aim of this study was to analyze the specificity of this autoantibody in sera from patients with and without hepatitis C virus infection. Fifteen anti-hepatitis C virus- and anti-liver kidney microsome-positive sera were compared with 11 sera from patients with autoimmune hepatitis, for reactivity against rat and human liver microsomal proteins, P450IID6 recombinant proteins, and various synthetic peptides spanning the 241-429 amino acids sequence of the P450IID6. Ten of 11 sera from patients with autoimmune hepatitis bound to recombinant proteins spanning the P450IID6 region between amino acids 72 and 458. These sera bound to the 254-271 peptide, and some also recognized the 321-351, 373-389 and 410-429 peptides. Four of 15 antihepatitis C virus recognized the fusion protein coded by the full-length P450IID6 complementary DNA; 3 of them also reacted with the P450IID6 region between amino acids 72-456. Only 1 sera recognized the 321-351 peptide. P450IID6 antigenic sites recognized by anti-hepatitis C virus-positive sera were different from those recognized by sera from patients with autoimmune hepatitis.

Role of metabolic activation by cytochrome P-450 in covalent binding of VP 16-213 to rat liver and HeLa cell microsomal proteins

Energy Technology Data Exchange (ETDEWEB)

van Maanen, J.M.; de Ruiter, C.; de Vries, J.; Kootstra, P.R.; Gobas, F.; Pinedo, H.M.

1985-09-01

Covalent binding of /sup 3/H-labeled VP 16-213 to rat liver and HeLa cell microsomal proteins was studied in vitro. Metabolic activation by cytochrome P-450 was found to play a role in the covalent binding of VP 16-213 to rat liver microsomal proteins, as shown by the need of NADPH cofactor, the increased binding after phenobarbital pretreatment and the inhibition by SFK-525A. Addition of ascorbic acid or alpha-phenyl-N-tert. butylnitrone to the incubation mixture depressed covalent binding by about 85%, suggesting that formation of a reactive metabolite from the phenolic structure may be involved in the binding process. VP 16-213 did not inhibit aminopyrine N-demethylase at the concentration used in the binding experiments (17 microM), indicating that metabolism of its methylenedioxy group does not play a role in binding to microsomal proteins. HeLa cell microsomes were found to possess aminopyrine N-demethylase activity. Covalent binding of radiolabeled VP 16-213 to HeLa cell microsomes decreased by about 64% if NADPH was omitted.

Microsomal metabolism of trenbolone acetate metabolites ...

Science.gov (United States)

Trenbolone acetate (TBA) is a synthetic growth promoter widely used in animal agriculture, and its metabolites are suspected endocrine disrupting compounds in agriculturally impacted receiving waters. However, beyond the three widely recognized TBA metabolites (17-trenbolone, 17-trenbolone and trendione), little is known about other metabolites formed in vivo and subsequently discharged into the environment, with some evidence suggesting these unknown metabolites comprise a majority of the TBA mass dosed to the animal. Here, we explored the metabolism of the three known TBA metabolites using rat liver microsome studies. All TBA metabolites are transformed into a complex mixture of monohydroxylated products. Based on product characterization, the majority are more polar than the parent metabolites but maintain their characteristic trienone backbone. A minor degree of interconversion between known metabolites was also observed, as were higher order hydroxylated products with a greater extent of reaction. Notably, the distribution and yield of products were generally comparable across a series of variably induced rat liver microsomes, as well as during additional studies with human and bovine liver microsomes. Bioassays conducted with mixtures of these transformation products suggest that androgen receptor (AR) binding activity is diminished as a result of the microsomal treatment, suggesting that the transformation products are generally less potent than

The effects of space flight on some rat liver enzymes regulating carbohydrate and lipid metabolism

Science.gov (United States)

Abraham, S.; Lin, C. Y.; Klein, H. P.; Volkmann, C.

1981-01-01

The effects of space flight conditions on the activities of certain enzymes regulating carbohydrate and lipid metabolism in rat liver are investigated in an attempt to account for the losses in body weight observed during space flight despite preflight caloric consumption. Liver samples were analyzed for the activities of 32 cytosolic and microsomal enzymes as well as hepatic glycogen and individual fatty acid levels for ground control rats and rats flown on board the Cosmos 936 biosatellite under normal space flight conditions and in centrifuges which were sacrificed upon recovery or 25 days after recovery. Significant decreases in the activities of glycogen phosphorylase, alpha-glycerol phosphate acyl transferase, diglyceride acyl transferase, aconitase and 6-phosphogluconate dehydrogenase and an increase in palmitoyl CoA desaturase are found in the flight stationary relative to the flight contrifuged rats upon recovery, with all enzymes showing alterations returning to normal values 25 days postflight. The flight stationary group is also observed to be characterized by more than twice the amount of liver glycogen of the flight centrifuged group as well as a significant increase in the ratio of palmitic to palmitoleic acid. Results thus indicate metabolic changes which may be involved in the mechanism of weight loss during weightlessness, and demonstrate the equivalence of centrifugation during space flight to terrestrial gravity.

CYP2B6, CYP2D6, and CYP3A4 catalyze the primary oxidative metabolism of perhexiline enantiomers by human liver microsomes.

Science.gov (United States)

Davies, Benjamin J; Coller, Janet K; Somogyi, Andrew A; Milne, Robert W; Sallustio, Benedetta C

2007-01-01

The cytochrome P450 (P450)-mediated 4-monohydroxylations of the individual enantiomers of the racemic antianginal agent perhexiline (PHX) were investigated in human liver microsomes (HLMs) to identify stereoselective differences in metabolism and to determine the contribution of the polymorphic enzyme CYP2D6 and other P450s to the intrinsic clearance of each enantiomer. The cis-, trans1-, and trans2-4-monohydroxylation rates of (+)- and (-)-PHX by human liver microsomes from three extensive metabolizers (EMs), two intermediate metabolizers (IMs), and two poor metabolizers (PMs) of CYP2D6 were measured with a high-performance liquid chromatography assay. P450 isoform-specific inhibitors, monoclonal antibodies directed against P450 isoforms, and recombinantly expressed human P450 enzymes were used to define the P450 isoform profile of PHX 4-monohydroxylations. The total in vitro intrinsic clearance values (mean +/- S.D.) of (+)- and (-)-PHX were 1376 +/- 330 and 2475 +/- 321, 230 +/- 225 and 482 +/- 437, and 63.4 +/- 1.6 and 54.6 +/- 1.2 microl/min/mg for the EM, IM, and PM HLMs, respectively. CYP2D6 catalyzes the formation of cis-OH-(+)-PHX and trans1-OH-(+)-PHX from (+)-PHX and cis-OH-(-)-PHX from (-)-PHX with high affinity. CYP2B6 and CYP3A4 each catalyze the trans1- and trans2-4-monohydroxylation of both (+)- and (-)-PHX with low affinity. Both enantiomers of PHX are subject to significant polymorphic metabolism by CYP2D6, although this enzyme exhibits distinct stereoselectivity with respect to the conformation of metabolites and the rate at which they are formed. CYP2B6 and CYP3A4 are minor contributors to the intrinsic P450-mediated hepatic clearance of both enantiomers of PHX, except in CYP2D6 PMs.

Rat liver microsomal cytochrome P450-dependent oxidation of 3,5-disubstituted analogues of paracetamol

NARCIS (Netherlands)

Bessems, J.G.M.; Koppele, J.M. te; Dijk, P.A. van; Stee, L.L.P. van; Commandeur, J.N.M.; Vermeulen, N.P.E.

1996-01-01

1. The cytochrome P450-dependent binding of paracetamol and a series of 3,5-disubstituted paracetamol analogues (R = -F, -Cl, -Br, -I, -C(H)3, -C2H5, -iC3H7) have been determined with β-naphthoflavone (βNF)-induced rat liver microsomes and produced reverse type I spectral changes. K(s,app) varied

Monkey liver cytochrome P450 2C19 is involved in R- and S-warfarin 7-hydroxylation.

Science.gov (United States)

Hosoi, Yoshio; Uno, Yasuhiro; Murayama, Norie; Fujino, Hideki; Shukuya, Mitsunori; Iwasaki, Kazuhide; Shimizu, Makiko; Utoh, Masahiro; Yamazaki, Hiroshi

2012-12-15

Cynomolgus monkeys are widely used as primate models in preclinical studies. However, some differences are occasionally seen between monkeys and humans in the activities of cytochrome P450 enzymes. R- and S-warfarin are model substrates for stereoselective oxidation in humans. In this current research, the activities of monkey liver microsomes and 14 recombinantly expressed monkey cytochrome P450 enzymes were analyzed with respect to R- and S-warfarin 6- and 7-hydroxylation. Monkey liver microsomes efficiently mediated both R- and S-warfarin 7-hydroxylation, in contrast to human liver microsomes, which preferentially catalyzed S-warfarin 7-hydroxylation. R-Warfarin 7-hydroxylation activities in monkey liver microsomes were not inhibited by α-naphthoflavone or ketoconazole, and were roughly correlated with P450 2C19 levels and flurbiprofen 4-hydroxylation activities in microsomes from 20 monkey livers. In contrast, S-warfarin 7-hydroxylation activities were not correlated with the four marker drug oxidation activities used. Among the 14 recombinantly expressed monkey P450 enzymes tested, P450 2C19 had the highest activities for R- and S-warfarin 7-hydroxylations. Monkey P450 3A4 and 3A5 slowly mediated R- and S-warfarin 6-hydroxylations. Kinetic analysis revealed that monkey P450 2C19 had high V(max) and low K(m) values for R-warfarin 7-hydroxylation, comparable to those for monkey liver microsomes. Monkey P450 2C19 also mediated S-warfarin 7-hydroxylation with V(max) and V(max)/K(m) values comparable to those for recombinant human P450 2C9. R-warfarin could dock favorably into monkey P450 2C19 modeled. These results collectively suggest high activities for monkey liver P450 2C19 toward R- and S-warfarin 6- and 7-hydroxylation in contrast to the saturation kinetics of human P450 2C9-mediated S-warfarin 7-hydroxylation. Copyright © 2012 Elsevier Inc. All rights reserved.

The effect of fenbuconazole on cell proliferation and enzyme induction in the liver of female CD1 mice

International Nuclear Information System (INIS)

Juberg, Daland R.; Mudra, Daniel R.; Hazelton, George A.; Parkinson, Andrew

2006-01-01

Fenbuconazole, a triazole fungicide, has been associated with an increase in the incidence of liver adenomas in female mice following long-term dietary exposure. The aim of this study was to evaluate whether the mode of action for liver tumor formation by fenbuconazole is similar to that of phenobarbital. Treatment of CD1 mice with 0, 20, 60, 180 or 1300 ppm fenbuconazole for up to 4 weeks caused a dose-dependent increase in liver weight that was associated with centrilobular hepatocellular hypertrophy, cytoplasmic eosinophilia and panlobular hepatocellular vacuolation, as well as an initial increase in the cell proliferation labeling index. Fenbuconazole also caused a dose-dependent increase in liver microsomal cytochromes b 5 and P450 and the levels of immunoreactive CYP2B10 and its associated activity 7-pentoxyresorufin O-dealkylation (PROD). Treatment of mice with 1000 ppm phenobarbital elicited the same effects as treatment of mice with 1300 ppm fenbuconazole, except that phenobarbital was more effective than fenbuconazole at inducing PROD activity, even though fenbuconazole induced CYP2B10 to the same extent as did phenobarbital. This difference was attributed to the ability of fenbuconazole to bind tightly to CYP2B10 and partially mask its catalytic activity in liver microsomes, which is characteristic of several azole-containing drugs. All hepatocellular changes and induced enzyme activity returned to control levels within 4 weeks of discontinuing treatment with fenbuconazole or phenobarbital, indicating that the observed changes were fully reversible. We conclude that fenbuconazole is a phenobarbital-type inducer of mouse liver cytochrome P450, and the mode of action by which fenbuconazole induces liver adenomas in mice is similar to that of phenobarbital

Human cytochrome-P450 enzymes metabolize N-(2-methoxyphenyl)hydroxylamine, a metabolite of the carcinogens o-anisidine and o-nitroanisole, thereby dictating its genotoxicity.

Science.gov (United States)

Naiman, Karel; Martínková, Markéta; Schmeiser, Heinz H; Frei, Eva; Stiborová, Marie

2011-12-24

N-(2-Methoxyphenyl)hydroxylamine is a component in the human metabolism of two industrial and environmental pollutants and bladder carcinogens, viz. 2-methoxyaniline (o-anisidine) and 2-methoxynitrobenzene (o-nitroanisole), and it is responsible for their genotoxicity. Besides its capability to form three deoxyguanosine adducts in DNA, N-(2-methoxyphenyl)-hydroxylamine is also further metabolized by hepatic microsomal enzymes. To investigate its metabolism by human hepatic microsomes and to identify the major microsomal enzymes involved in this process are the aims of this study. N-(2-Methoxyphenyl)hydroxylamine is metabolized by human hepatic microsomes predominantly to o-anisidine, one of the parent carcinogens from which N-(2-methoxyphenyl)hydroxylamine is formed, while o-aminophenol and two N-(2-methoxyphenyl)hydroxylamine metabolites, whose exact structures have not been identified as yet, are minor products. Selective inhibitors of microsomal CYPs, NADPH:CYP reductase and NADH:cytochrome-b(5) reductase were used to characterize human liver microsomal enzymes reducing N-(2-methoxyphenyl)hydroxylamine to o-anisidine. Based on these studies, we attribute the main activity for this metabolic step in human liver to CYP3A4, 2E1 and 2C (more than 90%). The enzymes CYP2D6 and 2A6 also partake in this N-(2-methoxyphenyl)hydroxylamine metabolism in human liver, but only to ∼6%. Among the human recombinant CYP enzymes tested in this study, human CYP2E1, followed by CYP3A4, 1A2, 2B6 and 2D6, were the most efficient enzymes metabolizing N-(2-methoxyphenyl)hydroxylamine to o-anisidine. The results found in this study indicate that genotoxicity of N-(2-methoxyphenyl)hydroxylamine is dictated by its spontaneous decomposition to nitrenium/carbenium ions generating DNA adducts, and by its susceptibility to metabolism by CYP enzymes. Copyright © 2011 Elsevier B.V. All rights reserved.

Isolation and purification of rat liver morphine UDP-glucuronosyltransferase

International Nuclear Information System (INIS)

Puig, J.F.; Tephly, T.R.

1986-01-01

The enhancement of rat liver microsomal morphine (M) and 4-hydroxybiphenyl (4-HBP) UDP-glucuronyltransferase (UDPGT) activities by phenobarbital treatment has been proposed to represent increased activity of a single enzyme form, GT-2. They have separated M and 4-HBP UDPGT activities from Emulgen 911-solubilized microsomes obtained from livers of phenobarbital-treated Wistar rats. A sensitive assay procedure was developed to quantify M-UDPGT and 4-HBP-UDPGT activities using 14 C-UDP-glucuronic acid (UDPGA) and reversed phase C-18 minicolumns whereby the radioactive glucuronides were differentially eluted from labeled UDPGA. Trisacryl DEAE, and chromatofocusing procedures were employed to separate M-UDPGT and 4-HBP-UDPGT in the presence of exogenous phosphatidylcholine (PC). The PC is necessary to stabilize UDPGT activities. M-UDPGT was isolated to apparent homogeneity and displayed a monomeric molecular weight of 56,000 daltons on SDS-PAGE. It reacted with M but not with 4-HBP, bilirubin, p-nitrophenol, testosterone, androsterone, estrone, 4-aminobiphenyl or α-naphthylamine. 4-HBP-UDPGT did not react with M. Therefore, M and 4-HBP glucuronidations are catalyzed by separate enzymes in rat liver microsomes

Glucuronidation of trans-resveratrol by human liver and intestinal microsomes and UGT isoforms.

Science.gov (United States)

Brill, Shirley S; Furimsky, Anna M; Ho, Mark N; Furniss, Michael J; Li, Yi; Green, Adam G; Bradford, Wallace W; Green, Carol E; Kapetanovic, Izet M; Iyer, Lalitha V

2006-04-01

Resveratrol (trans-resveratrol, trans-3,5,4'-trihydroxystilbene) is a naturally occurring stilbene analogue found in high concentrations in red wine. There is considerable research interest to determine the therapeutic potential of resveratrol, as it has been shown to have tumour inhibitory and antioxidant properties. This study was performed to investigate the glucuronidation of resveratrol and possible drug interactions via glucuronidation. Two glucuronide conjugates, resveratrol 3-O-glucuronide and resveratrol 4'-O-glucuronide, were formed by human liver and intestinal microsomes. UGT1A1 and UGT1A9 were predominantly responsible for the formation of the 3-O-glucuronide (Km = 149 microM) and 4'-O-glucuronide (Km = 365 microM), respectively. The glucuronide conjugates were formed at higher levels (up to 10-fold) by intestinal rather than liver microsomes. Resveratrol was co-incubated with substrates of UGT1A1 (bilirubin and 7-ethyl-10-hydroxycamptothecin (SN-38)) and UGT1A9 (7-hydroxytrifluoromethyl coumarin (7-HFC)). No major changes were noted in bilirubin glucuronidation in the presence of resveratrol. Resveratrol significantly inhibited the glucuronidation of SN-38 (Ki = 6.2 +/- 2.1 microM) and 7-HFC (Ki = 0.6 +/- 0.2 microM). Hence, resveratrol has the potential to inhibit the glucuronidation of concomitantly administered therapeutic drugs or dietary components that are substrates of UGT1A1 and UGT1A9.

Age dependent in vitro metabolism of bifenthrin in rat and human hepatic microsomes.

Science.gov (United States)

Nallani, Gopinath C; Chandrasekaran, Appavu; Kassahun, Kelem; Shen, Li; ElNaggar, Shaaban F; Liu, Zhiwei

2018-01-01

Bifenthrin, a pyrethroid insecticide, undergoes oxidative metabolism leading to the formation of 4'-hydroxy-bifenthrin (4'-OH-BIF) and hydrolysis leading to the formation of TFP acid in rat and human hepatic microsomes. In this study, age-dependent metabolism of bifenthrin in rats and humans were determined via the rates of formation of 4'-OH-BIF and TFP acid following incubation of bifenthrin in juvenile and adult rat (PND 15 and PND 90) and human (18years) liver microsomes. Furthermore, in vitro hepatic intrinsic clearance (CL int ) of bifenthrin was determined by substrate consumption method in a separate experiment. The mean V max (±SD) for the formation of 4'-OH-BIF in juvenile rat hepatic microsomes was 25.0±1.5pmol/min/mg which was significantly lower (pbifenthrin occurs primarily via oxidative pathway with relatively lesser contribution (~30%) from hydrolytic pathway in both rat and human liver microsomes. The CL int values for bifenthrin, determined by monitoring the consumption of substrate, in juvenile and adult rat liver microsomes fortified with NADPH were 42.0±7.2 and 166.7±20.5μl/min/mg, respectively, and the corresponding values for human liver microsomes were 76.0±4.0 and 21.3±1.2μl/min/mg, respectively. The data suggest a major species difference in the age dependent metabolism of bifenthrin. In human liver microsomes, bifenthrin is metabolized at a much higher rate in juveniles than in adults, while the opposite appears to be true in rat liver microsomes. Copyright © 2017 Elsevier Inc. All rights reserved.

Hepatic mitochondrial and microsomal recovery of rats intoxicated with CCl/sub 4/

Energy Technology Data Exchange (ETDEWEB)

Hasegawa, T.; Hirai, Y.; Koga, N.; Tomokuni, K.

1983-01-01

The hepatic mitochondrial and microsomal recovery of rats intoxicated with CCl/sub 4/ was investigated with specific reference to the oxygen utilization of liver slices. In control rats, the major oxygen utilization of the liver slices was attributed to mitochondrial particles. Since the mitochondrial oxygen utilization was inhibited by cyanide, the microsomal oxygen utilization was induced by NADPH and phenobarbital (a substrate for microsomal mixed function oxidase). Changes in oxygen utilization were observed in the recovery course of rats intoxicated with CCl/sub 4/, and the recovery of mitochondria was found to be faster than that of microsomes. A sex difference was present in the recovery mechanism of the microsomes.

Metabolism of ginger component [6]-shogaol in liver microsomes from mouse, rat, dog, monkey, and human.

Science.gov (United States)

Chen, Huadong; Soroka, Dominique; Zhu, Yingdong; Sang, Shengmin

2013-05-01

There are limited data on the metabolism of [6]-shogaol (6S), a major bioactive component of ginger. This study demonstrates metabolism of 6S in liver microsomes from mouse, rat, dog, monkey, and human. The in vitro metabolism of 6S was compared among five species using liver microsomes from mouse, rat, dog, monkey, and human. Following incubations with 6S, three major reductive metabolites 1-(4'-hydroxy-3'-methoxyphenyl)-4-decen-3-ol (M6), 1-(4'-hydroxy-3'-methoxyphenyl)-decan-3-ol (M9), and 1-(4'-hydroxy-3'-methoxyphenyl)-decan-3-one (M11), as well as two new oxidative metabolites (1E,4E)-1-(4'-hydroxy-3'-methoxyphenyl)-deca-1,4-dien-3-one (M14) and (E)-1-(4'-hydroxy-3'-methoxyphenyl)-dec-1-en-3-one (M15) were found in all species. The kinetic parameters of M6 in liver microsomes from each respective species were quantified using Michaelis-Menten theory. A broad CYP-450 inhibitor, 1-aminobenzotriazole, precluded the formation of oxidative metabolites, M14 and M15, and 18β-glycyrrhetinic acid, an aldo-keto reductase inhibitor, eradicated the formation of the reductive metabolites M6, M9, and M11 in all species. Metabolites M14 and M15 were tested for cancer cell growth inhibition and induction of apoptosis and both showed substantial activity, with M14 displaying greater potency than 6S. We conclude that 6S is metabolized extensively in mammalian species mouse, rat, dog, monkey, and human, and that there are significant interspecies differences to consider when planning preclinical trials toward 6S chemoprevention. © 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Monkey liver cytochrome P450 2C9 is involved in caffeine 7-N-demethylation to form theophylline.

Science.gov (United States)

Utoh, Masahiro; Murayama, Norie; Uno, Yasuhiro; Onose, Yui; Hosaka, Shinya; Fujino, Hideki; Shimizu, Makiko; Iwasaki, Kazuhide; Yamazaki, Hiroshi

2013-12-01

Caffeine (1,3,7-trimethylxanthine) is a phenotyping substrate for human cytochrome P450 1A2. 3-N-Demethylation of caffeine is the main human metabolic pathway, whereas monkeys extensively mediate the 7-N-demethylation of caffeine to form pharmacological active theophylline. Roles of monkey P450 enzymes in theophylline formation from caffeine were investigated using individual monkey liver microsomes and 14 recombinantly expressed monkey P450 enzymes, and the results were compared with those for human P450 enzymes. Caffeine 7-N-demethylation activity in microsomes from 20 monkey livers was not strongly inhibited by α-naphthoflavone, quinidine or ketoconazole, and was roughly correlated with diclofenac 4'-hydroxylation activities. Monkey P450 2C9 had the highest activity for caffeine 7-N-demethylation. Kinetic analysis revealed that monkey P450 2C9 had a high Vmax/Km value for caffeine 7-N-demethylation, comparable to low Km value for monkey liver microsomes. Caffeine could dock favorably with monkey P450 2C9 modeled for 7-N-demethylation and with human P450 1A2 for 3-N-demethylation. The primary metabolite theophylline was oxidized to 8-hydroxytheophylline in similar ways by liver microsomes and by recombinant P450s in both humans and monkeys. These results collectively suggest a high activity for monkey liver P450 2C9 toward caffeine 7-N-demethylation, whereas, in humans, P450 1A2-mediated caffeine 3-N-demethylation is dominant.

Liver/kidney microsomal antibody type 1 targets CYP2D6 on hepatocyte plasma membrane

OpenAIRE

Muratori, L; Parola, M; Ripalti, A; Robino, G; Muratori, P; Bellomo, G; Carini, R; Lenzi, M; Landini, M; Albano, E; Bianchi, F

2000-01-01

BACKGROUND—Liver/kidney microsomal antibody type 1 (LKM1) is the marker of type 2 autoimmune hepatitis (AIH) and is detected in up to 6% of patients with hepatitis C virus (HCV) infection. It recognises linear and conformational epitopes of cytochrome P450IID6 (CYP2D6) and may have liver damaging activity, provided that CYP2D6 is accessible to effector mechanisms of autoimmune attack.
METHODS—The presence of LKM1 in the plasma membrane was investigated by indirect immunofluorescence and confo...

Effect of a New Prokinetic Agent DA-9701 Formulated with Corydalis Tuber and Pharbitidis Semen on Cytochrome P450 and UDP-Glucuronosyltransferase Enzyme Activities in Human Liver Microsomes

Directory of Open Access Journals (Sweden)

Hye Young Ji

2012-01-01

Full Text Available DA-9701 is a new botanical drug composed of the extracts of Corydalis tuber and Pharbitidis semen, and it is used as an oral therapy for the treatment of functional dyspepsia in Korea. The inhibitory potentials of DA-9701 and its component herbs, Corydalis tuber and Pharbitidis semen, on the activities of seven major human cytochrome P450 (CYP enzymes and four UDP-glucuronosyltransferase (UGT enzymes in human liver microsomes were investigated using liquid chromatography-tandem mass spectrometry. DA-9701 and Corydalis tuber extract slightly inhibited UGT1A1-mediated etoposide glucuronidation, with 50% inhibitory concentration (IC50 values of 188 and 290 μg/mL, respectively. DA-9701 inhibited CYP2D6-catalyzed bufuralol 1′-hydroxylation with an inhibition constant (Ki value of 6.3 μg/mL in a noncompetitive manner. Corydalis tuber extract competitively inhibited CYP2D6-mediated bufuralol 1′-hydroxylation, with a Ki value of 3.7 μg/mL, whereas Pharbitidis semen extract showed no inhibition. The volume in which the dose could be diluted to generate an IC50 equivalent concentration (volume per dose index value of DA-9701 for inhibition of CYP2D6 activity was 1.16 L/dose, indicating that DA-9701 may not be a potent CYP2D6 inhibitor. Further clinical studies are warranted to evaluate the in vivo extent of the observed in vitro interactions.

Cyclic fatty acid monomers from dietary heated fats affect rat liver enzyme activity.

Science.gov (United States)

Lamboni, C; Sébédio, J L; Perkins, E G

1998-07-01

This study was conducted to investigate the effects of dietary cyclic fatty acid monomers (CFAM), contained in heated fat from a commercial deep-fat frying operation, on rat liver enzyme activity. A partially hydrogenated soybean oil (PHSBO) used 7 d (7-DH) for frying foodstuffs, or 0.15% methylated CFAM diets was fed to male weanling rats in comparison to a control group fed a nonheated PHSBO (NH) diet in a 10-wk experiment. All diets were isocaloric with 15% fat. Animals fed either CFAM or 7-DH diets showed increased hepatic content of cytochrome (cyt.) b5 and P450 and increased activity of (E.C. 1.6.2.4) NADPH-cyt. P450 reductase in comparison to the control rats. In addition, the activities of (E.C. 2.3.1.21) carnitine palmitoyltransferase-I and (E.C. 1.1.1.42) isocitrate dehydrogenase were significantly decreased when compared to that of rats fed the NH diet. A significantly depressed activity of (E.C. 1.1.1.49) glucose 6-phosphate dehydrogenase was also observed for these animals compared to the control rats fed NH diet. Moreover, liver and microsomal proteins were significantly increased when CFAM or 7-DH diets were fed to animals in comparison to controls while liver glycogen was decreased significantly in experimental groups of rats. The results obtained in this study indicate that the CFAM in the diet from either synthetic sources or used fats increase the activity of liver enzyme systems that detoxify them.

Activation and detoxification metabolism of urban air pollutants 2-nitrobenzanthrone and carcinogenic 3-nitrobenzanthrone by rat and mouse hepatic microsomes.

Science.gov (United States)

Stiborova, Marie; Cechova, Tereza; Borek-Dohalska, Lucie; Moserova, Michaela; Frei, Eva; Schmeiser, Heinz H; Paca, Jan; Arlt, Volker M

2012-01-01

2-Nitrobenzanthrone (2-NBA) has recently been detected in ambient air particulate matter. Its isomer 3-nitrobenzanthrone (3-NBA) is a potent mutagen and suspected human carcinogen identified in diesel exhaust. Understanding which enzymes are involved in metabolism of these toxicants is important in the assessment of individual susceptibility. Here, metabolism of 2-NBA and 3-NBA by rat and mouse hepatic microsomes containing cytochromes P450 (CYPs), their reductase (NADPH:CYP reductase), and NADH:cytochrome b5 reductase was investigated under anaerobic and aerobic conditions. In addition, using the same microsomal systems, 2-NBA and 3-NBA were evaluated to be enzymatically activated under anaerobic conditions to species generating 2-NBA- and 3-NBA-derived DNA adducts. High performance liquid chromatography (HPLC) with ultraviolet (UV) detection was employed for the separation and characterization of 2-NBA and 3-NBA metabolites formed by hepatic microsomes of rats and mice under the anaerobic and aerobic conditions. Microsomal systems isolated from the liver of the control (untreated) rats and rats pretreated with Sudan I, β-naphthoflavone (β-NF), phenobarbital (PB), ethanol and pregnenolon 16α-carbonitrile (PCN), the inducers of cytochromes P450 (CYP) 1A1, 1A1/2, 2B, 2E1 and 3A, respectively, were used in this study. Microsomes of mouse models, a control mouse line (wild-type, WT) and Hepatic Cytochrome P450 Reductase Null (HRN) mice with deleted gene of NADPH:CYP reductase in the liver, thus absenting this enzyme in their livers, were also employed. To detect and quantify the 2-NBA- and 3-NBA-derived DNA adducts, the 32P postlabeling technique was used. Both reductive metabolite of 3-NBA, 3-aminobenzanthrone (3-ABA), found to be formed predominantly under the anaerobic conditions, and two 3-NBA oxidative metabolites, whose structures have not yet been investigated, were formed by several microsomal systems used in the study. Whereas a 3-NBA reductive metabolite

Influence of acute and chronic administration of methadone hydrochloride on NADPH-cytochrome c reductase and cytochrome P-450 of mouse liver microsomes.

Science.gov (United States)

Datta, R K; Johnson, E A; Bhattacharjee, G; Stenger, R J

1976-03-01

Administration of a single acute dose (20 mg/kg body weight) of methadone hydrochloride to both male and female mice increased the specific activity of NADPH-cytochrome c reductase and did not change much the content of cytochrome P-450 of their liver microsomes. Administration of multiple acute doses of methadone in male mice increased the specific activity of cytochrome c reductase and the content of cytochrome P-450 of their liver microsomes. Chronic administration of progressively increasing doses of methadone (up to 40 mg/kg body weight) to male mice increased the specific activity of c reductase. Similar chronic administration of methadone up to 28 mg/kg body weight also increased the microsomal content of P-450, but with higher doses of methadone, the content of P-450 declined and finally dropped slightly below control levels. The levels of c reductase activity and P-450 content returned to normal about two weeks after discontinuation of methadone administration.

Effect of radio-detoxified endotoxin on the liver microsomal drug metabolizing enzyme system in rats

International Nuclear Information System (INIS)

Bertok, L.; Szeberenyi, S.

1983-01-01

E. coli endotoxin (LPS) depresses the hepatic microsomal mono-oxygenase activity. Radio-detoxified LPS (TOLERIN: 60 Co irradiated endotoxin preparation) decreases this biotransforming activity to a smaller extent. Phenobarbital, an inducer of this mono-oxygenase system, failed to induce in LPS-treated animals. In radio-detoxified LPS-treated rats, phenobarbital induced the mono-oxygenase and almost fully restored the biotransformation

Isolation and structural elucidation of tiamulin metabolites formed in liver microsomes of pigs

DEFF Research Database (Denmark)

Lykkeberg, Anne Kruse; Cornett, Claus; Halling-Sørensen, Bent

2006-01-01

Although the antimicrobial tiamulin is extensively metabolized in pigs, the metabolism is not well investigated. In this work the NADPH dependent metabolism of tiamulin in liver microsomes from pigs has been studied. The tiamulin metabolites formed in the incubations were analysed using LC-MS, an...... 20% of tiamulin was deethylated, 10% was hydroxylated in the 2beta-position and 7% was hydroxylated in the 8alpha-position. About 40% of tiamulin was metabolized during the incubation conditions used. The protein precipitation in the incubations was performed using perchloric acid...

Magnetic circular dichroism studies on microsomal aryl hydrocarbon hydroxylase: comparison with cytochrome b/sub 5/ and cytochrome P-450/sub cam/

Energy Technology Data Exchange (ETDEWEB)

Vickery, L; Salmon, A; Sauer, K

1975-01-01

Magnetic circular dichroism spectra are reported for the visible and near ultraviolet spectral regions of liver microsomes from dimethylbenzanthracene-treated rats. The sequential addition of NADH, dithionite, and carbon monoxide enables us to determine contributions to the magnetic circular dichroism by cytochromes b/sub 5/ and P-450, which dominate the spectra. The magnetic circular dichroism of the microsomal preparation is compared with that of purified oxidized and reduced cytochrome b/sub 5/ from pig liver and with the camphor-complexed and camphor-free oxidized, reduced, and reduced carbonmonoxy cytochrome P-450/sub cam/ from Pseudomonas putida. The magnetic circular dichroism spectra of the membrane bound cytochrome b/sub 5/ are similar to those of the purified protein, indicating that little or no alteration in the environment of the heme occurs during the isolation procedure. The soluble bacterial cytochrome P-450/sub cam/ also appears to be a suitable model for microsomal P-450, although differences in the magnetic circular dichroism intensity are observed for the two enzymes. No effect of dimethylbenzanthracene on the magnetic circular dichroism spectra of induced compared to control rat microsomes could be observed.

The role of CYP2D6 in primary and secondary oxidative metabolism of dextromethorphan: in vitro studies using human liver microsomes.

Science.gov (United States)

Kerry, N L; Somogyi, A A; Bochner, F; Mikus, G

1994-01-01

1. The enzyme kinetics of dextromethorphan O-demethylation in liver microsomes from three extensive metabolisers (EM) with respect to CYP2D6 indicated high (Km1 2.2-9.4 microM) and low (Km2 55.5-307.3 microM) affinity sites whereas microsomes from two poor metabolisers (PM) indicated a single site (Km 560 and 157 microM). Similar differences were shown for 3-methoxymorphinan O-demethylation to 3-hydroxymorphinan (Km 6.9-9.6 microM in EM subjects; Km 307 and 213 microM in PM subjects). 2. Dextromethorphan O-demethylation was inhibited competitively by quinidine (Ki 0.1 microM), rac-perhexiline (Ki 0.4 microM), dextropropoxyphene (Ki 6 microM), rac-methadone (Ki 8 microM), and 3-methoxymorphinan (Ki 15 microM). These compounds were also potent inhibitors of 3-methoxymorphinan O-demethylation with IC50 values ranging from 0.02-12 microM. Anti-LKM1 serum inhibited both dextromethorphan and 3-methoxymorphinan O-demethylations in a titre-dependent manner. 3. The Michaelis-Menten constant for dextromethorphan N-demethylation to 3-methoxymorphinan (Km 632-977 microM) and dextrorphan N-demethylation to 3-hydroxymorphinan (Km 1571-4286 microM) did not differ between EM and PM microsomes. These N-demethylation reactions were not inhibited by quinidine and rac-methadone or LKM1 antibodies. 4. Dextromethorphan and 3-methoxymorphinan are metabolised by the same P450 isoform, CYP2D6, whereas the N-demethylation reactions are not carried out by CYP2D6. PMID:7826826

Three conazoles increase hepatic microsomal retinoic acid metabolism and decrease mouse hepatic retinoic acid levels in vivo

International Nuclear Information System (INIS)

Chen, P.-J.; Padgett, William T.; Moore, Tanya; Winnik, Witold; Lambert, Guy R.; Thai, Sheau-Fung; Hester, Susan D.; Nesnow, Stephen

2009-01-01

Conazoles are fungicides used in agriculture and as pharmaceuticals. In a previous toxicogenomic study of triazole-containing conazoles we found gene expression changes consistent with the alteration of the metabolism of all trans-retinoic acid (atRA), a vitamin A metabolite with cancer-preventative properties (Ward et al., Toxicol. Pathol. 2006; 34:863-78). The goals of this study were to examine effects of propiconazole, triadimefon, and myclobutanil, three triazole-containing conazoles, on the microsomal metabolism of atRA, the associated hepatic cytochrome P450 (P450) enzyme(s) involved in atRA metabolism, and their effects on hepatic atRA levels in vivo. The in vitro metabolism of atRA was quantitatively measured in liver microsomes from male CD-1 mice following four daily intraperitoneal injections of propiconazole (210 mg/kg/d), triadimefon (257 mg/kg/d) or myclobutanil (270 mg/kg/d). The formation of both 4-hydroxy-atRA and 4-oxo-atRA were significantly increased by all three conazoles. Propiconazole-induced microsomes possessed slightly greater metabolizing activities compared to myclobutanil-induced microsomes. Both propiconazole and triadimefon treatment induced greater formation of 4-hydroxy-atRA compared to myclobutanil treatment. Chemical and immuno-inhibition metabolism studies suggested that Cyp26a1, Cyp2b, and Cyp3a, but not Cyp1a1 proteins were involved in atRA metabolism. Cyp2b10/20 and Cyp3a11 genes were significantly over-expressed in the livers of both triadimefon- and propiconazole-treated mice while Cyp26a1, Cyp2c65 and Cyp1a2 genes were over-expressed in the livers of either triadimefon- or propiconazole-treated mice, and Cyp2b10/20 and Cyp3a13 genes were over-expressed in the livers of myclobutanil-treated mice. Western blot analyses indicated conazole induced-increases in Cyp2b and Cyp3a proteins. All three conazoles decreased hepatic atRA tissue levels ranging from 45-67%. The possible implications of these changes in hepatic atRA levels

Inhibition of rat microsomal lipid peroxidation by the oral administration of D002

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Menéndez R.

2000-01-01

Full Text Available The effect of D002, a defined mixture of higher primary alcohols purified from bee wax, on in vivo and in vitro lipid peroxidation was studied. The extent of lipid peroxidation was measured on the basis of the levels of thiobarbituric acid reactive substances (TBARS. When D002 (5-100 mg/kg body weight was administered orally to rats for two weeks, a partial inhibition of the in vitro enzymatic and non-enzymatic lipid peroxidation was observed in liver and brain microsomes. Maximal protection (46% occurred at a dose of 25 mg/kg. D002 behaved differently depending on both the presence of NADPH and the integrity of liver microsomes, which suggests that under conditions where microsomal metabolism was favored the protective effect of D002 was increased. D002 (25 mg/kg also completely inhibited carbon tetrachloride- and toluene-induced in vivo lipid peroxidation in liver and brain. Also, D002 significantly lowered in a dose-dependent manner the basal level of TBARS in liver (19-40% and brain (28-44% microsomes. We conclude that the oral administration of D002 (5, 25 and 100 mg/kg for two weeks protected rat liver and brain microsomes against microsomal lipid peroxidation in vitro and in vivo. Thus, D002 could be useful as a dietary natural antioxidant supplement. More studies are required before these data can be extrapolated to the recommendation for the use of D002 as a dietary antioxidant supplement for humans.

Effects of orally applied butyrate bolus on histone acetylation and cytochrome P450 enzyme activity in the liver of chicken – a randomized controlled trial

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Mátis Gábor

2013-01-01

Full Text Available Abstract Background Butyrate is known as histone deacetylase inhibitor, inducing histone hyperacetylation in vitro and playing a predominant role in the epigenetic regulation of gene expression and cell function. We hypothesized that butyrate, endogenously produced by intestinal microbial fermentation or applied as a nutritional supplement, might cause similar in vivo modifications in the chromatin structure of the hepatocytes, influencing the expression of certain genes and therefore modifying the activity of hepatic microsomal drug-metabolizing cytochrome P450 (CYP enzymes. Methods An animal study was carried out in chicken as a model to investigate the molecular mechanisms of butyrate’s epigenetic actions in the liver. Broiler chicks in the early post-hatch period were treated once daily with orally administered bolus of butyrate following overnight starvation with two different doses (0.25 or 1.25 g/kg body weight per day for five days. After slaughtering, cell nucleus and microsomal fractions were separated by differential centrifugation from the livers. Histones were isolated from cell nuclei and acetylation of hepatic core histones was screened by western blotting. The activity of CYP2H and CYP3A37, enzymes involved in biotransformation in chicken, was detected by aminopyrine N-demethylation and aniline-hydroxylation assays from the microsomal suspensions. Results Orally added butyrate, applied in bolus, had a remarkable impact on nucleosome structure of hepatocytes: independently of the dose, butyrate caused hyperacetylation of histone H2A, but no changes were monitored in the acetylation state of H2B. Intensive hyperacetylation of H3 was induced by the higher administered dose, while the lower dose tended to increase acetylation ratio of H4. In spite of the observed modification in histone acetylation, no significant changes were observed in the hepatic microsomal CYP2H and CYP3A37 activity. Conclusion Orally added butyrate in bolus

Hepatic microsomal metabolism of BDE-47 and BDE-99 by lesser snow geese and Japanese quail.

Science.gov (United States)

Krieger, Lisa K; Szeitz, András; Bandiera, Stelvio M

2017-09-01

In the present study, we investigated the oxidative biotransformation of 2,2',4,4'-tetrabromodiphenyl ether (BDE-47) and 2,2',4,4',5-pentabromodiphenyl ether (BDE-99) by liver microsomes from wild lesser snow geese (Chen caerulescens caerulescens) and domesticated Japanese quail (Coturnix japonica). Formation of hydroxy-metabolites was analyzed using an ultra-high performance liquid chromatography-tandem mass spectrometry-based method. Incubation of BDE-47 with avian liver microsomes produced sixteen hydroxy-metabolites, eight of which were identified using authentic standards. The major metabolites formed by liver microsomes from individual lesser snow geese were 4-hydroxy-2,2',3,4'-tetrabromodiphenyl ether (4-OH-BDE-42), 3-hydroxy-2,2',4,4'-tetrabromodiphenyl ether (3-OH-BDE-47), and 4'-hydroxy-2,2',4,5'-tetrabromodiphenyl ether (4'-OH-BDE-49). By comparison, 4-OH-BDE-42 and 4'-OH-BDE-49, but not 3-OH-BDE-47, were major metabolites of Japanese quail liver microsomes. Unidentified metabolites included monohydroxy- and dihydroxy-tetrabromodiphenyl ethers. Incubation of BDE-99 with avian liver microsomes produced seventeen hydroxy-metabolites, twelve of which were identified using authentic standards. The major metabolites formed by lesser snow goose liver microsomes were 2,4,5-tribromophenol, 3-OH-BDE-47, 4'-OH-BDE-49, 4-hydroxy-2,2',3,4',5-pentabromodiphenyl ether (4-OH-BDE-90), and 5'-hydroxy-2,2',4,4',5-pentabromodiphenyl ether (5'-OH-BDE-99). By comparison, the major metabolites produced by liver microsomes from Japanese quail included 6-hydroxy-2,2',4,4'-tetrabromodiphenyl ether (6-OH-BDE-47) and 2-hydroxy-2',3,4,4',5-pentabromodiphenyl ether (2-OH-BDE-123), but not 3-OH-BDE-47. Unidentified metabolites consisted of monohydroxy-pentabromodiphenyl ethers, monohydroxy-tetrabromodiphenyl ethers and dihydroxy-tetrabromodiphenyl ethers. Another difference between the two species was that formation rates of BDE-47 and BDE-99 metabolites were greater with liver

Stereospecificity (ST) of the microsomal ethanol oxidizing system (MEOS)

International Nuclear Information System (INIS)

Alderman, J.; Kato, S.; Lasker, J.; Lieber, C.S.

1987-01-01

The ST of MEOS for the ethanol 1R hydrogen has been variously reported as absolute, partial or absent, with free radical involvement postulated in the latter case. To determine both the ST of MEOS and the participation of free radicals in the reaction, they investigated MEOS ST using 1R[1- 3 H] ethanol as substrate. ST is expressed as the fraction of 3 H labeling in acetaldehyde formed, relative to that in ethanol, and ranges from 0.5 to 0. Partial ST was observed using liver microsomes from both rats and hamsters; it significantly decreased after ethanol feeding. 0.1 mM desferrioxamine (dfx) did not increase ST in any of these microsomal preparations while ferric EDTA decreased it, suggesting that ethanol treatment induces a cytochrome P-450 with lower ST rather than increasing free radical involvement. This is supported by a virtual absence of ST observed in a reconstituted system containing purified hamster P-450/sub ALC/, a liver cytochrome P-450 isozyme induced in hamsters by ethanol treatment. Their results indicate that, unlike other enzymes that oxidize ethanol, MEOS has only partial ST. Thus, ST alone cannot be used as an index of free radical involvement but, when evaluated with the response of ST to dfx, it indicated that MEOS is unlikely to involve free radical attack on ethanol in solution

Liver/kidney microsomal antibody type 1 targets CYP2D6 on hepatocyte plasma membrane

Science.gov (United States)

Muratori, L; Parola, M; Ripalti, A; Robino, G; Muratori, P; Bellomo, G; Carini, R; Lenzi, M; Landini, M; Albano, E; Bianchi, F

2000-01-01

BACKGROUND—Liver/kidney microsomal antibody type 1 (LKM1) is the marker of type 2 autoimmune hepatitis (AIH) and is detected in up to 6% of patients with hepatitis C virus (HCV) infection. It recognises linear and conformational epitopes of cytochrome P450IID6 (CYP2D6) and may have liver damaging activity, provided that CYP2D6 is accessible to effector mechanisms of autoimmune attack.
METHODS—The presence of LKM1 in the plasma membrane was investigated by indirect immunofluorescence and confocal laser microscopy of isolated rat hepatocytes probed with 10 LKM1 positive sera (five from patients with AIH and five from patients with chronic HCV infection) and a rabbit polyclonal anti-CYP2D6 serum.
RESULTS—Serum from both types of patient stained the plasma membrane of non-permeabilised cells, where the fluorescent signal could be visualised as discrete clumps. Conversely, permeabilised hepatocytes showed diffuse submembranous/cytoplasmic staining. Adsorption with recombinant CYP2D6 substantially reduced plasma membrane staining and LKM1 immunoblot reactivity. Plasma membrane staining of LKM1 colocalised with that of anti-CYP2D6. Immunoprecipitation experiments showed that a single 50 kDa protein recognised by anti-CYP2D6 can be isolated from the plasma membrane of intact hepatocytes.
CONCLUSIONS—AIH and HCV related LKM1 recognise CYP2D6 exposed on the plasma membrane of isolated hepatocytes. This observation supports the notion that anti-CYP2D6 autoreactivity may be involved in the pathogenesis of liver damage.


Keywords: liver/kidney microsomal antibody type 1; autoimmunity; autoimmune hepatitis; hepatitis C virus infection; confocal microscopy PMID:10716687

Dietary saturated and monounsaturated fats protect against acute acetaminophen hepatotoxicity by altering fatty acid composition of liver microsomal membrane in rats

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Shim Eugene

2011-10-01

Full Text Available Abstract Background Dietary polyunsaturated fats increase liver injury in response to ethanol feeding. We evaluated the effect of dietary corn oil (CO, olive oil (OO, and beef tallow (BT on fatty acid composition of liver microsomal membrane and acute acetaminophen hepatotoxicity. Methods Male Sprague-Dawley rats were fed 15% (wt/wt CO, OO or BT for 6 weeks. After treatment with acetaminophen (600 mg/kg, samples of plasma and liver were taken for analyses of the fatty acid composition and toxicity. Results Treatment with acetaminophen significantly elevated levels of plasma GOT and GPT as well as hepatic TBARS but reduced hepatic GSH levels in CO compared to OO and BT groups. Acetaminophen significantly induced protein expression of cytochrome P450 2E1 in the CO group. In comparison with the CO diet, lower levels of linoleic acid, higher levels of oleic acids and therefore much lower ratios of linoleic to oleic acid were detected in rats fed OO and BT diets. Conclusions Dietary OO and BT produces similar liver microsomal fatty acid composition and may account for less severe liver injury after acetaminophen treatment compared to animals fed diets with CO rich in linoleic acid. These findings imply that types of dietary fat may be important in the nutritional management of drug-induced hepatotoxicity.

Effect of Curcuma longa on CYP2D6- and CYP3A4-mediated metabolism of dextromethorphan in human liver microsomes and healthy human subjects.

Science.gov (United States)

Al-Jenoobi, Fahad Ibrahim; Al-Thukair, Areej A; Alam, Mohd Aftab; Abbas, Fawkeya A; Al-Mohizea, Abdullah M; Alkharfy, Khalid M; Al-Suwayeh, Saleh A

2015-03-01

Effect of Curcuma longa rhizome powder and its ethanolic extract on CYP2D6 and CYP3A4 metabolic activity was investigated in vitro using human liver microsomes and clinically in healthy human subjects. Dextromethorphan (DEX) was used as common probe for CYP2D6 and CYP3A4 enzymes. Metabolic activity of CYP2D6 and CYP3A4 was evaluated through in vitro study; where microsomes were incubated with NADPH in presence and absence of Curcuma extract. In clinical study phase-I, six healthy human subjects received a single dose (30 mg) of DEX syrup, and in phase-II DEX syrup was administered with Curcuma powder. The enzyme CYP2D6 and CYP3A4 mediated O- and N-demethylation of dextromethorphan into dextrorphan (DOR) and 3-methoxymorphinan (3-MM), respectively. Curcuma extract significantly inhibited the formation of DOR and 3-MM, in a dose-dependent and linear fashion. The 100 μg/ml dose of curcuma extract produced highest inhibition, which was about 70 % for DOR and 80 % for 3-MM. Curcuma significantly increases the urine metabolic ratio of DEX/DOR but the change in DEX/3-MM ratio was statistically insignificant. Present findings suggested that curcuma significantly inhibits the activity of CYP2D6 in in vitro as well as in vivo; which indicates that curcuma has potential to interact with CYP2D6 substrates.

Influence of Chloramphenicol and Amoxicillin on Rat Liver ...

African Journals Online (AJOL)

This study examined the effect of chloramphenicol and amoxicillin on liver microsomal enzymes Ca2+-ATPase and Glucose-6-Phosphatase (G-6-P) and lipid peroxidation in rats. Male Wistar strain rats weighing 120 – 195 g were divided into four groups. Group one, the control group, received physiological saline, group ...

In vitro metabolism of the anti-androgenic fungicide vinclozolin by rat liver microsomes.

Science.gov (United States)

Sierra-Santoyo, Adolfo; Angeles-Soto, Esperanza; de Lourdes López-González, Ma; Harrison, Randy A; Hughes, Michael F

2012-03-01

Vinclozolin (V) is a fungicide used in agricultural settings. V administered to rats is hydrolyzed to 2-[[(3,5-dichlorophenyl)-carbamoyl]oxy]-2-methyl-3-butenoic acid (M1) and 3',5'-dichloro-2-hydroxy-2-methylbut-3-enanilide (M2). V, M1 and M2 have antiandrogenic properties by interacting with the androgen receptor. Data on V, M1 and M2 biotransformation are limited. Our objective was to characterize V metabolism by rat liver microsomes. V was incubated with non-treated adult male Long-Evans rat liver microsomes and NADPH. Several metabolites were detected following the extraction of incubate with acetonitrile and analysis by HPLC/DAD/MSD. One metabolite was identified as [3-(3,5-dichlorophenyl)-5-methyl-5-(1,2-dihydroxyethyl)-1,3-oxazolidine-2,4-dione] (M4), which was gradually converted to 3',5'-dichloro-2,3,4-trihydroxy-2-methylbutylanilide (M5). Both co-eluted in the same HPLC peak. Another metabolite ([M7]) was detected by UV but was unstable for mass spectral analysis. The K(M app) for co-eluted M4/M5 and [M7] was 53.7 and 135.4 μM, the V(max app) was 0.812 and 0.669 nmoles/min/mg protein, and CL(int) was 15.1 and 4.9 ml/min/g protein, respectively. Pilocarpine, orphenadrine and proadifen and anti-rat cytochrome P450 (CYP)2A, 2B and 3A antibodies inhibited M4/M5 and [M7] formation. These results indicate that V is efficiently metabolized by CYP. Determination of the metabolites of V will provide further insight into the relationship between toxicity and tissue dose of V and its metabolites.

Mitochondrial targeting of bilirubin regulatory enzymes: An adaptive response to oxidative stress

Energy Technology Data Exchange (ETDEWEB)

Muhsain, Siti Nur Fadzilah, E-mail: sitinurfadzilah077@ppinang.uitm.edu.my [The University of Queensland, National Research Centre for Environmental Toxicology (Entox), 4072 Brisbane, Queensland (Australia); Faculty of Pharmacy, University Teknologi Mara (Malaysia); Lang, Matti A., E-mail: m.lang@uq.edu.au [The University of Queensland, National Research Centre for Environmental Toxicology (Entox), 4072 Brisbane, Queensland (Australia); Abu-Bakar, A' edah, E-mail: a.abubakar@uq.edu.au [The University of Queensland, National Research Centre for Environmental Toxicology (Entox), 4072 Brisbane, Queensland (Australia)

2015-01-01

The intracellular level of bilirubin (BR), an endogenous antioxidant that is cytotoxic at high concentrations, is tightly controlled within the optimal therapeutic range. We have recently described a concerted intracellular BR regulation by two microsomal enzymes: heme oxygenase 1 (HMOX1), essential for BR production and cytochrome P450 2A5 (CYP2A5), a BR oxidase. Herein, we describe targeting of these enzymes to hepatic mitochondria during oxidative stress. The kinetics of microsomal and mitochondrial BR oxidation were compared. Treatment of DBA/2J mice with 200 mg pyrazole/kg/day for 3 days increased hepatic intracellular protein carbonyl content and induced nucleo-translocation of Nrf2. HMOX1 and CYP2A5 proteins and activities were elevated in microsomes and mitoplasts but not the UGT1A1, a catalyst of BR glucuronidation. A CYP2A5 antibody inhibited 75% of microsomal BR oxidation. The inhibition was absent in control mitoplasts but elevated to 50% after treatment. An adrenodoxin reductase antibody did not inhibit microsomal BR oxidation but inhibited 50% of mitochondrial BR oxidation. Ascorbic acid inhibited 5% and 22% of the reaction in control and treated microsomes, respectively. In control mitoplasts the inhibition was 100%, which was reduced to 50% after treatment. Bilirubin affinity to mitochondrial and microsomal CYP2A5 enzyme is equally high. Lastly, the treatment neither released cytochrome c into cytoplasm nor dissipated membrane potential, indicating the absence of mitochondrial membrane damage. Collectively, the observations suggest that BR regulatory enzymes are recruited to mitochondria during oxidative stress and BR oxidation by mitochondrial CYP2A5 is supported by mitochondrial mono-oxygenase system. The induced recruitment potentially confers membrane protection. - Highlights: • Pyrazole induces oxidative stress in the mouse liver. • Pyrazole-induced oxidative stress induces mitochondrial targeting of key bilirubin regulatory enzymes, HMOX1

Mitochondrial targeting of bilirubin regulatory enzymes: An adaptive response to oxidative stress

International Nuclear Information System (INIS)

Muhsain, Siti Nur Fadzilah; Lang, Matti A.; Abu-Bakar, A'edah

2015-01-01

The intracellular level of bilirubin (BR), an endogenous antioxidant that is cytotoxic at high concentrations, is tightly controlled within the optimal therapeutic range. We have recently described a concerted intracellular BR regulation by two microsomal enzymes: heme oxygenase 1 (HMOX1), essential for BR production and cytochrome P450 2A5 (CYP2A5), a BR oxidase. Herein, we describe targeting of these enzymes to hepatic mitochondria during oxidative stress. The kinetics of microsomal and mitochondrial BR oxidation were compared. Treatment of DBA/2J mice with 200 mg pyrazole/kg/day for 3 days increased hepatic intracellular protein carbonyl content and induced nucleo-translocation of Nrf2. HMOX1 and CYP2A5 proteins and activities were elevated in microsomes and mitoplasts but not the UGT1A1, a catalyst of BR glucuronidation. A CYP2A5 antibody inhibited 75% of microsomal BR oxidation. The inhibition was absent in control mitoplasts but elevated to 50% after treatment. An adrenodoxin reductase antibody did not inhibit microsomal BR oxidation but inhibited 50% of mitochondrial BR oxidation. Ascorbic acid inhibited 5% and 22% of the reaction in control and treated microsomes, respectively. In control mitoplasts the inhibition was 100%, which was reduced to 50% after treatment. Bilirubin affinity to mitochondrial and microsomal CYP2A5 enzyme is equally high. Lastly, the treatment neither released cytochrome c into cytoplasm nor dissipated membrane potential, indicating the absence of mitochondrial membrane damage. Collectively, the observations suggest that BR regulatory enzymes are recruited to mitochondria during oxidative stress and BR oxidation by mitochondrial CYP2A5 is supported by mitochondrial mono-oxygenase system. The induced recruitment potentially confers membrane protection. - Highlights: • Pyrazole induces oxidative stress in the mouse liver. • Pyrazole-induced oxidative stress induces mitochondrial targeting of key bilirubin regulatory enzymes, HMOX1

Are there differences in the catalytic activity per unit enzyme of recombinantly expressed and human liver microsomal cytochrome P450 2C9? A systematic investigation into inter-system extrapolation factors.

Science.gov (United States)

Crewe, H K; Barter, Z E; Yeo, K Rowland; Rostami-Hodjegan, A

2011-09-01

The 'relative activity factor' (RAF) compares the activity per unit of microsomal protein in recombinantly expressed cytochrome P450 enzymes (rhCYP) and human liver without separating the potential sources of variation (i.e. abundance of enzyme per mg of protein or variation of activity per unit enzyme). The dimensionless 'inter-system extrapolation factor' (ISEF) dissects differences in activity from those in CYP abundance. Detailed protocols for the determination of this scalar, which is used in population in vitro-in vivo extrapolation (IVIVE), are currently lacking. The present study determined an ISEF for CYP2C9 and, for the first time, systematically evaluated the effects of probe substrate, cytochrome b5 and methods for assessing the intrinsic clearance (CL(int) ). Values of ISEF for S-warfarin, tolbutamide and diclofenac were 0.75 ± 0.18, 0.57 ± 0.07 and 0.37 ± 0.07, respectively, using CL(int) values derived from the kinetic values V(max) and K(m) of metabolite formation in rhCYP2C9 + reductase + b5 BD Supersomes™. The ISEF values obtained using rhCYP2C9 + reductase BD Supersomes™ were more variable, with values of 7.16 ± 1.25, 0.89 ± 0.52 and 0.50 ± 0.05 for S-warfarin, tolbutamide and diclofenac, respectively. Although the ISEF values obtained from rhCYP2C9 + reductase + b5 for the three probe substrates were statistically different (p system, with the intrinsic clearance calculated from full kinetic data is recommended for generation of the CYP2C9 ISEF. Furthermore, as ISEFs have been found to be sensitive to differences in accessory proteins, rhCYP system specific ISEFs are recommended. Copyright © 2011 John Wiley & Sons, Ltd.

Influence of whole body irradiation on induction of the hepatic microsomal system metabolizing drugs

International Nuclear Information System (INIS)

Szyszko, A.; Bitny-Szlachto, S.

1977-01-01

Effects of whole body irradiation (600 R) on rat liver aminophenazone demethylase activities of the liver homogenate 10,000 X g supernatant and its microsomal fraction were compared. Either activities were found to be decreased by irradiation by some 35%. The phenobarbital treatment (3 x 100 mg/kg i.p.) has turned out to provide higher relative augmentation of the liver demethylase activity in irradiated than in unirradiated rats. The cytoplasmic activity was found to be augmented by phenobarbital treatment 2,21-fold in unirradiated, and 3,20-fold in irradiated rats, and the microsomal activity increased 3,28-fold and 3,77-fold, respectively. Microsomal levels of cytochrome P-450 were found to be not affected by irradiation. (author)

Stereoselective in vitro metabolism of rhynchophylline and isorhynchophylline epimers of Uncaria rhynchophylla in rat liver microsomes.

Science.gov (United States)

Wang, Xin; Qiao, Zhou; Liu, Jia; Zheng, Mei; Liu, Wenyuan; Wu, Chunyong

2017-11-10

1. The objective was to investigate the underlying mechanism of the stereoselectivity in the metabolism of rhynchophylline (RIN) and isorhynchophylline (IRN) epimers in rat liver microsomes (RLM). 2. After incubation, eight metabolites of RIN (M1-5) and IRN (M6-8) reacted at A- and C-ring were identified using LC-Q-TOF/MS. Metabolic pathways included oxidation, hydroxylation, N-oxidation and dehydrogenation. In addition, hydroxylation at A-ring was the major metabolic pathway for RIN whereas the oxidation at C-ring was the major one for IRN. 3. Enzyme kinetics showed that the intrinsic clearance (CL int ) for IRN elimination was 1.9-fold higher than RIN and the degradation half-life (T 1/2 ) of RIN was 4.7-fold higher than that of IRN, indicating IRN was more favorable to be metabolized than RIN in RLM. 4. Data from chemical inhibition study demonstrated CYP3A was the predominant isoform involved in the metabolic elimination of both epimers, as well as the formation of M1-8. 5. In conclusion, data revealed that due to the spatial configurations at C-7 position, RIN and IRN epimers possessed different hepatic metabolic pathways and elimination rates which were mainly mediated by CYP3A.

Binding of bilirubin and its structural analogues to hepatic microsomal bilirubin UDP glucuronyltransferase

International Nuclear Information System (INIS)

Vanstapel, F.; Blanckaert, N.

1987-01-01

Hepatic glucuronidation of the asymmetrical natural bilirubin molecule results in formation of two different positional isomers, bilirubin C-8 monoglucuronide and bilirubin C-12 monoglucuronide. In view of the existence of multiple isoforms of UDPglucuronyltransferase, which is the microsomal enzyme system responsible for bilirubin esterification, the authors performed kinetic analysis of microsomal glucuronidation of bilirubin and a number of its structural congeners to determine whether synthesis of the two monoglucuronide isomers involved two distinct substrate-binding sites or reflected two different modes of binding to a single catalytic site. Both isomers were found in all tested species (man, rat, guinea pig, sheep), but there were marked species differences in the C-8/C-12 ratio of monoglucuronide found in bile or formed by liver microsomes. Correspondence between in vivo and in vitro results for such regioselectivity of glucuronidation was excellent in each species. On the basis of these results of kinetic analysis of bilirubin esterification at variable pigment substrate concentrations and inhibition studies with alternative substrates, the authors postulate that both natural monoglucuronide isomers are synthesized at a single binding site. Possible mechanisms responsible for the markedly regioselective esterification of bilirubin by rat and sheep liver were investigated by study of glucuronidation of selected structural analgoues of the pigment. Collectively, their findings suggest that the molecular from(s) of bilirubin able to engage in catalytically effective binding to UDPglucuronyltransferase does (do) not correspond with intramolecularly hydrogen-bonded conformers and that the nature of the β-substituents of the outer pyrromethenone rings is a key determinant of glucuronidation rate

Activation versus inhibition of microsomal glutathione S-transferase activity by acrolein. Dependence on the concentration and time of acrolein exposure.

Science.gov (United States)

Sthijns, Mireille M J P E; den Hartog, Gertjan J M; Scasso, Caterina; Haenen, Jan P; Bast, Aalt; Haenen, Guido R M M

2017-09-25

The toxicity of acrolein, an α,β-unsaturated aldehyde, is due to its soft electrophilic nature and primarily involves the adduction of protein thiols. The thiol glutathione (GSH) forms the first line of defense against acrolein. The present study confirms that acrolein added to isolated rat liver microsomes can increase microsomal GSH transferase (MGST) activity 2-3 fold, which can be seen as a direct adaptive increase in the protection against acrolein. At a relatively high exposure level, acrolein appeared to inhibit MGST. The activation is due to adduction of thiol groups, and the inactivation probably involves adduction of amino groups in the enzyme by acrolein. The preference of acrolein to react with thiol groups over amino groups can explain why the enzyme is activated at a low exposure level and inhibited at a high exposure level of acrolein. These opposite forms of direct adaptation on the level of enzyme activity further narrow the thin line between survival and promotion of cell death, governed by the level of exposure. Copyright © 2017 The Authors. Published by Elsevier B.V. All rights reserved.

Characterization of the hepatic cytochrome P450 enzymes involved in the metabolism of 25I-NBOMe and 25I-NBOH

DEFF Research Database (Denmark)

Nielsen, Line Marie; Holm, Niels Bjerre; Leth-Petersen, Sebastian

2017-01-01

)ethylamino]methyl]phenol (25I-NBOH) and to characterize the metabolites. The following approaches were used to identify the main enzymes involved in primary metabolism: incubation with a panel of CYP and monoamine oxidase (MAO) enzymes and incubation in pooled human liver microsomes (HLM) with and without specific CYP...

Luminol-and lucigenin-amplified chemiluminescence with rat liver microsomes. Kinetics and influence of ascorbic acid, glutathione, dimethylsulfoxide, N-t-butyl-a-phenyl-nitrone, copper-ions and a copper complex, catalase, superoxide dismutase, hexobarbital and aniline.

