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Sample records for hepatic enzyme activities

  1. Antioxidant enzyme activities in hepatic tissue from children with chronic cholestatic liver disease

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    Ismail Nagwa

    2010-01-01

    Full Text Available Background/Aim: To study the oxidative stress status in children with cholestatic chronic liver disease by determining activities of glutathione peroxidase (GPx, superoxide dismutase (SOD and catalase (CAT in liver tissue. Materials and Methods: A total of 34 children suffering from cholestatic chronic liver disease were studied. They were selected from the Hepatology Clinic, Cairo University, and compared with seven children who happened to have incidental normal liver biopsy. The patients were divided into three groups: extrahepatic biliary atresia (n=13, neonatal hepatitis (n=15 and paucity of intrahepatic bile ducts (n=6; GPx, SOD and CAT levels were measured in fresh liver tissue using ELISA . Results: In the cholestatic patients, a significant increase was found in mean levels of SOD, GPx and CAT in hepatic tissue compared to control children. The three enzymes significantly increased in the extrahepatic biliary atresia group, whereas in the groups of neonatal hepatitis and paucity of intrahepatic bile ducts, only GPx and CAT enzymes were significantly increased. Conclusion: Oxidative stress could play a role in the pathogenesis of cholestatic chronic liver diseases. These preliminary results are encouraging to conduct more extensive clinical studies using adjuvant antioxidant therapy.

  2. [Effect of Arnica montana tincture on some hydrolytic enzyme activities of rat liver in experimental toxic hepatitis].

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    Iaremiĭ, I M; Meshchyshen, I F; Hrihor'ieva, N P; Kostiuk, L S

    1998-01-01

    Effects of tinctura arnica on arginase, adenosine triphosphatase, glucose-6-phosphatase and 5'-nucleotidase activities of rats liver in case of experimental toxic hepatitis have been studied. Toxic hepatitis was caused by 2 times interstomach administration of 0.25 ml oil solution of carbon tetrachloride per 100 g of animal weight. 20 mkl/100 g of tinctura arnica was administered every day per os for 14 days. The enzyme activities have been investigated at 3, 7 and 17 days. A significant demention of a studied hydrolytic enzyme activities in rats liver at intoxication of the body by CCI4 has been shown. It has been established that tinctura arnica administered per os to intoxicated animals sped up the normalization of hydrolytic enzyme activities in rat liver.

  3. D-pinitol attenuates the impaired activities of hepatic key enzymes in carbohydrate metabolism of streptozotocin-induced diabetic rats.

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    Sivakumar, Selvaraj; Subramanian, Sorimuthu P

    2009-09-01

    During diabetes mellitus, endogenous hepatic glucose production is increased as a result of impaired activities of the key enzymes of carbohydrate metabolism, which leads to the condition known as hyperglycemia. D-pinitol, a bioactive constituent isolated from soybeans, has been shown to reduce hyperglycemia in experimental diabetes. We therefore designed this study to investigate the effect of oral administration of D-pinitol (50 mg/kg b. w. for 30 days) on the activities of key enzymes in carbohydrate and glycogen metabolism in the liver tissues of streptozotocin-induced diabetic rats. The efficacy was compared with glyclazide, a standard hypoglycemic drug. Oral administration of D-pinitol to diabetic group of rats showed a marked decrease in the levels of blood glucose, glycosylated hemoglobin and an increase in plasma insulin and body weight. The activities of the hepatic enzymes such as hexokinase, pyruvate kinase, glucose-6-phosphate dehydrogenase, glycogen synthase and hepatic glycogen content were significantly (p pinitol. The results suggest that alterations in the activities of key metabolic enzymes of carbohydrate metabolism could be one of the biochemical rationale by which D-pinitol attenuates the hyperglycemic effect in diabetic rats.

  4. Di2-ethylhexyl phthalate disrupts thyroid hormone homeostasis through activating the Ras/Akt/TRHr pathway and inducing hepatic enzymes

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    Ye, Hanfeng; Ha, Mei; Yang, Min; Yue, Ping; Xie, Zhengyuan; Liu, Changjiang

    2017-01-01

    Di(2-ethylhexyl) phthalate (DEHP), as a widespread environmental pollutant and an endocrine disruptor, can disturb the homeostasis of thyroid hormones (THs). In order to elucidate roles of the MAPK and PI3K/Akt pathways and hepatic enzymes in thyroid-disrupting effects of DEHP, Sprague-Dawley rats were dosed with DEHP by gavage for 30 consecutive days; Nthy-ori 3-1 cells were treated with DEHP with NAC, k-Ras siRNA or inhibitors (U0126 and wortmannin). Results showed that DEHP led to histopathologic changes in rat thyroid and liver, such as the decrease in thyroid follicular cavity diameter, hepatocyte edema. Triiodothyronine (T3), thyroxine (T4) and thyrotropin releasing hormone (TRH) were reduced. DEHP caused ROS production, oxidative stress and k-Ras upregulation, thereby activating the ERK and Akt pathways in vivo and in vitro. Moreover, TRH receptor (TRHr) level was elevated after the activation of the Akt pathway and was downregulated after the inhibition of the Akt pathway. However, TRHr was not modulated by the ERK pathway. Additionally, hepatic enzymes, including Ugt1a1, CYP2b1, Sult1e1, and Sult2b1, were significantly induced after DEHP exposure. Taken together, DEHP can perturb TH homeostasis and reduce TH levels. The activated Ras/Akt/TRHr pathway and induced hepatic enzymes play vital roles in thyroid-disrupting effects of DEHP. PMID:28065941

  5. Effect of curcumin on hepatic antioxidant enzymes activities and gene expressions in rats intoxicated with aflatoxin B1.

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    El-Bahr, S M

    2015-01-01

    Twenty-eight rats were examined in a 5-week experiment to investigate the effect of curcumin on gene expression and activities of hepatic antioxidant enzymes in rats intoxicated with aflatoxin B1 (AFB1 ). The rats were divided into four groups. Rats in 1-4 groups served as control, oral curcumin treated (15 mg/kg body weight), single i.p. dose of AFB1 (3 mg/kg body weight) and combination of single i.p. dose of AFB1 with oral curcumin treated, respectively. AFB1 Liver damage and oxidative stress were evident in untreated AFB1 -intoxicated rats as indicated by a significant elevation in hepatic transaminases, elevation in lipid peroxide biomarkers (thiobarbituric acid reactive substances; TBARS), reduction of reduced glutathione (GSH) concentration, reduction in the activities of antioxidant enzymes namely catalase (CAT), total superoxide dismutase (SOD), glutathione peroxidase (GPX) and glutathione-S-transferase (GST) and down-regulation of gene expression of these antioxidant enzymes compared to control. Liver sections of rats intoxicated with AFB1 showed a disrupted lobular architecture, scattered necrotic cells and biliary proliferation. Administration of curcumin with AFB1 resulted in amelioration of AFB1 -induced effects compared to untreated AFB1 -intoxicated rats via an up-regulation of antioxidant enzyme gene expression, activation of the expressed genes and increase in the availability of GSH.

  6. Activity and mRNA Levels of Enzymes Involved in Hepatic Fatty Acid Synthesis in Rats Fed Naringenin.

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    Hashimoto, Toru; Ide, Takashi

    2015-11-04

    We investigated the physiological activity of naringenin in affecting hepatic lipogenesis and serum and liver lipid levels in rats. Rats were fed diets containing 0, 1, or 2.5 g/kg naringenin for 15 d. Naringenin at a dietary level of 2.5 g/kg significantly decreased the activities and the mRNA levels of various lipogenic enzymes and sterol regulatory element binding protein-1c (SREBP-1c) mRNA level. The activities and the mRNA levels were also 9-22% and 12-38% lower, respectively, in rats fed a 1 g/kg naringenin diet than in the animals fed a naringenin-free diet, although the differences were not significant in many cases. Naringenin at 2.5 g/kg significantly lowered serum triacylglycerol, cholesterol, and phospholipid and hepatic triacylglycerol and cholesterol. This flavonoid at 1.0 g/kg also significantly lowered these parameters except for serum triacylglycerol. Naringenin levels in serum and liver dose-dependently increased, and hepatic concentrations reached levels that can affect various signaling pathways.

  7. Clinical significance of activity of ALT enzyme in patients with hepatitis C virus

    Institute of Scientific and Technical Information of China (English)

    2007-01-01

    AIM: To investigate serum alanine aminotransferase (ALT) levels in relation to the clinical, biochemical,ultrasonographic and histological characteristics of patients with hepatitis C virus.METHODS: Duration of disease, HCV-RNA, liver steatosis, and the hepatitis activity index (HAI) were correlated with serum ALT in 36 patients with HCV. ALT values were also investigated in 16 control subjects without any liver diseases.RESULTS: In bivariate analyses, ALT levels correlated with duration of HCV infection (P< 0.01), HCV-RNA (P<0.05), and the HAI(P<0.01). Among the components of the HAI, ALT concentrations were significantly associated with pedportal bridging/necrosis (P<0.01) and fibrosis (P<0.05). In multivariate analysis, periportal bridging/necrosis (β = 0.508; P < 0.01), duration of HCV infection (β = 0.413; P < 0.01), and HCV-RNA (β= 0.253; P < 0.05)were independently associated with ALT activity. The normal ALT activity for men and women was < 23 IU/L and < 22 IU/L, respectively.CONCLUSION: In patients with HCV, alterations in the liver tissue as reflected by ALT elevation are mainly associated with periportal bridging/necrosis, viral load and duration of disease. A cut-off value < 23 IU/L distinguished with high diagnostic accuracy healthy controls from patients with HCV.

  8. Thyroid hormone levels and hepatic enzyme activity in lactating dams after gestational exposure to low dose PBDE 47

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    Kuriyama, S.N.; Grande, S.W.; Akkoc, Z.; Souza, C.A.M. de; Chahoud, I. [Charite Univ. Medical School Berlin (Germany). Inst. of Clinical Pharmacology and Toxicology, Dept. Toxicology, Campus Benjamin Franklin; Fidalgo-Neto, A.A. [Oswaldo Cruz Foundation, Rio de Janeiro (Brazil). Lab. of Environmental Toxicology

    2004-09-15

    Polybrominated diphenyl ethers (PBDEs), a class of widely used flame retardants, are found extensively in the environment (shown by several studies on sentinel animal species), as well as in humans. In rodents, technical commercial PBDE mixtures and individual congeners have shown to interfere with thyroid hormone homeostasis, produce a mix-type induction of hepatic microsomal enzymes, disrupt spontaneous behaviour, impair learning and memory and alter the cholinergic transmitter system. In rat and mice, some technical PBDE commercial mixtures such as DE-71 and Bromkal 70 and the congener PBDE 47 have shown to decrease circulating thyroid hormone levels. PBDEs are also able to induce both hepatic phase I and phase II detoxification enzymes, demonstrated by several investigations in laboratory animals. For example, induction of ethoxyresorufin-O-deethylase (EROD), pentoxyresorufin-Odespenthylase (PROD) and uridinediphospho-glucuronosyltransferase (UDPGT) has been shown in rodents and cell lines after exposure to technical mixtures or individual congeners. However, these studies deal with doses much higher than that found in human tissues, highlighting the importance of assessing the adverse effects of doses close to human exposure levels. PBDE 47 is the most predominant congener found in environmental and human samples (including human milk) and, therefore, hazard identification is extremely important for human risk assessment. We administered a single dose to gravid dams on gestation day 6 of either 140 {mu}g/kg BW or 700 {mu}g/kg BW of the congener, 2,2'4,4'-tetrabromo diphenyl ether (PBDE 47). These doses are pertinent to human exposure levels because a study by She et al. found a mean level of 33.3 {mu}g PBDE 47 /kg fat in human breast adipose tissue with a range from 7.01 to 196 {mu}g PBDE 47 /kg fat. In this study, thyroid hormone levels and hepatic enzyme activity were evaluated in lactating dams after in utero administration of low dose PBDE 47.

  9. Thalidomide increases human hepatic cytochrome P450 3A enzymes by direct activation of the pregnane X receptor.

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    Murayama, Norie; van Beuningen, Rinie; Suemizu, Hiroshi; Guguen-Guillouzo, Christiane; Shibata, Norio; Yajima, Kanako; Utoh, Masahiro; Shimizu, Makiko; Chesné, Christophe; Nakamura, Masato; Guengerich, F Peter; Houtman, René; Yamazaki, Hiroshi

    2014-02-17

    Heterotropic cooperativity of human cytochrome P450 (P450) 3A4/3A5 by the teratogen thalidomide was recently demonstrated by H. Yamazaki et al. ( ( 2013 ) Chem. Res. Toxicol. 26 , 486 - 489 ) using the model substrate midazolam in various in vitro and in vivo models. Chimeric mice with humanized liver also displayed enhanced midazolam clearance upon pretreatment with orally administered thalidomide, presumably because of human P450 3A induction. In the current study, we further investigated the regulation of human hepatic drug metabolizing enzymes. Thalidomide enhanced levels of P450 3A4 and 2B6 mRNA, protein expression, and/or oxidation activity in human hepatocytes, indirectly suggesting the activation of upstream transcription factors involved in detoxication, e.g., the nuclear receptors pregnane X receptor (PXR) and constitutive androstane receptor (CAR). A key event after ligand binding is an alteration of nuclear receptor conformation and recruitment of coregulator proteins that alter chromatin accessibility of target genes. To investigate direct engagement and functional alteration of PXR and CAR by thalidomide, we utilized a peptide microarray with 154 coregulator-derived nuclear receptor-interaction motifs and coregulator and nuclear receptor boxes, which serves as a sensor for nuclear receptor conformation and activity status as a function of ligand. Thalidomide and its human proximate metabolite 5-hydroxythalidomide displayed significant modulation of coregulator interaction with PXR and CAR ligand-binding domains, similar to established agonists for these receptors. These results collectively suggest that thalidomide acts as a ligand for PXR and CAR and causes enzyme induction leading to increased P450 enzyme activity. The possibilities of drug interactions during thalidomide therapy in humans require further evaluation.

  10. Effect of insecticides on vital activity, hepatic enzymes and red blood cell acetyl cholinesterase activity of rabbits in Makkah.

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    Sawas, A H

    1998-05-01

    Developing animals and invertebrate are markedly more sensitive to acute toxicity through exposure to insecticides. Varieties of insecticides are used for hygienic control in Makkah holy places. The present study examines the acute effects of commonly used insecticides in Makkah area. Rosfin as an organophosphorus, Airlen as a pyrethroid and Sulvac as a carbamate derivative were tested for their effects on vital activities, hepatic transaminases, serum triglycerides and acetyl cholinesterase activity of rabbits. The insecticides were tested in same and double concentration used for insect control by Makkah's municipal authorities. Compressed air was used as a source of pressure for spraying wooden boxes designed for habitation of animals during the experiments. There were no significant changes in vital activities of rabbits in both concentrations. However, serum glutamate pyuvate transaminase (SGPT) and serum glutamate oxaloacetate transaminase (SGOT) showed irregular changes (mild decrease or increase) in all groups, while triglyceride showed mild rise after six days exposure in case of double concentration. Acetyl cholinesterase showed mild increase in activity after five minutes incubation time, but there was unnoticed increase in activity after 15, 25, 35 and 45 minutes of incubation. In case of Airlen, the activity increased after five minutes of incubation and decreased thereafter. In conclusion, insecticides used in the holy places of Makkah area have no apparent effects on vital activity, acetyl cholinesterase activity and showed no significant effect on rabbit hepatic transaminases and serum triglyceride.

  11. Hepatic fatty acid oxidation : activity, localization and function of some enzymes involved

    NARCIS (Netherlands)

    A. van Tol (Arie)

    1971-01-01

    textabstractFatty acid oxidation is an important pathway for energy production in mammals and birds. In animal tissues the enzymes of fatty acid oxidation are located in the mitochondrion. Recent reports suggest that this is not the case in Castor bean endosperm. In this tissue the enzymes of B-oxid

  12. Hepatic fatty acid oxidation : activity, localization and function of some enzymes involved

    NARCIS (Netherlands)

    A. van Tol (Arie)

    1971-01-01

    textabstractFatty acid oxidation is an important pathway for energy production in mammals and birds. In animal tissues the enzymes of fatty acid oxidation are located in the mitochondrion. Recent reports suggest that this is not the case in Castor bean endosperm. In this tissue the enzymes of

  13. Hepatic fatty acid oxidation : activity, localization and function of some enzymes involved

    NARCIS (Netherlands)

    A. van Tol (Arie)

    1971-01-01

    textabstractFatty acid oxidation is an important pathway for energy production in mammals and birds. In animal tissues the enzymes of fatty acid oxidation are located in the mitochondrion. Recent reports suggest that this is not the case in Castor bean endosperm. In this tissue the enzymes of B-oxid

  14. Effect of Alcoholic Extract of Rosa canina on Hepatic Tissue and Hepatic Enzymes Activity in Diabetic Rats

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    Seyedmahdi Banan Khojasteh

    2017-04-01

    Full Text Available Background & aim: Nowadays because of difficulty in the supply and injection of insulin and blood sugar reducing drugs and considering the side effects of chemical drugs, researcher’s attention is drawn to using herbal medicines. Liver is one of the organs affected by diabetes in different functional aspects. The purpose of this research was studying the effect of Rosa canina extract on reducing the adverse effects and improving diabetes symptoms related to histophysiology, amount of biochemical serum factors and liver enzymes in male rats. Methods: 32 male Wistar rats, weighed 200-220 gr, were divided into 4 groups. Diabetes was induced by intraperitoneal injection of STZ (60mg/kg. The period of experiments was 4 weeks. At the end of experiments, the rats were sacrificed and their serums were collected for measurement of liver enzymes. Also, histological sampling of liver was done. Statistical analysis was performed using analysis of variance (one-way ANOVA. Results: The results of this study showed that treatment with ethanol extract of Rosa canina led to significant reduction of liver enzymes in the Rosa extract-treated groups in comparison with the diabetic group, indicating the protective effect of alcoholic extract of Rosa canina on liver tissue against diabetes-induced damages. In liver histopathology, a distinct fat change in the lobular center areas was created in the diabetic group. Pathological changes were not significant in the liver of the Rosa extract-treated groups; liver in this group was relatively healthier than in diabetic group. Conclusions: Rosa extract improved this organ's function in STZ-induced diabetic rats by reducing serum biomarkers of liver pathology and reducing histological damages.

  15. Characterization of hepatic enzyme activity in older adults with dementia: potential impact on personalizing pharmacotherapy

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    Campbell NL

    2015-01-01

    Full Text Available Noll L Campbell,1–4 Todd C Skaar,5 Anthony J Perkins,2 Sujuan Gao,2,3,6 Lang Li,7 Babar A Khan,2,3,5 Malaz A Boustani2,3,81College of Pharmacy, Purdue University, West Lafayette, 2Indiana University Center for Aging Research, 3Regenstrief Institute, 4Department of Pharmacy, Eskenazi Health Services, 5Division of Clinical Pharmacology, Department of Medicine, 6Department of Biostatistics, 7Department of Medical and Molecular Genetics, Indiana University School of Medicine, 8Center for Innovation and Implementation Science, Indiana University, Indianapolis, IN, USAObjective: To determine the frequency of pharmacogenomic variants and concurrent medications that may alter the efficacy and tolerability of acetylcholinesterase inhibitors (AChEIs.Materials and methods: A multisite cross-sectional study was carried out across four memory care practices in the greater Indianapolis area. Participants were adults aged 65 years and older with a diagnosis of probable or possible Alzheimer’s disease (AD (n=105. Blood samples and self-reported medication data were collected. Since two of the three AChEIs are metabolized by cytochrome P450 (CYP-2D6, we determined the frequency of functional genetic variants in the CYP2D6 gene and calculated their predicted CYP2D6-activity scores. Concurrent medication data were collected from self-reported medication surveys, and their predicted effect on the pharmacokinetics of AChEIs was determined based on their known effects on CYP2D6 and CYP3A4/5 enzyme activities.Results: Among the 105 subjects enrolled, 72% were female and 36% were African American. Subjects had a mean age of 79.6 years. The population used a mean of eight medications per day (prescription and nonprescription. The CYP2D6 activity score frequencies were 0 (3.8%, 0.5 (4.8%, 1.0 (36.2%, 1.5–2.0 (51.4%, and >2.0 (3.8%. Nineteen subjects (18.1% used a medication considered a strong or moderate inhibitor of CYP2D6, and eight subjects (7.6% used a

  16. Activity of the Respiratory Chain Enzymes of Blood Leucocytes’ Mitochondria Under the Conditions of Toxic Hepatitis Induced Against the Background Alimentary Deprivation of Protein

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    O.N. Voloshchuk

    2015-12-01

    Full Text Available Full functioning of the leucocytes’ energy supply system is one of the essential factors for the immune surveillance system effective work. The pivotal enzymes of the leucocytes’ energy biotransformation system are NADH-ubiquitin reductase, a marker of the Complex I of respiratory chain activity, and succinate dehydrogenase, key enzyme of the Complex II of respiratory chain. The aim of research – to study the NADH-ubiquitin reductase and succinate dehydrogenase activity of the blood leucocytes’ mitochondria under the conditions of toxic hepatitis induced against the background alimentary deprivation of protein. It is shown, that under the conditions of acetaminophen-induced hepatitis a reduction of the NADH-ubiquitin reductase enzymatic activity is observed on the background activation of the succinate-dependent way of the mitochondrial oxidation. Conclusion was made that alimentary deprivation or protein is a factor, aggravating the misbalance of the energy biotransformation system in the leucocytes of rats with toxic hepatitis. Established activity changes of the leucocytes’ mitochondria respiratory chain key enzymes may be considered as one of the mechanisms, directed on the maintenance of leucocytes energy supply on a level, sufficient for their functioning. Research results may be used for the biochemical rationale of the therapeutic approaches to the elimination and correction of the leucocytes’ energy metabolism disturbances consequences under the conditions of acetaminophen-induced hepatitis, aggravated by the alimentary protein deprivation.

  17. RESEARCHES ON THE ACTIVITY OF SOME ENZYMES WITH DIAGNOSIS VALUE IN SUBJECTS SUFFERING FROM HEPATIC CIRRHOSIS

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    Anca Mirela Amariei

    2012-06-01

    deviations from the physiologically normal values were noticed, all the other investigated subjects show values higher than the superior limit of the reference interval while, statistically, the values of the seric enzymes are highly significant (p<0.001, with the exception of those of aspartat-aminotransferase (in the 71-86 years group and of Ȗ-glutamyl- transferase (the 41-50 year group, which are insignificant.

  18. Hepatitis D in Chronic Active Hepatitis B: Prevalence, Liver Enzyme Levels and Histopathology- an Epidemiological Study in Shiraz, Southern Iran, 2003-2004

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    Farnaz Khademolhosseini

    2008-12-01

    Full Text Available Background and Aims: At least 5% of hepatitis B carriers worldwide are infected with Hepatitis D virus (HDV. This study aims to determine the prevalence, transaminase levels and histopathological findings of HDV among patients with chronic active hepatitis B in southern Iran.Methods: During 2003-2004, 93 patients >15 years with chronic active hepatitis B were enrolled from referrals to Shiraz University of Medical Sciences in southern Iran.Results: Nine (9.7% patients were seropositive for the anti HDV antibody. 76.3% of patients were male and among the HDV positive group, all subjects were male too. A significantly higher AST and more advanced grade and stage of liver disease were observed in the HDV positive group. The most common mode of transmission in the positive group was intravenous drug use.Conclusions: The risk of liver disease progression in chronic hepatitis B appears to be higher in HDV infected patients. Intravenous drug abuse is an important risk factor for acquiring HDV infection. Checking anti-HDV is suggested in any patient with positive HBsAg, especially in males or those with history of intravenous drug abuse.

  19. Enhanced expressions and activations of leukotriene C4 synthesis enzymes in D-galactosamine/lipopolysaccharide-induced rat fulminant hepatic failure model

    Institute of Scientific and Technical Information of China (English)

    Kui-Fen Ma; Hong-Yu Yang; Zhe Chen; Luo-Yang Qi; Dan-Yan Zhu; Yi-Jia Lou

    2008-01-01

    AIM:To investigate the expression and activity of leukotriene C4 (L-TC4) synthesis enzymes and their underlying relationship with cysteinyl leukotriene (cys-LT)generation in a rat fulminant hepatic failure (FHF) model induced by D-galactosamine/lipopolyseccharide (D-GaIN/LPS).METHODS:Rats were treated with D-GaIN (300 mg/kg)plus LPS (0.1 mg/kg) for 1,3,6,and 12 h.Enzyme immunoassay was used to determine the hepatic cys-LT content.Reverse transcription-polymerase chain reaction (RT-PCR),Western blot or immunohistochemical assay were employed to assess the expression or location of LTC4 synthesis enzymes,which belong to membrane associated proteins in eicosanoid and glutathione (MAPEG)metabolism superfamily.Activity of LTC4 synthesis enzymes was evaluated by determination of the products of LTA4 alter incubation with liver microsomes using high performance liquid chromatography (HPLC).RESULTS:Livers were injured after treatment with D-GalN/LPS,accompanied by cys-LT accumulation at the prophase of liver injury.Both LTC4 synthase (LTC4S) and microsomal glutathione-S-transferase (mGST) 2 were expressed in the rat liver,while the latter was specifically located in hepatocytes.Their mRNA and protein expressions were up-regulated at an earlier phase alter treatment with D-GalN/LPS.Meantime,a higher activity of LTC4 synthesis enzymes was detected,although the activity of LTC4S played the main role in this case.CONCLUSION:The expression and activity of both LTC4S and mGST2 are up regulated in a rat FHF model,which are,at least,partly responsible for cys-LT hepatic accumulation.

  20. Glutathione and antioxidant enzymes serve complementary roles in protecting activated hepatic stellate cells against hydrogen peroxide-induced cell death

    NARCIS (Netherlands)

    Dunning, Sandra; Rehman, Atta Ur; Tiebosch, Marjolein H.; Hannivoort, Rebekka A.; Haijer, Floris W.; Woudenberg, Jannes; van den Heuvel, Fiona A. J.; Buist-Homan, Manon; Faber, Klaas Nico; Moshage, Han

    2013-01-01

    Background: In chronic liver disease, hepatic stellate cells (HSCs) are activated, highly proliferative and produce excessive amounts of extracellular matrix, leading to liver fibrosis. Elevated levels of toxic reactive oxygen species (ROS) produced during chronic liver injury have been implicated i

  1. Comparison among Different Gilthead Sea Bream (Sparus aurata Farming Systems: Activity of Intestinal and Hepatic Enzymes and 13C-NMR Analysis of Lipids

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    Vincenzo Zonno

    2009-12-01

    Full Text Available In order to evaluate differences in general health and nutritional values of gilthead sea bream (Sparus aurata, the effects of semi-intensive, land-based tanks and sea-cages intensive rearing systems were investigated, and results compared with captured wild fish. The physiological state was determined by measuring the activity of three different intestinal digestive enzymes: alkaline phosphatase (ALP, leucine aminopeptidase (LAP and maltase; and the activity of the hepatic ALP. Also, the hepatic content in protein, cholesterol, and lipid were assessed. 13C-NMR analysis for qualitative and quantitative characterization of the lipid fraction extracted from fish muscles for semiintensive and land based tanks intensive systems was performed. The lipid fraction composition showed small but significant differences in the monounsaturated/saturated fatty acid ratio, with the semi-intensive characterized by higher monounsaturated and lower saturated fatty acid content with respect to land based tanks intensive rearing system.