Science.gov (United States)

Klinger, W; Karge, E; Kretzschmar, M; Rost, M; Schulze, H P; Dargel, R; Reinemann, C; Rein, H

1996-07-01

For the investigation of luminol (LM)-and lucigenin (LC)-amplified chemiluminescence (CL) in rat liver microsomes using both a liquid-scintillation counter (LKB/Wallac 1219 Rackbeta) and a Berthold luminometer (AutoLumat LB 953) optimal incubation mixtures and conditions and basic kinetics have been established. Whereas calibration curves for both LM- and LC-CL are performed with hydrogenperoxide (LC quantum yield is 6.25 fold higher as that of LM), distinct differences were revealed with microsomes, indicating that different reactive oxygen species (ROS) are determined: Both LM- and LC-CL follow the kinetics of enzymatic reactions in terms of dependence on protein and NADPH or NADH concentration, time course, temperature etc., but with differences. LM-CL does not work without addition of Fe2+, whereas LC-CL does. Both copper ions and copper bound in a complex abolish CL, LC-CL being much more sensitive. Isolated cytochrome P-450 (P450) and NADPH P450 reductase from liver of pheno-barbital treated rats alone proved to be inactive in LM-and LC-CL production, whereas te combination 1:1 without and with addition of lipid was highly active in both LM-and LC-CL. Ascorbic acid and glutathione as scavengers diminish both LM- and LC-CL in concentrations higher then 10(5). Dimethyl-sulfoxide (DMSO) was ineffective in LM-CL up to concentrations of 0.2 M, the very high concentration of 2 M diminished LM-CL only to 1/3. LC-CL was diminished starting at concentrations of 100 mM and at 2 M only 10% of maximum LC-CL was observed. The trap substance N-t-butyl-a-phenylnitrone (BNP) also diminished LC-CL more effectively than LM-CL. Clearcut differences were revealed by the addition of catalase and superoxide dismutase: both enzymes diminished LM-CL only, without any influence on LC-CL. Hexobarbital, a potent uncoupler of P450, enhances LM-CL fivefold, whereas LC-CL is barely influenced. Aniline (without uncoupling capability) decreased both LM-and LC-CL increasingly with increasing

Role of microsomal enzyme inducers in the reduction of misonidazole neurotoxicity

Energy Technology Data Exchange (ETDEWEB)

Jones, D.H.; Bleehen, N.M.; Workman, P.; Smith, N.C. (Cambridge Univ. (UK). Dept. of Clinical Oncology and Radiotherapeutics; Addenbrooke' s Hospital, Cambridge (UK))

1983-11-01

It has been shown that phenytoin, 300 mg daily for one week, produces consistent hepatic microsomal enzyme induction, resulting in a decrease of 25% in misonidazole half-life, without causing any toxicity per se. A longer period of administration gives only a slightly greater induction. Phenobarbitone in a daily dose of 90 mg causes a reduction of 18% and 23% in misonidazole half-life after 1 and 2 weeks' pre-treatment respectively, but is less suitable clinically because of its sedative effect. A further series of studies using phenytoin as the inducing agent has shown that, despite adequate enzyme induction and increased misonidazole metabolism, it is impossible to increase the total dose of misonidazole beyond the usually accepted value of 12 g/m/sup 2/ because of unacceptable neuropathy (a rate of 50% at a dose of 14 g/m/sup 2/ over three weeks). In single doses of above 3.0-4.0 g of misonidazole, severe nausea and vomiting are prominent, so that this side effect is a determining factor in the treatment fractionation. Audiometric studies show no correlation between the incidence of peripheral neuropathy and abnormal audiograms, and have no value in the early prediction of neurotoxicity.

Identification of cytochrome P450s involved in the metabolism of 6-benzyl-1-benzyloxymethyl-5-iodouracil (W-1) using human recombinant enzymes and rat liver microsomes in vitro.

Science.gov (United States)

Lu, Ying-Yuan; Cheng, Hai-Xu; Wang, Xin; Wang, Xiao-Wei; Liu, Jun-Yi; Li, Pu; Lou, Ya-Qing; Li, Jun; Lu, Chuang; Zhang, Guo-Liang

2017-08-01

1. The aim of this study was to identify the hepatic metabolic enzymes, which involved in the biotransformation of 6-benzyl-1-benzyloxymethyl-5-iodouracil (W-1), a novel non-nucleoside reverse transcriptase inhibitor (NNRTI) in rat and human in vitro. 2. The parent drug of W-1 was incubated with rat liver microsomes (RLMs) or recombinant CYPs (CYP1A2, CYP2A6, CYP2B6, CYP2C8, CYP2C9, CYP2C19, CYP2D6, CYP2E1, CYP3A4, and CYP3A5, respectively) in the presence or absence of nicotinamide adeninedinucleotide phosphate (NADPH)-regenerating system. The metabolites of W-1 were analyzed with liquid chromatography-ion trap-time of flight-mass spectrometry (LC-IT-TOF-MS). 3. The parent drug of W-1 was metabolized in a NADPH-dependent manner in RLMs. The kinetic parameters of prototype W-1 including K m , V max , and CL int were 2.3 μM, 3.3 nmol/min/mg protein, and 1.4 mL/min/mg protein, respectively. Two metabolites M1 and M2 were observed in shorter retention times (2.988 and 3.188 min) with a higher molecular ion at m/z 463.0160 (both M1 and M2) than that of the W-1 parent drug (6.158 min with m/z 447.0218). The CYP selective inhibition and recombinant enzymes also showed that two hydroxyl metabolites M1 and M2 are mainly mediated by CYP2C19 and CYP3A4. 4. The identification of CYPs involved in W-1 biotransformation is important to understand and minimize, if possible, the potential of drug-drug interactions.

The role of microsomal enzyme inducers in the reduction of misonidazole neurotoxicity

International Nuclear Information System (INIS)

Jones, D.H.; Bleehen, N.M.; Workman, P.; Smith, N.C.

1983-01-01

It has been shown that phenytoin, 300 mg daily for one week, produces consistent hepatic microsomal enzyme induction, resulting in a decrease of 25% in misonidazole half-life, without causing any toxicity per se. A longer period of administration gives only a slightly greater induction. Phenobarbitone in a daily dose of 90 mg causes a reduction of 18% and 23% in misonidazole half-life after 1 and 2 weeks' pre-treatment respectively, but is less suitable clinically because of its sedative effect. A further series of studies using phenytoin as the inducing agent has shown that, despite adequate enzyme induction and increased misonidazole metabolism, it is impossible to increase the total dose of misonidazole beyond the usually accepted value of 12 g/m 2 because of unacceptable neuropathy (a rate of 50% at a dose of 14 g/m 2 over three weeks). In single doses of above 3.0-4.0 g of misonidazole, severe nausea and vomiting are prominent, so that this side effect is a determining factor in the treatment fractionation. Audiometric studies show no correlation between the incidence of peripheral neuropathy and abnormal audiograms, and have no value in the early prediction of neurotoxicity. (author)

Characterisation of the cytochrome P450 enzymes involved in the in vitro metabolism of granisetron.

Science.gov (United States)

Bloomer, J C; Baldwin, S J; Smith, G J; Ayrton, A D; Clarke, S E; Chenery, R J

1994-01-01

1. The metabolism of granisetron was investigated in human liver microsomes to identify the specific forms of cytochrome P450 responsible. 2. 7-hydroxy and 9'-desmethyl granisetron were identified as the major products of metabolism following incubation of granisetron with human liver microsomes. At low, clinically relevant, concentrations of granisetron the 7-hydroxy metabolite predominated. Rates of granisetron 7-hydroxylation varied over 100-fold in the human livers investigated. 3. Enzyme kinetics demonstrated the involvement of at least two enzymes contributing to the 7-hydroxylation of granisetron, one of which was a high affinity component with a Km of 4 microM. A single, low affinity, enzyme was responsible for the 9'-desmethylation of granisetron. 4. Granisetron caused no inhibition of any of the cytochrome P450 activities investigated (CYP1A2, CYP2A6, CYP2B6, CYP2C9/8, CYP2C19, CYP2D6, CYP2E1 and CYP3A), at concentrations up to 250 microM. 5. Studies using chemical inhibitors selective for individual P450 enzymes indicated the involvement of cytochrome P450 3A (CYP3A), both pathways of granisetron metabolism being very sensitive to ketoconazole inhibition. Correlation data were consistent with the role of CYP3A3/4 in granisetron 9'-desmethylation but indicated that a different enzyme was involved in the 7-hydroxylation. PMID:7888294

2,2',3,3',6,6'-Hexachlorobiphenyl (PCB 136) is Enantioselectively Oxidized to Hydroxylated Metabolites by Rat Liver Microsomes

Science.gov (United States)

Wu, Xianai; Pramanik, Ananya; Duffel, Michael W.; Hrycay, Eugene G.; Bandiera, Stelvio M.; Lehmler, Hans-Joachim; Kania-Korwel, Izabela

2011-01-01

Developmental exposure to multiple-ortho substituted polychlorinated biphenyls (PCBs) causes adverse neurodevelopmental outcomes in laboratory animals and humans by mechanisms involving the sensitization of Ryanodine receptors (RyRs). In the case of PCB 136, the sensitization of RyR is enantiospecific, with only (-)-PCB 136 being active. However, the role of enantioselective metabolism in the developmental neurotoxicity of PCB 136 is poorly understood. The present study employed hepatic microsomes from phenobarbital (PB-), dexamethasone (DEX-) and corn oil (VEH-)treated male Sprague-Dawley rats to investigate the hypothesis that PCB 136 atropisomers are enantioselectively metabolized by P450 enzymes to potentially neurotoxic, hydroxylated PCB 136 metabolites. The results demonstrated the time- and isoform-dependent formation of three metabolites, with 5-OH-PCB 136 (2,2',3,3',6,6'-hexachlorobiphenyl-5-ol) being the major metabolite. The formation of 5-OH-PCB 136 increased with the activity of P450 2B enzymes in the microsomal preparation, which is consistent with PCB 136 metabolism by rat P450 2B1. The minor metabolite 4-OH-PCB 136 (2,2',3,3',6,6'-hexachlorobiphenyl-4-ol) was produced by a currently unidentified P450 enzymes. An enantiomeric enrichment of (-)-PCB 136 was observed in microsomal incubations due to the preferential metabolism of (+)-PCB 136 to the corresponding 5-OH-PCB 136 (2,2',3,3',6,6'-hexachlorobiphenyl-5-ol) atropisomer. 4-OH-PCB 136 displayed an enrichment of the atropisomer formed from (-)-PCB 136; however, the enrichment of this metabolite atropisomer didn't affect the enantiomeric enrichment of the parent PCB because 4-OH-PCB 136 is only a minor metabolite. Although the formation of 5- and 4-OH-PCB 136 atropisomers increased with time, the enantioselective formation of the OH-PCB metabolites resulted in constant enantiomeric enrichment, especially at later incubation times. These observations not only demonstrate that the chiral signatures of

Anti-liver-kidney microsome antibody type 1 recognizes human cytochrome P450 db1.

Science.gov (United States)

Gueguen, M; Yamamoto, A M; Bernard, O; Alvarez, F

1989-03-15

Anti-liver-kidney microsome antibody type 1 (LKM1), present in the sera of a group of children with autoimmune hepatitis, was recently shown to recognize a 50 kDa protein identified as rat liver cytochromes P450 db1 and db2. High homology between these two members of the rat P450 IID subfamily and human P450 db1 suggested that anti-LKM1 antibody is directed against this human protein. To test this hypothesis, a human liver cDNA expression library in phage lambda GT-11 was screened using rat P450 db1 cDNA as a probe. Two human cDNA clones were found to be identical to human P450 db1 by restriction mapping. Immunoblot analysis using as antigen, the purified fusion protein from one of the human cDNA clones showed that only anti-LKM1 with anti-50 kDa reactivity recognized the fusion protein. This fusion protein was further used to develop an ELISA test that was shown to be specific for sera of children with this disease. These results: 1) identify the human liver antigen recognized by anti-LKM1 auto-antibodies as cytochrome P450 db1, 2) allow to speculate that mutation on the human P450 db1 gene could alter its expression in the hepatocyte and make it auto-antigenic, 3) provide a simple and specific diagnostic test for this disease.

Evaluation of liver marker enzymes and biochemical indices of ...

African Journals Online (AJOL)

Liver marker enzymes, total protein, amylase and glucose were evaluated in alloxan-induced diabetic wistar rats treated with aqueous extract of Pennisetum purpureum. The liver marker enzymes evaluated were alanine aminotransferase (ALT) and aspartate aminotransferase (AST). Sixteen wistar rats were grouped into ...

Modulation of catechol estrogen synthesis by rat liver microsomes: effects of treatment with growth hormone or testosterone

International Nuclear Information System (INIS)

Quail, J.A.; Jellinck, P.H.

1987-01-01

The ability of GH from various mammalian species, administered to normal mature male rats by constant infusion, to decrease the hepatic 2-hydroxylation of estradiol (E2) to female levels, as measured by the release of 3 H 2 O from [2-3H]E2, was determined. Rat and human GH (hGH) showed the highest activity while ovine GH was inactive. PRL (0.6 IU/h X kg) administered together with hGH (0.02 IU/h X kg) did not antagonize the feminizing action of GH. Infusion of hGH into male rats decreased the affinity of estradiol 2-hydroxylase for its steroid substrate and altered the linear Lineweaver-Burk plot towards a nonlinear hyperbolic plot characteristic of the female. The apparent Michaelis-Menten constant (Km) for the reaction was 1.69 microM for males and 2.75 microM for testosterone-treated ovariectomized females. An equal mixture of liver microsomes from male and female rats gave kinetic values similar to those observed with males alone. Neonatal imprinting with androgen did not alter the magnitude of the response of female rats to treatment with testosterone and/or GH at maturity and the androgen effect could only be shown in ovariectomized animals. The results with rats of different endocrine status were corroborated by the kinetic data and by the pattern of metabolites obtained with [4- 14 C]E2 when examined by TLC and autoradiography. The hormonal control of estradiol 2-hydroxylase, the key enzyme in catechol estrogen formation, and the contribution of sex-specific multiple forms of the enzyme to this reaction are discussed

Age dependence of rat liver function measurements

DEFF Research Database (Denmark)

Fischer-Nielsen, A; Poulsen, H E; Hansen, B A

1989-01-01

Changes in the galactose elimination capacity, the capacity of urea-N synthesis and antipyrine clearance were studied in male Wistar rats at the age of 8, 20 and 44 weeks. Further, liver tissue concentrations of microsomal cytochrome P-450, microsomal protein and glutathione were measured. All...... liver function measurements increased from the age of 8 to 44 weeks when expressed in absolute values. In relation to body weight, these function measurements were unchanged or reduced from week 8 to week 20. At week 44, galactose elimination capacity and capacity of urea-N synthesis related to body...... weight were increased by 10% and 36%, respectively, and antipyrine plasma clearance was reduced to 50%. Liver tissue concentrations of microsomal cytochrome P-450 and microsomal protein increased with age when expressed in absolute values, but were unchanged per g liver, i.e., closely related to liver...

Comparison of epoxide and free-radical mechanisms for activation of benzo[a]pyrene by Sprague-Dawley rat liver microsomes

International Nuclear Information System (INIS)

Selkirk, J.K.

1980-01-01

Coincubation of [6- 3 H]benzo[a]pyrene ([6- 3 H]BP) and [ 14 C]BP with SD rat liver microsomes produced metabolic profiles that showed that the C-6 of BP was not affected by formation of 4,5-dihydro-4,5-dihydroxy-BP, 7,8-dihydro-7,8-dihydroxy-BP, and 9,10-dihydro-9,10-dihydroxy-BP nor the 3- and 9-phenols of BP. Complete retention of tritium at C-6, except in the three quinones, confirmed the radical-cation model for formation of the 6-oxo-radical followed by oxidation to quinone. Epoxide formation at the carcinogenically active regions of BP appeared to biochemically isolate from 6-position activation and suggested that the microsomal epoxide pathway is unrelated to the radicalcation scheme. These molar ratios derived from double-label experiments reinforced the current literature that indicates the epoxide mechanism as the major pathway toward carcinogenic forms of BP

Determination of the 4-monohydroxy metabolites of perhexiline in human plasma, urine and liver microsomes by liquid chromatography.

Science.gov (United States)

Davies, Benjamin J; Herbert, Megan K; Coller, Janet K; Somogyi, Andrew A; Milne, Robert W; Sallustio, Benedetta C

2006-11-07

The use of perhexiline (PHX) is limited by hepatic and neurological toxicity associated with elevated concentrations in plasma that are the result of polymorphism of the cytochrome P450 2D6 isoform (CYP2D6). PHX is cleared by hepatic oxidation that produces three 4-monohydroxy metabolites: cis-OH-PHX, trans1-OH-PHX and trans2-OH-PHX. The current study describes an HPLC-fluorescent method utilising pre-column derivatization with dansyl chloride. Following derivatization, the metabolites were resolved on a C18 column with a gradient elution using a mobile phase composed of methanol and water. The method described is suitable for the quantification of the metabolites in human plasma and urine following clinical doses and for kinetic studies using human liver microsomes. The method demonstrates sufficient sensitivity, accuracy and precision between 5.0 and 0.01, 50.0 and 0.2 and 1.0 and 0.005 mg/l in human plasma, urine and liver microsomes, respectively, with intra-assay coefficients of variation and bias D6 extensive metaboliser (EM) patients at steady state with respect to PHX dosing determined that the mean (+/-S.D.) renal clearances of trans1-OH-PHX and cis-OH-PHX were 1.58+/-0.35 and 0.16+/-0.06l/h, respectively. The mean (+/-S.D.) dose recovered in urine as free and glucuronidated 4-monohydroxy PHX metabolites was 20.6+/-11.6%.

Distinct ontogenic patterns of overt and latent DGAT activities of rat liver microsomes.

Science.gov (United States)

Waterman, Ian J; Price, Nigel T; Zammit, Victor A

2002-09-01

We have studied the ontogeny of the two functional diacylglycerol acyltransferase (DGAT) activities (overt and latent) during postnatal development in rat liver. We find that the ontogenic patterns of the two are highly distinct. Overt DGAT shows a transient rise in activity up to day 4 postnatally, after which it declines until weaning; thereafter, it increases steadily to reach high adult values that may contribute to the high rates of turnover of cytosolic triacylglycerol (TAG). By contrast, latent DGAT activity increases continuously during the suckling period but falls sharply upon weaning onto chow but not onto a high-fat diet. Rates of TAG secretion by hepatocytes are higher than in the adult during the first 7 days after birth, and are largely dependent on the mobilization of the abundant intrahepatocyte TAG as a source of acyl moieties. When the hepatic steatosis is cleared (after day 7) the TAG secretion rate declines by 80% to reach adult values. Quantification of the content of mRNA for the DGAT1 and DGAT2 genes does not show correlation with either of the DGAT activities. We conclude that post-translational modification may play an important role in the overt and latent distribution of DGAT activity in the liver microsomal membrane.

The rabbit liver microsomal biotransformation of 1,1-dialkylethylenes: enantioface selection of epoxidation and enantioselectivity of epoxide hydrolysis.

Science.gov (United States)

Bellucci, G; Chiappe, C; Cordoni, A; Marioni, F

1994-01-01

The rabbit liver microsomal biotransformation of alpha-methylstyrene (1a), 2-methyl-1-hexene (1b), 2,4,4-trimethyl-1-pentene (1c), and 1,3,3-trimethyl-1-butene (1d) has been investigated with the aim at establishing the enantioface selection of the cytochrome P-450-promoted epoxidation of the double bond and the enantioselectivity of microsomal epoxide hydrolase(mEH)-catalyzed hydrolysis of the resulting epoxides. GLC on a Chiraldex G-TA (ASTEC) column was used to determine the enantiomeric composition of the products. The epoxides 2 first produced in incubations carried out in the presence of an NADPH regenerating system were not detected, being rapidly hydrolyzed by mEH to diols 3. The enantiomeric composition of the latter showed that no enantioface selection occurred in the epoxidation of 1c and 1d, and a very low (8%) ee of the (R)-epoxide was formed from 1b. Incubation of racemic epoxides 2b-d with the microsomal fraction showed that the mEH-catalyzed hydrolysis of 2c and 2d was practically nonenantioselective, while that of 2b exhibited a selectivity E = 4.9 favoring the hydrolysis of the (S)-enantiomer. A comparison of these results with those previously obtained for linear and branched chain alkyl monosubstituted oxiranes shows that the introduction of the second alkyl substituent suppresses the selectivity of the mEH reaction of the latter and reverses that of the former substrates.

Microsomal lipid peroxidation as a mechanism of cellular damage. [Dissertation

Energy Technology Data Exchange (ETDEWEB)

Kornbrust, D.J.

1979-01-01

The NADPH/iron-dependent peroxidation of lipids in rat liver microsomes was found to be dependent on the presence of free ferrous ion and maintains iron in the reduced Fe/sup 2 +/ state. Chelation of iron by EDTA inhibited peroxidation. Addition of iron, after preincubation of microsomes in the absence of iron, did not enhance the rate of peroxidation suggesting that iron acts by initiating peroxidative decomposition of membrane lipids rather than by catalyzing the breakdown of pre-formed hydroperoxides. Liposomes also underwent peroxidation in the presence of ferrous iron at a rate comparable to intact microsomes and was stimulated by ascorbate. Carbon tetrachloride initiated lipid peroxidation in the absence of free metal ions. Rates of in vitro lipid peroxidation of microsomes and homogenates were found to vary widely between different tissues and species. The effects of paraquat on lipid peroxidation was also studied. (DC)

Prediction of interindividual variation in drug plasma levels in vivo from individual enzyme kinetic data and physiologically based pharmacokinetic modeling

NARCIS (Netherlands)

Bogaards, J.J.P.; Hissink, E.M.; Briggs, M.; Weaver, R.; Jochemsen, R.; Jackson, P.; Bertrand, M.; Bladeren, P. van

2000-01-01

A strategy is presented to predict interindividual variation in drug plasma levels in vivo by the use of physiologically based pharmacokinetic modeling and human in vitro metabolic parameters, obtained through the combined use of microsomes containing single cytochrome P450 enzymes and a human liver

Evaluating correlation between serum liver enzymes and toxocariasis: a case control study

Directory of Open Access Journals (Sweden)

Hosein Miladi

2016-06-01

Full Text Available Objective: To evaluate the prevalence of toxocariasis in individuals with the normal and abnormal level of liver enzymes. Methods: In this case-control study, the serum samples were collected in the individuals referred to diagnostic laboratories of Arak in Iran. A total of 144 sera with abnormal level of liver enzymes were selected as cases and the same numbers of sera with the normal level of liver enzymes also were selected as controls. The sera were examined for anti-Toxocara IgG. Results: Twelve (4.2% sera contained anti-Toxocara antibody and all of them were in the case group. Although the mean of all liver enzymes was significantly different in the two groups (P < 0.05, statistical test showed no relationship between the level of liver enzymes and toxocariasis. Conclusions: It was concluded that the liver enzyme alteration is not the valid indicator for predicting toxocariasis. Because the kind of liver dysfunction, that is caused by the larvae of Toxocara, is unspecified, and it seems factors such as the number of larvae can play a basic role for the emergence of alterations.

Rat Liver Enzyme Release Depends on Blood Flow-Bearing Physical Forces Acting in Endothelium Glycocalyx rather than on Liver Damage

Directory of Open Access Journals (Sweden)

Julieta A. Díaz-Juárez

2017-01-01

Full Text Available We have found selective elevation of serum enzyme activities in rats subjected to partial hepatectomy (PH, apparently controlled by hemodynamic flow-bearing physical forces. Here, we assess the involvement of stretch-sensitive calcium channels and calcium mobilization in isolated livers, after chemical modifications of the endothelial glycocalyx and changing perfusion directionality. Inhibiting in vivo protein synthesis, we found that liver enzyme release is influenced by de novo synthesis of endothelial glycocalyx components, and released enzymes are confined into a liver “pool.” Moreover, liver enzyme release depended on extracellular calcium entry possibly mediated by stretch-sensitive calcium channels, and this endothelial-mediated mechanotransduction in liver enzyme release was also evidenced by modifying the glycocalyx carbohydrate components, directionality of perfusing flow rate, and the participation of nitric oxide (NO and malondialdehyde (MDA, leading to modifications in the intracellular distribution of these enzymes mainly as nuclear enrichment of “mitochondrial” enzymes. In conclusion, the flow-induced shear stress may provide fine-tuned control of released hepatic enzymes through mediation by the endothelium glycocalyx, which provides evidence of a biological role of the enzyme release rather to be merely a biomarker for evaluating hepatotoxicity and liver damage, actually positively influencing progression of liver regeneration in mammals.

Effects of thiol antioxidants on the atropselective oxidation of 2,2',3,3',6,6'-hexachlorobiphenyl (PCB 136) by rat liver microsomes.

Science.gov (United States)

Wu, Xianai; Lehmler, Hans-Joachim

2016-02-01

Chiral polychlorinated biphenyl (PCB) congeners, such as PCB 136, are atropselectively metabolized to various hydroxylated PCB metabolites (HO-PCBs). The present study investigates the effect of two thiol antioxidants, glutathione and N-acetyl-cysteine (NAC), on profiles and chiral signatures of PCB 136 and its HO-PCB metabolites in rat liver microsomal incubations. Liver microsomes prepared from rats pretreated with phenobarbital were incubated with PCB 136 (5 μM) in the presence of the respective antioxidant (0-10 mM), and levels and chiral signatures of PCB 136 and its HO-PCB metabolites were determined. Three metabolites, 5-136 (2,2',3,3',6,6'-hexachlorobiphenyl-5-ol), 4-136 (2,2',3,3',6,6'-hexachlorobiphenyl-4-ol), and 4,5-136 (2,2',3,3',6,6'-hexachlorobiphenyl-4,5-diol), were detected in all incubations, with 5-136 being the major metabolite. Compared to microsomal incubations without antioxidant, levels of 4,5-136 increased with increasing antioxidant concentration, whereas levels of PCB 136 and both mono-HO-PCBs were not affected by the presence of either antioxidant. PCB 136, 4-136, and 5-136 displayed significant atropisomeric enrichment; however, the direction and extent of the atropisomeric enrichment was not altered in the presence of an antioxidant. Because 4,5-136 can either be conjugated to a sulfate or glucuronide metabolite that is readily excreted or further oxidized a potentially toxic PCB 136 quinone, the effect of both thiol antioxidants on 4,5-136 formation suggests that disruptions of glutathione homeostasis may alter the balance between both metabolic pathways and, thus, PCB 136 toxicity in vivo.

Clofibric acid increases the formation of oleic acid in endoplasmic reticulum of the liver of rats.

Science.gov (United States)

Hirose, Akihiko; Yamazaki, Tohru; Sakamoto, Takeshi; Sunaga, Katsuyoshi; Tsuda, Tadashi; Mitsumoto, Atsushi; Kudo, Naomi; Kawashima, Yoichi

2011-01-01

The effects of 2-(4-chlorophenoxy)-2-methylpropionic acid (clofibric acid) on the formation of oleic acid (18:1) from stearic acid (18:0) and utilization of the 18:1 formed for phosphatidylcholine (PC) formation in endoplasmic reticulum in the liver of rats were studied in vivo. [¹⁴C]18:0 was intravenously injected into control Wistar male rats and rats that had been fed on a diet containing 0.5% (w/w) clofibric acid for 7 days; and the distribution of radiolabeled fatty acids among subcellular organelles, microsomes, peroxisomes, and mitochondria, was estimated on the basis of correction utilizing the yields from homogenates of marker enzymes for these organelles. The radioactivity was mostly localized in microsomes and the radiolabeled fatty acids present in microsomes were significantly increased by the treatment of rats with clofibric acid. The formation of radiolabeled 18:1 in microsomes markedly increased and incorporations of the formed [¹⁴C]18:1 into PC and phosphatidylethanolamine in microsomes were augmented in response to clofibric acid. The [¹⁴C]18:1 incorporated into PC was mostly located at the C-2 position, but not the C-1 position, of PC, and the radioactivity in 18:1 at the C-2 position of PC was strikingly increased by clofibric acid. These results obtained from the in vivo experiments directly link the findings that clofibric acid treatment induces microsomal stearoyl-CoA desaturase and 1-acylglycerophosphocholine acyltransferase in the liver and the findings that the treatment with the drug elevated absolute mass and mass proportion of 18:1 at the C-2 position, but not the C-1 position, of PC in the liver together.

Hydrolysis of pyrethroids by human and rat tissues: Examination of intestinal, liver and serum carboxylesterases

International Nuclear Information System (INIS)

Crow, J. Allen; Borazjani, Abdolsamad; Potter, Philip M.; Ross, Matthew K.

2007-01-01

Hydrolytic metabolism of pyrethroid insecticides in humans is one of the major catabolic pathways that clear these compounds from the body. Rodent models are often used to determine the disposition and clearance rates of these esterified compounds. In this study the distribution and activities of esterases that catalyze pyrethroid metabolism have been investigated in vitro using several human and rat tissues, including small intestine, liver and serum. The major esterase in human intestine is carboxylesterase 2 (hCE2). We found that the pyrethroid trans-permethrin is effectively hydrolyzed by a sample of pooled human intestinal microsomes (5 individuals), while deltamethrin and bioresmethrin are not. This result correlates well with the substrate specificity of recombinant hCE2 enzyme. In contrast, a sample of pooled rat intestinal microsomes (5 animals) hydrolyze trans-permethrin 4.5-fold slower than the sample of human intestinal microsomes. Furthermore, it is demonstrated that pooled samples of cytosol from human or rat liver are ∼ 2-fold less hydrolytically active (normalized per mg protein) than the corresponding microsomal fraction toward pyrethroid substrates; however, the cytosolic fractions do have significant amounts (∼ 40%) of the total esteratic activity. Moreover, a 6-fold interindividual variation in carboxylesterase 1 protein expression in human hepatic cytosols was observed. Human serum was shown to lack pyrethroid hydrolytic activity, but rat serum has hydrolytic activity that is attributed to a single CE isozyme. We purified the serum CE enzyme to homogeneity to determine its contribution to pyrethroid metabolism in the rat. Both trans-permethrin and bioresmethrin were effectively cleaved by this serum CE, but deltamethrin, esfenvalerate, alpha-cypermethrin and cis-permethrin were slowly hydrolyzed. Lastly, two model lipase enzymes were examined for their ability to hydrolyze pyrethroids. However, no hydrolysis products could be detected

Influence of sex hormones on relative quantities of multiple species of cytochrome P-450 in rat liver microsomes

International Nuclear Information System (INIS)

Fujita, S.; Peisach, J.; Chevion, M.; Hebrew Univ., Jerusalem

1981-01-01

EPR spectra of rat liver microsomes from male, female and hormonally-treated castrated hepatectomized rats were studied. The spectra, especially in the region of gsub(max) suggested a heterogeneity of local environments of the low spin ferric heme indicative of multiple structures for cytochrome P-450. Certain features in the spectrum correlated with sexual differences. It is suggested that the changes in the relative amplitudes of the EPR features represent differences in the relative abundance of the individual proteins in the mixture that, in turn, are related to the sexual differences of metabolic patterns for reactions catalyzed by cytochrome P-450. (author)

Characterization of in vitro glucuronidation clearance of a range of drugs in human kidney microsomes: comparison with liver and intestinal glucuronidation and impact of albumin.

Science.gov (United States)

Gill, Katherine L; Houston, J Brian; Galetin, Aleksandra

2012-04-01

Previous studies have shown the importance of the addition of albumin for characterization of hepatic glucuronidation in vitro; however, no reports exist on the effects of albumin on renal or intestinal microsomal glucuronidation assays. This study characterized glucuronidation clearance (CL(int, UGT)) in human kidney, liver, and intestinal microsomes in the presence and absence of bovine serum albumin (BSA) for seven drugs with differential UDP-glucuronosyltransferase (UGT) 1A9 and UGT2B7 specificity, namely, diclofenac, ezetimibe, gemfibrozil, mycophenolic acid, naloxone, propofol, and telmisartan. The impact of renal CL(int, UGT) on accuracy of in vitro-in vivo extrapolation (IVIVE) of glucuronidation clearance was investigated. Inclusion of 1% BSA for acidic drugs and 2% for bases/neutral drugs in incubations was found to be suitable for characterization of CL(int, UGT) in different tissues. Although BSA increased CL(int, UGT) in all tissues, the extent was tissue- and drug-dependent. Scaled CL(int, UGT) in the presence of BSA ranged from 2.22 to 207, 0.439 to 24.4, and 0.292 to 23.8 ml · min(-1) · g tissue(-1) in liver, kidney, and intestinal microsomes. Renal CL(int, UGT) (per gram of tissue) was up to 2-fold higher in comparison with that for liver for UGT1A9 substrates; in contrast, CL(int, UGT) for UGT2B7 substrates represented approximately one-third of hepatic estimates. Scaled renal CL(int, UGT) (in the presence of BSA) was up to 30-fold higher than intestinal glucuronidation for the drugs investigated. Use of in vitro data obtained in the presence of BSA and inclusion of renal clearance improved the IVIVE of glucuronidation clearance, with 50% of drugs predicted within 2-fold of observed values. Characterization and consideration of kidney CL(int, UGT) is particularly important for UGT1A9 substrates.

Enzymes and Inhibitors in Neonicotinoid Insecticide Metabolism

Science.gov (United States)

Shi, Xueyan; Dick, Ryan A.; Ford, Kevin A.; Casida, John E.

2009-01-01

Neonicotinoid insecticide metabolism involves considerable substrate specificity and regioselectivity of the relevant CYP450, aldehyde oxidase, and phase II enzymes. Human CYP450 recombinant enzymes carry out the following conversions: CYP3A4, 2C19 and 2B6 for thiamethoxam (TMX) to clothianidin (CLO); 3A4, 2C19 and 2A6 for CLO to desmethyl-CLO; 2C19 for TMX to desmethyl-TMX. Human liver aldehyde oxidase reduces the nitro substituent of CLO to nitroso much more rapidly than that of TMX. Imidacloprid (IMI), CLO and several of their metabolites do not give detectable N-glucuronides but 5-hydroxy-IMI, 4,5-diol-IMI and 4-hydroxy-thiacloprid are converted to O-glucuronides in vitro with mouse liver microsomes and UDP-glucuronic acid or in vivo in mice. Mouse liver cytosol with S-adenosylmethionine converts desmethyl-CLO to CLO but not desmethyl-TMX to TMX. Two organophosphorus CYP450 inhibitors partially block IMI, thiacloprid and CLO metabolism in vivo in mice, elevating the brain and liver levels of the parent compounds while reducing amounts of the hydroxylated metabolites. PMID:19391582

Induction of biotransformation enzymes by the carcinogenic air-pollutant 3-nitrobenzanthrone in liver, kidney and lung, after intra-tracheal instillation in rats.

Science.gov (United States)

Mizerovská, Jana; Dračínská, Helena; Frei, Eva; Schmeiser, Heinz H; Arlt, Volker M; Stiborová, Marie

2011-02-28

3-Nitrobenzanthrone (3-NBA), a carcinogenic air pollutant, was investigated for its ability to induce cytochrome P450 (CYP) 1A1/2 and NAD(P)H:quinone oxidoreductase (NQO1) in liver, kidney and lung of rats treated by intra-tracheal instillation. The organs used were from a previous study performed to determine the persistence of 3-NBA-derived DNA adducts in target and non-target tissues (Bieler et al., Carcinogenesis 28 (2007) 1117-1121, [22]). NQO1 is the enzyme reducing 3-NBA to N-hydroxy-3-aminobenzanthrone (N-OH-3-ABA) and CYP1A enzymes oxidize a human metabolite of 3-NBA, 3-aminobenzanthrone (3-ABA), to yield the same reactive intermediate. 3-NBA and 3-ABA are both activated to species forming DNA adducts by cytosols and/or microsomes isolated from rat lung, the target organ for 3-NBA carcinogenicity, and from liver and kidney. Each compound generated the same five DNA adducts detectable by (32)P-postlabelling. When hepatic cytosols from rats treated with 0.2 or 2mg/kg body weight of 3-NBA were incubated with 3-NBA, DNA adduct formation was 3.2- and 8.6-fold higher, respectively, than in incubations with cytosols from control animals. Likewise, cytosols isolated from lungs and kidneys of rats exposed to 3-NBA more efficiently activated 3-NBA than those of control rats. This increase corresponded to an increase in protein levels and enzymatic activities of NQO1. Incubations of hepatic, pulmonary or renal microsomes of 3-NBA-treated rats with 3-ABA led to an 9.6-fold increase in DNA-adduct formation relative to controls. The highest induction in DNA-adduct levels was found in lung. The stimulation of DNA-adduct formation correlated with expression of CYP1A1/2 induced by the intra-tracheal instillation of 3-NBA. The results demonstrate that 3-NBA induces NQO1 and CYP1A1/2 in livers, lungs and kidneys of rats after intra-tracheal instillation, thereby enhancing its own genotoxic and carcinogenic potential. Copyright © 2010 Elsevier B.V. All rights reserved.

Hepatic microsomal phospholipids in rats exposed intratracheally to coal fly ash

International Nuclear Information System (INIS)

Srivastava, P.K.; Chauhan, S.S.; Misra, U.K.

1986-01-01

The effects of intratracheal administration of fly ash (50 mg/kg body weight, daily for 7 days) on hepatic microsomal phospholipid metabolism has been studied in rats using various phospholipid precursors, viz NaH 2 32 PO 4 , (methyl- 14 C)-choline, and (methyl- 14 C)-methionine. Fly ash administration significantly increased microsomal phosphatidylcholine (PC), and lysophosphatidylcholine (LPC). The incorporation of NaH 2 32 PO 4 into total liver phospholipids, PC and Phosphatidyl ethanolamine (PE) was significantly increased in fly ash-treated rats as compared to the control. Fly ash administration also increased the incorporation of (methyl- 14 C)-choline into microsomal PC. Incorporation of (methyl- 14 C)-methionine into microsomal PC was not affected. Fly ash administration decreased the per cent distribution of arachidonic acid in PC and PE and increased that of oleic acid in PC and of linoleic acid in PE. (orig.)

Effect of x-irradiation on the activity of the key enzymes of the synthesis and degradation of heme in rat liver

International Nuclear Information System (INIS)

Deev, L.I.; Topchishvili, G.I.; Akhalaya, M.Ya.; Platonov, A.G.

1985-01-01

For Wistar rat males the effect of X radiation on the activity of delta-aminolevuminatesynthase and chemoxygenase - the rate of limiting enzymes of synthesis and catabolism of heme, as well as the level of ''free'' (non-utilized) heme in the rat liver are studied. The animals were irradiated using the X-ray device with the dose rate 0.5 Gy/min. X irradiation in the minimum lethal dose (7 Gy) caused evident changes in the activity of delta-aminolevuminatesynthase and chemoxygenase in the rat liver. Already one day after irradiation a noticeable decrease in the activity of delta-aminoleviminatesynthase (up to 81% of the normal one) was observed, and the maximum decrease took place on the third day (49% of the norm). On the 5-7-th day partial normalization of the activity of delta-aminoleviminatesynthase occurred. The chemoxygenase activity at the initial period after irradiation (1 day after) increased, after that (on the 5-7-th day), the phase of more than twofold and prolonged increase in the enzyme activity followed. Irradiation of animals by the dose of 5 Gy did not induce reliable changes in the activity of chemoxygenase and caused a considerably lower decrease in the activity of delta-aminolevunatesynthase. The observed suppression of synthesis and increase in heme catabolism do not result in the decrease in the level of ''free'' heme in the liver of irradiated animals. The conclusion is made on the presence of considerable violations of heme metabolism in the liver of irradiated animals, however, there is no final answer to the question on the role of detected effects in the decrease of microsomal level of cytochrome R-450

Overexpression of Catalase Enhances Benzo(a)pyrene Detoxification in Endothelial Microsomes.

Science.gov (United States)

Yang, Fang; Yang, Hong; Ramesh, Aramandla; Goodwin, J Shawn; Okoro, Emmanuel U; Guo, ZhongMao

2016-01-01

We previously reported that overexpression of catalase upregulated xenobiotic- metabolizing enzyme (XME) expression and diminished benzo(a)pyrene (BaP) intermediate accumulation in mouse aortic endothelial cells (MAECs). Endoplasmic reticulum (ER) is the most active organelle involved in BaP metabolism. To examine the involvement of ER in catalase-induced BaP detoxification, we compared the level and distribution of XMEs, and the profile of BaP intermediates in the microsomes of wild-type and catalase transgenic endothelial cells. Our data showed that endothelial microsomes were enriched in cytochrome P450 (CYP) 1A1, CYP1B1 and epoxide hydrolase 1 (EH1), and contained considerable levels of quinone oxidoreductase-1 (NQO1) and glutathione S-transferase-pi (GSTP). Treatment of wild-type MAECs with 1μM BaP for 2 h increased the expression of microsomal CYP1A1, 1B1 and NQO1 by ~300, 64 and 116%, respectively. However, the same treatment did not significantly alter the expression of EH1 and GSTP. Overexpression of catalase did not significantly increase EH1, but upregulated BaP-induced expression of microsomal CYP1A1, 1B1, NQO1 and GSTP in the following order: 1A1>NQO1>GSTP>1B1. Overexpression of catalase did not alter the distribution of each of these enzymes in the microsomes. In contrast to our previous report showing lower level of BaP phenols versus BaP diols/diones in the whole-cell, this report demonstrated that the sum of microsomal BaP phenolic metabolites were ~60% greater than that of the BaP diols/diones after exposure of microsomes to BaP. Overexpression of catalase reduced the concentrations of microsomal BaP phenols and diols/diones by ~45 and 95%, respectively. This process enhanced the ratio of BaP phenol versus diol/dione metabolites in a potent manner. Taken together, upregulation of phase II XMEs and CYP1 proteins, but not EH1 in the ER might be the mechanism by which overexpression of catalase reduces the levels of all the BaP metabolites, and

Effect of inhibition of microsomal Ca(2+)-ATPase on cytoplasmic calcium and enzyme secretion in pancreatic acini.

Science.gov (United States)

Metz, D C; Pradhan, T K; Mrozinski, J E; Jensen, R T; Turner, R J; Patto, R J; Gardner, J D

1994-01-13

We used thapsigargin (TG), 2,5-di-tert-butyl-1,4-benzohydroquinone (BHQ) and cyclopiazonic acid (CPA), each of which inhibits microsomal Ca(2+)-ATPase, to evaluate the effects of this inhibition on cytoplasmic free calcium ([Ca2+]i) and secretagogue-stimulated enzyme secretion in rat pancreatic acini. Using single-cell microspectrofluorimetry of fura-2-loaded acini we found that all three agents caused a sustained increase in [Ca2+]i by mobilizing calcium from inositol-(1,4,5)-trisphosphate-sensitive intracellular calcium stores and by promoting influx of extracellular calcium. Concentrations of all three agents that increased [Ca2+]i potentiated the stimulation of enzyme secretion caused by secretagogues that activate adenylate cyclase but inhibited the stimulation of enzyme secretion caused by secretagogues that activate phospholipase C. With BHQ, potentiation of adenylate cyclase-mediated enzyme secretion occurred immediately whereas inhibition of phospholipase C-mediated enzyme secretion occurred only after several min of incubation. In addition, the effects of BHQ and CPA on both [Ca2+]i and secretagogue-stimulated enzyme secretion were reversed completely by washing whereas the actions of TG could not be reversed by washing. Concentrations of BHQ in excess of those that caused maximal changes in [Ca2+]i inhibited all modes of stimulated enzyme secretion by a mechanism that was apparently unrelated to changes in [Ca2+]i. Finally, in contrast to the findings with TG and BHQ, CPA inhibited bombesin-stimulated enzyme secretion over a range of concentrations that was at least 10-fold lower than the range of concentrations over which CPA potentiated VIP-stimulated enzyme secretion.

Prevalence of abnormal plasma liver enzymes in older people with Type 2 diabetes.

Science.gov (United States)

Morling, J R; Strachan, M W J; Hayes, P C; Butcher, I; Frier, B M; Reynolds, R M; Price, J F

2012-04-01

To determine the prevalence and distribution of abnormal plasma liver enzymes in a representative sample of older adults with Type 2 diabetes. Plasma concentrations of alanine aminotransferase, aspartate aminotransferase and γ-glutamyltransferase were measured in a randomly selected, population-based cohort of 1066 men and women aged 60-75 years with Type 2 diabetes (the Edinburgh Type 2 Diabetes Study). Overall, 29.1% (95% CI 26.1-31.8) of patients had one or more plasma liver enzymes above the upper limit of the normal reference range. Only 10.1% of these patients had a prior history of liver disease and a further 12.4% reported alcohol intake above recommended limits. Alanine aminotransferase was the most commonly raised liver enzyme (23.1% of patients). The prevalence of abnormal liver enzymes was significantly higher in men (odds ratio 1.40, 95% CI 1.07-1.83), in the youngest 5-year age band (odds ratio 2.02, 95% CI 1.44-2.84), in patients with diabetes duration enzyme abnormality. The prevalence of elevated liver enzymes in people with Type 2 diabetes is high, with only modest variation between clinically defined patient groups. Further research is required to determine the prognostic value of raised, routinely measured liver enzymes to inform decisions on appropriate follow-up investigations. © 2011 The Authors. Diabetic Medicine © 2011 Diabetes UK.

Peroxisomal enzymes in the liver of rats with experimental diabetes mellitus type 2.

Science.gov (United States)

Turecký, L; Kupčová, V; Uhlíková, E; Mojto, V

2014-01-01

Diabetes mellitus is relatively frequently associated with fatty liver disease. Increased oxidative stress probably plays an important role in the development of this hepatopathy. One of possible sources of reactive oxygen species in liver is peroxisomal system. There are several reports about changes of peroxisomal enzymes in experimental diabetes, mainly enzymes of fatty acid oxidation. The aim of our study was to investigate the possible changes of activities of liver peroxisomal enzymes, other than enzymes of beta-oxidation, in experimental diabetes mellitus type 2. Biochemical changes in liver of experimental animals suggest the presence of liver steatosis. The changes of serum parameters in experimental group are similar to changes in serum of patients with non-alcoholic fatty liver disease. We have shown that diabetes mellitus influenced peroxisomal enzymes by the different way. Despite of well-known induction of peroxisomal beta-oxidation, the activities of catalase, aminoacid oxidase and NADH-cytochrome b(5) reductase were not significantly changed and the activities of glycolate oxidase and NADP-isocitrate dehydrogenase were significantly decreased. The effect of diabetes on liver peroxisomes is probably due to the increased supply of fatty acids to liver in diabetic state and also due to increased oxidative stress. The changes of metabolic activity of peroxisomal compartment may participate on the development of diabetic hepatopathy.

In vitro biotransformation of tris(2-butoxyethyl) phosphate (TBOEP) in human liver and serum

Energy Technology Data Exchange (ETDEWEB)

Van den Eede, Nele, E-mail: nele.vandeneede@uantwerpen.be [Toxicological Center, Department of Pharmaceutical Sciences, University of Antwerp, Universiteitsplein 1, 2610 Wilrijk, Antwerp (Belgium); Erratico, Claudio [Toxicological Center, Department of Pharmaceutical Sciences, University of Antwerp, Universiteitsplein 1, 2610 Wilrijk, Antwerp (Belgium); Exarchou, Vassiliki [Natural Products & Food Research and Analysis (NatuRA), Department of Pharmaceutical Sciences, University of Antwerp, Universiteitsplein 1, 2610 Wilrijk, Antwerp (Belgium); Maho, Walid; Neels, Hugo [Toxicological Center, Department of Pharmaceutical Sciences, University of Antwerp, Universiteitsplein 1, 2610 Wilrijk, Antwerp (Belgium); Covaci, Adrian, E-mail: adrian.covaci@uantwerpen.be [Toxicological Center, Department of Pharmaceutical Sciences, University of Antwerp, Universiteitsplein 1, 2610 Wilrijk, Antwerp (Belgium)

2015-04-15

Tris(2-butoxyethyl) phosphate (TBOEP) is a plasticizer present in indoor dust, reaching levels of several micrograms per gram. Such levels could lead to significant daily exposure of adults and children. Currently, no toxicokinetic data are available to estimate TBOEP clearance in humans after uptake and therefore, one objective of this study was to investigate intrinsic clearance of TBOEP by human liver microsome (HLM) and serum enzymes. Another objective was to generate information to identify and prioritize several metabolites of TBOEP for investigation of human exposure by biomonitoring. 1D and 2D-NMR methodologies were successfully applied on a mixture of the metabolites to confirm the structure of 3-HO-TBOEP (bis(2-butoxyethyl) 3-hydroxyl-2-butoxyethyl phosphate) and to tentatively assign structures to 1-HO-TBOEP and 2-HO-TBOEP. HO-TBOEP isomers and bis(2-butoxyethyl) phosphate (BBOEP), bis(2-butoxyethyl) hydroxyethyl phosphate (BBOEHEP) were further monitored by liquid chromatography–tandem mass spectrometry. Rates of formation of BBOEHEP and HO-TBOEP metabolites by liver enzymes were best described by the Michaelis–Menten model. Apparent K{sub m} values for BBOEHEP, 3-HO-TBOEP, and sum of 1- and 2-HO-TBOEP isomer formation were 152, 197 and 148 μM, respectively. Apparent V{sub max} values for the formation of BBOEHEP, 3-HO-TBOEP, and the sum of 1- and 2-HO-TBOEP isomers were 2560, 643, and 254 pmol/min/mg protein, respectively. No detectable formation of BBOEP occurred with liver or serum enzymes. Our findings indicate that intrinsic clearance of TBOEP is mainly catalyzed by oxidative enzymes in the liver and that its major in vitro metabolite is BBOEHEP. These findings can be applied in human biomonitoring studies and risk assessment. - Highlights: • First steps in the elucidation of TBOEP toxicokinetics • Quantification of TBOEP metabolites in human serum and liver microsomes • No detectable formation of BBOEP occurred with liver or serum

Mechanism of microsomal metabolism of benzene to phenol

Energy Technology Data Exchange (ETDEWEB)

Hinson, J.A.; Freeman, J.P.; Potter, D.W.; Mitchum, R.K.; Evans, F.E.

1985-05-01

The mechanism of microsomal hydroxylation of benzene to phenol has been studied by examining the microsomal metabolism of the specifically deuterated derivative 1,3,5-(/sub 2/H/sup 3/)benzene. Evidence for the formation of the following four products was obtained: 2,3,5-(/sub 2/H/sup 3/)phenol, 3,5-(/sub 2/H/sup 2/)phenol, 2,4,6-(/sub 2/H/sup 3/)phenol, and 2,4-(/sub 2/H/sup 2/)phenol. The presence of 2,3,5-(2H3)phenol and 2,4-(/sub 2/H/sup 2/)phenol shows that, in the microsomal metabolism of benzene to phenol, a NIH shift had occurred. A deuterium isotope effect (kH/kD) of approximately 4 was detected in both the meta- and para-deuterated phenols. This finding indicates that cyclohexadienone, formed either by isomerization of the epoxide or directly from the enzyme-substrate complex, is a major intermediate in the metabolism of benzene to phenol.

Effects of dexamethasone on liver enzymes and some serum ...

African Journals Online (AJOL)

Concomitant usage of dexamethasone and other medications may alter electrolyte metabolism and increase the formation of potentially hepatotoxic reactive metabolites which can contribute to elevated liver enzymes. The role of dexamethasone in liver functions and electrolyte metabolism during pregnancy in Yankasa ...

Mutagenic activation and detoxification of benzo[a]pyrene in vitro by hepatic cytochrome P450 1A1 and phase II enzymes in three meat-producing animals.

Science.gov (United States)

Darwish, W; Ikenaka, Y; Eldaly, E; Ishizuka, M

2010-01-01

The mutagenic activation activity of hepatic microsomes from three meat-producing animals (cattle, deer and horses) was compared with those of rats as a reference species. In the Ames Salmonella typhimurium TA98 assay, the liver microsomes of all examined animals mutagenically activated benzo[a]pyrene, an ideal promutagens, in terms of production of histidine-independent revertant colonies. The microsomes of horses had the highest ability to produce revertant colonies of the examined animals under both low and high substrate concentrations. Inhibition of this mutagenic activity using alpha-naphthoflavone, anti-rat CYP1A1, CYP3A2 and CYP2E1 antibodies suggests that this activity was mainly because of CYP1A1 in these animals as well as in rats. The addition of co-factors for two phase II enzymes, microsomal UDP glucoronosyl transferase and cytosolic glutathione-S-transferase, reduced the production of the revertant colonies in a concentration-dependent manner. Interestingly, horses had the highest reduction rate among the examined animals, suggesting that phase II enzymes play a great role in producing a state of balance between the bioactivation and detoxification of xenobiotics in these meat-producing animals. This report is the first to investigate the mutagenic activation activity of the hepatic microsomes and the role of phase II enzymes against this activity in meat-producing animals. Copyright (c) 2010 Elsevier Ltd. All rights reserved.

Evidence of a genetic basis for the different geographic occurrences of liver/kidney microsomal antibody type 1 in hepatitis C.

Science.gov (United States)

Muratori, Paolo; Czaja, Albert J; Muratori, Luigi; Granito, Alessandro; Guidi, Marcello; Ferri, Silvia; Volta, Umberto; Mantovani, Wilma; Pappas, Georgios; Cassani, Fabio; Lenzi, Marco; Bianchi, Francesco B

2007-01-01

Antibodies to liver/kidney microsome type 1 occur in Italian patients with hepatitis C, but rarely develop in North American patients. Our goals were to compare the frequencies of the HLA markers associated with autoimmune expression in Italian and North American patients with chronic hepatitis C and to determine genetic bases for regional differences in antibody production. HLA B8, DR3, DR4, DR7, DR11, DR13, DQ2, and the B8-DR3-DQ2 haplotype were determined by microlymphocytotoxicity and polymerase chain reaction in 105 Italian patients (50 with microsomal antibodies), 100 North American patients (none with microsomal antibodies), and Italian and North American healthy control subjects. Italian patients with microsomal antibodies differed from North American patients without these antibodies by having a higher frequency of HLA DR7 (54% vs. 27%, P=0.002). HLA DR7 occurred more frequently in seropositive Italian patients than in seronegative counterparts (54% vs. 11% P < 0.0001), Italian healthy control subjects (54% vs. 29%, P=0.0009), and North American healthy control subjects (54% vs. 19%, P < 0.0001). The frequency of HLA DR7 was similar in North American patients and controls (27% vs. 19%, P=0.2), but it was lower than in Italian controls (19% vs. 29%, P=0.059). Seropositive Italian patients had a lower frequency of HLA DR11 than seronegative Italian patients and Italian controls (18% vs. 34%, P=0.07, and 18% vs. 35%, P=0.02, respectively). In contrast to seropositive Italian patients, North American patients had HLA DR4 (30% vs. 12%, P=0.02), HLA DR13 (29% vs. 10%, P=0.01), and the B8-DR3-DQ2 haplotype (23% vs. 6%, P=0.01) more often. Similarly, HLA DR4 and the B8-DR3-DQ2 phenotype were more frequent in North American patients than in Italian controls (30% vs. 16%, P=0.005, and 23% vs. 7%, P=0.00002, respectively). HLA DR7 is associated with the development of microsomal antibodies in Italian patients with chronic hepatitis C. The lower frequency of HLA DR7

Liver/kidney microsomal antibody type 1 targets CYP2D6 on hepatocyte plasma membrane.

Science.gov (United States)

Muratori, L; Parola, M; Ripalti, A; Robino, G; Muratori, P; Bellomo, G; Carini, R; Lenzi, M; Landini, M P; Albano, E; Bianchi, F B

2000-04-01

Liver/kidney microsomal antibody type 1 (LKM1) is the marker of type 2 autoimmune hepatitis (AIH) and is detected in up to 6% of patients with hepatitis C virus (HCV) infection. It recognises linear and conformational epitopes of cytochrome P450IID6 (CYP2D6) and may have liver damaging activity, provided that CYP2D6 is accessible to effector mechanisms of autoimmune attack. The presence of LKM1 in the plasma membrane was investigated by indirect immunofluorescence and confocal laser microscopy of isolated rat hepatocytes probed with 10 LKM1 positive sera (five from patients with AIH and five from patients with chronic HCV infection) and a rabbit polyclonal anti-CYP2D6 serum. Serum from both types of patient stained the plasma membrane of non-permeabilised cells, where the fluorescent signal could be visualised as discrete clumps. Conversely, permeabilised hepatocytes showed diffuse submembranous/cytoplasmic staining. Adsorption with recombinant CYP2D6 substantially reduced plasma membrane staining and LKM1 immunoblot reactivity. Plasma membrane staining of LKM1 colocalised with that of anti-CYP2D6. Immunoprecipitation experiments showed that a single 50 kDa protein recognised by anti-CYP2D6 can be isolated from the plasma membrane of intact hepatocytes. AIH and HCV related LKM1 recognise CYP2D6 exposed on the plasma membrane of isolated hepatocytes. This observation supports the notion that anti-CYP2D6 autoreactivity may be involved in the pathogenesis of liver damage.

Changes in Liver Function Enzymes of HIV/AIDS Patients Treated ...

African Journals Online (AJOL)

user

on liver enzyme markers (Aspartate aminotransferase, Alanine aminotransferase ... the diagnosis and advanced infection of the liver cells by HIV. ... recommended guideline for the treatment of HIV ... HIV-positive patients not on treatment and.

Elevated liver enzymes in women with a family history of diabetes.

Science.gov (United States)

Inoue, Kazuo; Matsumoto, Masatoshi; Miyoshi, Yuji; Kobayashi, Yasuki

2008-03-01

Both elevated liver enzymes and a family history of diabetes mellitus (FHDM) are independent risk factors for type 2 diabetes. This study evaluates the epidemiological association between elevated liver enzymes and FHDM. Subjects included 3512 women workers without diabetes, hepatitis, a smoking habit, or a history of alcohol intake. Blood samples and personal data were collected from all subjects. Subjects with FHDM had a higher mean body mass index (BMI: 23.9kg/m(2) vs. 23.4kg/m(2); p=0.003). Laboratory testing also revealed higher mean fasting plasma glucose (FPG: 5.67mmol/L vs. 5.22mmol/L; penzymes were associated with FHDM. In particular, elevated GGT was related to FHDM, independent of the other variables. Elevated liver enzymes, probably due to fat deposition in the liver, may play a role in increasing the risk of diabetes in individuals with FHDM.

Influence of nutrition on liver oxidative metabolism.

Science.gov (United States)

Jorquera, F; Culebras, J M; González-Gallego, J

1996-06-01

The liver plays a major role in the disposition of the majority of drugs. This is due to the presence of several drug-metabolizing enzyme systems, including a group of membrane-bound mixed-function oxidative enzymes, mainly the cytochrome P450 system. Hepatic oxidative capacity can be assessed by changes in antipyrine metabolism. Different drugs and other factors may induce or inhibit the cytochrome P450-dependent system. This effect is important in terms of the efficacy or toxicity of drugs that are substrates for the system. Microsomal oxidation in animals fed with protein-deficient diets is depressed. The mixed-function oxidase activity recovers after a hyperproteic diet or the addition of lipids. Similar findings have been reported in patients with protein-calorie malnutrition, although results in the elderly are conflicting. Different studies have revealed that microsomal oxidation is impaired by total parenteral nutrition and that this effect is absent when changing the caloric source from carbohydrates to a conventional amino acid solution or after lipid addition, especially when administered as medium-chain/long-chain triglyceride mixtures. Peripheral parenteral nutrition appears to increase antipyrine clearance.

Interaction of rocuronium with human liver cytochromes P450

OpenAIRE

Anzenbacherova, Eva; Spicakova, Alena; Jourova, Lenka; Ulrichova, Jitka; Adamus, Milan; Bachleda, Petr; Anzenbacher, Pavel

2015-01-01

Rocuronium is a neuromuscular blocking agent acting as a competitive antagonist of acetylcholine. Results of an inhibition of eight individual liver microsomal cytochromes P450 (CYP) are presented. As the patients are routinely premedicated with diazepam, possible interaction of diazepam with rocuronium has been also studied. Results indicated that rocuronium interacts with human liver microsomal CYPs by binding to the substrate site. Next, concentration dependent inhibition of liver micro...

Metabolic profiling of five flavonoids from Dragon's Blood in human liver microsomes using high-performance liquid chromatography coupled with high resolution mass spectrometry.

Science.gov (United States)

Li, Yujuan; Zhang, Yushi; Wang, Rui; Wei, Lizhong; Deng, Yulin; Ren, Wei

2017-05-01

Although much is known about the pharmacological activities of Dragon's Blood (DB, a traditional Chinese herb), its metabolism in human liver microsomes (HLMs) and the cytochrome P450 (CYP) enzymes has not been studied. This study aims to identify the metabolic profile of five flavonoids (loureirin A, loureirin B, loureirin C, 7,4'-dihydroxyflavone and 5,7,4'-trihydroxyflavanone) from DB in HLMs as well as the CYP enzymes that are involved in the metabolism of them. High-resolution mass spectrometry was used to characterize the structures of their metabolites and 10 cDNA-expressed CYP enzymes (CYP1A2, CYP2A6, CYP2B6, CYP2C8, CYP2C9, CYP2C19, CYP2D6, CYP2E1, CYP3A4 and CYP3A5) were used to verify which isozymes mediate in the metabolism of the metabolites. Totally, 29 metabolites including 10 metabolites of loureirin A, 10 metabolites of loureirin B, 4 metabolites of loureirin C, 2 metabolites of 7,4'-dihydroxyflavone and 3 metabolites of 5,7,4'-trihydroxyflavanone were elucidated and identified on the basis of the high-resolution MS n data. The metabolic profile of the five flavonoids in HLMs involved hydroxylation, oxidation and demethylation. Among them, hydroxylation was the predominant biotransformation of the five flavonoids in HLMs, occurring in combination with other metabolic reactions. Assay with recombinant P450s revealed that CYP2C9 and CYP2C19 played an important role in the hydroxylation of flavonoids in HLMs. To the best of our knowledge, this is the first in vitro evaluation of the metabolic profile of loureirin A, loureirin B, loureirin C, 7,4'-dihydroxyflavone and 5,7,4'-trihydroxyflavanone in HLMs. Copyright © 2017 Elsevier B.V. All rights reserved.

Biochemical assessement of liver enzymes in immunocompromised ...

African Journals Online (AJOL)

Aim: This study aims at the estimation of serum alkaline phosphatase (ALP), alanine aminotransferase (ALT), Aspartate aminotransferase (AST), and glutmyltransferase GGT (Liver enzymes) in Human immunodeficiency virus(HIV) and/or Acquired immune deficiency syndrome(AIDS) patients in parts of Edo State, Nigeria.

Radioprotection of liver lipids of whole-body gamma-irradiated female rats by cystamine

International Nuclear Information System (INIS)

Ramanathan, R.; Misra, U.K.

1976-01-01

The effect of administration of cystamine (5 mg/100 g body weight) before 1,200 R whole-body gamma irradiation has been studied on irradiation-induced changes in liver and its subcellular fractions'lipids of fasted female rats. Cystamine prevented the irradiation-induced increase in liver triglycerides and liver mitochondrial total phospholipids, but it decreased microsomal total phospholipids and proteins. Cystamine prevented the radiation-induced increased 32 P-radioactivity (counts/min/μmole phospholipid phosphorus) of microsomal phosphatidyl choline. Cystamine prevented the radiation-induced increased uptake of NaH 2 32 PO 4 (counts/min/g liver) in liver microsomal phosphatidyl ethanolamine and supernatant phosphatidyl choline; but in microsomal phosphatidyl choline, cystamine did not do so, but on the other hand it itself increased the uptake in control rats. Cystamine did not prevent the irradiation-induced decreased incorporation of (U- 14 C)glucose into liver triglycerides, total phospholipids and phosphatidyl choline. Cystamine itself decreased the incorporation of (U- 14 C)glucose into liver triglycerides and phosphoglycerides of control rats. (orig.) [de

Cytochrome P450 2C8 and flavin-containing monooxygenases are involved in the metabolism of tazarotenic acid in humans.

Science.gov (United States)

Attar, Mayssa; Dong, Dahai; Ling, Kah-Hiing John; Tang-Liu, Diane D-S

2003-04-01

Upon oral administration, tazarotene is rapidly converted to tazarotenic acid by esterases. The main circulating agent, tazarotenic acid is subsequently oxidized to the inactive sulfoxide metabolite. Therefore, alterations in the metabolic clearance of tazarotenic acid may have significant effects on its systemic exposure. The objective of this study was to identify the human liver microsomal enzymes responsible for the in vitro metabolism of tazarotenic acid. Tazarotenic acid was incubated with 1 mg/ml pooled human liver microsomes, in 100 mM potassium phosphate buffer (pH 7.4), at 37 degrees C, over a period of 30 min. The microsomal enzymes that may be involved in tazarotenic acid metabolism were identified through incubation with microsomes containing cDNA-expressed human microsomal isozymes. Chemical inhibition studies were then conducted to confirm the identity of the enzymes potentially involved in tazarotenic acid metabolism. Reversed-phase high performance liquid chromatography was used to quantify the sulfoxide metabolite, the major metabolite of tazarotenic acid. Upon incubation of tazarotenic acid with microsomes expressing CYP2C8, flavin-containing monooxygenase 1 (FMO1), or FMO3, marked formation of the sulfoxide metabolite was observed. The involvement of these isozymes in tazarotenic acid metabolism was further confirmed by inhibition of metabolite formation in pooled human liver microsomes by specific inhibitors of CYP2C8 or FMO. In conclusion, the in vitro metabolism of tazarotenic acid to its sulfoxide metabolite in human liver microsomes is mediated by CYP2C8 and FMO.

Stimulation of NADH-dependent microsomal DNA strand cleavage by rifamycin SV.

Science.gov (United States)

Kukiełka, E; Cederbaum, A I

1995-04-15

Rifamycin SV is an antibiotic anti-bacterial agent used in the treatment of tuberculosis. This drug can autoxidize, especially in the presence of metals, and generate reactive oxygen species. A previous study indicated that rifamycin SV can increase NADH-dependent microsomal production of reactive oxygen species. The current study evaluated the ability of rifamycin SV to interact with iron and increase microsomal production of hydroxyl radical, as detected by conversion of supercoiled plasmid DNA into the relaxed open circular state. The plasmid used was pBluescript II KS(-), and the forms of DNA were separated by agarose-gel electrophoresis. Incubation of rat liver microsomes with plasmid plus NADH plus ferric-ATP caused DNA strand cleavage. The addition of rifamycin SV produced a time- and concentration-dependent increase in DNA-strand cleavage. No stimulation by rifamycin SV occurred in the absence of microsomes, NADH or ferric-ATP. Stimulation occurred with other ferric complexes besides ferric-ATP, e.g. ferric-histidine, ferric-citrate, ferric-EDTA, and ferric-(NH4)2SO4. Rifamycin SV did not significantly increase the high rates of DNA strand cleavage found with NADPH as the microsomal reductant. The stimulation of NADH-dependent microsomal DNA strand cleavage was completely blocked by catalase, superoxide dismutase, GSH and a variety of hydroxyl-radical-scavenging agents, but not by anti-oxidants that prevent microsomal lipid peroxidation. Redox cycling agents, such as menadione and paraquat, in contrast with rifamycin SV, stimulated the NADPH-dependent reaction; menadione and rifamycin SV were superior to paraquat in stimulating the NADH-dependent reaction. These results indicate that rifamycin SV can, in the presence of an iron catalyst, increase microsomal production of reactive oxygen species which can cause DNA-strand cleavage. In contrast with other redox cycling agents, the stimulation by rifamycin SV is more pronounced with NADH than with NADPH as the

Effect of ethionine on hepatic mitochondrial and microsomal calcium uptake

International Nuclear Information System (INIS)

Agarwal, A.K.; Zinermon, W.D.; Latoni, L.

1988-01-01

Ethionine, an ethyl analog of methionine, produces a variety of physiological and pathological effects in animals. These range from acute effects in the liver, kidney, pancreas, and other organs to liver carcinogenesis. Female rats when injected with ethionine exhibit a rapid decrease in hepatic adenosine triphosphate levels followed by a marked inhibition of RNA and protein synthesis and accumulation of triglycerides. Since calcium transport in mitochondria and microsomes is ATP dependent, it becomes interesting to find out if ethionine administration has any effect on subcellular calcium transport. Calcium has recently gained an increased controversy regarding its role in chemical induced lethal cell damage. Certain groups believe that influx of extracellular calcium across the damaged plasma membrane might actually mediate the irreversible damage to the cell, whereas according to other, entry of calcium into the cell is secondary to the damage. The present study was carried out to investigate the calcium [ 45 Ca] transport in mitochondria and microsomes following ethionine administration. The effect of carbon tetrachloride on calcium uptake in ethionine treated rats was also studied

EFFECTS OF THIOL ANTIOXIDANTS ON THE ATROPSELECTIVE OXIDATION OF 2,2′,3,3′,6,6′-HEXACHLOROBIPHENYL (PCB 136) BY RAT LIVER MICROSOMES

Science.gov (United States)

Wu, Xianai; Lehmler, Hans-Joachim

2015-01-01

Chiral polychlorinated biphenyl (PCB) congeners, such as PCB 136, are atropselectively metabolized to various hydroxylated PCB metabolites (HO-PCBs). The present study investigates the effect of two thiol antioxidants, glutathione and N-acetyl-cysteine (NAC), on profiles and chiral signatures of PCB 136 and its HO-PCB metabolites in rat liver microsomal incubations. Liver microsomes prepared from rats pretreated with phenobarbital were incubated with PCB 136 (5 μM) in the presence of the respective antioxidant (0–10 mM), and levels and chiral signatures of PCB 136 and its HO-PCB metabolites were determined. Three metabolites, 5-136 (2,2′,3,3′,6,6′-hexachlorobiphenyl-5-ol), 4-136 (2,2′,3,3′,6,6′-hexachlorobiphenyl-4-ol) and 4,5-136 (2,2′,3,3′,6,6′-hexachlorobiphenyl-4,5-diol), were detected in all incubations, with 5-136 being the major metabolite. Compared to microsomal incubations without antioxidant, levels of 4,5-136 increased with increasing antioxidant concentration, whereas levels of PCB 136 and both mono-HO-PCBs were not affected by the presence of either antioxidant. PCB 136, 4-136 and 5-136 displayed significant atropisomeric enrichment; however, the direction and extent of the atropisomeric enrichment was not altered in the presence of an antioxidant. Because 4,5-136 can either be conjugated to a sulfate or glucuronide metabolite that is readily excreted or further oxidized a potentially toxic PCB 136 quinone, the effect of both thiol antioxidants on 4,5-136 formation suggests that disruptions of glutathione homeostasis may alter the balance between both metabolic pathways and, thus, PCB 136 toxicity in vivo. PMID:26155892

Immunological cross-reactivity to multiple autoantigens in patients with liver kidney microsomal type 1 autoimmune hepatitis.

Science.gov (United States)

Choudhuri, K; Gregorio, G V; Mieli-Vergani, G; Vergani, D

1998-11-01

We describe two patients with liver kidney microsomal antibody type 1 (LKM1)-positive autoimmune hepatitis (AIH) with associated endocrinopathies. The first patient had insulin-dependent diabetes (IDDM), and the second patient had Addison's disease and hypoparathyroidism, and is also positive for islet cell antibodies, without overt diabetes. To account for the existence of multiple endocrinopathy in these patients, we investigated whether there is sequence similarity between the target of LKM1 antibodies, cytochrome P4502D6 (CYP2D6), and other human proteins, and if so, whether this structural similarity produces a detectable cross-reactive immune response. Our database search identified two proteins, carboxypeptidase H, an autoantigen in insulin-dependent diabetes, and 21-hydroxylase, the major autoantigen in Addison's disease, that share sequence similarity to the second major LKM1 epitope on CYP2D6. We tested the reactivity of sera from these patients to the homologous regions of the three autoantigens using an enzyme-linked immunosorbent assay (ELISA). The cut-off for positivity was established by testing sera from 22 healthy children. To determine the significance of reactivity to the peptide homologues of the three autoantigens, we investigated 16 additional patients with LKM1 AIH and 20 children with chronic hepatitis B virus infection as pathological controls. We found that reactivity to the second major epitope of CYP2D6 is significantly associated with reactivity to the homologous regions of carboxypeptidase H (CPH) and 21-hydroxylase (21-OHase) in patients with LKM1 AIH, and that this simultaneous recognition is cross-reactive. We suggest that a cross-reactive immune response between homologous autoantigens may contribute to the development of multiple endocrinopathies in LKM1 AIH.

Sex-related difference in the inductions by perfluoro-octanoic acid of peroxisomal beta-oxidation, microsomal 1-acylglycerophosphocholine acyltransferase and cytosolic long-chain acyl-CoA hydrolase in rat liver.

Science.gov (United States)

Kawashima, Y; Uy-Yu, N; Kozuka, H

1989-01-01

Inductions by perfluoro-octanoic acid (PFOA) of hepatomegaly, peroxisomal beta-oxidation, microsomal 1-acylglycerophosphocholine acyltransferase and cytosolic long-chain acyl-CoA hydrolase were compared in liver between male and female rats. Marked inductions of these four parameters were seen concurrently in liver of male rats, whereas the inductions in liver of female rats were far less pronounced. The sex-related difference in the response of rat liver to PFOA was much more marked than that seen with p-chlorophenoxyisobutyric acid (clofibric acid) or 2,2'-(decamethylenedithio)diethanol (tiadenol). Hormonal manipulations revealed that this sex-related difference in the inductions is strongly dependent on sex hormones, namely that testosterone is necessary for the inductions, whereas oestradiol prevented the inductions by PFOA. PMID:2570571

Microsomal detoxication enzyme responses of the marine snail, Thais haemastoma, to laboratory oil exposure

International Nuclear Information System (INIS)

Livingstone, D.R.; Stickle, W.B.; Kapper, M.; Wang, S.

1986-01-01

The cytochrome P-450 monooxygenase or mixed function oxidase (MFO) system is a widely distributed enzyme system involved in the detoxication of foreign organic compounds (xenobiotics) taken up by organisms. Increases in the activities of the MFO system, occur with exposure of the organism to organic xenobiotics and such responses in the field have been proposed as a means of identifying biological impact by organic pollution. The carnivorous marine gastropod Thais haemastoma, or southern oyster drill, rapidly accumulated polynuclear aromatic and other hydrocarbons from the environment, through both the food source and the water-column. In laboratory experiments T. haemastoma were exposed to the water soluble fraction (WSF) of South Louisiana crude oil and the responses of the MFO system examined. Preliminary characterization of the snail MFO system was carried out using methodology developed from studies on the common mussel Mytilus edulis. Microsomal benz[a]pyrene hydroxylase (BPH), NADH- and NADPH- dependent cytochrome c reductase (NAD(P)H-CYTCRED) and NADH-dependent ferricyanide reductase (NADH-FERRIRED) activities were measured but it was not possible to determine cytochrome P-450 or b 5

A case of moderate liver enzyme elevation after acute acetaminophen overdose despite undetectable acetaminophen level and normal initial liver enzymes.

Science.gov (United States)

Bebarta, Vikhyat S; Shiner, Drew C; Varney, Shawn M

2014-01-01

Liver function test (LFT) increase is an early sign of acetaminophen (APAP) toxicity. Typically, when an acute overdose patient is evaluated and has an initial undetectable APAP level and normal liver enzymes, the patient is not treated with N-acetylcysteine, and liver enzymes are not expected to increase later. We report a case of moderate LFT increase despite normal LFTs and an undetectable APAP level after delayed presentation of an APAP ingestion. A 22-year-old male with no medical history ingested 15-25 hydrocodone/APAP tablets (5 mg/500 mg). His suicide note and his bunkmate corroborated the overdose time. He arrived at the emergency department 16 hours after ingestion. At that time, his APAP level was enzymes were normal [aspartate transaminase (AST) 31 U/L and alanine transaminase (ALT) 34 U/L]. Twenty-nine hours after ingestion, the psychiatry team obtained LFTs (AST 45, ALT 61). He had persistent nausea and diffuse abdominal pain. On repeat analysis, the APAP level at 36 hours was found to be <10 μg/mL, AST 150, and ALT 204. After 2 more days of increasing LFTs and persistent abdominal pain and nausea, the toxicology department was consulted, the patient was transferred to the medicine department, and intravenous N-acetylcysteine was started 66 hours after ingestion. He was treated for 16 hours and had a significant decline in LFTs and symptom resolution. His prothrombin time, bilirubin, lactate, creatinine, and mental status were normal throughout the admission. Other cases of LFT increase were excluded. Our case report illustrates that a moderate increase in liver transaminase may occur despite an initial undetectable APAP level and normal transaminases after a delayed presentation. In our case, no serious clinical effects were reported.

Overlapping but distinct specificities of anti-liver-kidney microsome antibodies in autoimmune hepatitis type II and hepatitis C revealed by recombinant native CYP2D6 and novel peptide epitopes

Science.gov (United States)

Klein, R; Zanger, U M; Berg, T; Hopf, U; Berg, P A

1999-01-01

Anti-liver-kidney microsome antibodies (anti-LKM) occur in autoimmune hepatitis (AIH) type II and in a subset of patients with hepatitis C. Anti-LKM1 in AIH are directed against cytochrome P4502D6 (CYP2D6), but conflicting data exist concerning the specificity of anti-LKM in hepatitis C. The aim of this study was to evaluate binding specificities of anti-LKM antibodies in both diseases using novel test antigens as well as their inhibitory capacity on CYP2D6 enzyme activity. Sera from 22 patients with AIH type II and 17 patients with hepatitis C being anti-LKM-positive in the immunofluorescence test were investigated for binding to native recombinant CYP2D6 and liver microsomes by ELISA and immunoblotting, and to synthetic peptides covering the region 254–339 (254–273, 257–269, 270–294, 291–310, 307–324, 321–339, 373–389) as well as the novel peptide 196–218 by ELISA. Furthermore, all sera were tested for inhibition of CYP2D6-dependent bufuralol 1′-hydroxylase activity. Twenty of the 22 AIH type II sera (91%) and nine of the 17 hepatitis C sera (53%) were positive for CYP2D6 by ELISA and/or immunoblotting. The previously described major peptide epitope comprising CYP2D6 amino acids 257–269 was recognized by 16 of the 22 AIH sera but by only one hepatitis C serum. A further epitope, 196–218, could be defined for the first time as another immunodominant epitope for AIH because it was recognized by 15 of the 22 AIH (68%) but only three of the 17 hepatitis C sera (18%). With the exception of the peptide 254–273, the other peptides showed no significant reactivity. Analysing the inhibitory properties of anti-LKM antibodies it emerged that 95% of AIH sera and 88% of hepatitis C sera inhibited enzyme function. These data indicate that anti-LKM antibodies in AIH and hepatitis C react with CYP2D6, as shown by their inhibitory activity, and that besides the known epitope 257–269 a further immunodominant epitope exists on CYP2D6 which is recognized

Altered carbohydrate, lipid, and xenobiotic metabolism by liver from rats flown on Cosmos 1887

Science.gov (United States)

Merrill, A. H. Jr; Hoel, M.; Wang, E.; Mullins, R. E.; Hargrove, J. L.; Jones, D. P.; Popova, I. A.; Merrill AH, J. r. (Principal Investigator)

1990-01-01

To determine the possible biochemical effects of prolonged weightlessness on liver function, samples of liver from rats that had flown aboard Cosmos 1887 were analyzed for protein, glycogen, and lipids as well as the activities of a number of key enzymes involved in metabolism of these compounds and xenobiotics. Among the parameters measured, the major differences were elevations in the glycogen content and hydroxymethylglutaryl-CoA (HMG-CoA) reductase activities for the rats flown on Cosmos 1887 and decreases in the amount of microsomal cytochrome P-450 and the activities of aniline hydroxylase and ethylmorphine N-demethylase, cytochrome P-450-dependent enzymes. These results support the earlier finding of differences in these parameters and suggest that altered hepatic function could be important during spaceflight and/or the postflight recovery period.

Association of liver enzymes and computed tomography markers of liver steatosis with familial longevity.

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Michiel Sala

Full Text Available OBJECTIVE: Familial longevity is marked by enhanced peripheral but not hepatic insulin sensitivity. The liver has a critical role in the pathogenesis of hepatic insulin resistance. Therefore we hypothesized that the extent of liver steatosis would be similar between offspring of long-lived siblings and control subjects. To test our hypothesis, we investigated the extent of liver steatosis in non-diabetic offspring of long-lived siblings and age-matched controls by measuring liver enzymes in plasma and liver fat by computed tomography (CT. RESEARCH DESIGN AND METHODS: We measured nonfasting alanine transaminase (ALT, aspartate aminotransferase (AST, and Υ-glutamyl transferase (GGT in 1625 subjects (736 men, mean age 59.1 years from the Leiden Longevity Study, comprising offspring of long-lived siblings and partners thereof. In a random subgroup, fasting serum samples (n = 230 were evaluated and CT was performed (n = 268 for assessment of liver-spleen (L/S ratio and the prevalence of moderate-to-severe non-alcoholic fatty liver disease (NAFLD. Linear mixed model analysis was performed adjusting for age, gender, body mass index, smoking, use of alcohol and hepatotoxic medication, and correlation of sibling relationship. RESULTS: Offspring of long-lived siblings had higher nonfasting ALT levels as compared to control subjects (24.3 mmol/L versus 23.2 mmol/L, p = 0.03, while AST and GGT levels were similar between the two groups. All fasting liver enzyme levels were similar between the two groups. CT L/S ratio and prevalence of moderate-to-severe NAFLD was similar between groups (1.12 vs 1.14, p = 0.25 and 8% versus 8%, p = 0.91, respectively. CONCLUSIONS: Except for nonfasting levels of ALT, which were slightly higher in the offspring of long-lived siblings compared to controls, no differences were found between groups in the extent of liver steatosis, as assessed with liver biochemical tests and CT. Thus, our data indicate that the extent of liver

Association of liver enzymes and computed tomography markers of liver steatosis with familial longevity.

Science.gov (United States)

Sala, Michiel; Kroft, Lucia J M; Röell, Boudewijn; van der Grond, Jeroen; Slagboom, P Eline; Mooijaart, Simon P; de Roos, Albert; van Heemst, Diana

2014-01-01

Familial longevity is marked by enhanced peripheral but not hepatic insulin sensitivity. The liver has a critical role in the pathogenesis of hepatic insulin resistance. Therefore we hypothesized that the extent of liver steatosis would be similar between offspring of long-lived siblings and control subjects. To test our hypothesis, we investigated the extent of liver steatosis in non-diabetic offspring of long-lived siblings and age-matched controls by measuring liver enzymes in plasma and liver fat by computed tomography (CT). We measured nonfasting alanine transaminase (ALT), aspartate aminotransferase (AST), and Υ-glutamyl transferase (GGT) in 1625 subjects (736 men, mean age 59.1 years) from the Leiden Longevity Study, comprising offspring of long-lived siblings and partners thereof. In a random subgroup, fasting serum samples (n = 230) were evaluated and CT was performed (n = 268) for assessment of liver-spleen (L/S) ratio and the prevalence of moderate-to-severe non-alcoholic fatty liver disease (NAFLD). Linear mixed model analysis was performed adjusting for age, gender, body mass index, smoking, use of alcohol and hepatotoxic medication, and correlation of sibling relationship. Offspring of long-lived siblings had higher nonfasting ALT levels as compared to control subjects (24.3 mmol/L versus 23.2 mmol/L, p = 0.03), while AST and GGT levels were similar between the two groups. All fasting liver enzyme levels were similar between the two groups. CT L/S ratio and prevalence of moderate-to-severe NAFLD was similar between groups (1.12 vs 1.14, p = 0.25 and 8% versus 8%, p = 0.91, respectively). Except for nonfasting levels of ALT, which were slightly higher in the offspring of long-lived siblings compared to controls, no differences were found between groups in the extent of liver steatosis, as assessed with liver biochemical tests and CT. Thus, our data indicate that the extent of liver steatosis is similar between offspring of long-lived siblings and

Identification of AKB-48 and 5F-AKB-48 Metabolites in Authentic Human Urine Samples Using Human Liver Microsomes and Time of Flight Mass Spectrometry

OpenAIRE

Vikingsson, Svante; Josefsson, Martin; Green, Henrik

2015-01-01

The occurrence of structurally related synthetic cannabinoids makes the identification of unique markers of drug intake particularly challenging. The aim of this study was to identify unique and abundant metabolites of AKB-48 and 5F-AKB-48 for toxicological screening in urine. Investigations of authentic urine samples from forensic cases in combination with human liver microsome (HLM) experiments were used for identification of metabolites. HLM incubations of AKB-48 and 5F-AKB-48 along with 3...

Simultaneous quantification of the abundance of several cytochrome P450 and uridine 5'-diphospho-glucuronosyltransferase enzymes in human liver microsomes using multiplexed targeted proteomics.

Science.gov (United States)

Achour, Brahim; Russell, Matthew R; Barber, Jill; Rostami-Hodjegan, Amin

2014-04-01

Cytochrome P450 (P450) and uridine 5'-diphospho-glucuronosyltransferase (UGT) enzymes mediate a major proportion of phase I and phase II metabolism of xenobiotics. In vitro-in vivo extrapolation (IVIVE) of hepatic clearance in conjunction with physiologically-based pharmacokinetics (PBPK) has become common practice in drug development. However, prediction of xenobiotic kinetics in virtual populations requires knowledge of both enzyme abundances and the extent to which these correlate. A multiplexed quantification concatemer (QconCAT) strategy was used in this study to quantify the expression of several P450 and UGT enzymes simultaneously and to establish correlations between various enzyme abundances in 24 individual liver samples (ages 27-66, 14 male). Abundances were comparable to previously reported values, including CYP2C9 (40.0 ± 26.0 pmol mg(-1)), CYP2D6 (11.9 ± 13.2 pmol mg(-1)), CYP3A4 (68.1 ± 52.3 pmol mg(-1)), UGT1A1 (33.6 ± 34.0 pmol mg(-1)), and UGT2B7 (82.9 ± 36.1 pmol mg(-1)), expressed as mean ± S.D. Previous reports of correlations in expression of various P450 (CYP3A4/CYP3A5*1/*3, CYP2C8/CYP2C9, and CYP3A4/CYP2B6) were confirmed. New correlations were demonstrated between UGTs [including UGT1A6/UGT1A9 (r(s) = 0.82, P enzymes were shown to be correlated [including CYP1A2/UGT2B4 (r(s) = 0.67, P = 0.0002)]. The expression of CYP3A5 in individuals with *1/*3 genotype (n = 11) was higher than those with *3/*3 genotype (n = 10) (P history of smoking or alcohol use on enzyme expression was observed; however, expression of several enzymes declined with age. The correlation matrix produced for the first time by this study can be used to generate more realistic virtual populations with respect to abundance of various enzymes.

Metabolism of polybrominated diphenyl ethers and tetrabromobisphenol A by fish liver subcellular fractions in vitro.

Science.gov (United States)

Shen, Mengnan; Cheng, Jie; Wu, Ruohan; Zhang, Shenghu; Mao, Liang; Gao, Shixiang

2012-06-15

Polybrominated diphenyl ethers (PBDEs) and tetrabromobisphenol A (TBBPA) are two major flame retardants that accumulate in fish tissues and are potentially toxic. Their debrominated and oxidated derivatives were also reported in fish tissues although the sources of theses derivatives were unidentified. Our study was to determine whether PBDEs and TBBPA could be metabolized by fish liver subcellular fractions in vitro and to identify what types of metabolites were formed. Liver microsomes and S9 fractions of crucian carp (Carassius auratus) were exposed to 4,4'-dibromodiphenyl ether (BDE 15), 2,2',4,4'-tetrabromodiphenyl ether (BDE 47) or TBBPA solutions for 4h. Exposure of liver subcellular fractions to BDE 15 resulted in the formation of bromophenol and two monohydroxylated dibromodiphenyl ether metabolites. Neither in microsomes nor in S9 studies has revealed the presence of hydroxylated metabolites with BDE 47 exposure which indicated that the oxidation reactions in vitro were hindered by the increased number of bromine substituents on the PBDEs. TBBPA underwent an oxidative cleavage near the central carbon of the molecule, which led to the production of 2,6-dibromo-4-isopropyl-phenol and three unidentified metabolites. Another metabolite of TBBPA characterized as a hexa-brominated compound with three aromatic rings was also found in the liver subcellular fractions. These results suggest that the biotransformation of BDE 15 and TBBPA in fish liver is mediated by cytochrome P450 (CYP450) enzymes, as revealed by the formation of hydroxylated metabolites and oxidative bond cleavage products. Moreover, further studies on the identification of specific CYP450 isozymes involved in the biotransformation revealed that CYP1A was the major enzyme responsible for the biotransformation of BDE 15 and TBBPA in fish liver subcellular fractions and CYP3A4 also played a major role in metabolism of TBBPA. Copyright © 2012 Elsevier B.V. All rights reserved.

Strategy for Hepatotoxicity Prediction Induced by Drug Reactive Metabolites Using Human Liver Microsome and Online 2D-Nano-LC-MS Analysis.

Science.gov (United States)

Zhuo, Yue; Wu, Jian-Lin; Yan, Xiaojing; Guo, Ming-Quan; Liu, Ning; Zhou, Hua; Liu, Liang; Li, Na

2017-12-19

Hepatotoxicity is a leading cause of drug withdrawal from the market; thus, the assessment of potential drug induced liver injury (DILI) in preclinical trials is necessary. More and more research has shown that the covalent modification of drug reactive metabolites (RMs) for cellular proteins is a possible reason for DILI. Unfortunately, so far no appropriate method can be employed to evaluate this kind of DILI due to the low abundance of RM-protein adducts in complex biological samples. In this study, we proposed a mechanism-based strategy to solve this problem using human liver microsomes (HLMs) and online 2D nano-LC-MS analysis. First, RM modification patterns and potential modified AA residues are determined using HLM and model amino acids (AAs) by UHPLC-Q-TOF-MS. Then, a new online 2D-nano-LC-Q-TOF-MS method is established and applied to separate the digested modified microsomal peptides from high abundance peptides followed by identification of RM-modified proteins using Mascot, in which RM modification patterns on specific AA residues are added. Finally, the functions and relationship with hepatotoxicity of the RM-modified proteins are investigated using ingenuity pathway analysis (IPA) to predict the possible DILI. Using this strategy, 21 proteins were found to be modified by RMs of toosendanin, a hepatotoxic drug with complex structure, and some of them have been reported to be associated with hepatotoxicity. This strategy emphasizes the identification of drug RM-modified proteins in complex biological samples, and no pretreatment is required for the drugs. Consequently, it may serve as a valuable method to predict potential DILI, especially for complex compounds.

Characterization of the human cytochrome P450 enzymes involved in the metabolism of dihydrocodeine

Science.gov (United States)

Kirkwood, L. C.; Nation, R. L.; Somogyi, A. A.

1997-01-01

Aims Using human liver microsomes from donors of the CYP2D6 poor and extensive metabolizer genotypes, the role of individual cytochromes P-450 in the oxidative metabolism of dihydrocodeine was investigated. Methods The kinetics of formation of N- and O-demethylated metabolites, nordihydrocodeine and dihydromorphine, were determined using microsomes from six extensive and one poor metabolizer and the effects of chemical inhibitors selective for individual P-450 enzymes of the 1A, 2A, 2C, 2D, 2E and 3A families and of LKM1 (anti-CYP2D6) antibodies were studied. Results Nordihydrocodeine was the major metabolite in both poor and extensive metabolizers. Kinetic constants for N-demethylation derived from the single enzyme Michaelis-Menten model did not differ between the two groups. Troleandomycin and erythromycin selectively inhibited N-demethylation in both extensive and poor metabolizers. The CYP3A inducer, α-naphthoflavone, increased N-demethylation rates. The kinetics of formation of dihydromorphine in both groups were best described by a single enzyme Michaelis-Menten model although inhibition studies in extensive metabolizers suggested involvement of two enzymes with similar Km values. The kinetic constants for O-demethylation were significantly different in extensive and poor metabolizers. The extensive metabolizers had a mean intrinsic clearance to dihydromorphine more than ten times greater than the poor metabolizer. The CYP2D6 chemical inhibitors, quinidine and quinine, and LKM1 antibodies inhibited O-demethylation in extensive metabolizers; no effect was observed in microsomes from a poor metabolizer. Conclusions CYP2D6 is the major enzyme mediating O-demethylation of dihydrocodeine to dihydromorphine. In contrast, nordihydrocodeine formation is predominantly catalysed by CYP3A. PMID:9431830

Photoaffinity labeling of rat liver microsomal morphine UDP-glucuronosyltransferase by ( sup 3 H)flunitrazepam

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Thomassin, J.; Tephly, T.R. (Univ. of Iowa, Iowa City (USA))

1990-09-01

Benzodiazepines have been shown to competitively inhibit morphine glucuronidation in rat and human hepatic microsomes. Flunitrazepam exerted a potent competitive inhibition of rat hepatic morphine UDP-glucuronosyltransferase (UDPGT) activity (Ki = 130 microM). It has no effect on the activity of p-nitrophenol, 17 beta-hydroxysteroid, 3 alpha-hydroxysteroid, or 4-hydroxybiphenyl UDPGTs. Because flunitrazepam is an effective photoaffinity label for benzodiazepine receptors, studied were performed in solubilized rat hepatic microsomes and with partially purified preparations of morphine UDPGT to determine the enhancement of flunitrazepam inhibition and binding to morphine UDPGT promoted by exposure to UV light. Under UV light, flunitrazepam inhibition was markedly enhanced. UV light exposure also led to a marked increase in binding of (3H)flunitrazepam to microsomal protein, which was protected substantially by preincubation with morphine. Testosterone, androsterone, and UDP-glucuronic acid did not protect against UV-enhanced flunitrazepam binding, and morphine did not reverse flunitrazepam binding once binding had occurred. As morphine UDPGT was purified, a good correlation was found between the increases in specific activity of morphine UDPGT and flunitrazepam binding to protein. Chromatofocusing chromatography showed that flunitrazepam bound only to fractions containing active morphine UDPGT, and no binding to 4-hydroxybiphenyl UDPGT was observed. Fluorography of a sodium dodecyl sulfate-polyacrylamide electrophoresis gel of solubilized hepatic microsomes that had been treated with (3H) flunitrazepam under UV light revealed a band with a monomeric molecular weight between 54,000 and 58,000. This monomeric molecular weight compares favorably with the reported monomeric molecular weight of homogeneous morphine UDPGT (56,000).

Microsomal receptor for steroid hormones: functional implications for nuclear activity.

Science.gov (United States)

Muldoon, T G; Watson, G H; Evans, A C; Steinsapir, J

1988-01-01

Target tissues for steroid hormones are responsive by virtue of and to the extent of their content of functional intracellular receptors. Recent years have seen a shift in considerations of the cellular dynamics and distribution of these receptors, with current views favoring predominant intranuclear localization in the intact cell. This paper summarizes our analyses of the microsomal estrogen and androgen binding capability of rat uterine and ventral prostate tissue, respectively; these studies have revealed a set of high affinity sites that may act as a conduit for estrogen traversing the cell en route to the nucleus. These sites have many properties in common with cytosolic receptors, with the salient difference of a failure to activate to a more avid DNA-binding form under conditions which permit such activation of cytosolic receptors. The microsomal estrogen-binding proteins also have appreciable affinity for progesterone, another distinction from other known cellular estrogen receptor species. Various experimental approaches were employed to demonstrate that the microsomal receptors were not simply cytosol contaminants; the most convincing evidence is the recent successful separation of the cytosolic and microsomal forms by differential ammonium sulfate precipitation. Discrete subfractionation of subcellular components on successive sucrose gradients, with simultaneous assessments of binding capability and marker enzyme concentrations, indicates that the major portion of the binding is localized within the vesicles of the endoplasmic reticulum free of significant plasma membrane contamination. The microsomal receptors are readily solubilized by extraction with high- or low-salt-containing buffers or with steroid. The residual microsomes following such extraction have the characteristics of saturable acceptor sites for cytosolic estrogen-receptor complexes. The extent to which these sites will accept the cytosolic complexes is equal to the concentration of

MATERNAL HEMOLYSIS, ELEVATED LIVER-ENZYMES AND LOW PLATELETS SYNDROME - SPECIFIC PROBLEMS IN THE NEWBORN

NARCIS (Netherlands)

EELTINK, CM; VANLINGEN, RA; AARNOUDSE, JG; DERKS, JB; OKKEN, A

To evaluate the effects of maternal haemolysis, elevated liver enzymes and low platelets (HELLP) syndrome on the fetus and neonate we retrospectively investigated the outcome of 87 pregnancies. All women showed thrombocytopenia, elevated liver enzymes and haemolysis. None of them died. Nine infants

PHOTOMETRIC EVIDENCE FOR THE OSMOTIC BEHAVIOR OF RAT LIVER MICROSOMES

Science.gov (United States)

Tedeschi, Henry; James, Joseph M.; Anthony, William

1963-01-01

Electron microscope observations are consistent with the interpretation that the elements of the endoplasmic reticulum are osmotically active in situ as well as after isolation. More recently, it has been reported that microsomal suspensions equilibrate almost completely with added C14-sucrose and that no osmotic behavior is evident from photometric data. These findings were considered at variance with the electron microscope data. However, equilibration with added label simply attests to a relatively high permeability, and, in addition, the photometric data need not be critical. Osmotic volume changes, measured photometrically, may be masked by concomitant events (e.g., changes in the refractive index of the test solutions at varying osmotic pressures, breakdown of the particles, and agglutination). For these reasons the photometric experiments were repeated. In this work, the reciprocal of optical density of microsomal suspensions was found to vary linearly with the reciprocal of concentration of the medium at constant refractive index. These changes probably correspond to osmotic volume changes, since the effect was found to be (a) independent of substance used and (b) osmotically reversible. The transmission of the suspension was found to vary with the refractive index of the medium, the concentration of particles, and the wavelength of incident light, according to relationships that are similar to or identical with those obtained for mitochondrial suspensions. PMID:14064105

2'-Deoxyguanosine as a surrogate trapping agent for DNA reactive drug metabolites.

Science.gov (United States)

Häkkinen, Merja R; Laine, Jaana E; Juvonen, Risto O; Auriola, Seppo; Häyrinen, Jukka; Pasanen, Markku

2011-11-10

Drug metabolism can result in the production of highly reactive metabolites that may form adducts with cellular macromolecules, and thus initiate adverse drug reactions, cause toxicity, and even require the withdrawal of drug from the market. In this study, a 2'-deoxyguanosine (dG)-based chemical trapping test system was developed for use as a fast screening tool for DNA adducting metabolites of new drug candidates. Reactive metabolites were generated from parent compounds in in vitro incubations with phenobarbital-induced mouse liver microsomes, human liver microsomes and different recombinant human CYP enzymes in the presence of dG. The formed dG-adducts were separated, characterized and their stability was studied by liquid chromatography-tandem mass spectrometry (LC-MS/MS). The method was evaluated with six test compounds, aflatoxin B1, estrone, clozapine, tolcapone, ticlopidine and imipramine. Estrone and aflatoxin B1 formed dG adducts with phenobarbital-induced mouse liver microsomes, human liver microsomes and human recombinant CYP enzymes. Adduct formation was also observed with tolcapone when phenobarbital-induced mouse liver microsomes were used as the enzyme source. The stability of each formed adduct was independent of the different enzyme sources. No dG-adducts were identified with ticlopidine, clozapine and imipramine. Compared to other classical DNA reactivity tests, e.g. Ames test, the present surrogate endpoint, the dG adduct, is faster, enables the characterization of the formed compounds, and also permits the investigation of more unstable adducts. Copyright © 2011 Elsevier Ireland Ltd. All rights reserved.

Reactivation in vitro of zinc-requiring apo-enzymes by rat liver zinc-thionein

OpenAIRE

Udom, Albert O.; Brady, Frank O.

1980-01-01

The ability of rat liver zinc-thionein to donate its metal to the apo-enzymes of the zinc enzymes horse liver alcohol dehydrogenase, yeast aldolase, thermolysin, Escherichia coli alkaline phosphatase and bovine erythrocyte carbonic anhydrase was investigated. Zinc-thionein was as good as, or better than, ZnSO4, Zn(CH3CO2)2 or Zn(NO3)2 in donating its zinc to these apo-enzymes. Apo-(alcohol dehydrogenase) could not be reactivated by zinc salts or by zinc-thionein. Incubation of the other apo-e...

Coupled motions direct electrons along human microsomal P450 Chains.

Directory of Open Access Journals (Sweden)

Christopher R Pudney

2011-12-01

Full Text Available Protein domain motion is often implicated in biological electron transfer, but the general significance of motion is not clear. Motion has been implicated in the transfer of electrons from human cytochrome P450 reductase (CPR to all microsomal cytochrome P450s (CYPs. Our hypothesis is that tight coupling of motion with enzyme chemistry can signal "ready and waiting" states for electron transfer from CPR to downstream CYPs and support vectorial electron transfer across complex redox chains. We developed a novel approach to study the time-dependence of dynamical change during catalysis that reports on the changing conformational states of CPR. FRET was linked to stopped-flow studies of electron transfer in CPR that contains donor-acceptor fluorophores on the enzyme surface. Open and closed states of CPR were correlated with key steps in the catalytic cycle which demonstrated how redox chemistry and NADPH binding drive successive opening and closing of the enzyme. Specifically, we provide evidence that reduction of the flavin moieties in CPR induces CPR opening, whereas ligand binding induces CPR closing. A dynamic reaction cycle was created in which CPR optimizes internal electron transfer between flavin cofactors by adopting closed states and signals "ready and waiting" conformations to partner CYP enzymes by adopting more open states. This complex, temporal control of enzyme motion is used to catalyze directional electron transfer from NADPH→FAD→FMN→heme, thereby facilitating all microsomal P450-catalysed reactions. Motions critical to the broader biological functions of CPR are tightly coupled to enzyme chemistry in the human NADPH-CPR-CYP redox chain. That redox chemistry alone is sufficient to drive functionally necessary, large-scale conformational change is remarkable. Rather than relying on stochastic conformational sampling, our study highlights a need for tight coupling of motion to enzyme chemistry to give vectorial electron

Metabolism of styrene to styrene oxide and vinylphenols in cytochrome P450 2F2- and P450 2E1-knockout mouse liver and lung microsomes.

Science.gov (United States)

Shen, Shuijie; Li, Lei; Ding, Xinxin; Zheng, Jiang

2014-01-21

Pulmonary toxicity of styrene is initiated by cytochromes P450-dependent metabolic activation. P450 2E1 and P450 2F2 are considered to be two main cytochrome P450 enzymes responsible for styrene metabolism in mice. The objective of the current study was to determine the correlation between the formation of styrene metabolites (i.e., styrene oxide and 4-vinylphenol) and pulmonary toxicity of styrene, using Cyp2e1- and Cyp2f2-null mouse models. A dramatic decrease in the formation of styrene glycol and 4-vinylphenol was found in Cyp2f2-null mouse lung microsomes relative to that in the wild-type mouse lung microsomes; however, no significant difference in the production of the styrene metabolites was observed between lung microsomes obtained from Cyp2e1-null and the wild-type mice. The knockout and wild-type mice were treated with styrene (6.0 mmol/kg, ip), and cell counts and LDH activity in bronchoalveolar lavage fluids were monitored to evaluate the pulmonary toxicity induced by styrene. Cyp2e1-null mice displayed a susceptibility to lung toxicity of styrene similar to that of the wild-type animals; however, Cyp2f2-null mice were resistant to styrene-induced pulmonary toxicity. In conclusion, both P450 2E1 and P450 2F2 are responsible for the metabolic activation of styrene. The latter enzyme plays an important role in styrene-induced pulmonary toxicity. Both styrene oxide and 4-vinylphenol are suggested to participate in the development of lung injury induced by styrene.

Oxidative metabolism of BDE-47, BDE-99, and HBCDs by cat liver microsomes: Implications of cats as sentinel species to monitor human exposure to environmental pollutants.

Science.gov (United States)

Zheng, Xiaobo; Erratico, Claudio; Luo, Xiaojun; Mai, Bixian; Covaci, Adrian

2016-05-01

The in vitro oxidative metabolism of 2,2',4,4'-tetrabromodiphenyl ether (BDE-47), 2,2',4,4',5-pentabromodiphenyl ether (BDE-99), and individual α-, β- and γ-hexabromocyclododecane (HBCD) isomers catalyzed by cytochrome P450 (CYP) enzymes was screened using cat liver microsomes (CLMs). Six hydroxylated metabolites, namely 4-hydroxy-2,2',3,4'-tetrabromodiphenyl ether (4-OH-BDE-42), 3-hydroxy-2,2',4,4'-tetrabromodiphenyl ether (3-OH-BDE-47), 5-hydroxy-2,2',4,4'-tetrabromodiphenyl ether (5-OH-BDE-47), 6-hydroxy-2,2',4,4'-tetrabromodiphenyl ether (6-OH-BDE-47), 4'-hydroxy-2,2',4,5'- tetrabromodiphenyl ether (4'-OH-BDE-49), and 2'-hydroxy-2,3',4,4'-tetrabromodiphenyl ether (2'-OH-BDE-66), were identified and quantified after incubation of BDE-47. A di-OH-tetra-BDE was also found as metabolite of BDE-47 with CLMs. 5-OH-BDE-47 was the major metabolite formed. Five hydroxylated metabolites (3'-hydroxy-2,2',4,4',5-pentabromodiphenyl ether (3'-OH-BDE-99), 5'-hydroxy-2,2',4,4',5-pentabromodiphenyl ether (5'-OH-BDE-99), 6-hydroxy-2,2',4,4',5-pentabromodiphenyl ether (6-OH-BDE-99), 6'-hydroxy-2,2',4,4',5-pentabromodiphenyl ether (6'-OH-BDE-99), and 4'-hydroxy-2,2',4,5,5'-pentabromodiphenyl ether (4'-OH-BDE-101) were formed from BDE-99 incubated with CLMs. Concentrations of BDE-99 metabolites were lower than those of BDE-47. Four or more mono-hydroxylated HBCD (OH-HBCDs), four or more di-hydroxylated HBCD (di-OH-HBCDs), five or more mono-hydroxylated pentabromocyclododecanes (OH-PBCDs), and five or more di-hydroxylated pentabromocyclododecenes (di-OH-PBCDs) were detected after incubation of α-, β-, or γ-HBCD with CLMs. No diastereoisomeric or enantiomeric enzymatic isomerisation was observed incubating α-, β- or γ-HBCD with CLMs. Collectively, our data suggest that (i) BDE-47 is metabolized at a faster rate than BDE-99 by CLMs, (ii) OH-HBCDs are the major hydroxylated metabolites of α-, β- and γ-HBCD produced by CLMs, and (iii) the oxidative metabolism of BDE-47 and

Sex-dependent differences in phenobarbitane-induced oestradiol-2-hydroxylase activity in rat liver

Energy Technology Data Exchange (ETDEWEB)

Theron, C.N.; Neethling, A.C.; Taljaard, J.J.F. (Stellenbosch Univ. (South Africa). Dept. of Chemical Pathology)

1981-08-15

Oestradiol-2-hydroxylase (E/sub 2/-OH) activity was measured in liver and brain microsomes of 6-8-week-old Wistar rats. Phenobarbitone (75 mg/kg daily for 3 days) significantly increased enzyme activity in the liver of males and females, but there were striking differences between the two sexes. In males the enzyme activity was increased by 37% over control values and in females by 200%. The total microsomol cytochrome P-450 content was increased by 75% in males and by 82% in females. The apparent Michaelis constant (K(m)) of E/sub 2/-OH for 17..beta..-oestradiol in untreated males (9,8 ..mu..M) and females (9,2 ..mu..M) did not differ significantly. Phenobarbitone treatment, however, tended to reduce the apparent K(m) in males (8,2 ..mu..M) and to increase it in females (18,7 ..mu..M). E/sub 2/-OH activity was also detected in brain tissue of both sexes, but it was 50-200-fold lower than in the liver and was not increased by phenobarbitone.

Sex-dependent differences in phenobarbitane-induced oestradiol-2-hydroxylase activity in rat liver

International Nuclear Information System (INIS)

Theron, C.N.; Neethling, A.C.; Taljaard, J.J.F.

1981-01-01

Oestradiol-2-hydroxylase (E 2 -OH) activity was measured in liver and brain microsomes of 6-8-week-old Wistar rats. Phenobarbitone (75 mg/kg daily for 3 days) significantly increased enzyme activity in the liver of males and females, but there were striking differences between the two sexes. In males the enzyme activity was increased by 37% over control values and in females by 200%. The total microsomol cytochrome P-450 content was increased by 75% in males and by 82% in females. The apparent Michaelis constant (K(m)) of E 2 -OH for 17β-oestradiol in untreated males (9,8 μM) and females (9,2 μM) did not differ significantly. Phenobarbitone treatment, however, tended to reduce the apparent K(m) in males (8,2 μM) and to increase it in females (18,7 μM). E 2 -OH activity was also detected in brain tissue of both sexes, but it was 50-200-fold lower than in the liver and was not increased by phenobarbitone

Biosynthesis of intestinal microvillar proteins. Processing of aminopeptidase N by microsomal membranes

DEFF Research Database (Denmark)

Danielsen, E M; Norén, Ove; Sjöström, H

1983-01-01

-bound rather than a soluble form, indicating that synthesis of the enzyme takes place on ribosomes attached to the rough endoplasmic reticulum. The microsomal fractions process the Mr-115 000 polypeptide, which is the primary translation product of aminopeptidase N, to a polypeptide of Mr 140 000...

Increased serum levels of hyaluronic acid in pregnancies complicated by preeclampsia or hemolysis, elevated liver enzymes, and low platelets syndrome.

Science.gov (United States)

Osmers, R G; Schütz, E; Diedrich, F; Wehry, B; Krauss, T; Oellerich, M; Kuhn, W

1998-02-01

Fifteen percent of patients who later have hemolysis, elevated liver enzymes, and low platelets syndrome develop initially have nonspecific symptoms. Early diagnosis could ensure adequate obstetric management; however, prognostic biochemical tests are lacking. We hypothesized that elevated hyaluronic acid serum levels might be an early indicator of hemolysis, elevated liver enzymes, and low platelets syndrome because it is known to be a sensitive marker of liver cell function. Hyaluronic acid in serum was measured in patients with normal pregnancies (n = 109) and in those patients with pregnancies complicated by preeclampsia (n = 14) or hemolysis, elevated liver enzymes, and low platelets syndrome (n = 11). A significant increase in hyaluronic acid serum concentrations was observed in patients with hemolysis, elevated liver enzymes, and low platelets syndrome or with preeclampsia (p hyaluronic acid serum levels in hemolysis, elevated liver enzymes, and low platelets syndrome correlated with the clinical severity of the individual course of disease as measured by intensive care unit time (r = 0.72; p hyaluronic acid in preeclampsia and hemolysis, elevated liver enzymes, and low platelets syndrome are significantly elevated and might play an important diagnostic and prognostic role in patients with hemolysis, elevated liver enzymes, and low platelets syndrome.

Streptozotocin-induced diabetes mellitus affects lysosomal enzymes in rat liver

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G.B. Peres

2014-06-01

Full Text Available It has been previously shown that dextran sulfate administered to diabetic rats accumulates in the liver and kidney, and this could be due to a malfunction of the lysosomal digestive pathway. The aim of the present study was to evaluate the expression and activities of lysosomal enzymes that act upon proteins and sulfated polysaccharides in the livers of diabetic rats. Diabetes mellitus was induced by streptozotocin in 26 male Wistar rats (12 weeks old, while 26 age-matched controls received only vehicle. The livers were removed on either the 10th or the 30th day of the disease, weighed, and used to evaluate the activity, expression, and localization of lysosomal enzymes. A 50-60% decrease in the specific activities of cysteine proteases, especially cathepsin B, was observed in streptozotocin-induced diabetes mellitus. Expression (mRNA of cathepsins B and L was also decreased on the 10th, but not on the 30th day. Sulfatase decreased 30% on the 30th day, while glycosidases did not vary (or presented a transitory and slight decrease. There were no apparent changes in liver morphology, and immunohistochemistry revealed the presence of cathepsin B in hepatocyte granules. The decrease in sulfatase could be responsible for the dextran sulfate build-up in the diabetic liver, since the action of sulfatase precedes glycosidases in the digestive pathway of sulfated polysaccharides. Our findings suggest that the decreased activities of cathepsins resulted from decreased expression of their genes, and not from general lysosomal failure, because the levels of glycosidases were normal in the diabetic liver.

Streptozotocin-induced diabetes mellitus affects lysosomal enzymes in rat liver

Energy Technology Data Exchange (ETDEWEB)

Peres, G.B. [Universidade Federal de São Paulo, Escola Paulista de Medicina, Departamento de Bioquímica, São Paulo, SP, Brasil, Departamento de Bioquímica, Escola Paulista de Medicina, Universidade Federal de São Paulo, São Paulo, SP (Brazil); Juliano, M.A. [Universidade Federal de São Paulo, Escola Paulista de Medicina, Departamento de Biofísica, São Paulo, SP, Brasil, Departamento de Biofísica, Escola Paulista de Medicina, Universidade Federal de São Paulo, São Paulo, SP (Brazil); Aguiar, J.A.K.; Michelacci, Y.M. [Universidade Federal de São Paulo, Escola Paulista de Medicina, Departamento de Bioquímica, São Paulo, SP, Brasil, Departamento de Bioquímica, Escola Paulista de Medicina, Universidade Federal de São Paulo, São Paulo, SP (Brazil)

2014-05-09

It has been previously shown that dextran sulfate administered to diabetic rats accumulates in the liver and kidney, and this could be due to a malfunction of the lysosomal digestive pathway. The aim of the present study was to evaluate the expression and activities of lysosomal enzymes that act upon proteins and sulfated polysaccharides in the livers of diabetic rats. Diabetes mellitus was induced by streptozotocin in 26 male Wistar rats (12 weeks old), while 26 age-matched controls received only vehicle. The livers were removed on either the 10{sup th} or the 30{sup th} day of the disease, weighed, and used to evaluate the activity, expression, and localization of lysosomal enzymes. A 50-60% decrease in the specific activities of cysteine proteases, especially cathepsin B, was observed in streptozotocin-induced diabetes mellitus. Expression (mRNA) of cathepsins B and L was also decreased on the 10{sup th}, but not on the 30{sup th} day. Sulfatase decreased 30% on the 30{sup th} day, while glycosidases did not vary (or presented a transitory and slight decrease). There were no apparent changes in liver morphology, and immunohistochemistry revealed the presence of cathepsin B in hepatocyte granules. The decrease in sulfatase could be responsible for the dextran sulfate build-up in the diabetic liver, since the action of sulfatase precedes glycosidases in the digestive pathway of sulfated polysaccharides. Our findings suggest that the decreased activities of cathepsins resulted from decreased expression of their genes, and not from general lysosomal failure, because the levels of glycosidases were normal in the diabetic liver.

Streptozotocin-induced diabetes mellitus affects lysosomal enzymes in rat liver

International Nuclear Information System (INIS)

Peres, G.B.; Juliano, M.A.; Aguiar, J.A.K.; Michelacci, Y.M.

2014-01-01

It has been previously shown that dextran sulfate administered to diabetic rats accumulates in the liver and kidney, and this could be due to a malfunction of the lysosomal digestive pathway. The aim of the present study was to evaluate the expression and activities of lysosomal enzymes that act upon proteins and sulfated polysaccharides in the livers of diabetic rats. Diabetes mellitus was induced by streptozotocin in 26 male Wistar rats (12 weeks old), while 26 age-matched controls received only vehicle. The livers were removed on either the 10 th or the 30 th day of the disease, weighed, and used to evaluate the activity, expression, and localization of lysosomal enzymes. A 50-60% decrease in the specific activities of cysteine proteases, especially cathepsin B, was observed in streptozotocin-induced diabetes mellitus. Expression (mRNA) of cathepsins B and L was also decreased on the 10 th , but not on the 30 th day. Sulfatase decreased 30% on the 30 th day, while glycosidases did not vary (or presented a transitory and slight decrease). There were no apparent changes in liver morphology, and immunohistochemistry revealed the presence of cathepsin B in hepatocyte granules. The decrease in sulfatase could be responsible for the dextran sulfate build-up in the diabetic liver, since the action of sulfatase precedes glycosidases in the digestive pathway of sulfated polysaccharides. Our findings suggest that the decreased activities of cathepsins resulted from decreased expression of their genes, and not from general lysosomal failure, because the levels of glycosidases were normal in the diabetic liver

Liver enzymes and markers of inflammation in Nigerian adults with metabolic syndrome

Directory of Open Access Journals (Sweden)

Udenze Ifeoma Christiana

2015-01-01

Full Text Available Aims and objectives: The aim of this study is to determine the plasma levels of the liver enzymes alanine aminotransferase (ALT, aspartate aminotransferase (AST, alkaline phosphatase (ALP, gamma-glutamyl transferase (GGT, and lactate dehydrogenase (LDH in people with metabolic syndrome and to determine the association between the liver enzymes and obesity, insulin resistance, interleukin 6 (IL-6, and C-reactive protein (CRP in adult Nigerians with metabolic syndrome. Materials and Methods: This was a case control study of 50 adult men and women with metabolic syndrome, and 50 age- and sex-matched males and females without metabolic syndrome. Metabolic syndrome was defined based on the National Cholesterol Education Program (NCEP-Adult Treatment Panel III (ATPIII criteria. Written informed consent was obtained from the participants. Sociodemographic and clinical data were collected using a structured questionnaire. Venous blood was collected after an overnight fast. The ethics committee of the Lagos University Teaching Hospital in Lagos, Nigeria, approved the study protocol. Comparison of continuous variables was done using the student′s t-test. Regression and correlation analysis were used to determine the associations between variables. Statistical significance was set at P < 0.05. Results: There was a statistically significant increase in the liver enzymes ALP (P = 0.031, ALT (P = 0.019, and GGT (P = 0.037, as well as in the inflammatory markers CRP (P = 0.019 and the cytokine IL-6 (P = 0.040 between the two study groups. ALP and ALT showed significant correlation with waist circumference, BMI, fasting insulin, and waist/hip ratio (P < 0.05. Multivariate regression also identified ALT, AST, and ALP to be associated with IL-6 and CRP (P < 0.05. Conclusion: Liver enzyme levels were increased in metabolic syndrome and associated with obesity, fasting insulin, and CRP. Elevated liver enzymes may indicate dysmetabolism and increased

Mutagenicity of anthraquinone and hydroxylated anthraquinones in the Ames/Salmonella microsome system.

Science.gov (United States)

Liberman, D F; Fink, R C; Schaefer, F L; Mulcahy, R J; Stark, A A

1982-01-01

The mutagenicity of anthracene, anthraquinone, and four structurally similar compounds of each was evaluated in the Ames/Salmonella microsome assay. Anthraquinone was shown to be mutagenic for strains TA1537, TA1538, and TA98 in the absence of rat liver homogenate. The four anthraquinone derivatives tested were mutagenic for TA1537 exclusively. None of the anthracenes exhibited mutagenic activity. PMID:7103489

Mutagenicity of anthraquinone and hydroxylated anthraquinones in the Ames/Salmonella microsome system.

OpenAIRE

Liberman, D F; Fink, R C; Schaefer, F L; Mulcahy, R J; Stark, A A

1982-01-01

The mutagenicity of anthracene, anthraquinone, and four structurally similar compounds of each was evaluated in the Ames/Salmonella microsome assay. Anthraquinone was shown to be mutagenic for strains TA1537, TA1538, and TA98 in the absence of rat liver homogenate. The four anthraquinone derivatives tested were mutagenic for TA1537 exclusively. None of the anthracenes exhibited mutagenic activity.

Superoxide Dismutase (SOD Enzyme Activity Assay in Fasciola spp. Para-sites and Liver Tissue Extract

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M Assady

2011-09-01

Full Text Available Background: The purpose of this comparative study was to detect superoxide dismutase (SOD activities in Fasciola hepatica, F. gigantica parasites, infected and healthy liver tissues in order to determine of species effects and liver infection on SODs activity level.Methods: Fasciola spp. parasites and sheep liver tissues (healthy and infected liver tissues, 10 samples for each, were collected, homogenized and investigated for protein measurement, protein detection and SOD enzyme activity assay. Protein concentration was measured by Bradford method and SODs band protein was detected on SDS-PAGE. SODs activity was determined by iodonitrotetrazolium chloride, INT, and xanthine substrates. Independent samples t-test was conducted for analysis of SODs activities difference.Results: Protein concentration means were detected for F. hepatica 1.3 mg/ ml, F. gigantica 2.9 mg/ml, healthy liver tissue 5.5 mg/ml and infected liver tissue 1.6 mg/ml (with similar weight sample mass. Specific enzyme activities in the samples were obtained 0.58, 0.57, 0.51, 1.43 U/mg for F. hepatica, F. gigantica, healthy liver and infected liver respectively. Gel electrophoresis of Fasciola spp. and sheep liver tissue extracts revealed a band protein with MW of 60 kDa. The statistical analysis revealed significant difference between SOD activities of Fasciola species and also between SOD activity of liver tissues (P<.05.Conclusion: Fasciola species and liver infection are effective causes on SOD enzyme activity level.

Lipid peroxidation in microsomes of murine bone marrow after low-dose γ-irradiation

International Nuclear Information System (INIS)

Schwenke, K.; Coslar, S.; Muehlensiepen, H.; Altman, K.I.; Feinendegen, L.E.

1994-01-01

The principal aim of the study was to investigate the effect of low-dose γ-irradiation on lipid peroxidation (LPO) in murine bone marrow. To this end, the degree of LPO in suspensions of microsomes of murine bone marrow cells (BMC) was determined in terms of malondialdehyde (MDA) formation after whole-body or in vitro exposure to various doses of γ-radiation. These effects were compared to some extent with similar effects in liver and spleen preparations. As to the effect of γ-irradiation on LPO in BMC, the response depends on the dose level and on whether whole-body or in vitro exposures are involved. Whole-body irradiation did not result in an increase in LPO in BMC microsomes, even at such high doses as 15 Gy, although hepatic microsomes showed a marked increase. In contrast, in vitro irradiation of BMC microsomes with 0.1, 10 and 50 Gy brought about an increase in LPO. This increase was already significant (P < 0.05) at 0.1 Gy following a post-irradiation incubation and substantial at 50 Gy, even without subsequent incubation. The results show that low doses of γ-irradiation are able to induce an elevation of LPO in murine BMC microsomes, but only after in vitro irradiation. In the case of whole-body irradiation cellular radical scavengers and other metabolic reactions may prevent a measurable increase in LPO. This is partly illustrated by the case of vitamin-E deficiency, where a substantial increase in LPO in BMC microsomes is observed even without γ-irradiation in comparison with euvitaminotic mice because normally occurring radicals are not scavenged sufficiently. (orig.)

In vitro inactivation of hepatic microsomal phospholipase A2 by the marine natural product manoalide

International Nuclear Information System (INIS)

Master, M.M.; Jacobs, R.S.

1986-01-01

The effects of manoalide (MLD) and several analogs (isolated from the sponge Luffariella variabilis) on mouse hepatic microsomal phospholipase A 2 (PLA 2 ) activity was investigated. Microsomal PLA 2 , a membrane bound, Ca ++ dependent enzyme with an alkaline pH optimum, functions in intracellular phospholipid turnover. In vitro PLA 2 activity was assayed by preincubating MLD or analogs (2.5-100μM) with microsomes for 60 min. at 37 0 C, combining this mixture with 14 C-phosphatidylcholine and CaCl 2 , and incubating at 37 0 C for 40 minutes. Enzyme activity was quantitated by measurement of the extracted 14 C-arachidonic acid product. MLD inhibited PLA 2 in a dose-dependent manner, with an IC 50 = 94μM. Lineweaver-Burk analysis suggests that MLD inhibits PLA 2 noncompetitively. One of the analogs, producing a comparable dose-response curve to MLD, was found to be more potent (IC 50 = 33μM). Another analog facilitated PLA 2 activity (15%) at 25μM, followed by inactivation at higher doses (IC 50 > 100 μM). Facilitation of PLA 2 activity was seen with concentrations as low as 2.5μM of a third analog, and significant inactivation of PLA 2 was evident. These results indicate that MLD is not as potent against microsomal PLA 2 as has been shown with purified bee venom and cobra venom PLA 2 's

Effect of treatment with cadmium on kinetic properties of Na+, K+-ATPase and glucose-6-phosphatase activity in rat liver microsomes

International Nuclear Information System (INIS)

Modi, Hiren R.; Patil, Nisha; Katyare, Surendra S.

2008-01-01

Studies on Cd hepatotoxicity have focused mainly on induction of cytochrome P 450 system and related enzymes. In the present study young adult male rats given a single intra-peritoneal injection of Cd (0.84 mg Cd/kg body weight) and effects on kinetic parameters rat liver microsomal Na + , K + -ATPase and G6Pase were evaluated at the end of 1 month and 1 week. The substrate and temperature kinetics parameters were examined and attempts were made to seek correlation with changes in lipid/phospholipid profiles. The Na + , K + ATPase activity decreased only in 1 week Cd-treated group but recovered at the end of 1 month. The activity resolved in two distinct kinetic components in control as well as the experimental groups. In 1 week Cd-treated group the K m value of both the components was unchanged, whereas V max value decreased. In 1-month Cd-treated group V max value only of component I increased. The catalytic efficiency of both the components was not affected in the experimental groups. In 1-week Cd-treated group the energy of activations at high-temperature range (E H ) and low-temperature range (E L ) decreased, whereas for 1-month Cd-treated group the energies of activations did not change. The G6Pase activity measured at 37 deg. C was high only in 1-month Cd-treated group. The activity resolved in two kinetically distinguishable components in control as well as in the experimental groups. K m value of component I decreased in both the Cd-treated groups. In 1-month Cd-treated group the V max value of component II increased. The catalytic efficiency of G6Pase was unchanged despite changes in K m and V max . In 1-week Cd-treated group the E H and E L decreased, whereas only E L showed decrease in 1-month Cd-treated group. Cholesterol (CHL) content increased in both the Cd-treated groups. Content of lysophospholipid (Lyso), spinghomyelin (SPM) and phosphatidic acid (PA) increased, whereas phosphatidylcholine (PC) and phosphatidylserine (PS) decreased in 1-week Cd

Microsomal protein synthesis inhibition: an early manifestation of gentamicin nephrotoxicity

International Nuclear Information System (INIS)

Bennett, W.M.; Mela-Riker, L.M.; Houghton, D.C.; Gilbert, D.N.; Buss, W.C.

1988-01-01

Aminoglycoside antibiotics achieve bacterial killing by binding to bacterial ribosomes and inhibiting protein synthesis. To examine whether similar mechanisms could be present in renal tubular cells prior to the onset of overt proximal tubular necrosis due to these drugs, we isolated microsomes from Fischer rats given 20 mg/kg gentamicin every 12 h subcutaneously for 2 days and from vehicle-injected controls. Concomitant studies of renal structure, function, and mitochondrial respiration were carried out. [3H]leucine incorporation into renal microsomes of treated animals was reduced by 21.9% (P less than 0.01), whereas brain and liver microsomes from the same animals were unaffected. Gentamicin concentration in the renal microsomal preparation was 56 micrograms/ml, a value 7- to 10-fold above concentrations necessary to inhibit bacterial growth. Conventional renal function studies were normal (blood urea, serum creatinine, creatinine clearance). Treated animals showed only a mild reduction of inulin clearance, 0.71 compared with 0.93 ml.min-1.100 g-1 in controls (P less than 0.05), and an increase in urinary excretion of N-acetylglucosaminidase of 20 compared with 14.8 units/l (P less than 0.05). Renal slice transport of p-aminohippuric acid, tetraethylammonium, and the fractional excretion of sodium were well preserved. There was no evidence, as seen by light microscopy, of proximal tubular necrosis. Mitochondrial cytochrome concentrations were normal and respiratory activities only slightly reduced. Processes similar to those responsible for bacterial killing could be involved in experimental gentamicin nephrotoxicity before overt cellular necrosis

Angiotensin converting enzyme (ACE) gene expression in experimentally induced liver cirrhosis in rats.

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Shahid, Syed Muhammad; Fatima, Syeda Nuzhat; Mahboob, Tabassum

2013-09-01

Angiotensin converting enzyme (ACE) is a key player of Renin Angiotensin System (RAS), involved in conversion of active product, angiotensin-II. Alterations in RAS have been implicated in the pathophysiology of various diseases involving heart, kidney, lung and liver. This study is designed to investigate the association of ACE gene expression in induction of liver cirrhosis in rats. Total 12 male albino Wistar rats were selected and divided in two groups. Control group received 0.9% NaCl, where as Test group received thioacidamide (TAA), dissolved in 0.9%NaCl, injected intraperitoneally at a dosage of 200mg/Kg of body weight, twice a week for 12 weeks. The rats were decapitated and blood sample was collected at the end of experimental period and used for liver functions, enzyme activity, antioxidant enzymes and lipid peroxidation estimations. Genomic DNA was isolated from excised tissue determine the ACE genotypes using specific primers. The ACE gene expression in liver tissue was assessed using the quantitative RT-PCR method. The activity of ALT, total and direct bilirubin, SOD and CAT levels were significantly high (pACE gene expression after 12 weeks TAA treatment in cirrhotic rats was significantly increased (pACE gene expression. The finding of major up-regulation of ACE in the experimental rat liver provides further insight into the complexities of the RAS and its regulation in liver injury. The development of specific modulators of ACE activity and function, in future, will help determine the role of ACE and its genetic variants in the pathophysiology of liver disease.

Effects of tin-protoporphyrin administration on hepatic xenobiotic metabolizing enzymes in the juvenile rat

International Nuclear Information System (INIS)

Stout, D.L.; Becker, F.F.