  2. Hepatic UDP-glucuronosyltransferase enzyme activity decreased during salt acclimation of sea lamprey juveniles from Minho river basin

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    Marta Candeias

    2015-11-01

    Full Text Available Sea lamprey (Petromyzon marinus L., 1758 is an anadromous species which migrates twice during its life cycle between freshwater and seawater. During downstream migration, the juveniles are subject to salinity changes ranging between 0 and 35. The aim of this study was to evaluate the influence of salinity in hepatic biomarkers of stress and biotransformation of juveniles from Minho river basin, Portugal, during trophic migration. Sampled juveniles (macrophthalmia were transported alive to the laboratory and maintained in 200 L tanks with LSS life support system. The specimens were separated in three groups of five pools (#8: i macrophthalmia, salinity 0 for 7 days; ii macrophthalmia, salinity 0 for 30 days and, iii macrophthalmia subjected to a salt gradient up to salinity 35, for 30 days. At final of the experiments the weight and the length of specimens were determined. Microsomes and cytosol obtained by centrifugation of liver homogenates were used for fluorimetric quantification of glutathione (GSH, glutathione disulphide (GSSG and malondialdehyde (MDA and, spectrophotometric determination of catalase (CTT1 and UDP-glucuronosyltransferase (UDPGT activities. Stats include ANOVA I and Duncan test. The results showed that salt exposure of the animals caused a decrease in the body weight and condition factor (K without changes the hepatic somatic index (p <0.05. It was also observed a decrease in the GSH/GSSG ratio and UDPGT activity, markers of oxidative stress and loss of biotransformation capacity of liver macrophthalmia (p <0.05. This disturbances may jeopardize the success of sea lamprey trophic migration, if occur a permanent or accidental exposure of juveniles to organic pollutants on the path to the sea. In addition, the salt exposure did not change the liver cytosolic MDA content (p <0.05. The significant increase in CTT1 activity may have contributed to prevent hepatic oxidative damages in the sea lamprey juveniles.

  3. Methadone induces the expression of hepatic drug-metabolizing enzymes through the activation of pregnane X receptor and constitutive androstane receptor.

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    Tolson, Antonia H; Li, Haishan; Eddington, Natalie D; Wang, Hongbing

    2009-09-01

    Methadone (MD) is the most established substance abuse pharmacotherapy of choice for the management of heroin dependence. To date, drug-drug interactions involving MD have been characterized asymmetrically among existing reports, which describe how other drugs affect the metabolic or pharmacokinetic profiles of MD; however, limited information is available regarding the potential for MD to influence similar fates of coadministered drugs. Moreover, little to no mechanistic evidence has been explored. Here, we show that MD induces hepatic drug-metabolizing enzymes (DMEs) through the activation of pregnane X receptor (PXR) and constitutive androstane receptor (CAR). Real-time polymerase chain reaction analysis of human hepatocyte cultures revealed that MD induces the mRNA expression of CYP2B6, CYP3A4, UGT1A1, and multidrug resistance 1 in a concentration-related manner, with the maximal induction of CYP2B6 challenging that of the induction by rifampicin. Furthermore, MD-mediated induction of CYP2B6 and CYP3A4 proteins was observed in Western blot analysis. In cell-based reporter assays, MD significantly increased human (h) PXR-mediated CYP2B6 reporter activities but exhibited minimal effect on hCAR activation as a result of the constitutive activity of hCAR in HepG2 cells. Further studies revealed that treatment with MD resulted in significant nuclear accumulation of adenovirus/enhanced yellow fluorescent protein tagged-hCAR in human hepatocytes, which has been regarded as the initial step of CAR activation. Additional analysis of the two enantiomers of MD, R-(-)-MD (active) and S-(+)-MD (inactive), indicates the lack of stereoselectivity pertaining to MD-mediated DME induction. Overall, our results show that MD induces the hepatic expression of multiple DMEs by activating PXR- and CAR-mediated pathways.

  4. Serum hepatic enzyme activity in relation to semen quality and serum reproductive hormone levels among Estonian fertile Men.

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    Ehala-Aleksejev, K; Punab, M

    2016-01-01

    The aim of this study was to investigate the relations of basic semen parameters and reproductive hormones with alanine aminotransferase (ALT) and gamma-glutamyl transferase (GGT). In addition, to examine possible interaction between adiposity, alcohol consumption, and liver tests in relation to male reproductive health, standard semen analysis was performed and serum levels of reproductive hormones and liver tests were measured in 245 male partners of pregnant women at a University Hospital Andrology Centres in Estonia. Quartile analysis revealed that after adjustment for covariates GGT was negatively related to sperm concentration and total sperm count. These significant changes appeared from a GGT >35.5 U/L. Next to these changes ALT was not related to sperm parameters. Both enzymes, GGT and ALT, were not related to reproductive hormones. Alcohol consumption was positively related to GGT and in cases with elevated GGT alcohol use was negatively related to sperm concentration and total sperm count. Alcohol consumption was positively related to body mass index (BMI) and waist circumference (WC). Our findings also confirm results of previous studies that BMI and WC are associated positively with ALT and GGT. According to the study, increased GGT activity might represent a possible connection between adiposity, alcohol consumption, and semen quality.

  5. Dietary chia seed induced changes in hepatic transcription factors and their target lipogenic and oxidative enzyme activities in dyslipidaemic insulin-resistant rats.

    Science.gov (United States)

    Rossi, Andrea S; Oliva, Maria E; Ferreira, Maria R; Chicco, Adriana; Lombardo, Yolanda B

    2013-05-01

    The present study analyses the effect of dietary chia seed rich in n-3 α-linolenic acid on the mechanisms underlying dyslipidaemia and liver steatosis developed in rats fed a sucrose-rich diet (SRD) for either 3 weeks or 5 months. The key hepatic enzyme activities such as fatty acid synthase (FAS), acetyl-CoA carboxylase (ACC), glucose-6-phosphate dehydrogenase (G-6-PDH), carnitine palmitoyltransferase-1 (CPT-1) and fatty acid oxidase (FAO) involved in lipid metabolism and the protein mass levels of sterol regulatory element-binding protein-1 (SREBP-1) and PPARα were studied. (1) For 3 weeks, Wistar rats were fed either a SRD with 11 % of maize oil (MO) as dietary fat or a SRD in which chia seed replaced MO (SRD+Chia). (2) A second group of rats were fed a SRD for 3 months. Afterwards, half the rats continued with the SRD while for the other half, MO was replaced by chia for 2 months (SRD+Chia). In a control group, maize starch replaced sucrose. Liver TAG and the aforementioned parameters were analysed in all groups. The replacement of MO by chia in the SRD prevented (3 weeks) or improved/normalised (5 months) increases in dyslipidaemia, liver TAG, FAS, ACC and G-6-PDH activities, and increased FAO and CPT-1 activities. Protein levels of PPARα increased, and the increased mature form of SREBP-1 protein levels in the SRD was normalised by chia in both protocols (1 and 2). The present study provides new data regarding some key mechanisms related to the fate of hepatic fatty acid metabolism that seem to be involved in the effect of dietary chia seed in preventing and normalising/improving dyslipidaemia and liver steatosis in an insulin-resistant rat model.

  6. Effect of High Dietary Carbohydrate on the Growth Performance, Blood Chemistry, Hepatic Enzyme Activities and Growth Hormone Gene Expression of Wuchang Bream ( at Two Temperatures

    Directory of Open Access Journals (Sweden)

    Chuanpeng Zhou

    2015-02-01

    Full Text Available The effects of high carbohydrate diet on growth, serum physiological response, and hepatic heat shock protein 70 expression in Wuchang bream were determined at 25°C and 30°C. At each temperature, the fish fed the control diet (31% CHO had significantly higher weight gain, specific growth rate, protein efficiency ratio and hepatic glucose-6-phosphatase activities, lower feed conversion ratio and hepatosomatic index (HSI, whole crude lipid, serum glucose, hepatic glucokinase (GK activity than those fed the high-carbohydrate diet (47% CHO (p<0.05. The fish reared at 25°C had significantly higher whole body crude protein and ash, serum cholesterol and triglyceride, hepatic G-6-Pase activity, lower glycogen content and relative levels of hepatic growth hormone (GH gene expression than those reared at 30°C (p<0.05. Significant interaction between temperature and diet was found for HSI, condition factor, hepatic GK activity and the relative levels of hepatic GH gene expression (p<0.05.

  7. Effect of thiabendazole on some rat hepatic xenobiotic metabolising enzymes

    NARCIS (Netherlands)

    Price, R.J.; Scott, M.P.; Walters, D.G.; Stierum, R.H.; Groten, J.P.; Meredith, C.; Lake, B.G.

    2004-01-01

    The effect of thiabendazole (TB) on some rat hepatic xenobiotic metabolising enzymes has been investigated. Male Sprague-Dawley rats were fed control diet or diets containing 102-5188 ppm TB for 28 days. As a positive control for induction of hepatic xenobiotic metabolism, rats were also fed diets c

  8. Effect of thiabendazole on some rat hepatic xenobiotic metabolising enzymes

    NARCIS (Netherlands)

    Price, R.J.; Scott, M.P.; Walters, D.G.; Stierum, R.H.; Groten, J.P.; Meredith, C.; Lake, B.G.

    2004-01-01

    The effect of thiabendazole (TB) on some rat hepatic xenobiotic metabolising enzymes has been investigated. Male Sprague-Dawley rats were fed control diet or diets containing 102-5188 ppm TB for 28 days. As a positive control for induction of hepatic xenobiotic metabolism, rats were also fed diets

  9. Effects of Cyclamen trochopteranthum on hepatic drug-metabolizing enzymes

    Directory of Open Access Journals (Sweden)

    Arslan Sevki

    2011-01-01

    Full Text Available The modulatory effects of the Cyclamen trochopterantum tuber extract on hepatic drug-metabolizing enzymes, including aniline 4-hydroxylase (A4H; CYP2E1, ethoxyresorufin O-deethylase (EROD; CYP1A, methoxyresorufin O-demethylase (MROD; CYP1A, caffeine N-demethylase (C3ND; CYP1A2 aminopyrene N-demethylase (APND; CYP2C6, and erythromycin N-demethylase (ERND; CYP3A1, were examined in vivo in rats. The activities of all of these enzymes were induced by the cyclamen extract. In addition, Western-blot and RT-PCR results clearly showed that CYP2E1, CYP1A1/CYP1A2 and CYP2C6 protein and mRNA levels were substantially increased by four different doses of cyclamen. Although, the CYP3A1 protein level was increased significantly, the mRNA level was not changed. These results indicate that cyclamen tuber extract might have a potential not only to inhibit and/or induce the metabolism of certain co-administered drugs but also influence the development of toxicity and carcinogenesis due to the induction of the cytochrome P450-dependent drug-metabolizing enzymes.

  10. Interferon augments the anti-fibrotic activity of an angiotensin-converting enzyme inhibitor in patients with refractory chronic hepatitis C

    Institute of Scientific and Technical Information of China (English)

    Hitoshi Yoshiji; Masaharu Yamazaki; Masahisa Toyohara; Akira Mitoro; Hiroshi Fukui; Ryuichi Noguchi; Hideyuki Kojima; Yasuhide Ikenaka; Mitsuteru Kitade; Kosuke Kaji; Masahito Uemura; Junichi Yamao; Masao Fujimoto

    2006-01-01

    AIM:To evaluate the effect of combination treatment with the interferon (IFN) and angiotensin-converting enzyme inhibitor (ACE-I ) on several fibrotic indices in patients with refractory chronic hepatitis C (CHC).METHODS: Perindopril (an ACE-I; 4 mg/d) and/or natural IFN (3 MU/L; 3 times a week) were administered for 12 mo to refractory CHC patients, and several indices of serum fibrosis markers were analyzed.RESULTS:ACE-Ⅰ decreased the serum fibrosis markers,whereas single treatment with IFN did not exert these inhibitory effects. However, IFN significantly augmented the effects of ACE-Ⅰ, and the combination treatment exerted the most potent inhibitory effects. The serum levels of alanine transaminase and HCV-RNA were not significantly different between the groups, whereas the plasma level of transforming growth factor-β was significantly attenuated almost in parallel with suppression of the serum fibrosis markers.CONCLUSION:The combination therapy of an ACE-Ⅰand IFN may have a diverse effect on disease progression in patients with CHC refractory to IFN therapy through its anti-fibrotic effect.

  11. An enzyme with rhamnogalacturonase activity.

    NARCIS (Netherlands)

    Kovod, L.V.; Dalboge, H.; Andersen, L.N.; Kauppinen, M.; Christgan, S.; Heldt-Hansen, H.P.; Christophersen, C.; Nielsen, P.M.; Voragen, A.G.J.; Schols, H.A.

    1994-01-01

    An enzyme exhibiting rhamnogalacturonase activity, which enzyme: a) is encoded by the DNA sequence shown in SEQ ID No. 1 or a sequence homologous thereto encoding a polypeptide with RGase activity, b) has the amino acid sequence shown in SEQ ID No. 2 or an analogue thereof, c) is reactive with an

  12. Hepatic Rupture Caused by Hemolysis, Elevated Liver Enzyme, and Low Platelet Count Syndrome: A Case Report with Computed Tomographic and Conventional Angiographic Findings

    Energy Technology Data Exchange (ETDEWEB)

    Lee, Cheong Bok; Ahn, Jae Hong; Choi, Soo Jung; Lee, Jong Hyeog; Park, Man Soo; Jung, Seung Mun; Ryu, Dae Sik [Dept. of Radiology, Asan Foundation, Gangneung Asan Hospital, University of Ulsan College of Medicine, Gangneung (Korea, Republic of)

    2013-03-15

    The authors recently obtained successful clinical outcome after embolization of the hepatic artery and right inferior phrenic artery in a pregnant patient with hemolysis, elevated liver enzyme, and low platelet count (HELLP) syndrome causing hepatic rupture. We report the computed tomographic and conventional angiographic findings in a case of HELLP syndrome, resulting in hepatic infarction and rupture with active bleeding.

  13. Effect of pretreatment with hepatic microsomal enzyme inducers on the toxicity of diazinon in calves.

    Science.gov (United States)

    Abdelsalam, E B; Ford, E J

    1986-11-01

    The pretreatment of calves with a single dose of 10 mg kg-1 dieldrin or 21 daily doses of 10 mg kg-1 phenobarbitone increased the toxicity of diazinon as reflected by the development of more severe clinical signs and greater depression in whole blood cholinesterase activity in the pretreated calves. Induction by dieldrin or phenobarbitone of the hepatic microsomal enzyme amidopyrine-N-demethylase was also accompanied by a concurrent rise in the liver carboxylesterase activity.

  14. Leflunomide Induces Pulmonary and Hepatic CYP1A Enzymes via Aryl Hydrocarbon Receptor.

    Science.gov (United States)

    Patel, Ananddeep; Zhang, Shaojie; Paramahamsa, Maturu; Jiang, Weiwu; Wang, Lihua; Moorthy, Bhagavatula; Shivanna, Binoy

    2015-12-01

    Emerging evidence indicates that the aryl hydrocarbon receptor (AhR) plays a crucial role in normal physiologic homeostasis. Additionally, aberrant AhR signaling leads to several pathologic states in the lung and liver. Activation of AhR transcriptionally induces phase I (CYP1A) detoxifying enzymes. Although the effects of the classic AhR ligands such as 3-methylcholanthrene and dioxins on phase 1 enzymes are well studied in rodent lung, liver, and other organs, the toxicity profiles limit their use as therapeutic agents in humans. Hence, there is a need to identify and investigate nontoxic AhR ligands not only to understand the AhR biology but also to develop the AhR as a clinically relevant therapeutic target. Leflunomide is a Food and Drug Administration-approved drug in humans that is known to have AhR agonist activity in vitro. Whether it activates AhR and induces phase 1 enzymes in vivo is unknown. Therefore, we tested the hypothesis that leflunomide will induce pulmonary and hepatic CYP1A enzymes in C57BL/6J wild-type mice, but not in AhR-null mice. We performed real-time reverse-transcription polymerase chain reaction analyses for CYP1A1/2 mRNA expression, western blot assays for CYP1A1/2 protein expression, and ethoxyresorufinO-deethylase assay for CYP1A1 catalytic activity. Leflunomide increased CYP1A1/A2 mRNA, protein, and enzymatic activities in wild-type mice. In contrast, leflunomide failed to increase pulmonary and hepatic CYP1A enzymes in AhR-null mice. In conclusion, we provide evidence that leflunomide induces pulmonary and hepatic CYP1A enzymes via the AhR.

  15. Expression of hepatic antioxidant enzymes in non-obese type-2 diabetic Goto-Kakizaki rats.

    Science.gov (United States)

    Ryu, Chang Seon; Oh, Soo Jin; Oh, Jung Min; Lee, Sang Yoon; Kwak, Hui Chan; Yun, Kang Uk; Lee, Ji-Yoon; Park, Song-Kyu; Kim, Bong-Hee; Ma, Jin Yeul; Kim, Sang Kyum

    2014-10-01

    Diabetes mellitus and its complications have been attributed in part to oxidative stress, against which antioxidant enzymes constitute a major protective mechanism. The present study was performed to investigate the effects of early stage type 2 diabetes in the absence of obesity and liver damage on hepatic antioxidant enzyme expression and oxidative stress using 9-week-old Goto-Kakizaki (GK) rats. Hepatic total antioxidant capacity determined by total oxygen radical scavenging capacity and lipid peroxidation determined by malondialdehyde in plasma and liver were not significantly different between normal Wistar rats and GK rats. These results indicated that oxidative stress is not evident in these type 2 diabetic rats. Hepatic expression levels of antioxidant enzymes, including superoxide dismutase-1, catalase, glutathione peroxidase and reductase, thioredoxin-1, mu- and pi-class glutathione S-transferase (GST), and the gamma-glutamylcysteine ligase catalytic subunit, were not different between normal rats and GK rats. But, hepatic level and activity of alpha-class GST were decreased and peroxiredoxin-1 level was increased in GK rats, suggesting that upregulation of peroxiredoxin-1 compensates for downregulation of alpha-class GST. These results suggest that alpha-class GST and peroxiredoxin-1 in liver can be altered during the early stages of type 2 diabetes in the absence of obesity and severe oxidative stress.

  16. Chemoprotective activity of boldine: modulation of drug-metabolizing enzymes.

    Science.gov (United States)

    Kubínová, R; Machala, M; Minksová, K; Neca, J; Suchý, V

    2001-03-01

    Possible chemoprotective effects of the naturally occurring alkaloid boldine, a major alkaloid of boldo (Peumus boldus Mol.) leaves and bark, including in vitro modulations of drug-metabolizing enzymes in mouse hepatoma Hepa-1 cell line and mouse hepatic microsomes, were investigated. Boldine manifested inhibition activity on hepatic microsomal CYP1A-dependent 7-ethoxyresorufin O-deethylase and CYP3A-dependent testosterone 6 beta-hydroxylase activities and stimulated glutathione S-transferase activity in Hepa-1 cells. In addition to the known antioxidant activity, boldine could decrease the metabolic activation of other xenobiotics including chemical mutagens.

  17. Hepatic enzyme concentrations as indicators of nonalcoholic fatty liver disease

    Directory of Open Access Journals (Sweden)

    Alvina Alvina

    2016-02-01

    Full Text Available Nonalcoholic fatty liver disease (NAFLD has emerged as a world-wide problem because it runs an asymptomatic course, ultimately leading to cirrhosis of the liver and portal hypertension, resulting in death. The prevalence of the disease accounts for 3-24% of the population in several countries. Generally there are increased concentrations of hepatic enzymes as markers of liver damage, such as serum glutamic oxaloacetic transaminase (SGOT, serum glutamic pyruvic transaminase (SGPT and gamma glutamyl transferase (GGT. The aim of the present study was to determine the concentrations of hepatic enzymes as markers of NAFLD. The study design was cross-sectional, involving 90 subjects meeting the inclusion and exclusion criteria. The degree of severity NAFLD was determined by ultrasonography and the concentrations of SGOT, SGPT and GGT by automated clinical chemistry analyzer. The results indicated that there were 32 subjects with mild NAFLD (35.6%, 35 subjects with moderate NAFLD (38.9% and 23 subjects with severe NAFLD (25.6%. There was a significant difference in degree of NAFLD by gender (p<0.05, where severe NAFLD was more frequent in males than in females. Concentrations of SGOT, SGPT and GGT were significantly different between degrees of NAFLD (p<0.05. The conclusion is that SGOT, SGPT and GGT concentrations are indicators of degree of NAFLD.

  18. Comparison of amplification enzymes for Hepatitis C Virus quasispecies analysis

    Directory of Open Access Journals (Sweden)

    Bonkovsky Herbert L

    2005-04-01

    Full Text Available Abstract Background Hepatitis C virus (HCV circulates as quasispecies (QS, whose evolution is associated with pathogenesis. Previous studies have suggested that the use of thermostable polymerases without proofreading function may contribute to inaccurate assessment of HCV QS. In this report, we compared non-proofreading (Taq with proofreading (Advantage High Fidelity-2; HF-2 polymerases in the sensitivity, robustness, and HCV QS diversity and complexity in the second envelope glycoprotein gene hypervariable region 1 (E2-HVR1 on baseline specimens from 20 patients in the HALT-C trial and in a small cohort of 12 HCV/HIV co-infected patients. QS diversity and complexity were quantified using heteroduplex mobility assays (HMA. Results The sensitivities of both enzymes were comparable at 50 IU/ml, although HF-2 was more robust and slightly more sensitive than Taq. Both enzymes generated QS diversity and complexity scores that were correlated (r = 0.68; p Conclusion The data suggest that while Taq may overestimate HCV QS complexity, its use does not significantly affect results in cohort-based studies of HCV QS analyzed by HMA. However, the use of proofreading enzymes such as HF-2 is recommended for more accurate characterization of HCV QS in vivo.

  19. 营养限制与补偿对蒙古育成羯羊肝脏关键酶活性的影响%Feed Restriction and Realimentation Affect Activities of Hepatic Key Enzymes in Mongolia Growing Wethers

    Institute of Scientific and Technical Information of China (English)

    徐永霞; 侯先志; 王海荣; 高爱武; 李俊良; 杨金丽

    2012-01-01

    This experiment was conducted to study the effects of feed restriction and realimentation on the activities of hepatic key enzymes in Mongolia growing wethers. Forty three-month-old growing wethers with an average body weight of (14. 72 ± 1. 10) kg were selected and divided into 2 groups (control group and restricted group). The feeding trial included a 60-d feed restriction period and a 90-d feed realimentation period. The metabolizable energy (ME) and crude protein (CP) content in diets were 8. 62 MJ/kg and 5.7% for restricted group and were 10. 88 MJ/kg and 15. 0% for the control group during the feed restriction period. All growing wethers were offered the same diet (ME: 9. 75 MJ/kg; CP: 12% ) during the feed realimentation period. At the end of feed restriction and realimentation period, the activities of hexokinase (HK) , pyruvate ki-nase ( PK) , lactate dehydrogenase ( LDH) , fructose-1, 6-bisphosphatase ( FBP1) , malate dehydrogenase (MDH) , α-glycerophosphate dehydrogenase ( GAPDH) , alanine aminotransferase ( ALT) , aspartate amin-otransferase (AST) and glutamate dehydrogenase (GLDH) in liver in each group were examined. The results showed that the activities of hepatic HK, ALT, AST, GLDH of restricted group were significantly lower than those of the control group (P 0. 05). These results indicate that the activities of key enzymes in the glycolysis and protein metabolism are decreased, and the activities of key enzymes in gluconeogenesis and lipolysis are increased. That may be related to the strengthening of fat mobilization during the feed restriction period. The activities of key enzymes in protein metabolism are decreased which means to decrease protein metabolism during the feed restriction period. At the end of realimentation period, the activities of key enzymes in glycolysis, gluconeogenesis and lipid metabolism are recovered to the level of control group, while the activities of hepatic PK and key enzymes in fat synthesis and protein

  20. Measuring the Enzyme Activity of Arabidopsis Deubiquitylating Enzymes.

    Science.gov (United States)

    Kalinowska, Kamila; Nagel, Marie-Kristin; Isono, Erika

    2016-01-01

    Deubiquitylating enzymes, or DUBs, are important regulators of ubiquitin homeostasis and substrate stability, though the molecular mechanisms of most of the DUBs in plants are not yet understood. As different ubiquitin chain types are implicated in different biological pathways, it is important to analyze the enzyme characteristic for studying a DUB. Quantitative analysis of DUB activity is also important to determine enzyme kinetics and the influence of DUB binding proteins on the enzyme activity. Here, we show methods to analyze DUB activity using immunodetection, Coomassie Brilliant Blue staining, and fluorescence measurement that can be useful for understanding the basic characteristic of DUBs.

  1. The angiotensin-converting enzyme inhibitor captopril rescues mice from endotoxin-induced lethal hepatitis.

    Science.gov (United States)

    Ge, Pu; Jiang, Rong; Yao, Xin; Li, Jing; Dai, Jie; Zhang, Li; Ye, Bin

    2017-02-01

    The renin-angiotensin system is classically regarded as a crucial regulator of circulatory homeostasis, but recent studies also revealed its pro-inflammatory roles. The beneficial effects of the angiotensin-converting enzyme inhibitor (ACEI) in severe inflammatory injury in the lung and heart have been previously reported, but its potential effects on lethal hepatitis were unknown. In this study, a mouse model with LPS/d-galactosamine (GalN)-induced fulminant hepatitis were used to test the protective potential of captopril, a representative ACEI. The results indicated that treatment with captopril significantly decreased the plasma level of alanine aminotransferase and aspartate aminotransferase, alleviated the histopathological damage of the liver tissue and improve the survival rate of LPS/GalN-challenged mice. These effects were accompanied by reduced mRNA levels of TNF-α and IL-6 in the liver, and decreased protein level of TNF-α and IL-6 in the plasma. In addition, the activation of caspases 3, 8 and 9, and the presence of TUNEL-positive apoptotic cells, were also suppressed by captopril treatment. The above evidence suggested that the renin-angiotensin system might be involved in the development of LPS/GalN-induced fulminant hepatitis and ACEI might have potential value in lethal hepatitis.

  2. Modulatory effects of Kaempferia parviflora extract on mouse hepatic cytochrome P450 enzymes.

    Science.gov (United States)

    Mekjaruskul, Catheleeya; Jay, Michael; Sripanidkulchai, Bungorn

    2012-06-14

    Kaempferia parviflora is a herbal plant, the extracts of which are commonly used as alternative medicines. It widely uses as aphrodisiac, anti-inflammation, anti-microbacterial, and anti-peptic ulcer. In order to obtain an effective utilization and safety of the herb, the influence of Kaempferia parviflora on hepatic CYP450 metabolizing enzymes including CYP1A1, CYP1A2, CYP2B, CYP2E1, and CYP3A was investigated. The impact of Kaempferia parviflora on CYP450 both in vitro and in vivo was examined by using ethoxyresorufin O-dealkylation, methoxyresorufin O-dealkylation, pentoxyresorufin O-dealkylation, p-nitrophenol hydroxylation, and erythromycin N-demethylation assays, respectively. In vitro studies using non-induced mouse hepatic microsomes in the presence or absence of Kaempferia parviflora extract showed that Kaempferia parviflora extract altered CYP1A1, CYP1A2, CYP2B, and CYP2E1 activities by non-competitive, mixed-competitive, competitive, and uncompetitive mechanisms, respectively. Among these enzymes, CYP1A2 was affected by Kaempferia parviflora based on the highest value of V(max) (15.276±0.206 nmol/min) and lowest of K(i) value (0.008±0.002 μg/ml). In addition, the plant extract also modulated CYP2B activity based on the low K(m) value (1.599±0.147 pmol). For in vivo studies, mice were orally treated with 250 mg/kg of Kaempferia parviflora extract for 7, 14, and 21 days. The results demonstrated that Kaempferia parviflora extract significantly induced CYP1A1, CYP1A2 enzyme activities following short-term treatment. CYP2B enzyme activities were markedly increased all Kaempferia parviflora extract treatment timepoints, whereas Kaempferia parviflora extract significantly enhanced CYP2E1 activity only after long-term treatment. However, Kaempferia parviflora extract did not affect the CYP3A enzyme activity. Kaempferia parviflora extract modulated several CYP450 enzyme activities, thus, its utilization with drugs or other herbs should raise concern for

  3. Plasma metabolites, ions and thyroid hormones levels, and hepatic enzymes׳ activity in Caspian roach (Rutilus rutilus caspicus) exposed to waterborne manganese.