1988-01-01

The heme analogue tin-protoporphyrin IX (SnP) is a potent inhibitor of microsomal heme oxygenase. Administration of SnP to neonatal rats can prevent hyperbilirubinemia by blocking the postnatal increase of heme oxygenase activity. Apparently innocuous at therapeutic doses, it is of potential clinical value for chemoprevention of neonatal jaundice. We found that when 50-g male Sprague-Dawley rats were treated daily with 50 mumol of SnP/kg sc for 6 days, hepatic microsomal cytochromes b5 and P-450 were significantly diminished. Cytochrome P-450 reductase, two P-450-dependent monooxygenases, aminopyrine demethylase and benzo(a)pyrene hydroxylase, and catalase, a peroxisomal hemoprotein, were also significantly diminished. These results suggested that SnP might significantly affect the metabolism of other xenobiotics. This possibility was confirmed by the finding that hexobarbital-induced sleep lasted 4 times longer in SnP-treated rats than in controls. Inhibition of protein synthesis by SnP was ruled out as the cause of hemoprotein loss when administration of [ 3 H]leucine to SnP-treated and control rats demonstrated that proteins of the microsomal, cytosolic, and plasma membrane fractions of the livers from both groups incorporated similar levels of leucine. When 55 FeCl 3 and [2- 14 C]glycine were administered to measure heme synthesis, heme extract from the livers of SnP-treated rats contained 4 times more label from iron and glycine than did heme from control livers. Despite the apparent increased rate of heme synthesis in SnP-treated rats, each of the three cell fractions demonstrated a significant loss of heme but contained sizable amounts of SnP. These findings suggest that SnP causes a decrease of functional hemoprotein and partial loss of enzymic activity by displacing intracellular heme

The effect of aqueous extract of Avicennia marina (Forsk. Vierh. leaves on liver enzymes' activity, oxidative stress parameters and liver histopathology in male diabetic rats

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Akram Hamzevi

2017-12-01

Full Text Available Background: Avicennia marina has antioxidant and anti-diabetic properties. This study was conducted to examine the effect of aqueous extract of A. marina on liver enzymes' activity, oxidative stress parameters and liver histopathology in diabetic rats. Materials and Methods: In this experimental study, 28 male rats were allocated into the equal groups of control, diabetic control and experimental diabetic 1 and 2. The diabetes in diabetic control and experimental diabetic groups was induced using an intraperitoneal injection of 120 mg/kg alloxan. The experimental diabetic groups received the aqueous extract of A. marina (100 and 200 mg/kg, i.p. in alternate days for one month. Sterile distilled water was injected to the animals of control and diabetic control groups. At the end of the treatment period, serum levels of ALT, AST, GGT and ALP were measured. Then, levels of SOD, GST, CAT and MDA were measured in the liver tissue. The liver sections were prepared and examined by an optical microscope. Results: Results showed that administration of the A. marina extract (100 and 300 mg/kg, ip to the diabetic rats significantly decreased the serum levels of liver enzymes and tissue level of MDA. Also, the activity of the liver tissue's antioxidant enzymes was increased (P<0.05. The A. marina extract dose-dependently decreased liver damages in diabetic rats. Conclusion: Administration of the A. marina extract improves liver tissue oxidative stress indices and decreases the serum level of liver enzymes. Also, A. marina extract improves liver tissue injuries induced by diabetes.

Isolation and structural elucidation of tiamulin metabolites formed in liver microsomes of pigs.

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Lykkeberg, Anne Kruse; Cornett, Claus; Halling-Sørensen, Bent; Hansen, Steen Honoré

2006-09-18

Although the antimicrobial tiamulin is extensively metabolized in pigs, the metabolism is not well investigated. In this work the NADPH dependent metabolism of tiamulin in liver microsomes from pigs has been studied. The tiamulin metabolites formed in the incubations were analysed using LC-MS, and three major metabolites were isolated using solid phase extraction and preparative HPLC. The final structure elucidations were performed by tandem mass spectrometry and (1)H and (13)C NMR. The structures of the metabolites were found to be 2beta-hydroxy-tiamulin, 8alpha-hydroxy-tiamulin and N-deethyl-tiamulin. In addition, the LC-MS chromatograms revealed two other minor metabolites. From their chromatography and from MS(2) analysis the structures were estimated to be 2beta-hydroxy-N-deethyl-tiamulin and 8alpha-hydroxy-N-deethyl-tiamulin, but the structures were not confirmed by NMR. In these studies approximately 20% of tiamulin was deethylated, 10% was hydroxylated in the 2beta-position and 7% was hydroxylated in the 8alpha-position. About 40% of tiamulin was metabolized during the incubation conditions used. The protein precipitation in the incubations was performed using perchloric acid, and the preparative purification was performed under alkaline conditions. Therefore, the stability of the metabolites under these conditions was studied. The metabolites were found to be stable in the acid solution, but under alkaline conditions, particularly at room temperature, the stability of especially 8alpha-hydroxy-tiamulin was considerably reduced (40% loss after 1 week).

Effects of dibutyl phthalate on lipid metabolism and drug metabolising enzyme system in rats

International Nuclear Information System (INIS)

Arakaki, Mitsuo; Ariyoshi, Toshihiko.

1976-01-01

Effects of dibutyl phthalate (DBP) on the liver constituents and the drug metabolizing enzyme system were investigated in rats. 1. In the experiments at a single oral dose of DBP (630 or 1260 mg/kg), the glycogen content was decreased only at the high dose, but no effects were observed on the contents of glycogen, triglyceride, microsomal protein and cytochromes, and on the activities of drug metabolizing enzymes. 2. In the repeated oral dose of DBP (630 or 1260 mg/kg/day) for 5 days, the ratio of liver weight to body weight was increased in both female and male rats, whereas the increases of cytochrome P-450 content and aniline hydroxylase activity were noted only in male rats. However, the contents of liver triglyceride, phospholipids, and cholesterol were unchanged. On the other hand, serum cholesterol content which showed the tendency to be decreased at the low dose was significantly decreased at the high dose. 3. In the incorporation of 1- 14 C-acetate into liver and serum lipids after repeated oral dose of DBP (630 mg/kg/day) for 5 days in male rats, the incorporation into triglyceride showed tendency to be increased, whereas the incorporation into cholesterol and cholesterol ester remained unchanged in vivo and in vitro. (auth.)

[The effect of alpha-tocopherol and ionol on the physical structure of the membranes of rat liver microsomes under conditions of antioxidant insufficiency].

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Gubskiĭ, Iu I; Boldeskul, A E; Primak, R G; Zadorina, O V

1989-01-01

Physiochemical conformity of the alpha-tocopherol interaction with hepatic microsomal membranes has been studied by means of fluorescent probes (pyrene and 1-anilinonaphthalene-8-sulphonate). The microsomal membrane microviscosity is shown to sharply decrease under conditions of the antioxidant deficiency with vitamin E expelled into animals normalizes microviscosity, but feebly influences the microsomal surface charge. Microcalorimetry has been used to establish that penetration of tocopherol into microsomal membranes was accompanied by the exothermic effect.

Associated liver enzymes with hyperlipidemic profile in type 2 diabetes patients.

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Al-Jameil, Noura; Khan, Farah A; Arjumand, Sadia; Khan, Mohammad F; Tabassum, Hajera

2014-01-01

Type 2 diabetes mellitus (T2DM) is characterized by hyperglycemia and is associated with dyslipidemia and disturbed liver function. Aim of the present work is to assess the liver enzymes and to find its association with hyperlipidemic profile in T2DM. Total of 157 subjects were studied and divided into two groups; diabetes (n=81) and non-diabetes (n=76). Various biochemical parameters like fasting glucose, post prandial glucose, HbA1c, total cholesterol (TC), triglycerides (Tg), high density lipoprotein cholesterol (HDL-C), alanine amino transferase (ALT), aspartate amino transferase (AST) and gamma-glutamyl transferase (GGT) were analyzed by ROCHE module Cobas 6000 (C501 & C601) analyzer, kits were procured by ROCHE diagnostics. Low density lipoprotein cholesterol (LDL-C) was estimated by Freidwald's formula. Statistical analysis was performed by applying student t test and Pearson's correlation coefficient, at 0.0001 and 0.05 level of significance, respectively. All the glycemic control parameters, lipid profile parameters except HDL-C and liver enzymes were found increased in diabetes group and significantly differ from non-diabetes group (p>0.0001). ALT showed significant positive correlation with fasting glucose, post prandial glucose, HbA1c, TC, Tg, LDL-C and GGT at p>0.05. AST showed very weak relation with all parameters while GGT was positively associated with fasting glucose, post prandial glucose, HbA1c, TC, Tg, LDL-C and ALT at p>0.05. In conclusion, T2DM incline to elevate liver enzymes, especially ALT and GGT were of significance. Routine screening of ALT and GGT in T2DM patients may assists early detection of liver abnormalities and to arrest the progress of disease.

Health behavior associated with liver enzymes among obese Korean adolescents, 2009-2014.

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Lee, Eun-Young; Choi, Hwa Young; Cho, Hyunsoon; Kim, Bo Hyun; Ki, Moran

2018-01-01

Obesity is major risk factor for liver health. This study aimed to clarify whether specific health behaviors were associated with liver function in obese adolescents in Korea. Based on national school health examination data from 2009 to 2014, 25,142 obese or overweight students were examined for aspartate transaminase and alanine transaminase levels, and health behaviors. Multiple logistic regression was used to calculate the odds ratio for liver enzyme elevation. Subjects who thought of themselves as "very fat" had a 1.6 times higher odds ratio for liver enzyme elevations than those who thought of themselves as "normal." Those who consumed fast food 3 to 5 times weekly had 1.3 times higher odds ratio (OR = 1.27, 95% confidence interval = 1.05-1.54) for the elevation of ALT than those who did not consume fast food. Those who took sugar-sweetened beverage 3 to 5 times weekly had 1.2 times higher odds ratio (OR = 1.24, 95% confidence interval = 1.07-1.42) for the elevation of ALT than those who did not take it. Those who played computer game more than 2 hours a day showed 1.1 times higher odds ratio (OR = 1.10, 95% confidence interval = 1.01-1.21) for the elevation of ALT than those who did not. Specific food item and its frequency of consumption were identified for the positive and negative association with the elevation of liver enzymes. Self-image of body shape, sleeping time and need of help for alcohol or smoking problems also showed substantial association with the elevation.

Comparing Effects of Medication Therapy and Exercise Training with Diet on Liver enzyme Levels and Liver Sonography in Patients with Non-Alcoholic Fatty Liver Disease (NAFLD

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Azadeh Nabizadeh Haghighi

2016-03-01

Full Text Available Background & Objectives: Non-alcoholic fatty liver disease, characterized by the deposition of fat in liver cells, can cause fibrosis, cirrhosis, and liver cell damage if not controlled. The aim of this study is to compare the effects of medication therapy and exercise training with diet on liver enzyme levels and liver sonography in patients with non-alcoholic fatty liver disease (NAFLD. Materials & Methods :In this quasi-experimental study, female patients with non-alcoholic fatty liver were randomly divided into two groups: medication therapy (n = 10 and exercise therapy (n = 10 for 8 weeks. During this period, the exercise group performed exercise training three days a week for 90 minutes per session. The drug was given to the medication group. In both groups, the diet was 500 calories less than their daily energy. Before and after intervention, blood tests and liver sonography were executed. All statistical analyses were done using SPSS for Windows version 20. Comparisons between and within groups were performed by Student's t-test and Wilcoxon test on paired and unpaired data. P < 0.05 was considered statistically significant. Results :In both groups, liver enzyme levels and disease severity in sonography reduced significantly (p<0.05. Conclusion: The findings of the present research showed that both methods of therapy have the same effect on reducing the severity of NAFLD.

Effects of phenobarbital pretreatment on the in vivo metabolism of carbaryl in rats

International Nuclear Information System (INIS)

Knight, E.V.; Alvares, A.P.; Chin, B.H.

1987-01-01

Phenobarbital (PB) pretreatment of animals is known to induce the activity of drug-metabolizing enzymes in liver microsomes. Previous studies showed that incubation of carbaryl with microsomes obtained from livers of untreated or PB-treated rats resulted in little or no oxidative metabolism of the substrate. In addition, no spectral interactions were observed when carbaryl was added to hepatic microsomal suspensions. The present study was carried out to determine the effect of PB pretreatment on the in vivo metabolism of carbaryl in rats

Induction of phosphatidylcholine biosynthesis via CDPcholine pathway in lung and liver of rats following intratracheal administration of DDT and endosulfan

International Nuclear Information System (INIS)

Narayan, S.; Dani, H.M.; Misra, U.K.

1989-01-01

The induction of phosphatidylcholine (PC) biosynthesis via the CDPcholine pathway in lung and liver of rats has been shown following the intratracheal administration of 1,1,1-trichloro-2m2-bis(p-chlorophenyl) ethane (DDT) (5 mg/100 g body weight) and endosulfan (1 mg/100 g body weight) for 3 days. Controls received only the vehicle solution (groundnut oil, 0.1 m1/100 g body weight). The treatment of DDT and endosulfan significantly increased the PC contents and the incorporation of radioactive [methyl-3H]choline into PC of lung and liver microsomes. The incorporation of radioactive [methyl-14C]methionine into microsomal PC of lung and liver was not affected significantly by treatment with either of the insecticides. 1,4,5,6,7-hexachloro-5-norbornene-2,3-dimethano cyclic sulfite (endosulfan) administration significantly increased the activity of choline kinase and phosphocholine cytidylyltransferase (both cytosolic and microsomal) of lung, whereas DDT increased the activity of only latter. In liver, both DDT and endosulfan administration significantly increased the activity of choline kinase and phosphocholine cytidylyltransferase (both cytosolic and microsomal). However, the activity of phosphocholinetransferase was not affected in both lung and liver microsomes of rats treated with these insecticides. The PC precursor pool sizes, choline and phosphorylcholine, of lung and liver tissues were not altered by DDT and endosulfan treatments. The present results suggest that the increased level of PC and incorporation of radioactive [methyl-3H]choline into microsomal PC could be the result of increased activity of choline kinase and phosphocholine cytidylyltransferase of lung and liver of rats following intratracheal administration of DDT and endosulfan

[Detection and the production mechanism of antinuclear antibodies (ANA) and anti-liver/kidney microsomal tpe 1 antibodies (anti-LKM1) in patients with chronic hepatitis C].

Science.gov (United States)

Bai, Li; Lu, Hai-Ying; Feng, Zhen-Ru; Yu, Min; Li, Wen-Gang; Gong, Wei-Bo; Zhao, Nu-en-ji-ya; Xu, Xiao-Yuan

2009-08-01

To investigate the prevalence of antinuclear antibodies (ANA) and anti-liver/ kidney microsomal type 1 antibodies (anti-LKM1) in patients with chronic hepatitis C (CHC)and to explore the mechanism of production of these autoantibodies. Serum samples were collected from 360 patients with CHC (case group), 69 patients with chronic hepatitis B (CHB) and 69 patients with autoimmune hepatitis (AIH) (control group). Serum ANA and anti-LKM1 were detected by indirect immunofluorescence (HF) technique and enzyme-linked immunosorbent assay (ELISA), respectively. Multi-factor analysis was performed to explore the correlations of the production of autoantibodies with some factors such as age, sex, viral loads, HCV genotype, biochemical parameters and clinical characteristics. Fifty-four (15%) of 360 patients infected with HCV were positive in autoantibodies. The prevalence of ANA and anti-LKM1 were 12.5% (45/360) and 2.5% (9/ 360), respectively. The positive rate of autoantibodies in patients with CHC was significantly higher than that in patients with CHB (15% vs 2.9%, P = 0.006), but significantly lower than that in patients with AIH (15% vs 47.9%, P 0.05). Autoantibodies related to AIH can be detected in CHC patients; interferon may not induce the production of autoantibodies; it is very likely that HCV infection induces the autoimmune reaction and the production of autoantibodies.

Liver enzymes and psychological well-being response to aerobic ...

African Journals Online (AJOL)

2014-06-02

Jun 2, 2014 ... Background: Chronic hepatitis C (CHC) is a medical condition that has broad implications for a person's physical and ... Objective: The aim of this study was to detect changes in liver enzymes and psychological well-being in response to aerobic .... of mood that can be used to calculate a Total Mood.

[Glucose-6-phosphatase from nuclear envelope in rat liver].

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González-Mujica, Freddy

2008-06-01

Nuclear envelope (NE) and microsomal glucosa-6-phosphatase (G-6-Pase) activities were compared. Intact microsomes were unable to hydrolyze mannose-6-phosphate (M-6-P), on the other hand, intact NE hydrolyzes this substrate. Galactose-6-phosphate showed to be a good substrate for both NE and microsomal enzymes, with similar latency to that obtained with M-6-P using microsomes. In consequence, this substrate was used to measure the NE integrity. The kinetic parameters (Kii and Kis) of the intact NE G-6-Pase for the phlorizin inhibition using glucose-6-phosphate (G-6-P) and M-6-P as substrates, were very similar. The NE T1 transporter was more sensitive to amiloride than the microsomal T1. The microsomal system was more sensitive to N-ethylmalemide (NEM) than the NE and the latter was insensitive to anion transport inhibitors DIDS and SITS, which strongly affect the microsomal enzyme. The above results allowed to postulate the presence of a hexose-6-phosphate transporter in the NE which is able to carry G-6-P and M-6-P, and perhaps other hexose-6-phosphate which could be different from that present in microsomes or, if it is the same, its activity could by modified by the membrane system where it is included. The higher PPi hydrolysis activity of the intact NE G-6-Pase in comparison to the intact microsomal, suggests differences between the Pi/PPi transport (T2) of both systems. The lower sensitivity of the NE G-6-Pase to NEM suggests that the catalytic subunit of this system has some differences with the microsomal isoform.

Studies to further investigate the inhibition of human liver microsomal CYP2C8 by the acyl-β-glucuronide of gemfibrozil.

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Jenkins, S M; Zvyaga, T; Johnson, S R; Hurley, J; Wagner, A; Burrell, R; Turley, W; Leet, J E; Philip, T; Rodrigues, A D

2011-12-01

In previous studies, gemfibrozil acyl-β-glucuronide, but not gemfibrozil, was found to be a mechanism-based inhibitor of cytochrome P450 2C8. To better understand whether this inhibition is specific for gemfibrozil acyl-β-glucuronide or whether other glucuronide conjugates are potential substrates for inhibition of this enzyme, we evaluated several pharmaceutical compounds (as their acyl glucuronides) as direct-acting and metabolism-dependent inhibitors of CYP2C8 in human liver microsomes. Of 11 compounds that were evaluated as their acyl glucuronide conjugates, only gemfibrozil acyl-β-glucuronide exhibited mechanism-based inhibition, indicating that CYP2C8 mechanism-based inhibition is very specific to certain glucuronide conjugates. Structural analogs of gemfibrozil were synthesized, and their glucuronide conjugates were prepared to further examine the mechanism of inhibition. When the aromatic methyl groups on the gemfibrozil moiety were substituted with trifluoromethyls, the resulting glucuronide conjugate was a weaker inhibitor of CYP2C8 and mechanism-based inhibition was abolished. However, the glucuronide conjugates of monomethyl gemfibrozil analogs were mechanism-based inhibitors of CYP2C8, although not as potent as gemfibrozil acyl-β-glucuronide itself. The ortho-monomethyl analog was a more potent inhibitor than the meta-monomethyl analog, indicating that CYP2C8 favors the ortho position for oxidation and potential inhibition. Molecular modeling of gemfibrozil acyl-β-glucuronide in the CYP2C8 active site is consistent with the ortho-methyl position being the favored site of covalent attachment to the heme. Moreover, hydrogen bonding to four residues (Ser100, Ser103, Gln214, and Asn217) is implicated.

In vitro formation of metabolic-intermediate cytochrome P450 complexes in rabbit liver microsomes by tiamulin and various macrolides.

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Carletti, Monica; Gusson, Federica; Zaghini, Anna; Dacasto, Mauro; Marvasi, Luigi; Nebbia, Carlo

2003-01-01

Tiamulin and a number of macrolides were evaluated as to their ability in forming metabolic-intermediate (MI) complexes with cytochrome P450 in liver microsomes from rabbits bred for meat production. Complex formation, which occurred only in preparations where the expression of P450 3A was increased as the result of rifampicin pre-treatment and with different kinetics, was in the order tiamulin > erythromycin > TAO approximately roxithromycin approximately tylosin and did not take place with tilmicosin and spiramycin. Most of the tested compounds underwent an oxidative N-dealkylation and a good relationship could be found between the rate of N-dealkylase activity in induced preparations and the aptitude in generating MI complexes. Although the results from in vitro studies should be interpreted with caution, it is suggested that the potential for in vivo drug interactions also exists in the rabbit for tiamulin and for four out of the six tested macrolides.

Correlation of secretory phospholipase-A2 activity and fatty acids in cerebrospinal fluid with liver enzymes tests

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Sepideh Ghodoosifar

2016-02-01

Full Text Available Introduction: The aim was to determine whether secretory phospholipase-A2 (sPLA2 activity and fatty acids in cerebrospinal fluid (CSF are correlated with liver enzymes tests. Methods: CSF and serum samples were collected from 49 patients (age 18-65 as part of routine diagnostic testing. Along with serum liver enzymes aspartate aminotransferase (AST, alanine aminotransferase (ALT and alkaline phosphatase (ALP, the fatty acid composition of CSF was measured by gas liquid chromatography. CSF enzyme activities of sPLA2 were measured using the standard assay with diheptanoyl thio-phosphatidylcholin as substrate. Results: The saturated fatty acids (SFAs including palmitic acid and stearic acid were positively, and the unsaturated fatty acids including oleic acid and linoleic acid were negatively correlated with liver enzymes tests. In regression analysis with adjustment for body mass index (BMI, the elevated liver enzymes tests were positively associated with activity of sPLA2 (β > 0.31, P 0.38, P < 0.010 and negatively with total monounsaturated fatty acids (MUFAs (β < -0.40, P < 0.001 contents of CSF. Conclusion: CSF activity of sPLA2 and fatty acids may be linked to peripheral markers of liver function, suggesting an indirect impact of central fatty acids on hepatocytes function and metabolism.

Health behavior associated with liver enzymes among obese Korean adolescents, 2009-2014.

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Eun-Young Lee

Full Text Available Obesity is major risk factor for liver health. This study aimed to clarify whether specific health behaviors were associated with liver function in obese adolescents in Korea.Based on national school health examination data from 2009 to 2014, 25,142 obese or overweight students were examined for aspartate transaminase and alanine transaminase levels, and health behaviors. Multiple logistic regression was used to calculate the odds ratio for liver enzyme elevation.Subjects who thought of themselves as "very fat" had a 1.6 times higher odds ratio for liver enzyme elevations than those who thought of themselves as "normal." Those who consumed fast food 3 to 5 times weekly had 1.3 times higher odds ratio (OR = 1.27, 95% confidence interval = 1.05-1.54 for the elevation of ALT than those who did not consume fast food. Those who took sugar-sweetened beverage 3 to 5 times weekly had 1.2 times higher odds ratio (OR = 1.24, 95% confidence interval = 1.07-1.42 for the elevation of ALT than those who did not take it. Those who played computer game more than 2 hours a day showed 1.1 times higher odds ratio (OR = 1.10, 95% confidence interval = 1.01-1.21 for the elevation of ALT than those who did not.Specific food item and its frequency of consumption were identified for the positive and negative association with the elevation of liver enzymes. Self-image of body shape, sleeping time and need of help for alcohol or smoking problems also showed substantial association with the elevation.

Clinical features and effect of antiviral therapy on anti-liver/kidney microsomal antibody type 1 positive chronic hepatitis C.

Science.gov (United States)

Ferri, Silvia; Muratori, Luigi; Quarneti, Chiara; Muratori, Paolo; Menichella, Rita; Pappas, Georgios; Granito, Alessandro; Ballardini, Giorgio; Bianchi, Francesco B; Lenzi, Marco

2009-06-01

Anti-liver/kidney microsomal antibody type 1 (anti-LKM1), a serological marker of type 2 autoimmune hepatitis, is also detected in a small proportion of patients with hepatitis C. This study aimed to evaluate clinical features and effect of antiviral therapy in patients with hepatitis C who are anti-LKM1 positive. Sixty consecutive anti-LKM1 positive and 120 age and sex-matched anti-LKM1 negative chronic hepatitis C patients were assessed at diagnosis and during follow-up. Of these, 26 anti-LKM1 positive and 72 anti-LKM1 negative received antiviral therapy. Anti-LKM1 was detected by indirect immunofluorescence and immunoblot. Number of HCV-infected hepatocytes and intrahepatic CD8+ lymphocytes was determined by immunohistochemistry. At diagnosis anti-LKM1 positive patients had higher IgG levels and more intrahepatic CD8+ lymphocytes (p 0.022 and 0.046, respectively). Viral genotypes distribution and response to therapy were identical. Hepatic flares during antiviral treatment only occurred in a minority of patients in concomitance with anti-LKM1 positivity. Immune system activation is more pronounced in anti-LKM1 positive patients with hepatitis C, possibly representing the expression of autoimmune mechanisms of liver damage. Antiviral treatment is as beneficial in these patients as in anti-LKM1 negative patients, and the rare necroinflammatory flares are effectively controlled by corticosteroids, allowing subsequent resumption of antiviral therapy.

Persistently and asymptomatic raised liver enzymes as a form of presentation of Wilson's disease at pediatric age

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Catarina Matos

2015-06-01

Full Text Available Background: Wilson`s disease is a rare autosomal recessive disorder characterized by a change in the transport of copper in the liver, with progressive accumulation in this and other organs such as brain, kidney and cornea. Phenotypic expression of the disease varies widely and can range from elevated liver enzymes, fatty liver or gallstones in asymptomatic patients, to cirrhosis and fulminant hepatic failure, or disabling neuropsychiatric disease. Aim: To characterize a sample of patients with Wilson´s disease. Patients and Methods: Retrospective survey including children diagnosed with Wilson´s disease between 2002 and 2011 according to the criteria of the European Association for the Study of the Liver (2012. We analyzed family history; age, clinical data, imaging and histology at the time of diagnosis; genetic analysis; treatment and side effects; follow-up and current status. Results: We identified five patients. Three had a family history of disease. All were asymptomatic and had maintained raised liver enzymes. No patient had clinical stigmata of chronic liver disease. One female patient had overweight. All were treated with D-penicillamine, withdrawn in two patients because of side effects. Currently all patients remain asymptomatic, without evidence of progression of liver disease, with a median follow-up of 5 years and 3 months. Discussion: Our series show that Wilson´s disease may be present with raised liver enzymes in asymptomatic children. The overweight patient alerts us to screen the disease in overweight/obese patients with raised liver enzymes and/or steatosis persisting for more than six months after weight loss.

Microsomal prostaglandin E synthase-1 in rheumatic diseases

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Marina eKorotkova

2011-01-01

Full Text Available Microsomal prostaglandin E synthase-1 (mPGES-1 is a well recognized target for the development of novel anti-inflammatory drugs that can reduce symptoms of inflammation in rheumatic diseases and other inflammatory conditions. In this review, we focus on mPGES-1 in rheumatic diseases with the aim to cover the most recent advances in the understanding of mPGES-1 in rheumatoid arthritis, osteoarthritis and inflammatory myopathies. Novel findings regarding regulation of mPGES1 cell expression as well as enzyme inhibitors are also summarized.

Decrease in Activities of Selected Rat Liver Enzymes following ...

African Journals Online (AJOL)

The effects of the chemical effluent from Soap and Detergent Industry on some rat liver enzymes were investigated. Chemical analyses of both the effluent and tap water which served as the control were carried out before various concentrations of the effluent (5%v/v, 25%v/v, 50%v/v and 100%v/v) were made. The effluent ...

Modulation of liver enzymes by an Iranian preparation of Echinacea purpurea

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A. Manayi

2015-10-01

Full Text Available Hepatitis B, a common infectious disease of liver, is transmitted by blood and body fluids like semen and vaginal fluid that carry hepatitis B virus (HBV.  In chronic infection, medical care is required to decrease possibility of cirrhosis and liver cancer. In the present report, the hepatoprotective effect of an Echinacea purpurea preparation (Echiherb® has been described in a patient who suffered from HBV infection. The levels of both enzymes of aspartate aminotransferase (AST and alanine aminotransferase (ALT decreased to their normal level after 6 weeks of treatment. Therefore, this report may provide a new perspective for protection of liver in patients with HBV infection along with other diseases which damage liver cells using E. purpurea preparations.

Hepatoprotective effects of Nigella sativa L and Urtica dioica L on lipid peroxidation, antioxidant enzyme systems and liver enzymes in carbon tetrachloride-treated rats

Science.gov (United States)

Kanter, Mehmet; Coskun, Omer; Budancamanak, Mustafa

2005-01-01

AIM: To investigate the effects of Nigella sativa L (NS) and Urtica dioica L (UD) on lipid peroxidation, antioxidant enzyme systems and liver enzymes in CCl4-treated rats. METHODS: Fifty-six healthy male Wistar albino rats were used in this study. The rats were randomly allotted into one of the four experimental groups: A (CCl4-only treated), B (CCl4+UD treated), C (CCl4+NS treated) and D (CCl4+UD+NS treated), each containing 14 animals. All groups received CCl4 (0.8 mL/kg of body weight, sc, twice a week for 60 d). In addition, B, C and D groups also received daily i.p. injections of 0.2 mL/kg NS or/and 2 mL/kg UD oils for 60 d. Group A, on the other hand, received only 2 mL/kg normal saline solution for 60 d. Blood samples for the biochemical analysis were taken by cardiac puncture from randomly chosen-seven rats in each treatment group at beginning and on the 60th d of the experiment. RESULTS: The CCl4 treatment for 60 d increased the lipid peroxidation and liver enzymes, and also decreased the antioxidant enzyme levels. NS or UD treatment (alone or combination) for 60 d decreased the elevated lipid peroxidation and liver enzyme levels and also increased the reduced antioxidant enzyme levels. The weight of rats decreased in group A, and increased in groups B, C and D. CONCLUSION: NS and UD decrease the lipid per-oxidation and liver enzymes, and increase the anti-oxidant defense system activity in the CCl4-treated rats. PMID:16425366

Defining the optimal cut-off values for liver enzymes in diagnosing blunt liver injury.

Science.gov (United States)

Koyama, Tomohide; Hamada, Hirohisa; Nishida, Masamichi; Naess, Paal A; Gaarder, Christine; Sakamoto, Tetsuya

2016-01-25

Patients with blunt trauma to the liver have elevated levels of liver enzymes within a short time post injury, potentially useful in screening patients for computed tomography (CT). This study was performed to define the optimal cut-off values for serum aspartate aminotransferase (AST) and alanine aminotransferase (ALT) in patients with blunt liver injury diagnosed with contrast enhanced multi detector-row CT (CE-MDCT). All patients admitted from May 2006 to July 2013 to Teikyo University Hospital Trauma and Critical Care Center, and who underwent abdominal CE-MDCT within 3 h after blunt trauma, were retrospectively enrolled. Using receiver operating characteristic (ROC) curve analysis, the optimal cut-off values for AST and ALT were defined, and sensitivity and specificity were calculated. Of a total of 676 blunt trauma patients 64 patients were diagnosed with liver injury (Group LI+) and 612 patients without liver injury (Group LI-). Group LI+ and LI- were comparable for age, Revised Trauma Score, and Probability of survival. The groups differed in Injury Severity Score [median 21 (interquartile range 9-33) vs. 17 (9-26) (p tool for CT scan in patients otherwise eligible for observation only or as a transfer criterion to a facility with CT scan capability.

In vitro inactivation of hepatic microsomal phospholipase A/sub 2/ by the marine natural product manoalide

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Master, M.M.; Jacobs, R.S.

1986-03-01

The effects of manoalide (MLD) and several analogs (isolated from the sponge Luffariella variabilis) on mouse hepatic microsomal phospholipase A/sub 2/ (PLA/sub 2/) activity was investigated. Microsomal PLA/sub 2/, a membrane bound, Ca/sup + +/ dependent enzyme with an alkaline pH optimum, functions in intracellular phospholipid turnover. In vitro PLA/sub 2/ activity was assayed by preincubating MLD or analogs (2.5-100..mu..M) with microsomes for 60 min. at 37/sup 0/C, combining this mixture with /sup 14/C-phosphatidylcholine and CaCl/sub 2/, and incubating at 37/sup 0/C for 40 minutes. Enzyme activity was quantitated by measurement of the extracted /sup 14/C-arachidonic acid product. MLD inhibited PLA/sub 2/ in a dose-dependent manner, with an IC/sub 50/ = 94..mu..M. Lineweaver-Burk analysis suggests that MLD inhibits PLA/sub 2/ noncompetitively. One of the analogs, producing a comparable dose-response curve to MLD, was found to be more potent (IC/sub 50/ = 33..mu..M). Another analog facilitated PLA/sub 2/ activity (15%) at 25..mu..M, followed by inactivation at higher doses (IC/sub 50/ > 100 ..mu..M). Facilitation of PLA/sub 2/ activity was seen with concentrations as low as 2.5..mu..M of a third analog, and significant inactivation of PLA/sub 2/ was evident. These results indicate that MLD is not as potent against microsomal PLA/sub 2/ as has been shown with purified bee venom and cobra venom PLA/sub 2/'s.

Development of a high-throughput screening assay for stearoyl-CoA desaturase using rat liver microsomes, deuterium labeled stearoyl-CoA and mass spectrometry

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Soulard, Patricia; McLaughlin, Meg; Stevens, Jessica; Connolly, Brendan; Coli, Rocco; Wang Leyu [Research Technology Center, Pfizer Global Research and Development, Cambridge, MA (United States); Moore, Jennifer; Kuo, Ming-Shang T. [Pfizer Global Research and Development, San Diego, CA (United States); LaMarr, William A.; Ozbal, Can C. [Biotrove, Inc., Woburn, MA (United States); Bhat, B. Ganesh [Pfizer Global Research and Development, San Diego, CA (United States)], E-mail: gbhat@gnf.org

2008-10-03

Several recent reports suggest that stearoyl-CoA desaturase 1 (SCD1), the rate-limiting enzyme in monounsaturated fatty acid synthesis, plays an important role in regulating lipid homeostasis and lipid oxidation in metabolically active tissues. As several manifestations of type 2 diabetes and related metabolic disorders are associated with alterations in intracellular lipid partitioning, pharmacological manipulation of SCD1 activity might be of benefit in the treatment of these disease states. In an effort to identify small molecule inhibitors of SCD1, we have developed a mass spectrometry based high-throughput screening (HTS) assay using deuterium labeled stearoyl-CoA substrate and induced rat liver microsomes. The methodology developed allows the use of a nonradioactive substrate which avoids interference by the endogenous SCD1 substrate and/or product that exist in the non-purified enzyme source. Throughput of the assay was up to twenty 384-well assay plates per day. The assay was linear with protein concentration and time, and was saturable for stearoyl-CoA substrate (K{sub m} = 10.5 {mu}M). The assay was highly reproducible with an average Z' value = 0.6. Conjugated linoleic acid and sterculic acid, known inhibitors of SCD1, exhibited IC{sub 50} values of 0.88 and 0.12 {mu}M, respectively. High-throughput mass spectrometry screening of over 1.7 million compounds in compressed format demonstrated that the enzyme target is druggable. A total of 2515 hits were identified (0.1% hit rate), and 346 were confirmed active (>40% inhibition of total SCD activity at 20 {mu}M - 14% conformation rate). Of the confirmed hits 172 had IC{sub 50} values of <10 {mu}M, including 111 <1 {mu}M and 48 <100 nM. A large number of potent drug-like (MW < 450) hits representing six different chemical series were identified. The application of mass spectrometry to high-throughput screening permitted the development of a high-quality screening protocol for an otherwise intractable

Development of a high-throughput screening assay for stearoyl-CoA desaturase using rat liver microsomes, deuterium labeled stearoyl-CoA and mass spectrometry

International Nuclear Information System (INIS)

Soulard, Patricia; McLaughlin, Meg; Stevens, Jessica; Connolly, Brendan; Coli, Rocco; Wang Leyu; Moore, Jennifer; Kuo, Ming-Shang T.; LaMarr, William A.; Ozbal, Can C.; Bhat, B. Ganesh

2008-01-01

Several recent reports suggest that stearoyl-CoA desaturase 1 (SCD1), the rate-limiting enzyme in monounsaturated fatty acid synthesis, plays an important role in regulating lipid homeostasis and lipid oxidation in metabolically active tissues. As several manifestations of type 2 diabetes and related metabolic disorders are associated with alterations in intracellular lipid partitioning, pharmacological manipulation of SCD1 activity might be of benefit in the treatment of these disease states. In an effort to identify small molecule inhibitors of SCD1, we have developed a mass spectrometry based high-throughput screening (HTS) assay using deuterium labeled stearoyl-CoA substrate and induced rat liver microsomes. The methodology developed allows the use of a nonradioactive substrate which avoids interference by the endogenous SCD1 substrate and/or product that exist in the non-purified enzyme source. Throughput of the assay was up to twenty 384-well assay plates per day. The assay was linear with protein concentration and time, and was saturable for stearoyl-CoA substrate (K m = 10.5 μM). The assay was highly reproducible with an average Z' value = 0.6. Conjugated linoleic acid and sterculic acid, known inhibitors of SCD1, exhibited IC 50 values of 0.88 and 0.12 μM, respectively. High-throughput mass spectrometry screening of over 1.7 million compounds in compressed format demonstrated that the enzyme target is druggable. A total of 2515 hits were identified (0.1% hit rate), and 346 were confirmed active (>40% inhibition of total SCD activity at 20 μM - 14% conformation rate). Of the confirmed hits 172 had IC 50 values of <10 μM, including 111 <1 μM and 48 <100 nM. A large number of potent drug-like (MW < 450) hits representing six different chemical series were identified. The application of mass spectrometry to high-throughput screening permitted the development of a high-quality screening protocol for an otherwise intractable target, SCD1. Further medicinal

Frequency and significance of antibodies to liver/kidney microsome type 1 in adults with chronic active hepatitis.

Science.gov (United States)

Czaja, A J; Manns, M P; Homburger, H A

1992-10-01

To assess the frequency of antibodies to liver/kidney microsome type 1 (anti-LKM1) in patients with chronic active hepatitis, 131 such patients were tested by an indirect immunofluorescence assay. Of 62 patients with type 1 autoimmune hepatitis, none were seropositive. In contrast, 3 of 11 patients with autoimmune hepatitis and antimitochondrial antibodies (27%) were seropositive for anti-LKM1. Each had responded to corticosteroid therapy, and retesting of sera confirmed that each had been misclassified as antimitochondrial antibody positive. None of the patients with chronic active hepatitis B (14 patients) or C (24 patients) had anti-LKM1. Similarly, none of the 20 patients with cryptogenic disease had these antibodies. It is concluded that anti-LKM1 is specific for type 2 autoimmune hepatitis and is infrequent in adult patients seen at a referral center in the United States for chronic active hepatitis. Anti-LKM1 reactivity may be misinterpreted as antimitochondrial antibody reactivity by indirect immunofluorescence. Chronic hepatitis B and C virus infections are not important stimuli for the production of anti-LKM1, and testing for anti-LKM 1 is unlikely to clarify the nature of cryptogenic disease.

"Nifedipine in the treatment of liver toxicity induced by Acetaminophen overdose in mice "

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Kalantari H

2000-11-01

Full Text Available Acetaminophen is an analgesic and antipyretic drug, which is widely used by public and poisoning with this drug, is common. One of the most important adverse effects of acetaminophen poisoning is centrilobullar necrosis in hepatic cells, which depends on activity of microsomal cytochrome P-450 (CYP enzymes. The aim of this investigation was to find out the protective effect of nifedipine against liver toxicity caused by acetaminophen overdose (700 mg/kg as calcium channel blocker. In this study doses of 5, 50, 100, 250, 500 mg/kg of nifedipine were administered to mice orally one hour before acetaminophen administration. The negative control group receive normal saline. The positive control group was administered with acetaminophen at a dose of 700 mg/kg one hour after nifedipine administration. After 24 hours, enzyme activity (ALT, AST, histopathological examination and liver weight were compared with the control groups. The results revealed that nifedipine at dose of 500 mg/kg was the most effective and protected damage from acetaminophen toxicity.

Inhibition of the phospholipid transfer within the organelles of cells in the irradiated rat liver

Energy Technology Data Exchange (ETDEWEB)

Kaznacheev, Yu S; Kolomiytseva, I K [AN SSSR, Pushchino-na-Oke. Inst. Biologicheskoj Fiziki

1975-01-01

Phosphatidyl choline (PC) and phosphatidyl ethanolamine (PE) exchange between different subcellular fractions of liver has been studied in normal rats and 1 hr after gamma-irradiation of rats at a dose of 1200 R. The rate of PC transfer in microsome-mitochondrion and microsome-nucleus systems is 1.5 to 2 times higher than that of PE. Early after irradiation the rates of PE and PC transfer decrease in both microsome-mitochondrion and microsome-nucleus systems.

Molecular cloning of cDNAs of human liver and placenta NADH-cytochrome b5 reductase

International Nuclear Information System (INIS)

Yubisui, T.; Naitoh, Y.; Zenno, S.; Tamura, M.; Takeshita, M.; Sakaki, Y.

1987-01-01

A cDNA coding for human liver NADH-cytochrome b 5 reductase was cloned from a human liver cDNA library constructed in phage λgt11. The library was screened by using an affinity-purified rabbit antibody against NADH-cytochrome b 5 reductase of human erythrocytes. A cDNA about 1.3 kilobase pairs long was isolated. By using the cDNA as a probe, another cDNA (pb 5 R141) of 1817 base pairs was isolated that hybridized with a synthetic oligonucleotide encoding Pro-Asp-Ile-Lys-Tyr-Pro, derived from the amino acid sequence at the amino-terminal region of the enzyme from human erythrocytes. Furthermore, by using the pb 5 R141 as a probe, cDNA clones having more 5' sequence were isolated from a human placenta cDNA library. The amino acid sequences deduced from the nucleotide sequences of these cDNA clones overlapped each other and consisted of a sequence that completely coincides with that of human erythrocytes and a sequence of 19 amino acid residues extended at the amino-terminal side. The latter sequence closely resembles that of the membrane-binding domain of steer liver microsomal enzyme

Evaluation of liver marker enzymes in diabetic subjects in the ...

African Journals Online (AJOL)

The activity levels of the liver marker enzymes; Aspartate and Alanine amino transferases (AST and ALT), Alkaline Phosphatase (ALP) and Gamma glutamyltransferase (GGT) in diabetic and non-diabetic subjects within the ages of 25 to 86 were assayed. This study was for two weeks. The results indicated that the plasma ...

RATE AND CAPACITY OF HEPATIC MICROSOMAL RING HYDROXYLATION OF PHENOL TO HYDROQUINONE AND CATECHOL IN RAINBOW TROUT (ONCORHYNCHUS MYKISS)

Science.gov (United States)

Rainbow trout liver microsomes were used to study the rate of ring-hydroxylation of phenol (PH) by directly measuring the production of hydroquinone (HQ), the primary metabolite, and catechol (CAT), a secondary metabolite. An HPLC method with integrated ultroviolet (UV) and elect...

Acetanilide 4-hydroxylase and acetanilide 2-hydroxylase activity in hepatic microsomes from induced mice.

Science.gov (United States)

Lewandowski, M; Chui, Y C; Levi, P; Hodgson, E

1991-02-01

A simple and sensitive method for the separation of 14C-labelled acetanilide, 4-hydroxyacetanilide, 3-hydroxyacetanilide and 2-hydroxyacetanilide was developed using thin-layer chromatography. This separation is the basis for the assay of acetanilide 4-hydroxylase and acetanilide 2-hydroxylase activity in liver microsomes from DBA2/N male mice that had been treated with phenobarbital, 3-methylcholanthrene, isosafrole or n-butylbenzodioxole. Microsomes were incubated with [14C]acetanilide and extracted with benzene and ethyl acetate. The extract was applied to silica gel plates and developed with a hexane/isopropanol/ammonium hydroxide/water solvent system. The radiolabelled phenolic metabolites and the parent compound were detected using a Berthold Automatic TLC Linear Analyzer. Although the 4-hydroxylated metabolite was the primary product detected, this method can be used to detect other phenolic metabolites.

Liver enzymes in diabetic and non diabetic subjects with clinically diagnosed hepatitis

Directory of Open Access Journals (Sweden)

Bidhan Chandra Sarkar

2011-07-01

Full Text Available The occurrence of liver disease and raised liver enzymes is common in diabetic patients and the increasing level of enzymes indicates the severity of hepatic injury. Very few studies have addressed this issue in Bangladesh though Bangladeshi population is very much susceptible to diabetes. This study investigated a total of 1400 diabetic patients and 100 non diabetic individuals to compare the level of liver enzymes between diabetic and non-diabetic subjects. The comparisons were made among subjects who were referred to the department of Gastro-hepato-pancreatic diseases (GHPD of BIRDEM with the clinical diagnosis of chronic hepatitis and other gastro-intestinal disorders. The investigations included alanine aminotransferase (ALT, aspartate aminotransferase (AST, alkaline phosphatase (ALP and bilirubin levels. The subjects were categorized with and without hepatitis based on these investigations. The biochemical markers (ALT, AST, ALP, bilirubin did not differ significantly between non-diabetic male and female subjects. Neither the differences were significant between diabetic males and females though the diabetic patients had higher level of markers. In contrast, when compared between diabetic and non-diabetic subjects there were striking differences in either sex. Compared with the non-diabetic the diabetic subjects had significantly higher level of ALT (48.3 vs. 277.0, AST (42.0 vs. 213.0 and ALP (148 vs. 302 in males (p<0.005 for all. Similarly, these values were found significantly higher in diabetic females than their non-diabetic counterparts (p<0.01. For bilirubin, it was also found significant in males (p<0.001. The study revealed that the liver enzymes were found elevated in both diabetic and non-diabetic subjects who were referred with clinically diagnosed hepatitis. The enzymes were found markedly elevated among the diabetic than non diabetic patients, which indicate hepatic injury was more marked among the diabetic patients. Further

In vitro inhibitory effects of plumbagin, the promising antimalarial candidate, on human cytochrome P450 enzymes.

Science.gov (United States)

Sumsakul, Wiriyaporn; Chaijaroenkul, Wanna; Na-Bangchang, Kesara

2015-11-01

To investigate the propensity of plumbagin to inhibit the three isoforms of human cytochrome P450 (CYP), i.e., CYP1A2, CYP2C19, and CYP3A4 using human liver microsomes in vitro. Inhibitory effects of plumbagin on the three human CYP isoforms were investigated using pooled human liver microsomes. Phenacetin O-deethylation, omeprazole hydroxylation and nifedipine oxidation were used as selective substrates for CYP1A2, CYP2C19 and CYP3A4 activities, respectively. Concentrations of paracetamol, 5-hydroxyomeprazole, and oxidized nifedipine were determined in microsomal incubation mixture using high-performance liquid chromatography. Plumbagin showed significant inhibitory effects on all CYP isoforms, but with the most potent activity on CYP2C19-mediated omeprazole hydroxylation. The IC50 (concentration that inhibits enzyme activity by 50%) values of plumbagin and nootkatone (selective inhibitor) for CYP2C19 were (0.78 ± 0.01) and (27.31 ± 0.66) μM, respectively. The inhibitory activities on CYP1A2-mediated phenacetin O-deethylation and CYP3A4-mediated nifedipine oxidation were moderate. The IC50 values of plumbagin and α-naphthoflavone (selective inhibitor) for CYP1A2 were (1.39 ± 0.01) and (0.02 ± 0.36) μM, respectively. The corresponding IC50 values of plumbagin and ketoconazole (selective inhibitor) for CYP3A4 were (2.37 ± 0.10) and (0.18 ± 0.06) μM, respectively. Clinical relevance of the interference of human drug metabolizing enzymes should be aware of for further development scheme of plumbagin as antimalarial drug when used in combination with other antimalarial drugs which are metabolized by these CYP isoforms. Copyright © 2015 Hainan Medical College. Production and hosting by Elsevier B.V. All rights reserved.

Chromatographic separation of piracetam and its metabolite in a mixture of microsomal preparations, followed by an MS/MS analysis.

Science.gov (United States)

Sahu, Kapendra; Siddiqui, Anees A; Shaharyar, Mohammad; Ahmad, Niyaz; Anwar, Mohammad; Ahmad, Farhan J

2013-07-01

A rapid bioanalytical method was evaluated for the simultaneous determination of piracetam and its metabolite (M1) in human microsomal preparations by fast ultra-performance liquid chromatography/tandem mass spectrometry (UPLC-MS/MS). In addition, a validated method of M1 in rat plasma was developed and successfully applied on pharmacokinetic studies. The present study was carried out to determine the metabolic pathways of piracetam for phase I metabolism and used cytochrome P450 isoforms responsible for the piracetam metabolism in human liver microsomes (HLMs). While additional potential metabolites of piracetam were suggested by computer-modeling. The resulting 2-(2-oxopyrrolidin-1-yl) acetic acid was the sole metabolite detected after the microsomal treatment. The amide hydrolysis mainly underwent to form a metabolite i.e., 2-(2-oxopyrrolidin-1-yl) acetic acid (M1). Copyright © 2013 Elsevier Masson SAS. All rights reserved.

In vitro enantioselective human liver microsomal metabolism and prediction of in vivo pharmacokinetic parameters of tetrabenazine by DLLME-CE.

Science.gov (United States)

Bocato, Mariana Zuccherato; de Lima Moreira, Fernanda; de Albuquerque, Nayara Cristina Perez; de Gaitani, Cristiane Masetto; de Oliveira, Anderson Rodrigo Moraes

2016-09-05

A new capillary electrophoresis method for the enantioselective analysis of cis- and trans- dihydrotetrabenazine (diHTBZ) after in vitro metabolism by human liver microsomes (HLMs) was developed. The chiral electrophoretic separations were performed by using tris-phosphate buffer (pH 2.5) containing 1% (w/v) carboxymethyl-β-CD as background electrolyte with an applied voltage of +15kV and capillary temperature kept at 15°C. Dispersive liquid-liquid microextraction was employed to extract the analytes from HLMs. Dichloromethane was used as extraction solvent (75μL) and acetone as disperser solvent (150μL). The method was validated according to official guidelines and showed to be linear over the concentration range of 0.29-19.57μmolL(-1) (r=0.9955) for each metabolite enantiomer. Within- and between-day precision and accuracy evaluated by relative standard deviation and relative error were lower than 15% for all enantiomers. The stability assay showed that the analytes kept stable under handling, storage and in metabolism conditions. After method validation, an enantioselective in vitro metabolism and in vivo pharmacokinetic prediction was carried out. This study showed a stereoselective metabolism and the observed kinetic profile indicated a substrate inhibition behavior. DiHTBZ enantiomers were catalyzed mainly by CYP2C19 and the predicted clearance suggests that liver metabolism is the main route for TBZ elimination which supports the literature data. Copyright © 2016 Elsevier B.V. All rights reserved.

Interaction of rocuronium with human liver cytochromes P450.

Science.gov (United States)

Anzenbacherova, Eva; Spicakova, Alena; Jourova, Lenka; Ulrichova, Jitka; Adamus, Milan; Bachleda, Petr; Anzenbacher, Pavel

2015-02-01

Rocuronium is a neuromuscular blocking agent acting as a competitive antagonist of acetylcholine. Results of an inhibition of eight individual liver microsomal cytochromes P450 (CYP) are presented. As the patients are routinely premedicated with diazepam, possible interaction of diazepam with rocuronium has been also studied. Results indicated that rocuronium interacts with human liver microsomal CYPs by binding to the substrate site. Next, concentration dependent inhibition of liver microsomal CYP3A4 down to 42% (at rocuronium concentration 189 μM) was found. This effect has been confirmed with two CYP3A4 substrates, testosterone (formation of 6β-hydroxytestosterone) and diazepam (temazepam formation). CYP2C9 and CYP2C19 activities were inhibited down to 75-80% (at the same rocuronium concentration). Activities of other microsomal CYPs have not been inhibited by rocuronium. To prove the possibility of rocuronium interaction with other drugs (diazepam), the effect of rocuronium on formation of main diazepam metabolites, temazepam (by CYP3A4) and desmethyldiazepam, (also known as nordiazepam; formed by CYP2C19) in primary culture of human hepatocytes has been examined. Rocuronium has caused inhibition of both reactions by 20 and 15%, respectively. The results open a possibility that interactions of rocuronium with drugs metabolized by CYP3A4 (and possibly also CYP2C19) may be observed. Copyright © 2014 Japanese Pharmacological Society. Production and hosting by Elsevier B.V. All rights reserved.

Embryonic turkey liver: activities of biotransformation enzymes and activation of DNA-reactive carcinogens

International Nuclear Information System (INIS)

Perrone, Carmen E.; Duan, Jian Dong; Jeffrey, Alan M.; Williams, Gary M.; Ahr, Hans-Juergen; Schmidt, Ulrich; Enzmann, Harald H.

2004-01-01

Avian embryos are a potential alternative model for chemical toxicity and carcinogenicity research. Because the toxic and carcinogenic effects of some chemicals depend on bioactivation, activities of biotransformation enzymes and formation of DNA adducts in embryonic turkey liver were examined. Biochemical analyses of 22-day in ovoturkey liver post-mitochondrial fractions revealed activities of the biotransformation enzymes 7-ethoxycoumarin de-ethylase (ECOD), 7-ethoxyresorufin de-ethylase (EROD), aldrin epoxidase (ALD), epoxide hydrolase (EH), glutathione S-transferase (GST), and UDP-glucuronyltransferase (GLUT). Following the administration of phenobarbital (24 mg/egg) on day 21, enzyme activities of ECOD, EROD, ALD, EH and GLUT, but not of GST, were increased by two-fold or higher levels by day 22. In contrast, acute administration of 3-methylcholanthrene (5 mg/egg) induced only ECOD and EROD activities. Bioactivation of structurally diverse pro-carcinogens was also examined using 32 P-postlabeling for DNA adducts. In ovoexposure of turkey embryos on day 20 of gestation to 2-acetylaminofluorene (AAF), 4,4'-methylenebis(2-chloroaniline) (MOCA), benzo[a]pyrene (BaP), and 2-amino-3,8-dimethylimidazo[4,5-f]quinoxaline (MeIQx) resulted in the formation of DNA adducts in livers collected by day 21. Some of the DNA adducts had 32 P-postlabeling chromatographic migration patterns similar to DNA adducts found in livers from Fischer F344 rats exposed to the same pro-carcinogens. We conclude that 21-day embryonic turkey liver is capable of chemical biotransformation and activation of genotoxic carcinogens to form DNA adducts. Thus, turkey embryos could be utilized to investigate potential chemical toxicity and carcinogenicity. (orig.)

Embryonic turkey liver: activities of biotransformation enzymes and activation of DNA-reactive carcinogens

Energy Technology Data Exchange (ETDEWEB)

Perrone, Carmen E.; Duan, Jian Dong; Jeffrey, Alan M.; Williams, Gary M. [New York Medical College, Department of Pathology, Valhalla (United States); Ahr, Hans-Juergen; Schmidt, Ulrich [Bayer AG, Institute of Toxicology, Wuppertal (Germany); Enzmann, Harald H. [Federal Institute for Drugs and Medical Devices, Bonn (Germany)

2004-10-01

Avian embryos are a potential alternative model for chemical toxicity and carcinogenicity research. Because the toxic and carcinogenic effects of some chemicals depend on bioactivation, activities of biotransformation enzymes and formation of DNA adducts in embryonic turkey liver were examined. Biochemical analyses of 22-day in ovoturkey liver post-mitochondrial fractions revealed activities of the biotransformation enzymes 7-ethoxycoumarin de-ethylase (ECOD), 7-ethoxyresorufin de-ethylase (EROD), aldrin epoxidase (ALD), epoxide hydrolase (EH), glutathione S-transferase (GST), and UDP-glucuronyltransferase (GLUT). Following the administration of phenobarbital (24 mg/egg) on day 21, enzyme activities of ECOD, EROD, ALD, EH and GLUT, but not of GST, were increased by two-fold or higher levels by day 22. In contrast, acute administration of 3-methylcholanthrene (5 mg/egg) induced only ECOD and EROD activities. Bioactivation of structurally diverse pro-carcinogens was also examined using {sup 32}P-postlabeling for DNA adducts. In ovoexposure of turkey embryos on day 20 of gestation to 2-acetylaminofluorene (AAF), 4,4'-methylenebis(2-chloroaniline) (MOCA), benzo[a]pyrene (BaP), and 2-amino-3,8-dimethylimidazo[4,5-f]quinoxaline (MeIQx) resulted in the formation of DNA adducts in livers collected by day 21. Some of the DNA adducts had {sup 32}P-postlabeling chromatographic migration patterns similar to DNA adducts found in livers from Fischer F344 rats exposed to the same pro-carcinogens. We conclude that 21-day embryonic turkey liver is capable of chemical biotransformation and activation of genotoxic carcinogens to form DNA adducts. Thus, turkey embryos could be utilized to investigate potential chemical toxicity and carcinogenicity. (orig.)

Effect of ethylene glycol monomethyl ether and diethylene glycol monomethyl ether on hepatic metabolizing enzymes.

Science.gov (United States)

Kawamoto, T; Matsuno, K; Kayama, F; Hirai, M; Arashidani, K; Yoshikawa, M; Kodama, Y

1990-06-01

Glycol ethers have been extensively used in industry over the past 40-50 years. Numerous studies on the toxicity of glycol ethers have been performed, however, the effects of glycol ethers on the hepatic drug metabolizing enzymes are still unknown. We studied the changes of the putative metabolic enzymes, that is, the hepatic microsomal mixed function oxidase system and cytosolic alcohol dehydrogenase, by the oral administration of diEGME and EGME. Adult male Wistar rats were used. DiEGME was administered orally; 500, 1000, 2000 mg/kg for 1, 2, 5 or 20 days and EGME was 100, 300 mg/kg for 1, 2, 5 or 20 days. Decreases in liver weights were produced by highest doses of diEGME (2000 mg/kg body wt/day for 20 days) and EGME (300 mg/kg body wt/day for 20 days). DiEGME increased hepatic microsomal protein contents and induced cytochrome P-450, but not cytochrome b5 or NADPH-cytochrome c reductase. The activity of cytosolic ADH was not affected by diEGME administration. On the other hand, EGME did not change cytochrome P-450, cytochrome b5 or NADPH-cytochrome c reductase. The activity of cytosolic ADH was increased by repeated EGME treatment. Therefore it is suspected that the enzyme which takes part in the metabolism of diEGME is different from that of EGME, although diEGME is a structural homologue of EGME.

Activation of the microsomal glutathione-S-transferase and reduction of the glutathione dependent protection against lipid peroxidation by acrolein

NARCIS (Netherlands)

Haenen, G R; Vermeulen, N P; Tai Tin Tsoi, J N; Ragetli, H M; Timmerman, H; Blast, A

1988-01-01

Allyl alcohol is hepatotoxic. It is generally believed that acrolein, generated out of allyl alcohol by cytosolic alcohol dehydrogenase, is responsible for this toxicity. The effect of acrolein in vitro and in vivo on the glutathione (GSH) dependent protection of liver microsomes against lipid

The effect of trimethoprim on CYP2C8 mediated rosiglitazone metabolism in human liver microsomes and healthy subjects

Science.gov (United States)

Hruska, M W; Amico, J A; Langaee, T Y; Ferrell, R E; Fitzgerald, S M; Frye, R F

2005-01-01

Aims Rosiglitazone, a thiazolidinedione antidiabetic medication used in the treatment of Type 2 diabetes mellitus, is predominantly metabolized by the cytochrome P450 (CYP) enzyme CYP2C8. The anti-infective drug trimethoprim has been shown in vitro to be a selective inhibitor of CYP2C8. The purpose of this study was to evaluate the effect of trimethoprim on the CYP2C8 mediated metabolism of rosiglitazone in vivo and in vitro. Methods The effect of trimethoprim on the metabolism of rosiglitazone in vitro was assessed in pooled human liver microsomes. The effect in vivo was determined by evaluating rosiglitazone pharmacokinetics in the presence and absence of trimethoprim. Eight healthy subjects (four men and four women) completed a randomized, cross-over study. Subjects received single dose rosiglitazone (8 mg) in the presence and absence of trimethoprim 200 mg given twice daily for 5 days. Results Trimethoprim inhibited rosiglitazone metabolism both in vitro and in vivo. Inhibition of rosiglitazone para-hydroxylation by trimethoprim in vitro was found to be competitive with apparent Ki and IC50 values of 29 µm and 54.5 µm, respectively. In the presence of trimethoprim, rosiglitazone plasma AUC was increased by 31% (P = 0.01) from 2774 ± 645 µg l−1 h to 3643 ± 1051 µg l−1 h (95% confidence interval (Cl) for difference 189, 1549), and half-life was increased by 27% (P = 0.006) from 3.3 ± 0.5 to 4.2 ± 0.8 h (95% Cl for difference 0.36, 1.5). Trimethoprim reduced the para-O-sulphate rosiglitazone/rosiglitazone and the N-desmethylrosiglitazone/rosiglitazone AUC(0–24) ratios by 22% and 38%, respectively. Conclusions These results indicate that trimethoprim is a competitive inhibitor of CYP2C8-mediated rosiglitazone metabolism in vitro and that trimethoprim administration increases plasma rosiglitazone concentrations in healthy subjects. PMID:15606443

Absence of cross-reactivity to myeloperoxidase of anti-thyroid microsomal antibodies in patients with autoimmune thyroid diseases

NARCIS (Netherlands)

Freire, BA; Paula, ID; Paula, F; Kallenberg, GGM; Limburg, PC; Queluz, TT

Background: Thyroperoxidase is the major antigen of the thyroid microsomal antibodies (TMA) detected in autoimmune thyroid diseases. Its amino acid sequence has 44% homology with myeloperoxidase (MPO), an enzyme present in the primary granules of neutrophils and one of the major antineutrophil

Perfluorodecanoic acid enhances the formation of oleic acid in rat liver.

Science.gov (United States)

Yamamoto, A; Kawashima, Y

1997-01-01

The feeding of perfluorodecanoic acid (PFDA) to male rats at a dietary concentration of 0.005% (w/w) for 7 days resulted in a marked increase in the activity of microsomal stearoyl-CoA desaturation in the liver. This increase in the overall desaturation activity was due to the induction of terminal desaturase among the components comprising the desaturation system. In contrast, PFDA inhibited desaturation in vitro, seemingly due to interference with electron transport through the desaturation system. Accordingly, PFDA can be an inducer and also an inhibitor of delta9-desaturation. PFDA feeding enhanced the conversion of radioactive stearic acid into oleic acid in the liver in vivo, indicating that the induction of delta9-desaturase by PFDA functions in vivo. PFDA feeding increased the mass of octadecenoic acid (C18:1) in the liver and the proportion of C18:1 in microsomal lipid. A highly significant linear correlation existed between the microsomal desaturase activity and the proportion of C18:1 in microsomal lipid when compared using rats in five different physiological states: control, PFDA-fed, p-chlorophenoxyisobutyric acid (clofibric acid)-fed, starved and starved/refed. These results suggest that the increase in the hepatic level of C18:1 caused by feeding of PFDA to rats can be explained by the common concept of regulation, i.e. the hepatic level of C18:1 is under the control of delta9-desaturase. The dietary administration of PFDA also increased the content of cytochrome P-450 and the activity of 7-ethoxycoumarin O-de-ethylase in the liver. PMID:9230124

Enzyme immunoassay of oestrogen receptors in needle biopsies from human liver

DEFF Research Database (Denmark)

Becker, U; Andersen, J; Poulsen, H S

1991-01-01

For quantitative assessments of sex hormone receptors in liver tissue, ligand binding assays are inconvenient, as they require large biopsies (0.5-1.0 g). The present study shows that it is possible to measure oestrogen receptors (ER) quantitatively in needle biopsy specimens as small as 10 mg...... by modifications of a commercial enzyme immunoassay employing monoclonal antibodies. Sucrose gradient centrifugation and the dextran charcoal method served as reference methods. A consecutive series of needle biopsies from patients suspected of liver disease were investigated. The biopsies (n = 37) had a median...

Liver Enzymes and Risk of Ischemic Heart Disease and Type 2 Diabetes Mellitus: A Mendelian Randomization Study

OpenAIRE

Liu, Junxi; Au Yeung, Shiu Lun; Lin, Shi Lin; Leung, Gabriel M.; Schooling, C. Mary

2016-01-01

We used Mendelian randomization to estimate the causal effects of the liver enzymes, alanine aminotransferase (ALT), alkaline phosphatase (ALP) and gamma glutamyltransferase (GGT), on diabetes and cardiovascular disease, using genetic variants predicting these liver enzymes at genome wide significance applied to extensively genotyped case-control studies of diabetes (DIAGRAM) and coronary artery disease (CAD)/myocardial infarction (MI) (CARDIoGRAMplusC4D 1000 Genomes). Genetically higher ALT ...

Quantitative aspects of enzyme histochemistry on sections of freeze-substituted glycol methacrylate-embedded rat liver

NARCIS (Netherlands)

Frederiks, W. M.; Bosch, K. S.