    Science.gov (United States)

    Hoseini, Seyyed Morteza; Hedayati, Aliakbar; Ghelichpour, Melika

    2014-09-01

    To investigate the effect of waterborne manganese on plasma biochemical characteristics in fish, Caspian roach (Rutilus rutilus caspicus) was exposed to 0 (control), 60 (M60), 150 (M150) and 300 (M300) mg/L water manganese for 96h. Thereafter, plasma biochemical characteristics were studied. Plasma glucose level significantly increased in M60 and decreased in M150 and M300 groups, compared to the control. M300 had significantly lower hematocrit compared to the control. Albumin remained unchanged after manganese exposure, however, the manganese-exposed fish showed significant increase in plasma total protein levels. M150 and M300 showed significant increase in the plasma cholesterol and triglyceride levels compared to the control and M60. M60 and M150 had significantly higher alkaline phosphatase (ALP) activity compared to the control. The manganese-exposed groups had significantly higher alanine aminotransferase (ALT) activity compared to the control. M150 and M300 had aspartate aminotransferase (AST) activity significantly higher than those of the control and M60. M300 had significantly higher triiodothyronine (T3) levels than the other groups. All manganese-exposed fish had significantly higher thyroxin (T4) levels than the control. The plasma levels of chloride showed a significant decrease in the manganese-exposed fish, compared to the control. M150 and M300 had significantly lower sodium levels, compared to the control. M60 and M150 had significantly lower plasma calcium levels compared to the other groups. It is concluded that clinical chemistry along with thyroid hormones levels can be the useful tools to monitor manganese toxicity in fish. The possible mechanisms involving in the biochemical changes were discussed.

  4. Activation of thiamin diphosphate in enzymes.

    Science.gov (United States)

    Hübner, G; Tittmann, K; Killenberg-Jabs, M; Schäffner, J; Spinka, M; Neef, H; Kern, D; Kern, G; Schneider, G; Wikner, C; Ghisla, S

    1998-06-29

    Activation of the coenzyme ThDP was studied by measuring the kinetics of deprotonation at the C2 carbon of thiamin diphosphate in the enzymes pyruvate decarboxylase, transketolase, pyruvate dehydrogenase complex, pyruvate oxidase, in site-specific mutant enzymes and in enzyme complexes containing coenzyme analogues by proton/deuterium exchange detected by 1H-NMR spectroscopy. The respective deprotonation rate constant is above the catalytic constant in all enzymes investigated. The fast deprotonation requires the presence of an activator in pyruvate decarboxylase from yeast, showing the allosteric regulation of this enzyme to be accomplished by an increase in the C2-H dissociation rate of the enzyme-bound thiamin diphosphate. The data of the thiamin diphosphate analogues and of the mutant enzymes show the N1' atom and the 4'-NH2 group to be essential for the activation of the coenzyme and a conserved glutamate involved in the proton abstraction mechanism of the enzyme-bound thiamin diphosphate.

  5. Determining Enzyme Activity by Radial Diffusion

    Science.gov (United States)

    Davis, Bill D.

    1977-01-01

    Discusses advantages of radial diffusion assay in determining presence of enzyme and/or rough approximation of amount of enzyme activities. Procedures are included for the preparation of starch-agar plates, and the application and determination of enzyme. Techniques using plant materials (homogenates, tissues, ungerminated embryos, and seedlings)…

  6. Determining Enzyme Activity by Radial Diffusion

    Science.gov (United States)

    Davis, Bill D.

    1977-01-01

    Discusses advantages of radial diffusion assay in determining presence of enzyme and/or rough approximation of amount of enzyme activities. Procedures are included for the preparation of starch-agar plates, and the application and determination of enzyme. Techniques using plant materials (homogenates, tissues, ungerminated embryos, and seedlings)…

  7. Enzyme Activity Experiments Using a Simple Spectrophotometer

    Science.gov (United States)

    Hurlbut, Jeffrey A.; And Others

    1977-01-01

    Experimental procedures for studying enzyme activity using a Spectronic 20 spectrophotometer are described. The experiments demonstrate the effect of pH, temperature, and inhibitors on enzyme activity and allow the determination of Km, Vmax, and Kcat. These procedures are designed for teaching large lower-level biochemistry classes. (MR)

  8. Enzyme Activity Experiments Using a Simple Spectrophotometer

    Science.gov (United States)

    Hurlbut, Jeffrey A.; And Others

    1977-01-01

    Experimental procedures for studying enzyme activity using a Spectronic 20 spectrophotometer are described. The experiments demonstrate the effect of pH, temperature, and inhibitors on enzyme activity and allow the determination of Km, Vmax, and Kcat. These procedures are designed for teaching large lower-level biochemistry classes. (MR)

  9. Transporter-Enzyme Interplay: Deconvoluting Effects of Hepatic Transporters and Enzymes on Drug Disposition Using Static and Dynamic Mechanistic Models.

    Science.gov (United States)

    Varma, Manthena V; El-Kattan, Ayman F

    2016-07-01

    A large body of evidence suggests hepatic uptake transporters, organic anion-transporting polypeptides (OATPs), are of high clinical relevance in determining the pharmacokinetics of substrate drugs, based on which recent regulatory guidances to industry recommend appropriate assessment of investigational drugs for the potential drug interactions. We recently proposed an extended clearance classification system (ECCS) framework in which the systemic clearance of class 1B and 3B drugs is likely determined by hepatic uptake. The ECCS framework therefore predicts the possibility of drug-drug interactions (DDIs) involving OATPs and the effects of genetic variants of SLCO1B1 early in the discovery and facilitates decision making in the candidate selection and progression. Although OATP-mediated uptake is often the rate-determining process in the hepatic clearance of substrate drugs, metabolic and/or biliary components also contribute to the overall hepatic disposition and, more importantly, to liver exposure. Clinical evidence suggests that alteration in biliary efflux transport or metabolic enzymes associated with genetic polymorphism leads to change in the pharmacodynamic response of statins, for which the pharmacological target resides in the liver. Perpetrator drugs may show inhibitory and/or induction effects on transporters and enzymes simultaneously. It is therefore important to adopt models that frame these multiple processes in a mechanistic sense for quantitative DDI predictions and to deconvolute the effects of individual processes on the plasma and hepatic exposure. In vitro data-informed mechanistic static and physiologically based pharmacokinetic models are proven useful in rationalizing and predicting transporter-mediated DDIs and the complex DDIs involving transporter-enzyme interplay.

  10. Regulation of hepatic lipase activity by sphingomyelin in plasma lipoproteins

    Science.gov (United States)

    Yang, Peng; Subbaiah, Papasani V.

    2015-01-01

    Hepatic lipase (HL) is an important enzyme in the clearance of triacylglycerol (TAG) from the circulation, and has been proposed to have pro-atherogenic as well as anti-atherogenic properties. It hydrolyzes both phospholipids and TAG of lipoproteins, and its activity is negatively correlated with HDL levels. Although it is known that HL acts preferentially on HDL lipids, the basis for this specificity is not known, since it does not require any specific apoprotein for activity. In this study, we tested the hypothesis that sphingomyelin (SM), whose concentration is much higher in VLDL and LDL compared to HDL, is an inhibitor of HL, and that this could explain the lipoprotein specificity of the enzyme. The results presented show that the depletion of SM from normal lipoproteins activated the HL roughly in proportion to their SM content. SM depletion stimulated the hydrolysis of both phosphatidylcholine (PC) and TAG, although the PC hydrolysis was stimulated more. In the native lipoproteins, HL showed specificity for PC species containing polyunsaturated fatty acids at sn-2 position, and produced more unsaturated lyso PC species. The enzyme also showed preferential hydrolysis of certain TAG species over others. SM depletion affected the specificity of the enzyme towards PC and TAG species modestly. These results show that SM is a physiological inhibitor of HL activity in lipoproteins and that the specificity of the enzyme towards HDL is at least partly due to its low SM content. PMID:26193433

  11. Activation of interfacial enzymes at membrane surfaces

    DEFF Research Database (Denmark)

    Mouritsen, Ole G.; Andresen, Thomas Lars; Halperin, Avi;

    2006-01-01

    A host of water-soluble enzymes are active at membrane surfaces and in association with membranes. Some of these enzymes are involved in signalling and in modification and remodelling of the membranes. A special class of enzymes, the phospholipases, and in particular secretory phospholipase A2 (s......PLA2), are only activated at the interface between water and membrane surfaces, where they lead to a break-down of the lipid molecules into lysolipids and free fatty acids. The activation is critically dependent on the physical properties of the lipid-membrane substrate. A topical review is given...

  12. A survey of orphan enzyme activities

    Directory of Open Access Journals (Sweden)

    Pouliot Yannick

    2007-07-01

    Full Text Available Abstract Background Using computational database searches, we have demonstrated previously that no gene sequences could be found for at least 36% of enzyme activities that have been assigned an Enzyme Commission number. Here we present a follow-up literature-based survey involving a statistically significant sample of such "orphan" activities. The survey was intended to determine whether sequences for these enzyme activities are truly unknown, or whether these sequences are absent from the public sequence databases but can be found in the literature. Results We demonstrate that for ~80% of sampled orphans, the absence of sequence data is bona fide. Our analyses further substantiate the notion that many of these enzyme activities play biologically important roles. Conclusion This survey points toward significant scientific cost of having such a large fraction of characterized enzyme activities disconnected from sequence data. It also suggests that a larger effort, beginning with a comprehensive survey of all putative orphan activities, would resolve nearly 300 artifactual orphans and reconnect a wealth of enzyme research with modern genomics. For these reasons, we propose that a systematic effort to identify the cognate genes of orphan enzymes be undertaken.

  13. [Serum gamma-glutamyl transpeptidase activity in children with chronic hepatitis B].

    Science.gov (United States)

    Lebensztejn, Dariusz Marek; Skiba, Elzbieta; Sobaniec-Lotowska, Maria Elzbieta; Kaczmarski, Maciej

    2005-03-01

    THE AIM OF THE STUDY was evaluation of serum activity of chosen enzymes (ALT, AST, GGT and ALP) in assessment of fibrosis degree in children with chronic hepatitis B. We determined serum activity of liver enzymes in 47 children aged 4-16 with biopsy-verified chronic hepatitis B, prior to interferon alpha treatment. Fibrosis stage was assessed in a blinded fashion according to Batts and Ludwig. We defined advanced fibrosis as a score =3. Receiver operating characteristics (ROC) analysis was used to calculate the power of the assays to detect advanced liver fibrosis (AccuROC, Canada). Serum GGT activity of 19 IU/I had a sensitivity of 50% and a specificity of 90% (AUC=0.7324, p=0.0391) to predict advanced fibrosis. All other enzymes did not allow a useful prediction. GGT is suitable serum marker to predict advanced liver fibrosis in children with chronic hepatitis B.

  14. Visualization of enzyme activities inside earthworm pores

    Science.gov (United States)

    Hoang, Duyen; Razavi, Bahar S.

    2015-04-01

    In extremely dynamic microhabitats as bio-pores made by earthworm, the in situ enzyme activities are assumed as a footprint of complex biotic interactions. Our study focused on the effect of earthworm on the enzyme activities inside bio-pores and visualizing the differences between bio-pores and earthworm-free soil by zymography technique (Spohn and Kuzyakov, 2013). For the first time, we aimed at quantitative imaging of enzyme activities in bio-pores. Lumbricus terrestris L. was placed into transparent box (15×20×15cm). After two weeks when bio-pore systems were formed by earthworms, we visualized in situ enzyme activities of five hydrolytic enzymes (β-glucosidase, cellobiohydrolase, chitinase, xylanase, leucine-aminopeptidase, and phosphatase. Zymography showed higher activity of β-glucosidase, chitinase, xylanase and phosphatase in biopores comparing to bulk soil. However, the differences in activity of cellobiohydrolase and leucine aminopeptidase between bio-pore and bulk soil were less pronounced. This demonstrated an applicability of zymography approach to monitor and to distinguish the in situ activity of hydrolytic enzymes in soil biopores.

  15. How thiamine diphosphate is activated in enzymes.

    Science.gov (United States)

    Kern, D; Kern, G; Neef, H; Tittmann, K; Killenberg-Jabs, M; Wikner, C; Schneider, G; Hübner, G

    1997-01-03

    The controversial question of how thiamine diphosphate, the biologically active form of vitamin B1, is activated in different enzymes has been addressed. Activation of the coenzyme was studied by measuring thermodynamics and kinetics of deprotonation at the carbon in the 2-position (C2) of thiamine diphosphate in the enzymes pyruvate decarboxylase and transketolase by use of nuclear magnetic resonance spectroscopy, proton/deuterium exchange, coenzyme analogs, and site-specific mutant enzymes. Interaction of a glutamate with the nitrogen in the 1'-position in the pyrimidine ring activated the 4'-amino group to act as an efficient proton acceptor for the C2 proton. The protein component accelerated the deprotonation of the C2 atom by several orders of magnitude, beyond the rate of the overall enzyme reaction. Therefore, the earlier proposed concerted mechanism or stabilization of a C2 carbanion can be excluded.

  16. Viral hepatitis prevalence in patients with active and latent tuberculosis

    Science.gov (United States)

    Nooredinvand, Hesam Ahmadi; Connell, David W; Asgheddi, Mahmoud; Abdullah, Mohammed; O’Donoghue, Marie; Campbell, Louise; Wickremasinghe, Melissa I; Lalvani, Ajit; Kon, Onn Min; Khan, Shahid A

    2015-01-01

    AIM: To assess the prevalence of hepatitis B virus (HBV) and hepatitis C virus (HCV) infection and association with drug induced liver injury (DILI) in patients undergoing anti-tuberculosis (TB) therapy. METHODS: Four hundred and twenty nine patients with newly diagnosed TB - either active disease or latent infection - who were due to commence anti-TB therapy between September 2008 and May 2011 were included. These patients were prospectively tested for serological markers of HBV, HCV and human immunodeficiency virus (HIV) infections - hepatitis B core antigen (HBcAg), hepatitis B surface antigen (HBsAg), hepatitis B e antigen, IgG and IgM antibody to HBcAg (anti-HBc), HCV IgG antibody and HIV antibody using a combination of enzyme-linked immunosorbent assay, Western blot assay and polymerase chain reaction techniques. Patients were reviewed at least monthly during the TB treatment initiation phase. Liver function tests were measured prior to commencement of anti-TB therapy and 2-4 wk later. Liver function tests were also performed at any time the patient had significant nausea, vomiting, rash, or felt non-specifically unwell. Fisher’s exact test was used to measure significance in comparisons of proportions between groups. A P value of less than 0.05 was considered statistically significant. RESULTS: Of the 429 patients, 270 (62.9%) had active TB disease and 159 (37.1%) had latent TB infection. 61 (14.2%) patients had isolated anti-HBc positivity, 11 (2.6%) were also HBsAg positive and 7 (1.6%) were HCV-antibody positive. 16/270 patients with active TB disease compared to 2/159 patients with latent TB infection had markers of chronic viral hepatitis (HBsAg or HCV antibody positive; P = 0.023). Similarly the proportion of HBsAg positive patients were significantly greater in the active vs latent TB infection group (10/43 vs 1/29, P = 0.04). The prevalence of chronic HBV or HCV was significantly higher than the estimated United Kingdom prevalence of 0.3% for each

  17. Effects of phenoxyherbicides and glyphosate on the hepatic and intestinal biotransformation activities in the rat.

    Science.gov (United States)

    Hietanen, E; Linnainmaa, K; Vainio, H

    1983-08-01

    The effects of phenoxyacid herbicides 2,4-D (2,4-dichlorophenoxyacetic acid) and MCPA (4-chloro-2-methylphenoxyacetic acid), clofibrate, and glyphosate on hepatic and intestinal drug metabolizing enzyme activities were studied in rats intragastrically exposed for 2 weeks. The hepatic ethoxycoumarin O-deethylase activity increased about 2-fold with MCPA. Both 2,4-D and MCPA increased the hepatic epoxide hydrolase activity and decreased the hepatic glutathione S-transferase activity. MCPA also increased the intestinal activities of ethoxycoumarin O-deethylase and epoxide hydrolase. Glyphosate decreased the hepatic level of cytochrome P-450 and monooxygenase activities and the intestinal activity of aryl hydrocarbon hydroxylase. Clofibrate decreased the hepatic activities of UDPglucuronosyltransferase with p-nitrophenol or methylumbelliferone as the substrate. Also 2,4-D decreased the hepatic activity of UDPglucuronosyltransferase with p-nitrophenol as the substrate. MCPA decreased the intestinal activities of UDPglucuronosyltransferase with either p-nitrophenol or methylumbelliferone as the substrate. The results indicate that phenoxyacetic acids, especially MCPA, may have potent effects on the metabolism of xenobiotics. Glyphosate, not chemically related to phenoxyacids, seems to inhibit monooxygenases. Whether these changes are related to the toxicity of these xenobiotics remains to be clarified in further experiments.

  18. Soil Enzyme Activities with Greenhouse Subsurface Irrigation

    Institute of Scientific and Technical Information of China (English)

    ZHANG Yu-Long; WANG Yao-Sheng

    2006-01-01

    Various environmental conditions determine soil enzyme activities, which are important indicators for changes of soil microbial activity, soil fertility, and land quality. The effect of subsurface irrigation scheduling on activities of three soil enzymes (phosphatase, urease, and catalase) was studied at five depths (0-10, 10-20, 20-30, 30-40, and 40-60 cm) of a tomato greenhouse soil. Irrigation was scheduled when soil water condition reached the maximum allowable depletion(MAD) designed for different treatments (-10, -16, -25, -40, and -63 kPa). Results showed that soil enzyme activities had significant responses to the irrigation scheduling during the period of subsurface irrigation. The neutral phosphatase activity and the catalase activity were found to generally increase with more frequent irrigation (MAD of -10 and -16kPa). This suggested that a higher level of water content favored an increase in activity of these two enzymes. In contrast,the urease activity decreased under irrigation, with less effect for MAD of -40 and -63 kPa. This implied that relatively wet soil conditions were conducive to retention of urea N, but relatively dry soil conditions could result in increasing loss of urea N. Further, this study revealed that soil enzyme activities could be alternative natural bio-sensors for the effect of irrigation on soil biochemical reactions and could help optimize irrigation management of greenhouse crop production.

  19. Enzyme activity in dialkyl phosphate ionic liquids

    Energy Technology Data Exchange (ETDEWEB)

    Thomas, M.F.; Dunn, J.; Li, L.-L.; Handley-Pendleton, J. M.; van der lelie, D.; Wishart, J. F.

    2011-12-01

    The activity of four metagenomic enzymes and an enzyme cloned from the straw mushroom, Volvariellavolvacea were studied in the following ionic liquids, 1,3-dimethylimidazolium dimethyl phosphate, [mmim][dmp], 1-ethyl-3-methylimidazolium dimethyl phosphate, [emim][dmp], 1-ethyl-3-methylimidazolium diethyl phosphate, [emim][dep] and 1-ethyl-3-methylimidazolium acetate, [emim][OAc]. Activity was determined by analyzing the hydrolysis of para-nitrobenzene carbohydrate derivatives. In general, the enzymes were most active in the dimethyl phosphate ionic liquids, followed by acetate. Generally speaking, activity decreased sharply for concentrations of [emim][dep] above 10% v/v, while the other ionic liquids showed less impact on activity up to 20% v/v.

  20. Enzyme activity in dialkyl phosphate ionic liquids.

    Science.gov (United States)

    Thomas, Marie F; Li, Luen-Luen; Handley-Pendleton, Jocelyn M; van der Lelie, Daniel; Dunn, John J; Wishart, James F

    2011-12-01

    The activity of four metagenomic enzymes and an enzyme cloned from the straw mushroom, Volvariella volvacea were studied in the following ionic liquids, 1,3-dimethylimidazolium dimethyl phosphate, [mmim][dmp], 1-ethyl-3-methylimidazolium dimethyl phosphate, [emim][dmp], 1-ethyl-3-methylimidazolium diethyl phosphate, [emim][dep] and 1-ethyl-3-methylimidazolium acetate, [emim][OAc]. Activity was determined by analyzing the hydrolysis of para-nitrobenzene carbohydrate derivatives. In general, the enzymes were most active in the dimethyl phosphate ionic liquids, followed by acetate. Generally speaking, activity decreased sharply for concentrations of [emim][dep] above 10% v/v, while the other ionic liquids showed less impact on activity up to 20% v/v.

  1. [Hepatic manifestation of a macrophage activation syndrome (MAS)].

    Science.gov (United States)

    Nagel, Michael; Schwarting, Andreas; Straub, Beate K; Galle, Peter R; Zimmermann, Tim

    2017-05-01

    Background Elevated liver values are the most common pathological laboratory result in Germany. Frequent findings, especially in younger patients, are nutritive- or medicamentous- toxic reasons, viral or autoimmune hepatitis. A macrophage activation syndrome (MAS) may manifest like a viral infectious disease with fever, hepatosplenomegaly and pancytopenia and is associated with a high mortality. It is based on an enhanced activation of macrophages with increased cytokine release, leading to organ damage and multi-organ failure. In addition to genetic causes, MAS is commonly associated with infections and rheumatic diseases. We report the case of a 26-year-old female patient suffering from MAS as a rare cause of elevated liver enzymes. Methods Patient characteristics, laboratory values, liver histology, bone marrow and radiological imaging were documented and analyzed. Case Report After an ordinary upper airway infection with bronchitis, a rheumatic arthritis appeared and was treated with leflunomide und methotrexate. In the further course of the disease, the patient developed an acute hepatitis with fever, pancytopenia and massive hyperferritinemia. Immunohistochemistry of the liver biopsy revealed hemophagocytosis and activation of CD68-positive macrophages. In the radiological and histological diagnostics of the liver and bone marrow, an MAS was diagnosed as underlying disease of the acute hepatitis. Under therapy with prednisolone, the fever disappeared and transaminases and ferritin rapidly normalized. Conclusion Aside from the frequent causes of elevated liver values in younger patients, such as nutritive toxic, drug induced liver injury, viral or autoimmune hepatitis, especially in case of massive hyperferritinemia, a MAS should be considered as a rare cause of acute liver disease. © Georg Thieme Verlag KG Stuttgart · New York.

  2. Dietary phytic acid prevents fatty liver by reducing expression of hepatic lipogenic enzymes and modulates gut microflora in rats fed a high-sucrose diet.

    Science.gov (United States)

    Sekita, Ayaka; Okazaki, Yukako; Katayama, Tetsuyuki

    2016-06-01

    The aim of this study was to investigate the effect of phytic acid (PA) on fatty liver and gut microflora in rats fed a high-sucrose (HSC) diet. Three groups of rats were fed a high-starch (HSR) diet or an HSC diet with or without 1.02% sodium PA for 12 d. We evaluated hepatic weight, total lipids, and triacylglycerol (TG) levels, the activities and expression of hepatic lipogenic enzymes (glucose-6-phosphate dehydrogenase, malic enzyme 1, and fatty acid synthetase), and fecal microflora. The HSC diet significantly increased hepatic total lipids and TG levels, and the activities and expression of the hepatic lipogenic enzymes compared with the HSR diet. These upregulations were clearly suppressed by dietary PA. Consumption of PA elevated the fecal ratio of Lactobacillus spp. and depressed the ratio of Clostridium cocoides, and suppressed the elevation in the ratio of C. leptum induced by the HSC diet. This work showed that dietary PA ameliorates sucrose-induced fatty liver through reducing the expression of hepatic lipogenesis genes and modulates gut microflora in rats. Copyright © 2016 Elsevier Inc. All rights reserved.

  3. Central Insulin Action Activates Kupffer Cells by Suppressing Hepatic Vagal Activation via the Nicotinic Alpha 7 Acetylcholine Receptor

    Directory of Open Access Journals (Sweden)

    Kumi Kimura

    2016-03-01

    Full Text Available Central insulin action activates hepatic IL-6/STAT3 signaling, which suppresses the gene expression of hepatic gluconeogenic enzymes. The vagus nerve plays an important role in this centrally mediated hepatic response; however, the precise mechanism underlying this brain-liver interaction is unclear. Here, we present our findings that the vagus nerve suppresses hepatic IL-6/STAT3 signaling via α7-nicotinic acetylcholine receptors (α7-nAchR on Kupffer cells, and that central insulin action activates hepatic IL-6/STAT3 signaling by suppressing vagal activity. Indeed, central insulin-mediated hepatic IL-6/STAT3 activation and gluconeogenic gene suppression were impeded in mice with hepatic vagotomy, pharmacological cholinergic blockade, or α7-nAchR deficiency. In high-fat diet-induced obese and insulin-resistant mice, control of the vagus nerve by central insulin action was disturbed, inducing a persistent increase of inflammatory cytokines. These findings suggest that dysregulation of the α7-nAchR-mediated control of Kupffer cells by central insulin action may affect the pathogenesis of chronic hepatic inflammation in obesity.

  4. Predictors of hepatitis B cure using gene therapy to deliver DNA cleavage enzymes: a mathematical modeling approach.

    Directory of Open Access Journals (Sweden)

    Joshua T Schiffer

    Full Text Available Most chronic viral infections are managed with small molecule therapies that inhibit replication but are not curative because non-replicating viral forms can persist despite decades of suppressive treatment. There are therefore numerous strategies in development to eradicate all non-replicating viruses from the body. We are currently engineering DNA cleavage enzymes that specifically target hepatitis B virus covalently closed circular DNA (HBV cccDNA, the episomal form of the virus that persists despite potent antiviral therapies. DNA cleavage enzymes, including homing endonucleases or meganucleases, zinc-finger nucleases (ZFNs, TAL effector nucleases (TALENs, and CRISPR-associated system 9 (Cas9 proteins, can disrupt specific regions of viral DNA. Because DNA repair is error prone, the virus can be neutralized after repeated cleavage events when a target sequence becomes mutated. DNA cleavage enzymes will be delivered as genes within viral vectors that enter hepatocytes. Here we develop mathematical models that describe the delivery and intracellular activity of DNA cleavage enzymes. Model simulations predict that high vector to target cell ratio, limited removal of delivery vectors by humoral immunity, and avid binding between enzyme and its DNA target will promote the highest level of cccDNA disruption. Development of de novo resistance to cleavage enzymes may occur if DNA cleavage and error prone repair does not render the viral episome replication incompetent: our model predicts that concurrent delivery of multiple enzymes which target different vital cccDNA regions, or sequential delivery of different enzymes, are both potentially useful strategies for avoiding multi-enzyme resistance. The underlying dynamics of cccDNA persistence are unlikely to impact the probability of cure provided that antiviral therapy is given concurrently during eradication trials. We conclude by describing experiments that can be used to validate the model, which

  5. Activity assessment of microbial fibrinolytic enzymes.

    Science.gov (United States)

    Kotb, Essam

    2013-08-01

    Conversion of fibrinogen to fibrin inside blood vessels results in thrombosis, leading to myocardial infarction and other cardiovascular diseases. In general, there are four therapy options: surgical operation, intake of antiplatelets, anticoagulants, or fibrinolytic enzymes. Microbial fibrinolytic enzymes have attracted much more attention than typical thrombolytic agents because of the expensive prices and the side effects of the latter. The fibrinolytic enzymes were successively discovered from different microorganisms, the most important among which is the genus Bacillus. Microbial fibrinolytic enzymes, especially those from food-grade microorganisms, have the potential to be developed as functional food additives and drugs to prevent or cure thrombosis and other related diseases. There are several assay methods for these enzymes; this may due to the insolubility of substrate, fibrin. Existing assay methods can be divided into three major groups. The first group consists of assay of fibrinolytic activity with natural proteins as substrates, e.g., fibrin plate methods. The second and third groups of assays are suitable for kinetic studies and are based on the determination of hydrolysis of synthetic peptide esters. This review will deal primarily with the microorganisms that have been reported in literature to produce fibrinolytic enzymes and the first review discussing the methods used to assay the fibrinolytic activity.

  6. Influence of dietary macronutrients on induction of hepatic drug metabolizing enzymes by Coleus forskohlii extract in mice.

    Science.gov (United States)

    Yokotani, Kaori; Chiba, Tsuyoshi; Sato, Yoko; Nakanishi, Tomoko; Murata, Masatsune; Umegaki, Keizo

    2013-01-01

    From studies in mice, we have reported that Coleus forskohlii extract (CFE), a popular herbal weight-loss ingredient, markedly induced hepatic drug metabolizing enzymes, especially cytochrome P450 (CYP), and interacted with co-administered drugs. This study was designed to examine how the induction of drug metabolizing enzymes by CFE was influenced by different levels of macronutrients in the diet. Mice were fed a non-purified diet or semi-purified diet with and without CFE (0.3-0.5%) for 14-18 d, and changes in the ratio of liver weight to body weight, an indicator of hepatic CYP induction, and hepatic drug metabolizing enzymes were analyzed. The ratio of liver weight to body weight, content and activities of CYPs, and activity of glutathione S-transferase were higher in a semi-purified standard diet (AIN93G formula) group than in high sucrose (62.9%) and high fat (29.9%) diet groups. Different levels of protein (7%, 20%, and 33%) in the diets did not influence CFE-induced CYP induction or increase the ratio of liver weight to body weight. The effect of CFE on the ratio of liver weight to body weight was higher with a semi-purified diet than with a non-purified diet, and was similar between dietary administration and intragastric gavage when the CFE dose and the diet were the same. There was a positive correlation between CFE-induced CYP induction and the content of starch in the diets, suggesting that dietary starch potentiates CFE-induced CYP induction in mice. The mechanism of enhanced CYP induction remains unclear.

  7. Effect of the combined probiotics with aflatoxin B₁-degrading enzyme on aflatoxin detoxification, broiler production performance and hepatic enzyme gene expression.

    Science.gov (United States)

    Zuo, Rui-yu; Chang, Juan; Yin, Qing-qiang; Wang, Ping; Yang, Yu-rong; Wang, Xiao; Wang, Guo-qiang; Zheng, Qiu-hong

    2013-09-01

    In order to degrade aflatoxin B₁ (AFB₁), AFB₁-degrading microbes (probiotics) such as Lactobacillus casei, Bacillus subtilis and Pichia anomala, and the AFB₁-degrading enzyme from Aspergillus oryzae were selected and combined to make feed additive. Seventy-five 43-day-old male Arbor Acres broilers were randomly divided into 5 groups, 15 broilers for each group. The broilers were given with 5 kinds of diets such as the basal diet, 400 μg/kg AFB₁ supplement without feed additive, and 200, 400, 800 μg/kg AFB₁ supplement with 0.15% feed additive. The feeding experimental period was 30 d, which was used to determine production performance of broilers. In addition, serum, liver and chest muscle were selected for measuring AFB₁ residues, gene expressions, microscopic and antioxidant analyses. The results showed that adding 0.15% feed additive in broiler diets could significantly relieve the negative effect of AFB₁ on chicken's production performance and nutrient metabolic rates (P<0.05). It could also improve AFB₁ metabolism, hepatic cell structure, antioxidant activity, and many hepatic enzyme gene expressions involved in oxidoreductase, apoptosis, cell growth, immune system and metabolic process (P<0.05). It could be concluded that the feed additive was able to degrade AFB₁ and improve animal production.

  8. Exploratory Studies of (-)-Epicatechin, a Bioactive Compound of Phyllanthus niruri, on the Antioxidant Enzymes and Oxidative Stress Markers in D-galactosamine-induced Hepatitis in Rats: A Study with Reference to Clinical Prospective.

    Science.gov (United States)

    Shanmugam, Bhasha; Shanmugam, Kondeti Ramudu; Ravi, Sahukari; Subbaiah, Ganjikunta Venkata; Ramakrishana, Chilakala; Mallikarjuna, Korivi; Reddy, Kesireddy Sathyavelu

    2017-01-01

    Hepatitis is a health problem affecting millions of people worldwide and it is the major risk factor for liver cirrhosis. In India, many plants are used to treat hepatitis. But little is known about the effects of (-)-epicatechin a bioactive compound of Phyllanthus niruri (PN) in hepatitis rats. The present study was designed to explore the antioxidant property of (-)-epicatechin isolated from PN in D-Galactosamine (D-GalN) induced hepatitis rats. The rats are divided into five groups as per the experimental design. (-)-Epicatchin pretreatment was given to the hepatitis rats for 21 days and biochemical analysis was carried out. The hepatic antioxidant enzymes like superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPx), and glutathione reductase (GR), glutathione S-transferase (GST), reduced glutathione (GSH), and malondialdehyde (MDA) and serum markers aspartate aminotransferase (AST), alanine aminotransferase (ALT), alkaline phosphatase (ALP), albumin, and bilirubin are estimated. All the antioxidant enzymes activities and albumin levels are depleted in hepatitic rats. Whereas GST, ALP, AST, ALT activities and MDA, and bilirubin levels are elevated in hepatitis rats, (-)-epicatechin pretreatment increased all the antioxidant enzymes and decreased the GST, ALP, AST, ALT, and MDA levels in hepatitis rats. However, histopatholoigic studies also proves that (-)-epicatechin pretreatment decreased the tissue damage in hepatitis condition. This is the first report on the antioxidant enzymes and hepatoprotective effect of (-)-epicatechin in hepatitis rats. From this study, we conclude that (-)-epicatechin treatment decreased the oxidative damage in hepatitis rats. The present study was carried out to know the impact of (-)-epicatechin on antioxidant enzymes activities in hepatitis rats. From this study, we found that the antioxidant enzymes SOD, CAT, GPx, GR, GSH depleted in hepatitis rats and increased with (-)-epicatechin in hepatitis rats.MDA levels

  9. A modern mode of activation for nucleic acid enzymes.

    Directory of Open Access Journals (Sweden)

    Dominique Lévesque

    Full Text Available Through evolution, enzymes have developed subtle modes of activation in order to ensure the sufficiently high substrate specificity required by modern cellular metabolism. One of these modes is the use of a target-dependent module (i.e. a docking domain such as those found in signalling kinases. Upon the binding of the target to a docking domain, the substrate is positioned within the catalytic site. The prodomain acts as a target-dependent module switching the kinase from an off state to an on state. As compared to the allosteric mode of activation, there is no need for the presence of a third partner. None of the ribozymes discovered to date have such a mode of activation, nor does any other known RNA. Starting from a specific on/off adaptor for the hepatitis delta virus ribozyme, that differs but has a mechanism reminiscent of this signalling kinase, we have adapted this mode of activation, using the techniques of molecular engineering, to both catalytic RNAs and DNAs exhibiting various activities. Specifically, we adapted three cleaving ribozymes (hepatitis delta virus, hammerhead and hairpin ribozymes, a cleaving 10-23 deoxyribozyme, a ligating hairpin ribozyme and an artificially selected capping ribozyme. In each case, there was a significant gain in terms of substrate specificity. Even if this mode of control is unreported for natural catalytic nucleic acids, its use needs not be limited to proteinous enzymes. We suggest that the complexity of the modern cellular metabolism might have been an important selective pressure in this evolutionary process.

  10. Strain differences in the induction of hepatic microsomal cytochrome P450 and drug-metabolizing enzymes by climbazole in rats

    National Research Council Canada - National Science Library

    Michiya Suzuki; Yasuna Kobayashi; Takashi Sunagawa; Takiko Oguro; Shogo Tokuyama; Toshinori Yamamoto; Takemi Yoshida

    2000-01-01

    .... Climbazole is one of imidazole-ring containing antifungal agents. However, there is not enough data concerned with their effect on hepatic drug metabolizing enzymes with respect to the strain differences...

  11. FLUCONAZOLE-INDUCED HEPATIC CYTOCHROME P450 GENE EXPRESSION AND ENZYMATIC ACTIVITIES IN RATS AND MICE

    Science.gov (United States)

    This study was undertaken to examine the effects of the triazole antifungal agent fluconazole on the expression of hepatic cytochrome P450 (Cyp) genes and the activities of Cyp enzymes in male Sprague-Dawley rats and male CD-1 mice. Alkoxyresorufin O-dealkylation (AROD) methods w...

  12. An NMR Study of Enzyme Activity.

    Science.gov (United States)

    Peterman, Keith E.; And Others

    1989-01-01

    A laboratory experiment designed as a model for studying enzyme activity with a basic spectrometer is presented. Included are background information, experimental procedures, and a discussion of probable results. Stressed is the value of the use of Nuclear Magnetic Resonance in biochemistry. (CW)

  13. An NMR Study of Enzyme Activity.

    Science.gov (United States)

    Peterman, Keith E.; And Others

    1989-01-01

    A laboratory experiment designed as a model for studying enzyme activity with a basic spectrometer is presented. Included are background information, experimental procedures, and a discussion of probable results. Stressed is the value of the use of Nuclear Magnetic Resonance in biochemistry. (CW)

  14. Arabinogalactan proteins: focus on carbohydrate active enzymes

    Directory of Open Access Journals (Sweden)

    Eva eKnoch

    2014-06-01

    Full Text Available Arabinogalactan proteins (AGPs are a highly diverse class of cell surface proteoglycans that are commonly found in most plant species. AGPs play important roles in many cellular processes during plant development, such as reproduction, cell proliferation, pattern formation and growth, and in plant-microbe interaction. However, little is known about the molecular mechanisms of their function. Numerous studies using monoclonal antibodies that recognize different AGP glycan epitopes have shown the appearance of a slightly altered AGP glycan in a specific stage of development in plant cells. Therefore, it is anticipated that the biosynthesis and degradation of AGP glycan is tightly regulated during development. Until recently, however, little was known about the enzymes involved in the metabolism of AGP glycans. In this review, we summarize recent discoveries of carbohydrate active enzymes (CAZy; http://www.cazy.org/ involved in the biosynthesis and degradation of AGP glycans, and we discuss the biological role of these enzymes in plant development.

  15. Effects of arecoline on hepatic cytochrome P450 activity and oxidative stress.

    Science.gov (United States)

    Run-mei, Xiao; Jun-jun, Wang; Jing-ya, Chen; Li-juan, Sun; Yong, Chen

    2014-08-01

    Betel-quid use is associated with the risk of liver cirrhosis and hepatocellular carcinoma. The aim of the present work was to evaluate the impact of arecoline on human hepatic cytochrome P450 (CYP) enzymes in vitro and rat hepatic CYP enzymes, as well as the hepatic oxidative stress and liver injury of rats in vivo. The in vitro results indicated that arecoline hydrobromide (AH) has no significant effect on the activities of CYP2B, 2C9, 3A4, 1A2, 2E1 and 2D6 in human liver microsome (HLM). However, oral administration of AH at 4 and 20 mg/kg/d for seven consecutive days significantly increased the activities of rat hepatic CYP2B, 2E1, 2D, 3A, 2C and 1A2. In addition, AH at 100 mg/kg/d significantly increased the levels of ALT, AST and MDA, decreased the levels of SOD, CAT, GSH-Px and GSH, in rat liver. The in vivo induction of AH on rat hepatic CYP isoforms suggested that the high risk of metabolic interaction should be existed when the substrate drugs of the six kinds of CYP isoforms was administered in betel-quid use human. Furthermore, the in vivo results also suggested that AH-induced hepatoxicity should be associated with the induction of AH on rat hepatic CYP2E1 and 2B.

  16. The effect of bamboo extract on hepatic biotransforming enzymes – Findings from an obese–diabetic mouse model

    Science.gov (United States)

    Koide, Cheryl L.K.; Collier, Abby C.; Berry, Marla J.; Panee, Jun

    2012-01-01

    Aim of the study Bamboo leaves are used as a component in traditional Chinese medicine for the anti-inflammatory function. Our previous studies have demonstrated that an ethanol/water extract from Phyllostachys edulis ameliorated obesity-associated chronic systemic inflammation in mice, and therefore relieving the symptoms of type 2 diabetes. The aim of this project was to further investigate the effects of this bamboo extract on hepatic biotransformation enzymes in both lean and obese mice, as an initial step in the toxicological evaluation of using this traditional medicine in obese/diabetic population. Materials and methods Male C57BL/6J mice were randomized to 4 groups and fed standard (10% kcal from fat) diet with or without bamboo extract supplementation at a dose of 10 gram per kilogram diet (n = 10 and n = 9, respectively), or high fat (45% kcal from fat) diet with or without bamboo extract (n = 8 and N = 7, respectively). The dietary treatment lasted for 6 months. Subsequently, the activities and expression of the major Phase I and II hepatic biotransformation enzymes were assessed in subcellular fractions from murine livers. Results Three groups of mice, lean bamboo extract-supplemented, obese/diabetic, and bamboo extract-supplemented obese/diabetic, showed greater activities of cytochromes P450 1a2 and 3a11 compared to control but no changes in the expression level of these proteins. For Phase II enzymes, bamboo extract supplementation in lean mice caused decreased glutathione-S-transferase activity (−12%) and greater uridine diphosphate glucuronosyltransferase activity (+46%), but had no effect on sulfotransferase activity. Conversely, the obese/diabetic condition itself increased glutathione-S-transferase and uridine diphosphate glucuronosyltransferase activities, but decreased total sulfotransferase activity and sulfotransferase 2a1 expression. Conclusions Bamboo extract and obesity/diabetes show significant independent effects on hepatic bio

  17. Enzyme Activities in Waste Water and Activated Sludge

    DEFF Research Database (Denmark)

    Nybroe, Ole; Jørgensen, Per Elberg; Henze, Mogens

    1992-01-01

    The purpose of the present study was to evaluate the potential of selected enzyme activity assays to determine microbial abundance and heterotrophic activity in waste water and activated sludge. In waste water, esterase and dehydrogenase activities were found to correlate with microbial abundance...... measured as colony forming units of heterotrophic bacteria. A panel of four enzyme activity assays, α-glucosidase, alanine-aminopeptidase, esterase and dehydrogenase were used to characterize activated sludge and anaerobic hydrolysis sludge from a pilot scale plant. The enzymatic activity profiles were...... distinctly different, suggesting that microbial populations were different, or had different physiological properties, in the two types of sludge. Enzyme activity profiles in activated sludge from four full-scale plants seemed to be highly influenced by the composition of the inlet. Addition of hydrolysed...

  18. Hydrolytic enzyme activity enhanced by Barium supplementation

    Directory of Open Access Journals (Sweden)

    Camilo Muñoz

    2016-10-01

    Full Text Available Hydrolysis of polymers is a first and often limiting step during the degradation of plant residues. Plant biomass is generally a major component of waste residues and a major renewable resource to obtain a variety of secondary products including biofuels. Improving the performance of enzymatic hydrolysis of plant material with minimum costs and limiting the use of additional microbial biomass or hydrolytic enzymes directly influences competitiveness of these green biotechnological processes. In this study, we cloned and expressed a cellulase and two esterases recovered from environmental thermophilic soil bacterial communities and characterize their optimum activity conditions including the effect of several metal ions. Results showed that supplementing these hydrolytic reactions with Barium increases the activity of these extracellular hydrolytic enzymes. This observation represents a simple but major improvement to enhance the efficiency and competitiveness of this process within an increasingly important biotechnological sector.

  19. Chronic Elevation of Liver Enzymes in Acute Intermittent Porphyria Initially Misdiagnosed as Autoimmune Hepatitis

    Directory of Open Access Journals (Sweden)

    A. González Estrada

    2011-01-01

    Full Text Available Autoimmune hepatitis is a disease characterized by an elevation of liver enzymes, as well as specific autoantibodies. It is more common in women than men. We describe a 32-year-old woman with elevated transaminases, autoantibodies, and a liver biopsy result suggestive of autoimmune hepatitis. The indicated treatment was administered without showing a satisfactory response. The patient had a family history of acute intermittent porphyria (AIP so we decided to begin treatment with hematin, achieving a complete remission of the symptoms. Acute intermittent porphyria is a rare condition characterized by neurovisceral symptoms, abdominal pain being the most common of them. The disease has a higher prevalence among young women and certain European countries such as Sweden, Great Britain, and Spain. A correct diagnosis and prompt treatment are essential because patients affected by AIP must have a strict followup due to the fatal outcome of the outbreaks.

  20. Effect of combined pharmacotherapy lipid peroxidation and activity of enzymes antioxidant protection in rat livergepatoсancerogeneze

    OpenAIRE

    KASYMOVA GULMIRA GAFUROVNA

    2016-01-01

    When hepatitis carcinogenesis marked imbalance in the system of lipid peroxidation and antioxidant enzyme system. Pharmacotherapy cytostatic leads to even greater intensification of lipid peroxidation. Thus activity of antioxidant defense enzymes even more oppressed. Roncoleukin results to some rebalancing of the system of lipid peroxidation and antioxidant enzyme systems. Which has a temporary nature. The combined use of doxorubicin and ronkolejkin some what reduces the marked of doxorubicin...

  1. Hepatitis

    Institute of Scientific and Technical Information of China (English)

    2008-01-01

    2008312 Impact of hepatitis B virus infection on the activity of hematopoietic stem cell.SHI Yanmei(石雁梅),et al.Dept Infect Dis,1st Clin Coll,Harbin Med Univ,Harbin 150001.Chin J Infect Dis 2008;26(4):197-201.Objective To study the impact of hepatitis B virus (HBV)infection on the activity of cord hematopoieticstem cells.Methods CD34+cells were isolated from healthy human cord blood by mini MACS.Cells were

  2. Successful treatment of activated occult hepatitis B in a non-responder chronic hepatitis C patient

    OpenAIRE

    Emara Mohamed H; Radwan Mohamed I

    2011-01-01

    Abstract We reported a 23 years old male with chronic hepatitis C virus infection, discontinued from pegylated interferon/ribavirin combination therapy due to a lack of early virological response. He has developed activation of occult hepatitis B virus that was successfully treated by a one year of lamivudine therapy.

  3. Effects of Arctium lappa aqueous extract on lipid profile and hepatic enzyme levels of sucrose-induced metabolic syndrome in female rats

    Directory of Open Access Journals (Sweden)

    Akram Ahangarpour

    Full Text Available ABSTRACT Arctium lappa is known to have antioxidant and antidiabetic effects in traditional medicine. Objectives: The aim of this paper was to study the effects of A. lappa root extract (AE on lipid profile and hepatic enzyme levels in sucrose-induced metabolic syndrome (MS in female rats. The study used 40 adult female Wistar rats weighing 150 g-250 g randomly divided into five groups: control, metabolic syndrome (MS, metabolic syndrome+AE at 50,100, 200 mg/kg. MS was induced by administering 50% sucrose in drinking water for 6 weeks. AE was intra-peritoneally administered daily at doses of 50,100, and 200 mg/kg for two sequential weeks at the end of the fourth week in metabolic syndrome rats. Twenty-four hours after the last administration of AE, blood was collected and centrifuged, and then the serum was used for the measurement of lipid profile and hepatic enzyme. Serum glucose, insulin, fasting insulin resistance index, body weight, water intake, lipid profile, and hepatic enzymes were significantly increased although food intake was decreased in MS rats compared to the control rats. The lipids and liver enzymes were reduced by AE extracts in the MS group. This study showed that the A. lappa root aqueous extract exhibits a hypolipidemic activity of hyperlipidemic rats. This activity is practically that of a triple-impact antioxidant, hypolipidemic, and hepatoprotective.

  4. Following Enzyme Activity with Infrared Spectroscopy

    Directory of Open Access Journals (Sweden)

    Saroj Kumar

    2010-03-01

    Full Text Available Fourier transform infrared (FTIR spectroscopy provides a direct, "on-line" monitor of enzymatic reactions. Measurement of enzymatic activity is based on the fact that the infrared spectra of reactants and products of an enzymatic reaction are usually different. Several examples are given using the enzymes pyruvate kinase, fumarase and alcohol dehydrogenase. The main advantage of the infrared method is that it observes the reaction of interest directly, i.e.,no activity assay is required to convert the progress of the reaction into an observable quantity.

  5. Effect of alachlor on hepatic cytochrome P450 enzymes in rats.

    Science.gov (United States)

    Hanioka, Nobumitsu; Watanabe, Kayoko; Yoda, Reiko; Ando, Masanori

    2002-02-01

    Alachlor ((2-chloro-N-methoxymethyl)-N-(2,6-diethylphenyl)acetamide) is a widely used preemergence herbicide which has been classified by the USEPA as a probable human carcinogen. The herbicide has been suggested to be metabolized by hepatic cytochrome P450 system. We examined the effects of alachlor on cytochrome P450 enzymes in rat liver microsomes. Rats were treated intraperitoneally with alachlor daily for 5 days, at doses of 25, 50 and 100 mg/kg. Among the cytochrome P450-dependent monooxygenase activities, 7-pentoxyresorufin O-depentylase, which is associated with CYP2B1, was dose-dependently increased by alachlor. The induction relative to control activity was 1.7-4.2-fold. The activities of CYP1A-dependent monooxygenases such as 7-ethoxy-resorufin O-deethylase and acetanilide 4-hydroxylase were also significantly increased by alachlor at doses of 50 and 100 mg/kg (1.7-2.1-fold). Furthermore, immunoblotting showed that alachlor significantly increased CYP2B1/2 and CYP1A1/2 protein levels by 4.2-6.3- and 1.8-fold, respectively. Although 7-ethoxycoumarin O-deethylase, bufuralol 1'-hydroxylase and 4-nitrophenol 2-hydroxylase activities were significantly increased by alachlor at higher doses (> or = 50 mg/kg), the induction ratios were less than 1.6-fold. The activities of other cytochrome P450-dependent monooxygenases, namely testosterone 7 alpha-hydroxylase, testosterone 2 alpha-hydroxylase, testosterone 6 beta-hydroxylase and lauric acid omega-hydroxylase, were not affected by alachlor at any dose. In addition, there was no significant change in the protein levels of CYP2C11/6, CYP2D1, CYP2E1, CYP3A2/1 and CYP4A1/2/3. These results suggest that alachlor selectively induces cytochrome P450 isoforms of the CYP1A and CYP2B subfamilies in rat liver microsomes, and that the expression of these isoforms is closely related to the toxicity of alachlor.

  6. The trans-10,cis-12 isomer of conjugated linoleic acid reduces hepatic triacylglycerol content without affecting lipogenic enzymes in hamsters.

    Science.gov (United States)

    Zabala, Amaia; Churruca, Itziar; Macarulla, M Teresa; Rodríguez, Víctor M; Fernández-Quintela, Alfredo; Martínez, J Alfredo; Portillo, María P

    2004-09-01

    Conjugated linoleic acid (CLA) refers to the positional and geometric dienoic isomers of linoleic acid. The dietary intake of CLA has been associated with changes in lipid metabolism. The aim of the present work was to assess the effects of the two main isomers of CLA on sterol regulatory element binding protein (SREBP)-1a and SREBP-1c mRNA levels, as well as on mRNA levels and the activities of several lipogenic enzymes in liver. For this purpose hamsters were fed an atherogenic diet supplemented with 5 g linoleic acid, cis-9,trans-11 or trans-10,cis-12 CLA/kg diet for 6 weeks. The trans-10,cis-12 isomer intake produced significantly greater liver weight, but also significantly decreased liver fat accumulation. No changes in mRNA levels of SREBP-1a, SREBP-1c and lipogenic enzymes, or in the activities of these enzymes, were observed. There was no effect of feeding cis-9,trans-11 CLA. These results suggest that increased fat accumulation in liver does not occur on the basis of liver enlargement produced by feeding the trans-10,cis-12 isomer of CLA in hamsters. The reduction in hepatic triacylglycerol content induced by this isomer was not attributable to changes in lipogenesis.

  7. The Effects of Subacute Exposure of Peracetic Acid on Lipid Peroxidation and Hepatic Enzymes in Wistar Rats

    Directory of Open Access Journals (Sweden)

    Abdoljalal Marjani

    2010-10-01

    Full Text Available Objectives: This study was undertaken to determine the effect of subacute exposure of peracetic acid on lipid peroxidation and hepatic enzymes in Wistar rats.Methods: 48 male animals in Treatment Group I, II and III received 0.2%, 2% and 20% peracetic acid daily for 2 and 4 weeks.Results: Serum malondialdehyde increased and Alanine Transaminase and Aspartate Transaminase decreased significantly in groups 2 and 3, compared to the control group. The malondialdehyde, Alanine Transaminase and Aspartate Transaminase with 0.2% and 2% doses of peracetic acid for 2 weeks do not lead to the alteration of malondialdehyde and enzyme activities.Conclusion: This study demonstrated that the enhancement of malondialdehyde could provide an oxidative damage induced by disinfectant peroxidation at 20% and 2% doses at 2 and 4 weeks. The consumption of peroxidation with 20% for 2 weeks and 2% for 4 weeks can cause the increase of malondialdehyde and the decrease of enzyme activities, respectively.

  8. The Action of Antidiabetic Plants of the Canadian James Bay Cree Traditional Pharmacopeia on Key Enzymes of Hepatic Glucose Homeostasis

    Directory of Open Access Journals (Sweden)

    Abir Nachar

    2013-01-01

    Full Text Available We determined the capacity of putative antidiabetic plants used by the Eastern James Bay Cree (Canada to modulate key enzymes of gluconeogenesis and glycogen synthesis and key regulating kinases. Glucose-6-phosphatase (G6Pase and glycogen synthase (GS activities were assessed in cultured hepatocytes treated with crude extracts of seventeen plant species. Phosphorylation of AMP-dependent protein kinase (AMPK, Akt, and Glycogen synthase kinase-3 (GSK-3 were probed by Western blot. Seven of the seventeen plant extracts significantly decreased G6Pase activity, Abies balsamea and Picea glauca, exerting an effect similar to insulin. This action involved both Akt and AMPK phosphorylation. On the other hand, several plant extracts activated GS, Larix laricina and A. balsamea, far exceeding the action of insulin. We also found a significant correlation between GS stimulation and GSK-3 phosphorylation induced by plant extract treatments. In summary, three Cree plants stand out for marked effects on hepatic glucose homeostasis. P. glauca affects glucose production whereas L. laricina rather acts on glucose storage. However, A. balsamea has the most promising profile, simultaneously and powerfully reducing G6Pase and stimulating GS. Our studies thus confirm that the reduction of hepatic glucose production likely contributes to the therapeutic potential of several antidiabetic Cree traditional medicines.

  9. Epigenetic Changes during Hepatic Stellate Cell Activation.

    Directory of Open Access Journals (Sweden)

    Silke Götze

    Full Text Available Hepatic stellate cells (HSC, which can participate in liver regeneration and fibrogenesis, have recently been identified as liver-resident mesenchymal stem cells. During their activation HSC adopt a myofibroblast-like phenotype accompanied by profound changes in the gene expression profile. DNA methylation changes at single genes have been reported during HSC activation and may participate in the regulation of this process, but comprehensive DNA methylation analyses are still missing. The aim of the present study was to elucidate the role of DNA methylation during in vitro activation of HSC.The analysis of DNA methylation changes by antibody-based assays revealed a strong decrease in the global DNA methylation level during culture-induced activation of HSC. To identify genes which may be regulated by DNA methylation, we performed a genome-wide Methyl-MiniSeq EpiQuest sequencing comparing quiescent and early culture-activated HSC. Approximately 400 differentially methylated regions with a methylation change of at least 20% were identified, showing either hypo- or hypermethylation during activation. Further analysis of selected genes for DNA methylation and expression were performed revealing a good correlation between DNA methylation changes and gene expression. Furthermore, global DNA demethylation during HSC activation was investigated by 5-bromo-2-deoxyuridine assay and L-mimosine treatment showing that demethylation was independent of DNA synthesis and thereby excluding a passive DNA demethylation mechanism.In summary, in vitro activation of HSC initiated strong DNA methylation changes, which were associated with gene regulation. These results indicate that epigenetic mechanisms are important for the control of early HSC activation. Furthermore, the data show that global DNA demethylation during activation is based on an active DNA demethylation mechanism.

  10. Serum enzymes activities in Plasmodium falciparum infection in Southern Pakistan

    Directory of Open Access Journals (Sweden)

    Koay Yen Chin

    2011-05-01

    Full Text Available Objective: Serum levels of lactate dehydrogenase (LDH,aspartate aminotranferase (AST, alanine aminotransferase(ALT, and alkaline phosphatase (ALP were assessed todetermine the liver functions of patients infected withPlasmodium falciparum. The enzyme activities were assessedin 60 malarial patients and a control group of 44 people.Materials and Methods: The data for the study was collectedfrom the survey conducted from Liaquat University of medicaland health sciences Hospital, Hyderabad, Pakaistan. Sample of60 patients aged between 20 and 50 years were collected. Acontrol group of 44 healthy individual adults was also assessedfor comparative purposes. All the malaria patients who visitedthe OPD during the study period enrolled in the study.Results: The LDH activity in male patients was found to be674.89 ± 33.354 IU/L. This is above the control LDH activity of296.59 ± 14.476 IU/L. Similarly, in female patients, the serumLDH activity of 580.25 ± 24.507 IU/L is over twice the controlfemale serum LDH activity of 302.18 ± 18.082 IU/L. Furtherone-way anova test was performed to find any significance ininfected and control male and female.Conclusion: Hepatic dysfunction was found to be associated toP. falciparum malaria infection.

  11. Effect of Pterocarpus santalinus bark, on blood glucose, serum lipids, plasma insulin and hepatic carbohydrate metabolic enzymes in streptozotocin-induced diabetic rats.