1993-01-01

Freeze-substituted rat liver embedded in glycol methacrylate (GMA) has been used to demonstrate the activities of several enzymes. The following enzymes could be detected in GMA-sections by the indicated histochemical procedure(s): 5'-nucleotidase (lead salt, cerium-diaminobenzidine), alkaline

The Effect of Chlorella vulgaris Supplementation on Liver Enzymes, Serum Glucose and Lipid Profile in Patients with Non-Alcoholic Fatty Liver Disease

Directory of Open Access Journals (Sweden)

Mehrangiz Ebrahimi-Mameghani

2014-07-01

Full Text Available Background: Non-alcoholic fatty liver disease (NAFLD is becoming a public health problem worldwide and using microalgae is a new approach on its treatment. The aim of this study was to investigate the effect of Chlorella vulgaris supplementation on liver enzymes, serum glucose and lipid profile in patients with NAFLD. Methods: This double-blind randomized placebo-controlled clinical trial was conducted on 60 NAFLD patients from specialized clinics of Tabriz University of Medical Sciences from December 2011 to July 2012. The subjects were randomly allocated into 2 groups: 1 “intervention” (n=30 received 400 mg/day vitamin E plus four 300 mg tablets of Chlorella vulgaris and, 2 “placebo” (n=30 received 400 mg/day vitamin E and four placebo tablets per day for 8 weeks. Weight, liver enzymes and metabolic factors were assessed in fasting serum and dietary data was collected at baseline and end of the study. Results: Weight, liver enzymes, fasting blood sugar (FBS and lipid profile decreased significantly in both groups (P<0.05. The differences in weight, ALP and FBS between the two groups were statistically significant (P=0.01, P=0.04 and P=0.02, respectively. Conclusion: C. vulgaris seems to improve FBS and lipid profile and therefore could be considered as an effective complementary treatment in NAFLD.

Biliary excretion of ouabain in isolated perfused rat liver after treatment with microsomal enzyme inducers

International Nuclear Information System (INIS)

Nevasaari, K.; Alakare, B.; Kaerki, N.T.

1976-01-01

The effect of pretreatment with spironolactone, phenobarbital and 3,4-benzpyrene on biliary excretion of ouabain was studied in isolated perfused rat liver system after a single dose of 3 H-ouabain. Spironolactone pretreatment (100 mg/kg intraperitoneally for 4 days) changed the time course of the excretion, thus accelerating the transport of ouabain into the bile. Phenobarbital pretreatment (75 mg/kg intraperitoneally for 4 days) enhanced bile flow and increased biliary excretion of ouabain only after 15 min. At longer time periods the increase in bile flow diluted the bile level of ouabain there being no difference in the amounts excreted into the bile between the treated and untreated groups. 3,4-benzpyrene pretreatment (20 mg/kg intraperitoneally for 4 days) was without efffect on biliary excretion of ouabain. The results suggest that spironolactone differs from phenobarbital in its enhancing effect on biliary excretion of ouabain, possibly through a specific effect on an unknown hepatic transport mechanism. (author)

[The effect of monensin, tiamulin and the simultaneous administration of both substances on the microsomal mixed function oxidases and on the peroxide formation in broilers].

Science.gov (United States)

Laczay, P; Simon, F; Lehel, J

1990-09-01

The influence of Monensin, Tiamulin and the simultaneous administration of the two substances on the microsomal, mixed function oxidases was studied on cockerels. Monensin was seen to cause a slight depression in the amount of cytochrome P-450 and cytochrome b5 as well as in the activities of aniline-p-hydroxylase, p-nitrophenol-hydroxylase and p-nitroanisole-O-demethylase. Tiamulin induced a moderate increase in the amount of cytochrome P-450 and in the activities of aniline-p-hydroxylase, p-nitrophenol-hydroxylase and aminopyrine-N-demethylase. The combined administration of monensin and tiamulin resulted in marked induction of the microsomal enzymes; the amount of cytochrome P-450 reduced by metyrapone or carbon monoxide increased 2.5 or 2-times, respectively, and the activities of the tested microsomal hydroxylases and demethylases showed also an expressed increase. At the same time the formation of lipid peroxides also markedly increased and the GSH concentration was reduced. In conclusion, the results of the investigations indicate that the simultaneous application of monensin and tiamulin cause a marked induction of the drug-metabolizing microsomal enzymes and a significant increase in the lipid peroxide formation.

Investigation of the associations between low-dose serum perfluorinated chemicals and liver enzymes in US adults.

Science.gov (United States)

Lin, Chien-Yu; Lin, Lian-Yu; Chiang, Chih-Kang; Wang, Wei-Jie; Su, Yi-Ning; Hung, Kuan-Yu; Chen, Pau-Chung

2010-06-01

Perfluorinated chemicals (PFCs) have been largely used for years in a variety of products worldwide. However, the toxic effect of PFCs on exposure to the liver in the general population has not yet been determined. In this study, 2,216 adults (18 years of age or older) were recruited in a National Health and Nutrition Examination Survey (NHANES) in 1999-2000 and 2003-2004 to determine the relationship between serum level of PFCs and the levels of liver enzymes. The data were adjusted for all other confounding variants. After performing mathematical analysis, we determined when serum log-perfluorooctanoic acid (PFOA) increases in one unit, the serum alanine aminotransferase (ALT) concentration (U/l) increases by 1.86 units (95% confidence interval (CI), 1.24-2.48; P=0.005), and the serum log-gamma-glutamyltransferase (GGT) concentration (U/l) is 0.08 unit higher (95% CI, 0.05-0.11; P=0.019). The association between PFOA and liver enzymes was more evident in obese subjects, as well as subjects with insulin resistance and/or metabolic syndromes. When dividing the serum PFOA into quartiles in the fully adjusted models in subjects with a body mass index>or=30 kg/m2, the ALT level trend across the serum PFOA quartiles was significant (P=0.003). On the basis of these data, we conclude that a higher serum concentration of PFOA may cause liver enzymes to increase abnormally in the general population, particularly in obese individuals. Further studies are warranted to clarify the casual relationship between PFCs and these liver enzymes.

Potential role of liver enzymes levels as predictor markers of glucose metabolism disorders in Tunisian population.

Science.gov (United States)

Bouhajja, Houda; Abdelhedi, Rania; Amouri, Ali; Hadj Kacem, Faten; Marrakchi, Rim; Safi, Wajdi; Mrabet, Houcem; Chtourou, Lassaad; Charfi, Nadia; Fourati, Mouna; Bensassi, Salwa; Jamoussi, Kamel; Abid, Mohamed; Ayadi, Hammadi; Feki, Mouna Mnif; Elleuch, Noura Bougacha

2018-03-10

The relationship between liver enzymes and type 2 diabetes (T2D) risk is inconclusive. We aimed to evaluate the association between liver markers and risk of carbohydrate metabolism disorders and their discriminatory power for T2D prediction. This cross-sectional study enrolled 216 participants classified as normoglycemic, prediabetes, newly-diagnosed diabetes and diagnosed diabetes. All participants underwent anthropometric and biochemical measurements. The relationship between hepatic enzymes and glucose metabolism markers was evaluated by ANCOVA analyses. The associations between liver enzymes and incident carbohydrate metabolism disorders were analyzed through logistic regression and their discriminatory capacity for T2D by receiver operating characteristic (ROC) analysis. High alkaline phosphatase (AP), alanine aminotransferase (ALT), γ-glutamyltransferase (γGT) and aspartate aminotrasferase (AST) levels were independently related to decreased insulin sensitivity. Interestingly, higher AP level was significantly associated with increased risk of prediabetes (p=0.017), newly-diagnosed diabetes (p=0.004) and T2D (p=0.007). Elevated γGT level was an independent risk factor for T2D (p=0.032) and undiagnosed-T2D (p=0.010) in prediabetic and normoglycemic subjects, respectively. In ROC analysis, AP was a powerful predictor of incident diabetes and significantly improved T2D prediction. Liver enzymes within normal range, specifically AP levels, are associated with increased risk of carbohydrate metabolism disorders and significantly improved T2D prediction.

Early radiation impairment of the cholesterol metabolism in organelles of rat liver cells

Energy Technology Data Exchange (ETDEWEB)

Kaznacherev, Yu S; Kolomiitseva, I K [AN SSSR, Pushchino-na-Oke. Inst. Biologicheskoj Fiziki

1975-05-01

The incorporation of 1-C/sup 14/-acetate into cholesterol of the nuclear, mitochondrial, microsomal and 105000 g-supernatant fractions isolated from the rat liver in norm and 60 min after the exposure to 1200 r has been investigated. An increase has been observed in the label uptake into liver cholesterol of irradiated rats. Intracellular distribution of the newly synthesized (labelled) cholesterol is substantially changed after irradiation: maximum label incorporation into the cholesterol is observed in the 105000 g-supernatant fraction, whereas, normally, the cholesterol of microsomal fraction has the highest specific activity.

Effect of pH 5 enzyme from liver on the protein synthesis by mammary gland subcellular fractions in vitro

International Nuclear Information System (INIS)

Singh, Jaspal; Singh, Ajit; Ganguli, N.C.

1976-01-01

The effect of pH 5 enzyme fraction of liver on the protein synthesizing activity of the subcellular fractions of the mammary gland has been investigated. Results indicate that (1) lactating liver pH 5 enzyme stimulates protein synthesis which is enhanced by the addition of ATP-generating system and (2) the enzyme fractions from the non-lactating liver inhibits the protein synthesis by mammary fractions, but in some cases like mitochondrial and supernatant fractions of mammary it elevates the synthesis when supplemented with ATP-generating system. Chlorella protein hydrolysate- 14 C was used as a tracer and rabits were used as experimental animals. (M.G.B.)

Profile of liver enzymes in non-alcoholic fatty liver disease in patients with impaired glucose tolerance and newly detected untreated type 2 diabetes

Directory of Open Access Journals (Sweden)

Debmalya Sanyal

2015-01-01

Full Text Available Context: The perception of non-alcoholic fatty liver disease (NAFLD as an uncommon and benign condition is rapidly changing. Approximately, 70% type 2 diabetes mellitus (T2DM patients have a fatty liver, which may follow an aggressive course with necroinflammation and fibrosis. Aims: To assess the profile of liver enzymes in subjects with impaired glucose tolerance (IGT, new onset treatment naive T2DM and normal glucose tolerance (NGT with and without NAFLD. Settings and Design: Cross-sectional clinic-based study. Subjects and Methods: 152 IGT and 158 recently detected T2DM subjects aged between 30 and 69 years, along with 160 age and gender matched controls with NGT. An ultrasonography scan of the upper abdomen was done in all patients in order to examine presence of fatty liver. Anthropometry, lipid profile, liver enzymes were also analyzed in all patients. Statistical Analysis Used: Unpaired t-test, Chi-square/Fisher Exact test (for categorical variables, Pearson/Spearmen correlation test to find significant difference, association and correlation between two or more groups respectively. Results: NAFLD was significantly associated with higher alanine aminotransferase (ALT and gamma-glutamyl transferase (GGT but not ALP levels in IGT and T2DM patients. ALT, GGT significant correlated with waist circumference, body mass index, fasting insulin, homeostatic model assessment- insulin resistance, fasting blood glucose, high density lipoprotein cholesterol, triglyceride. 57% of NAFLD patients had normal ALT between 25 and 40 U/L, 53% of NAFLD subjects had normal GGT between 15 and 30 U/L. ALT 40 U/L and GGT > 30 U/L had highest positive predictivity for presence of NAFLD in our study sample. Conclusions: Mild elevations of liver enzymes in the upper normal range are associated with features of metabolic syndrome and NAFLD even in IGT and recently detected T2DM patients. Novel cut-offs for liver enzymes are warranted in order to prevent unnecessary

Posttraumatic levels of liver enzymes can reduce the need for CT in children

DEFF Research Database (Denmark)

Bruhn, Peter James; Østerballe, Lene; Hillingsø, Jens

2016-01-01

alternative diagnostic modalities. Aspartate aminotransferase (AST) and alanine aminotransferase (ALT) are hepatic enzymes, which are elevated in peripheral blood in relation to liver injury. The aim of the present study was to investigate a potential role of normal liver transaminase levels in the decision......BACKGROUND: Computed tomography (CT) is the gold standard in the initial evaluation of the hemodynamically stable patient with suspected liver trauma. However, the adverse effects of radiation exposure are of specific concern in the pediatric population. It is therefore desirable to explore...... algorithm in suspected pediatric blunt liver trauma. METHODS: Retrospective analysis of consecutively collected data from children (0-17 years) with blunt liver trauma, admitted to a single trauma centre in Denmark, between 2000 and 2013. Patients underwent abdominal CT during initial evaluation...

Short-term hepatic effects of depleted uranium on xenobiotic and bile acid metabolizing cytochrome P450 enzymes in the rat

International Nuclear Information System (INIS)

Gueguen, Y.; Souidi, M.; Baudelin, C.; Dudoignon, N.; Grison, S.; Dublineau, I.; Marquette, C.; Voisin, P.; Gourmelon, P.; Aigueperse, J.

2006-01-01

The toxicity of uranium has been demonstrated in different organs, including the kidneys, skeleton, central nervous system, and liver. However, few works have investigated the biological effects of uranium contamination on important metabolic function in the liver. In vivo studies were conducted to evaluate its effects on cytochrome P450 (CYP) enzymes involved in the metabolism of cholesterol and xenobiotics in the rat liver. The effects of depleted uranium (DU) contamination on Sprague-Dawley were measured at 1 and 3 days after exposure. Biochemical indicators characterizing liver and kidney functions were measured in the plasma. The DU affected bile acid CYP activity: 7α-hydroxycholesterol plasma level decreased by 52% at day 3 whereas microsomal CYP7A1 activity in the liver did not change significantly and mitochondrial CYP27A1 activity quintupled at day 1. Gene expression of the nuclear receptors related to lipid metabolism (FXR and LXR) also changed, while PPARα mRNA levels did not. The increased mRNA levels of the xenobiotic-metabolizing CYP3A enzyme at day 3 may be caused by feedback up-regulation due to the decreased CYP3A activity at day 1. CAR mRNA levels, which tripled on day 1, may be involved in this up-regulation, while mRNA levels of PXR did not change. These results indicate that high levels of depleted uranium, acting through modulation of the CYP enzymes and some of their nuclear receptors, affect the hepatic metabolism of bile acids and xenobiotics. (orig.)

Correlation of Endothelin-1 Concentration and Angiotensin-Converting Enzyme Activity with the Staging of Liver Fibrosis

OpenAIRE

Kardum, Duško; Fabijanić, Damir; Lukić, Anita; Romić, Željko; Petrovečki, Mladen; Bogdanović, Zoran; Jurić, Klara; Urek-Crnčević, Marija; Banić, Marko

2012-01-01

Increased serum angiotensin-converting enzyme (SACE) activity and serum concentration of endothelin-1 (ET-1) were found in liver cirrhosis. We investigated a correlation between the different stages of liver fibrosis and SACE activity and serum ET-1 concentration. Seventy patients with pathohistologically established chronic liver disease were divided in three groups according to Ishak criteria for liver fibrosis: minimal fibrosis (Ishak score 0–1, n=20), medium fibrosis (Ishak sc...

The participation of soluble factors in the omega-oxidation of fatty acids in the liver of the sheep

International Nuclear Information System (INIS)

Hare, W.R.; Wahle, K.W.

1991-01-01

The removal of soluble components from an ovine hepatic microsomal preparation decreased the omega-hydroxylation of dodecanoic and hexadecanoic acids. The results suggest that one or more soluble components play a role in the microsomal omega-hydroxylation of fatty acids. The possible roles in the reaction of catalase (known to stimulate the microsomal desaturations of fatty acids and alkylglycerols) and superoxide dismutase were investigated. The addition of these enzymes to the complete (but not the washed) microsomal preparation stimulated both the initial omega-hydroxylation reaction and the subsequent dehydrogenation reactions of the omega-oxidation pathway. The similarity of the effects of catalase and superoxide dismutase and stimulation of two different steps of the omega-oxidation pathway suggest that these agents are acting indirectly by removing active oxygen species rather than directly on the enzymes of microsomal fatty acid omega-hydroxylation

The effect of steroidal contraceptives on liver enzymes and serum ...

African Journals Online (AJOL)

This study assessed the influence of steroidal contraceptives on liver enzymes and serum total protein using 48 adult female rats in four groups -A as control and B, C and D as tests. The animals were further divided into two subgroups - treatment (A1 - D1; n=6 each) and reversal (A2 - D2; n=6 each). Groups A1&A2 ...

Antithyroid microsomal antibody

Science.gov (United States)

Thyroid antimicrosomal antibody; Antimicrosomal antibody; Microsomal antibody; Thyroid peroxidase antibody; TPOAb ... Granulomatous thyroiditis Hashimoto thyroiditis High levels of these antibodies have also been linked with an increased risk ...

Studies on the metabolism of chlorotrianisene to a reactive intermediate and subsequent covalent binding to microsomal proteins

International Nuclear Information System (INIS)

Juedes, M.J.

1989-01-01

The studies on chlorotrianisene were conducted to determine whether metabolism of chlorotrianisene occurs via the cytochrome P450 monooxygenase system and whether a reactive intermediate is being formed that is capable of binding covalently to microsomal proteins. [ 3 H]-chlorotrianisene was incubated with liver microsomes supplemented with NADPH. At the termination of the incubation, the protein was trapped on a glass filter and the unbound chlorotrianisene was removed by extensive washing of the protein with organic solvent. A dramatic stimulation of covalent binding was demonstrated in microsomes from rats treated with methylcholanthrene (60 fold increase) versus control or phenobarbital treatment. Verification of covalent binding was achieved by localization of radiolabeled bands following sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) of the macromolecules in the incubation mixture. Further analysis of the radiolabeled macromolecules separated on SDS-PAGE revealed that these macromolecules were degraded by protease degradation indicating that the macromolecules were proteins. Further investigations were done to determine the cause of the dramatic stimulation of covalent binding detected in microsomes from methylcholanthrene treated rats versus control or phenobarbital treated rats. Further evidence for the participation of P-450c was obtained with a reconstituted cytochrome P-450 system. Incubations of chlorotrianisene with reconstituted P-450c and NADPH-cytochrome P-450 reductase exhibited covalent binding characteristics comparable to those seen in microsomal incubations. Investigations into the nature of the binding site and the reactive intermediate are currently being conducted. By analyzing the BSA adduct, the author intends to isolate the specific amino acid binding site(s)

Inhibition of rat mammary microsomal oxidation of ethanol to acetaldehyde by plant polyphenols.

Science.gov (United States)

Maciel, María Eugenia; Castro, José Alberto; Castro, Gerardo Daniel

2011-07-01

We previously reported that the microsomal fraction from rat mammary tissue is able to oxidize ethanol to acetaldehyde, a mutagenic-carcinogenic metabolite, depending on the presence of NADPH and oxygen but not inhibited by carbon monoxide or other cytochrome P450 inhibitors. The process was strongly inhibited by diphenyleneiodonium, a known inhibitor of NADPH oxidase, and by nordihydroguaiaretic acid, an inhibitor of lipoxygenases. This led us to suggest that both enzymes could be involved. With the purpose of identifying natural compounds present in food with the ability to decrease the production of acetaldehyde in mammary tissue, in the present studies, several plant polyphenols having inhibitory effects on lipoxygenases and of antioxidant nature were tested as potential inhibitors of the rat mammary tissue microsomal pathway of ethanol oxidation. We included in the present screening study 32 polyphenols having ready availability and that were also tested against the rat mammary tissue cytosolic metabolism of ethanol to acetaldehyde. Several polyphenols were also able to inhibit the microsomal ethanol oxidation at concentrations as low was 10-50 μM. The results of these screening experiments suggest the potential of several plant polyphenols to prevent in vivo production and accumulation of acetaldehyde in mammary tissue.

Microsomal aryl hydrocarbon hydroxylase comparison of the direct, indirect and radiometric assays

International Nuclear Information System (INIS)

Denison, M.S.; Murray, M.; Wilkinson, C.F.

1983-01-01

The direct fluorometric assay of aryl hydrocarbon hydroxlyase has been compared to the more commonly used indirect fluorometric and radiometric assays. Although rat hepatic microsomal activities measured by the direct assay were consistently higher than those obtained by the other assays, the relative changes in activity following enzyme induction and/or inhibition were similar. The direct assay provides an accurate and rapid measure of aryl hydrocarbon hydroxylase activity and avoids several problems inherent in the indirect and radiometric assays. 2 tables

N-Hydroxylation of 4-Aminobiphenyl by CYP2E1 Produces Oxidative Stress in a Mouse Model of Chemically Induced Liver Cancer

Science.gov (United States)

Wang, Shuang; Sugamori, Kim S.; Tung, Aveline; McPherson, J. Peter; Grant, Denis M.

2015-01-01

4-Aminobiphenyl (ABP) is a trace component of cigarette smoke and hair dyes, a suspected human carcinogen and a potent rodent liver carcinogen. Postnatal exposure of mice to ABP results in a higher incidence of liver tumors in males than in females, paralleling the sex difference in human liver cancer incidence. A traditional model of ABP tumorigenesis involves initial CYP1A2-mediated N-hydroxylation, which eventually leads to production of mutagenic ABP-DNA adducts that initiate tumor growth. However, several studies have found no correlation between sex or CYP1A2 function and the DNA-damaging, mutagenic, or tumorigenic effects of ABP. Oxidative stress may be an important etiological factor for liver cancer, and it has also been linked to ABP exposure. The goals of this study were to identify novel enzyme(s) that contribute to ABP N-oxidation, and to investigate a potential role for oxidative stress in ABP liver tumorigenicity. Isozyme-selective inhibition experiments using liver microsomes from wild-type and genetically modified mice identified CYP2E1 as a major ABP N-hydroxylating enzyme. The N-hydroxylation of ABP by transiently expressed CYP2E1 produced oxidative stress in cultured mouse hepatoma cells. In vivo postnatal exposure of mice to a tumorigenic dose of ABP also produced oxidative stress in male wild-type mice, but not in male Cyp2e1(−/−) mice or in female mice. However, a stronger NRF2-associated antioxidant response was observed in females. Our results identify CYP2E1 as a novel ABP-N-oxidizing enzyme, and suggest that sex differences in CYP2E1-dependent oxidative stress and antioxidant responses to ABP may contribute to the observed sex difference in tumor incidence. PMID:25601990

Autoimmune hepatitis-specific antibodies against soluble liver antigen and liver cytosol type 1 in patients with chronic viral hepatitis

OpenAIRE

Rigopoulou, Eirini I; Mytilinaiou, Maria; Romanidou, Ourania; Liaskos, Christos; Dalekos, George N

2007-01-01

Background Non-organ specific autoantibodies are highly prevalent in patients with chronic hepatitis C (HCV). Among them, anti-liver kidney microsomal type 1 (LKM1) antibody – the serological marker of type 2 autoimmune hepatitis (AIH-2)- is detected in up to 11% of the HCV-infected subjects. On the other hand, anti-liver cytosol type 1 antibodies (anti-LC1) – either in association with anti-LKM1, or in isolation- and anti-soluble liver antigen antibodies (anti-SLA) have been considered as us...

Binding of 7-dehydrocholesterol to sterol carrier protein and vitamin D3 effect

International Nuclear Information System (INIS)

Takase, Sachiko; Oizumi, Kumiko; Moriuchi, Sachiko; Hosoya, Norimasa

1975-01-01

It was confirmed that deltasup(5,7)-sterol delta 7 -reductase activity was suppressed by cholecalciferol (vitamin D 3 ) in the enzyme system consisted of microsomes and sterol carrier protein (SCP). The enzyme activity was significantly decreased in the combination with microsomes obtained from either vitamin D-deficient or vitamin D 3 -treated rat liver and with SCP obtained from vitamin D 3 -treated rat. It was also demonstrated by the binding assay of the dextran-charcoal technique that 7-dehydrocholesterol binding to SCP could be specifically displaced by vitamin D 3 . The inhibition of cholecalciferol on 7-dehydro-cholesterol binding to liver SCP was confirmed to be non-competitive inhibition. (auth.)

Metabolism of tributyltin and triphenyltin by rat, hamster and human hepatic microsomes

Energy Technology Data Exchange (ETDEWEB)

Ohhira, Shuji; Watanabe, Masatomo; Matsui, Hisao [Department of Hygiene, Dokkyo University School of Medicine, Mibu-machi, 321-0293, Tochigi (Japan)

2003-03-01

Tributyltin and triphenyltin are metabolized by cytochrome P-450 system enzymes, and their metabolic fate may contribute to the toxicity of the chemicals. In the current study, the in vitro metabolism of tributyltin and triphenyltin by rat, hamster and human hepatic microsomes was investigated to elucidate the metabolic competence for these compounds in humans. The metabolic reaction using microsome-NADPH system that is usually conducted was not applicable to in vitro metabolism of organotins, especially triphenyltin. We therefore examined the effects of dithiothreitol (DTT), one of the antioxidants for sulfhydryl groups, to determine the in vitro metabolism of tributyltin and triphenyltin. As a result, the treatment with 0.1 mM DTT in vitro increased the activity of the microsomal monooxygenase system for metabolism of tributyltin as well as triphenyltin; the total yield of tributyltin and triphenyltin metabolites as tin increased, respectively, by approximately 1.8 and 8.9 times for rat, 2.1 and 1.2 times for hamster, and 1.6 and 1.5 times for human. It is suggested that the organotins directly inactivate cytochrome P-450 because of the interaction with critical sulfhydryl groups of the hemoprotein. We confirmed the utility of this in vitro metabolic system using DTT in the hepatic microsomes of phenobarbital (PB)-pretreated and untreated hamsters. Thus, the in vitro metabolic system described here was applied to a comparative study of the metabolism of organotins in rats, hamsters and humans. Tributyltin was metabolized more readily than triphenyltin in all the species. In humans, the in vitro metabolic pattern resembled that of hamsters, which were susceptible to in vivo triphenyltin toxicity because of incompetent metabolism. It is possible that the hamster is a qualitatively and quantitatively suitable animal model for exploring the influence of tributyltin and triphenyltin in humans. (orig.)

Rat-liver cholesterol 7α-hydroxylase. Pt. 1

International Nuclear Information System (INIS)

Cantfort, J. van; Renson, J.; Gielen, J.

1975-01-01

A new assay is described to measure the activity of cholesterol 7α-hydroxylase and compared to the conventional 14 C method used by other investigators. This method is based on the mechanism of the enzymic hydroxylation, i.e. a direct and stereospecific substitution of the 7α-hydrogen by a hydroxyl group. [7α- 3 H]cholesterol is incubated at 37 0 C and in the presence of molecular O 2 , in a medium buffered by potassium phosphate at pH 7.4 and containing liver microsomes (or 9,000 x g supernatant), NADPH, MgCl 2 and cysteamine. Tween-80 (1.5 mg/ml) is used to introduce enough substrate (300 μM) in the incubation mixture to saturate the ezyme (K(m) = 100 μM). Under these conditions the tritiated water released into the incubation medium reflects accurately the enzymic activity. The results obtained with this method are similar to the one obtained with a [4- 14 C]cholesterol technique (r = 0.96; P 3 H]cholesterol method is a complete independence from further metabolism of the first enzymic product, the 7α-hydroxycholesterol, the tritiated water representing the entire cholesterol 7α-hydroxylase activity. (orig.) [de

Mechanistic studies with solubilized rat liver steroid 5 alpha-reductase: Elucidation of the kinetic mechanism

International Nuclear Information System (INIS)

Levy, M.A.; Brandt, M.; Greway, A.T.

1990-01-01

A solubilized preparation of steroid 5 alpha-reductase from rat liver has been used in studies focused toward an understanding of the kinetic mechanism associated with enzyme catalysis. From the results of analyses with product and dead-end inhibitors, a preferentially ordered binding of substrates and release of products from the surface of the enzyme is proposed. The observations from these experiments were identical with those using the steroid 5 alpha-reductase activity associated with rat liver microsomes. The primary isotope effects on steady-state kinetic parameters when [4S-2H]NADPH was used also were consistent with an ordered kinetic mechanism. Normal isotope effects were observed for all three kinetic parameters (Vm/Km for both testosterone and NADPH and Vm) at all substrate concentrations used experimentally. Upon extrapolation to infinite concentration of testosterone, the isotope effect on Vm/Km for NADPH approached unity, indicating that the nicotinamide dinucleotide phosphate is the first substrate binding to and the second product released from the enzyme. The isotope effects on Vm/Km for testosterone at infinite concentration of cofactor and on Vm were 3.8 +/- 0.5 and 3.3 +/- 0.4, respectively. Data from the pH profiles of these three steady-state parameters and the inhibition constants (1/Ki) of competitive inhibitors versus both substrates indicate that the binding of nicotinamide dinucleotide phosphate involves coordination of its anionic 2'-phosphate to a protonated enzyme-associated base with an apparent pK near 8.0. From these results, relative limits have been placed on several of the internal rate constants used to describe the ordered mechanism of the rat liver steroid 5 alpha-reductase

First Trimester Hemolysis, Elevated Liver Enzymes, Low Platelets Syndrome in a Surrogate Pregnancy.

Science.gov (United States)

Myer, Emily; Hill, James

2015-10-01

Background The occurrence of hemolysis, elevated liver enzymes, low platelets (HELLP) syndrome before 20 weeks of gestation is rare. HELLP is a possible but rare syndrome in gestational surrogate pregnancies for surrogates with risk factors for development of preeclampsia. Case A 32-year-old patient with chronic hypertension and positive antinuclear antibody presented for prenatal care at 13 weeks and 1 day. She was a surrogate for the embryo of a 43-year-old couple. By 15 weeks she developed uncontrolled hypertension requiring hospitalization. She was expectantly managed until her condition deteriorated. At 16 weeks and 1 day she developed hemolysis, elevated liver enzymes, thrombocytopenia, and fetal demise. Conclusions HELLP syndrome is rare and carries a significant morbidity and mortality for the mother and fetus. Clinicians should encourage the surrogate to share her medical history with the embryo donor for appropriate counseling on pregnancy risks.

In vitro screening of reversible and time-dependent inhibition on CYP3A by TM208 and TM209 in rat liver microsomes

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Miaoran Ning

2012-04-01

Full Text Available TM208 and TM209, dithiocarbamate derivatives with potential anti-cancer effects, were evaluated in reversible and time-dependent cytochrome P450 (CYP 3A inhibition assays in rat liver microsomes using testosterone as probe substrate. Both compounds were found to be weak reversible inhibitors and moderate mechanism-based inhibitors of rat CYP3A. For reversible inhibition on rat CYP3A, the Ki values of competitive inhibition model were 12.10±1.75 and 13.94±1.31 μM, respectively. For time-dependent inhibition, the inactivation constants (Kl were 31.93±12.64 and 32.91±15.58 μM, respectively, and the maximum inactivation rates (kinact were 0.03497±0.0069 and 0.07259±0.0172 min−1 respectively. These findings would provide useful in vitro information for future in vivo DDI studies on TM208 or TM209.

Carbonyl Reduction of NNK by Recombinant Human Lung Enzymes. Identification of HSD17β12 as the Reductase important in (R)-NNAL formation in Human Lung.

Science.gov (United States)

Ashmore, Joseph H; Luo, Shaman; Watson, Christy J W; Lazarus, Philip

2018-05-17

4-(Methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK) is the most abundant and carcinogenic tobacco-specific nitrosamine in tobacco and tobacco smoke. The major metabolic pathway for NNK is carbonyl reduction to form the (R) and (S) enantiomers of 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanol (NNAL) which, like NNK, is a potent lung carcinogen. The goal of the present study was to characterize NNAL enantiomer formation in human lung and identify the enzymes responsible for this activity. While (S)-NNAL was the major enantiomer of NNAL formed in incubations with NNK in lung cytosolic fractions, (R)-NNAL comprised ~60 and ~95% of the total NNAL formed in lung whole cell lysates and microsomes, respectively. In studies examining the role of individual recombinant reductase enzymes in lung NNAL enantiomer formation, AKR1C1, AKR1C2, AKR1C3, AKR1C4 and CBR1 all exhibited (S)-NNAL formation activity. To identify the microsomal enzymes responsible for (R)-NNAL formation, 28 microsomal reductase enzymes were screened for expression by real-time PCR in normal human lung. HSD17β6, HSD17β12, KDSR, NSDHL, RDH10, RDH11 and SDR16C5 were all expressed at levels >HSD11β1, the only previously reported microsomal reductase enzyme with NNK-reducing activity, with HSD17β12 the most highly expressed. Of these lung-expressing enzymes, only HSD17β12 exhibited activity against NNK, forming primarily (>95%) (R)-NNAL, a pattern consistent with that observed in lung microsomes. siRNA knockdown of HSD17β12 resulted in significant decreases in (R)-NNAL formation activity in HEK293 cells. These data suggest that both cytosolic and microsomal enzymes are active against NNK and that HSD17β12 is the major active microsomal reductase that contributes to (R)-NNAL formation in human lung.

Relationships among alcoholic liver disease, antioxidants, and antioxidant enzymes.

Science.gov (United States)

Han, Kyu-Ho; Hashimoto, Naoto; Fukushima, Michihiro

2016-01-07

Excessive consumption of alcoholic beverages is a serious cause of liver disease worldwide. The metabolism of ethanol generates reactive oxygen species, which play a significant role in the deterioration of alcoholic liver disease (ALD). Antioxidant phytochemicals, such as polyphenols, regulate the expression of ALD-associated proteins and peptides, namely, catalase, superoxide dismutase, glutathione, glutathione peroxidase, and glutathione reductase. These plant antioxidants have electrophilic activity and may induce antioxidant enzymes via the Kelch-like ECH-associated protein 1-NF-E2-related factor-2 pathway and antioxidant responsive elements. Furthermore, these antioxidants are reported to alleviate cell injury caused by oxidants or inflammatory cytokines. These phenomena are likely induced via the regulation of mitogen-activating protein kinase (MAPK) pathways by plant antioxidants, similar to preconditioning in ischemia-reperfusion models. Although the relationship between plant antioxidants and ALD has not been adequately investigated, plant antioxidants may be preventive for ALD because of their electrophilic and regulatory activities in the MAPK pathway.

Mammalian folylpoly-γ-glutamate synthetase. 1. Purification and general properties of the hog liver enzyme

International Nuclear Information System (INIS)

Cichowicz, D.J.; Shane, B.

1987-01-01

Folylpolyglutamate synthetase was purified 30,000-150,000-fold from hog liver. Purification required the use of protease inhibitors, and the protein was purified to homogeneity in two forms. Both forms of the enzyme were monomers of M/sub r/ 62,000 and had similar specific activities. The specific activity of the homogeneous protein was over 2000-fold higher than reported for partially purified folylpolyglutamate synthetases from other mammalian sources. Enzyme activity was absolutely dependent on the presence of a reducing agent and a monovalent cation, of which K + was most effective. The purified enzyme catalyzed a MgATP-dependent addition of glutamate to tetrahydrofolate with the concomitant stoichiometric formation of MgADP and phosphate. Under conditions that resembled the expected substrate and enzyme concentrations in hog liver, tetrahydrofolate was metabolized to long glutamate chain length derivatives with the hexaglutamate, the major in vivo folate derivative, predominating. Enzyme activity was maximal at about pH 9.5. The high-pH optimum was primarily due to an increase in the K/sub m/ value for the L-glutamate substrate at lower pH values, and the reaction proceeded effectively at physiological pH provided high levels of glutamate were supplied

Acute cadmium intoxication induces alpha-class glutathione S-transferase protein synthesis and enzyme activity in rat liver

International Nuclear Information System (INIS)

Casalino, Elisabetta; Sblano, Cesare; Calzaretti, Giovanna; Landriscina, Clemente

2006-01-01

Acute cadmium intoxication affects glutathione S-transferase (GST) in rat liver. It has been found that 24 h after i.p. cadmium administration to rats, at a dose of 2.5 mg CdCl 2 kg -1 body weight, the activity of this enzyme in liver cytosol increased by 40%. A less stimulatory effect persisted till 48 h and thereafter the enzyme activity normalized. Since, GST isoenzymes belong to different classes in mammalian tissues, we used quantitative immunoassays to verify which family of GST isoenzymes is influenced by this intoxication. Only alpha-class glutathione S-transferase (α-GST) proteins were detected in rat liver cytosol and their level increased by about 25%, 24 h after cadmium treatment. No pi-GST isoforms were found in liver cytosol from either normal or cadmium-treated rats. Co-administration of actinomycin D with cadmium normalized both the protein level and the activity of α-GST, suggesting that some effect occurs on enzyme transcription of these isoenzymes by this metal. On the other hand, it seems unlikely that the stimulatory effect is due to the high level of peroxides caused by lipid peroxidation, since Vitamin E administration strongly reduced the TBARS level, but did not cause any GST activity decrease

Selective inhibition by chloramphenicol of pregnenolone-16 α-carbonitrile-inducible rat liver cytochrome P-450 isozymes

International Nuclear Information System (INIS)

Graves, P.E.; Kaminsky, L.S.; Halpert, J.

1986-01-01

Pregnenolone-16 α-carbonitrile (PCN) has been shown to induce, in male rats, cytochrome P-450 isozymes responsible for the formation of R-10-hydroxywarfarin and R-dehydrowarfarin. Antibodies to the major PCN-inducible isozyme (PB/PCN-E) inhibit both activities in microsomal preparations. Recently the authors have shown that PCN treatment of female rats also induces the formation of both R-warfarin metabolites. However, in both sexes chloramphenicol (CAP) treatment selectively inhibits only the rate of formation of the R-dehydrowarfarin. A decrease in microsomal P-450 content occurs after in vivo administration of CAP to PCN-treated rats of both sexes. This is in contrast to the lack of effect of CAP on P-450 levels in phenobarbital-treated rats. Covalent binding of 14 C-CAP to microsomal protein in vitro was increased 3 to 4-fold following PCN treatment. Chromatographic evidences suggests the presence of at least two PCN-induced isozymes of similar molecular weights in both male and female rat liver microsomes. These data are consistent with the multiplicity of PCN-inducible P-450 in rat liver

Effect of parenteral serum plant sterols on liver enzymes and cholesterol metabolism in a patient with short bowel syndrome.

Science.gov (United States)

Hallikainen, Maarit; Huikko, Laura; Kontra, Kirsi; Nissinen, Markku; Piironen, Vieno; Miettinen, Tatu; Gylling, Helena

2008-01-01

Hepatobiliary complications are common during parenteral nutrition. Lipid moiety in commercially available solutions contains plant sterols. It is not known whether plant sterols in parenteral nutrition interfere with hepatic function in adults. We detected how different amounts of plant sterols in parenteral nutrition solution affected serum plant sterol concentrations and liver enzymes during a 1.5-year follow-up in a patient with short bowel syndrome. Serum lipid, plant sterol, and liver enzyme levels were measured regularly during the transition from Intralipid (100% soy-based intravenous fat emulsion) to ClinOleic (an olive oil-based intravenous fat emulsion with 80% olive oil, 20% soy oil and lower plant sterols); the lipid supply was also gradually increased from 20 to 35 g/d. Plant sterols in parenteral nutrition solution and serum were measured with gas-liquid chromatography. During infusion of soy-based intravenous fat emulsion (30 g/d, total plant sterols 87 mg/d), the concentrations of sitosterol, campesterol, and stigmasterol were 4361, 1387, and 378 microg/dL, respectively, and serum liver enzyme values were >or= 2.5 times above upper limit of normal. After changing to olive oil-based intravenous fat emulsion (20-35 g/d, plant sterols 37-65 mg/d), concentrations decreased to 2148 to 2251 microg/dL for sitosterol, 569-297 microg/dL for campesterol, and 95-55 microg/dL for stigmasterol. Concomitantly, liver enzyme values decreased to 1.4 to 1.8 times above upper limit of normal at the end of follow-up. The nutrition status of the patient improved. The amount of plant sterols in lipid emulsion affects serum liver enzyme levels more than the amount of lipid.

Gd-EOB-DTPA enhanced MRI of the liver: Correlation of relative hepatic enhancement, relative renal enhancement, and liver to kidneys enhancement ratio with serum hepatic enzyme levels and eGFR

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Talakic, Emina; Steiner, Jürgen; Kalmar, Peter; Lutfi, Andre [Division of General Radiology, Department of Radiology, Medical University of Graz, Auenbruggerplatz 9, 8036 Graz (Austria); Quehenberger, Franz [Institute for Medical Informatics, Statistics and Documentation, Medical University of Graz, Auenbruggerplatz 2, 8036 Graz (Austria); Reiter, Ursula; Fuchsjäger, Michael [Division of General Radiology, Department of Radiology, Medical University of Graz, Auenbruggerplatz 9, 8036 Graz (Austria); Schöllnast, Helmut, E-mail: helmut.schoellnast@medunigraz.at [Division of General Radiology, Department of Radiology, Medical University of Graz, Auenbruggerplatz 9, 8036 Graz (Austria)

2014-04-15

Objectives: To assess the correlation of relative hepatic enhancement (RHE), relative renal enhancement (RRE) and liver to kidneys enhancement ratio (LKR) with serum hepatic enzyme levels and eGFR in Gd-EOB-DTPA enhanced MRI of the liver and to assess threshold levels for predicting enhancement of the liver parenchyma. Methods: Data of 75 patients who underwent Gd-EOB-DTPA enhanced MRI of the liver were collected. Images were obtained before contrast injection, during the early arterial phase, late arterial phase, venous phase, delayed phase, and hepatobiliary phase which was 20 min after Gd-EOB-DTPA administration. Signal intensity of the liver and the kidneys in all phases was defined using region-of-interest measurements for relative enhancement calculation. Serum hepatic enzyme levels and eGFR were available in all patients. Spearman correlation test was used to test the correlation of RHE, RRE and LKR with serum hepatic enzyme levels and eGFR. Results: In the hepatobiliary phase all serum hepatic enzymes were significantly correlated with RHE; total bilirubin (TBIL) and cholin esterase (CHE) showed strongest correlations. TBIL and CHE were significantly correlated with RRE in the arterial phases. TBIL and CHE were significantly correlated with LKR in the arterial phase and hepatobiliary phase. eGFR showed no correlation. Conclusions: In Gd-EOB-DTPA enhanced MRI, TBIL and CHE levels may predict RHE, RRE and LKR.

A cross-sectional study of the relationship between serum liver enzymes level and the incidence of impaired fasting glucose in males and females.

Science.gov (United States)

Qin, Guangming; Lu, Lihong; Xiao, Yufei; Zhu, Yimiao; Pan, Wensheng; Xu, Xiang; Shen, Shengrong; Das, Undurti N

2014-07-28

The aim of this study was to investigate the possible correlation between levels of serum liver enzymes and impaired fasting glucose (IFG) in Chinese adults and to provide a new perspective for the prevention of pre-diabetes. Serum liver enzymes of the samples including alanine aminotransferase (ALT), aspartate aminotransferase (AST), and g-glutamyl transferase (GGT), as well as plasma glucose, blood lipids, and insulin, were measured. The cumulative incidences of IFG between different quartiles of liver enzymes were compared by the chi-square test. A logistic regression model (binary regression) was used to calculate the odds ratio (OR) of IFG with 95% confidence interval (95% CI). The total incidence of IFG was 20.3% and the cumulative incidence of IFG was higher in men compared to women. In both sexes, IFG is more prevalent in higher quartiles of liver enzymes. After adjusting for age, BMI, blood pressure, triglycerides (TG), high-density lipoprotein cholesterol (HDL-C), low-density lipoprotein cholesterol (LDL-C), and total cholesterol (TC), the cumulative incidences of IFG were significantly higher in the highest quartiles of liver enzymes than in the lowest quartiles. A significantly higher cumulative incidence of IFG was found in the highest GGT quartile than in the lowest quartile for woman. The results of this study suggest that serum liver enzymes are related to the risk of IFG in Chinese adults. We infer that preserving the hepatic function may be an efficient way to prevent the development of IFG, especially in males.

[Effect of low-intensity 900 MHz frequency electromagnetic radiation on rat liver and blood serum enzyme activities].

Science.gov (United States)

Nersesova, L S; Petrosian, M S; Gazariants, M G; Mkrtchian, Z S; Meliksetian, G O; Pogosian, L G; Akopian, Zh I

2014-01-01

The comparative analysis of the rat liver and blood serum creatine kinase, alanine aminotransferase, aspartate aminotransferase, alkaline phosphatase and purine nucleoside phosphorylase post-radiation activity levels after a total two-hour long single and fractional exposure of the animals to low-intensity 900 MHz frequency electromagnetic field showed that the most sensitive enzymes to the both schedules of radiation are the liver creatine kinase, as well as the blood serum creatine kinase and alkaline phosphatase. According to the comparative analysis of the dynamics of changes in the activity level of the liver and blood serum creatine kinase, alanine aminotransferase, aspartate aminotransferase and purine nucleoside phosphorylase, both single and fractional radiation schedules do not affect the permeability of a hepatocyte cell membrane, but rather cause changes in their energetic metabolism. The correlation analysis of the post-radiation activity level changes of the investigated enzymes did not reveal a clear relationship between them. The dynamics of post-radiation changes in the activity of investigated enzyme levels following a single and short-term fractional schedules of radiation did not differ essentially.

Liver microsomal fraction is known to participate in:

African Journals Online (AJOL)

Abdullahi Balarabe

Bayero Journal of Pure and Applied Sciences, 5(1): 11 – 16. Received: November 2011 ... of diet rich in fruit and vegetable may decrease the risk of cancer ((Steinmetz .... during analysis and experiment. The differences .... the liver mitochondrial membrane (Balzan et al.,. 1999). .... and Plasma Malondialdehyde in Human.

Sera of children with hepatitis C infection and anti-liver-kidney microsome-1 antibodies recognize different CYP2D6 epitopes than adults with LKM+/HCV+ sera.

Science.gov (United States)

Herzog, D; Yamamoto, A M; Jara, P; Maggiore, G; Sarles, J; Alvarez, F

1999-11-01

Liver-kidney microsome type 1 (LKM1) antibodies are specific markers of autoimmune hepatitis (AIH) type 2. Antibodies to LKM1 have been found in 2% to 3% of adults infected with hepatitis C virus (HCV) without AIH. Thirty percent of these antibodies are directed against linear sequences of CYP2D6 protein. LKM1 antibodies in HCV+/LKM1+ sera and in sera of AIH patients do not recognize the same CYP2D6 epitopes. The current study was conducted to determine whether LKM1 antibodies in HCV+/LKM1+ children's sera are the result of the same immune response as the antibodies described in AIH type 2 and in HCV+/LKM1+ adult patients. Sera from 10 HCV+/LKM1+ children were tested against human liver microsomal and cytosolic proteins by Western blot analysis and against synthetic peptides of the CYP2D6 sequence between amino acids 200 and 429 by dot blot. The same sera were tested against radiolabeled CYP2D6 by immunoprecipitation. Four of 10 sera tested by Western blot analysis showed immunoglobulin (Ig) G-type antibodies against CYP2D6, and 2 had antibodies against proteins of 58, 66, and 84 kDa. One of the sera also contained IgM-type anti-66-kDa and 84-kDa proteins. The radioligand test detected anti-CYP2D6 antibodies in 9 of 10 patients. Five of the anti-CYP2D6-positive sera recognized a peptide between amino acids 200 and 429 including amino acids 254-271. Most HCV+/LKM1+ sera from children recognize conformational epitopes of the CYP2D6 antigen, and half recognize linear epitopes. Some HCV+/LKM1+ sera demonstrated antibodies against the AIH type 2 main antigenic site of the CYP2D6. Screening of HCV RNA should be performed before starting treatment of presumed autoimmune hepatitis associated with LKM1.

The effect of ZnO Nps of 20 nm on changes of enzyme and liver tissues of pregnant NMRI mice

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bager Seyed Alipour

2015-01-01

Full Text Available Background :Nowadays, nanotechnology has been developing rapidly and may have considerable effects on industry, society and the environment. In this research the toxicity properties of zinc oxide nanoparticles with a size of 20 nm on enzyme and liver tissue of NMRI mice were studied. Materials and Methods: This experimental study was performed in standard conditions on 25 NMRI mice with an average weight of 30 ± 3 g so that they received different doses of zinc oxide nanoparticles, an a days, for 15 days intraperitoneally. Then, blood samples were taken on day 17 of NMRI mice. The collected tissues were washed with saline and fixed in Boin΄s fluied buffer and stained with hematoxylin and eosin for histopathology evaluation. After data collection, statistical analysis was done using SAS software. Results: The results showed that activity of ALT enzyme at concentrations 50, 100, 150 and 200 mg/kg ZnO Nps at a significant level (p<0.05 increased in comparison with the control group. Histopathological investigation showed that zinc oxide nanoparticles caused severe damage in liver. Damaged liver cells develop leaky membranes and escape of intracellular enzymes into the bloodstream. Conclusion: Our findings showed that using different concentration of zinc oxide nanoparticles could be caused undesirable effects on liver with damage to hepatocyte and level elevation of liver enzymes.

2-Hexadecynoic acid inhibits plasmodial FAS-II enzymes and arrests erythrocytic and liver stage Plasmodium infections.

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Tasdemir, Deniz; Sanabria, David; Lauinger, Ina L; Tarun, Alice; Herman, Rob; Perozzo, Remo; Zloh, Mire; Kappe, Stefan H; Brun, Reto; Carballeira, Néstor M

2010-11-01

Acetylenic fatty acids are known to display several biological activities, but their antimalarial activity has remained unexplored. In this study, we synthesized the 2-, 5-, 6-, and 9-hexadecynoic acids (HDAs) and evaluated their in vitro activity against erythrocytic (blood) stages of Plasmodium falciparum and liver stages of Plasmodium yoelii infections. Since the type II fatty acid biosynthesis pathway (PfFAS-II) has recently been shown to be indispensable for liver stage malaria parasites, the inhibitory potential of the HDAs against multiple P. falciparum FAS-II (PfFAS-II) elongation enzymes was also evaluated. The highest antiplasmodial activity against blood stages of P. falciparum was displayed by 5-HDA (IC(50) value 6.6 μg/ml), whereas the 2-HDA was the only acid arresting the growth of liver stage P. yoelii infection, in both flow cytometric assay (IC(50) value 2-HDA 15.3 μg/ml, control drug atovaquone 2.5 ng/ml) and immunofluorescence analysis (IC(50) 2-HDA 4.88 μg/ml, control drug atovaquone 0.37 ng/ml). 2-HDA showed the best inhibitory activity against the PfFAS-II enzymes PfFabI and PfFabZ with IC(50) values of 0.38 and 0.58 μg/ml (IC(50) control drugs 14 and 30 ng/ml), respectively. Enzyme kinetics and molecular modeling studies revealed valuable insights into the binding mechanism of 2-HDA on the target enzymes. All HDAs showed in vitro activity against Trypanosoma brucei rhodesiense (IC(50) values 3.7-31.7 μg/ml), Trypanosoma cruzi (only 2-HDA, IC(50) 20.2 μg/ml), and Leishmania donovani (IC(50) values 4.1-13.4 μg/ml) with generally low or no significant toxicity on mammalian cells. This is the first study to indicate therapeutic potential of HDAs against various parasitic protozoa. It also points out that the malarial liver stage growth inhibitory effect of the 2-HDA may be promoted via PfFAS-II enzymes. The lack of cytotoxicity, lipophilic nature, and calculated pharmacokinetic properties suggests that 2-HDA could be a useful compound to

2-Hexadecynoic Acid Inhibits Plasmodial FAS-II Enzymes and Arrest Erythrocytic and Liver Stage Plasmodium Infections

Science.gov (United States)

Tasdemir, Deniz; Sanabria, David; Lauinger, Ina L.; Tarun, Alice; Herman, Rob; Perozzo, Remo; Zloh, Mire; Kappe, Stefan H.; Brun, Reto; Carballeira, Néstor M.

2010-01-01

Acetylenic fatty acids are known to display several biological activities, but their antimalarial activity has remained unexplored. In this study, we synthesized the 2-, 5-, 6-, and 9-hexadecynoic acids (HDAs) and evaluated their in vitro activity against erythrocytic (blood) stages of Plasmodium falciparum and liver stages of P. yoelii infections. Since the type II fatty acid biosynthesis pathway (PfFAS-II) has recently been shown to be indispensable for liver stage malaria parasites, the inhibitory potential of the HDAs against multiple P. falciparum FAS-II (PfFAS-II) elongation enzymes was also evaluated. The highest antiplasmodial activity against blood stages of P. falciparum was displayed by 5-HDA (IC50 value 6.6. μg/ml), whereas the 2-HDA was the only acid arresting the growth of liver stage P. yoelii infection, in both flow cytometric assay (IC50 value 2-HDA 15.3 μg/ml, control drug atovaquone 2.5 ng/ml) and immunofluorescense analysis (IC50 2-HDA 4.88 μg/ml, control drug atovaquone 0.37 ng/ml). 2-HDA showed the best inhibitory against the PfFAS-II enzymes PfFabI and PfFabZ with IC50 values of 0.38 and 0.58 μg/ml (IC50 control drugs 14 and 30 ng/ml) respectively. Enzyme kinetics and molecular modeling studies revealed valuable insights into the binding mechanism of 2-HDA on the target enzymes. All HDAs showed in vitro activity against Trypanosoma brucei rhodesiense (IC50 values 3.7–31.7 μg/ml), Trypanosoma cruzi (only 2-HDA, IC50 20.2 μg/ml), and Leishmania donovani (IC50 values 4.1–13.4 μg/ml) with generally low or no significant toxicity on mammalian cells. This is the first study to indicate therapeutic potential of HDAs against various parasitic protozoa. It also points out that the malarial liver stage growth inhibitory effect of the 2-HDA may be promoted via PfFAS-II enzymes. The lack of cytotoxicity, lipophilic nature and calculated pharmacokinetic properties suggest that 2-HDA could be a useful compound to study the interaction of fatty

Short-term calorie restriction feminizes the mRNA profiles of drug metabolizing enzymes and transporters in livers of mice

International Nuclear Information System (INIS)

Fu, Zidong Donna; Klaassen, Curtis D.

2014-01-01

Calorie restriction (CR) is one of the most effective anti-aging interventions in mammals. A modern theory suggests that aging results from a decline in detoxification capabilities and thus accumulation of damaged macromolecules. The present study aimed to determine how short-term CR alters mRNA profiles of genes that encode metabolism and detoxification machinery in the liver. Male C57BL/6 mice were fed CR (0, 15, 30, or 40%) diets for one month, followed by mRNA quantification of 98 xenobiotic processing genes (XPGs) in the liver, including 7 uptake transporters, 39 phase-I enzymes, 37 phase-II enzymes, 10 efflux transporters, and 5 transcription factors. In general, 15% CR did not alter mRNAs of most XPGs, whereas 30 and 40% CR altered over half of the XPGs (32 increased and 29 decreased). CR up-regulated some phase-I enzymes (fold increase), such as Cyp4a14 (12), Por (2.3), Nqo1 (1.4), Fmo2 (5.4), and Fmo3 (346), and numerous number of phase-II enzymes, such as Sult1a1 (1.2), Sult1d1 (2.0), Sult1e1 (33), Sult3a1 (2.2), Gsta4 (1.3), Gstm2 (1.3), Gstm3 (1.7), and Mgst3 (2.2). CR feminized the mRNA profiles of 32 XPGs in livers of male mice. For instance, CR decreased the male-predominantly expressed Oatp1a1 (97%) and increased the female-predominantly expressed Oatp1a4 (11). In conclusion, short-term CR alters the mRNA levels of over half of the 98 XPGs quantified in livers of male mice, and over half of these alterations appear to be due to feminization of the liver. - Highlights: • Utilized a graded CR model in male mice • The mRNA profiles of xenobiotic processing genes (XPGs) in liver were investigated. • CR up-regulates many phase-II enzymes. • CR tends to feminize the mRNA profiles of XPGs

Short-term calorie restriction feminizes the mRNA profiles of drug metabolizing enzymes and transporters in livers of mice

Energy Technology Data Exchange (ETDEWEB)

Fu, Zidong Donna [Department of Pharmacology, Toxicology, and Therapeutics, University of Kansas Medical Center, Kansas City, KS 66160 (United States); Klaassen, Curtis D., E-mail: cklaasse@kumc.edu [Department of Internal Medicine, University of Kansas Medical Center, Kansas City, KS 66160 (United States)

2014-01-01

Calorie restriction (CR) is one of the most effective anti-aging interventions in mammals. A modern theory suggests that aging results from a decline in detoxification capabilities and thus accumulation of damaged macromolecules. The present study aimed to determine how short-term CR alters mRNA profiles of genes that encode metabolism and detoxification machinery in the liver. Male C57BL/6 mice were fed CR (0, 15, 30, or 40%) diets for one month, followed by mRNA quantification of 98 xenobiotic processing genes (XPGs) in the liver, including 7 uptake transporters, 39 phase-I enzymes, 37 phase-II enzymes, 10 efflux transporters, and 5 transcription factors. In general, 15% CR did not alter mRNAs of most XPGs, whereas 30 and 40% CR altered over half of the XPGs (32 increased and 29 decreased). CR up-regulated some phase-I enzymes (fold increase), such as Cyp4a14 (12), Por (2.3), Nqo1 (1.4), Fmo2 (5.4), and Fmo3 (346), and numerous number of phase-II enzymes, such as Sult1a1 (1.2), Sult1d1 (2.0), Sult1e1 (33), Sult3a1 (2.2), Gsta4 (1.3), Gstm2 (1.3), Gstm3 (1.7), and Mgst3 (2.2). CR feminized the mRNA profiles of 32 XPGs in livers of male mice. For instance, CR decreased the male-predominantly expressed Oatp1a1 (97%) and increased the female-predominantly expressed Oatp1a4 (11). In conclusion, short-term CR alters the mRNA levels of over half of the 98 XPGs quantified in livers of male mice, and over half of these alterations appear to be due to feminization of the liver. - Highlights: • Utilized a graded CR model in male mice • The mRNA profiles of xenobiotic processing genes (XPGs) in liver were investigated. • CR up-regulates many phase-II enzymes. • CR tends to feminize the mRNA profiles of XPGs.

Effect of Electromagnetic Waves Generated by Base Transiver Station on Liver Enzymes in Female Rats

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Gholam-Ali Jelodar

2013-07-01

Full Text Available Background: This study was investigating the effect of electromagnetic wave generated by mobile and base transceiver station (900 MHz on liver enzymes in both mature and immature female age.Materials and Methods: In this study, 20 rats Sprague Dawley white mature female age 8 to 9 weeks and weight 180 to 200 g and 20 rats immature age 3 to 4 weeks, weight 80 to 100 g, each age group were randomly divided in two groups (control and test. Test groups, were daily for four hours and four different times exposed to electromagnetic waves with signal generator (900 MHz, 5-meter intervals. Control groups were kept at equal condition (themperature and light in laboratory during experiment. After at the end experimental period, blood was collected by heart puncture of all animal. Exposure to EMF generated by BTS had significant effect on liver enzymes composition in mature and immature rats.Results: AST, ALT and ALP in immature-test groups decreased significantly compared with their respective control groups (p<0.05. ALP in mature-test groups increased significantly compared with their respective control groups (p<0.05.Conclusion: These result suggest that exposure to EMF generated by BTS has a deleterious effect on liver enzymes and that this effect is more sever in immature animals.

CHANGES IN SERUM ENZYMES LEVELS ASSOCIATED WITH LIVER FUNCTIONS IN STRESSED MARWARI GOAT

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Kataria N.

2011-03-01

Full Text Available Serum enzyme levels were determined in goats of Marwari breed belonging to farmers’ stock of arid tract of Rajasthan state, India. The animals were grouped into healthy and stressed comprising of gastrointestinal parasiticised, pneumonia affected, and drought affected. The serum enzymes determined were sorbitol dehydrogenase, malate dehydrogenase, glucose-6-phosphate dehydrogenase, glutamate dehydrogenase, ornithine carbamoyl transferase, gamma-glutamayl transferase, 5’nucleotidase, glucose-6-phosphatase, arginase, and aldolase. In stressed group the mean values of all the enzymes increased significantly (p≤0.05 as compared to respective healthy mean value. All the enzymes showed highest values in the gastrointestinal parasiticised animals and least values in the animals having pneumonia. In gastrointestinal parasiticised animals maximum change was observed in G-6-Pase activity and minimum change was observed in malate dehydrogenase mean value. It was concluded that Increased activity of all the serum enzymes was due to modulation of liver functions directly or indirectly.

Role of Liver Function Enzymes in Diagnosis of Choledocholithiasis in Biliary Colic Patients

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Mohammad Hussein Mirshamsi

2011-10-01

Full Text Available Liver functional tests due to inflammatory process which induced by cholecystitis might changed and some clinicians suggested that these changes might help us to stone prediction in common bile ducts and decrease hazards of performing ERCP and other invasive procedures. Present study was performed for assessment of role of liver functional test in diagnosis of common bile duct stone in patients with cholecystitis and help in their management. Present prospective study was performed between April 2010 and March 2011 on 350 patients who come to our hospital with cholecystitis or biliary colic diagnosis. Patients with cholesistitis diagnosis were underwent operation for removing gall bladder stone and retrograde cholangiopancreatography (ERCP was performed for patients with suspicious to biliary colic and common bile duct (CBD stones. Ultrasonography, Aspartate Aminotransferases (AST, Alanine Aminotransferases (ALT, Alkaline Phosphatase (ALP and direct and total serum bilirubin were measured for all of participated patients. Mean of AST. ALT, ALP and total and direct bilirubin were had no significant differences between two study groups. In logistic regression analysis, after entering into the model only CBD diameter (OR: 20; P=0.00 and elevated serum level of ALT (OR: 2; P=0.04 were remained into the model and were known as independent predictor of cholelithiasis. Elevated level of liver enzymes had not main role in CBD diagnosis and ERCP had no to perform for suspicious CBD stone only with elevated liver enzyme and even with normal ultrasonography findings. Endosonography as non invasive procedure recommend for patients before ERCP.

The iron-responsive microsomal proteome of Aspergillus fumigatus.

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Moloney, Nicola M; Owens, Rebecca A; Meleady, Paula; Henry, Michael; Dolan, Stephen K; Mulvihill, Eoin; Clynes, Martin; Doyle, Sean

2016-03-16

Aspergillus fumigatus is an opportunistic fungal pathogen. Siderophore biosynthesis and iron acquisition are essential for virulence. Yet, limited data exist with respect to the adaptive nature of the fungal microsomal proteome under iron-limiting growth conditions, as encountered during host infection. Here, we demonstrate that under siderophore biosynthetic conditions--significantly elevated fusarinine C (FSC) and triacetylfusarinine C (TAFC) production (pproteome remodelling occurs. Specifically, a four-fold enrichment of transmembrane-containing proteins was observed with respect to whole cell lysates following ultracentrifugation-based microsomal extraction. Comparative label-free proteomic analysis of microsomal extracts, isolated following iron-replete and -deplete growth, identified 710 unique proteins. Scatterplot analysis (MaxQuant) demonstrated high correlation amongst biological replicates from each growth condition (Pearson correlation >0.96 within groups; biological replicates (n=4)). Quantitative and qualitative comparison revealed 231 proteins with a significant change in abundance between the iron-replete and iron-deplete conditions (pAspergillus fumigatus must acquire iron to facilitate growth and pathogenicity. Iron-chelating non-ribosomal peptides, termed siderophores, mediate iron uptake via membrane-localised transporter proteins. Here we demonstrate for the first time that growth of A. fumigatus under iron-deplete conditions, concomitant with siderophore biosynthesis, leads to an extensive remodelling of the microsomal proteome which includes significantly altered levels of 231 constituent proteins (96 increased and 135 decreased in abundance), many of which have not previously been localised to the microsome. We also demonstrate the first synthesis of a fluorescent version of fusarinine C, an extracellular A. fumigatus siderophore, and its uptake and localization under iron-restricted conditions. This infers the use of an A. fumigatus

The effects of space flight on some rat liver enzymes regulating carbohydrate and lipid metabolism

Science.gov (United States)

Abraham, S.; Lin, C. Y.; Klein, H. P.; Volkmann, C.

We have examined, in the livers of rats carried aboard the Cosmos 936 biosatellite, the activities of about 30 enzymes concerned with carbohydrate and lipid metabolism. In addition to the enzyme studies, the levels of glycogen and of the individual fatty acids in hepatic lipids were determined. Livers from flight and ground control rats at recovery (R0) and 25 days after recovery (R25) were used for these analyses. For all parameters measured, the most meaningful comparisons are those made between flight stationary (FS) and flight centrifuged (FC) animals at R0. When these two groups of flight rats were compared at R0, statistically significant decreases in the activity levels of glycogen phosphorylase, α-glycerol phosphate acyl transferase, diglyceride acyl transferase, aconitase and 6-phosphogluconate dehydrogenase and an increase in the palmitoyl CoA desaturase were noted in the weightless group (FS). The significance of these findings was strengthened by the fact that all enzyme activities showing alterations at R0 returned to normal 25 days postflight. When liver glycogen and total fatty acids of the two sets of flight animals were determined, significant differences that could be attributed to reduced gravity were observed. The weightless group (FS) at R0 contained, on the average, more than twice the amount of glycogen than did the centrifuged controls (FC) and a remarkable shift in the ratio of palmitate to palmitoleate was noted. These metabolic alterations, both in enzyme levels and in hepatic constituents, appear to be characteristic of the weightless condition. Our data seem to justify the conclusion that centrifugation during flight is equivalent to terrestrial gravity.

Grapefruit juice intake does not enhance but rather protects against aflatoxin B1-induced liver DNA damage through a reduction in hepatic CYP3A activity.