    Science.gov (United States)

    Kondeti, Vinay Kumar; Badri, Kameswara Rao; Maddirala, Dilip Rajasekhar; Thur, Sampath Kumar Mekala; Fatima, Shaik Sameena; Kasetti, Ramesh Babu; Rao, Chippada Appa

    2010-05-01

    The present study was designed to investigate the effect of bark of Pterocarpus santalinus, an ethnomedicinal plant, on blood glucose, plasma insulin, serum lipids and the activities of hepatic glucose metabolizing enzymes in streptozotocin-induced diabetic rats. Streptozotocin-induced diabetic rats were treated (acute/short-term and long-term) with ethyl acetate:methanol fractions of ethanolic extract of the bark of P. santalinus. Fasting blood glucose, HbA(1C), plasma insulin and protein were estimated before and after the treatment, along with hepatic glycogen, and activities of hexokinase, glucose-6-phosphatase, fructose-1,6-bisphosphatase and glucose-6-phosphate dehydrogenase. Further anti-hyperlipidemic activity was studied by measuring the levels of serum lipids and lipoproteins. Phytochemical analysis of active fraction showed the presence of flavonoids, glycosides and phenols. Biological testing of the active fraction demonstrated a significant antidiabetic activity by reducing the elevated blood glucose levels and glycosylated hemoglobin, improving hyperlipidemia and restoring the insulin levels in treated experimental induced diabetic rats. Further elucidation of mechanism of action showed improvement in the hepatic carbohydrate metabolizing enzymes after the treatment. Our present investigation suggests that active fraction of ethanolic extract of bark of P. santalinus decreases streptozotocin induced hyperglycemia by increasing glycolysis and decreasing gluconeogenesis. Copyright (c) 2010 Elsevier Ltd. All rights reserved.

  12. Epigenetics of dominance for enzyme activity

    Indian Academy of Sciences (India)

    Kuldip S Trehan; Kulbir S Gill

    2002-03-01

    We have isolated and purified two parental homodimers and a unique heterodimer of acid phosphatase [coded by Acph-11.05() and Acph-10.95()] from isogenic homozygotes and heterozygotes of Drosophila malerkotliana. and produce qualitatively different allozymes and the two alleles are expressed equally within and across all three genotypes and and play an equal role in the epigenetics of dominance. Subunit interaction in the heterodimer over a wide range of H+ concentrations accounts for the epigenetics of dominance for enzyme activity.

  13. Enzyme activity assay of glycoprotein enzymes based on a boronate affinity molecularly imprinted 96-well microplate.

    Science.gov (United States)

    Bi, Xiaodong; Liu, Zhen

    2014-12-16

    Enzyme activity assay is an important method in clinical diagnostics. However, conventional enzyme activity assay suffers from apparent interference from the sample matrix. Herein, we present a new format of enzyme activity assay that can effectively eliminate the effects of the sample matrix. The key is a 96-well microplate modified with molecularly imprinted polymer (MIP) prepared according to a newly proposed method called boronate affinity-based oriented surface imprinting. Alkaline phosphatase (ALP), a glycoprotein enzyme that has been routinely used as an indicator for several diseases in clinical tests, was taken as a representative target enzyme. The prepared MIP exhibited strong affinity toward the template enzyme (with a dissociation constant of 10(-10) M) as well as superb tolerance for interference. Thus, the enzyme molecules in a complicated sample matrix could be specifically captured and cleaned up for enzyme activity assay, which eliminated the interference from the sample matrix. On the other hand, because the boronate affinity MIP could well retain the enzymatic activity of glycoprotein enzymes, the enzyme captured by the MIP was directly used for activity assay. Thus, additional assay time and possible enzyme or activity loss due to an enzyme release step required by other methods were avoided. Assay of ALP in human serum was successfully demonstrated, suggesting a promising prospect of the proposed method in real-world applications.

  14. Lycopene attenuated hepatic tumorigenesis via differential mechanisms depending on carotenoid cleavage enzyme in mice.

    Science.gov (United States)

    Ip, Blanche C; Liu, Chun; Ausman, Lynne M; von Lintig, Johannes; Wang, Xiang-Dong

    2014-12-01

    Obesity is associated with increased liver cancer risks and mortality. We recently showed that apo-10'-lycopenoic acid, a lycopene metabolite generated by beta-carotene-9',10'-oxygenase (BCO2), inhibited carcinogen-initiated, high-fat diet (HFD)-promoted liver inflammation, and hepatic tumorigenesis development. The present investigation examined the outstanding question of whether lycopene could suppress HFD-promoted hepatocellular carcinoma (HCC) progression, and if BCO2 expression is important using BCO2-knockout (BCO2-KO) and wild-type male mice. Results showed that lycopene supplementation (100 mg/kg diet) for 24 weeks resulted in comparable accumulation of hepatic lycopene (19.4 vs. 18.2 nmol/g) and had similar effects on suppressing HFD-promoted HCC incidence (19% vs. 20%) and multiplicity (58% vs. 62%) in wild-type and BCO2-KO mice, respectively. Intriguingly, lycopene chemopreventive effects in wild-type mice were associated with reduced hepatic proinflammatory signaling (phosphorylation of NK-κB p65 and STAT3; IL6 protein) and inflammatory foci. In contrast, the protective effects of lycopene in BCO2-KO but not in wild-type mice were associated with reduced hepatic endoplasmic reticulum stress-mediated unfolded protein response (ER(UPR)), through decreasing ER(UPR)-mediated protein kinase RNA-activated like kinase-eukaryotic initiation factor 2α activation, and inositol requiring 1α-X-box-binding protein 1 signaling. Lycopene supplementation in BCO2-KO mice suppressed oncogenic signals, including Met mRNA, β-catenin protein, and mTOR complex 1 activation, which was associated with increased hepatic microRNA (miR)-199a/b and miR214 levels. These results provided novel experimental evidence that dietary lycopene can prevent HFD-promoted HCC incidence and multiplicity in mice, and may elicit different mechanisms depending on BCO2 expression. ©2014 American Association for Cancer Research.

  15. High-Throughput Analysis of Enzyme Activities

    Energy Technology Data Exchange (ETDEWEB)

    Lu, Guoxin [Iowa State Univ., Ames, IA (United States)

    2007-01-01

    High-throughput screening (HTS) techniques have been applied to many research fields nowadays. Robot microarray printing technique and automation microtiter handling technique allows HTS performing in both heterogeneous and homogeneous formats, with minimal sample required for each assay element. In this dissertation, new HTS techniques for enzyme activity analysis were developed. First, patterns of immobilized enzyme on nylon screen were detected by multiplexed capillary system. The imaging resolution is limited by the outer diameter of the capillaries. In order to get finer images, capillaries with smaller outer diameters can be used to form the imaging probe. Application of capillary electrophoresis allows separation of the product from the substrate in the reaction mixture, so that the product doesn't have to have different optical properties with the substrate. UV absorption detection allows almost universal detection for organic molecules. Thus, no modifications of either the substrate or the product molecules are necessary. This technique has the potential to be used in screening of local distribution variations of specific bio-molecules in a tissue or in screening of multiple immobilized catalysts. Another high-throughput screening technique is developed by directly monitoring the light intensity of the immobilized-catalyst surface using a scientific charge-coupled device (CCD). Briefly, the surface of enzyme microarray is focused onto a scientific CCD using an objective lens. By carefully choosing the detection wavelength, generation of product on an enzyme spot can be seen by the CCD. Analyzing the light intensity change over time on an enzyme spot can give information of reaction rate. The same microarray can be used for many times. Thus, high-throughput kinetic studies of hundreds of catalytic reactions are made possible. At last, we studied the fluorescence emission spectra of ADP and obtained the detection limits for ADP under three different

  16. Hepatic expression of detoxification enzymes is decreased in human obstructive cholestasis due to gallstone biliary obstruction.

    Directory of Open Access Journals (Sweden)

    Jin Chai

    Full Text Available Levels of bile acid metabolic enzymes and membrane transporters have been reported to change in cholestasis. These alterations (e.g. CYP7A1 repression and MRP4 induction are thought to be adaptive responses that attenuate cholestatic liver injury. However, the molecular mechanisms of these adaptive responses in human obstructive cholestasis due to gallstone biliary obstruction remain unclear.We collected liver samples from cholestatic patients with biliary obstruction due to gallstones and from control patients without liver disease (n = 22 per group. The expression levels of bile acid synthetic and detoxification enzymes, membrane transporters, and the related nuclear receptors and transcriptional factors were measured.The levels of bile acid synthetic enzymes, CYP7B1 and CYP8B1, and the detoxification enzyme CYP2B6 were increased in cholestatic livers by 2.4-fold, 2.8-fold, and 1.9-fold, respectively (p<0.05. Conversely, the expression levels of liver detoxification enzymes, UGT2B4/7, SULT2A1, GSTA1-4, and GSTM1-4, were reduced by approximately 50% (p<0.05 in human obstructive cholestasis. The levels of membrane transporters, OSTβ and OCT1, were increased 10.4-fold and 1.8-fold, respectively, (p<0.05, whereas those of OSTα, ABCG2 and ABCG8 were all decreased by approximately 40%, (p<0.05 in human cholestatic livers. Hepatic nuclear receptors, VDR, HNF4α, RXRα and RARα, were induced (approximately 2.0-fold, (p<0.05 whereas FXR levels were markedly reduced to 44% of control, (p<0.05 in human obstructive cholestasis. There was a significantly positive correlation between the reduction in FXR mRNA and UGT2B4/7, SULT2A1, GSTA1, ABCG2/8 mRNA levels in livers of obstructive cholestatic patients (p<0.05.The levels of hepatic detoxification enzymes were significantly decreased in human obstructive cholestasis, and these decreases were positively associated with a marked reduction of FXR levels. These findings are consistent with impaired

  17. Modulation of insulin degrading enzyme activity and liver cell proliferation.

    Science.gov (United States)

    Pivovarova, Olga; von Loeffelholz, Christian; Ilkavets, Iryna; Sticht, Carsten; Zhuk, Sergei; Murahovschi, Veronica; Lukowski, Sonja; Döcke, Stephanie; Kriebel, Jennifer; de las Heras Gala, Tonia; Malashicheva, Anna; Kostareva, Anna; Lock, Johan F; Stockmann, Martin; Grallert, Harald; Gretz, Norbert; Dooley, Steven; Pfeiffer, Andreas F H; Rudovich, Natalia

    2015-01-01

    Diabetes mellitus type 2 (T2DM), insulin therapy, and hyperinsulinemia are independent risk factors of liver cancer. Recently, the use of a novel inhibitor of insulin degrading enzyme (IDE) was proposed as a new therapeutic strategy in T2DM. However, IDE inhibition might stimulate liver cell proliferation via increased intracellular insulin concentration. The aim of this study was to characterize effects of inhibition of IDE activity in HepG2 hepatoma cells and to analyze liver specific expression of IDE in subjects with T2DM. HepG2 cells were treated with 10 nM insulin for 24 h with or without inhibition of IDE activity using IDE RNAi, and cell transcriptome and proliferation rate were analyzed. Human liver samples (n = 22) were used for the gene expression profiling by microarrays. In HepG2 cells, IDE knockdown changed expression of genes involved in cell cycle and apoptosis pathways. Proliferation rate was lower in IDE knockdown cells than in controls. Microarray analysis revealed the decrease of hepatic IDE expression in subjects with T2DM accompanied by the downregulation of the p53-dependent genes FAS and CCNG2, but not by the upregulation of proliferation markers MKI67, MCM2 and PCNA. Similar results were found in the liver microarray dataset from GEO Profiles database. In conclusion, IDE expression is decreased in liver of subjects with T2DM which is accompanied by the dysregulation of p53 pathway. Prolonged use of IDE inhibitors for T2DM treatment should be carefully tested in animal studies regarding its potential effect on hepatic tumorigenesis.

  18. Lymphocyte activation and hepatic cellular infiltration in immunocompetent mice infected by dengue virus.

    Science.gov (United States)

    Chen, Hsuen-Chin; Lai, Show-Yun; Sung, Jui-Min; Lee, Shu-Hwae; Lin, Yu-Chin; Wang, Wei-Kung; Chen, Yee-Chun; Kao, Chuan-Liang; King, Chwan-Chuen; Wu-Hsieh, Betty A

    2004-07-01

    Activation and expansion of dengue virus-specific T cells and abnormal liver functions in dengue patients have been documented. However, it remains to be determined whether T cells are involved in the pathogenic mechanism of dengue virus infection. In this study, immunocompetent C57BL/6 mice were employed to study dengue virus-induced T cell activation. Mice were inoculated with 10(8) PFU dengue virus serotype 2 strain 16681 by the intravenous route. Dengue viral core RNA was detected by RT-PCR in mouse serum, liver, spleen, and brain at different time points after infection. Splenic T cells were activated as evidenced by their expression of CD69 and O-glycosylated CD43 at as early as day 3 after infection. Splenic T cell expression of O-glycosylated CD43 and IFN-gamma production coordinately peaked at day 5. Coincided with the peak of splenic T cell activation was hepatic lymphocyte infiltration and elevation of liver enzymes. Flow cytometric analysis revealed the infiltrating CD8(+) T cell to CD4(+) T cell ratio was 5/3. After a second inoculation of dengue virus, hepatic T cell infiltration and liver enzyme levels increased sharply. The infiltrating hepatic CD8(+) T cell to CD4(+) T cell ratio increased to 5.8/1. A strong correlation was found between T cell activation and hepatic cellular infiltration in immunocompetent mice infected with dengue virus. The kinetics of liver enzyme elevation also correlated with that of T cell activation. These data suggest a relationship between T cell infiltration and elevation of liver enzymes.

  19. Exploration of the spontaneous fluctuating activity of single enzyme molecules

    NARCIS (Netherlands)

    Schwabe, A.; Maarleveld, T.R.; Bruggeman, F.J.

    2013-01-01

    Single enzyme molecules display inevitable, stochastic fluctuations in their catalytic activity. In metabolism, for instance, the stochastic activity of individual enzymes is averaged out due to their high copy numbers per single cell. However, many processes inside cells rely on single enzyme activ

  20. Efeitos da farinha de folhas de mandioca sobre a atividade das enzimas AST, ALT, FA e lipídios hepáticos de ratos Wistar Effects of cassava leaves flour on the AST, ALT, ALP enzymes activity and hepatic lipids of Wistar rats

    Directory of Open Access Journals (Sweden)

    Daniela Séfora de Melo

    2008-12-01

    Aspartate Aminotransferase (AST and Alkaline Phosphatase (ALP enzymes activity, but they significantly increased the Alanine Aminotransferase (ALT enzyme activity. The histopathologic study showed hepatocytes cytoplasm vacuolization for all groups. However, the number of animals with sharp vacuolization was higher in the groups that received diets with CLF , which also showed higher levels of hepatic lipids, total cholesterol, and greater ratio liver weight / body weight. These results indicate that the antinutrients in the cassava leaves, such as tannins, cyanide, and saponins are probably responsible for the reduction in the hepatic function of animals fed with CLF.

  1. Coleus forskohlii extract induces hepatic cytochrome P450 enzymes in mice.

    Science.gov (United States)

    Virgona, Nantiga; Yokotani, Kaori; Yamazaki, Yuko; Shimura, Fumio; Chiba, Tsuyoshi; Taki, Yuko; Yamada, Shizuo; Shinozuka, Kazumasa; Murata, Masatsune; Umegaki, Keizo

    2012-03-01

    Coleus forskohlii root extract (CFE) is popular for use as a weight loss dietary supplement. In this study, the influence of standardized CFE containing 10% active component forskolin on the hepatic drug metabolizing system was investigated to evaluate the safety through its drug interaction potential. Male ICR mice were fed AIN93G-based diets containing 0-5% CFE or 0.05% pure forskolin for 2-3 weeks. Intake of two different sources of 0.5% CFE significantly increased the relative liver weight, total content of hepatic cytochrome P450 (CYP) and induced CYPs (especially 2B, 2C, 3A types) and glutathione S-transferase (GST) activities. CFE significantly increased mRNA expression of CYPs and GST with dose related responses. However, unlike the CFE, intake of 0.05% pure forskolin was found to be associated with only weak induction in CYP3A and GST activities with no significant increases in relative liver weight, total hepatic content or other CYPs activities. The inductions of CYPs and GST by CFE were observed at 1 week of feeding and rapidly recovered by discontinuation of CFE. These results indicated the induction potential of CFE on CYPs, and that this effect was predominantly due to other, as yet unidentified constituents, and not forskolin contained in CFE. Copyright © 2011 Elsevier Ltd. All rights reserved.

  2. Evaluation of an enzyme immunoassay for hepatitis C virus antibody detection using a recombinant protein derived from the core region of hepatitis C virus genome

    Directory of Open Access Journals (Sweden)

    Lopes EPA

    2000-01-01

    Full Text Available This study was undertaken to evaluate an enzyme immunoassay (EIA for hepatitis C virus antibody detection (anti-HCV, using just one antigen. Anti-HCV EIA was designed to detect anti-HCV IgG using on the solid-phase a recombinant C22 antigen localized at the N-terminal end of the core region of HCV genome, produced by BioMérieux. The serum samples diluted in phosphate buffer saline were added to wells coated with the C22, and incubated. After washings, the wells were loaded with conjugated anti-IgG, and read in a microtiter plate reader (492 nm. Serum samples of 145 patients were divided in two groups: a control group of 39 patients with non-C hepatitis (10 acute hepatitis A, 10 acute hepatitis B, 9 chronic hepatitis B, and 10 autoimmune hepatitis and a study group consisting of 106 patients with chronic HCV hepatitis. In the study group all patients had anti-HCV detected by a commercially available EIA (Abbott®, specific for HCV structural and nonstructural polypeptides, alanine aminotransferase elevation or positive serum HCV-RNA detected by nested-PCR. They also had a liver biopsy compatible with chronic hepatitis. The test was positive in 101 of the 106 (95% sera from patients in the study group and negative in 38 of the 39 (97% sera from those in the control group, showing an accuracy of 96%. According to these results, our EIA could be used to detect anti-HCV in the serum of patients infected with hepatitis C virus.

  3. Synthetic Polymer with a Structure-Driven Hepatic Deposition and Curative Pharmacological Activity in Hepatic Cells

    DEFF Research Database (Denmark)

    Riber, Camilla Frich; Halling Folkmar Andersen, Anna; Anegaard Rolskov, Lærke

    2017-01-01

    Synthetic polymers make strong contributions as tools for delivery of biological drugs and chemotherapeutics. The most praised characteristic of polymers in these applications is complete lack of pharmacological function such as to minimize the side effects within the human body. In contrast......, and pharmacokinetic properties not observed in close structural analogues. Specifically, PEAA reveals capacity to bind to albumin with ensuing natural hepatic deposition in vivo and exhibits concurrent inhibitory activity against the hepatitis C virus and inflammation in hepatic cells. Our findings provide a view...... on synthetic polymers as curative, functional agents and present PEAA as a unique biomedical tool with applications related to health of the human liver....

  4. The Impact of Enzyme Orientation and Electrode Topology on the Catalytic Activity of Adsorbed Redox Enzymes

    Science.gov (United States)

    McMillan, Duncan G. G.; Marritt, Sophie J.; Kemp, Gemma L.; Gordon-Brown, Piers; Butt, Julea N.; Jeuken, Lars J. C.

    2014-01-01

    It is well established that the structural details of electrodes and their interaction with adsorbed enzyme influences the interfacial electron transfer rate. However, for nanostructured electrodes, it is likely that the structure also impacts on substrate flux near the adsorbed enzymes and thus catalytic activity. Furthermore, for enzymes converting macro-molecular substrates it is possible that the enzyme orientation determines the nature of interactions between the adsorbed enzyme and substrate and therefore catalytic rates. In essence the electrode may impede substrate access to the active site of the enzyme. We have tested these possibilities through studies of the catalytic performance of two enzymes adsorbed on topologically distinct electrode materials. Escherichia coli NrfA, a nitrite reductase, was adsorbed on mesoporous, nanocrystalline SnO2 electrodes. CymA from Shewanella oneidensis MR-1 reduces menaquinone-7 within 200 nm sized liposomes and this reaction was studied with the enzyme adsorbed on SAM modified ultra-flat gold electrodes. PMID:24634538

  5. The Impact of Enzyme Orientation and Electrode Topology on the Catalytic Activity of Adsorbed Redox Enzymes.

    Science.gov (United States)

    McMillan, Duncan G G; Marritt, Sophie J; Kemp, Gemma L; Gordon-Brown, Piers; Butt, Julea N; Jeuken, Lars J C

    2013-11-01

    It is well established that the structural details of electrodes and their interaction with adsorbed enzyme influences the interfacial electron transfer rate. However, for nanostructured electrodes, it is likely that the structure also impacts on substrate flux near the adsorbed enzymes and thus catalytic activity. Furthermore, for enzymes converting macro-molecular substrates it is possible that the enzyme orientation determines the nature of interactions between the adsorbed enzyme and substrate and therefore catalytic rates. In essence the electrode may impede substrate access to the active site of the enzyme. We have tested these possibilities through studies of the catalytic performance of two enzymes adsorbed on topologically distinct electrode materials. Escherichia coli NrfA, a nitrite reductase, was adsorbed on mesoporous, nanocrystalline SnO2 electrodes. CymA from Shewanella oneidensis MR-1 reduces menaquinone-7 within 200 nm sized liposomes and this reaction was studied with the enzyme adsorbed on SAM modified ultra-flat gold electrodes.

  6. [Detection of enzyme activity in decontaminated spices in industrial use].

    Science.gov (United States)

    Müller, R; Theobald, R

    1995-03-01

    A range of decontaminated species of industrial use have been examined for their enzymes (catalase, peroxidase, amylase, lipase activity). The genuine enzymes remain fully active in irradiated spices, whereas the microbial load is clearly reduced. In contrast steam treated spices no longer demonstrate enzyme activities. Steam treatment offers e.g. black pepper without lipase activity, which can no longer cause fat deterioration. Low microbial load in combination with clearly detectable enzyme activity in spices is an indication for irradiation, whereas, reduced microbial contamination combined with enzyme inactivation indicate steam treatment of raw material.

  7. Effects of Different Lipid and Glucose Levels on Growth Performance, Hepatic Glycolysis and Gluconeogenic Key Enzyme Activities of Large Yellow Croaker (Larmichthys crocea Richardson)%不同脂肪和葡萄糖水平对大黄鱼生长性能、肝脏糖酵解和糖异生关键酶活性的影响

    Institute of Scientific and Technical Information of China (English)

    马红娜; 周飘苹; 陆游; 袁野; 侯迎梅; 孙蓬; 丁立云; 周歧存

    2016-01-01

    本试验旨在研究不同脂肪和葡萄糖水平对大黄鱼生长性能、肝脏糖酵解和糖异生关键酶活性、血清生化指标、糖原含量及消化酶活性等的影响。采用2×3双因素试验设计,其中脂肪设5%、10%2个水平,葡萄糖设10%、20%、30%3个水平,共配制6种试验饲料。每种饲料设3个重复,每个重复放养平均体重为(14.79±0.13) g的大黄鱼幼鱼50尾。试验期为8周。结果表明:饲料脂肪和葡萄糖水平的交互作用对大黄鱼增重率( WGR)、特定生长率( SGR)和饲料效率( FE)的影响不显著( P>0.05)。在饲料脂肪水平为5%时,WGR和SGR随饲料葡萄糖水平增加而降低,30%葡萄糖组的WGR和SGR显著低于10%葡萄糖组( P0.05)。饲料脂肪和葡萄糖水平的交互作用对大黄鱼肝糖原、肌糖原含量有显著影响( P0.05);然而,在饲料脂肪水平相同时,血清葡萄糖含量随饲料葡萄糖水平的升高而升高,30%葡萄糖组血清葡萄糖含量显著高于10%葡萄糖组( P0.05)。在饲料脂肪水平为10%时,随饲料葡萄糖水平的升高,肝脏GK、PFK活性升高,肝脏PEPCK活性先升高后降低。由结果可知,与饲料脂肪水平为10%时相比,在饲料脂肪水平为5%时,随饲料葡萄糖的水平的升高,大黄鱼能够通过调节糖代谢关键酶活性及肝糖原含量来维持血糖含量的平衡,有效利用饲料中的葡萄糖。综合本试验结果,建议大黄鱼幼鱼阶段饲料适宜的脂肪和糖水平分别为10%和20%。%An 8-week feeding trial was conducted to evaluate the effects of different lipid and glucose levels on growth performance, hepatic glycolysis and gluconeogenic key enzyme activities, serum biochemical indices, glycogen contents and digestive enzyme activities etc. of large yellow croaker. Six experimental diets were for-mulated to contain two lipid levels ( 5% and 10%) and three glucose levels ( 10%, 20% and 30%) using 2 × 3 double-factor experimental design. Each diet was

  8. Liquiritigenin, a flavonoid aglycone from licorice, has a choleretic effect and the ability to induce hepatic transporters and phase-II enzymes.

    Science.gov (United States)

    Kim, Young Woo; Kang, Hee Eun; Lee, Myung Gull; Hwang, Se Jin; Kim, Sang Chan; Lee, Chang Ho; Kim, Sang Geon

    2009-02-01

    Liquiritigenin (LQ), an active component of licorice, has an inhibitory effect on LPS-induced inhibitory nitric oxide synthase expression. This study investigated the effects of LQ on choleresis, the expression of hepatic transporters and phase-II enzymes, and fulminant hepatitis. The choleretic effect and the pharmacokinetics of LQ and its glucuronides were monitored in rats. After intravenous administration of LQ, the total area under the plasma concentration-time curve of glucuronyl metabolites was greater than that of LQ in plasma, which accompanied elevations in bile flow rate and biliary excretion of bile acid, glutathione, and bilirubin. The expressions of hepatocellular transporters and phase-II enzymes were assessed by immunoblots, real-time PCR, and immunohistochemistry. In the livers of rats treated with LQ, the protein and mRNA levels of multidrug resistance protein 2 and bile salt export pump were increased in the liver, which was verified by their increased localizations in canalicular membrane. In addition, LQ treatment enhanced the expression levels of major hepatic phase-II enzymes. Consistent with these results, LQ treatments attenuated galactosamine/LPS-induced hepatitis in rats, as supported by decreases in the plasma alanine aminotransferase, liver necrosis, and plasma TNF-alpha. These results demonstrate that LQ has a choleretic effect and the ability to induce transporters and phase-II enzymes in the liver, which may be associated with a hepatoprotective effect against galactosamine/LPS. Our findings may provide insight into understanding the action of LQ and its therapeutic use for liver disease.

  9. Changes in activities of tissues enzymes in rats administered Ficus ...

    African Journals Online (AJOL)

    enzymes in the serum, liver, kidney and heart of albino rats. ... significant alteration in enzyme activities in the serum and tissues as well as relative organ ... powder using an electrical blender. ... (average weight 160 g) at the Central Animal.

  10. [Chronic active hepatitis: clinical, biochemical, and histopathologic correlation].

    Science.gov (United States)

    Subauste, M C

    1989-01-01

    A retrospective study over 26 female patients with chronic active hepatitis was made. The mean age was 39 years old, the mean length of illness of 8 months; 5 patients had positive markers for hepatitis B. Patients were selected with the grade of histological activity: 8 patients had a mild form from disease (2A) and 16 with a severe one (2B). The predominant group was 2B. Severe inflammatory infiltration was the hallmark and multiobulillar necrosis, bridging, eosinophils and hiperplasia of kuppfer cells were found only in this group. Clinical features range from hepatic manifestations to systemic ones. Chronic active hepatitis may present with cholestasis, but the latter is not always related with the grade of activity. Group 2B had elevated aminotransferases and a low concentration for protrobine.

  11. [Hepatitis non-A, non-B: epidemiological significance in acute viral hepatitis and chronic active hepatitis of hepatological consultation].