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Miyata, Masaaki; Takano, Hiroki; Guo, Lian Q; Nagata, Kiyoshi; Yamazoe, Yasushi

2004-02-01

Influence of grapefruit juice intake on aflatoxin B1 (AFB1)-induced liver DNA damage was examined using a Comet assay in F344 rats given 5 mg/kg AFB1 by gavage. Rats allowed free access to grapefruit juice for 5 days prior to AFB1 administration resulted in clearly reduced DNA damage in liver, to 65% of the level in rats that did not receive grapefruit juice. Furthermore, rats treated with grapefruit juice extract (100 mg/kg per os) for 5 days prior to AFB1 treatment also reduced the DNA damage to 74% of the level in rats that did not receive grapefruit juice. No significant differences in the portal blood and liver concentrations of AFB1 were observed between grapefruit juice intake rats and the controls. In an Ames assay with AFB1 using Salmonella typhimurium TA98, lower numbers of revertant colonies were detected with hepatic microsomes prepared from rats administered grapefruit juice, compared with those from control rats. Microsomal testosterone 6beta-hydroxylation was also lower with rats given grapefruit juice than with control rats. Immunoblot analyses showed a significant decrease in hepatic CYP3A content, but not CYP1A and CYP2C content, in microsomes of grapefruit juice-treated rats than in non-treated rats. No significant difference in hepatic glutathione S-transferase (GST) activity and glutathione content was observed in the two groups. GSTA5 protein was not detected in hepatic cytosol of the two groups. In microsomal systems, grapefruit juice extract inhibited AFB1-induced mutagenesis in the presence of a microsomal activation system from livers of humans as well as rats. These results suggest that grapefruit juice intake suppresses AFB1-induced liver DNA damage through inactivation of the metabolic activation potency for AFB1 in rat liver.

Single administration of recombinant IL-6 restores the gene expression of lipogenic enzymes in liver of fasting IL-6-deficient mice

DEFF Research Database (Denmark)

Gavito, A L; Cabello, R; Suarez, J

2016-01-01

BACKGROUND AND PURPOSE: Lipogenesis is intimately controlled by hormones and cytokines as well as nutritional conditions. IL-6 participates in the regulation of fatty acid metabolism in the liver. We investigated the role of IL-6 in mediating fasting/re-feeding changes in the expression of hepatic...... lipogenic enzymes. EXPERIMENTAL APPROACH: Gene and protein expression of lipogenic enzymes were examined in livers of wild-type (WT) and IL-6-deficient (IL-6(-/-) ) mice during fasting and re-feeding conditions. Effects of exogenous IL-6 administration on gene expression of these enzymes were evaluated...

Identification of the Metabolic Enzyme Involved Morusin Metabolism and Characterization of Its Metabolites by Ultraperformance Liquid Chromatography Quadrupole Time-of-Flight Mass Spectrometry (UPLC/Q-TOF-MS/MS

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Xianbao Shi

2016-01-01

Full Text Available Morusin, the important active component of a traditional Chinese medicine, Morus alba L., has been shown to exhibit many vital pharmacological activities. In this study, six recombinant CYP450 supersomes and liver microsomes were used to perform metabolic studies. Chemical inhibition studies and screening assays with recombinant human cytochrome P450s were also used to characterize the CYP450 isoforms involved in morusin metabolism. The morusin metabolites identified varied greatly among different species. Eight metabolites of morusin were detected in the liver microsomes from pigs (PLMs, rats (RLMs, and monkeys (MLMs by LC-MS/MS and six metabolites were detected in the liver microsomes from humans (HLMs, rabbits (RAMs, and dogs (DLMs. Four metabolites (M1, M2, M5, and M7 were found in all species and hydroxylation was the major metabolic transformation. CYP1A2, CYP2C9, CYP2D6, CYP2E1, CYP3A4, and CYP2C19 contributed differently to the metabolism of morusin. Compared to other CYP450 isoforms, CYP3A4 played the most significant role in the metabolism of morusin in human liver microsomes. These results are significant to better understand the metabolic behaviors of morusin among various species.

Aetiology and pathogenesis of alcoholic liver disease.

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Lieber, C S

1993-09-01

carcinogens and even nutritional factors such as vitamin A. Ethanol causes not only vitamin A depletion but it also enhances its hepatotoxicity. Furthermore, induction of the microsomal pathway contributes to increased acetaldehyde generation, with formation of protein adducts, resulting in antibody production, enzyme inactivation and decreased DNA repair; it is also associated with a striking impairment of the capacity of the liver to utilize oxygen. Moreover, acetaldehyde promotes glutathione depletion, free-radical mediated toxicity and lipid peroxidation. In addition, acetaldehyde affects hepatic collagen synthesis: both in vivo and in vitro (in cultured myofibroblasts and lipocytes), ethanol and its metabolite acetaldehyde were found to increase collagen accumulation and mRNA levels for collagen. This new understanding of the pathogenesis of alcoholic liver disease may eventually improve therapy with drugs and nutrients.

Subcellular distribution of histone-degrading enzyme activities from rat liver

International Nuclear Information System (INIS)

Heinrich, P.C.; Raydt, G.; Puschendorf, B.; Jusic, M.

1976-01-01

Chromatin prepared from liver tissue contains a histone-degrading enzyme activity with a pH optimum of 7.5-8.0, whereas chromatin isolated from purified nuclei is devoid of it. The histone-degrading enzyme activity was assayed with radioactively labelled total histones from Ehrlich ascites tumor cells. Among the different subcellular fractions assayed, only lysosomes and mitochondria exhibited histone-degrading enzymes. A pH optimum around 4.0-5.0 was found for the lysosomal fraction, whereas 7.5-8.0 has been found for mitochondria. Binding studies of frozen and thawed lysosomes or mitochondria to proteinase-free chromatin demonstrate that the proteinase associated with chromatin isolated from frozen tissue originates from damaged mitochondria. The protein degradation patterns obtained after acrylamide gel electrophoresis are similar for the chromatin-associated and the mitochondrial proteinase and different from that obtained after incubation with lysosomes. The chromatin-associated proteinase as well as the mitochondrial proteinase are strongly inhibited by 1.0 mM phenylmethanesulfonyl fluoride. Weak inhibition is found for lysosomal proteinases at pH 5. Kallikrein-trypsin inhibitor, however, inhibits lysosomal proteinase activity and has no effect on either chromatin-associated or mitochondrial proteinases. The higher template activity of chromatin isolated from a total homogenate compared to chromatin prepared from nuclei may be due to the presence of this histone-degrading enzyme activity. (orig.) [de

SUPRESSION OF MICROSOMAL OXIDATION WEAKENS HISTOCHROME’S DIURETIC EFFECT AT RATS

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O. S. Talalaeva

2013-01-01

Full Text Available Histochrome is the medicinal form of echinochrome (2, 3, 5, 6, 8-pentahydroxy-7-ethyl-1,4-naphthoquinone. Arisen during clinical application of the drug questions concerning its biotransformation have predetermined the aim of this research: to study participation liver monooxygenase system in maintenance of histochrome’s pharmacological activity.Simple and informative method of the lifetime control of liver monooxygenase systems influence on a metabolism of a medical product is the estimation of changes of pharmacological effect of a r esearched preparation on a background microsomal oxidations i nhibitor. In experiments on rats chloramphenicol action on diuretic effect of histochrome, as the most convenient for screening, was i nvestigated.To control group of animals during 10 days were hypodermically entered by histochrome in a doze of 10 mg/kg (n = 15. Experimental animals preliminary oral received 50 mg/kg of chloramphenicol before three hours of histochrome introduction (n = 16. In both groups of animals measured volume daily excretion of water, creathinin, sodium and potassium ions excretions in experimental rats each two days. The initial level of parameters of excretory kidneys functions were estimated before introduction of preparations at animals.Long-term histochrome’s injection was followed by a fivefold increasing of water excretion and simultaneously creathinin growth one. Allocation of ions of sodium was statistically significantly increased by 11-th day of experiment, and potassium ions – since the ninth day of histochrome injection. In conditions preliminary chloramphenicol applications volume daily daily urine output and creathinin excretion were essentially less control parameters. Allocation with urine of ions of sodium was decreased almost twice in comparison with the values, fixed at introduction histochrome. Excretion potassium ions ware corresponded to an initial level during all period of supervision.Taking into

Impaired rate of microsomal fatty acid elongation in undernourished neonatal rat brain

International Nuclear Information System (INIS)

Yeh, Y.Y.

1986-01-01

Hypomyelination caused by undernourishment in characterized by low concentrations of myelin lipids and marked reduction in lignocerate (C/sub 24:0/) and nervonate (C/sub 24:1/) moiety of cerebroside and sulfatide. Since microsomal elongation is the major source of long chain (22 to 24 carbons) fatty acids in the brain, the effect of neonatal undernourishment on acyl elongation was investigated. Undernourishment of suckling rats were induced after birth by restricting maternal dietary intake to 40% of that consumed by dams fed ad libitum. Neonates suckled by the normally fed dams served as controls. Microsomal elongation was measured as nmol from [2- 14 C] malonyl CoA incorporated/h per mg of protein. At 19 days of age, rates of behenoyl CoA (C/sub 22:0/) and erucoyl CoA (C/sub 22:1/) elongation in whole brain of undernourished neonates were 30-40% lower than that of the control, whereas the elongation rates of acyl CoA 16, 18 and 20 carbons in length either saturated or monounsaturated were similar in both groups. Undernourishment had no effect on cytoplasmic de novo fatty acid synthesis from acetyl CoA. If there are multiple elongation factors, the results indicate that the depressed activity of elongating enzyme(s) for C/sub 22:0/ and C/sub 22:1/ is an important contributing factor in lowering S/sub 24:0/ and C/sub 24:1/ content in cerebroside and sulfatide. This impairment may be a specific lesion leading to hypomyelination in undernourished rats

Role of CYP2B6 and CYP3A4 in the in vitro N-dechloroethylation of (R)- and (S)-ifosfamide in human liver microsomes.

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Granvil, C P; Madan, A; Sharkawi, M; Parkinson, A; Wainer, I W

1999-04-01

The central nervous system toxicity of ifosfamide (IFF), a chiral antineoplastic agent, is thought to be dependent on its N-dechloroethylation by hepatic cytochrome P-450 (CYP) enzymes. The purpose of this study was to identify the human CYPs responsible for IFF-N-dechloroethylation and their corresponding regio- and enantioselectivities. IFF exists in two enantiomeric forms, (R) - and (S)-IFF, which can be dechloroethylated at either the N2 or N3 positions, producing the corresponding (R,S)-2-dechloroethyl-IFF [(R, S)-2-DCE-IFF] and (R,S)-3-dechloroethyl-IFF [(R,S)-3-DCE-IFF]. The results of the present study suggest that the production of (R)-2-DCE-IFF and (S)-3-DCE-IFF from (R)-IFF is catalyzed by different CYPs as is the production of (S)-2-DCE-IFF and (R)-3-DCE-IFF from (S)-IFF. In vitro studies with a bank of human liver microsomes revealed that the sample-to-sample variation in the production of (S)-3-DCE-IFF from (R)-IFF and (S)-2-DCE-IFF from (S)-IFF was highly correlated with the levels of (S)-mephenytoin N-demethylation (CYP2B6), whereas (R)-2-DCE-IFF production from (R)-IFF and (R)-3-DCE-IFF production from (S)-IFF were both correlated with the activity of testosterone 6beta-hydroxylation (CYP3A4/5). Experiments with cDNA-expressed P-450 and antibody and chemical inhibition studies supported the conclusion that the formation of (S)-3-DCE-IFF and (S)-2-DCE-IFF is catalyzed primarily by CYP2B6, whereas (R)-2-DCE-IFF and (R)-3-DCE-IFF are primarily the result of CYP3A4/5 activity.

Effects of Curcuma xanthorrhiza Extracts and Their Constituents on Phase II Drug-metabolizing Enzymes Activity.

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Salleh, Nurul Afifah Mohd; Ismail, Sabariah; Ab Halim, Mohd Rohaimi

2016-01-01

Curcuma xanthorrhiza is a native Indonesian plant and traditionally utilized for a range of illness including liver damage, hypertension, diabetes, and cancer. The study determined the effects of C. xanthorrhiza extracts (ethanol and aqueous) and their constituents (curcumene and xanthorrhizol) on UDP-glucuronosyltransferase (UGT) and glutathione transferase (GST) activities. The inhibition studies were evaluated both in rat liver microsomes and in human recombinant UGT1A1 and UGT2B7 enzymes. p-nitrophenol and beetle luciferin were used as the probe substrates for UGT assay while 1-chloro-2,4-dinitrobenzene as the probe for GST assay. The concentrations of extracts studied ranged from 0.1 to 1000 μg/mL while for constituents ranged from 0.01 to 500 μM. In rat liver microsomes, UGT activity was inhibited by the ethanol extract (IC 50 =279.74 ± 16.33 μg/mL). Both UGT1A1 and UGT2B7 were inhibited by the ethanol and aqueous extracts with IC 50 values ranging between 9.59-22.76 μg/mL and 110.71-526.65 μg/Ml, respectively. Rat liver GST and human GST Pi-1 were inhibited by ethanol and aqueous extracts, respectively (IC 50 =255.00 ± 13.06 μg/mL and 580.80 ± 18.56 μg/mL). Xanthorrhizol was the better inhibitor of UGT1A1 (IC 50 11.30 ± 0.27 μM) as compared to UGT2B7 while curcumene did not show any inhibition. For GST, both constituents did not show any inhibition. These findings suggest that C. xanthorrhiza have the potential to cause herb-drug interaction with drugs that are primarily metabolized by UGT and GST enzymes. Findings from this study would suggest which of Curcuma xanthorrhiza extracts and constituents that would have potential interactions with drugs which are highly metabolized by UGT and GST enzymes. Further clinical studies can then be designed if needed to evaluate the in vivo pharmacokinetic relevance of these interactions Abbreviations Used : BSA: Bovine serum albumin, CAM: Complementary and alternative medicine, cDNA: Complementary

Metabolism of styrene in the human liver in vitro: interindividual variation and enantioselectivity

NARCIS (Netherlands)

Wenker, M. A.; Kezić, S.; Monster, A. C.; de Wolff, F. A.

2001-01-01

1. The interindividual variation and enantioselectivity of the in vitro styrene oxidation by cytochrome P450 have been investigated in 20 human microsomal liver samples. Liver samples were genotyped for the CYP2E1*6 and CYP2E1*5B alleles. 2. Kinetic analysis indicated the presence of at least two

Distribution of sterol carrier protein2 (SCP2) in rat tissues and evidence for slow turnover in liver and adrenal cortex

International Nuclear Information System (INIS)

Kharroubi, A.; Chanderbhan, R.; Fiskum, G.; Noland, B.J.; Scallen, T.J.; Vahouny, G.V.

1986-01-01

Sterol carrier protein 2 (SCP 2 ) has been implicated in the regulation of the terminal stages of hepatic cholesterol biosynthesis, and in sterol utilization for adrenal steroid hormone and hepatic bile acid synthesis. In the present studies, a highly sensitive radioimmunoassay, using [ 125 I] SCP 2 , has been developed. Highest levels of SCP 2 were found in rat liver with progressively lower levels in intestinal mucosa, adrenal, kidney, lung and testis. SCP 2 levels were low or absent in heart, brain, skeletal muscle and serum. Liver SCP 2 was largely (44%) associated with the microsomal fraction, while in adrenal, 46% was associated with mitochondria, a distribution which is consistent with the proposed roles for SCP 2 in these tissues. Levels of SCP 2 in AS 30D hepatoma cells were only 5% of those in normal liver. In liver there was no indication of diurnal rhythm of SCP 2 in the cytosol and only slight variation of the microsomal SCP 2 levels. Fasting has only slight effects on SCP 2 concentration of rat liver microsomes and cytosol. Neither ACTH nor cycloheximide treatment of rats had a significant effect on SCP 2 distribution in the adrenal. In general, these findings indicate that SCP 2 has a low turn-over rate

Metabolism of methylstenbolone studied with human liver microsomes and the uPA⁺/⁺-SCID chimeric mouse model.

Science.gov (United States)

Geldof, Lore; Lootens, Leen; Polet, Michael; Eichner, Daniel; Campbell, Thane; Nair, Vinod; Botrè, Francesco; Meuleman, Philip; Leroux-Roels, Geert; Deventer, Koen; Eenoo, Peter Van

2014-07-01

Anti-doping laboratories need to be aware of evolutions on the steroid market and elucidate steroid metabolism to identify markers of misuse. Owing to ethical considerations, in vivo and in vitro models are preferred to human excretion for nonpharmaceutical grade substances. In this study the chimeric mouse model and human liver microsomes (HLM) were used to elucidate the phase I metabolism of a new steroid product containing, according to the label, methylstenbolone. Analysis revealed the presence of both methylstenbolone and methasterone, a structurally closely related steroid. Via HPLC fraction collection, methylstenbolone was isolated and studied with both models. Using HLM, 10 mono-hydroxylated derivatives (U1-U10) and a still unidentified derivative of methylstenbolone (U13) were detected. In chimeric mouse urine only di-hydroxylated metabolites (U11-U12) were identified. Although closely related, neither methasterone nor its metabolites were detected after administration of isolated methylstenbolone. Administration of the steroid product resulted mainly in the detection of methasterone metabolites, which were similar to those already described in the literature. Methylstenbolone metabolites previously described were not detected. A GC-MS/MS multiple reaction monitoring method was developed to detect methylstenbolone misuse. In one out of three samples, previously tested positive for methasterone, methylstenbolone and U13 were additionally detected, indicating the applicability of the method. Copyright © 2014 John Wiley & Sons, Ltd.

Irreversible binding of 14C-labelled trichloroethylene to mice liver constituents in vivo and in vitro

International Nuclear Information System (INIS)

Uehleke, H.; Poplawski-Tabarelli, S.

1977-01-01

1) 14 C-labelled trichloroethylene was injected i.p. into male mice (10 μmole/g of b.w.). The radioactivity irreversibly bound to hepatic protein reached highest levels after 6 h : 2 nmole/mg in cytosol protein, 4.4 nmole/mg in mitochondrial protein, and 7.6 nmole/mg in microsomal protein. 2) The commercial trichloroethylene contained radioactive impurities binding to proteins without metabolic activation. Purification by various extraction removed 60-70% of those materials. In aerobic incubates of mice hepatic microsomes and NADPH the covalent binding rate of the purified trichloroethylene was 1.4 nmole/mg protein in 60 min. The activity of rat liver microsomes was approximately 40% less. Covalent binding increased 2-fold with microsomes of mice pretreated with phenobarbital. (orig.) [de

Time-dependent inhibition of CYP3A4 by gallic acid in human liver microsomes and recombinant systems.

Science.gov (United States)

Pu, Qiang-Hong; Shi, Liang; Yu, Chao

2015-03-01

1.Gallic acid is a main polyphenol in various fruits and plants. Inhibitory characteristics of gallic acid on CYP3A4 were still unclear. The objective of this work is hence to investigate inhibitory characteristics of gallic acid on CYP3A4 using testosterone as the probe substrate in human liver microsomes (HLMs) and recombinant CYP3A4 (rCYP3A4) systems. 2.Gallic acid caused concentration-dependent loss of CYP3A4 activity with IC50 values of 615.2 μM and 669.5 μM in HLM and rCYP3A4 systems, respectively. IC50-shift experiments showed that pre-incubation with gallic acid in the absence of NADPH contributed to 12- or 14-fold reduction of IC50 in HLM and rCYP3A4 systems, respectively, supporting a time-dependent inhibition. In HLM, time-dependent inactivation variables KI and Kinact were 485.8 μM and 0.05 min(-1), respectively. 3.Compared with the presence of NADPH, pre-incubation of gallic acid in the absence of NADPH markedly increased its inhibitory effects in HLM and rCYP3A4 systems. Those results indicate that CYP3A4 inactivation by gallic acid was independent on NADPH and was mainly mediated its oxidative products. 4.In conclusion, we showed that gallic acid weakly and time-dependently inactivated CYP3A4 via its oxidative products.

Selective inhibition of CYP2C8 by fisetin and its methylated metabolite, geraldol, in human liver microsomes.

Science.gov (United States)

Shrestha, Riya; Kim, Ju-Hyun; Nam, Wongshik; Lee, Hye Suk; Lee, Jae-Mok; Lee, Sangkyu

2018-04-01

Fisetin is a flavonol compound commonly found in edible vegetables and fruits. It has anti-tumor, antioxidant, and anti-inflammatory effects. Geraldol, the O-methyl metabolite of fisetin in mice, is reported to suppress endothelial cell migration and proliferation. Although the in vivo and in vitro effects of fisetin and its metabolites are frequently reported, studies on herb-drug interactions have not yet been performed. This study was designed to investigate the inhibitory effect of fisetin and geraldol on eight isoforms of human cytochrome P450 (CYP) by using cocktail assay and LC-MS/MS analysis. The selective inhibition of CYP2C8-catalyzed paclitaxel hydroxylation by fisetin and geraldol were confirmed in pooled human liver microsomes (HLMs). In addition, an IC 50 shift assay under different pre-incubation conditions confirmed that fisetin and geraldol shows a reversible concentration-dependent, but not mechanism-based, inhibition of CYP2C8. Moreover, Michaelis-Menten, Lineweaver-burk plots, Dixon and Eadie-Hofstee showed a non-competitive inhibition mode with an equilibrium dissociation constant of 4.1 μM for fisetin and 11.5 μM for geraldol, determined from secondary plot of the Lineweaver-Burk plot. In conclusion, our results indicate that fisetin showed selective reversible and non-competitive inhibition of CYP2C8 more than its main metabolite, geraldol, in HLMs. Copyright © 2018 The Japanese Society for the Study of Xenobiotics. Published by Elsevier Ltd. All rights reserved.

Measure of enzymatic activity coincident with 2450 MHz microwave exposure

Energy Technology Data Exchange (ETDEWEB)

Ward, T R; Allis, J W; Elder, J A

1975-09-01

Enzyme preparations were exposed to microwave radiation at 2450 MHz and enzymatic activity was simultaneously monitored spectrophotometrically with a crossed-beam exposure detection system. Enzymes studied were glucose 6-phosphate dehydrogenase from human red blood cells and yeast, adenylate kinase from rat liver mitochondria and rabbit muscle, and rat liver microsomal NADPH cytochrome c reductase. No difference was found between the specific activity at 25/sup 0/C of unirradiated controls and enzyme preparations irradiated at an absorbed dose rate of 42 W/kg.

Chimeric mice with humanized liver: Application in drug metabolism and pharmacokinetics studies for drug discovery.

Science.gov (United States)

Naritomi, Yoichi; Sanoh, Seigo; Ohta, Shigeru

2018-02-01

Predicting human drug metabolism and pharmacokinetics (PK) is key to drug discovery. In particular, it is important to predict human PK, metabolite profiles and drug-drug interactions (DDIs). Various methods have been used for such predictions, including in vitro metabolic studies using human biological samples, such as hepatic microsomes and hepatocytes, and in vivo studies using experimental animals. However, prediction studies using these methods are often inconclusive due to discrepancies between in vitro and in vivo results, and interspecies differences in drug metabolism. Further, the prediction methods have changed from qualitative to quantitative to solve these issues. Chimeric mice with humanized liver have been developed, in which mouse liver cells are mostly replaced with human hepatocytes. Since human drug metabolizing enzymes are expressed in the liver of these mice, they are regarded as suitable models for mimicking the drug metabolism and PK observed in humans; therefore, these mice are useful for predicting human drug metabolism and PK. In this review, we discuss the current state, issues, and future directions of predicting human drug metabolism and PK using chimeric mice with humanized liver in drug discovery. Copyright © 2017 The Japanese Society for the Study of Xenobiotics. Published by Elsevier Ltd. All rights reserved.

Effect of Simulated Microgravity on the Activity of Regulatory Enzymes of Glycolysis and Gluconeogenesis in Mice Liver

Science.gov (United States)

Ramirez, Joaquin; Periyakaruppan, Adaikkappan; Sarkar, Shubhashish; Ramesh, Govindarajan T.; Sharma, S. Chidananda

2014-02-01

Gravity supports all the life activities present on earth. Microgravity environments have effect on the biological functions and physiological status of an individual. The present study was undertaken to investigate the effect of simulated microgravity on important regulatory enzymes of carbohydrate metabolism in liver using HLS mice model. Following hind limb unloading of mice for 11 days the animal's average body weights were found to be not different, while the liver weights were decreased and found to be significantly different ( p gluconeogenesis in liver and reciprocally regulated.

Coffee bean extracts rich and poor in kahweol both give rise to elevation of liver enzymes in healthy volunteers

Directory of Open Access Journals (Sweden)

Schouten Evert G

2004-07-01

Full Text Available Abstract Background Coffee oil potently raises serum cholesterol levels in humans. The diterpenes cafestol and kahweol are responsible for this elevation. Coffee oil also causes elevation of liver enzyme levels in serum. It has been suggested that cafestol is mainly responsible for the effect on serum cholesterol levels and that kahweol is mainly responsible for the effect on liver enzyme levels. The objective of this study was to investigate whether coffee oil that only contains a minute amount of kahweol indeed does not cause elevation of liver enzyme levels. Methods The response of serum alanine aminotransferase (ALAT and aspartate aminotransferase (ASAT to Robusta coffee oil (62 mg/day cafestol, 1.6 mg/day kahweol was measured in 18 healthy volunteers. Results After nine days one subject was taken off Robusta oil treatment due to an ALAT level of 3.6 times the upper limit of normal (ULN. Another two subjects stopped treatment due to other reasons. After 16 days another two subjects were taken off Robusta oil treatment. One of those subjects had levels of 5.8 ULN for ALAT and 2.0 ULN for ASAT; the other subject had an ALAT level of 12.4 ULN and an ASAT level of 4.7 ULN. It was then decided to terminate the study. The median response of subjects to Robusta oil after 16 days was 0.27 ULN (n = 15, 25th,75th percentile: 0.09;0.53 for ALAT and 0.06 ULN (25th,75th percentile -0.06;0.22 for ASAT. Conclusions We conclude that the effect on liver enzyme levels of coffee oil containing hardly any kahweol is similar to that of coffee oil containing high amounts of kahweol. Therefore it is unlikely that kahweol is the component of coffee oil that is responsible for the effect. Furthermore, we conclude that otherwise unexplained elevation of liver enzyme levels observed in patients might be caused by a switch from consumption of filtered coffee to unfiltered coffee.

Autoimmune liver serology: current diagnostic and clinical challenges.

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Bogdanos, Dimitrios-P; Invernizzi, Pietro; Mackay, Ian-R; Vergani, Diego

2008-06-07

Liver-related autoantibodies are crucial for the correct diagnosis and classification of autoimmune liver diseases (AiLD), namely autoimmune hepatitis types 1 and 2 (AIH-1 and 2), primary biliary cirrhosis (PBC), and the sclerosing cholangitis variants in adults and children. AIH-1 is specified by anti-nuclear antibody (ANA) and smooth muscle antibody (SMA). AIH-2 is specified by antibody to liver kidney microsomal antigen type-1 (anti-LKM1) and anti-liver cytosol type 1 (anti-LC1). SMA, ANA and anti-LKM antibodies can be present in de-novo AIH following liver transplantation. PBC is specified by antimitochondrial antibodies (AMA) reacting with enzymes of the 2-oxo-acid dehydrogenase complexes (chiefly pyruvate dehydrogenase complex E2 subunit) and disease-specific ANA mainly reacting with nuclear pore gp210 and nuclear body sp100. Sclerosing cholangitis presents as at least two variants, first the classical primary sclerosing cholangitis (PSC) mostly affecting adult men wherein the only (and non-specific) reactivity is an atypical perinuclear antineutrophil cytoplasmic antibody (p-ANCA), also termed perinuclear anti-neutrophil nuclear antibodies (p-ANNA) and second the childhood disease called autoimmune sclerosing cholangitis (ASC) with serological features resembling those of type 1 AIH. Liver diagnostic serology is a fast-expanding area of investigation as new purified and recombinant autoantigens, and automated technologies such as ELISAs and bead assays, become available to complement (or even compete with) traditional immunofluorescence procedures. We survey for the first time global trends in quality assurance impacting as it does on (1) manufacturers/purveyors of kits and reagents, (2) diagnostic service laboratories that fulfill clinicians' requirements, and (3) the end-user, the physician providing patient care, who must properly interpret test results in the overall clinical context.

Evaluation of coagulation parameters and liver enzymes among alcohol drinkers in Port Harcourt, Nigeria

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Adias TC

2013-06-01

Full Text Available Teddy Charles Adias,1 Everton Egerton,2 Osaro Erhabor3 1Bayelsa College of Health Technology, Bayelsa State, Nigeria; 2Department of Medical Laboratory Science, Rivers State University of Science and Technology, Port Harcourt, Nigeria; 3Faculty of Medical Laboratory Science, Department of Haematology and Transfusion Medicine, Usmanu Danfodiyo University, Sokoto, Nigeria Abstract: Alcohol is a major contributor to the global burden of disease, disability, and death in high, middle, and low-income countries. Harmful use of alcohol is one of the main factors contributing to premature deaths and avoidable disease burden worldwide and has a major impact on public health. The aim of this present cross-sectional study was to investigate the effect of alcohol consumption on coagulation parameters and liver enzymes of subjects in Port Harcourt, Nigeria. Two hundred adults consisting of 120 alcohol dependent subjects and 80 age, gender-matched nondrinkers aged 25–65 years (mean age 45.25 ± 11.50 years were enrolled in this study. Of the 120 chronic alcohol drinkers, 37 were dependent on local dry gin, while 83 were dependent on other alcoholic beverages. The mean values of the liver enzymes, aspartate aminotransferase and gamma glutamyl transferase, were significantly higher (P = 0.002 and P = 0.02 respectively among the chronic alcohol consumers compared with their nondrinker counterparts. Although the value of alanine aminotransferase was higher in the chronic drinkers, it did not reveal any significant difference (P = 0.11. The coagulation parameters, prothrombin time and activated partial thromboplastin time were investigated among chronic drinkers and nondrinkers. The mean value of prothrombin time and activated partial thromboplastin time was significantly higher in the chronic alcohol drinkers compared to the nondrinkers (P = 0.04 and P = 0.02 respectively. We observed a positive and significant correlation between values of liver enzymes, serum

Short-term calorie restriction feminizes the mRNA profiles of drug metabolizing enzymes and transporters in livers of mice.

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Fu, Zidong Donna; Klaassen, Curtis D

2014-01-01

Calorie restriction (CR) is one of the most effective anti-aging interventions in mammals. A modern theory suggests that aging results from a decline in detoxification capabilities and thus accumulation of damaged macromolecules. The present study aimed to determine how short-term CR alters mRNA profiles of genes that encode metabolism and detoxification machinery in the liver. Male C57BL/6 mice were fed CR (0, 15, 30, or 40%) diets for one month, followed by mRNA quantification of 98 xenobiotic processing genes (XPGs) in the liver, including 7 uptake transporters, 39 phase-I enzymes, 37 phase-II enzymes, 10 efflux transporters, and 5 transcription factors. In general, 15% CR did not alter mRNAs of most XPGs, whereas 30 and 40% CR altered over half of the XPGs (32 increased and 29 decreased). CR up-regulated some phase-I enzymes (fold increase), such as Cyp4a14 (12), Por (2.3), Nqo1 (1.4), Fmo2 (5.4), and Fmo3 (346), and numerous number of phase-II enzymes, such as Sult1a1 (1.2), Sult1d1 (2.0), Sult1e1 (33), Sult3a1 (2.2), Gsta4 (1.3), Gstm2 (1.3), Gstm3 (1.7), and Mgst3 (2.2). CR feminized the mRNA profiles of 32 XPGs in livers of male mice. For instance, CR decreased the male-predominantly expressed Oatp1a1 (97%) and increased the female-predominantly expressed Oatp1a4 (11). In conclusion, short-term CR alters the mRNA levels of over half of the 98 XPGs quantified in livers of male mice, and over half of these alterations appear to be due to feminization of the liver. Copyright © 2013 Elsevier Inc. All rights reserved.

The mouse liver displays daily rhythms in the metabolism of phospholipids and in the activity of lipid synthesizing enzymes.

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Gorné, Lucas D; Acosta-Rodríguez, Victoria A; Pasquaré, Susana J; Salvador, Gabriela A; Giusto, Norma M; Guido, Mario Eduardo

2015-02-01

The circadian system involves central and peripheral oscillators regulating temporally biochemical processes including lipid metabolism; their disruption leads to severe metabolic diseases (obesity, diabetes, etc). Here, we investigated the temporal regulation of glycerophospholipid (GPL) synthesis in mouse liver, a well-known peripheral oscillator. Mice were synchronized to a 12:12 h light-dark (LD) cycle and then released to constant darkness with food ad libitum. Livers collected at different times exhibited a daily rhythmicity in some individual GPL content with highest levels during the subjective day. The activity of GPL-synthesizing/remodeling enzymes: phosphatidate phosphohydrolase 1 (PAP-1/lipin) and lysophospholipid acyltransferases (LPLATs) also displayed significant variations, with higher levels during the subjective day and at dusk. We evaluated the temporal regulation of expression and activity of phosphatidylcholine (PC) synthesizing enzymes. PC is mainly synthesized through the Kennedy pathway with Choline Kinase (ChoK) as a key regulatory enzyme or through the phosphatidylethanolamine (PE) N-methyltransferase (PEMT) pathway. The PC/PE content ratio exhibited a daily variation with lowest levels at night, while ChoKα and PEMT mRNA expression displayed maximal levels at nocturnal phases. Our results demonstrate that mouse liver GPL metabolism oscillates rhythmically with a precise temporal control in the expression and/or activity of specific enzymes.

Molecular, cellular, and tissue impact of depleted uranium on xenobiotic-metabolizing enzymes.

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Gueguen, Yann; Rouas, Caroline; Monin, Audrey; Manens, Line; Stefani, Johanna; Delissen, Olivia; Grison, Stéphane; Dublineau, Isabelle

2014-02-01

Enzymes that metabolize xenobiotics (XME) are well recognized in experimental models as representative indicators of organ detoxification functions and of exposure to toxicants. As several in vivo studies have shown, uranium can alter XME in the rat liver or kidneys after either acute or chronic exposure. To determine how length or level of exposure affects these changes in XME, we continued our investigation of chronic rat exposure to depleted uranium (DU, uranyl nitrate). The first study examined the effect of duration (1-18 months) of chronic exposure to DU, the second evaluated dose dependence, from a level close to that found in the environment near mining sites (0.2 mg/L) to a supra-environmental dose (120 mg/L, 10 times the highest level naturally found in the environment), and the third was an in vitro assessment of whether DU exposure directly affects XME and, in particular, CYP3A. The experimental in vivo models used here demonstrated that CYP3A is the enzyme modified to the greatest extent: high gene expression changed after 6 and 9 months. The most substantial effects were observed in the liver of rats after 9 months of exposure to 120 mg/L of DU: CYP3A gene and protein expression and enzyme activity all decreased by more than 40 %. Nonetheless, no direct effect of DU by itself was observed after in vitro exposure of rat microsomal preparations, HepG2 cells, or human primary hepatocytes. Overall, these results probably indicate the occurrence of regulatory or adaptive mechanisms that could explain the indirect effect observed in vivo after chronic exposure.

Hepatoprotective effect of kaempferol against alcoholic liver injury in mice.

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Wang, Meng; Sun, Jianguo; Jiang, Zhihui; Xie, Wenyan; Zhang, Xiaoying

2015-01-01

Kaempferol is a biologically active component present in various plants. The hepatoprotective effect of kaempferol in drug-induced liver injury has been proven, while its effect against alcoholic liver injury (ALI) remains unclear. Hence, the present study aimed to evaluate the effect of kaempferol against ALI in mice. The experimental ALI mice model was developed and the mice were treated with different doses of kaempferol for 4 weeks. The liver functions were observed by monitoring the following parameters: Aspartate aminotransferase (AST/GOT) and alanine aminotransferase (ALT/GPT) levels in serum; histopathological studies of liver tissue; oxidative stress by hydrogen peroxide (H2O2), superoxide dismutase (SOD) and glutathione (GSH); the lipid peroxidation status by malondialdehyde (MDA) and lipid accumulation by triglyceride (TG) level in serum; and the expression levels and activities of a key microsomal enzyme cytochrome 2E1 (CYP2E1), by both in vitro and in vivo methods. The ALI mice (untreated) showed clear symptoms of liver injury, such as significantly increased levels of oxidative stress, lipid peroxidation and excessive CYP2E1 expression and activity. The mice treated with different kaempferol dosages exhibited a significant decrease in the oxidative stress as well as lipid peroxidation, and increased anti-oxidative defense activity. The kaempferol treatment has significantly reduced the expression level and activity of hepatic CYP2E1, thus indicating that kaempferol could down regulate CYP2E1. These findings show the hepatoprotective properties of kaempferol against alcohol-induced liver injury by attenuating the activity and expression of CYP2E1 and by enhancing the protective role of anti-oxidative defense system.

Detection on immunoblot of new proteins from the soluble fraction of the cell recognized either by anti-liver-kidney microsome antibodies type 1 or by anti-liver cytosol antibodies type 1--relationship with hepatitis C virus infection.

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Ballot, E; Desbos, A; Monier, J C

1996-09-01

Antibodies directed against liver cytosol protein, called anti-liver cytosol type 1 (LC1 Ab), have been described by both immunofluorescence (IF) and immunodiffusion techniques in sera from patients with autoimmune hepatitis (AIH). They have never been found in association with antibodies directed against the hepatitis C virus (HCV), unlike the anti-liver-kidney microsome antibodies type 1 (LKM1 Ab), the serological marker of AIH type 2. This suggests that there are two subgroups of AIH type 2, i.e., HCV-related and non-HCV-related. In this study, immunoblotting experiments were performed using proteins from the soluble phase of the rat liver cell; 141 sera which tested positive for LKM1 Ab by IF, 24 identified as having LC1 Ab by IF, and 50 from blood donors as controls were analyzed. Three bands were stained by LC1 Ab sera more often than by the control sera, and with a statistically significant frequency. These 3 proteins were located at apparent Mr 50,000, 55,000, and 60,000. The LKM1 Ab-positive sera as defined by IF stained six bands with a statistically significant frequency compared to the controls. Their apparent Mr were 35,000, 39,000, 47,000, 50,000, 55,000, and 60,000. LKM1 Ab-positive sera which were anti-HCV negative recognized a 60,000 protein belonging to the soluble phase of the cell, with a statistically significant frequency compared to LKM1 Ab-positive sera which were anti-HCV positive. This 60,000 protein was also recognized by LC1 Ab-positive sera, which were almost always anti-HCV negative. The presence of antibodies against a 60,000 protein from the soluble phase of the cell is discussed in terms of the anti-HCV serological markers found in the sera from patients with AIH.

Disruption of thyroid hormone homeostasis in Ugt1a-deficient Gunn rats by microsomal enzyme inducers is not due to enhanced thyroxine glucuronidation

International Nuclear Information System (INIS)

Richardson, Terrilyn A.; Klaassen, Curtis D.

2010-01-01

Microsomal enzyme inducers (MEI) that increase UDP-glucuronosyltransferases (UGTs) are thought to increase glucuronidation of thyroxine (T 4 ), thus reducing serum T 4 , and subsequently increasing thyroid stimulating hormone (TSH). Ugt1a1 and Ugt1a6 mediate T 4 glucuronidation. Therefore, this experiment determined the involvement of Ugt1a enzymes in increased T 4 glucuronidation, decreased serum T 4 , and increased TSH after MEI treatment. Male Wistar and Ugt1a-deficient Wistar (Gunn) rats were fed a control diet or diet containing pregnenolone-16α-carbonitrile (PCN; 800 ppm), 3-methylcholanthrene (3-MC; 200 ppm), or Aroclor 1254 (PCB; 100 ppm) for 7 days. Serum T 4 , triiodothyronine (T 3 ), and TSH concentrations, hepatic T 4 /T 3 glucuronidation, and thyroid histology and follicular cell proliferation were investigated. PCN, 3-MC, and PCB treatments decreased serum T 4 , whereas serum T 3 was maintained in both Gunn and Wistar rats (except for PCB treatment). TSH was increased in Wistar and Gunn rats after PCN (130 and 277%) or PCB treatment (72 and 60%). T 4 glucuronidation in Wistar rats was increased after PCN (298%), 3-MC (85%), and PCB (450%), but was extremely low in Gunn rats, and unchanged after MEI. T 3 glucuronidation was increased after PCN (121%) or PCB (58%) in Wistar rats, but only PCN increased T 3 glucuronidation in Gunn rats (43%). PCN treatment induced thyroid morphological changes and increased follicular cell proliferation in both strains. These data demonstrate that T 4 glucuronidation cannot be increased in Ugt1a-deficient Gunn rats. Thus, the decrease in serum T 4 , increase in TSH, and increase in thyroid cell proliferation after MEI are not dependent on increased T 4 glucuronidation, and cannot be attributed to Ugt1a enzymes.

Activities of acyl-CoA:diacylglycerol acyltransferase (DGAT) and phospholipid:diacylglycerol acyltransferase (PDAT) in microsomal preparations of developing sunflower and safflower seeds.

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Banaś, Walentyna; Sanchez Garcia, Alicia; Banaś, Antoni; Stymne, Sten

2013-06-01

The last step in triacylglycerols (TAG) biosynthesis in oil seeds, the acylation of diacylglycerols (DAG), is catalysed by two types of enzymes: the acyl-CoA:diacylglycerol acyltransferase (DGAT) and phospholipid:diacylglycerol acyltransferase (PDAT). The relative contribution of these enzymes in the synthesis of TAG has not yet been defined in any plant tissue. In the presented work, microsomal preparations were obtained from sunflower and safflower seeds at different stages of development and used in DGAT and PDAT enzyme assays. The ratio between PDAT and DGAT activity differed dramatically between the two different species. DGAT activities were measured with two different acyl acceptors and assay methods using two different acyl-CoAs, and in all cases the ratio of PDAT to DGAT activity was significantly higher in safflower than sunflower. The sunflower DGAT, measured by both methods, showed significant higher activity with 18:2-CoA than with 18:1-CoA, whereas the opposite specificity was seen with the safflower enzyme. The specificities of PDAT on the other hand, were similar in both species with 18:2-phosphatidylcholine being a better acyl donor than 18:1-PC and with acyl groups at the sn-2 position utilised about fourfold the rate of the sn-1 position. No DAG:DAG transacylase activity could be detected in the microsomal preparations.

Identification of UGT2B9*2 and UGT2B33 isolated from female rhesus monkey liver.

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Dean, Brian; Arison, Byron; Chang, Steve; Thomas, Paul E; King, Christopher

2004-06-01

Two UDP-glucuronosyltransferases (UGT2B9(*)2 and UGT2B33) have been isolated from female rhesus monkey liver. Microsomal preparations of the cell lines expressing the UGTs catalyzed the glucuronidation of the general substrate 7-hydroxy-4-(trifluoromethyl)coumarin in addition to selected estrogens (beta-estradiol and estriol) and opioids (morphine, naloxone, and naltrexone). UGT2B9(*)2 displayed highest efficiency for beta-estradiol-17-glucuronide production and did not catalyze the glucuronidation of naltrexone. UGT2B33 displayed highest efficiency for estriol and did not catalyze the glucuronidation of beta-estradiol. UGT2B9(*)2 was found also to catalyze the glucuronidation of 4-hydroxyestrone, 16-epiestriol, and hyodeoxycholic acid, while UGT2B33 was capable of conjugating 4-hydroxyestrone, androsterone, diclofenac, and hyodeoxycholic acid. Three glucocorticoids (cortisone, cortisol, and corticosterone) were not substrates for glucuronidation by liver or kidney microsomes or any expressed UGTs. Our current data suggest the use of beta-estradiol-3-glucuronidation, beta-estradiol-17-glucuronidation, and estriol-17-glucuronidation to assay UGT1A01, UGT2B9(*)2, and UGT2B33 activity in rhesus liver microsomes, respectively.

Metabolic Syndrome and Serum Liver Enzymes Level at Patients with Type 2 Diabetes Mellitus

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Music, Miralem; Dervisevic, Amela; Pepic, Esad; Lepara, Orhan; Fajkic, Almir; Ascic-Buturovic, Belma; Tuna, Enes

2015-01-01

Objectives: The aim of this study was to evaluate liver function in patients with type 2 diabetes mellitus (T2DM) with and without metabolic syndrome (MS) by determining serum levels of gamma glutamyltransferase (GGT), alanine aminotransferase (ALT) and aspartate aminotransferase (AST). We also investigated correlation between levels of liver enzymes and some components of MS in both groups of patients. Methods: This cross-sectional study included 96 patients (age 47–83 years) with T2DM. All patients were divided according to the criteria of the National Cholesterol Education Program (NCEP) in two groups: 50 patients with T2 DM and MS (T2DM-MS) and 46 patients with T2DM without MS (T2DM-Non MS). The analysis included blood pressure monitoring and laboratory tests: fasting blood glucose (FBG), total lipoprotein cholesterol (TC), high-density lipoprotein cholesterol (HDL-C), triglyceride (TG), fibrinogen and liver enzymes: GGT, ALT and AST. T2DM-MS group included patients which had FBG ≥ 6,1 mmol/L, TG ≥ 1,7 mmol/L and blood pressure ≥ 130/85 mm Hg. Results: T2DM-MS patients had significant higher values of systolic blood pressure, diastolic blood pressure and medium arterial pressure compared to T2DM-Non MS patients. Serum levels of TC, TG, LDL-C, VLDL-C and FBG were significantly higher in the T2DM-MS group compared to the T2DM-Non MS group. Serum fibrinogen level and GGT level were significantly higher in patients with T2DM-MS compared to the serum fibrinogen level and GGT level in T2DM-Non MS patients. Mean serum AST and ALT level were higher, but not significantly, in patients with T2DM and MS compared to the patients with T2DM without MS. Significant negative correlations were observed between TC and AST (r= -0,28, p<0,05), as well as between TC and ALT level (r= -0,29, p<0,05) in T2DM-MS group of patients. Conclusion: These results suggest that patients with T2DM and MS have markedly elevated liver enzymes. T2DM and MS probably play a role in

The effect of ghee (clarified butter) on serum lipid levels and microsomal lipid peroxidation.

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Sharma, Hari; Zhang, Xiaoying; Dwivedi, Chandradhar

2010-04-01

Ghee, also known as clarified butter, has been utilized for thousands of years in Ayurveda as a therapeutic agent. In ancient India, ghee was the preferred cooking oil. In the last several decades, ghee has been implicated in the increased prevalence of coronary artery disease (CAD) in Asian Indians due to its content of saturated fatty acids and cholesterol and, in heated ghee, cholesterol oxidation products. Our previous research on Sprague-Dawley outbred rats, which serve as a model for the general population, showed no effect of 5 and 10% ghee-supplemented diets on serum cholesterol and triglycerides. However, in Fischer inbred rats, which serve as a model for genetic predisposition to diseases, results of our previous research showed an increase in serum total cholesterol and triglyceride levels when fed a 10% ghee-supplemented diet. In the present study, we investigated the effect of 10% dietary ghee on microsomal lipid peroxidation, as well as serum lipid levels in Fischer inbred rats to assess the effect of ghee on free radical mediated processes that are implicated in many chronic diseases including cardiovascular disease. Results showed that 10% dietary ghee fed for 4 weeks did not have any significant effect on levels of serum total cholesterol, but did increase triglyceride levels in Fischer inbred rats. Ghee at a level of 10% in the diet did not increase liver microsomal lipid peroxidation or liver microsomal lipid peroxide levels. Animal studies have demonstrated many beneficial effects of ghee, including dose-dependent decreases in serum total cholesterol, low density lipoprotein (LDL), very low density lipoprotein (VLDL), and triglycerides; decreased liver total cholesterol, triglycerides, and cholesterol esters; and a lower level of nonenzymatic-induced lipid peroxidation in liver homogenate. Similar results were seen with heated (oxidized) ghee which contains cholesterol oxidation products. A preliminary clinical study showed that high doses of

Evaluation of the effects of hydroalcoholic extract of Berberis vulgaris root on the activity of liver enzymes in male hypercholesterolemic rats

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Soheila Taheri

2012-06-01

Conclusion: Noticing the antioxidant properties of B. vulgaris root extract  and its effects on reducing the activity of liver enzymes, the extract of this plant can be a good choice for improving the function of liver.

Metabolism of fatty acids in rat brain in microsomal membranes

International Nuclear Information System (INIS)

Aeberhard, E.E.; Gan-Elepano, M.; Mead, J.F.

1980-01-01

Using a technique in which substrate fatty acids are incorporated into microsomal membranes followd by comparison of their rates of desaturation or elongation with those of exogenous added fatty acids it has been found that the desaturation rate is more rapid for the membrane-bound substrate than for the added fatty acid. Moreover, the product of the membrane-bound substrate is incorporated into membrane phospholipid whereas the product of the exogenous substrate is found in di- and triacyl glycerols and in free fatty acids as well. These and other findings point to a normal sequence of reaction of membrane liqids with membrane-bound substrates involving transfer of fatty acid from phospholipid to the coupled enzyme systems without ready equilibration with the free fatty acid pool

Serum activities of liver enzymes in workers exposed to sub-TLV levels of dimethylformamide

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Jinjiang He

2015-04-01

Full Text Available Objectives: The aim of this study has been to investigate serum activities of liver enzymes in workers exposed to sub-TLV levels of dimethylformamide (DMF. Material and Methods: Seventy-two workers and 72 healthy controls participated in the study. All subjects underwent complete physical examinations and abdominal ultrasound examination. Serum aspartate aminotransferase (AST, alanine aminotransferase (ALT, and c-glutamyl transpeptidase (c-GT were determined by an auto-chemistry analyzer. The data of airborne concentrations of DMF was obtained from the local Center of Disease Control and Prevention. The level of urine N-acetyl-S-(N-methylcarbamoylcysteine (AMCC was measured by means of high-performance liquid chromatography. Results: Time weighted average (TWA concentration of the DMF in workplace was 18.6 (range: 9.8–36.2 mg/m3. The concentration of the AMCC in workers’ urine was 28.32 (range: 1.8–58.6 mg/l and 9 workers’ AMCC exceeded the biological exposure index (40 mg/l. Thirty-one workers reported gastrointestinal symptoms (abdominal pain, nausea, anorexia and 10 workers reported headache, dizziness and/or palpitation in the exposed group. Serum analysis revealed that both the mean of serum activities of liver enzymes (ALT, AST and c-GT and the percentage of workers with abnormal liver function were significantly higher in the exposed group as compared to the controls. Conclusions: Dimethylformamide can cause liver damage even if air concentration is in the sub-threshold limit value (sub-TLV level. The protection of skin contact against the exposure to the DMF might be a critical issue as far as the occupational health is concerned.

Serum levels of angiotensin converting enzyme as a biomarker of liver fibrosis

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Miranda, Aline Silva; Simões e Silva, Ana Cristina

2017-01-01

The renin angiotensin system (RAS) is classically conceived as a circulating hormonal system involved in blood pressure control and hydroelectrolyte balance. The discovery that RAS components are locally expressed in a wide range of organs and tissues, including the liver, pointed to a role for this system in the pathogenesis of several conditions including hepatic fibrosis and cirrhosis. It has been widely reported that the classical RAS axis composed by the angiotensin converting enzyme (ACE)-angiotensin (Ang) II-Ang type 1 (AT1) receptor mediates pro-inflammatory, pro-thrombotic, and pro-fibrotic processes. On the other hand, the alternative axis comprising ACE2-Ang-(1-7)-Mas receptor seems to play a protective role by frequently opposing Ang II action. Chronic hepatitis B (CHB) is one of the leading causes of liver fibrosis, accounting for the death of nearly one million people worldwide. Liver fibrosis is a key factor to determine therapeutic interventions for patients with CHB. However, the establishment of non-invasive and accurate methods to detect reversible stages of liver fibrosis is still a challenge. In an elegant study published in the 36th issue of the World Journal of Gastroenterology, Noguchi et al showed the predictive value of serum ACE levels in detecting not only advanced stages of liver fibrosis but also initial and intermediate fibrotic stages. The serum levels of ACE might represent an accurate, non-invasive, widely available, and easy method to evaluate fibrosis related to CHB. Moreover, therapies involving the inhibition of the classical RAS axis components might be promising in the control of CHB-related liver fibrosis. PMID:29358853

Oxidative metabolism of monensin in rat liver microsomes and interactions with tiamulin and other chemotherapeutic agents: evidence for the involvement of cytochrome P-450 3A subfamily.

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Nebbia, C; Ceppa, L; Dacasto, M; Carletti, M; Nachtmann, C

1999-09-01

Monensin (MON) is an ionophore antibiotic widely used in veterinary practice as a coccidiostatic or a growth promoter. The aims of this study were to characterize the P-450 isoenzyme(s) involved in the biotransformation of the ionophore and to investigate how this process may be affected by tiamulin and other chemotherapeutic agents known to produce toxic interactions with MON when administered concurrently in vivo. In liver microsomes from untreated rats (UT) or from rats pretreated, respectively, with ethanol (ETOH), beta-naphthoflavone (betaNAF), phenobarbital (PB), pregnenolone 16alpha-carbonitrile (PCN), or dexamethasone (DEX), the rate of MON O-demethylation was the following: DEX > PCN > PB > UT = ETOH > betaNAF; similar results were obtained by measuring total MON metabolism. In addition, the extent of triacetyloleandomycin-mediated P-450 complexes was greatly reduced by the prior addition of 100 microM MON. In DEX-treated microsomes, MON O-demethylation was found to fit monophasic Michaelis-Menten kinetics (K(M) = 67.6 +/- 0.01 microM; V(max) = 4.75 +/- 0.76 nmol/min/mg protein). Tiamulin markedly inhibited this activity in an apparent competitive manner, with a calculated K(i) (Dixon plot) of 8.2 microM and an IC(50) of about 25 microM. At the latter concentration, only ketoconazole or metyrapone, which can bind P-450 3A, inhibited MON O-demethylase to a greater extent than tiamulin, whereas alpha-naphthoflavone, chloramphenicol, or sulphametasine was less effective. These results suggest that P-450 3A plays an important role in the oxidative metabolism of MON and that compounds capable of binding or inhibiting this isoenzyme could be expected to give rise to toxic interactions with the ionophore.

Carbamoyl-phosphate synthase (ammonia) of rat and axolotl liver: determination of immunological cross-reactivity without purification of the axolotl enzyme

NARCIS (Netherlands)

Lamers, W. H.; de Graaf, A.; Mooren, P. G.; Moorman, A. F.; Charles, R.

1982-01-01

A method has been developed to establish the degree of cross-reactivity of an antiserum raised against purified carbamoyl-phosphate synthase (ammonia) from adult rat liver, toward a homologous enzyme from another species without purification of the latter enzyme. For that purpose the ratio between

Microsomal epoxide hydrolase gene polymorphisms and risk of chronic obstructive pulmonary disease: A comprehensive meta-analysis

OpenAIRE

LI, HUI; FU, WEI-PING; HONG, ZE-HUI

2012-01-01

Microsomal epoxide hydrolase (EPHX1) is an enzyme involved in the detoxification the products of smoking and is proposed to be a genetic factor for the development of chronic obstructive pulmonary disease (COPD). Two functional polymorphisms of EPHX1, T113C and A139G, have been analyzed in numerous studies to assess the COPD risk attributed to these variants. However, the conclusions were controversial. We performed a comprehensive meta-analysis to clarify these findings. A total of 24 studie...

Urinary Elimination of Bile Acid Glucuronides under Severe Cholestatic Situations: Contribution of Hepatic and Renal Glucuronidation Reactions

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Martin Perreault

2018-01-01

Full Text Available Biliary obstruction, a severe cholestatic complication, causes accumulation of toxic bile acids (BAs in liver cells. Glucuronidation, catalyzed by UDP-glucuronosyltransferase (UGT enzymes, detoxifies cholestatic BAs. Using liquid chromatography coupled to tandem mass spectrometry, 11 BA glucuronide (-G species were quantified in prebiliary and postbiliary stenting serum and urine samples from 17 patients with biliary obstruction. Stenting caused glucuronide- and fluid-specific changes in BA-G levels and BA-G/BA metabolic ratios. In vitro glucuronidation assays with human liver and kidney microsomes revealed that even if renal enzymes generally displayed lower KM values, the two tissues shared similar glucuronidation capacities for BAs. By contrast, major differences between the two tissues were observed when four human BA-conjugating UGTs 1A3, 1A4, 2B4, and 2B7 were analyzed for mRNA and protein levels. Notably, the BA-24G producing UGT1A3 enzyme, abundant in the liver, was not detected in kidney microsomes. In conclusion, the circulating and urinary BA-G profiles are hugely impacted under severe cholestasis. The similar BA-glucuronidating abilities of hepatic and renal extracts suggest that both the liver and kidney may contribute to the urine BA-G pool.

The Effect of Symbiotic Supplementation on Liver Enzymes, C-reactive Protein and Ultrasound Findings in Patients with Non-alcoholic Fatty Liver Disease: A Clinical Trial.

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Asgharian, Atefe; Askari, Gholamreza; Esmailzade, Ahmad; Feizi, Awat; Mohammadi, Vida

2016-01-01

Regarding to the growing prevalence of nonalcoholic fatty liver disease (NAFLD), concentrating on various strategies to its prevention and management seems necessary. The aim of this study was to determine the effects of symbiotic on C-reactive protein (CRP), liver enzymes, and ultrasound findings in patients with NAFLD. Eighty NAFLD patients were enrolled in this randomized, double-blind, placebo-controlled clinical trial. Participants received symbiotic in form of a 500 mg capsule (containing seven species of probiotic bacteria and fructooligosaccharides) or a placebo capsule daily for 8 weeks. Ultrasound grading, CRP, and liver enzymes were evaluated at the baseline and the end of the study. In the symbiotic group, ultrasound grade decreased significantly compared to baseline (P symbiotic supplementation was not associated with changes in alanine aminotransferase (ALT) and aspartate transaminase (AST) levels. In the placebo group, there was no significant change in steatosis grade whereas ALT and AST levels were significantly increased (P = 0.002, P = 0.02, respectively). CRP values remained static in either group. Symbiotic supplementation improved steatosis in NAFLD patients and might be useful in the management of NAFLD or protective against its progression.

[Pharmacological analysis of the effect of natural double-helical nucleic acids on the detoxifying function of the liver].

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Masycheva, V I; Morozova, E N; Nadolinnaia, I G

1988-10-01

The effect of interferon inductors i.e. double stranded RNAs from S. cerevisiae and phage F6 on the liver detoxicating function was studied on noninbred albino mice. The liver detoxicating function was tested by duration of hexenal sleep. It was shown that intraperitoneal administration of the yeast and phage RNAs in doses of 1/5 LD50 for three times led to increasing of the narcotic sleep duration in the animals by 65 and 207 per cent, respectively. The effect was of the dose-dependent nature. The doses not inducing reliable inhibition of hexenal metabolism were equal to 1/10 LD50 for the yeast dsRNA and 1/27 LD50 for the phage dsRNA. The inhibitory effect of the dsRNAs was retained for 2-3 days after discontinuation of the drug use. When the dsRNAs were administered simultaneously with nembutal, an inductor of the liver microsomal enzymes, the dsRNAs eliminated its inducing effect. Simultaneous administration of alpha-tocopherol lowered the dsRNA effect on hexenal metabolism. The findings suggested that the dsRNA inhibitory effect on the liver detoxicating function was grounded on the mechanisms associated with inhibition of syntheses and activation of lipid peroxidation specific of the monooxygenase system under the action of the dsRNAs.

Role of cytochrome P450 IA2 in acetanilide 4-hydroxylation as determined with cDNA expression and monoclonal antibodies.

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Liu, G; Gelboin, H V; Myers, M J

1991-02-01

The role of P450 IA2 in the hydroxylation of acetanilide was examined using an inhibitory monoclonal antibody (MAb) 1-7-1 and vaccinia cDNA expression producing murine P450 IA1 (mIA1), murine P450 IA2 (mIA2), or human P450 IA2 (hIA2). Acetanilide hydroxylase (AcOH) activity was measured using an HPLC method with more than 500-fold greater sensitivity than previously described procedures. This method, which does not require the use of radioactive acetanilide, was achieved by optimizing both the gradient system and the amount of enzyme needed to achieve detection by uv light. MAb 1-7-1 inhibits up to 80% of the AcOH activity in both rat liver microsomes and cDNA expressed mouse and human P450 IA2. MAb 1-7-1, which recognizes both P450 IA1 and P450 IA2, completely inhibits the aryl hydrocarbon hydroxylase (AHH) activity of cDNA expressed in IA1. The inhibition of only 80% of the AHH activity present in MC liver microsomes by MAb 1-7-1 suggests that additional P450 forms are contributing to the overall AHH activity present in methylcholanthrene (MC)-liver microsomes as MAb 1-7-1 almost completely inhibits the AHH activity of expressed mIA1. Maximal inhibition of IA2 by 1-7-1 results in an 80% decrease in acetanilide hydroxylase activity in both liver microsomes and expressed mouse and human IA2. The capacity of MAb 1-7-1 to produce identical levels of inhibition of acetanilide hydroxylase activity in rat MC microsomes (80%) and in expressed mouse (81%) and human P450 IA2 (80%) strongly suggests that P450 IA2 is the major and perhaps the only enzyme responsible for the metabolism of acetanilide. These results demonstrate the complementary utility of monoclonal antibodies and cDNA expression for defining the contribution of specific P450 enzymes to the metabolism of a given substrate. This complementary approach allows for a more precise determination of the inhibitory capacity of MAb with respect to the metabolic capacity of the target P450.

Adenosinetriphosphate content and adenosinetriphosphatase activity in cell fractions of the liver and brain of chick embryos and birds treated with gamma-rays

International Nuclear Information System (INIS)

Todorov, B.

1977-01-01

Studies are conducted on the level of ADP and the adenosinetriphosphatase in nuclei, mitochondria, and microsomes taken from the brain and liver of singly gamma-irradiated (1000 rd) chick embryos and birds. As a result of the treatment the ADP content dropped, while the activity of ADP rose. These changes were more strongly expressed in the nuclei, than in the mitochondria, and to a lesser extent - in the microsomes. Twelve-day chick embryos showed more markedly expressed radiosensitivity than newly hatched chicks. This embryonal stage is characterized by intense growth, differentiation and metabolic processes in the liver, which substantiate not only the higher radiosensitivity of this age group but the more strongly expressed changes in the liver as compared with the brain. (author)

Dipeptidyl peptidase-4 greatly contributes to the hydrolysis of vildagliptin in human liver.

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Asakura, Mitsutoshi; Fujii, Hideaki; Atsuda, Koichiro; Itoh, Tomoo; Fujiwara, Ryoichi

2015-04-01

The major metabolic pathway of vildagliptin in mice, rats, dogs, and humans is hydrolysis at the cyano group to produce a carboxylic acid metabolite M20.7 (LAY151), whereas the major metabolic enzyme of vildagliptin has not been identified. In the present study, we determined the contribution rate of dipeptidyl peptidase-4 (DPP-4) to the hydrolysis of vildagliptin in the liver. We performed hydrolysis assay of the cyano group of vildagliptin using mouse, rat, and human liver samples. Additionally, DPP-4 activities in each liver sample were assessed by DPP-4 activity assay using the synthetic substrate H-glycyl-prolyl-7-amino-4-methylcoumarin (Gly-Pro-AMC). M20.7 formation rates in liver microsomes were higher than those in liver cytosol. M20.7 formation rate was significantly positively correlated with the DPP-4 activity using Gly-Pro-AMC in liver samples (r = 0.917, P vildagliptin hydrolysis in the liver. Additionally, we established stable single expression systems of human DPP-4 and its R623Q mutant, which is the nonsynonymous single-nucleotide polymorphism of human DPP-4, in human embryonic kidney 293 (HEK293) cells to investigate the effect of R623Q mutant on vildagliptin-hydrolyzing activity. M20.7 formation rate in HEK293 cells expressing human DPP-4 was significantly higher than that in control HEK293 cells. Interestingly, R623Q mutation resulted in a decrease of the vildagliptin-hydrolyzing activity. Our findings might be useful for the prediction of interindividual variability in vildagliptin pharmacokinetics. Copyright © 2015 by The American Society for Pharmacology and Experimental Therapeutics.

Effects of Curcuma xanthorrhiza Extracts and Their Constituents on Phase II Drug-metabolizing Enzymes Activity

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Salleh, Nurul Afifah Mohd; Ismail, Sabariah; Ab Halim, Mohd Rohaimi

2016-01-01

Background: Curcuma xanthorrhiza is a native Indonesian plant and traditionally utilized for a range of illness including liver damage, hypertension, diabetes, and cancer. Objective: The study determined the effects of C. xanthorrhiza extracts (ethanol and aqueous) and their constituents (curcumene and xanthorrhizol) on UDP-glucuronosyltransferase (UGT) and glutathione transferase (GST) activities. Materials and Methods: The inhibition studies were evaluated both in rat liver microsomes and in human recombinant UGT1A1 and UGT2B7 enzymes. p-nitrophenol and beetle luciferin were used as the probe substrates for UGT assay while 1-chloro-2,4-dinitrobenzene as the probe for GST assay. The concentrations of extracts studied ranged from 0.1 to 1000 μg/mL while for constituents ranged from 0.01 to 500 μM. Results: In rat liver microsomes, UGT activity was inhibited by the ethanol extract (IC50 =279.74 ± 16.33 μg/mL). Both UGT1A1 and UGT2B7 were inhibited by the ethanol and aqueous extracts with IC50 values ranging between 9.59–22.76 μg/mL and 110.71–526.65 μg/Ml, respectively. Rat liver GST and human GST Pi-1 were inhibited by ethanol and aqueous extracts, respectively (IC50 =255.00 ± 13.06 μg/mL and 580.80 ± 18.56 μg/mL). Xanthorrhizol was the better inhibitor of UGT1A1 (IC50 11.30 ± 0.27 μM) as compared to UGT2B7 while curcumene did not show any inhibition. For GST, both constituents did not show any inhibition. Conclusion: These findings suggest that C. xanthorrhiza have the potential to cause herb-drug interaction with drugs that are primarily metabolized by UGT and GST enzymes. SUMMARY Findings from this study would suggest which of Curcuma xanthorrhiza extracts and constituents that would have potential interactions with drugs which are highly metabolized by UGT and GST enzymes. Further clinical studies can then be designed if needed to evaluate the in vivo pharmacokinetic relevance of these interactions Abbreviations Used: BSA: Bovine serum albumin

Effects of whole body x-ray irradiation on induction by phenobarbital of rat liver glucose-6-phosphate dehydrogenase and glutathione reductase

Energy Technology Data Exchange (ETDEWEB)

Bitny-Szlachto, S.; Szyszko, A. (Wojskowy Inst. Higieny i Epidemiologii, Warsaw (Poland))

1979-01-01

In rats treated with phenobarbital (3x100 mg/kg, i.p.), liver G-6-P dehydrogenase activity increased by 70% in the cytosol and in the 9.000xg supernatant, and only by 20% in microsomes. Moreover, the phenobarbital treatment increased rat liver GSSG reductase activity by 30%. On the other hand, activity of the liver microsomal G-6-P dehydrogenase was found to increase by some 20% in whole body irradiated, both control and phenobarbital treated rats. In rats irradiated with 600 R prior to the first dose of the inducer there was not noted any increase in G-6-P dehydrogenase of the 9.000xg supernatant, and increase in the cytosol activity dropped to 38%. Thus, induction of the soluble liver G-6-P dehydrogenase by phenobarbital has turned out to be radiosensitive, whereas phenobarbital induction of GSSG reductase was unaffected by irradiation.

L-malate enhances the gene expression of carried proteins and antioxidant enzymes in liver of aged rats.

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Zeng, X; Wu, J; Wu, Q; Zhang, J

2015-01-01

Previous studies in our laboratory reported L-malate as a free radical scavenger in aged rats. To investigate the antioxidant mechanism of L-malate in the mitochondria, we analyzed the change in gene expression of two malate-aspartate shuttle (MAS)-related carried proteins (AGC, aspartate/glutamate carrier and OMC, oxoglutarate/malate carrier) in the inner mitochondrial membrane, and three antioxidant enzymes (CAT, SOD, and GSH-Px) in the mitochondria. The changes in gene expression of these proteins and enzymes were examined by real-time RT-PCR in the heart and liver of aged rats treated with L-malate. L-malate was orally administered in rats continuously for 30 days using a feeding atraumatic needle. We found that the gene expression of OMC and GSH-Px mRNA in the liver increased by 39 % and 38 %, respectively, in the 0.630 g/kg L-malate treatment group than that in the control group. The expression levels of SOD mRNA in the liver increased by 39 %, 56 %, and 78 % in the 0.105, 0.210, and 0.630 g/kg L-malate treatment groups, respectively. No difference were observed in the expression levels of AGC, OMC, CAT, SOD, and GSH-Px mRNAs in the heart of rats between the L-malate treatment and control groups. These results predicted that L-malate may increase the antioxidant capacity of mitochondria by enhancing the expression of mRNAs involved in the MAS and the antioxidant enzymes.

Lack of evidence for metabolism of p-phenylenediamine by human hepatic cytochrome P450 enzymes

International Nuclear Information System (INIS)

Stanley, Lesley A.; Skare, Julie A.; Doyle, Edward; Powrie, Robert; D'Angelo, Diane; Elcombe, Clifford R.

2005-01-01

p-Phenylenediamine (PPD) is a widely used ingredient in permanent hair dyes; however, little has been published on its metabolism, especially with respect to hepatic cytochrome P450 (CYP)-mediated oxidation. This is regarded as a key step in the activation of carcinogenic arylamines that ultimately leads to the development of bladder cancer. Most epidemiology studies show no significant association between personal use of hair dyes and bladder cancer, but one recent study reported an increased risk of bladder cancer in women who were frequent users of permanent hair dyes. The aim of the present study was to use intact human hepatocytes, human liver microsomes, and heterologously expressed human CYPs to determine whether PPD is metabolised by hepatic CYPs to form an N-hydroxylamine. p-Phenylenediamine was N-acetylated by human hepatocytes to form N-acetylated metabolites, but there was no evidence for the formation of mono-oxygenated metabolites or for enzyme-mediated covalent binding of 14 C-PPD to microsomal protein. In contrast, 2-aminofluorene underwent CYP-mediated metabolism to ≥4 different hydroxylated metabolites. The lack of evidence for hepatic CYP-mediated metabolism of PPD is inconsistent with the hypothesis that this compound plays a causal role in the development of bladder cancer via a mode of action involving hepatic metabolism to an N-hydroxyarylamine

Comparative liver accumulation of dioxin-like compounds in sheep and cattle: Possible role of AhR-mediated xenobiotic metabolizing enzymes.

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Girolami, F; Spalenza, V; Benedetto, A; Manzini, L; Badino, P; Abete, M C; Nebbia, C

2016-11-15

PCDDs, PCDFs, and PCBs are persistent organic pollutants (POPs) that accumulate in animal products and may pose serious health problems. Those able to bind the aryl hydrocarbon receptor (AhR), eliciting a plethora of toxic responses, are defined dioxin-like (DL) compounds, while the remainders are called non-DL (NDL). An EFSA opinion has highlighted the tendency of ovine liver to specifically accumulate DL-compounds to a greater extent than any other farmed ruminant species. To examine the possible role in such an accumulation of xenobiotic metabolizing enzymes (XME) involved in DL-compound biotransformation, liver samples were collected from ewes and cows reared in an area known for low dioxin contamination. A related paper reported that sheep livers had about 5-fold higher DL-compound concentrations than cattle livers, while the content of the six marker NDL-PCBs did not differ between species. Specimens from the same animals were subjected to gene expression analysis for AhR, AhR nuclear translocator (ARNT) and AhR-dependent oxidative and conjugative pathways; XME protein expression and activities were also investigated. Both AhR and ARNT mRNA levels were about 2-fold lower in ovine samples and the same occurred for CYP1A1 and CYP1A2, being approximately 3- and 9-fold less expressed in sheep compared to cattle, while CYP1B1 could be detectable in cattle only. The results of the immunoblotting and catalytic activity (most notably EROD) measurements of the CYP1A family enzymes were in line with the gene expression data. By contrast, phase II enzyme expression and activities in sheep were higher (UGT1A) or similar (GSTA1, NQO1) to those recorded in cattle. The overall low expression of CYP1 family enzymes in the sheep is in line with the observed liver accumulation of DL-compounds and is expected to affect the kinetics and the dynamics of other POPs such as many polycyclic aromatic hydrocarbons, as well as of toxins (e.g. aflatoxins) or drugs (e.g. benzimidazole

Shotgun proteomics of Aspergillus niger microsomes upon D-xylose induction.

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Ferreira de Oliveira, José Miguel P; van Passel, Mark W J; Schaap, Peter J; de Graaff, Leo H

2010-07-01

Protein secretion plays an eminent role in cell maintenance and adaptation to the extracellular environment of microorganisms. Although protein secretion is an extremely efficient process in filamentous fungi, the mechanisms underlying protein secretion have remained largely uncharacterized in these organisms. In this study, we analyzed the effects of the d-xylose induction of cellulase and hemicellulase enzyme secretion on the protein composition of secretory organelles in Aspergillus niger. We aimed to systematically identify the components involved in the secretion of these enzymes via mass spectrometry of enriched subcellular microsomal fractions. Under each condition, fractions enriched for secretory organelles were processed for tandem mass spectrometry, resulting in the identification of peptides that originate from 1,081 proteins, 254 of which-many of them hypothetical proteins-were predicted to play direct roles in the secretory pathway. d-Xylose induction led to an increase in specific small GTPases known to be associated with polarized growth, exocytosis, and endocytosis. Moreover, the endoplasmic-reticulum-associated degradation (ERAD) components Cdc48 and all 14 of the 20S proteasomal subunits were recruited to the secretory organelles. In conclusion, induction of extracellular enzymes results in specific changes in the secretory subproteome of A. niger, and the most prominent change found in this study was the recruitment of the 20S proteasomal subunits to the secretory organelles.

Protective Effect of Prosopis cineraria Against N-Nitrosodiethylamine Induced Liver Tumor by Modulating Membrane Bound Enzymes and Glycoproteins

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Naina Mohamed Pakkir Maideen

2012-06-01

Full Text Available Purpose: The objective of the present study was to evaluate the protective effect of methanol extract of Prosopis cineraria (MPC against N-nitrosodiethylamine (DEN, 200mg/kg induced Phenobarbital promoted experimental liver tumors in male Wistar rats. Methods: The rats were divided into four groups, each group consisting of six animals. Group 1 served as control animals. Liver tumor was induced in group 2, 3, and 4 and Group 3 animals received MPC 200mg/kg and Group 4 animals received MPC 400mg/kg. Results: Administration of DEN has brought down the levels of membrane bound enzymes like Na+/ K+ ATPase, Mg2+ ATPase and Ca2+ATPase which were later found to be increased by the administration of Prosopis cineraria (200 and 400mg/kg in dose dependent manner. The MPC extract also suppressed the levels of glycoproteins like Hexose, Hexosamine and Sialic acid when compared to liver tumor bearing animals. Conclusions: Our study suggests that MPC may extend its protective role by modulating the levels of membrane bound enzymes and suppressing glycoprotein levels.

Porcine foetal and neonatal CYP3A liver expression

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Marie Louise Hiort Hermann

2011-05-01

Full Text Available Human cytochrome P450 3A7 (CYP3A7 and cytochrome P450 3A4 (CYP3A4 are hepatic metabolising enzymes which participates in the biotransformation of endo- and exogenous substances in foetuses and neonates respectively. These CYP3A enzymes display an inverse relationship: CYP3A7 is the dominant enzyme in the foetal liver, whereas the expression of CYP3A4 is low. After parturition there is a shift in the expression, thus CYP3A7 is down regulated, while the level of CYP3A4 gradually increases and becomes the dominant metabolising CYP3A enzyme in the adult. The minipig is increasingly being used as a model for humans in biomedical studies, because of its many similarities with the human physiology and anatomy. The aim of this study was to examine whether, as in humans, a shift is seen in the hepatic expression of a CYP3A7- like enzyme to cytochrome P450 3A29 (CYP3A29 (an orthologue to the human CYP3A4 in minipigs. This was elucidated by examining the hepatic mRNA expression of CYP3A7 and CYP3A29 in 39 foetuses and newborn Göttingen minipigs using quantitative real time polymerase chain reaction (qPCR. Furthermore the immunochemical level of CYP3A7-LE and CYP3A29 was measured in liver microsomes using western blotting. The expression of CYP3A29 was approximately 9- fold greater in neonates compared to foetuses, and a similar difference was reflected on the immunochemical level. It was not possible to detect a significant level of foetal CYP3A7 mRNA, but immunoblotting showed a visible difference depending on age. This study demonstrates an increase in the expression of CYP3A29, the CYP3A4 orthologue in perinatal minipigs as in humans, which suggests that the minipig could be a good model when testing for human foetal toxicity towards CYP3A4 substrates.

Positional specificity of saturated and unsaturated fatty acids in phosphatidic acid from rat liver

NARCIS (Netherlands)

Possmayer, F.; Scherphof, G.L.; Dubbelman, T.M.A.R.; Golde, L.M.G. van; Deenen, L.L.M. van

1969-01-01

1. 1. The relative incorporation of a number of radioactive fatty acids into the different glycerolipids of rat liver microsomes has been investigated. 2. 2. Studies on the distribution of the radioactivity incorporated into phosphatidylcholine, phosphatidylethanolamine and phosphatidic acid

Species Differences in the Oxidative Desulfurization of a Thiouracil-Based Irreversible Myeloperoxidase Inactivator by Flavin-Containing Monooxygenase Enzymes.

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Eng, Heather; Sharma, Raman; Wolford, Angela; Di, Li; Ruggeri, Roger B; Buckbinder, Leonard; Conn, Edward L; Dalvie, Deepak K; Kalgutkar, Amit S

2016-08-01

N1-Substituted-6-arylthiouracils, represented by compound 1 [6-(2,4-dimethoxyphenyl)-1-(2-hydroxyethyl)-2-thioxo-2,3-dihydropyrimidin-4(1H)-one], are a novel class of selective irreversible inhibitors of human myeloperoxidase. The present account is a summary of our in vitro studies on the facile oxidative desulfurization in compound 1 to a cyclic ether metabolite M1 [5-(2,4-dimethoxyphenyl)-2,3-dihydro-7H-oxazolo[3,2-a]pyrimidin-7-one] in NADPH-supplemented rats (t1/2 [half-life = mean ± S.D.] = 8.6 ± 0.4 minutes) and dog liver microsomes (t1/2 = 11.2 ± 0.4 minutes), but not in human liver microsomes (t1/2 > 120 minutes). The in vitro metabolic instability also manifested in moderate-to-high plasma clearances of the parent compound in rats and dogs with significant concentrations of M1 detected in circulation. Mild heat deactivation of liver microsomes or coincubation with the flavin-containing monooxygenase (FMO) inhibitor imipramine significantly diminished M1 formation. In contrast, oxidative metabolism of compound 1 to M1 was not inhibited by the pan cytochrome P450 inactivator 1-aminobenzotriazole. Incubations with recombinant FMO isoforms (FMO1, FMO3, and FMO5) revealed that FMO1 principally catalyzed the conversion of compound 1 to M1. FMO1 is not expressed in adult human liver, which rationalizes the species difference in oxidative desulfurization. Oxidation by FMO1 followed Michaelis-Menten kinetics with Michaelis-Menten constant, maximum rate of oxidative desulfurization, and intrinsic clearance values of 209 μM, 20.4 nmol/min/mg protein, and 82.7 μl/min/mg protein, respectively. Addition of excess glutathione essentially eliminated the conversion of compound 1 to M1 in NADPH-supplemented rat and dog liver microsomes, which suggests that the initial FMO1-mediated S-oxygenation of compound 1 yields a sulfenic acid intermediate capable of redox cycling to the parent compound in a glutathione-dependent fashion or undergoing further oxidation to a more

Formation of glutathione conjugates by reactive metabolites of vinylidene chloride in microsomes and isolated hepatocytes

International Nuclear Information System (INIS)

Liebler, D.C.; Meredith, M.J.; Guengerich, F.P.

1985-01-01

Oxidation of the vinyl halide carcinogen and hepatotoxin vinylidene chloride (VDC) by microsomal cytochrome P-450 yields 2,2-dichloroacetaldehyde, 2-chloroacetyl chloride, 2-chloroacetic acid, and 1,1-dichloroethylene oxide. The roles of these metabolites in covalent modification of proteins and reduced glutathione (GSH) were examined. 2-Chloroacetyl chloride reacted with model thiols at least 10(3)-fold faster than did 1,1-dichloroethylene oxide and at least 10(5)-fold faster than did 2,2-dichloroacetaldehyde or 2-chloroacetic acid. Microsomal covalent binding of [ 14 C]VDC was inhibited by GSH but not by lysine, suggesting that protein thiols, rather than amino groups, are major targets. Liver microsomes catalyzed the formation of three GSH:VDC metabolite conjugates, identified as S-(2,2-dichloro-1-hydroxy)ethylglutathione, 2-(S-glutathionyl)acetate, and S-(2-glutathionyl)acetylglutathione, a novel conjugate containing both stable (thioether) and labile (thioester) linkages. The latter two conjugates also were formed in isolated rat hepatocytes and measurable amounts of 2-(S-glutathionyl)acetate were released into the incubation medium. Both 2-(S-glutathionyl)acetate and S-(2-glutathionyl)acetylglutathione were formed with [ 35 S]GSH added to the hepatic medium, indicating that reactive VDC metabolites are capable of crossing the plasma membrane to react with extracellular targets. Unlabeled S-(2-glutathionyl)-acetylglutathione underwent carbonyl substitution with added [ 35 S]GSH, suggesting that this conjugate may participate in modification of protein thiols. This conjugate also underwent hydrolysis with a half-life of approximately 3 hr. GSH:VDC metabolite conjugates may serve as accessible models for labile covalent adducts formed between VDC metabolites and protein thiols

Evaluation of liver enzyme levels in workers exposed to vinyl chloride vapors in a petrochemical complex: a cross-sectional study

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Dolati Mandana

2007-08-01

Full Text Available Abstract Background Polyvinyl chloride is used in production and manufacturing of many essential tools (e.g. plastic pipes, photography films, etc.. Its production is impossible without the use of vinyl chloride monomer (VCM, which can cause liver damage in long-term. In this study we intend to assess the effects of mild to moderate long term exposure to VCM on liver and to assess the importance of liver enzyme measurements as a screening tool. Methods In this study, liver enzyme levels of 52 workers were compared to 48 control workers using the T-test. The cases all worked in a PVC production unit in a petrochemical complex and the controls were randomly selected from office personnel of the same complex. A questionnaire was also filled in about information such as age, weight, work history, etc. in both groups. Results Mean comparisons for ALP and GGT using T-test showed statistically significant differences between the two groups. For AST, ALT and bilirubin (total, direct the mean was higher in the case group but this difference was not statistically significant. Discussion This study showed that mild exposure to VCM can cause mild liver cholestasis. So, using cholestasis assessment tests such as ALP and GGT should be considered in periodic assessment of liver function in PVC producing units.

Reductive metabolism of oxymatrine is catalyzed by microsomal CYP3A4

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Liu W

2015-10-01

Full Text Available Wenqin Liu,1,2,* Jian Shi,1,2,* Lijun Zhu,2 Lingna Dong,1 Feifei Luo,2 Min Zhao,2 Ying Wang,2 Ming Hu,2,3 Linlin Lu,2 Zhongqiu Liu1,2 1Department of Pharmaceutics, School of Pharmaceutical Sciences, Southern Medical University, Guangzhou, Guangdong, People’s Republic of China; 2International Institute for Translational Chinese Medicine, Guangzhou University of Chinese Medicine, Guangzhou, Guangdong, People’s Republic of China; 3Department of Pharmacological and Pharmaceutical Sciences, College of Pharmacy, University of Houston, Houston, TX, USA *These authors contributed equally to this work Abstract: Oxymatrine (OMT is a pharmacologically active primary quinolizidine alkaloid with various beneficial and toxic effects. It is confirmed that, after oral administration, OMT could be transformed to the more toxic metabolite matrine (MT, and this process may be through the reduction reaction, but the study on the characteristics of this transformation is limited. The aim of this study was to investigate the characteristics of this transformation of OMT in the human liver microsomes (HLMs and human intestinal microsomes (HIMs and the cytochrome P450 (CYP isoforms involved in this transformation. The current studies demonstrated that OMT could be metabolized to MT rapidly in HLMs and HIMs and CYP3A4 greatly contributed to this transformation. All HLMs, HIMs, and CYP3A4 isoform mediated reduction reaction followed typical biphasic kinetic model, and Km, Vmax, and CL were significant higher in HLMs than those in HIMs. Importantly, different oxygen contents could significantly affect the metabolism of OMT, and with the oxygen content decreased, the formation of metabolite was increased, suggesting this transformation was very likely a reduction reaction. Results of this in vitro study elucidated the metabolic pathways and characteristics of metabolism of OMT to MT and would provide a theoretical basis and guidance for the safe application of OMT

Polyphenols of Salix aegyptiaca modulate the activities of drug metabolizing and antioxidant enzymes, and level of lipid peroxidation.

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Nauman, Mohd; Kale, R K; Singh, Rana P

2018-03-07

Salix aegyptiaca is known for its medicinal properties mainly due to the presence of salicylate compounds. However, it also contains other beneficial phytochemicals such as gallic acid, quercetin, rutin and vanillin. The aim of the study was to examine the redox potential, antioxidant and anti-inflammatory activity of these phytochemicals along with acetylsalicylic acid. The redox potential and antioxidant activity of gallic acid, quercetin, rutin, vanillin and acetylsalicylic acid were determined by oxidation-reduction potential electrode method and 1,1-diphenyl-2-picrylhydrazyl (DPPH) assay, respectively. In ex vivo studies, antioxidant activity of these phytochemicals was determined by lipid peroxidation and carbonyl content assay in the liver of mice. Anti-inflammatory activity was determined by protein denaturation method. Six-week old C57BL/6 mice treated with gallic acid (100 mg/kg body weight) and acetylsalicylic acid (25 and 50 mg/kg body weight) to investigate their in vivo modulatory effects on the specific activities of drug metabolizing phase I and phase II enzymes, antioxidant enzymes and level of lipid peroxidation in liver. The order of ability to donate electron and antioxidant activity was found to be: gallic acid > quercetin > rutin > vanillin > acetylsalicylic acid. In ex vivo studies, the similar pattern and magnitude of inhibitory effects of these phytochemicals against peroxidative damage in microsomes and protein carbonyl in cytosolic fraction were observed. In in vivo studies, gallic acid and acetylsalicylic acid alone or in combination, enhanced the specific activities of drug metabolizing phase I and phase II enzymes as well as antioxidant enzymes and also inhibited lipid peroxidation in liver. These findings show a close link between the electron donation and antioxidation potential of these phytochemicals, and in turn their biological activity. Gallic acid, quercetin, rutin and vanillin were found to be better electron donors and

Effects of coenzyme Q10 supplementation on the anthropometric variables, lipid profiles and liver enzymes in patients with non-alcoholic fatty liver disease

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Elnaz Jafarvand

2016-03-01

Full Text Available This randomized double-blind placebo-controlled trial was conducted on 41 patients with non-alcoholic fatty liver disease. Patients in intervention group received 100 mg/day coenzyme Q10 (CoQ10 for four weeks. There was a significant reduction in waist circumference and aspartate aminotransferase concentrations after CoQ10 supplementation (p<0.05. Dietary fiber was in negative correlation with change in serum alanine aminotransferase (ALT concentrations (r = -410, p = 0.04, and dietary fat intake was in positive relation with serum triglyceride (r = 463, p = 0.04 and in negative relation with serum high-density lipoprotein cholesterol (HDL-C (r = -533, p = 0.02 in CoQ10-treated group. CoQ10 supplement is able to reduce central obesity and improve liver function in non-alcoholic fatty liver disease. Dietary factors were also significant determinants of change in liver-specific enzyme ALT and lipid profile in these patients. Further trials with higher dose of CoQ10 and longer treatment periods are warranted to better clarify these findings.

Autoimmune liver disease 2007.

Science.gov (United States)

Muratori, Paolo; Granito, Alessandro; Pappas, Georgios; Muratori, Luigi; Lenzi, Marco; Bianchi, Francesco B

2008-01-01

Autoimmune liver disease (ALD) includes a spectrum of diseases which comprises both cholestatic and hepatitic forms: autoimmune hepatitis (AIH), primary biliary cirrhosis (PBC), primary sclerosing cholangitis (PSC) and the so called "overlap" syndromes where hepatitic and cholestatic damage coexists. All these diseases are characterized by an extremely high heterogeneity of presentation, varying from asymptomatic, acute (as in a subset of AIH) or chronic (with aspecific symptoms such as fatigue and myalgia in AIH or fatigue and pruritus in PBC and PSC). The detection and characterization of non organ specific autoantibodies plays a major role in the diagnostic approach of autoimmune liver disease; anti nuclear reactivities (ANA) and anti smooth muscle antibodies (SMA) mark type 1 AIH, liver kidney microsomal antibody type 1 (LKM1) and liver cytosol type 1 (LC1) are the serological markers of type 2 AIH; antimitochondrial antibodies (AMA) are associated with PBC, while no specific marker is found in PSC, since anticytoplasmic neutrophil antibodies with perinuclear pattern (atypical p-ANCA or p-ANNA) are also detected in a substantial proportion of type 1 AIH cases. Treatment options rely on immunosoppressive therapy (steroids and azathioprine) in AIH and on ursodeoxycholic acid in cholestatic conditions; in all these diseases liver transplantation remains the only therapeutical approach for the end stage of liver disease.

Functioning of Microsomal Cytochrome P450s: Murburn Concept Explains the Metabolism of Xenobiotics in Hepatocytes.

Science.gov (United States)

Manoj, Kelath Murali; Parashar, Abhinav; Gade, Sudeep K; Venkatachalam, Avanthika

2016-01-01

murburn as the operative concept. The mechanism of uncoupling (peroxide/water formation) was found to be dependent on multiple one and two electron equilibriums amongst the reaction components. The investigation explains the evolutionary implications of xenobiotic metabolism, confirms the obligatory role of diffusible reactive species in routine redox metabolism within liver microsomes and establishes that a redox enzyme like CYP enhances reaction rates (achieves catalysis) via a novel (hitherto unknown) modality.

Vitamin B2 content determination in liver paste by using acid and acid-enzyme hydrolysis

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Basić Zorica

2007-01-01

the samples (r = 0.9994, and r = 0.99987. Hydrolysis procedures make a sample suitable for vitamin B2 determination. In the liver paste samples a high content of vitamin B2 was determined: 0.83 mg/100 g after acid hydrolysis, and 0.909 mg/100 g after acid-enzyme hydrolysis. There were statistically significantly higher values determined after the acid-enzyme hydrolysis (p < 0.05. Conclusion. Using acid-enzyme hydrolysis and separation instrument technique (liquid chromatography with a fluorescent detector as detection system, statistically significantly greater vitamin B2 quantities were determined than after using acid hydrolysis procedure. Vitamin B2 content determined in ten liver paste samples was high (0.881 − 0.936 mg/100g indicating that this meat product is a good vitamin B2 source.

Hydrioxylation of sesquiterpenes by enzymes from chicory (Cichorium intybus L.) roots

NARCIS (Netherlands)

Kraker, de J.W.; Schurink, M.; Franssen, M.C.R.; König, W.A.; Groot, de Æ.; Bouwmeester, H.J.

2003-01-01

A microsomal enzyme preparation of chicory roots catalyses the hydroxylation of various sesquiterpene olefins in the presence of NADPH. Most of these hydroxylations take place at an isopropenyl or isopropylidene group. The number of products obtained from any of the substrates is confined to one or,

Failure of Chemotherapy in Hepatocellular Carcinoma Due to Impaired and Dysregulated Primary Liver Drug Metabolizing Enzymes and Drug Transport Proteins: What to Do?

Science.gov (United States)

Ul Islam, Salman; Ahmed, Muhammad Bilal; Shehzad, Adeeb; Ul-Islam, Mazhar; Lee, Young Sup

2018-05-28

Most of the drugs are metabolized in the liver by the action of drug metabolizing enzymes. In hepatocellular carcinoma (HCC), primary drug metabolizing enzymes are severely dysregulated, leading to failure of chemotherapy. Sorafenib is the only standard systemic drug available, but it still presents certain limitations, and much effort is required to understand who is responsive and who is refractory to the drug. Preventive and therapeutic approaches other than systemic chemotherapy include vaccination, chemoprevention, liver transplantation, surgical resection, and locoregional therapies. This review details the dysregulation of primary drug metabolizing enzymes and drug transport proteins of the liver in HCC and their influence on chemotherapeutic drugs. Furthermore, it emphasizes the adoption of safe alternative therapeutic strategies to chemotherapy. The future of HCC treatment should emphasize the understanding of resistance mechanisms and the finding of novel, safe, and efficacious therapeutic strategies, which will surely benefit patients affected by advanced HCC. Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.

Effects of Cinnamon extract on biochemical enzymes, TNF-α and NF-κB gene expression levels in liver of broiler chickens inoculated with Escherichia coli

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Seyed Mahmoud Tabatabaei

2015-09-01

Full Text Available Abstract: Infection with Escherichia coli (E. coli is a common disease in poultry industry. The use of antibiotics to treat diseases is facing serious criticism and concerns. The medicinal plants may be effective alternatives because of their multiplex activities. The aim of this study was to investigate the effects of cinnamon extract on the levels of liver enzymes, tumor necrosis factor-alpha (TNF-α and nuclear factor-kappa B (NF-κB gene expressions in liver of broiler chickens infected with E. coli. Ninety Ross-308 broilers were divided into healthy or E. coli-infected groups, receiving normal or cinnamon extract (in concentrations of 100 or 200mg/kg of food supplemented diets. E. coli suspension (108cfu was injected subcutaneously after 12 days cinnamon administration. Seventy-two hours after E. coli injection, the blood samples were taken for biochemical analysis of liver enzymes in serum (spectrophotometrically, and liver tissue samples were obtained for detection of gene expression of inflammatory markers TNF-α and NF-κB, using real-time PCR. Infection with E. coli significantly increased the levels of TNF-α and NF-κB gene expressions as well as some liver enzymes including creatine-kinase (CK, lactate-dehydrogenase (LDH, alanine-transferase (ALT and aspartate-transferase (AST as compared with control group (P<0.05. Pre-administration of cinnamon extract in broilers diet (in both concentrations significantly reduced the tissue levels of TNF-α and NF-κB gene expressions and enzymes CK and ALT in serum of broiler chickens inoculated with E. coli in comparison with E. coli group (P<0.05 and P<0.01. The levels of LDH and AST were significantly decreased only by 200mg/kg cinnamon extract in infected broilers. The level of alkaline-phosphatase (ALP was not affected in any groups. Pre-administration of cinnamon extract in diets of broiler chickens inoculated with E. coli could significantly reduce the gene expression levels of pro

[The effect of berberine administration of evaluation of the functional state of rat liver after ligation of common bile duct].

Science.gov (United States)

Zverinskiĭ, I V; Mel'nichenko, N G; Poplavskiĭ, V A; Sut'ko, I P; Telegin, P G; Shliakhtun, A G

2013-01-01

On the eighth day after ligation of the common bile duct in rats a significant increase in the serum content of total lipids, cholesterol bilirubin and ALT, alkaline phosphatase, and gamma-glutamyltransferase was observed. In the microsomal fraction there was a marked decrease in the content and activity of microsomal monooxygenases. Introperitoneal injection of berberine (10 mg/kg) for 6 days caused a partial normalization of permeability of hepatocytes plasma membranes and activity microsomal flavin-containing monooxygenases. It is suggested that berberine is a substrate and inducer of flavin-containing monooxygenases. Membrane-stabilizing effect of berberine is probably realized at the level of inhibition of prooxidant status of liver cells.

A new liver function test using the asialoglycoprotein-receptor system on the liver cell membrane, 3

International Nuclear Information System (INIS)

Hazama, Hiroshi; Kawa, Soukichi; Kubota, Yoshitsugu

1986-01-01

We evaluated the vilidity of a new liver function test using liver scintigraphy based on the asialoglycoprotein (ASGP) receptor system on the liver cell membrane in rats with galactosamine-induced acute liver disorder and those with carbon tetra-chloride-induced chronic liver disorder. Neoglycoprotein (GHSA) produced by combining human serum albumin with 32 galactose units was labeled with 99m Tc and administered (50 μg/100 g body weight) to rats with acute or chronic liver disorder. Clearance curves were produced based on liver scintigrams and analysed using the two-compartment model to obtain parameters. In acute liver disorder, the prolongation of 99m Tc-GHSA clearance and the decrease in ASGP receptor activities correlated well to the increase in serum GOT and the decrease in the esterified to total cholesterol ratio (E/T ratio); in chronic liver disorder, they correlated significantly to the increase in the content of liver hydroxyproline (Hyp) which increased in proportion to the severity of liver fibrosis studied histologically, and to the decrease in the contents of cytochrome P-450 and cytochrome b 5 in liver microsomes. Significant correlation was observed between the prolongation of 99m Tc-GHSA clearance and the decrease in ASGP receptor activities in both acute and chronic liver disorders. These findings indicate that the measurement of 99m Tc-GHSA clearance can be a new liver function test sensitively reflecting the severity of liver damage. (author)

First Trimester Hemolysis, Elevated Liver Enzymes, Low Platelets Syndrome in a Surrogate Pregnancy

OpenAIRE

Myer, Emily; Hill, James

2015-01-01

Background The occurrence of hemolysis, elevated liver enzymes, low platelets (HELLP) syndrome before 20 weeks of gestation is rare. HELLP is a possible but rare syndrome in gestational surrogate pregnancies for surrogates with risk factors for development of preeclampsia. Case A 32-year-old patient with chronic hypertension and positive antinuclear antibody presented for prenatal care at 13 weeks and 1 day. She was a surrogate for the embryo of a 43-year-old couple. By 15 weeks she developed...

Pyrethroid activity-based probes for profiling cytochrome P450 activities associated with insecticide interactions.

Science.gov (United States)

Ismail, Hanafy M; O'Neill, Paul M; Hong, David W; Finn, Robert D; Henderson, Colin J; Wright, Aaron T; Cravatt, Benjamin F; Hemingway, Janet; Paine, Mark J I

2013-12-03

Pyrethroid insecticides are used to control diseases spread by arthropods. We have developed a suite of pyrethroid mimetic activity-based probes (PyABPs) to selectively label and identify P450s associated with pyrethroid metabolism. The probes were screened against pyrethroid-metabolizing and nonmetabolizing mosquito P450s, as well as rodent microsomes, to measure labeling specificity, plus cytochrome P450 oxidoreductase and b5 knockout mouse livers to validate P450 activation and establish the role for b5 in probe activation. Using PyABPs, we were able to profile active enzymes in rat liver microsomes and identify pyrethroid-metabolizing enzymes in the target tissue. These included P450s as well as related detoxification enzymes, notably UDP-glucuronosyltransferases, suggesting a network of associated pyrethroid-metabolizing enzymes, or "pyrethrome." Considering the central role P450s play in metabolizing insecticides, we anticipate that PyABPs will aid in the identification and profiling of P450s associated with insecticide pharmacology in a wide range of species, improving understanding of P450-insecticide interactions and aiding the development of unique tools for disease control.

Differential induction of enzymes and genes involved in lipid metabolism in liver and visceral adipose tissue of juvenile yellow catfish Pelteobagrus fulvidraco exposed to copper

International Nuclear Information System (INIS)

Chen, Qi-Liang; Luo, Zhi; Pan, Ya-Xiong; Zheng, Jia-Lang; Zhu, Qing-Ling; Sun, Lin-Dan; Zhuo, Mei-Qin; Hu, Wei

2013-01-01

Highlights: •Cu downregulates lipogenesis and reduces lipid deposition in liver and adipose tissue. •Mechanism of Cu affecting lipid metabolism is determined at the enzymatic and molecular levels. •Cu exposure differentially influences lipid metabolism between liver and adipose tissue. -- Abstract: The present study was conducted to determine the mechanism of waterborne Cu exposure influencing lipid metabolism in liver and visceral adipose tissue (VAT) of juvenile yellow catfish Pelteobagrus fulvidraco. Yellow catfish were exposed to four waterborne copper (Cu) concentrations (2 (control), 24 (low), 71 (medium), 198 (high) μg Cu/l, respectively) for 6 weeks. Waterborne Cu exposure had a negative effect on growth and several condition indices (condition factor, viscerosomatic index, hepatosomatic index and visceral adipose index). In liver, lipid content, activities of lipogenic enzymes (6-phosphogluconate dehydrogenase (6PGD), glucose-6-phosphate dehydrogenase (G6PD), malic enzyme (ME), isocitrate dehydrogenase (ICDH), and fatty acid synthase (FAS)) as well as mRNA levels of 6PGD, G6PD, FAS and sterol-regulator element-binding protein-1 (SREBP-1) genes decreased with increasing Cu concentrations. However, activity and mRNA level of lipoprotein lipase (LPL) gene in liver increased. In VAT, G6PD, ME and LPL activities as well as the mRNA levels of FAS, LPL and PPARγ genes decreased in fish exposed to higher Cu concentrations. The differential Pearson correlations between transcription factors (SREBP-1 and peroxisome proliferators-activated receptor-γ (PPARγ)), and the activities and mRNA expression of lipogenic enzymes and their genes were observed between liver and VAT. Thus, our study indicated that reduced lipid contents in liver and VAT after Cu exposure were attributable to the reduced activities and mRNA expression of lipogenic enzymes and their genes in these tissues. Different response patterns of several tested enzymes and genes to waterborne Cu

Differential induction of enzymes and genes involved in lipid metabolism in liver and visceral adipose tissue of juvenile yellow catfish Pelteobagrus fulvidraco exposed to copper

Energy Technology Data Exchange (ETDEWEB)

Chen, Qi-Liang; Luo, Zhi, E-mail: luozhi99@yahoo.com.cn; Pan, Ya-Xiong; Zheng, Jia-Lang; Zhu, Qing-Ling; Sun, Lin-Dan; Zhuo, Mei-Qin; Hu, Wei

2013-07-15

Highlights: •Cu downregulates lipogenesis and reduces lipid deposition in liver and adipose tissue. •Mechanism of Cu affecting lipid metabolism is determined at the enzymatic and molecular levels. •Cu exposure differentially influences lipid metabolism between liver and adipose tissue. -- Abstract: The present study was conducted to determine the mechanism of waterborne Cu exposure influencing lipid metabolism in liver and visceral adipose tissue (VAT) of juvenile yellow catfish Pelteobagrus fulvidraco. Yellow catfish were exposed to four waterborne copper (Cu) concentrations (2 (control), 24 (low), 71 (medium), 198 (high) μg Cu/l, respectively) for 6 weeks. Waterborne Cu exposure had a negative effect on growth and several condition indices (condition factor, viscerosomatic index, hepatosomatic index and visceral adipose index). In liver, lipid content, activities of lipogenic enzymes (6-phosphogluconate dehydrogenase (6PGD), glucose-6-phosphate dehydrogenase (G6PD), malic enzyme (ME), isocitrate dehydrogenase (ICDH), and fatty acid synthase (FAS)) as well as mRNA levels of 6PGD, G6PD, FAS and sterol-regulator element-binding protein-1 (SREBP-1) genes decreased with increasing Cu concentrations. However, activity and mRNA level of lipoprotein lipase (LPL) gene in liver increased. In VAT, G6PD, ME and LPL activities as well as the mRNA levels of FAS, LPL and PPARγ genes decreased in fish exposed to higher Cu concentrations. The differential Pearson correlations between transcription factors (SREBP-1 and peroxisome proliferators-activated receptor-γ (PPARγ)), and the activities and mRNA expression of lipogenic enzymes and their genes were observed between liver and VAT. Thus, our study indicated that reduced lipid contents in liver and VAT after Cu exposure were attributable to the reduced activities and mRNA expression of lipogenic enzymes and their genes in these tissues. Different response patterns of several tested enzymes and genes to waterborne Cu

Rapid and accurate liquid chromatography and tandem mass spectrometry method for the simultaneous quantification of ten metabolic reactions catalyzed by hepatic cytochrome P450 enzymes.

Science.gov (United States)

Shi, Rong; Ma, Bingliang; Wu, Jiasheng; Wang, Tianming; Ma, Yueming

2015-10-01

The hepatic cytochrome P450 enzymes play a central role in the biotransformation of endogenous and exogenous substances. A sensitive high-throughput liquid chromatography with tandem mass spectrometry assay was developed and validated for the simultaneous quantification of the products of ten metabolic reactions catalyzed by hepatic cytochrome P450 enzymes. After the substrates were incubated separately, the samples were pooled and analyzed by liquid chromatography with tandem mass spectrometry using an electrospray ionization source in the positive and negative ion modes. The method exhibited linearity over a broad concentration range, insensitivity to matrix effects, and high accuracy, precision, and stability. The novel method was successfully applied to study the kinetics of phenacetin-O deethylation, coumarin-7 hydroxylation, bupropion hydroxylation, taxol-6 hydroxylation, omeprazole-5 hydroxylation, dextromethorphan-O demethylation, tolbutamide-4 hydroxylation, chlorzoxazone-6 hydroxylation, testosterone-6β hydroxylation, and midazolam-1 hydroxylation in rat liver microsomes. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Effect of Vitamin B5 on Liver Enzyme Levels in Bile Duct Ligation Cholestatic Rat Model

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Fatemeh Sadat Emami

2017-01-01

Full Text Available Background and Objectives: Accumulation of toxic bile salts in a bile duct ligation (BDL animal model plays a pivotal role in the induction of liver fibrosis. Vitamin B5 is an essential nutrient, which acts as a cofactor in many detoxification system enzymes. In the present research, the antifibrotic effect of vitamin B5 was investigated on liver cholestasis induced by BDL in rats. Methods: In this experimental study, 72 male Wistar rats were divided into 9 groups: Control, sham-operated, vitamin B5 (5, 50, and 100mg/kg bw, BDL, and BDL+vitamin B5 (5, 50, and 100mg/kg bw. After BDL, rats were given vitamin B5 via intragastric gavage for 28 consecutive days. At the end of the experiment, blood was collected from heart and activity of aspartate aminotransferase (AST, alanine aminotransferase (ALT, and alkaline phosphatase (ALP enzymes, were measured. The data were analyzed using one-way ANOVA test. Results: In the BDL animals, the serum activities of AST, ALT, and ALP significantly increased (p<0.001. Treatment of BDL rats with vitamin B5 significantly attenuated these changes. Conclusion: The results of this study indicated that vitamin B5 has hepatoprotective and antifibrotic effects in the cholestatic liver, which is likely due to the antioxidative and free radical scavenging effects of this vitamin.

Effects of Angiotensin Converting Enzyme Inhibitors on Liver Fibrosis in HIV and Hepatitis C Coinfection

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Lindsey J. Reese

2012-01-01

Full Text Available Background. Liver fibrosis is accelerated in HIV and hepatitis C coinfection, mediated by profibrotic effects of angiotensin. The objective of this study was to determine if angiotensin converting enzyme inhibitors (ACE-Is attenuate liver fibrosis in coinfection. Methods. A retrospective review of 156 coinfected subjects was conducted to analyze the association between exposure to ACE-Is and liver fibrosis. Noninvasive indices of liver fibrosis (APRI, FIB-4, Forns indices were compared between subjects who had taken ACE-Is and controls who had not taken them. Linear regression was used to evaluate ACE-I use as an independent predictor of fibrosis. Results. Subjects taking ACE-Is for three years were no different than controls on the APRI and the FIB-4 but had significantly higher scores than controls on the Forns index, indicating more advanced fibrosis. The use of ACE-Is for three years remained independently associated with an elevated Forns score when adjusted for age, race, and HIV viral load (P<0.001. There were significant associations between all of the indices and significant fibrosis, as determined clinically and radiologically. Conclusions. There was not a protective association between angiotensin inhibition and liver fibrosis in coinfection. These noninvasive indices may be useful for ruling out significant fibrosis in coinfection.

Monolignol biosynthesis in microsomal preparations from lignifying stems of alfalfa (Medicago sativa L.).

Science.gov (United States)

Guo, Dianjing; Chen, Fang; Dixon, Richard A

2002-11-01

Microsomal preparations from lignifying stems of alfalfa (Medicago sativa L.) contained coniferaldehyde 5-hydroxylase activity and immunodetectable caffeic acid 3-O-methyltransferase (COMT), and catalyzed the S-adenosyl L-methionine (SAM) dependent methylation of caffeic acid, caffeyl aldehyde and caffeyl alcohol. When supplied with NADPH and SAM, the microsomes converted caffeyl aldehyde to coniferaldehyde, 5-hydroxyconiferaldehyde, and traces of sinapaldehyde. Coniferaldehyde was a better precursor of sinapaldehyde than was 5-hydroxyconiferaldehyde. The alfalfa microsomes could not metabolize 4-coumaric acid, 4-coumaraldehyde, 4-coumaroyl CoA, or ferulic acid. No metabolism of monolignol precursors was observed in microsomal preparations from transgenic alfalfa down-regulated in COMT expression. In most microsomal preparations, the level of the metabolic conversions was independent of added recombinant COMT. Taken together, the data provide only limited support for the concept of metabolic channeling in the biosynthesis of S monolignols via coniferaldehyde.

Promoting effects of phenobarbital on the enzyme-altered foci induced by intrahepatic γ-ray-irradiation in the rat liver

International Nuclear Information System (INIS)

Ida, Katsuya; Nakamura, Satoshi; Muro, Hiroyuki; Takai, Michikatsu; Kaneko, Masao

1995-01-01

Radiation-induced carcinogenesis of the rat liver using iridium-192 seeds as an intrahepatic radioactive source was studied by enzyme histochemical means. Rats were divided into six groups according to various combinations of one or two iridium-192 or stainless steel seeds and whether they were given a diet containing 0.05% phenobarbital (PB) or a basal diet (BD). Each group were sacrificed at 20, 40, and 60 weeks after intrahepatic insertion of the iridium-192 or stainless steel seeds. γ-Glutamyl transpeptidase (GGT), glucose-6-phosphatase (G6Pase), and adenosine triphosphatase (ATPase) were stained in the liver tissues, and GGT-positive foci were quantified. Liver neoplasm was not evident, but enzyme-altered foci (EAF) were induced by γ-ray irradiation. At every point (20, 40, and 60 weeks) after the insertion of the seeds, the GGT-positive area was larger in the rats given PB than those given BD. Moreover, despite the iridium-192 radioactivity decay, EAF developed continuously in the rats given PB, and persisted in those given BD from 40 to 60 weeks after insertion. These results indicated that phenobarbital promotes the development of EAF initiated by irradiation, as it promotes the process of chemical carcinogenesis in the rat liver. (author)

Enzymatic conversion of bilirubin monoglucuronide to diglucuronide by rat liver plasma membranes

NARCIS (Netherlands)

Jansen, P. L.; Chowdhury, J. R.; Fischberg, E. B.; Arias, I. M.

1977-01-01

Formation of bilirubin monoglucuronide from unconjugated bilirubin requires a microsomal enzyme, UDP-glucuronate glucuronyltransferase (EC 2.4.1.17). Conversion of bilirubin monoglucuronide to bilirubin diglucuronide, the major bilirubin conjugate in bile, was studied in subcellular fractions of rat

Properties of 5-aminolaevulinate synthetase and its relationship to microsomal mixed-function oxidation in the southern armyworm (Spodoptera eridania).

Science.gov (United States)

Brattsten, L B; Wilkinson, C F

1975-07-01

1. Activity of 5-aminolaevulinate synthetase was measured in the midgut and other tissues of the last larval instar of the southern armyworm (Spodoptera eridania Cramer, formerly Prodenia eridania Cramer). 2. Optimum conditions for measuring the activity were established with respect to all variables involved and considerable differences from those reported for mammalian enzyme preparations were found. 3. Maximum activity (20 nmol/h per mg of protein) occurs 18-24 h after the fifth moult and thereafter decreases to trace amounts as the larvae age and approach pupation. 4. Synthetase activity was rapidly induced by oral administration (in the diet) of pentamethylbenzene, phenobarbital, diethyl 1,4-dihydro-2,4,6-trimethylpyridine-3, 5-dicarboxylate, and 2-allyl-2-isopropylacetamide. 5. Puromycin inhibited the induction of synthetase by pentamethylbenzene. 6. Induction of 5-aminolaevulinate synthetase correlated well with the induction of microsomal N-demethylation of p-chloro-N-methylaniline, except for phenobarbital, which induced the microsomal oxidase relatively more than the synthetase.

Effect of propylthiouracil on 125I-L-triiodothyronine binding to the nuclei and on malic enzyme activity in rat liver cytosol

International Nuclear Information System (INIS)

Knopp, J.

1980-01-01

The effects of propylthiouracil (PTU), triiodothyronine (T 3 ) and thyroxine (T 4 ) on the malic enzyme activity of rat liver cytosol and on the binding of 125 I-L-triiodothyronine to the nuclear fraction were examined. A significant decrease in in vitro binding of 125 I-T 3 to the liver nuclei was found in rats fed PTU for 12 h, 24 h and 120 h. The hepatic malic enzyme activity was unchanged after 12 h, but markedly decreased after 24 h and 120 h of PTU treatment. In 120 h PTU fed animals the effect of T 3 on the malic enzyme activity was five times higher in comparison with an equimolar dose of T 4 . Finally, the effect of T 4 on the malic enzyme activity was observed which might result from the intrinsic activity of T 4 under the conditions of a decreased T 4 to T 3 conversion due to the PTU feeding. The obtained results show that there is a certain correlation between the nuclear T 3 binding and the cytosol malic enzyme activity which is presumably induced directly through the m-RNA synthesis stimulated by thyroid hormones. (author)

Regulation of rat liver cytochrome P450j, a high affinity N-nitrosodimethylamine demethylase (NDMAD)

International Nuclear Information System (INIS)

Thomas, P.E.; Bandiera, S.; Maines, S.L.; Ryan, D.E.; Levin, W.

1987-01-01

Purified IgG from sera of rabbits immunized with homogeneous P450j was absorbed to produce monospecific anti-P450j. Results using anti-P450j in ELISA show that rat liver microsomal P450j content decreases between 3 and 6 wks of age in both sexes. Several xenobiotics (Aroclor 1254, mirex and 3-methylcholanthrene) repressed P450j levels when administered to male rats. In contrast, hepatic levels of P450j were induced by isoniazid, dimethylsulfoxide, pyrazole, 4-methylpyrazole, ethanol and chemically-induced diabetes. P450j levels were measurable in kidney, whereas this isozyme was barely detectable in lung, ovaries and testes; however, extra-hepatic P450j was inducible by isoniazid. Between 80-90% of microsomal NDMAD was inhibited by anti-P450j whether the microsomes were isolated from untreated rats or animals administered inducers or repressors of P450j. Results obtained with the reconstituted system suggest that the remaining microsomal NDMAD resistant to antibody inhibition is the result of the inaccessibility of a certain proportion of P450j due to interference by NADPH-P450 reductase. P450j content and NDMAD activity correlated well in microsomes from rats of all treatment groups. The evidence indicates that P450j is the primary, and possibly only, microsomal catalyst of NDMAD at substrate concentrations relevant to hepatocarcinogenesis induced by NDMA

Shotgun Proteomics of Aspergillus niger Microsomes upon d-Xylose Induction▿ †

Science.gov (United States)

de Oliveira, José Miguel P. Ferreira; van Passel, Mark W. J.; Schaap, Peter J.; de Graaff, Leo H.

2010-01-01

Protein secretion plays an eminent role in cell maintenance and adaptation to the extracellular environment of microorganisms. Although protein secretion is an extremely efficient process in filamentous fungi, the mechanisms underlying protein secretion have remained largely uncharacterized in these organisms. In this study, we analyzed the effects of the d-xylose induction of cellulase and hemicellulase enzyme secretion on the protein composition of secretory organelles in Aspergillus niger. We aimed to systematically identify the components involved in the secretion of these enzymes via mass spectrometry of enriched subcellular microsomal fractions. Under each condition, fractions enriched for secretory organelles were processed for tandem mass spectrometry, resulting in the identification of peptides that originate from 1,081 proteins, 254 of which—many of them hypothetical proteins—were predicted to play direct roles in the secretory pathway. d-Xylose induction led to an increase in specific small GTPases known to be associated with polarized growth, exocytosis, and endocytosis. Moreover, the endoplasmic-reticulum-associated degradation (ERAD) components Cdc48 and all 14 of the 20S proteasomal subunits were recruited to the secretory organelles. In conclusion, induction of extracellular enzymes results in specific changes in the secretory subproteome of A. niger, and the most prominent change found in this study was the recruitment of the 20S proteasomal subunits to the secretory organelles. PMID:20453123

Metabolism of the fungicide Denmert (S-n-butyl S'-p-tert-butyl-benzyl N-3-pyridyldithiocarbonimidate, S-1358) and Denmert sulfoxides in liver enzyme systems

International Nuclear Information System (INIS)

Ohkawa, Hideo; Okihara, Yukiko; Miyamoto, Junshi

1976-01-01

On incubation with rat liver microsomes plus MADPH, Denmert (S-n-butyl S'-p-tert-butylbenzyl N-3-pyridyldithiocarbonimidate) underwent at least two different types of oxidation; hydroxylation and sulfoxidation. Hydroxylation of Denmert at the tert-butyl group was one of the major metabolic attacks in mammalian metabolism. Sulfoxidation of Denmert gave two isomers of Denmert sulfoxides which were intermediates in the metabolism and readily transformed into 2-(3'-pyridylimino)-4-carboxylthiazolidine (HM) in the presence of L-cysteine without enzymatic mediation. This type of conjugation with cysteine appears to be a new class of metabolic reactions in mammals. Denmert S-oxide showed increased fungicidal activity when assayed in liquid cultures, but not on plant leaves. (auth.)

Association between hepatic steatosis and serum liver enzyme levels with atrial fibrillation in the general population: The Study of Health in Pomerania (SHIP).

Science.gov (United States)

Markus, Marcello Ricardo Paulista; Meffert, Peter J; Baumeister, Sebastian Edgar; Lieb, Wolfgang; Siewert, Ulrike; Schipf, Sabine; Koch, Manja; Kors, Jan A; Felix, Stephan Burkhard; Dörr, Marcus; Targher, Giovanni; Völzke, Henry

2016-02-01

Hepatic steatosis (HS) affects up to 35% of adults in the general population. Atrial fibrillation (AF) is the most prevalent sustained arrhythmia and has a substantial impact on healthcare costs. We analyzed cross-sectional associations of HS and serum liver enzyme levels with prevalent AF in a general population sample. We analyzed data from 3090 women and men, aged 20-81 years, from the population-based Study of Health in Pomerania. HS was determined by ultrasonography. Serum levels of alanine aminotransferase (ALT), aspartate aminotransferase (AST) and gamma-glutamyltranspeptidase (GGT) were measured photometrically. AF was determined by automatic electrocardiographic analysis software. The prevalences of HS and AF were 30.3% and 1.49%, respectively. ALT, AST and GGT showed a positive linear association with the risk of prevalent AF, after multivariable adjustment. The adjusted odds ratios for AF per 1-standard deviation increment in log-transformed serum liver enzyme levels were 1.65 (95% confidence interval [CI]: 1.16 to 2.35; p = 0.006) for ALT, 1.47 (95%CI: 1.07 to 2.02; p = 0.017) for AST and 2.17 (95%CI: 1.64 to 2.87; p < 0.001) for GGT. In contrast, ultrasonographic HS was not associated with AF. Our findings indicate that moderately elevated serum liver enzymes, but not sonographic liver hyperechogenicity, were associated with increased AF prevalence in the general adult population. The hepatic release of increased levels of serum liver enzymes might be accompanied by higher levels of pro-inflammatory, pro-coagulant and pro-fibronogenic mediators that might lead to structural and electrical remodeling of the atrium resulting in the development and persistence of AF. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

The relationship between 25-hydroxyvitamin D concentration and liver enzymes in overweight or obese adults: Cross-sectional and interventional outcomes.

Science.gov (United States)

Naderpoor, Negar; Mousa, Aya; de Courten, Maximilian; Scragg, Robert; de Courten, Barbora

2018-03-01

Vitamin D deficiency is prevalent in individuals with non-alcoholic fatty liver disease (NAFLD) and non-alcoholic steatohepatitis. However, there is limited and inconsistent data on the effect of vitamin D supplementation on liver function. Hepatic enzymes have been used as surrogate markers for NAFLD and have been associated with metabolic syndrome. We examined the relationships between 25-hydroxyvitamin D (25(OH)D) and γ-glutamyl transferase (GGT), alanine aminotransferase (ALT), alkaline phosphatase (ALP) in 120 drug-naïve individuals with no history of liver disease. In addition, the effect of vitamin D supplementation (100,000 loading dose of cholecalciferol followed by 4000IU daily for 16 weeks) on hepatic enzymes was investigated in a subgroup of 54 vitamin D-deficient overweight or obese individuals (28 randomised to cholecalciferol and 26 to placebo). Hepatic enzymes, anthropometric parameters, lipid profile, insulin sensitivity (hyperinsulinaemic-euglycaemic clamp, M value) and high sensitivity C-reactive protein (hs-CRP) were measured before and after the intervention. In the cross-sectional study, levels of GGT and ALT were higher in men compared to women (both p=0.001). There were no significant differences in GGT, ALT and ALP between vitamin D categories (25(OH)D50nmol/L) and no relationships were found between the three enzymes and 25(OH)D before and after adjustment for age, sex, BMI, WHR, and insulin sensitivity (all p>0.5). In the randomised trial, 25(OH)D concentrations increased in the vitamin D group (mean change 57.0±21.3nmol/L) compared to the placebo group (mean change 1.9±15.1nmol/L). Mean changes in GGT, ALT and ALP were not significantly different between vitamin D and placebo groups (all p>0.2). Change in 25(OH)D concentration was not correlated with changes in GGT, ALT and ALP before and after adjustments for age and sex (all p>0.1). In summary, 25(OH)D concentrations were not related to hepatic enzymes in drug-naive adults with no

The Effect of Myrtus communis Extract on Liver Enzymes and Blood Biochemical Factors in Diabetic Adult Male Rats

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Habiballah Johari

2014-10-01

Full Text Available Background: The aim of this study was the effect of Myrtus communis extract on liver enzymes and blood biochemical factors in diabetic adult male rats. Materials and Methods: This study has been carried out experimentally and completely random. Seventy adult male Wistar rats were divided in 7 groups including: control which received no treatment, sham who received 2 mL of distilled water, the 1st, 2nd and 3rd experimental groups which received 0.75, 1.5 and 3 mg/kg Myrtus communis leaf extract respectively, the 4th experimental group as the diabetic control group who received streptozotocin (60 mg/kg and the 5th experimental group as the diabetic treatment group who received 3 mg/kg of extract. This experiment lasted 14 days with prescript orally. After this period, all the rats, were weighted, anesthetized and blood samples were taken from the heart centrifuged and sera were evaluated for the concentration of various factors. In addition liver were removed and sliced. Results: According to the obtained results, the plasma concentration of liver enzyme (alanine aminotransferase, alkaline phosphatase, aspartate aminotransferase, cholesterol and glucose presented a significant decrease at (p≤0.05. Whereas no significant change were seen in body weight, triglyceride, urea, albumin and total protein. Histological studies of the liver tissue showed no significant difference among various groups. Conclusion: Myrtus communis is comprise of collections of flavonoids and other various components with antioxidant and anti inflammatory properties. Thence it can effective in treatment of liver diseases and decrease of blood sugar and cholesterol in diabetes mellitus patients.

Dynamics of some conjugated enzymes of aminonitrogen metabolism in the liver of the irradiated body

International Nuclear Information System (INIS)

Savitskij, V.I.

1976-01-01

Changes in the activity of five conjugated enzymes of the aminonitrogen metabolism in subcellular fractions of liver tissue have been studied on irradiated (450 R) rabbits during thirty days after exposure. These changes are peculiar for their manifestation in time, their depth and trend. It is suggested that in the early period of radiation damage, gluconeogenesis is enhanced, and in the later period, biosynthesis of pyrimidine bases is intensified

The Role of Liver Biopsy in the Management of Patients with Liver Disease

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Florence Wong

2003-01-01

Full Text Available The role of liver biopsy in the diagnosis and management of liver disease is a controversial issue even among hepatologists. Although most causes of elevated liver enzymes can be determined, or at least suspected, on the basis of a careful history and laboratory tests, histological assessment remains the gold standard for most liver diseases. Histological evaluation can either confirm or refute clinical diagnoses and can provide information about the severity and stage of disease. Occasionally, the liver biopsy also provides an additional diagnosis. The spectrum of nonalcoholic fatty liver disease accounts for a substantial proportion of cases of chronically elevated liver enzymes and can be reliably diagnosed only by liver biopsy. Prognostic information can be obtained in patients with this disorder, as well as in those with alcoholic liver disease and viral hepatitis, and liver biopsy can be used as a guide to their management.

Microsomal biotransformation of chlorpyrifos, parathion and fenthion in rainbow trout (Oncorhynchus mykiss) and coho salmon (Oncorhynchus kisutch): mechanistic insights into interspecific differences in toxicity

Science.gov (United States)

Lavado, Ramon

2010-01-01

Rainbow trout often serve as a surrogate species evaluating xenobiotic toxicity in cold-water species including other salmonids of the same genus, which are listed as threatened or endangered. Biotransformation tends to show species-specific patterns that influence susceptibility to xenobiotic toxicity, particularly organophoshpate insecticides (OPs). To evaluate the contribution of biotransformation in the mechanism of toxicity of three organophosphate (phosphorothionate) insecticides, chlorpyrifos, parathion and fenthion, microsomal bioactivation and detoxification pathways were measured in gills, liver and olfactory tissues in juvenile rainbow trout (Oncorhynchus mykiss) and compared to juvenile coho salmon (Oncorhynchus kisutch). Consistent with species differences in acute toxicity, significantly higher chlorpyrifos bioactivation was found in liver microsomes of rainbow trout (up to 2-fold) when compared with coho salmon. Although bioactivation to the oxon was observed, the catalytic efficiency towards chlorpyrifos dearylation (detoxification) was significantly higher in liver for both species (1.82 and 0.79 for trout and salmon, respectively) when compared to desulfuration (bioactivation). Bioactivation of parathion to paraoxon was significantly higher (up to 2.2-fold) than detoxification to p-nitrophenol in all tissues of both species with rates of conversion in rainbow trout, again significantly higher than coho salmon. Production of fenoxon and fenthion sulfoxides from fenthion was detected only in liver and gills of both species with activities in rainbow trout significantly higher than coho salmon. NADPH-Dependent hydrolysis of fenthion was observed in all tissues, and was the only activity detected in olfactory tissues. These results indicate rainbow trout are more sensitive than coho salmon to the acute toxicity of OP pesticides because trout have higher catalytic rates of oxon formation. Thus, rainbow trout may serve as a conservative surrogate

Enantioselective N-demethylation and hydroxylation of sibutramine in human liver microsomes and recombinant cytochrome p-450 isoforms.

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Shinde, Dhananjay D; Kim, Min-Jung; Jeong, Eun-Sook; Kim, Yang-Weon; Lee, Ji-Woo; Shin, Jae-Gook; Kim, Dong-Hyun

2014-01-01

The enantioselective metabolism of sibutramine was examined using human liver microsomes (HLM) and recombinant cytochrome P-450 (CYP) isoforms. This drug is metabolized to N-mono-desmethyl- (M1) and N,N-di-desmethylsibutramine (M2), and subsequent hydroxylation results in hydroxyl M1 (HM1) and hydroxyl M2 (HM2). No significant difference was noted in formation of M1from sibutramine between R- and S-sibutramine in HLM. However, S-enantiomers of M1 and M2 were preferentially metabolized to M2, HM1, and HM2compared to R-enantiomers in HLM, and intrinsic clearance (Clint) ratios of S-enantiomers/R-enantiomers were 1.97, 4.83, and 9.94 for M2, HM1, and HM2, respectively. CYP3A4 and CYP3A5 were only involved in the formation of M1, whereas CYP2B6 and CYP2C19 were responsible for all metabolic reactions of sibutramine. CYP2C19 and CYP3A5 displayed catalytic preference for S-sibutramine to S-M1, whereas CYP2B6 and CYP3A4 showed little or no stereoselectivity in metabolism of sibutramine to M1. In the case of M2 formation, CYP2B6 metabolized S-M1 more rapidly than R-M1 with a Clint ratio of 2.14. However, CYP2C19 catalyzed less S-M1 than R-M1 and the Clint ratio of S-M1 to R-M1 was 0.65. The most significant enantioselectivity was observed in formation of HM1 from M1, and HM2 from M2. CYP2B6 and CYP2C19 exhibited preferential catalysis of formation of hydroxyl metabolites from S-enantiomers rather than R-enantiomers. These results indicate that S-sibutramine was more rapidly metabolized by CYP isoforms than R-sibutramine, and that enantioselective metabolism needs to be considered in drug interactions involving sibutramine and co-administered drugs.

Maintenance of adult primate liver in organ culture: Potential use in toxicity testing

International Nuclear Information System (INIS)

Smith, P.F.; O'Brien, K.A.; Allen, L.; DeLuca, J.; Norman, B.; Keenan, K.P.

1991-01-01

Adult Rhesus monkey liver slices were incubated using a dynamic organ culture method to determine hepatocyte viability, drug biotransformation potential and the in vitro response to the hepatotoxicant, allyl alcohol (AA). After 1, 2, 4, or 8 hr, slices were removed from culture and analyzed for incorporation of [ 3 H]-leucine into acid-precipitable material, and medium alanine aminotransferase (ALT) activity was determined. Separate slices were taken for histological evaluation and for evaluation of microsomal 7-ethoxy-4-trifluoromethyl coumarin-O-deethylase (EFCOD) activity. Incorporation of [ 3 H]-leucine into slices was linear over the period of incubation and was specifically inhibited by cycloheximide (10 uM) at all time points. In the absence of AA, enzyme leakage was minimal over 8 hr. Marked ALT leakage occurred with 1 mM AA. Control slices had an initial fall to 55% of in vivo EFCOD activity that stabilized at 40-50% control slices indicated that there was minimal cellular degeneration and that, in PAS-stained sections, glycogen accumulation occurred over the incubation period. This system allows for maintenance and viability of adult primate liver slices in culture for at least 8 hr and may be useful for in vitro toxicity and biotransformation studies

Transmission electron microscopy of heart and liver tissues from rats fed with gums arabic and tragacanth.