    Science.gov (United States)

    Jmelnitzky, A C; Basualdo, J A; Belloni, P O; Ponce de León, H H; García, C; Curciarello, J

    1987-01-01

    157 acute viral hepatitis and 60 chronic active ones have been analyzed focusing on NANB etiology. HAV was implicated in 36.3% of the hole acute viral hepatitis sample, HBV in 29.3%, and HNANBV was presumed as etiology in 31.2%, 5 patients (3.2%) had acute infection by HAV, on previous one by HBV, except for Epstein-Barr virus, no other test for viruses were determined (CMV, HSV, etc.). Male/female ratio was 1.4:1, 1.9:1, and 1.4:1 for HAV, HBV and HNANBV acute hepatitis respectively; HAV was the main etiology in the 0-9 age group (72.2%) although it only represents 11.5% of the sample; small occurrence of HAV hepatitis were found in patients over 40 (8.8%); HBV was clearly prevalent in patients over 50 (65.2%); the highest concentration of NANB etiology was found between 20-39 years old, but it was represented in all age-groups. Out of 49 NANB acute hepatitis, 12.2% had related transfusional antecedents, 12.2% belonged to health care worker group, and 4.1% had a close family NANB hepatitis contact; 71.5% had no reported antecedent. Viral source was presumably implicated in 75.0% of chronic active hepatitis, 25.0% attributable to HNANBV. Results seem not feasible to transfer to general population due to the facts that most patients were of specialized consult, and pediatric assistance is unusual to the authors practice.

  12. Novel matrine derivative MD-1 attenuates hepatic fibrosis by inhibiting EGFR activation of hepatic stellate cells.

    Science.gov (United States)

    Feng, Yi; Ying, Hai-Yan; Qu, Ying; Cai, Xiao-Bo; Xu, Ming-Yi; Lu, Lun-Gen

    2016-09-01

    Matrine (MT), the effective component of Sophora flavescens Ait, has been shown to have anti-inflammation, immune-suppressive, anti-tumor, and anti-hepatic fibrosis activities. However, the pharmacological effects of MT still need to be strengthened due to its relatively low efficacy and short half-life. In the present study, we report a more effective thio derivative of MT, MD-1, and its inhibitory effects on the activation of hepatic stellate cells (HSCs) in both cell culture and animal models. Cytological experiments showed that MD-1 can inhibit the proliferation of HSC-T6 cells with a half-maximal inhibitory concentration (IC50) of 62 μmol/L. In addition, MD-1 more strongly inhibits the migration of HSC-T6 cells compared to MT and can more effectively induce G0/G1 arrest and apoptosis. Investigating the biological mechanisms underlying anti-hepatic fibrosis in the presence of MD-1, we found that MD-1 can bind the epidermal growth factor receptor (EGFR) on the surface of HSC-T6 cells, which can further inhibit the phosphorylation of EGFR and its downstream protein kinase B (Akt), resulting in decreased expression of cyclin D1 and eventual inhibition of the activation of HSC-T6 cells. Furthermore, in rats with dimethylnitrosamine (DMN)-induced hepatic fibrosis, MD-1 slowed the development and progression of hepatic fibrosis, protecting hepatic parenchymal cells and improving hepatic functions. Therefore, MD-1 is a potential drug for anti-hepatic fibrosis.

  13. Glycyl radical activating enzymes: structure, mechanism, and substrate interactions.

    Science.gov (United States)

    Shisler, Krista A; Broderick, Joan B

    2014-03-15

    The glycyl radical enzyme activating enzymes (GRE-AEs) are a group of enzymes that belong to the radical S-adenosylmethionine (SAM) superfamily and utilize a [4Fe-4S] cluster and SAM to catalyze H-atom abstraction from their substrate proteins. GRE-AEs activate homodimeric proteins known as glycyl radical enzymes (GREs) through the production of a glycyl radical. After activation, these GREs catalyze diverse reactions through the production of their own substrate radicals. The GRE-AE pyruvate formate lyase activating enzyme (PFL-AE) is extensively characterized and has provided insights into the active site structure of radical SAM enzymes including GRE-AEs, illustrating the nature of the interactions with their corresponding substrate GREs and external electron donors. This review will highlight research on PFL-AE and will also discuss a few GREs and their respective activating enzymes. Copyright © 2014. Published by Elsevier Inc.

  14. Antioxidant activity of new aramide nanoparticles containing redox-active N-phthaloyl valine moieties in the hepatic cytochrome P450 system in male rats.

    Science.gov (United States)

    Hassan, Hammed H A M; El-Banna, Sabah G; Elhusseiny, Amel F; Mansour, El-Sayed M E

    2012-07-10

    We report the synthesis of aramide nanoparticles containing a chiral N-phthaloyl valine moiety and their antioxidant activities on hepatic contents of cytochrome P₄₅₀, amidopyrene N-demethylase, aniline-4-hyroxylase and induced the hepatic content of cytochrome b5 and nicotinamide adenine dinucleotide phosphate (NADPH) cytochrome C-reductase. Polymers were obtained as well-separated spherical nanoparticles while highly aggregated particles via H-bonding organization of the aramide-containing pyridine led to a thin layer formation. The effects of the nanoparticles and CCl₄ on enzyme activities and thiobarbituric acid reactive substances (TBARS) levels of male rat liver were studied. Pretreatments of rats with the polyamides prior to the administration of CCl₄ decreased the hepatic content of the tested enzymes. Doses reduced the toxic effects exerted by (•CCl₃) upon the liver through inhibition of the cytochrome P₄₅₀ system. Inhibition of such metabolizing enzymes could reduce the carcinogenic effects of chemical carcinogens.

  15. Induction of hepatic CYP3A enzymes by pregnancy-related hormones: studies in human hepatocytes and hepatic cell lines.

    Science.gov (United States)

    Papageorgiou, Ioannis; Grepper, Susan; Unadkat, Jashvant D

    2013-02-01

    CYP3A activity is induced by approximately 2-fold during the third trimester of human pregnancy. Placental growth hormone (PGH), estrogens (primarily 17β-estradiol), cortisol, and progesterone have the potential to modulate CYP3A activity. Therefore, we determined whether the elevated plasma concentrations of these hormones during pregnancy induce hepatic CYP3A expression. We incubated sandwich-cultured human hepatocytes (SCHH) from premenopausal female donors (n = 2) with the physiologic (unbound, 1× total) and the 10× total third trimester hormone plasma concentrations (individually and in combination) and determined their effect on CYP3A activity and the transcripts of CYP3A4, CYP3A5, and the respective hormone receptors (growth hormone receptor, glucocorticoid receptor, and estrogen receptor alpha). Of all the hormones, cortisol was the most potent inducer of CYP3A activity and CYP3A4, CYP3A5 mRNA expression. The combination of PGH/growth hormone and cortisol induced CYP3A activity and expression significantly more than did cortisol alone. When incubated with the unbound or total plasma concentration of all the hormones, CYP3A activity in SCHH was induced to an extent comparable to that observed in vivo during the third trimester. These hormones had only a modest effect on the mRNA expression of the hormone receptors. The pattern of induction observed in SCHH was reproduced in HepaRG cells but not in HuH7/HepG2 cells. SCHH or HepaRG cells could be used to determine the mechanistic basis of CYP3A induction during pregnancy and to predict the magnitude of induction likely to be observed during the first and second trimesters, when phenotyping studies to measure in vivo CYP3A activity are logistically difficult to perform.

  16. Histidine Augments the Suppression of Hepatic Glucose Production by Central Insulin Action

    OpenAIRE

    KIMURA, Kumi; Nakamura, Yusuke; Inaba, Yuka; Matsumoto, Michihiro; Kido, Yoshiaki; Asahara, Shun-ichiro; Matsuda, Tomokazu; Watanabe, Hiroshi; Maeda, Akifumi; Inagaki, Fuyuhiko; Mukai, Chisato; Takeda, Kiyoshi; Akira, Shizuo; Ota, Tsuguhito; Nakabayashi, Hajime

    2013-01-01

    Glucose intolerance in type 2 diabetes is related to enhanced hepatic glucose production (HGP) due to the increased expression of hepatic gluconeogenic enzymes. Previously, we revealed that hepatic STAT3 decreases the expression of hepatic gluconeogenic enzymes and suppresses HGP. Here, we show that increased plasma histidine results in hepatic STAT3 activation. Intravenous and intracerebroventricular (ICV) administration of histidine-activated hepatic STAT3 reduced G6Pase protein and mRNA le...

  17. Influence of Piper betle on hepatic marker enzymes and tissue antioxidant status in D-galactosamine-induced hepatotoxic rats.

    Science.gov (United States)

    Pushpavalli, Ganesan; Veeramani, Chinnadurai; Pugalendi, Kodukkur Viswanathan

    2008-01-01

    D-galactosamine is a well-established hepatotoxicant that induces a diffuse type of liver injury closely resembling human viral hepatitis. D-galactosamine by its property of generating free radicals causes severe damage to the membrane and affects almost all organs of the human body. The leaves of Piper betle L., a commonly used masticatory in Asian countries, possess several biological properties. Our aim is to investigate the in vivo antioxidant potential of P. betle leaf-extract against oxidative stress induced by D-galactosamine intoxication in male albino Wistar rats. Toxicity was induced by an intraperitoneal injection of D-galactosamine, 400 mg/kg body weight (BW) for 21 days. Rats were treated with P. betle extract (200 mg/kg BW) via intragastric intubations. We assessed the activities of liver marker enzymes (aspartate amino-transferase, alanine aminotransferase, alkaline phosphatase, gamma glutamyl transpeptidase) and levels of thiobarbituric acid reactive substances (TBARS), lipid hydroperoxides, superoxide dismutase, catalase, glutathione peroxidase, vitamin C, vitamin E, and reduced glutathione. The extract significantly improved the status of antioxidants and decreased TBARS, hydroperoxides, and liver marker enzymes when compared with the D-galactosamine treated group, demonstrating its hepatoprotective and antioxidant properties.

  18. Modulation of CYP1A1 and CYP1A2 hepatic enzymes after oral administration of Chios mastic gum to male Wistar rats.

    Science.gov (United States)

    Katsanou, Efrosini S; Kyriakopoulou, Katerina; Emmanouil, Christina; Fokialakis, Nikolas; Skaltsounis, Alexios-Leandros; Machera, Kyriaki

    2014-01-01

    Chios mastic gum (CMG), a resin derived from Pistacia lentiscus var. chia, is known since ancient times for its pharmacological activities. CYP1A1 and CYP1A2 enzymes are among the most involved in the biotransformation of chemicals and the metabolic activation of pro-carcinogens. Previous studies referring to the modulation of these enzymes by CMG have revealed findings of unclear biological and toxicological significance. For this purpose, the modulation of CYP1A1 and CYP1A2 enzymes in the liver of male Wistar rats following oral administration of CMG extract (CMGE), at the levels of mRNA and CYP1A1 enzyme activity, was compared to respective enzyme modulation following oral administration of a well-known bioactive natural product, caffeine, as control compound known to involve hepatic enzymes in its metabolism. mRNA levels of Cyp1a1 and Cyp1a2 were measured by reverse transcription real-time polymerase chain reaction and their relative quantification was calculated. CYP1A1 enzyme induction was measured through the activity of ethoxyresorufin-O-deethylase (EROD). The results indicated that administration of CMGE at the recommended pharmaceutical dose does not induce significant transcriptional modulation of Cyp1a1/2 and subsequent enzyme activity induction of CYP1A1 while effects of the same order of magnitude were observed in the same test system following the administration of caffeine at the mean daily consumed levels. The outcome of this study further confirms the lack of any toxicological or biological significance of the specific findings on liver following the administration of CMGE.

  19. Modulation of CYP1A1 and CYP1A2 hepatic enzymes after oral administration of Chios mastic gum to male Wistar rats.

    Directory of Open Access Journals (Sweden)

    Efrosini S Katsanou

    Full Text Available Chios mastic gum (CMG, a resin derived from Pistacia lentiscus var. chia, is known since ancient times for its pharmacological activities. CYP1A1 and CYP1A2 enzymes are among the most involved in the biotransformation of chemicals and the metabolic activation of pro-carcinogens. Previous studies referring to the modulation of these enzymes by CMG have revealed findings of unclear biological and toxicological significance. For this purpose, the modulation of CYP1A1 and CYP1A2 enzymes in the liver of male Wistar rats following oral administration of CMG extract (CMGE, at the levels of mRNA and CYP1A1 enzyme activity, was compared to respective enzyme modulation following oral administration of a well-known bioactive natural product, caffeine, as control compound known to involve hepatic enzymes in its metabolism. mRNA levels of Cyp1a1 and Cyp1a2 were measured by reverse transcription real-time polymerase chain reaction and their relative quantification was calculated. CYP1A1 enzyme induction was measured through the activity of ethoxyresorufin-O-deethylase (EROD. The results indicated that administration of CMGE at the recommended pharmaceutical dose does not induce significant transcriptional modulation of Cyp1a1/2 and subsequent enzyme activity induction of CYP1A1 while effects of the same order of magnitude were observed in the same test system following the administration of caffeine at the mean daily consumed levels. The outcome of this study further confirms the lack of any toxicological or biological significance of the specific findings on liver following the administration of CMGE.

  20. The hepatitis B virus ribonuclease H is sensitive to inhibitors of the human immunodeficiency virus ribonuclease H and integrase enzymes.

    Directory of Open Access Journals (Sweden)

    John E Tavis

    2013-01-01

    Full Text Available Nucleos(tide analog therapy blocks DNA synthesis by the hepatitis B virus (HBV reverse transcriptase and can control the infection, but treatment is life-long and has high costs and unpredictable long-term side effects. The profound suppression of HBV by the nucleos(tide analogs and their ability to cure some patients indicates that they can push HBV to the brink of extinction. Consequently, more patients could be cured by suppressing HBV replication further using a new drug in combination with the nucleos(tide analogs. The HBV ribonuclease H (RNAseH is a logical drug target because it is the second of only two viral enzymes that are essential for viral replication, but it has not been exploited, primarily because it is very difficult to produce active enzyme. To address this difficulty, we expressed HBV genotype D and H RNAseHs in E. coli and enriched the enzymes by nickel-affinity chromatography. HBV RNAseH activity in the enriched lysates was characterized in preparation for drug screening. Twenty-one candidate HBV RNAseH inhibitors were identified using chemical structure-activity analyses based on inhibitors of the HIV RNAseH and integrase. Twelve anti-RNAseH and anti-integrase compounds inhibited the HBV RNAseH at 10 µM, the best compounds had low micromolar IC(50 values against the RNAseH, and one compound inhibited HBV replication in tissue culture at 10 µM. Recombinant HBV genotype D RNAseH was more sensitive to inhibition than genotype H. This study demonstrates that recombinant HBV RNAseH suitable for low-throughput antiviral drug screening has been produced. The high percentage of compounds developed against the HIV RNAseH and integrase that were active against the HBV RNAseH indicates that the extensive drug design efforts against these HIV enzymes can guide anti-HBV RNAseH drug discovery. Finally, differential inhibition of HBV genotype D and H RNAseHs indicates that viral genetic variability will be a factor during drug

  1. Liver Allograft Its Use in Chronic Active Hepatitis With Macronodular Cirrhosis, Hepatitis B Surface Antigen

    Science.gov (United States)

    Corman, Jarques L.; Putnam, Charles W.; Iwatsuki, Shunzaburo; Redeker, Allan G.; Porter, K. A.; Peters, Robert L.; Schröter, Gerhard; Starzl, Thomas E.

    2010-01-01

    A patient suffering from chronic active hepatitis with macronodular cirrhosis, positive for hepatitis B surface antigen (HB,Ag), was treated with an orthoiopic liver allograft. The HB, antigenemia, as measured with several precipltation tests and by complement fixation, became negative after transplantation and remained so for about 2½ months. During the interval, very low Iters of the antigen were detectable by, radioimmunoassay. At about three months after transplantation, she had an attack of acute hepatitis, at which time HB,Ag became detectable by all tests. She recovered, but progressive liver disease developed during the remaining 1½ years of her life. She died of disseminaled nocardiosis and candidiasis with deteriorating hepatic function. The homograft at autopsy, showed no evidence of rejection, but was the site of chronic active liver disease, although of a different pathologic pattern than that affecting her native liver. The differences in histology may reflect the influence of chronic Immunosuppression on the features of chronic active hepatitis. PMID:365134

  2. Antioxidant activity of white rice, brown rice and germinated brown rice (in vivo and in vitro) and the effects on lipid peroxidation and liver enzymes in hyperlipidaemic rabbits.

    Science.gov (United States)

    Mohd Esa, Norhaizan; Abdul Kadir, Khairul-Kamilah; Amom, Zulkhairi; Azlan, Azrina

    2013-11-15

    Antioxidant activity of different rice extract and the effect on the levels of antioxidant enzyme activity, superoxide dismutase (SOD) and glutathione peroxidase (GPx), vitamin E, lipid peroxidation and liver enzymes in hyperlipidaemia rabbits were investigated. Germinated brown rice (GBR) has the highest antioxidant activity compared to white rice (WR) and brown rice (BR). All rice grains increased the activity of SOD and GPx. However, vitamin E levels increased only in the groups that received the BR and GBR diets. The reduction of lipid peroxidation levels and activity of hepatic enzymes (alanine transferase, ALT and aspartate transaminase, AST) were only significantly observed in the GBR group. In conclusion, GBR supplementation has the greatest impact on increasing antioxidant enzyme activity and vitamin E level and on reducing lipid peroxidation in hypercholesterolaemia rabbit, thereby preventing the formation of atherosclerotic plaques. Furthermore, GBR diet can also reduce the level of hepatic enzymes. Copyright © 2013 Elsevier Ltd. All rights reserved.

  3. Hepatic glycogen supercompensation activates AMP-activated protein kinase, impairs insulin signaling, and reduces glycogen deposition in the liver.

    Science.gov (United States)

    Winnick, Jason J; An, Zhibo; Ramnanan, Christopher J; Smith, Marta; Irimia, Jose M; Neal, Doss W; Moore, Mary Courtney; Roach, Peter J; Cherrington, Alan D

    2011-02-01

    The objective of this study was to determine how increasing the hepatic glycogen content would affect the liver's ability to take up and metabolize glucose. During the first 4 h of the study, liver glycogen deposition was stimulated by intraportal fructose infusion in the presence of hyperglycemic-normoinsulinemia. This was followed by a 2-h hyperglycemic-normoinsulinemic control period, during which the fructose infusion was stopped, and a 2-h experimental period in which net hepatic glucose uptake (NHGU) and disposition (glycogen, lactate, and CO(2)) were measured in the absence of fructose but in the presence of a hyperglycemic-hyperinsulinemic challenge including portal vein glucose infusion. Fructose infusion increased net hepatic glycogen synthesis (0.7 ± 0.5 vs. 6.4 ± 0.4 mg/kg/min; P vs. 100 ± 3 mg/g; P fructose infusion group) did not alter NHGU, but it reduced the percent of NHGU directed to glycogen (79 ± 4 vs. 55 ± 6; P vs. 29 ± 5; P = 0.01) and oxidation (9 ± 3 vs. 16 ± 3; P = NS). This change was associated with increased AMP-activated protein kinase phosphorylation, diminished insulin signaling, and a shift in glycogenic enzyme activity toward a state discouraging glycogen accumulation. These data indicate that increases in hepatic glycogen can generate a state of hepatic insulin resistance, which is characterized by impaired glycogen synthesis despite preserved NHGU.

  4. Detection of Extracellular Enzyme Activities in Ganoderma neo-japonicum

    OpenAIRE

    Jo, Woo-Sik; Park, Ha-Na; Cho, Doo-Hyun; Yoo, Young-Bok; Park, Seung-Chun

    2011-01-01

    The ability of Ganoderma to produce extracellular enzymes, including β-glucosidase, cellulase, avicelase, pectinase, xylanase, protease, amylase, and ligninase was tested in chromogenic media. β-glucosidase showed the highest activity, among the eight tested enzymes. In particular, Ganoderma neo-japonicum showed significantly stronger activity for β-glucosidase than that of the other enzymes. Two Ganoderma lucidum isolates showed moderate activity for avicelase; however, Ganoderma neo-japonic...

  5. Coffee modulates transcription factor Nrf2 and highly increases the activity of antioxidant enzymes in rats.

    Science.gov (United States)

    Vicente, Silvio J V; Ishimoto, Emília Y; Torres, Elizabeth A F S

    2014-01-08

    This study investigated the effect of a 28 day administration of coffee brew on the activity of antioxidant enzymes in rats. After this period of 2.0 mL/day dosages of this beverage, the activities of hepatic superoxide dismutase, catalase, and glutathione peroxidase increased 74.8, 59.4, and 135.2%, respectively, whereas the cytosolic level of Nrf2 increased 131.3%. At the same time, the total antioxidant capacity of the hepatic tissue increased 25.1%, improving the defensive status against oxidative stress. At the end of the experiment, the levels of biomarkers alanine transaminase and aspartate transaminase remained equal to the control group, and no changes were observed in the hepatic histoarchiteture of the animals, suggesting that the liver tissue was not impaired by the exposure to coffee. The changes in enzyme activities and antioxidant capacity were statistically significant (p < 0.05), indicating that coffee could be considered an important alternative against oxidative stress and its correlated degenerative diseases.

  6. Crystallographic B factor of critical residues at enzyme active site

    Institute of Scientific and Technical Information of China (English)

    张海龙; 宋时英; 林政炯

    1999-01-01

    Thirty-seven sets of crystallographic enzyme data were selected from Protein Data Bank (PDB, 1995). The average temperature factors (B) of the critical residues at the active site and the whole molecule of those enzymes were calculated respectively. The statistical results showed that the critical residues at the active site of most of the enzymes had lower B factors than did the whole molecules, indicating that in the crystalline state the critical residues at the active site of the natural enzymes possess more stable conformation than do the whole molecules. The flexibility of the active site during the unfolding by denaturing was also discussed.

  7. Apolipoprotein D Transgenic Mice Develop Hepatic Steatosis through Activation of PPARγ and Fatty Acid Uptake.

    Directory of Open Access Journals (Sweden)

    Marilyne Labrie

    Full Text Available Transgenic mice (Tg overexpressing human apolipoprotein D (H-apoD in the brain are resistant to neurodegeneration. Despite the use of a neuron-specific promoter to generate the Tg mice, they expressed significant levels of H-apoD in both plasma and liver and they slowly develop hepatic steatosis and insulin resistance. We show here that hepatic PPARγ expression in Tg mice is increased by 2-fold compared to wild type (WT mice. Consequently, PPARγ target genes Plin2 and Cide A/C are overexpressed, leading to increased lipid droplets formation. Expression of the fatty acid transporter CD36, another PPARgamma target, is also increased in Tg mice associated with elevated fatty acid uptake as measured in primary hepatocytes. Elevated expression of AMPK in the liver of Tg leads to phosphorylation of acetyl CoA carboxylase, indicating a decreased activity of the enzyme. Fatty acid synthase expression is also induced but the hepatic lipogenesis measured in vivo is not significantly different between WT and Tg mice. In addition, expression of carnitine palmitoyl transferase 1, the rate-limiting enzyme of beta-oxidation, is slightly upregulated. Finally, we show that overexpressing H-apoD in HepG2 cells in presence of arachidonic acid (AA, the main apoD ligand, increases the transcriptional activity of PPARγ. Supporting the role of apoD in AA transport, we observed enrichment in hepatic AA and a decrease in plasmatic AA concentration. Taken together, our results demonstrate that the hepatic steatosis observed in apoD Tg mice is a consequence of increased PPARγ transcriptional activity by AA leading to increased fatty acid uptake by the liver.

  8. Early hepatic stellate cell activation predicts severe hepatitis C recurrence after liver transplantation.

    Science.gov (United States)

    Gawrieh, Samer; Papouchado, Bettina G; Burgart, Lawrence J; Kobayashi, Shogo; Charlton, Michael R; Gores, Gregory J

    2005-10-01

    Only a subset of hepatitis C virus (HCV)-infected patients develop progressive hepatic fibrosis after liver transplantation (LT). Hepatic stellate cell (HSC) activation is a pivotal step in hepatic fibrosis and precedes clinically apparent fibrosis. We determined whether early HSC activation, measured in 4-month protocol post-LT biopsies, is predictive of subsequent development of more histologically severe recurrence of HCV. Early (4 month) post-LT HSC activation, as measured by alpha-smooth muscle actin (alpha-SMA) staining, was determined in liver biopsies from recipients with severe (fibrosis score > or = 2, n = 13) and with mild (fibrosis score of 0, n = 13) recurrence of HCV at one-year post-LT. Immunohistochemical staining for alpha-smooth muscle actin (alpha-SMA) was used to generate HSC activation scores (regional and total). Total HSC activation scores at 4 months were similar in patients with severe and mild HCV recurrence (3.9 +/- 2.0 vs. 2.7 +/- 2.2, P = 0.2). Regional HSC activation, assessed as parenchymal (zones 1, 2, and 3) or mesenchymal (portal tracts and fibrous septa), was different between the study groups, with higher mesenchymal scores predictive of progression. No patients in the mild recurrence group had detectable mesenchymal alpha-SMA staining vs. 46% (6/13) of patients with severe recurrence (P HCV or HSC-targeted therapy.

  9. Angiotensin receptor blockade recovers hepatic UCP2 expression and aconitase and SDH activities and ameliorates hepatic oxidative damage in insulin resistant rats.

    Science.gov (United States)

    Montez, Priscilla; Vázquez-Medina, José Pablo; Rodríguez, Rubén; Thorwald, Max A; Viscarra, José A; Lam, Lisa; Peti-Peterdi, Janos; Nakano, Daisuke; Nishiyama, Akira; Ortiz, Rudy M

    2012-12-01

    Metabolic syndrome (MetS) is commonly associated with elevated renin-angiotensin system, oxidative stress, and steatohepatitis with down-regulation of uncoupling proteins (UCPs). However, the mechanisms linking renin-angiotensin system, steatosis, and UCP2 to hepatic oxidative damage during insulin resistance are not described. To test the hypothesis that angiotensin receptor activation contributes to decreased hepatic UCP2 expression and aconitase activity and to increased oxidative damage after increased glucose intake in a model of MetS, lean and obese Long Evans rats (n = 10/group) were randomly assigned to the following groups: 1) untreated Long Evans Tokushima Otsuka (lean, strain control), 2) untreated Otsuka Long Evans Tokushima Fatty (OLETF) (MetS model), 3) OLETF + angiotensin receptor blocker (ARB) (10 mg olmesartan/kg·d × 6 wk), 4) OLETF + high glucose (HG) (5% in drinking water × 6 wk), and 5) OLETF + ARB + HG (ARB/HG × 6 wk). HG increased body mass (37%), plasma triglycerides (TGs) (35%), plasma glycerol (87%), plasma free fatty acids (28%), and hepatic nitrotyrosine (74%). ARB treatment in HG decreased body mass (12%), plasma TG (15%), plasma glycerol (23%), plasma free fatty acids (14%), and hepatic TG content (42%), suggesting that angiotensin receptor type 1 (AT1) activation and increased adiposity contribute to the development of obesity-related dyslipidemia. ARB in HG also decreased hepatic nitrotyrosine and increased hepatic UCP2 expression (59%) and aconitase activity (40%), as well as antioxidant enzyme activities (50-120%), suggesting that AT1 activation also contributes to protein oxidation, impaired lipid metabolism, and antioxidant metabolism in the liver. Thus, in addition to promoting obesity-related hypertension, AT1 activation may also impair lipid metabolism and antioxidant capacity, resulting in steatosis via decreased UCP2 and tricarboxylic acid cycle activity.

  10. Dietary sesamin and docosahexaenoic and eicosapentaenoic acids synergistically increase the gene expression of enzymes involved in hepatic peroxisomal fatty acid oxidation in rats.