Science.gov (United States)

Anderson, D M; Ashby, P; Busuttil, A; Kempson, S A; Lawson, M E

1984-04-01

Transmission electron microscopy has been used to examine the ultrastructure of rat hearts and livers after diet supplementation with (a) 0, 0.5, 1.5, 2.5 and 3.5% (w/w) gum tragacanth (GT) for 91 days, (b) 0 and 1% GT for 5 days (c) 0, 1, 4 and 8% (w/w) gum arabic (GA) for 28 days. The preparation and scrutiny of the electron micrographs was undertaken by two independent teams of specialists. There were no detectable abnormalities in any of the organelles in the heart and liver specimens from any of the test animals and no inclusions nor other pathological changes were observed. All micrographs showed normal, healthy tissues; particular attention was given to the mitochondria in hepatocytes as they serve as sensitive indicators of the health and state of activity of cells. In addition, the data obtained from assays of the microsomal protein and cytochrome P-450 content of the livers showed that GA and GT did not cause inductive effects. These results do not support earlier suggestions, based on in vitro assays, that GA and GT cause changes in the function of rat heart and liver mitochondria and liver microsomes; however, they confirm a report by Zbinden that the ingestion of GT does not produce abnormalities in the cardiac function of rats.

ENZYME MARKERS ACTIVITY AND BILE FORMATION FUNCTION OF LIVER IN CASES OF TUBERCULOSTATICS AND HEXAVALENT CHROMIUM COMPOUNDS AFFECTION IN RATS

OpenAIRE

N. I. Burmas; L. S. Fira; P. H. Lyhackyy

2016-01-01

Background. Currently, the growing incidence of toxic lesions of the liver is associated with industrial chemicalization and uncontrolled use of hepatotoxic drugs in everyday life. There are about one thousand drugs with high or low hepatotoxicity, such as anti-TB drugs. Objective. In this research we studied the intracellular enzymes activity and bile formation function of the liver in rats of different ages in cases of tuberculostatic (isoniazid and rifampicin) affection and chromium (p...

Environmentally relevant organophosphate triesters in herring gulls: In vitro biotransformation and kinetics and diester metabolite formation using a hepatic microsomal assay

International Nuclear Information System (INIS)

Greaves, Alana K.; Su, Guanyong; Letcher, Robert J.

2016-01-01

The in vitro biotransformation and kinetics of six organophosphate triester (OPE) flame retardants were investigated in herring gulls (Larus argentatus) from the Great Lakes using a hepatic microsomal metabolism assay. Administration of each individual OPE (tri-n-butyl phosphate (TNBP), tris(2-butoxyethyl) phosphate (TBOEP), triphenyl phosphate (TPHP), triethyl phosphate (TEP), tris(1,3-dichloro-2-propyl) phosphate (TDCIPP) and tris(2-chloroisopropyl) phosphate (TCIPP)) to the in vitro assay (concentration range 0.01 to 10 μM) resulted in rapid depletion with the exception of TEP. Following the Michaelis-Menten enzyme kinetics model, a preliminary 2-minute incubation period was used to estimate the V max (± SE) values (i.e., the maximal rate of reaction for a saturated enzyme system), which ranged from 5.0 ± 0.4 (TPHP) to 29 ± 18 pmol/min/mg protein (TBOEP), as well as the K M (± SE) values (i.e., the OPE concentration corresponding to one half of the V max ), which ranged from 9.8 ± 1 (TPHP) to 189 ± 135 nM (TBOEP). Biotransformation assays over a 100-minute incubation period revealed that TNBP was metabolized most rapidly (with a depletion rate of 73 ± 4 pmol/min/mg protein), followed by TBOEP (53 ± 8 pmol/min/mg), TCIPP (27 ± 1 pmol/min/mg), TPHP (22 ± 2 pmol/min/mg) and TDCIPP (8 ± 1 pmol/min/mg). In vitro biotransformation of OP triesters was clearly structure-dependent where non-halogenated alkyl OP triesters were metabolized more rapidly than halogenated alkyl triesters. Halogenated OP triesters were transformed to their respective diesters more efficiently relative to non-halogenated OP triesters. To our knowledge, this is the first study to investigate OP triester metabolism and OP diester formation in an avian or wildlife model system, which is important to understand the fate and biological activity of OPEs in an exposed organism. - Highlights: • The metabolism and kinetics of 6 OPEs were examined in herring gull liver microsomes. • The

Environmentally relevant organophosphate triesters in herring gulls: In vitro biotransformation and kinetics and diester metabolite formation using a hepatic microsomal assay

Energy Technology Data Exchange (ETDEWEB)

Greaves, Alana K. [Wildlife and Landscape Directorate, Science and Technology Branch, Environment and Climate Change Canada, National Wildlife Research Centre, Carleton University, Ottawa, ON K1A 0H3 (Canada); Department of Chemistry, Carleton University, Ottawa, ON K1S 5B6 (Canada); Su, Guanyong, E-mail: guanyong.su85@gmail.com [Wildlife and Landscape Directorate, Science and Technology Branch, Environment and Climate Change Canada, National Wildlife Research Centre, Carleton University, Ottawa, ON K1A 0H3 (Canada); Department of Chemistry, Carleton University, Ottawa, ON K1S 5B6 (Canada); Letcher, Robert J., E-mail: robert.letcher@canada.ca [Wildlife and Landscape Directorate, Science and Technology Branch, Environment and Climate Change Canada, National Wildlife Research Centre, Carleton University, Ottawa, ON K1A 0H3 (Canada); Department of Chemistry, Carleton University, Ottawa, ON K1S 5B6 (Canada)

2016-10-01

The in vitro biotransformation and kinetics of six organophosphate triester (OPE) flame retardants were investigated in herring gulls (Larus argentatus) from the Great Lakes using a hepatic microsomal metabolism assay. Administration of each individual OPE (tri-n-butyl phosphate (TNBP), tris(2-butoxyethyl) phosphate (TBOEP), triphenyl phosphate (TPHP), triethyl phosphate (TEP), tris(1,3-dichloro-2-propyl) phosphate (TDCIPP) and tris(2-chloroisopropyl) phosphate (TCIPP)) to the in vitro assay (concentration range 0.01 to 10 μM) resulted in rapid depletion with the exception of TEP. Following the Michaelis-Menten enzyme kinetics model, a preliminary 2-minute incubation period was used to estimate the V{sub max} (± SE) values (i.e., the maximal rate of reaction for a saturated enzyme system), which ranged from 5.0 ± 0.4 (TPHP) to 29 ± 18 pmol/min/mg protein (TBOEP), as well as the K{sub M} (± SE) values (i.e., the OPE concentration corresponding to one half of the V{sub max}), which ranged from 9.8 ± 1 (TPHP) to 189 ± 135 nM (TBOEP). Biotransformation assays over a 100-minute incubation period revealed that TNBP was metabolized most rapidly (with a depletion rate of 73 ± 4 pmol/min/mg protein), followed by TBOEP (53 ± 8 pmol/min/mg), TCIPP (27 ± 1 pmol/min/mg), TPHP (22 ± 2 pmol/min/mg) and TDCIPP (8 ± 1 pmol/min/mg). In vitro biotransformation of OP triesters was clearly structure-dependent where non-halogenated alkyl OP triesters were metabolized more rapidly than halogenated alkyl triesters. Halogenated OP triesters were transformed to their respective diesters more efficiently relative to non-halogenated OP triesters. To our knowledge, this is the first study to investigate OP triester metabolism and OP diester formation in an avian or wildlife model system, which is important to understand the fate and biological activity of OPEs in an exposed organism. - Highlights: • The metabolism and kinetics of 6 OPEs were examined in herring gull liver

Antibody to liver cytosol (anti-LC1) in patients with autoimmune chronic active hepatitis type 2.

Science.gov (United States)

Martini, E; Abuaf, N; Cavalli, F; Durand, V; Johanet, C; Homberg, J C

1988-01-01

A new autoantibody was detected by immunoprecipitation in the serum of 21 patients with chronic active hepatitis. The antibody reacted against a soluble cytosolic antigen in liver. The antibody was organ specific but not species specific and was therefore called anti-liver cytosol antibody Type 1 (anti-LC1). In seven of 21 cases, no other autoantibody was found; the remaining 14 cases had anti-liver/kidney microsome antibody Type 1 (anti-LKM1). With indirect immunofluorescence, a distinctive staining pattern was observed with the seven sera with anti-LC1 and without anti-LKM1. The antibody stained the cytoplasm of hepatocytes from four different animal species and spared the cellular layer around the central veins of mouse and rat liver that we have called juxtavenous hepatocytes. The immunofluorescence pattern disappeared after absorption of sera by a liver cytosol fraction. The 14 sera with both antibodies displayed anti-LC1 immunofluorescent pattern after absorption of anti-LKM1 by the liver microsomal fraction. The anti-LC1 was found in the serum only in patients with chronic active hepatitis of unknown cause. Anti-LC1 antibody was not found in sera from 100 patients with chronic active hepatitis associated with anti-actin antibody classic chronic active hepatitis Type 1, 100 patients with primary biliary cirrhosis, 157 patients with drug-induced hepatitis and a large number of patients with liver and nonliver diseases. This new antibody was considered a second marker of chronic active hepatitis associated with anti-LKM1 (anti-LKM1 chronic active hepatitis) or autoimmune chronic active hepatitis Type 2.

Association between elevated coffee consumption and daily chocolate intake with normal liver enzymes in HIV-HCV infected individuals: results from the ANRS CO13 HEPAVIH cohort study.

Science.gov (United States)

Carrieri, M Patrizia; Lions, Caroline; Sogni, Philippe; Winnock, Maria; Roux, Perrine; Mora, Marion; Bonnard, Philippe; Salmon, Dominique; Dabis, François; Spire, Bruno

2014-01-01

We used longitudinal data from the ANRS CO13 HEPAVIH cohort study of HIV-HCV co-infected individuals to investigate whether polyphenol rich food intake through coffee and/or daily chocolate consumption could play a role in reducing liver enzymes levels. Longitudinal data collection included self-administered questionnaires and medical data (aspartate aminotransferase (AST) and alanine aminotransferase (ALT) liver enzymes). Two analyses were performed to assess the association between coffee (≥3 cups a day) and daily chocolate intake and abnormal values of AST and ALT (AST or ALT >2.5 × upper normal limit (UNL)) (N=990) over time, after adjustment for known correlates. Logistic regression models based on generalized estimating equations were used to take into account the correlations between repeated measures and estimate adjusted odds ratio. After adjustment, patients reporting elevated coffee consumption and daily chocolate intake were less likely to present abnormal ALT (OR=0.65; p=0.04 and OR=0.57; p=0.04, for coffee and chocolate respectively), while only patients reporting elevated coffee consumption were less likely to have abnormal AST values (p=0.05). Nevertheless, the combined indicator of coffee and chocolate intake was most significantly associated with approximately 40% reduced risk of abnormal liver enzymes (p=0.003 for AST; p=0.002 for ALT). Elevated coffee consumption and daily chocolate intake appear to be associated with reduced levels of liver enzymes in HIV-HCV co-infected patients. Further experimental and observational research is needed to better understand the role that polyphenol intake or supplementation can play on liver disease and liver injury. Copyright © 2013 European Association for the Study of the Liver. Published by Elsevier B.V. All rights reserved.

Diagnostic performances of serum liver enzymes and cytokines in non-alcoholic fatty liver disease

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Hakan Turkon

2015-03-01

Full Text Available Objective:Non-alcoholic fatty liver disease (NAFLD is affecting people worldwide with increasing prevalence. Non-invasive tests are required for both diagnosis and staging of the disease. We aimed to evaluate diagnostic accuracy of routine liver enzymes and cytokines in NAFLD. Methods:A total of 88 cases, aged between 20 and 62 years, were included in the study. Serum ALT, AST, GGT, triglyceride, TNF-alpha, IL-6 and IL-8 were measured in 40 patients with NAFLD and in 48 healthy control patients with similar BMI and demographic characteristics. Diagnostic performances of serum biomarkers for diagnosis of NAFLD were evaluated with ROC analysis. Results:ALT and AST showed good diagnostic performance in predicting patients with NAFLD in the overall group (AUC=0.817; 95% CI[0.721-0.913], AUC=0.815;95% CI[0.718-0.911] respectively but in obese subjects ALT and AST showed poor performance (AUC=0.659;95% CI[0.478-0.841], AUC=0.680; 95% CI[0.498-0.861] respectively. Among cytokines TNF-alpha showed best performance in the diagnosis of NAFLD in both overall group and obese subjects (AUC=0.892; 95% CI[0.824- 0.959], AUC=0.858; 95% CI[0.739-0.977] respectively. The optimal cut off value for TNF-alpha was 10.65pg/ml with a sensitivity of 75% and a specificity of 93% in the overall group. IL-6 and IL-8 showed poor performance. Conclusion: TNF-alpha may be a good parameter for predicting patients with NAFLD. J Clin Exp Invest 2015;6 (1: 16-20

Characteristics and significance of D-tagatose-induced liver enlargement in rats: An interpretative review.

Science.gov (United States)

Bär, A

1999-04-01

This review addresses the issue of asymptomatic liver enlargement in rats. It was necessitated by the observation of significantly increased liver weights in rats fed diets with 10 to 20% D-tagatose, a potential new bulk sweetener, for between 28 and 90 days. Increases of liver size without accompanying histopathological changes or impairment of organ function have been observed in rats in response to the ingestion of various xenobiotic compounds (including some food additives), changes of dietary composition (e.g. , high doses of fructose and sucrose), metabolic aberrations (e.g., diabetes), as well as normal pregnancy and lactation. The underlying mechanism(s) are not yet understood in detail but peroxisome proliferation, microsomal enzyme induction, increased storage of glycogen or lipids, and hyperfunction due to an excessive workload are well-established causes of hepatomegaly in rats. In D-tagatose- and fructose-fed rats, a treatment-related increase of hepatic glycogen storage was identified as a likely cause of the liver enlargement. Dietary levels of 5% and about 15-20% were determined as no-effect levels (NOEL) for D-tagatose- and fructose-induced liver enlargement, respectively. At doses above the NOEL, D-tagatose is about four times more efficient than fructose in inducing liver enlargement. On the other hand, the estimated intake of D-tagatose from its intended uses in food is about four times lower than the actual fructose intake. Consequently, a similar safety margin would apply for both sugars. Considering the similarity of the liver effects in rats of fructose, a safe food ingredient, and D-tagatose, the absence of histopathological changes in rats fed a diet with 20% D-tagatose for 90 days, and the absence of adverse long-term consequences of glycogen-induced liver enlargement in rats, it is concluded that the observed liver enlargement in D-tagatose-fed rats has no relevance for the assessment of human safety of this substance. Copyright 1999

Changes induced by gamma radiation in microsomal membranes of storage of garlic

International Nuclear Information System (INIS)

Perez, M.B.; Croci, C.A.; Aveldano, M.I.

2003-01-01

This study evaluates the effects of the radio inhibition process on garlic bulbs in terms of phase properties of microsomal membranes and their lipid and fatty acid composition. Garlic bulbs were irradiated with an average dose of 60 Gy of 60 Co gamma rays 30-40 days after harvest. The treatment was carried out in the facilities of the National Atomic Energy Commission (CNEA). Rough and smooth microsomal membranes were isolated by ultracentrifugation from tissues of irradiated and non-irradiated storage leaves. Wide angle X-ray diffractograms of both fractions were recorded along 270 days of storage. Lipids were separated by thin layer chromatography. The fatty acid composition of major lipid fractions was studied by gas-liquid chromatography. The diffractograms featured peaks at Bragg spacing of 4.15 Armstrong and 3.75 Armstrong, revealing the presence of a gel (crystalline) phase, while the characteristic peak of the liquid-crystalline phase (4.6 Armstrong) was not observed in both sorts of membranes. Irradiation was found to bring about modifications in the intensity of 4.15 Armstrong and 3.75 Armstrong peaks from smooth microsomal membranes, but not in the behaviour along the studied period. Data from the rough microsomal fraction were erratic. Parallel to these changes, radiation induced significant modifications in the level of smooth microsomal membrane triacylglycerols in relation to phospholipids and their fatty acids. These findings indicate that the storage leaf tissues of garlic are radiosensitive both in terms of physical and chemical properties of their microsomal membranes. From the practical point of view, these results could be the basis for the development of techniques to be applied to storage garlic to evaluate if it was irradiated. (author)

A new factor from enteric bacteria of rats amplifying induction of liver enzyme by glucocorticoid. Pt. 2

International Nuclear Information System (INIS)

Kido, Hiroshi; Higashi, Takao; Katanuma, Nobuhiko

1977-01-01

1) An amplifier of the action of glucocorticoid was purified from Proteus mirabilis as described previously. It was found that it amplified the induction of liver tyrosine aminotransferase by dexamethasone markedly with doses of dexamethasone that caused minimal enzyme induction, but had little effect with doses that caused maximal induction. Thus the amplification may represent a saving of glucocorticoid. The amplification of enzyme activity was brought about by increase in amount of enzyme. 2) The amplification was observed when the amplifier was administered before or with dexamethasone, but not when it was given 2 h after dexamethasone. These results and the finding that actinomycin D inhibited the amplification indicate that the amplifier does not act on the translational level of enzyme induction. 3) It was found that the amplifier increased both incorporation of [ 3 H]dexamethasone into the cytosol and binding of [ 3 H]dexamethasone to cytosol protein and that it decreased decay of the [ 3 H]dexamethasone protein complex. (orig.) [de

Study of Cytochrome P450 2E1 and its allele Variants in Liver Injury of Nondiabetic, Nonalcoholic Steatohepatitis Obese Women

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NELSON M VARELA

2008-01-01

Full Text Available CYP2E1 enzyme is related to nonalcoholic steatohepatitis (NASH due to its ability for reactive oxygen species production, which can be influenced by polymorphisms in the gene. The aim of this study was to investigate hepatic levels, activity, and polymorphisms of the CYP2E1 gene to correlate it with clinical and histological features in 48 female obese NASH patients. Subjects were divided into three groups: (i normal; (ii steatosis; and (iii steatohepatitis. CYP2E1 protein level was assayed in microsomes from liver biopsies, and in vivo chlorzoxazone hydroxylation was determined by HPLC. Genomic DNA was isolated for genotype analysis through PCR. The results showed that liver CYP2E1 content was significantly higher in the steatohepatitis (45%; p=0.024 and steatosis (22%; p=0.032 group compared with normal group. Chlorzoxazone hydroxylase activity showed significant enhancement in the steatohepatitis group (15%, p=0.027 compared with the normal group. c2 rare allele of RsallPstl polymorphisms but no C allele of Dral polymorphism was positively associated with CHZ hydroxylation, which in turn is correlated with liver CYP2E1 content (r=0.59; p=0.026. In conclusion, c2 allele is positively associated with liver injury in NASH. This allele may determine a higher transcriptional activity of the gene, with consequent enhancement in pro-oxidant activity of CYP2E1 thus affording liver toxicity

The relationship between HbA(1c) and fasting plasma glucose in patients with increased plasma liver enzyme measurements

DEFF Research Database (Denmark)

Christiansen, R; Rasmussen, L Melholt; Nybo, H

2012-01-01

levels of increased liver enzyme concentrations. Methods:  Data from 10 065 patients with simultaneous measurement of HbA(1c) , venous fasting plasma glucose, alanine aminotransferase and γ-glutamyl transferase were extracted from our laboratory database. Correlations were investigated in four patient...

Soybean meal fermented by Aspergillus awamori increases the cytochrome P-450 content of the liver microsomes of mice.

Science.gov (United States)

Kishida, T; Ataki, H; Takebe, M; Ebihara, K

2000-04-01

The effect of soybean meal fermented by Aspergillus awamori on the acute lethality of acetaldehyde, pentobarbital sleeping time, and cytochrome P-450 content of the hepatic microsomes was studied in mice. Most of the daidzin and genistin in soybean meal (SBM) were converted into the respective aglycones, daidzein and genistein, by fermentation. In experiment 1, mice were fed isonitrogenic test diets with one of the following five protein sources for 28 d: casein, SBM, fermented and hot-air-dried SBM (FSBM-HD), fermented and freeze-dried SBM (FSBM-FD), or methanol-extracted FSBM-FD (FSMB-FD-R). The acute lethality of acetaldehyde in mice fed the FSBM-FD diet was significantly lower than that in mice fed the SBM, FSBM-HD, or FSBM-FD-R diet. In experiments 2 and 3, mice were fed isonitrogenic test diets with one of the following four protein sources for 28 d: casein, SBM, FSBM-FD, and FSBM-FD-R. The pentobarbital sleeping time was significantly shorter and the cytochrome P-450 content was significantly higher in the mice fed the FSBM-FD diet than the respective value in mice fed the other test diets. In experiment 4, mice were fed one of eight diets which contained different levels of aglycone obtained by varying the proportion of FSBM-FD and FSBM-FD-R, for 28 d. The cytochrome P-450 content in hepatic microsomes increased as the dietary level of isoflavonoid aglycones increased, but there was a saturation phenomenon. These results suggest that soy isoflavonoid aglycones are more potent inducers of cytochrome P-450 than isoflavonoid glycosides.

Stereoselective formation of a cholesterol ester conjugate from fenvalerate by mouse microsomal carboxyesterase(s).

Science.gov (United States)

Miyamoto, J; Kaneko, H; Takamatsu, Y

1986-06-01

In accordance with in vivo findings, of the four chiral isomers of fenvalerate (S-5602 Sumicidin, Pydrin, [RS]-alpha-cyano-3-phenoxybenzyl [RS]-2-(4-chlorophenyl)isovalerate), only the [2R, alpha S]-isomer (B-isomer) yielded cholesteryl [2R]-2-(4-chlorophenyl)isovalerate (CPIA-cholesterol ester) in the in vitro study using several tissue homogenates of mice, rats, dogs, and monkeys. There were species differences in the extent of CPIA-cholesterol-ester formation, with mouse tissues showing relatively higher activity than those of other animals. The kidney, brain, and spleen of mice showed relatively higher capacities to form this ester compared to other tissues, and the enzyme activity was mainly localized in microsomal fractions. The CPIA-cholesterol ester did not seem to be produced by three known biosynthetic pathways of endogenous cholesterol esters--acyl-CoA:cholesterol O-acyltransferase (ACAT), lecithin:cholesterol O-acyltransferase (LCAT), and cholesterol esterase. Carboxyesterase(s) of mouse kidney microsomes solubilized by digitonin hydrolyzed only the B alpha-isomer of fenvalerate, yielding CPIA, whereas they yielded the corresponding cholesterol ester in the presence of artificial liposomes containing cholesterol. Thus, it appears that the stereoselective formation of the CPIA-cholesterol ester results from the stereoselective formation of the CPIA-carboxyesterase complex only from the B alpha-isomer, which subsequently undergoes cleavage by cholesterol to yield the CPIA-cholesterol ester.

Prediction of biotransformation products of the fungicide fluopyram by electrochemistry coupled online to liquid chromatography-mass spectrometry and comparison with in vitro microsomal assays.

Science.gov (United States)

Mekonnen, Tessema F; Panne, Ulrich; Koch, Matthias

2018-04-01

Biotransformation processes of fluopyram (FLP), a new succinate dehydrogenase inhibitor (SDHI) fungicide, were investigated by electrochemistry (EC) coupled online to liquid chromatography (LC) and electrospray mass spectrometry (ESI-MS). Oxidative phase I metabolite production was achieved using an electrochemical flow-through cell equipped with a boron-doped diamond (BDD) electrode. Structural elucidation and prediction of oxidative metabolism pathways were assured by retention time, isotopic patterns, fragmentation, and accurate mass measurements using EC/LC/MS, LC-MS/MS, and/or high-resolution mass spectrometry (HRMS). The results obtained by EC were compared with conventional in vitro studies by incubating FLP with rat and human liver microsomes (RLM, HLM). Known phase I metabolites of FLP (benzamide, benzoic acid, 7-hydroxyl, 8-hydroxyl, 7,8-dihydroxyl FLP, lactam FLP, pyridyl acetic acid, and Z/E-olefin FLP) were successfully simulated by EC/LC/MS. New metabolites including an imide, hydroxyl lactam, and 7-hydroxyl pyridyl acetic acid oxidative metabolites were predicted for the first time in our study using EC/LC/MS and liver microsomes. We found oxidation by dechlorination to be one of the major metabolism mechanisms of FLP. Thus, our results revealed that EC/LC/MS-based metabolic elucidation was more advantageous on time and cost of analysis and enabled matrix-free detection with valuable information about the mechanisms and intermediates of metabolism processes. Graphical abstract Oxidative metabolism of fluopyram.

Changes in activities of adaptive liver enzymes in rats after non-lethal x-irradiation

International Nuclear Information System (INIS)

Toropila, M.; Ahlersova, E.; Ahlers, I.; Benova, K.

1998-01-01

The effect of a single dose of whole-body X-irradiation of 2.39 Gy (250 R) on the activities of selected adaptive rat liver enzymes and blood serum corticosterone concentrations was followed for a period of 28 days. Rats of Wistar strain SPF breeding (VELAZ Prague) were used. Both irradiated and control animals were fed in pairs with the same amount of feed as was consumed by irradiated animals in the pilot experiment. The feed intake of irradiated animals decreased significantly until the fourth day. During the rest of the experimental period no significant differences were recorded in feed intake between the experimental and control groups. The activity of tyrosine aminotransferase (TAT) in the liver of irradiated animals increased, with the exception of the initial period. Similar changes were recorded in the activity of tryptophane-2-3 dioxygenase (TO). A significant increase on the third day and a significant decrease from the seventh day after irradiation was recorded in the activity of aspartate aminotransferase (AST). Similar changes were observed with alanine aminotransferase (ALT). It is necessary to stress that the activity of this enzyme decreased also on the first day after irradiation. Until the third day there was a marked increase of serum corticosterone in the irradiated animals. The results point not only towards significant changes to the parameters observed, caused by a non-lethal irradiation dose, but also towards the importance of the nutritional regime, so-called paired feeding

In vitro metabolism studies of 18F-labeled 1-phenylpiperazine using mouse liver S9 fraction

International Nuclear Information System (INIS)

Ryu, Eun Kyoung; Choe, Yearn Seong; Kim, Dong Hyun; Ko, Bong-Ho; Choi, Yong; Lee, Kyung-Han; Kim, Byung-Tae

2006-01-01

The in vitro metabolism of 1-(4-[ 18 F]fluoromethylbenzyl)-4-phenylpiperazine ([ 18 F]1) and 1-(4-[ 18 F]fluorobenzyl)-4-phenylpiperazine ([ 18 F]2) was investigated using mouse liver S9 fraction. Results were compared to those of in vivo metabolism using mouse blood and bone and to in vitro metabolism using mouse liver microsomes. Defluorination was the main metabolic pathway for [ 18 F]1 in vitro and in vivo. Based on TLC, HPLC and LC-MS data, [ 18 F]fluoride ion and less polar radioactive metabolites derived from aromatic ring oxidation were detected in vitro, and the latter metabolites were rapidly converted into the former with time, whereas only the [ 18 F]fluoride ion was detected in vivo. Similarly, the in vitro metabolism of [ 18 F]2 using either S9 fraction or microsomes showed the same pattern as the in vivo method using blood; however, the radioactive metabolites derived from aromatic ring oxidation were not detected in vivo. These results demonstrate that liver S9 fraction can be widely used to investigate the intermediate radioactive metabolites and to predict the in vivo metabolism of radiotracers

ENZYME MARKERS ACTIVITY AND BILE FORMATION FUNCTION OF LIVER IN CASES OF TUBERCULOSTATICS AND HEXAVALENT CHROMIUM COMPOUNDS AFFECTION IN RATS

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N. I. Burmas

2016-05-01

Full Text Available Background. Currently, the growing incidence of toxic lesions of the liver is associated with industrial chemicalization and uncontrolled use of hepatotoxic drugs in everyday life. There are about one thousand drugs with high or low hepatotoxicity, such as anti-TB drugs. Objective. In this research we studied the intracellular enzymes activity and bile formation function of the liver in rats of different ages in cases of tuberculostatic (isoniazid and rifampicin affection and chromium (potassium dichromate intoxication. Methods. The experimental affection of rats of different ages was performed by combined injection of hexavalent chromium compounds (a solution of potassium dichromate, 3 mg/kg, isoniazid (0.05 g/kg and rifampicin (0.25 g/kg. On the 7th and 14th days the rats were injected with enterosorbent Sorbex (150 mg/kg. Enzyme markers activity of the liver was evaluated due to alanine and aspartate aminotransferases (ALT and AST and alkaline phosphatase (ALP rates. Bile formation function of the liver was evaluated by total bilirubin and bile acids content in blood. Results. The disorders in hepatocytes plasma membranes permeability were defined by the increased rates of ALT, AST and alkaline phosphatase in blood serum which were decreased in the liver. It was determined that total bilirubin and bile acids content in blood serum of the affected animals increased. It influenced hepatocytes excretion in bile capillaries and caused cholestasis and revenues decrease in bile. Conclusions. The most significant metabolic disorders in cases of chrome-isoniazid-rifampicin affection were defined in immature and senior animals in comparison with mature animals.

Studies on the toxic interaction between monensin and tiamulin in rats: effects on P450 activities.

Science.gov (United States)

Szücs, G; Laczay, P; Bajnógel, J; Móra, Z

2000-01-01

Studies were carried out to investigate the effects of monensin and tiamulin, and the simultaneous administration of both compounds on microsomal enzymes in rats. In Phase I of the experiments the effects of monensin and tiamulin were studied separately (monensin 10, 30, and 50 mg/kg or tiamulin 40, 120, and 200 mg/kg body weight, respectively), while in Phase II the two compounds were administered simultaneously (monesin 10 mg/kg and tiamulin 40 mg/kg b.w., respectively). When monensin was administered by itself, it exerted no significant effect on microsomal liver enzymes. In a few cases, slight inhibition of certain enzyme activities was seen. Tiamulin provoked a dose-dependent hepatic enzyme induction. The combined administration of monensin and tiamulin at low doses (10 and 40 mg/kg, respectively) resulted in marked elevation of P450-related enzyme activities. The enzyme induction was more pronounced in females than in males. The results suggest that the simultaneous administration of tiamulin may influence the biotransformation of monensin, possibly increasing the amount of reactive metabolite(s) of the ionophore antibiotic.

The Effects of Exercise on Abdominal Fat and Liver Enzymes in Pediatric Obesity: A Systematic Review and Meta-Analysis.

Science.gov (United States)

González-Ruiz, Katherine; Ramírez-Vélez, Robinson; Correa-Bautista, Jorge Enrique; Peterson, Mark D; García-Hermoso, Antonio

2017-08-01

Despite the prevalence of obesity and the multiple position stands promoting exercise for the treatment of obesity and hepatic function, a meta-analytic approach has not previously been used to examine the effects in the pediatric population. The aim of the study was to determine the effectiveness of exercise interventions on abdominal fat, liver enzymes, and intrahepatic fat in overweight and obese youth. A computerized search was made using three databases. The analysis was restricted to studies that examined the effect of supervised exercise interventions on abdominal fat (visceral and subcutaneous fat), liver enzymes (alanine aminotransferase, aspartate aminotransferase, and gamma-glutamyl transferase), and intrahepatic fat. Fourteen clinical trials (1231 youths) were eligible for inclusion in this systematic review and meta-analysis. Standardized mean difference [SMD] and 95% confidence intervals (CIs) were calculated. Exercise was associated with a significant reduction in visceral (SMD = -0.661; 95% CI, -0.976 to -0.346; p exercise programs that involve aerobic exercise longer than three sessions per week. This meta-analysis supports current recommendation for physical exercise, mainly aerobic, as an effective intervention for nonalcoholic fatty liver disease progression by targeting hepatic lipid composition, visceral and subcutaneous adipose tissue. Systematic review registration: PROSPERO CRD42016042163.

Pyrethroid Activity-Based Probes for Profiling Cytochrome P450 Activities Associated with Insecticide Interactions

Energy Technology Data Exchange (ETDEWEB)

Ismail, Hanafy M.; O' Neill, Paul M.; Hong, David; Finn, Robert; Henderson, Colin; Wright, Aaron T.; Cravatt, Benjamin; Hemingway, Janet; Paine, Mark J.

2014-01-18

Pyrethroid insecticides are used to control a diverse spectrum of diseases spread by arthropods. We have developed a suite of pyrethroid mimetic activity based probes (PyABPs) to selectively label and identify P450s associated with pyrethroid metabolism. The probes were screened against pyrethroid metabolizing and non-metabolizing mosquito P450s, as well as rodent microsomes to measure labeling specificity, plus CPR and b5 knockout mouse livers to validate P450 activation and establish the role for b5 in probe activation. Using a deltamethrin mimetic PyABP we were able to profile active enzymes in rat liver microsomes and identify pyrethroid metabolizing enzymes in the target tissue. The most reactive enzyme was a P450, CYP2C11, which is known to metabolize deltamethrin. Furthermore, several other pyrethroid metabolizers were identified (CYPs 2C6, 3A4, 2C13 and 2D1) along with related detoxification enzymes, notably UDP-g’s 2B1 - 5, suggesting a network of associated pyrethroid metabolizing enzymes, or ‘pyrethrome’. Considering the central role that P450s play in metabolizing insecticides, we anticipate that PyABPs will aid the identification and profiling of P450s associated with insecticide pharmacology in a wide range of species, improving understanding of P450-insecticide interactions and aiding the development of new tools for disease control.

Effect of diethyldithiocarbamate (DDC) and ticlopidine on CYP1A2 activity and caffeine metabolism: an in vitro comparative study with human cDNA-expressed CYP1A2 and liver microsomes.

Science.gov (United States)

Kot, Marta; Daniel, Władysława A

2009-01-01

The aim of the present study was to test the effect of diethyldithiocarbamate (DDC), which is regarded as a cytochrome P450 (CYP) CYP2A6 and CYP2E1 inhibitor, and ticlopidine, an efficient CYP2B6, CYP2C19 and CYP2D6 inhibitor, on the activity of human CYP1A2 and the metabolism of caffeine (1-N-, 3-N- and 7-N-demethylation, and C-8-hydroxylation). The experiment was carried out in vitro using human cDNA-expressed CYP1A2 (Supersomes) and human pooled liver microsomes. The effects of DDC and ticlopidine were compared to those of furafylline (a strong CYP1A2 inhibitor). A comparative in vitro study provides clear evidence that ticlopidine and DDC, applied at concentrations that inhibit the above-mentioned CYP isoforms, potently (as compared to furafylline) inhibit human CYP1A2 and caffeine metabolism, in particular 1-N- and 3-N-demethylation.

Correlation of endothelin-1 concentration and angiotensin-converting enzyme activity with the staging of liver fibrosis.

Science.gov (United States)

Kardum, Dusko; Fabijanić, Damir; Lukić, Anita; Romić, Zeljko; Petrovecki, Mladen; Bogdanović, Zoran; Jurić, Klara; Urek-Crncević, Marija; Banić, Marko

2012-06-01

Increased serum angiotensin-converting enzyme (SACE) activity and serum concentration of endothelin-1 (ET-1) were found in liver cirrhosis. We investigated a correlation between the different stages of liver fibrosis and SACE activity and serum ET-1 concentration. Seventy patients with pathohistologically established chronic liver disease were divided in three groups according to Ishak criteria for liver fibrosis: minimal fibrosis (Ishak score 0-1, n =20), medium fibrosis (Ishak score 2-5, n=20) and cirrhosis (Ishak score 6, n=30). SACE activity and ET-1 concentration were determined using commercial ELISA kits. SACE activity and ET-1 concentrations were proportional to the severity of disease, the highest being in patients with liver cirrhosis. Maximal increase in SACE activity was found between minimal and medium fibrosis while maximal increase in ET-1 concentration was revealed between medium fibrosis and cirrhosis. The analysis of the Receiver Operating Characteristic (ROC) curve for SACE activity suggested a cut-off value to separate minimal from medium fibrosis at 59.00 U/L (sensitivity 100%, specificity 64.7%). The cut-off value for serum ET-1 concentration to separate medium fibrosis from cirrhosis was 12.4 pg/mL (sensitivity 96.8%, specificity 94.4%). A positive correlation between SACE activity and ET-1 concentration was registered (Spearman's ñ = 0.438, p = 0.004). Both SACE activity and ET-1 concentration were increased in all stages of liver fibrosis. Cut-off points for SACE activity and ET-1 concentration could be a biochemical marker for the progression of fibrosis. Positive correlation between SACE activity and ET-1 concentration might indicate their interaction in the development of liver cirrhosis.

Translation of Angiotensin-Converting Enzyme 2 upon Liver- and Lung-Targeted Delivery of Optimized Chemically Modified mRNA

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Eva Schrom

2017-06-01

Full Text Available Changes in lifestyle and environmental conditions give rise to an increasing prevalence of liver and lung fibrosis, and both have a poor prognosis. Promising results have been reported for recombinant angiotensin-converting enzyme 2 (ACE2 protein administration in experimental liver and lung fibrosis. However, the full potential of ACE2 may be achieved by localized translation of a membrane-anchored form. For this purpose, we advanced the latest RNA technology for liver- and lung-targeted ACE2 translation. We demonstrated in vitro that transfection with ACE2 chemically modified messenger RNA (cmRNA leads to robust translation of fully matured, membrane-anchored ACE2 protein. In a second step, we designed eight modified ACE2 cmRNA sequences and identified a lead sequence for in vivo application. Finally, formulation of this ACE2 cmRNA in tailor-made lipidoid nanoparticles and in lipid nanoparticles led to liver- and lung-targeted translation of significant amounts of ACE2 protein, respectively. In summary, we provide evidence that RNA transcript therapy (RTT is a promising approach for ACE2-based treatment of liver and lung fibrosis to be tested in fibrotic disease models.

Curcumin Prevents Aflatoxin B1 Hepatoxicity by Inhibition of Cytochrome P450 Isozymes in Chick Liver

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Ni-Ya Zhang

2016-11-01

Full Text Available This study was designed to establish if Curcumin (CM alleviates Aflatoxin B1 (AFB1-induced hepatotoxic effects and to determine whether alteration of the expression of cytochrome P450 (CYP450 isozymes is involved in the regulation of these effects in chick liver. One-day-old male broilers (n = 120 were divided into four groups and used in a two by two factorial trial in which the main factors included supplementing AFB1 (< 5 vs. 100 μg/kg and CM (0 vs. 150 mg/kg in a corn/soybean-based diet. Administration of AFB1 induced liver injury, significantly decreasing albumin and total protein concentrations and increasing alanine aminotransferase and aspartate aminotransferase activities in serum, and induced hepatic histological lesions at week 2. AFB1 also significantly decreased hepatic glutathione peroxidase, catalase, and glutathione levels, while increasing malondialdehyde, 8-hydroxydeoxyguanosine, and exo-AFB1-8,9-epoxide (AFBO-DNA concentrations. In addition, the mRNA and/or activity of enzymes responsible for the bioactivation of AFB1 into AFBO—including CYP1A1, CYP1A2, CYP2A6, and CYP3A4—were significantly induced in liver microsomes after 2-week exposure to AFB1. These alterations induced by AFB1 were prevented by CM supplementation. Conclusively, dietary CM protected chicks from AFB1-induced liver injury, potentially through the synergistic actions of increased antioxidant capacities and inhibition of the pivotal CYP450 isozyme-mediated activation of AFB1 to toxic AFBO.

The antioxidant effects of vitamin C on liver enzymes: aspartate aminotransferase, alanine aminotranferease, alkaline phosphatase and gamma-glutamyltransferase activities in rats under Paraquat insult

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Benjamin Nnamdi Okolonkwo

2013-06-01

Full Text Available Paraquat (PQ is a bipyridylium herbicide; applied around trees in orchards and between crop rows to control broad-leaved and grassy weeds. Its oxidation results in the formation of superoxides which causes damage to cellular components. In this study, we determined the antioxidant effect vitamin C has on the liver enzymes [aspartate aminotransferase (SGOT, alanine aminotranferease (SGPT, alkaline phosphatase (ALP, and gamma-glutamyltransferase (GGT] of rats under this toxic insult. Male rats in groups (A, B, C and D were intraperitoneally injected with different sublethal increasing doses (0, 0.02, 0.04 and 0.06 g/kg body weigh of PQ respectively on monthly basis. Subsequently, the subgroups (A2, B2, C2 and D2 were given orally, 200 mg/L vitamin C, while the subgroups A1, B1, C1, and D1, received only water. Four animals per subgroup were decapitated on monthly basis and blood samples taken for enzyme assay. The parameters studied were - SGOT, SGPT, ALP and GGT - liver enzymes. The dose and time dependent PQ toxicity effect resulted in highly elevated Liver enzymes activities. The subgroups on vitamin C had significantly lower enzyme activities when compared to the same subgroups on only PQ insult. But the values were high when compared to the control subgroups (A1 and A2. These results were indication that vitamin C when given at moderate doses and maintained for a longer period could be a life saving adjunct to toxic insult.

Hemolysis, Elevated Liver Enzymes, and Low Platelets, Severe Fetal Growth Restriction, Postpartum Subarachnoid Hemorrhage, and Craniotomy: A Rare Case Report and Systematic Review

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Shadi Rezai

2017-01-01

Full Text Available Introduction. Hemolysis, elevated liver enzymes, and low platelets (HELLP syndrome is a relatively uncommon but traumatic condition occurring in the later stage of pregnancy as a complication of severe preeclampsia or eclampsia. Prompt brain computed tomography (CT or magnetic resonance imaging (MRI and a multidisciplinary management approach are required to improve perinatal outcome. Case. A 37-year-old, Gravida 6, Para 1-0-4-1, Hispanic female with a history of chronic hypertension presented at 26 weeks and 6 days of gestational age. She was noted to have hemolysis, elevated liver enzymes, and low platelets (HELLP syndrome accompanied by fetal growth restriction (FGR, during ultrasound evaluation, warranting premature delivery. The infant was delivered in stable condition suffering no permanent neurological deficit. Conclusion. HELLP syndrome is an uncommon and traumatic obstetric event which can lead to neurological deficits if not managed in a responsive and rapid manner. The central aggravating factor seems to be hypertension induced preeclamptic or eclamptic episode and complications thereof. The syndrome itself is manifested by hemolytic anemia, increased liver enzymes, and decreasing platelet counts with a majority of neurological defects resulting from hemorrhagic stroke or subarachnoid hemorrhage (SAH. To minimize adverse perinatal outcomes, obstetric management of this medical complication must include rapid clinical assessment, diagnostic examination, and neurosurgery consultation.

Atorvastatin induces bile acid-synthetic enzyme Cyp7a1 by suppressing FXR signaling in both liver and intestine in mice[S

Science.gov (United States)

Fu, Zidong Donna; Cui, Julia Yue; Klaassen, Curtis D.

2014-01-01

Statins are effective cholesterol-lowering drugs to treat CVDs. Bile acids (BAs), the end products of cholesterol metabolism in the liver, are important nutrient and energy regulators. The present study aims to investigate how statins affect BA homeostasis in the enterohepatic circulation. Male C57BL/6 mice were treated with atorvastatin (100 mg/kg/day po) for 1 week, followed by BA profiling by ultra-performance LC-MS/MS. Atorvastatin decreased BA pool size, mainly due to less BA in the intestine. Surprisingly, atorvastatin did not alter total BAs in the serum or liver. Atorvastatin increased the ratio of 12α-OH/non12α-OH BAs. Atorvastatin increased the mRNAs of the BA-synthetic enzymes cholesterol 7α-hydroxylase (Cyp7a1) (over 10-fold) and cytochrome P450 27a1, the BA uptake transporters Na+/taurocholate cotransporting polypeptide and organic anion transporting polypeptide 1b2, and the efflux transporter multidrug resistance-associated protein 2 in the liver. Noticeably, atorvastatin suppressed the expression of BA nuclear receptor farnesoid X receptor (FXR) target genes, namely small heterodimer partner (liver) and fibroblast growth factor 15 (ileum). Furthermore, atorvastatin increased the mRNAs of the organic cation uptake transporter 1 and cholesterol efflux transporters Abcg5 and Abcg8 in the liver. The increased expression of BA-synthetic enzymes and BA transporters appear to be a compensatory response to maintain BA homeostasis after atorvastatin treatment. The Cyp7a1 induction by atorvastatin appears to be due to suppressed FXR signaling in both the liver and intestine. PMID:25278499

Stimulation of NADH-dependent microsomal DNA strand cleavage by rifamycin SV.

OpenAIRE

Kukiełka, E; Cederbaum, A I

1995-01-01

Rifamycin SV is an antibiotic anti-bacterial agent used in the treatment of tuberculosis. This drug can autoxidize, especially in the presence of metals, and generate reactive oxygen species. A previous study indicated that rifamycin SV can increase NADH-dependent microsomal production of reactive oxygen species. The current study evaluated the ability of rifamycin SV to interact with iron and increase microsomal production of hydroxyl radical, as detected by conversion of supercoiled plasmid...

Effects of Artemisia dracunculus Aqueous Extract on Blood Sugar, Serum Insulin, Triglyceride and Liver Enzymes in Fructose Drinking Water Male Rats

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Mohammad Reza Shahraki

2017-02-01

Full Text Available Background Artemisia are various groups of plants which are used as an herbal medicine in all countries; the present study was designed to evaluate the effects of Artemisia dracunculus (AD leaves aqueous extract on blood sugar, serum insulin, and triglyceride and liver enzymes in Fructose Drinking water (FDW male rats. Methods At the beginning of experiment, 48 Wistar-albino male rats, weighing 200 - 250g were divided into control (C and FDW groups (n = 24. FDW group received FDW (10%, w/v for a month but control group did not receive any agents during the trial period. A half of control and FDW groups received AD L aqueous extract daily during trial period. At the end, animals were anesthetized, sacrificed and blood samples were collected from cervical vessels. Serum insulin, Blood glucose, insulin resistance index, triglyceride and liver enzymes were measured by ordinary methods. Obtained data were analyzed using SPSS-17 via one way ANOVA and Tukey tests. Results Our results showed that serum insulin, blood sugar, insulin resistance index, triglyceride, Aspartate amino transferase (AST and Alanine amino transferase (ALT values in FDW group significantly increased compared to C and C + E groups but these values in group FDW + E were significantly decreases compared to group FDW (P < 0.001. Conclusions Our findings demonstrated that AD L aqueous extract improves blood sugar, serum insulin, insulin resistance index and liver enzymes in rat model.

Subcellular distribution of styrene oxide in rat liver

International Nuclear Information System (INIS)

Pacifici, G.M.; Cuoci, L.; Rane, A.

1984-01-01

The subcellular distribution of ( 3 H)-styrene-7,8-oxide was studied in the rat liver. The compound was added to liver homogenate to give a final concentration of 2 X 10(-5); 2 X 10(-4) and 2 X 10(-3) M. Subcellular fractions were obtained by differential centrifugation. Most of styrene oxide (59-88%) was associated with the cytosolic fraction. Less than 15 percent of the compound was retrieved in each of the nuclear, mitochondrial and microsomal fractions. A considerable percentage of radioactivity was found unextractable with the organic solvents, suggesting that styrene oxide reacted with the endogenous compounds. The intracellular distribution of this epoxide was also studied in the perfused rat liver. Comparable results with those previously described were obtained. The binding of styrene oxide to the cytosolic protein was investigated by equilibrium dialysis and ultrafiltration. Only a small percentage of the compound was bound to protein

Modification of radiation-induced oxidative damage in liposomal and microsomal membrane by eugenol

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Pandey, B.N. [Radiation Biology and Health Sciences Division, Bhabha Atomic Research Centre, Mumbai 400 085 (India); Lathika, K.M. [Radiation Biology and Health Sciences Division, Bhabha Atomic Research Centre, Mumbai 400 085 (India); Mishra, K.P. [Radiation Biology and Health Sciences Division, Bhabha Atomic Research Centre, Mumbai 400 085 (India)]. E-mail: kpm@magnum.barc.ernet.in

2006-03-15

Radiation-induced membrane oxidative damage, and their modification by eugenol, a natural antioxidant, was investigated in liposomes and microsomes. Liposomes prepared with DPH showed decrease in fluorescence after {gamma}-irradiation, which was prevented significantly by eugenol and correlated with magnitude of oxidation of phospholipids. Presence of eugenol resulted in substantial inhibition in MDA formation in irradiated liposomes/microsomes, which was less effective when added after irradiation. Similarly, the increase in phospholipase C activity observed after irradiation in microsomes was inhibited in samples pre-treated with eugenol. Results suggest association of radio- oxidative membrane damage with alterations in signaling molecules, and eugenol significantly prevented these membrane damaging events.

Quantitative liver functions in Turner syndrome with and without hormone replacement therapy

DEFF Research Database (Denmark)

Gravholt, Claus Højbjerg; Poulsen, H.E.; Ott, Peter

2007-01-01

Studies have documented elevated levels of liver enzymes in many females with Turner syndrome (TS). Histology has shown a range of changes. Treatment with female hormone replacement therapy (HRT) reduces liver enzymes.......Studies have documented elevated levels of liver enzymes in many females with Turner syndrome (TS). Histology has shown a range of changes. Treatment with female hormone replacement therapy (HRT) reduces liver enzymes....

An in-vitro cocktail assay for assessing compound-mediated inhibition of six major cytochrome P450 enzymes

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Jing-Jing Wang

2014-08-01

Full Text Available An efficient screening assay was developed and validated for simultaneous assessment of compound-mediated inhibition of six major human cytochrome P450 (CYP enzymes. This method employed a cocktail of six probe substrates (i.e., phenacetin, amodiaquine, diclofenac, S-mephenytoin, dextromethorphan and midazolam for CYP1A2, 2C8, 2C9, 2C19, 2D6 and 3A4, respectively as well as individual prototypical inhibitors of the six CYP enzymes in human liver microsomes under optimized incubation conditions. The corresponding marker metabolites (i.e., acetaminophen, N-desethylamodiaquine, 4-OH-diclofenac, 4-OH-S-mephenytoin, dextrorphan and 1-OH-midazolam in the incubates were quantified using LC–MS/MS methods either by an internal standard (IS calibration curve or a simplified analyte-to-IS peak area ratio approach. The results showed that the IC50 values determined by the cocktail approach were in good agreement with those obtained by the individual substrate approach as well as those reported in the literature. Besides, no remarkable difference was observed between the two quantification approaches. In conclusion, this new cocktail assay can be used for reliable screening of compound-mediated CYP inhibition. Keywords: LC–MS/MS, Cytochrome P450, Cocktail-probe, Inhibition assessment, Drug screenning

Regular exercise is associated with a reduction in the risk of NAFLD and decreased liver enzymes in individuals with NAFLD independent of obesity in Korean adults.

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Ji Cheol Bae

Full Text Available BACKGROUND: We evaluated the association of regular physical exercise with the presence of non-alcoholic fatty liver disease (NAFLD and liver enzymes in relation to obesity and insulin resistance. METHODOLOGY/PRINCIPAL FINDINGS: A cross-sectional analysis was conducted in 72,359 healthy Korean adults without diabetes who participated in a comprehensive health check-up. Subjects who have been exercising regularly (more than 3 times per week, at least for 30 minutes each time and for consecutive 3 month were categorized into exercise group. All subjects were categorized into deciles based on their body mass index (BMI and we estimated the odds ratios (ORs for having NAFLD according to exercise regularity in each decile. The diagnosis of NAFLD was based on ultrasonography findings. Individuals with NAFLD (n = 19,921 were analyzed separately to evaluate ORs for having elevated liver enzymes based on regularity of exercise. The risk for NAFLD was significantly reduced in exercise group with age- and sex-adjusted ORs of 0.53-0.72 for all BMI deciles except at BMI categories of <19.6 and 20.7-21.6 kg/m(2. While no difference was seen in BMI between subjects in exercise and non-exercise group across the BMI deciles, the values of body fat percentage and metabolic risk factors differed. Among NAFLD patients, subjects in exercise group had a lower risk for having elevated liver enzymes with multivariable adjusted OR of 0.85 (95% CI 0.74-0.99, for AST and 0.74 (95% CI 0.67-0.81, for ALT than did subjects in non-exercise group. CONCLUSIONS/SIGNIFICANCE: Regular exercise was associated with a reduced risk for having NAFLD and decreased liver enzymes in patients with NAFLD, and this relationship was also independent of obesity.

Vitamin D status, liver enzymes, and incident liver disease and mortality

DEFF Research Database (Denmark)

Skaaby, Tea; Husemoen, Lise Lotte Nystrup; Borglykke, Anders

2014-01-01

, alcohol consumption, smoking, physical activity, dietary habits, education, body mass index, and ALT). The risk of having a high level of ALT, AST, or GGT tended to be higher for lower vitamin D levels, although not statistically significant. In this general population study, vitamin D status...... was inversely associated with incident liver disease. Further studies are needed to determine whether patients in risk of developing impaired liver function should be screened for vitamin D deficiency for preventive purposes....

Photoeffects of near ultraviolet light upon a polycyclic aromatic hydrocarbon exposed to mouse skin microsomes

International Nuclear Information System (INIS)

Peirano, W.B.

1991-01-01

Near ultraviolet (UV) light has been reported to both enhance and inhibit the tumor incidence in mice dermally exposed to benzo(a)pyrene (BaP) or polycyclic aromatic hydrocarbon (PAH) mixtures. Near UV light interacts with PAHs producing a variety of oxygenated products such as phenols, endoperoxides and quinones. However, little is known about BaP products formed from near UV irradiation of BaP-exposed mouse skin. Therefore, 14 C-BaP was incubated with 3-methylcholanthrene (3-MC) induced C 3 H/HeJ and DBA/2J mouse skin microsomes with or without a 365 nm light source. The results indicated that the concurrent 365 nm light irradiation of induced mouse skin microsomes and BaP greatly enhanced the total conversion of BaP to its products, approximately 3-fold for the C 3 H/HeJ and approximately 7-fold for the DBA/2J mouse microsomes, compared to the induced mouse skin microsomes and BaP alone. HPLC analyses of organic extracts indicated a more than additive enhancement of the formation of most of the individual cochromatographed BaP metabolites due to the combined interaction of 365 nm light with BaP and skin microsomes. Similar interactions were observed using benz(a)anthracene (BaA) in this system. These data show that near UV light alters the metabolic profile of PAHs produced by mouse skin microsomes

Responses of the antioxidative and biotransformation enzymes in the aquatic fungus Mucor hiemalis exposed to cyanotoxins.

Science.gov (United States)

Balsano, Evelyn; Esterhuizen-Londt, Maranda; Hoque, Enamul; Lima, Stephan Pflugmacher

2017-08-01

To investigate antioxidative and biotransformation enzyme responses in Mucor hiemalis towards cyanotoxins considering its use in mycoremediation applications. Catalase (CAT), glutathione reductase (GR), and glutathione peroxidase (GPx) in M. hiemalis maintained their activities at all tested microcystin-LR (MC-LR) exposure concentrations. Cytosolic glutathione S-transferase (GST) activity decreased with exposure to 100 µg MC-LR l -1 while microsomal GST remained constant. Cylindrospermopsin (CYN) at 100 µg l -1 led to an increase in CAT activity and inhibition of GR, as well as to a concentration-dependent GPx inhibition. Microsomal GST was inhibited at all concentrations tested. β-N-methylamino-L-alanine (BMAA) inhibited GR activity in a concentration-dependent manner, however, CAT, GPx, and GST remained unaffected. M. hiemalis showed enhanced oxidative stress tolerance and intact biotransformation enzyme activity towards MC-LR and BMAA in comparison to CYN, confirming its applicability in bioreactor technology in terms of viability and survival in their presence.

Metabolic activation of the bladder carcinogen 4-nitrobiphenyl (NBP)

International Nuclear Information System (INIS)

Swaminathan, S.

1986-01-01

The metabolism of NBP, a dog bladder carcinogen, was examined in vitro using rat liver tissues. NBP was metabolized by enzymes localized both in the microsomes and cytosol. The microsomal enzyme activity was inducible by Aroclor 1254 and phenobarbital. High pressure liquid chromatography analysis of the ethyl acetate extract of the reaction mixture, following incubation of [ 3 H]NBP with NADPH and microsomes, revealed four radioactive and UV absorbing peaks with retention times of 5, 8, 14 and 28 min. The peaks at 8, 14 and 28 min corresponded with 4-aminobiphenyl (ABP), NBP and azoxy biphenyl, respectively. The early eluting component with a retention time of 5 min has been tentatively identified as a ring hydroxylated derivative. In contrast to microsomal metabolism, cytosol-mediated metabolism yielded only one major metabolite identified as ABP. Cytosol-mediate reduction was inhibited by the xanthine oxidase inhibitor allopurinol. In vitro incubation of NBP with NADH and commercial preparations of xanthine oxidase also yielded ABP and the formation of the latter was blocked by allopurinol. Xanthine oxidase catalyzed also the binding of [ 3 H]NBP to DNA and proteins; the binding was inhibited by allopurinol. These data support the hypothesis that the nitro reduction step is involved in the activation of the bladder carcinogen NBP, and that the nitroreductases occur in both the microsomes and cytosol. The cytosolic activity is primarily due to xanthine oxidase

Drug-induced liver injury associated with HIV medications.

Science.gov (United States)

Jain, Mamta K

2007-08-01

Antiretroviral therapy (ART) for HIV infection frequently has been associated with elevated liver enzyme levels. Determining the cause of elevated liver enzyme levels in patients who have HIV is difficult because ART usually consists of three different drugs, patients may be taking additional hepatotoxic medications and patients who have HIV often suffer from other liver diseases. Several agents, however, are recognized as having noteworthy and specific patterns of toxicity. This article reviews the different HIV drug classes, incidence of elevated liver enzyme values by class and by individual drug, risk factors, specific toxicities, and possible mechanisms of injury.

Abetalipoproteinemia: A novel mutation of microsomal triglyceride ...

African Journals Online (AJOL)

Hager Barakizou

2016-01-25

Jan 25, 2016 ... Abetalipoproteinemia: A novel mutation of microsomal triglyceride transfer protein (MTP) gene in a young Tunisian patient. Hager Barakizou a,. *, Souha Gannouni a. , Khalil Messaoui a. , Mathilde Difilippo b. ,. Agne`s Sassolas b. , Fethi Bayoudh a a Department of Pediatrics, Military Hospital of Tunis, ...

Inhibitory activity of diacylglycerol acyltransferase (DGAT) and microsomal triglyceride transfer protein (MTP) by the flavonoid, taxifolin, in HepG2 cells: potential role in the regulation of apolipoprotein B secretion.

Science.gov (United States)

Casaschi, Adele; Rubio, Brent K; Maiyoh, Geoffrey K; Theriault, Andre G

2004-10-01

The purpose of the present study was to examine the role of taxifolin, a plant flavonoid, on several aspects involving apolipoprotein B (apoB) secretion and triglyceride (TG) availability in HepG2 cells. Taxifolin was shown by ELISA to markedly reduce apoB secretion under basal and lipid-rich conditions up to 63% at 200 micromol/L. As to the mechanism underlying this effect, we examined whether taxifolin exerted its effect by limiting TG availability in the microsomal lumen essential for lipoprotein assembly. Taxifolin was shown to inhibit microsomal TG synthesis by 37% and its subsequent transfer into the lumen (-26%). The reduction in synthesis was due to a decrease in diacylglycerol acyltransferase (DGAT) activity (-35%). The effect on DGAT activity was found to be non-competitive and non-transcriptional in nature. Both DGAT-1 and DGAT-2 mRNA expression remained essentially unchanged suggesting the point of regulation may be at the post-transcriptional level. Evidence is accumulating that microsomal triglyceride transfer protein (MTP) is also involved in determining the amount of lumenal TG available for lipoprotein assembly and secretion. Taxifolin was shown to inhibit this enzyme by 41%. Whether the reduction in TG accumulation in the microsomal lumen is predominantly due to DGAT and/or MTP activity remains to be addressed. In summary, taxifolin reduced apoB secretion by limiting TG availability via DGAT and MTP activity.

Interrelationship of dietary lipids and ascorbic acid with hepatic enzymes of cholesterol metabolic pathway.

Science.gov (United States)

Sen, S; Mukherjee, S

1997-01-01

Effect of unsaturated and saturated fats on cholesterol metabolism was studied in ascorbate sufficient and deficient guineapigs. Experimental animals were made chronic ascorbic acid deficient by allowing oral intake of 0.5 mg ascorbic acid/day/animal. Elevation in serum and liver cholesterol and triglyceride along with depression in cholesterol oxidation and 7 alpha-hydroxylation in liver was observed in unsaturated fat fed guineapigs with ascorbate deficiency. Liver microsomal cytochrome P-450 level was found to be low in ascorbate deficient animals. Polyunsaturated fat intake could not lower the serum cholesterol level in ascorbate deficiency. Today polyunsaturated fat in the diet is encouraged all over the world for its hypocholesterolemic effect. This study indicates that polyunsaturated fat intake with ascorbic acid deficiency may produce hypercholesterolemia.

Dichloromethane and carbon monoxide inhalation: carboxyhemoglobin addition, and drug metabolizing enzymes in rat

Energy Technology Data Exchange (ETDEWEB)

Kurppa, K.; Kivistoe, H.; Vainio, H.

1981-09-01

Male Wistar rats were exposed for 3 h to 100 ppm CO, 1,000 ppm dichloromethane, or to their combination. Exposure to dichloromethane alone or in combination with CO doubled the ethoxycoumarin O-deethylase activity in the kidney microsomes but not in the liver. An additive effect on blood COHb concentration by simultaneous exposure to CO and dichloromethane was observed. The mechanism of the additive effect is discussed.

Development of monoclonal antibodies to human microsomal epoxide hydrolase and analysis of “preneoplastic antigen”-like molecules

Energy Technology Data Exchange (ETDEWEB)

Duan, Hongying [Department of Microbiology, Faculty of Medicine, Saitama Medical University, Moroyama-cho, Iruma-gun, Saitama 350-0495 (Japan); Yoshimura, Kazunori [Department of Physiology, Faculty of Medicine, Saitama Medical University, Moroyama-cho, Iruma-gun, Saitama 350-0495 (Japan); Kobayashi, Nobuharu; Sugiyama, Kazuo [Department of Microbiology, Faculty of Medicine, Saitama Medical University, Moroyama-cho, Iruma-gun, Saitama 350-0495 (Japan); Sawada, Jun-ichi; Saito, Yoshiro [Division of Biochemistry and Immunochemistry, National Institute of Health Sciences, Kamiyoga 1-18-1, Setagaya-ku, Tokyo 158-8501 (Japan); Morisseau, Christophe; Hammock, Bruce D. [Department of Entomology and Cancer Center, University of California, Davis, One Shields Avenue, Davis, CA 95616-8584 (United States); Akatsuka, Toshitaka, E-mail: akatsuka@saitama-med.ac.jp [Department of Microbiology, Faculty of Medicine, Saitama Medical University, Moroyama-cho, Iruma-gun, Saitama 350-0495 (Japan)

2012-04-01

Microsomal epoxide hydrolase (mEH) is a drug metabolizing enzyme which resides on the endoplasmic reticulum (ER) membrane and catalyzes the hydration of reactive epoxide intermediates that are formed by cytochrome P450s. mEH is also thought to have a role in bile acid transport on the plasma membrane of hepatocytes. It is speculated that efficient execution of such multiple functions is secured by its orientation and association with cytochrome P450 enzymes on the ER membrane and formation of a multiple transport system on the plasma membrane. In certain disease status, mEH loses its association with the membrane and can be detected as distinct antigens in the cytosol of preneoplastic foci of liver (preneoplastic antigen), in the serum in association with hepatitis C virus infection (AN antigen), or in some brain tumors. To analyze the antigenic structures of mEH in physiological and pathological conditions, we developed monoclonal antibodies against different portions of mEH. Five different kinds of antibodies were obtained: three, anti-N-terminal portions; one anti-C-terminal; and one, anti-conformational epitope. By combining these antibodies, we developed antigen detection methods which are specific to either the membrane-bound form or the linearized form of mEH. These methods detected mEH in the culture medium released from a hepatocellular carcinoma cell line and a glioblastoma cell line, which was found to be a multimolecular complex with a unique antigenic structure different from that of the membrane-bound form of mEH. These antibodies and antigen detection methods may be useful to study pathological changes of mEH in various human diseases. -- Highlights: ► Monoclonal antibodies against different portions of mEH were developed. ► They discriminate between the membrane-bound and the linearized forms of mEH. ► We analyze the antigenic structure of the altered form of mEH in tumor cells. ► Preneoplastic antigen is a multimolecular complex of mEH with

Effects of sh-reagents on rat hepatic aldehyde dehydrogenase activity

Energy Technology Data Exchange (ETDEWEB)

Konoplitskaya, K.L.; Kuz' mina, G.I.; Grigor' yeva, M.V.; Poznyakova, T.N.

The liver serves as the primary organ for the oxidation of ingested ethanol via a pathway involving alcohol- and aldehyde dehydrogenase. In view of the problem of alcoholism, three enzymes are of particular interest in understanding the biochemical mechanism that may be involved in alcohol addiction and in the formulation of therapeutic approaches. While alcohol dehydrogenase has been studied in considerable detail, current attention is centered on aldehyde dehydrogenase. A comparative analysis of the effects of a series of SH-active reagents - tetraethylthiuram disulfide (TETD), 5,5-dithiobisnitrobenzoic acid (DTNB), p-chloromercurybenzoate (PCMB), and N-ethylmaleimide (NEM) - were tested for their effects on the activity of aldehyde dehydrogenase of the hepatic mitochondrial (isozymes I and II) and microsomal (isozyme II) fractions of outbred albino rats. DTNB was found to be inhibited by 100 and 50% mitochondrial isozymes I and II, respectively, and by 20%, the microsomal enzyme under the conditions employed. DTNB and NEM inhibited by 30 and 50% isozymes I and II of the mitochondria, but had no effect on the microsomal isozyme. 24 references, 3 figures.