    Science.gov (United States)

    Arachchige, Premakumara G; Takahashi, Yoko; Ide, Takashi

    2006-03-01

    The interaction of sesamin, one of the most abundant lignans in sesame seed, and highly purified docosahexaenoic acid (DHA) or eicosapentaenoic acid (EPA) in the form of ethyl ester in affecting hepatic fatty acid oxidation was examined in rats. In the first experiment, 3 groups of rats were fed with purified experimental diets free of n-3 fatty acid ethyl ester and containing 0%, 0.2%, and 0.4% sesamin (1:1 mixture of sesamin and episesamin), and 2 groups of animals were fed with a 2% DHA ethyl ester diet containing either 0% or 0.2% sesamin. In the second trial, 4 groups of rats were fed with either a 0% or a 2% EPA ethyl ester diet containing 0% or 0.2% sesamin. After 15 days of feeding, DHA and EPA ethyl esters added to a sesamin-free diet little affected the activity and messenger RNA (mRNA) levels of various enzymes involved in fatty acid oxidation. Sesamin increased the activity levels of various hepatic enzymes involved in fatty acid oxidation irrespective of the presence or absence of n-3 fatty acid ethyl ester in diets. However, the diet containing sesamin and DHA or EPA ethyl ester in combination increased many of these parameters synergistically. In particular, the peroxisomal palmitoyl-coenzyme A oxidation rate and acyl-coenzyme A oxidase activity level were much higher in rats fed with sesamin and DHA or EPA in combination than in animals fed with a diet free of n-3 fatty acid ethyl ester and containing sesamin. Analyses of mRNA levels revealed that a diet simultaneously containing sesamin and n-3 fatty acid ethyl ester increased the gene expression of various enzymes involved in peroxisomal fatty acid oxidation in a synergistic manner. However, the combination of sesamin and n-3 fatty acid ethyl esters was ineffective in causing a synergistic increase in mRNA levels of enzymes of mitochondrial fatty acid oxidation, microsomal cytochrome P-450 IV A1, and cytosolic liver-type fatty acid-binding protein. It was concluded that sesamin and DHA or EPA

  11. Spatial distribution of enzyme activities in the rhizosphere

    Science.gov (United States)

    Razavi, Bahar S.; Zarebanadkouki, Mohsen; Blagodatskaya, Evgenia; Kuzyakov, Yakov

    2015-04-01

    The rhizosphere, the tiny zone of soil surrounding roots, certainly represents one of the most dynamic habitat and interfaces on Earth. Activities of enzymes produced by both plant roots and microbes are the primary biological drivers of organic matter decomposition and nutrient cycling. That is why there is an urgent need in spatially explicit methods for the determination of the rhizosphere extension and enzyme distribution. Recently, zymography as a new technique based on diffusion of enzymes through the 1 mm gel plate for analysis has been introduced (Spohn & Kuzyakov, 2013). We developed the zymography technique to visualize the enzyme activities with a higher spatial resolution. For the first time, we aimed at quantitative imaging of enzyme activities as a function of distance from the root tip and the root surface in the soil. We visualized the two dimensional distribution of the activity of three enzymes: β-glucosidase, phosphatase and leucine amino peptidase in the rhizosphere of maize using fluorogenically labelled substrates. Spatial-resolution of fluorescent images was improved by direct application of a substrate saturated membrane to the soil-root system. The newly-developed direct zymography visualized heterogeneity of enzyme activities along the roots. The activity of all enzymes was the highest at the apical parts of individual roots. Across the roots, the enzyme activities were higher at immediate vicinity of the roots (1.5 mm) and gradually decreased towards the bulk soil. Spatial patterns of enzyme activities as a function of distance from the root surface were enzyme specific, with highest extension for phosphatase. We conclude that improved zymography is promising in situ technique to analyze, visualize and quantify spatial distribution of enzyme activities in the rhizosphere hotspots. References Spohn, M., Kuzyakov, Y., 2013. Phosphorus mineralization can be driven by microbial need for carbon. Soil Biology & Biochemistry 61: 69-75

  12. Effects of human diets of two different Japanese populations on cancer incidence in rat hepatic drug-metabolizing and antioxidant enzyme systems.

    Science.gov (United States)

    Kanke, Y; Iitoi, Y; Iwasaki, M; Iwase, Y; Iwama, M; Kimira, M; Takahashi, T; Tsugane, S; Watanabe, S; Akabane, M

    1996-01-01

    Hepatic enzyme systems of drug metabolism and antioxidation were investigated in rats fed the complete human diets consumed in the two Japanese prefectures, Akita and Okinawa, where the incidence of cancers was quite different: Okinawa had the lowest and Akita the highest age-adjusted mortality rate. In rats fed the human diet consumed in Okinawa, hepatic glutathione S-transferase activity was higher and lipid peroxide content was lower than in rats fed the diet consumed in Akita. These data might indicate that the number and/or quantity of the dietary components attributed to the detoxification of carcinogens and the scavenging reactive carcinogen species was much higher in the foods consumed in the population having lower cancer mortality rate.

  13. Effect of geniposide, a hypoglycemic glucoside, on hepatic regulating enzymes in diabetic mice induced by a high-fat diet and streptozotocin

    Institute of Scientific and Technical Information of China (English)

    Shaoyu WU; Guangfa WANG; Zhongqiu LIU; Jinjun RAO; Lin LU; Wei XU; Shuguang WU; Jiajie ZHANG

    2009-01-01

    Aim:Hepatic glycogen phosphorylase (GP) and glucose-6-phosphatase (G6Pase) play an important role in the control of blood glucose homeostasis and are proposed to be potential targets for anti-diabetic drugs.Geniposide is an iridoid gluco-side extracted from Gardenia jasminoides Ellis fruits and has been reported to have a hypoglycemic effect.However,little is known about the biochemical mechanisms by which geniposide regulates hepatic glucose-metabolizing enzymes.The pres-ent study investigates whether the hypoglycemic effect of geniposide is mediated by GP or G6Pase.Methods: Type 2 diabetic mice,induced by a high-fat diet and streptozotocin injection,were treated with or without geni-poside for 2 weeks.Blood glucose levels were monitored by a glucometer.Insulin concentrations were analyzed by the ELISA method.Total cholesterol (TC) and triglyceride (TG) levels were measured using LabassayTM kits.Activities of hepatic GP and G6Pase were measured by glucose-6-phosphate dehydrogenase-coupled reaction.Real-time RT-PCR and Western blotting were used to determine the mRNA and protein levels of both enzymes.Results: Geniposide (200 and 400 mg/kg) significantly decreased the blood glucose,insulin and TG levels in diabetic mice in a dose-dependent manner.This compound also decreased the expression of GP and G6Pase at mRNA and immunoreac-tive protein levels,as well as enzyme activity.Conclusion: Geniposide is an effective hypoglycemic agent in diabetic mice.The hypoglycemic effect of this compound may be mediated,at least in part,by inhibiting the GP and G6Pase activities.

  14. Type IV collagen-degrading enzyme activity in human serum.

    OpenAIRE

    Hashimoto, Noriaki; Kobayashi, Michio; Watanabe,Akiharu; Higashi, Toshiro; Tsuji, Takao

    1988-01-01

    Type IV collagen-degrading enzyme activity was detected in human serum. Serum was preincubated with 4-aminophenylmercuric acetate and trypsin to activate the enzyme prior to assay. Type IV collagen, purified from human placentas and radiolabeled with [1-14C] acetic anhydride, was used as the substrate. The enzyme activity was measured at pH 7.5 and inhibited by treatment with ethylenediaminetetraacetic acid or heat. The assay of type IV collagen-degrading enzyme in human serum might be useful...

  15. Type IV collagen-degrading enzyme activity in human serum.

    Directory of Open Access Journals (Sweden)

    Hashimoto,Noriaki

    1988-02-01

    Full Text Available Type IV collagen-degrading enzyme activity was detected in human serum. Serum was preincubated with 4-aminophenylmercuric acetate and trypsin to activate the enzyme prior to assay. Type IV collagen, purified from human placentas and radiolabeled with [1-14C] acetic anhydride, was used as the substrate. The enzyme activity was measured at pH 7.5 and inhibited by treatment with ethylenediaminetetraacetic acid or heat. The assay of type IV collagen-degrading enzyme in human serum might be useful for estimating the degradation of type IV collagen.

  16. Hepatic inflammation mediated by hepatitis C virus core protein is ameliorated by blocking complement activation

    Directory of Open Access Journals (Sweden)

    Hsu Chen-Ming

    2009-08-01

    Full Text Available Abstract Background The pathogenesis of inflammation and fibrosis in chronic hepatitis C virus (HCV infection remains unclear. Transgenic mice with constitutive HCV core over-expression display steatosis only. While the reasons for this are unclear, it may be important that core protein production in these models begins during gestation, in contrast to human hepatitis C virus infection, which occurs post-natally and typically in adults. AIMS: To more realistically model the effect of core protein production in the adult liver, we developed a mouse with conditional expression of HCV core and examined the effect of core protein production in the adult liver. Methods Liver biopsy samples from transgenic mice with tetracycline(tet-regulated conditional core protein expression were evaluated immunohistologically. Microarray analysis of HCV core transgenic mice with steatohepatitis pointed to a role of the complement pathway. This was further explored by blocking complement activation by in vivo administration of CD55 (decay accelerating factor for complement, which inhibits activation of C3. Results Transgenic mice exhibited low, intermediate, or high HCV core protein expression when fed a permissive diet of standard chow. Aside from hepatic steatosis, hepatic inflammation and fibrosis were seen in mice with intermediate levels of core protein. Microarray analyses of inflamed liver demonstrated activation of both the complement (C3 up-regulation and coagulation pathways (fibrinogen B up-regulation. Administration of CD55 reduced hepatic inflammation. Conclusion Transgenic mice that conditionally express intermediate HCV core protein develop inflammation, steatosis, and fibrosis. These effects mediated by HCV core are reduced by administration of CD55, a regulator of the complement pathway. The model may be valuable in investigating the pathogenesis of liver inflammation in chronic hepatitis C.

  17. Increase in sphingolipid catabolic enzyme activity during aging

    Institute of Scientific and Technical Information of China (English)

    Santosh J SACKET; Hae-young CHUNG; Fumikazu OKAJIMA; Dong-soon IM

    2009-01-01

    Aim:To understand the contribution of sphingolipid metabolism and its metabolites to development and aging.Methods: A systemic analysis on the changes in activity of sphingolipid metabolic enzymes in kidney, liver and brain tissues during development and aging was conducted. The study was conducted using tissues from 1-day-old to 720-day-old rats.Results: Catabolic enzyme activities as well as the level of sphingomyelinase (SMase) and ceramidase (CDase) were higher than that of anabolic enzyme activities, sphingomyelin synthase and ceramide synthase. This suggested an accumulation of ceramide and sphingosine during development and aging. The liver showed the highest neutral-SMase activity among the tested enzymes while the kidney and brain exhibited higher neutral-SMase and ceramidase activities, indicating a high production of ceramide in liver and ceramide/sphingosine in the kidney and brain. The activities of sphingolipid metabolic enzymes were significantly elevated in all tested tissues during development and aging, although the onset of significant increase in activity varied on the tissue and enzyme type. During aging, 18 out of 21 enzyme activities were further increased on day 720 compared to day 180.Conclusion: Differential increases in sphingolipid metabolic enzyme activities suggest that sphingolipids including ceramide and sphingosine might play important and dynamic roles in proliferation, differentiation and apoptosis during development and aging.

  18. Effects of Lanthanum on Hydrolytic Enzyme Activities in Red Soil

    Institute of Scientific and Technical Information of China (English)

    褚海燕; 朱建国; 谢祖彬; 李振高; 曹志洪; 曾青; 林先贵

    2002-01-01

    The effects of La on some hydrolytic enzyme activities in red soil were studied in incubation and pot culture experiments. In the incubation experiment, La slightly stimulates the activities of urease and acidic phosphatase in soil and strongly stimulates sucrase activity in soil. In the pot culture experiment, La stimulates the activities of urease, acidic phosphatase and sucrase to different degrees. The stimulative effects of rare earth elements (REE) on hydrolytic enzyme activities in soil may result in increasing yield of crops.

  19. Microbial Enzyme Activity and Carbon Cycling in Grassland Soil Fractions

    Science.gov (United States)

    Allison, S. D.; Jastrow, J. D.

    2004-12-01

    Extracellular enzymes are necessary to degrade complex organic compounds present in soils. Using physical fractionation procedures, we tested whether old soil carbon is spatially isolated from degradative enzymes across a prairie restoration chronosequence in Illinois, USA. We found that carbon-degrading enzymes were abundant in all soil fractions, including macroaggregates, microaggregates, and the clay fraction, which contains carbon with a mean residence time of ~200 years. The activities of two cellulose-degrading enzymes and a chitin-degrading enzyme were 2-10 times greater in organic matter fractions than in bulk soil, consistent with the rapid turnover of these fractions. Polyphenol oxidase activity was 3 times greater in the clay fraction than in the bulk soil, despite very slow carbon turnover in this fraction. Changes in enzyme activity across the restoration chronosequence were small once adjusted for increases in soil carbon concentration, although polyphenol oxidase activity per unit carbon declined by 50% in native prairie versus cultivated soil. These results are consistent with a `two-pool' model of enzyme and carbon turnover in grassland soils. In light organic matter fractions, enzyme production and carbon turnover both occur rapidly. However, in mineral-dominated fractions, both enzymes and their carbon substrates are immobilized on mineral surfaces, leading to slow turnover. Soil carbon accumulation in the clay fraction and across the prairie restoration chronosequence probably reflects increasing physical isolation of enzymes and substrates on the molecular scale, rather than the micron to millimeter scale.

  20. A Simple and Accurate Method for Measuring Enzyme Activity.

    Science.gov (United States)

    Yip, Din-Yan

    1997-01-01

    Presents methods commonly used for investigating enzyme activity using catalase and presents a new method for measuring catalase activity that is more reliable and accurate. Provides results that are readily reproduced and quantified. Can also be used for investigations of enzyme properties such as the effects of temperature, pH, inhibitors,…

  1. A Simple and Accurate Method for Measuring Enzyme Activity.

    Science.gov (United States)

    Yip, Din-Yan

    1997-01-01

    Presents methods commonly used for investigating enzyme activity using catalase and presents a new method for measuring catalase activity that is more reliable and accurate. Provides results that are readily reproduced and quantified. Can also be used for investigations of enzyme properties such as the effects of temperature, pH, inhibitors,…

  2. Exploration of the spontaneous fluctuating activity of single enzyme molecules

    NARCIS (Netherlands)

    A. Schwabe (Anne); T.R. Maarleveld (Timo); F.J. Bruggeman (Frank)

    2013-01-01

    htmlabstractSingle enzyme molecules display inevitable, stochastic fluctuations in their catalytic activity. In metabolism, for instance, the stochastic activity of individual enzymes is averaged out due to their high copy numbers per single cell. However, many processes inside cells rely on single

  3. Effects of Angiotensin Converting Enzyme Inhibitors on Liver Fibrosis in HIV and Hepatitis C Coinfection

    Directory of Open Access Journals (Sweden)

    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.

  4. Epigenetic regulation of hepatic stellate cell activation and liver fibrosis.

    Science.gov (United States)

    El Taghdouini, Adil; van Grunsven, Leo A

    2016-12-01

    Chronic liver injury to hepatocytes or cholangiocytes, when left unmanaged, leads to the development of liver fibrosis, a condition characterized by the excessive intrahepatic deposition of extracellular matrix proteins. Activated hepatic stellate cells constitute the predominant source of extracellular matrix in fibrotic livers and their transition from a quiescent state during fibrogenesis is associated with important alterations in their transcriptional and epigenetic landscape. Areas covered: We briefly describe the processes involved in hepatic stellate cell activation and discuss our current understanding of alterations in the epigenetic landscape, i.e DNA methylation, histone modifications and the functional role of non-coding RNAs that accompany this key event in the development of chronic liver disease. Expert commentary: Although great progress has been made, our understanding of the epigenetic regulation of hepatic stellate cell activation is limited and, thus far, insufficient to allow the development of epigenetic drugs that can selectively interrupt liver fibrosis.

  5. Hepatoprotective activity of Tridax procumbens against d-galactosamine/lipopolysaccharide-induced hepatitis in rats.

    Science.gov (United States)

    Ravikumar, Vilwanathan; Shivashangari, Kanchi Subramanian; Devaki, Thiruvengadam

    2005-10-03

    The hepatoprotective activity of aerial parts of Tridax procumbens was investigated against d-Galactosamine/Lipopolysaccharide (d-GalN/LPS) induced hepatitis in rats. d-GalN/LPS (300 mg/kg body weight/30 microg/kg body weight)-induced hepatic damage was manifested by a significant increase in the activities of marker enzymes (aspartate transaminase, alanine transaminase, alkaline phosphatase, lactate dehydrogenase and gamma glutamyl transferase) and bilirubin level in serum and lipids both in serum and liver. Pretreatment of rats with a chloroform insoluble fraction from ethanolic extract of Tridax procumbens reversed these altered parameters to normal values. The biochemical observations were supplemented by histopathological examination of liver sections. Results of this study revealed that Tridax procumbens could afford a significant protection in the alleviation of d-GalN/LPS-induced hepatocellular injury.

  6. Performance and Serum Hepatic Enzymes of Hy-Line W-36 Laying Hens Intoxicated with Dietary Carbon Tetrachloride

    Directory of Open Access Journals (Sweden)

    Hadavi A

    2015-12-01

    Full Text Available An experiment was conducted to study the effects of carbon tetrachloride (CCl4 on post-peak performance and serum enzymes of Hy-Line W-36 laying hens from 32-36 weeks of age. The experiment was carried out with a total of 192 laying hens in a completely randomized block design. During the experiment laying hens were allocated to 4 groups consisted of T1 no CCl4 as control diet, T2, T3 and T4 control diet supplemented with 1, 3 and 5 mL CCl4/100 g diet, respectively. Each experimental group was divided into 6 blocks of 8 hens each. Egg production, cracked egg percentage and feed intake were recorded weekly. Blood samples were taken from wing veins of hens at the middle and end of the experiment to measure serum hepatic enzymes of alkaline phosphatase, alanine aminotransferase and aspartate aminotransferase. Data showed that in comparison with the control group, the inclusion of CCl4 to the diets had no significant effect on performance parameters. However, by increasing the level of CCl4, egg production was linearly decreased and feed intake was linearly increased (P < 0.05. The effect of CCl4 on cracked eggs was significant and this effect was linearly increased (P < 0.05. Dietary supplementation of 3 and 5 mL CCl4 elevated the serum concentration of hepatic enzymes of alkaline phosphatase, aspartate aminotransferase and alanine aminotransferase, linearly (P < 0.0001. In conclusion, the dietary supplementation of CCl4 has the ability to decrease the performance and egg quality. CCl4 is also a potent hepatic toxicity inducer and may damage liver hepatocytes. Therefore, the level of 3 mL CCl4 was assigned as the one had the maximum negative effect on serum hepatic enzymes concentration (maximum liver damage alongside the minimum negative effect on laying hen performance for further studies.

  7. Operating Conditions Effects Onenzyme Activity: Case Enzyme Protease

    Directory of Open Access Journals (Sweden)

    Adel Oueslati

    2014-09-01

    Full Text Available The Proteases an enzyme added to detergents to degrade the protein spots origin.Their action is manifested through its activity the middle of washing clothes. This activity depends on the operating conditions. In this article, the effects of temperature and pH of the reaction and the substrate concentration and time of washing medium on the enzyme activity were studied. There action mechanism has been shown. The activity measurements were made by absorption spectrometry

  8. Diffusional correlations among multiple active sites in a single enzyme.

    Science.gov (United States)

    Echeverria, Carlos; Kapral, Raymond

    2014-04-07

    Simulations of the enzymatic dynamics of a model enzyme containing multiple substrate binding sites indicate the existence of diffusional correlations in the chemical reactivity of the active sites. A coarse-grain, particle-based, mesoscopic description of the system, comprising the enzyme, the substrate, the product and solvent, is constructed to study these effects. The reactive and non-reactive dynamics is followed using a hybrid scheme that combines molecular dynamics for the enzyme, substrate and product molecules with multiparticle collision dynamics for the solvent. It is found that the reactivity of an individual active site in the multiple-active-site enzyme is reduced substantially, and this effect is analyzed and attributed to diffusive competition for the substrate among the different active sites in the enzyme.

  9. Effect of inositol requiring enzyme 1-mediated endoplasmic reticulum stress in liver cell apoptosis of experimental fulminant hepatic failure and its significance

    Institute of Scientific and Technical Information of China (English)

    甄真

    2013-01-01

    Objective To study the role of inositol requiring enzyme 1(IRE1)-mediated endoplasmic reticulum stress on hepatocyte apoptosis of experimental fulminant hepatic failure(FHF). Methods Thirty male depuratory Wistar

  10. Hepatic steroid inactivating enzymes, hepatic portal blood flow, and corpus luteum blood perfusion in lactating dairy cattle

    Science.gov (United States)

    In ruminants, a decrease in pregnancy rates may be due to decreased concentrations of progesterone (P4). It is important to note that both production from the corpus luteum and/or hepatic steroid inactivation impacts peripheral concentrations of P4. Cattle with an elevated dry matter intake have inc...

  11. Enzyme

    Science.gov (United States)

    Enzymes are complex proteins that cause a specific chemical change in all parts of the body. For ... use them. Blood clotting is another example of enzymes at work. Enzymes are needed for all body ...

  12. assessment of some hepatic enzyme activities in adult rabbits ...

    African Journals Online (AJOL)

    Uwaifoh

    2012-04-30

    Apr 30, 2012 ... At the end of the experiment, a dose depended significant ... It is a known fact that plants serve as sources of novel drug compounds and as such, medicines derived from plants ... protective potentials couple with reproductive power of G. kola, there is this ..... Clinical biochemistry of domestic animals.

  13. [Gallbladder motor activity in patients with virus hepatitis B].

    Science.gov (United States)

    Mamos, Arkadiusz; Wichan, Paweł; Chojnacki, Jan; Grzegorczyk, Krzysztof

    2003-12-01

    In acute stage of virus hepatitis B patients often complain of dyspeptic discomfort. They may be a consequence of alimentary tract motor activity disorders including these of gallbladder. Routine ultrasonography in an early phase of virus hepatitis often reveals gallbladder wall thickening what may confirm the above thesis. Thus, a group of 15 patients in an acute phase of virus hepatitis B was subjected to examinations. Gallbladder motor activity was assessed by ultrasonographic method determining its total volume and ejection fraction and volume after test meal stimulus. First examination was performed in the first week since the appearance of yellowing of the walls, successive in 4 and 8 week of the disease. Obtained results were compared to the values obtained in the group of 25 healthy volunteers. It was found out that gallbladder volume was significantly decreased and ejection fraction increased in the acute phase of virus hepatitis B than in the controls. This may speak for gallbladder hyperreactivity in patients in the course of virus hepatitis B. These disorders decreased during two-month observation but even in the 8 week the investigated parameters differed from those found in the control group.

  14. Effect of Xin-Jiang-Tang Granules on Activity of Hepatic Glycometabolic Key Enzymes and Liver Function in Type 2 Diabetic Rats%新降糖颗粒对2型糖尿病大鼠糖代谢关键酶活性及肝功能的影响

    Institute of Scientific and Technical Information of China (English)

    吕小龙; 杨元生; 陈垦; 王晖; 周文; 冯英巧; 刘少波

    2015-01-01

    目的:探讨新降糖颗粒对链脲佐菌素诱导的2型糖尿病大鼠糖代谢关键酶活性及肝功能的影响。方法:50只雄性SD大鼠除8只给予普通饲料喂养作为正常组外,其余大鼠均采用高脂饲料喂养4周,一次性腹腔注射链脲佐菌素(STZ)40 mg·kg-1建立2型糖尿病大鼠模型,随机分为模型组、二甲双胍组(0.15 g·kg-1)和新降糖颗粒高、低剂量组(12.64、6.32 g·kg-1),每天灌胃给药1次,连续8周。干预8周后,检测大鼠空腹血糖(FBG)、空腹胰岛素(FINS)、糖化血红蛋白(HbA1c)、肝糖原,检测肝功能指标丙氨酸氨基转移酶(ALT)、天门冬氨酸氨基转移酶(AST)、碱性磷酸酶(ALP)、γ-谷氨酰转移酶(γ-GT)及糖代谢关键酶己糖激酶(HK)、果糖-6-磷酸激酶(PFK)、丙酮酸激酶(PK)及葡萄糖-6-磷酸脱氢酶(G6PDH)活性。结果:与模型组比较,新降糖颗粒能明显降低FBG、FINS、内环境稳态模型法胰岛素抵抗指数(HOMA-IR)、HbA1c及肝功能指标ALT、AST、ALP及γ-GT(P<0.05,P<0.01),增加肝糖原含量(P<0.01),增强糖代谢关键酶HK、PFK、PK及G6PDH的活性(P<0.05,P<0.01)。结论:新降糖颗粒能有效改善2型糖尿病大鼠的症状,推测其机制可能与新降糖颗粒增强肝脏糖代谢关键酶HK、PFK、PK及G6PDH活性,促进肝糖原合成,改善肝功能,下调血清FINS水平,改善胰岛素抵抗等多因素有关,最终降低了2型糖尿病大鼠的FBG、HbA1c水平。%This article was aimed to investigate the effect of theXin-Jiang-Tang(XJT) Granules on activity of hepatic glycometabolic key enzymes and liver function in streptozotocin (STZ)-induced type 2 diabetes mellitus (T2DM) rats. Fifty male SD rats were randomly divided into the normal control group with 8 rats fed with normal diet, and other rats in the model group fed with high-fat diet for 4 weeks. And then, STZ

  15. Hepatitis B virus e antigen induces activation of rat hepatic stellate cells

    Energy Technology Data Exchange (ETDEWEB)

    Zan, Yanlu [Center for Molecular Virology, CAS Key Laboratory of Pathogenic Microbiology and Immunology, Institute of Microbiology, Chinese Academy of Sciences, Beijing 100101 (China); University of Chinese Academy of Sciences, Beijing 100049 (China); Zhang, Yuxia, E-mail: yzhang@wehi.edu.au [Center for Molecular Virology, CAS Key Laboratory of Pathogenic Microbiology and Immunology, Institute of Microbiology, Chinese Academy of Sciences, Beijing 100101 (China); Tien, Po, E-mail: tienpo@sun.im.ac.cn [Center for Molecular Virology, CAS Key Laboratory of Pathogenic Microbiology and Immunology, Institute of Microbiology, Chinese Academy of Sciences, Beijing 100101 (China)

    2013-06-07

    Highlights: •HBeAg expression in HSCs induced production of ECM protein and liver fibrotic markers. •The activation and proliferation of HSCs were mediated by TGF-β. •HBeAg protein purified from cell medium directly activated HSCs. -- Abstract: Chronic hepatitis B virus infection is a major cause of hepatic fibrosis, leading to liver cirrhosis and hepatocellular carcinoma. Hepatitis B virus e antigen (HBeAg) is an accessory protein of HBV, not required for viral replication but important for natural infection in vivo. Hepatic stellate cells (HSCs) are the major producers of excessive extracellular matrix during liver fibrogenesis. Therefore, we examined the influence of HBeAg on HSCs. The rat HSC line HSC-T6 was transfected with HBeAg plasmids, and expression of α-smooth muscle actin, collagen I, transforming growth factor-β1 (TGF-β), and tissue inhibitors of metalloproteinase 1 (TIMP-1) was investigated by quantitative real-time PCR. The proliferation of HSCs was determined by MTS analysis. HBeAg transduction induced up-regulation of these fibrogenic genes and proliferation of HSCs. We found that HBeAg induced TGF-β secretion in HSCs, and the activation of HSCs was prevented by a neutralizing anti-TGF-β antibody. Depletion and addition of HBeAg protein in conditioned medium from HSC-T6 cells transduced with HBeAg indicated that HBeAg directly induced the activation and proliferation of rat primary HSCs. Taken together, HBeAg induces the activation and proliferation of HSCs, mainly mediated by TGF-β, and HBeAg protein purified from cell medium can directly activate HSCs.

  16. Eucommia ulmoides Oliver extract, aucubin, and geniposide enhance lysosomal activity to regulate ER stress and hepatic lipid accumulation.

    Directory of Open Access Journals (Sweden)

    Hwa-Young Lee

    Full Text Available Eucommia ulmoides Oliver is a natural product widely used as a dietary supplement and medicinal plant. Here, we examined the potential regulatory effects of Eucommia ulmoides Oliver extracts (EUE on hepatic dyslipidemia and its related mechanisms by in vitro and in vivo studies. EUE and its two active constituents, aucubin and geniposide, inhibited palmitate-induced endoplasmic reticulum (ER stress, reducing hepatic lipid accumulation through secretion of apolipoprotein B and associated triglycerides and cholesterol in human HepG2 hepatocytes. To determine how EUE diminishes the ER stress response, lysosomal and proteasomal protein degradation activities were analyzed. Although proteasomal activity was not affected, lysosomal enzyme activities including V-ATPase were significantly increased by EUE as well as aucubin and geniposide in HepG2 cells. Treatment with the V-ATPase inhibitor, bafilomycin, reversed the inhibition of ER stress, secretion of apolipoprotein B, and hepatic lipid accumulation induced by EUE or its component, aucubin or geniposide. In addition, EUE was determined to regulate hepatic dyslipidemia by enhancing lysosomal activity and to regulate ER stress in rats fed a high-fat diet. Together, these results suggest that EUE and its active components enhance lysosomal activity, resulting in decreased ER stress and hepatic dyslipidemia.

  17. Why do crown ethers activate enzymes in organic solvents?

    NARCIS (Netherlands)

    Unen, van Dirk-Jan; Engbersen, Johan F.J.; Reinhoudt, David N.

    2002-01-01

    One of the major drawbacks of enzymes in nonaqueous solvents is that their activity is often dramatically low compared to that in water. This limitation can be largely overcome by crown ether treatment of enzymes. In this paper, we describe a number of carefully designed new experiments that have im

  18. Ecological effects of atmospheric nitrogen deposition on soil enzyme activity

    Institute of Scientific and Technical Information of China (English)

    WANG Cong-yan; Lv Yan-na; LIU Xue-yan Liu; WANG Lei

    2013-01-01

    The continuing increase in human activities is causing global changes such as increased deposition of atmospheric nitrogen.There is considerable interest in understanding the effects of increasing atmospheric nitrogen deposition on soil enzyme activities,specifically in terms of global nitrogen cycling and its potential future contribution to global climate change.This paper summarizes the ecological effects of atmospheric nitrogen deposition on soil enzyme activities,including size-effects,stage-effects,site-effects,and the effects of different levels and forms of atmospheric nitrogen deposition.We discuss needs for further research on the relationship between atmospheric nitrogen deposition and soil enzymes.

  19. Hepatitis

    Institute of Scientific and Technical Information of China (English)

    1993-01-01

    930140 Hepatocyte stimulator peptide and itsclinical significance in viral hepatitis.ZHOUWeiping(周卫平),et al.Instit Viral Hepatitis,Chongqing Med Univ,630010.Chin J InternMed 1992;31(10):626-628.Hepatocyte stimulator peptide(HSP)is anewly developed hepatic stimulator substance.Its monoclonal antibodies have been obtained inour laboratory.In this study,HSP was deter-mined in the sera of 315 subjects including pa-

  20. Hepatitis

    Institute of Scientific and Technical Information of China (English)

    2010-01-01

    2010349 Relationships between serum hepatitis B virus load in mothers,free maternal DNA in peripheral blood of newborns and hepatitis B virus infection of newborns. WEI Junni(魏俊妮),et al. Dept Epidemiol,Shanxi Med Univ,Taiyuan 030001. Chin J Infect Dis 2010;28(5):297-300. Objective To study the relationships between serum hepatitis B virus (HBV) DNA level

  1. Enzyme activity in banana fruits rotted by Botryodiplodia theobromae Pat.

    Directory of Open Access Journals (Sweden)

    Nityananda Chakraborty

    2015-06-01

    Full Text Available Peroxidase and polyphenol oxidase activities in fruits of two cultivars of banana, 'champa' and 'kanthali' rotted by Botryodiplodia theobromae Pat. was studied. The enzymes showed much higher activities in infected than that in uninfected 'tissues. Increase in peroxidase activity was evidently inhibited by cycloheximide. Polyphenol oxidase activity was also inhibited in presence of phenylthiourea and Na-diethyldithiocarbamate more strongly by the former. Increase in activities seemed to be due to increased sytheses of the enzymes. In an in vitro culture, the fungus exhibited some peroxidase but no polyphenoloxidase activity.

  2. Investigation of Antioxidant and Hepatoprotective Activity of Standardized Curcuma xanthorrhiza Rhizome in Carbon Tetrachloride-Induced Hepatic Damaged Rats

    Directory of Open Access Journals (Sweden)

    Sutha Devaraj

    2014-01-01

    Full Text Available Curcuma xanthorrhiza (CX has been used for centuries in traditional system of medicine to treat several diseases such as hepatitis, liver complaints, and diabetes. It has been consumed as food supplement and “jamu” as a remedy for hepatitis. Hence, CX was further explored for its potential as a functional food for liver related diseases. As such, initiative was taken to evaluate the antioxidant and hepatoprotective potential of CX rhizome. Antioxidant activity of the standardized CX fractions was determined using in vitro assays. Hepatoprotective assay was conducted against carbon tetrachloride- (CCl4- induced hepatic damage in rats at doses of 125, 250, and 500 mg/kg of hexane fraction. Highest antioxidant activity was found in hexane fraction. In the case of hepatoprotective activity, CX hexane fraction showed significant improvement in terms of a biochemical liver function, antioxidative liver enzymes, and lipid peroxidation activity. Good recovery was observed in the treated hepatic tissues histologically. Hence, the results concluded that CX hexane fraction possessed prominent hepatoprotective activities which might be due to its in vitro antioxidant activity. These findings also support the use of CX as a functional food for hepatitis remedy in traditional medicinal system.

  3. Metformin inhibits glutaminase activity and protects against hepatic encephalopathy.

    Science.gov (United States)

    Ampuero, Javier; Ranchal, Isidora; Nuñez, David; Díaz-Herrero, María del Mar; Maraver, Marta; del Campo, José Antonio; Rojas, Ángela; Camacho, Inés; Figueruela, Blanca; Bautista, Juan D; Romero-Gómez, Manuel

    2012-01-01

    To investigate the influence of metformin use on liver dysfunction and hepatic encephalopathy in a retrospective cohort of diabetic cirrhotic patients. To analyze the impact of metformin on glutaminase activity and ammonia production in vitro. Eighty-two cirrhotic patients with type 2 diabetes were included. Forty-one patients were classified as insulin sensitizers experienced (metformin) and 41 as controls (cirrhotic patients with type 2 diabetes mellitus without metformin treatment). Baseline analysis included: insulin, glucose, glucagon, leptin, adiponectin, TNFr2, AST, ALT. HOMA-IR was calculated. Baseline HE risk was calculated according to minimal hepatic encephalopathy, oral glutamine challenge and mutations in glutaminase gene. We performed an experimental study in vitro including an enzymatic activity assay where glutaminase inhibition was measured according to different metformin concentrations. In Caco2 cells, glutaminase activity inhibition was evaluated by ammonia production at 24, 48 and 72 hours after metformina treatment. Hepatic encephalopathy was diagnosed during follow-up in 23.2% (19/82): 4.9% (2/41) in patients receiving metformin and 41.5% (17/41) in patients without metformin treatment (logRank 9.81; p=0.002). In multivariate analysis, metformin use [H.R.11.4 (95% CI: 1.2-108.8); p=0.034], age at diagnosis [H.R.1.12 (95% CI: 1.04-1.2); p=0.002], female sex [H.R.10.4 (95% CI: 1.5-71.6); p=0.017] and HE risk [H.R.21.3 (95% CI: 2.8-163.4); p=0.003] were found independently associated with hepatic encephalopathy. In the enzymatic assay, glutaminase activity inhibition reached 68% with metformin 100 mM. In Caco2 cells, metformin (20 mM) decreased glutaminase activity up to 24% at 72 hours post-treatment (p<0.05). Metformin was found independently related to overt hepatic encephalopathy in patients with type 2 diabetes mellitus and high risk of hepatic encephalopathy. Metformin inhibits glutaminase activity in vitro. Therefore, metformin use seems

  4. Metformin Inhibits Glutaminase Activity and Protects against Hepatic Encephalopathy

    Science.gov (United States)

    Ampuero, Javier; Ranchal, Isidora; Nuñez, David; Díaz-Herrero, María del Mar; Maraver, Marta; del Campo, José Antonio; Rojas, Ángela; Camacho, Inés; Figueruela, Blanca; Bautista, Juan D.; Romero-Gómez, Manuel

    2012-01-01

    Aim To investigate the influence of metformin use on liver dysfunction and hepatic encephalopathy in a retrospective cohort of diabetic cirrhotic patients. To analyze the impact of metformin on glutaminase activity and ammonia production in vitro. Methods Eighty-two cirrhotic patients with type 2 diabetes were included. Forty-one patients were classified as insulin sensitizers experienced (metformin) and 41 as controls (cirrhotic patients with type 2 diabetes mellitus without metformin treatment). Baseline analysis included: insulin, glucose, glucagon, leptin, adiponectin, TNFr2, AST, ALT. HOMA-IR was calculated. Baseline HE risk was calculated according to minimal hepatic encephalopathy, oral glutamine challenge and mutations in glutaminase gene. We performed an experimental study in vitro including an enzymatic activity assay where glutaminase inhibition was measured according to different metformin concentrations. In Caco2 cells, glutaminase activity inhibition was evaluated by ammonia production at 24, 48 and 72 hours after metformina treatment. Results Hepatic encephalopathy was diagnosed during follow-up in 23.2% (19/82): 4.9% (2/41) in patients receiving metformin and 41.5% (17/41) in patients without metformin treatment (logRank 9.81; p = 0.002). In multivariate analysis, metformin use [H.R.11.4 (95% CI: 1.2–108.8); p = 0.034], age at diagnosis [H.R.1.12 (95% CI: 1.04–1.2); p = 0.002], female sex [H.R.10.4 (95% CI: 1.5–71.6); p = 0.017] and HE risk [H.R.21.3 (95% CI: 2.8–163.4); p = 0.003] were found independently associated with hepatic encephalopathy. In the enzymatic assay, glutaminase activity inhibition reached 68% with metformin 100 mM. In Caco2 cells, metformin (20 mM) decreased glutaminase activity up to 24% at 72 hours post-treatment (p<0.05). Conclusions Metformin was found independently related to overt hepatic encephalopathy in patients with type 2 diabetes mellitus and high risk of hepatic encephalopathy. Metformin

  5. Metformin inhibits glutaminase activity and protects against hepatic encephalopathy.

    Directory of Open Access Journals (Sweden)

    Javier Ampuero

    Full Text Available AIM: To investigate the influence of metformin use on liver dysfunction and hepatic encephalopathy in a retrospective cohort of diabetic cirrhotic patients. To analyze the impact of metformin on glutaminase activity and ammonia production in vitro. METHODS: Eighty-two cirrhotic patients with type 2 diabetes were included. Forty-one patients were classified as insulin sensitizers experienced (metformin and 41 as controls (cirrhotic patients with type 2 diabetes mellitus without metformin treatment. Baseline analysis included: insulin, glucose, glucagon, leptin, adiponectin, TNFr2, AST, ALT. HOMA-IR was calculated. Baseline HE risk was calculated according to minimal hepatic encephalopathy, oral glutamine challenge and mutations in glutaminase gene. We performed an experimental study in vitro including an enzymatic activity assay where glutaminase inhibition was measured according to different metformin concentrations. In Caco2 cells, glutaminase activity inhibition was evaluated by ammonia production at 24, 48 and 72 hours after metformina treatment. RESULTS: Hepatic encephalopathy was diagnosed during follow-up in 23.2% (19/82: 4.9% (2/41 in patients receiving metformin and 41.5% (17/41 in patients without metformin treatment (logRank 9.81; p=0.002. In multivariate analysis, metformin use [H.R.11.4 (95% CI: 1.2-108.8; p=0.034], age at diagnosis [H.R.1.12 (95% CI: 1.04-1.2; p=0.002], female sex [H.R.10.4 (95% CI: 1.5-71.6; p=0.017] and HE risk [H.R.21.3 (95% CI: 2.8-163.4; p=0.003] were found independently associated with hepatic encephalopathy. In the enzymatic assay, glutaminase activity inhibition reached 68% with metformin 100 mM. In Caco2 cells, metformin (20 mM decreased glutaminase activity up to 24% at 72 hours post-treatment (p<0.05. CONCLUSIONS: Metformin was found independently related to overt hepatic encephalopathy in patients with type 2 diabetes mellitus and high risk of hepatic encephalopathy. Metformin inhibits glutaminase

  6. Fish oil at low dietary levels enhances physiological activity of sesamin to increase hepatic fatty acid oxidation in rats.

    Science.gov (United States)

    Ide, Takashi

    2012-11-01

    We previously demonstrated that a diet containing fish oil at a level of 80 g/kg strongly stimulated the physiological activity of a sesame sesamin preparation containing sesamin and episesamin at equal amounts to increase hepatic fatty acid oxidation. This study was conducted to clarify whether fish oil at lower dietary levels enhances the physiological activity of sesamin to increase hepatic fatty acid oxidation. Rats were fed experimental diets supplemented with 0 or 2 g sesamin/kg, and containing 0, 15 or 30 g fish oil/kg for 15 days. Among rats fed sesamin-free diets, diets containing 15 and 30 g fish oil/kg slightly increased the activity of enzymes involved in hepatic fatty acid oxidation. Sesamin increased these values irrespective of the presence or absence of fish oil in diets; however, the extent of the increase of many parameters was much greater in rats given fish oil-containing diets than in those fed a fish oil-free diet. Diets simultaneously containing sesamin and fish oil increased the gene expression of various peroxisomal fatty acid oxidation enzymes in a synergistic manner; but they were ineffective in causing a synergistic increase in mRNA levels of mitochondrial fatty acid oxidation enzymes. The extent of the synergistic increase in the activity of hepatic fatty acid oxidation enzymes and mRNA levels of the peroxisomal enzymes was indistinguishable between diets containing 15 and 30 g fish oil/kg and appeared comparable to that observed previously with a diet containing 80 g fish oil/kg.

  7. Is autoimmune chronic active hepatitis a HCV-related disease?

    Science.gov (United States)

    Magrin, S; Craxì, A; Fiorentino, G; Fabiano, C; Provenzano, G; Pinzello, G B; Palazzo, U; Almasio, P; Pagliaro, L

    1991-07-01

    We evaluated the specificity and clinical relevance of anti-hepatitis C virus antibody positivity in 22 HBsAg-negative patients with autoimmune (anti-nuclear, anti-actin or anti-liver-kidney microsomal antibody positive) chronic active hepatitis. An ELISA anti-HCV test and a recombinant immunoblot assay (RIBA-HCV) were used. Thirteen patients (59%) were anti-HCV positive and five (23%) anti-HCV negative by both ELISA and RIBA-HCV tests. Four patients (18%) were borderline positive by ELISA (OD less than 1.0), and three of them (all with severe disease) were negative by RIBA. Histologic necroinflammation, AST/ALT and gamma-globulins levels were higher and response to prednisolone treatment was better in RIBA anti-HCV-negative than in anti-HCV-positive cases. We confirmed with both RIBA and ELISA tests the high prevalence of anti-HCV already reported by ELISA in anti-nuclear and anti-liver-kidney microsomal antibody positive chronic active hepatitis. False positive for anti-HCV (i.e., a positive ELISA test not confirmed by RIBA) occurred only among patients with severe disease. Since RIBA-negative subjects showed the best response to corticosteroid, they might represent the only subset of cases of 'true' autoimmune chronic active hepatitis.

  8. Successful Interferon Therapy Reverses Enhanced Hepatic Progenitor Cell Activation in Patients with Chronic Hepatitis C.

    Science.gov (United States)

    Noritake, Hidenao; Kobayashi, Yoshimasa; Ooba, Yukimasa; Matsunaga, Erika; Ohta, Kazuyoshi; Shimoyama, Shin; Yamazaki, Satoru; Chida, Takeshi; Kawata, Kazuhito; Sakaguchi, Takanori; Suda, Takafumi

    2015-12-01

    The enhanced accumulation of hepatic progenitor cells (HPCs) is related to the risk of progression to hepatocellular carcinoma (HCC). Interferon (IFN) treatment reduces HCC risk in patients with chronic hepatitis C virus (HCV) infection. However, the underlying mechanisms remain unclear. The aim of this study was to examine the effects of IFN treatment on HPC activation in HCV patients. Immunohistochemical detection and computer-assisted quantitative image analyses of cytokeratin 7 (CK7) were performed to evaluate HPC activation in paired pre- and post-treatment liver biopsies from 18 HCV patients with sustained virological response (SVR) to IFN-based therapy and from 23 patients without SVR, as well as normal liver tissues obtained from surgical resection specimens of 10 patients. Pretreatment HCV livers showed increased CK7 immunoreactivity, compared with normal livers (HCV: median, 1.38%; normal: median, 0.69%, P=0.006). IFN treatment reduced hepatic CK7 immunoreactivity (median, 1.57% pre-IFN vs. 0.69% post-IFN, P=0.006) in SVR patients, but not in non-SVR patients. The development of HCC following IFN treatment was encountered in 3 non-SVR patients who showed high post-IFN treatment CK7 immunoreactivity (>4%). Successful IFN therapy can reverse enhanced HPC activation in HCV patients, which may contribute to the reduced risk of HCC development in these patients.

  9. activity of enzyme trypsin immobilized onto macroporous poly(epoxy ...

    African Journals Online (AJOL)

    dell

    The immobilization yield and the influence of pH and temperature conditions on the activity of the ... immobilized enzyme that might change the .... trypsin was packed in a rubber stoppered ..... elastic polyacrylamide gels prepared at subzero ...

  10. Fluorogenic Peptide Substrate for Quantification of Bacterial Enzyme Activities

    National Research Council Canada - National Science Library

    Ismail H Al-abdullah; Karine Bagramyan; Shiela Bilbao; Meirigeng Qi; Markus Kalkum

    2017-01-01

    A novel peptide substrate (A G G P L G P P G P G G) was developed for quantifying the activities of bacterial enzymes using a highly sensitive Fluorescence Resonance Energy Transfer (FRET) based assay...

  11. Diced electrophoresis gel assay for screening enzymes with specified activities.

    Science.gov (United States)

    Komatsu, Toru; Hanaoka, Kenjiro; Adibekian, Alexander; Yoshioka, Kentaro; Terai, Takuya; Ueno, Tasuku; Kawaguchi, Mitsuyasu; Cravatt, Benjamin F; Nagano, Tetsuo

    2013-04-24

    We have established the diced electrophoresis gel (DEG) assay as a proteome-wide screening tool to identify enzymes with activities of interest using turnover-based fluorescent substrates. The method utilizes the combination of native polyacrylamide gel electrophoresis (PAGE) with a multiwell-plate-based fluorometric assay to find protein spots with the specified activity. By developing fluorescent substrates that mimic the structure of neutrophil chemoattractants, we could identify enzymes involved in metabolic inactivation of the chemoattractants.

  12. Enzyme activity measurement via spectral evolution profiling and PARAFAC

    DEFF Research Database (Denmark)

    Baum, Andreas; Meyer, Anne S.; Garcia, Javier Lopez

    2013-01-01

    The recent advances in multi-way analysis provide new solutions to traditional enzyme activity assessment. In the present study enzyme activity has been determined by monitoring spectral changes of substrates and products in real time. The method relies on measurement of distinct spectral...... fingerprints of the reaction mixture at specific time points during the course of the whole enzyme catalyzed reaction and employs multi-way analysis to detect the spectral changes. The methodology is demonstrated by spectral evolution profiling of Fourier Transform Infrared (FTIR) spectral fingerprints using...

  13. Relationship between differential hepatic microRNA expression and decreased hepatic cytochrome P450 3A activity in cirrhosis.

    Directory of Open Access Journals (Sweden)

    Raj Vuppalanchi

    Full Text Available BACKGROUND AND AIM: Liver cirrhosis is associated with decreased hepatic cytochrome P4503A (CYP3A activity but the pathogenesis of this phenomenon is not well elucidated. In this study, we examined if certain microRNAs (miRNA are associated with decreased hepatic CYP3A activity in cirrhosis. METHODS: Hepatic CYP3A activity and miRNA microarray expression profiles were measured in cirrhotic (n=28 and normal (n=12 liver tissue. Hepatic CYP3A activity was measured via midazolam hydroxylation in human liver microsomes. Additionally, hepatic CYP3A4 protein concentration and the expression of CYP3A4 mRNA were measured. Analyses were conducted to identify miRNAs which were differentially expressed between two groups but also were significantly associated with lower hepatic CYP3A activity. RESULTS: Hepatic CYP3A activity in cirrhotic livers was 1.7-fold lower than in the normal livers (0.28 ± 0.06 vs. 0.47 ± 0.07mL* min(-1*mg protein(-1 (mean ± SEM, P=0.02. Six microRNAs (miR-155, miR-454, miR-582-5p, let-7f-1*, miR-181d, and miR-500 had >1.2-fold increase in cirrhotic livers and also had significant negative correlation with hepatic CYP3A activity (range of r = -0.44 to -0.52, P <0.05. Notably, miR-155, a known regulator of liver inflammation, had the highest fold increase in cirrhotic livers (2.2-fold, P=4.16E-08 and significantly correlated with hepatic CYP3A activity (r=-0.50, P=0.017. The relative expression (2(-ΔΔCt mean ± SEM of hepatic CYP3A4 mRNA was significantly higher in cirrhotic livers (21.76 ± 2.65 vs. 5.91 ± 1.29, P=2.04E-07 but their levels did not significantly correlate with hepatic CYP3A activity (r=-0.43, P=0.08. CONCLUSION: The strong association between certain miRNAs, notably miR-155, and lower hepatic CYP3A activity suggest that altered miRNA expression may regulate hepatic CYP3A activity.

  14. Hepatitis

    Science.gov (United States)

    ... inflammation of the liver.” This inflammation can be caused by a wide variety of toxins, drugs, and metabolic diseases, as well as infection. There are at least 5 hepatitis viruses. Hepatitis A is contracted when a child eats food or drinks water that is contaminated with the virus or has ...

  15. Hepatitis

    Institute of Scientific and Technical Information of China (English)

    1997-01-01

    970349 Primary structure and variability of partialsequences in nonstructural gene 5 region of hepatitis Gvirus, CHANG Jinhong(常锦红), et al. Hepatol Instis,People’s Hosp, Beijing Med Univ, Beijing, 100044. NatlMed J China 1997; 77(3): 178-182. Objective: To sequence partial genome of hepatitis G

  16. Hepatitis

    Institute of Scientific and Technical Information of China (English)

    2009-01-01

    2009209 Effects of chronic hepatitis B virus infection on human hepatic cytochrome P450 2C9.ZHO Fuping(周福平),et al.Dept Infect Dis,Shanghai Changzheng Hosp,Shanghai 200003.Chin J Infect Dis,2009;27(2):94-98.

  17. Hepatitis

    Institute of Scientific and Technical Information of China (English)

    1992-01-01

    920691 The determination of serum hepa-titis B virus DNA by polymerase chain rea-ction in hepatitis B patients treated withalpha-interferon. XU. Jianye(徐建业), et al.Centr Lab, Chongqing Cancer Instit, 630030.Chin J Intern Med, 1992; 31(5): 278-280. To clarify the status of HBV in serum of

  18. Effects of Dietary Lipid Source and Emulsifier on Growth Performance, Intestinal Digestive Enzyme Activities and Hepatic Biochemical Indexes of Bullfrog, Rana (Lithobates) catesbeiana%饲料脂肪源和乳化剂对牛蛙生长性能、肠道消化酶活力及肝脏生化指标的影响

    Institute of Scientific and Technical Information of China (English)

    冯伟; 王玲; 张春晓; 宋凯; 张璐

    2015-01-01

    本试验旨在研究以猪油或鱼油为脂肪源的饲料中添加乳化剂对牛蛙生长性能、肠道消化酶活力及肝脏生化指标的影响。试验采用2×2因子完全随机区组试验设计,2种脂肪源(猪油和鱼油)和2个乳化剂添加水平(0和300 mg/kg),配成4种等氮等能的试验饲料。将初始平均体重为(19.01±0.01) g的180只牛蛙随机分为4组,每组3个重复,每个重复15只蛙,进行为期8周的饲养试验。结果表明:未添加乳化剂的鱼油组增重率( WGR)和摄食量( FI)显著高于未添加乳化剂的猪油组( P0.05)。添加乳化剂的鱼油组的各生长性能指标与未添加乳化剂的鱼油组均无显著差异(P>0.05)。各组牛蛙胴体水分、粗蛋白质、粗脂肪和粗灰分含量差异不显著(P>0.05),而添加乳化剂的猪油组全体粗脂肪含量显著高于未添加乳化剂的猪油组( P0.05),饲料中添加乳化剂显著提高以猪油为主要脂肪源牛蛙的肠道脂肪酶活力(P0.05)。各组牛蛙肝脏过氧化氢酶( CAT)和超氧化物歧化酶( SOD)活力无显著差异( P>0.05)。在以猪油为主要脂肪源时,添加乳化剂显著提高肝脏丙二醛( MDA)含量( P0.05)。根据结果得出,在以猪油为主要脂肪源的饲料中添加乳化剂可提高牛蛙的生长性能。%The objective of this study was to investigate the effects of using lard oil or fish oil as main lipid source supplemented with emulsifier on growth performance, intestinal digestive enzyme activities and hepatic biochemical indexes of bullfrog, Rana (Lithobates) catesbeiana. A randomized complete block design with 2× 2 factors was adopted in the experiment. Four iso-nitrogenous and iso-energetic diets were formulated with diet-ary lipid source including pork lard and fish oil, and the supplemental level of emulsifier was 0 and 300 mg/kg, respectively. One hundred and eighty bullfrogs with the initial average body weight of ( 19. 01 ± 0.01) g were randomly assigned to 4

  19. Effects of cadium, zinc and lead on soil enzyme activities

    Institute of Scientific and Technical Information of China (English)

    YANG Zhi-xin; LIU Shu-qing; ZHENG Da-wei; FENG Sheng-dong

    2006-01-01

    Heavy metal (HM) is a major hazard to the soil-plant system. This study investigated the combined effects of cadium (Cd),zinc (Zn) and lead (Pb) on activities of four enzymes in soil, including calatase, urease, invertase and alkalin phosphatase. HM content in tops of canola and four enzymes activities in soil were analyzed at two months after the metal additions to the soil. Pb was not significantly inhibitory than the other heavy metals for the four enzyme activities and was shown to have a protective role on calatase activity in the combined presence of Cd, Zn and Pb; whereas Cd significantly inhibited the four enzyme activities, and Zn only inhibited urease and calatase activities. The inhibiting effect of Cd and Zn on urease and calatase activities can be intensified significantly by the additions of Zn and Cd. There was a negative synergistic inhibitory effect of Cd and Zn on the two enzymes in the presence of Cd, Zn and Pb. The urease activity was inhibited more by the HM combinations than by the metals alone and reduced approximately 20%-40% of urease activity. The intertase and alkaline phosphatase activities significantly decreased only with the increase of Cd concentration in the soil. It was shown that urease was much more sensitive to HM than the other enzymes. There was a obvious negative correlation between the ionic impulsion of HM in soil, the ionic impulsion of HM in canola plants tops and urease activity. It is concluded that the soil urease activity may be a sensitive tool for assessing additive toxic combination effect on soil biochemical parameters.

  20. Sensitive enzyme immunoassay for hepatitis B virus core-related antigens and their correlation to virus load.

    Science.gov (United States)

    Kimura, Tatsuji; Rokuhara, Akinori; Sakamoto, Yoko; Yagi, Shintaro; Tanaka, Eiji; Kiyosawa, Kendo; Maki, Noboru

    2002-02-01

    A sensitive enzyme immunoassay (EIA) specific for hepatitis B virus core antigen (HBcAg) and hepatitis B e antigen (HBeAg) was developed. We designated the precore/core gene products as hepatitis B virus (HBV) core-related antigens (HBcrAg). In order to detect HBcrAg even in anti-HBc/e antibody-positive specimens, the specimens were pretreated in detergents. The antibodies are inactivated by this pretreatment and, simultaneously, the antigens are released and the epitopes are exposed. The assay demonstrated 71 to 112% recovery using HBcrAg-positive sera. We observed no interference from the tested anticoagulants or blood components. When the cutoff value was tentatively set at 10(3) U/ml, all healthy control (HBsAg/HBV-DNA negative; n = 108) and anti-HCV antibody-positive (n = 59) sera were identified as negative. The assay showed a detection limit of 4 x 10(2) U/ml using recombinant antigen. Detection limits were compared in four serially diluted HBV high-titer sera. The HBcrAg assay demonstrated higher sensitivity than HBV-DNA transcription-mediated amplification (TMA) or HBeAg radio immunoassay (RIA) in the dilution test. HBcrAg concentrations correlated well with HBV-DNA TMA (r = 0.91, n = 29) and in-house real-time detection-PCR (r = 0.93, n = 47) in hepatitis B patients. On HBeAg/anti-HBe antibody seroconversion panels, the HBcrAg concentration changed in accordance with HBV-DNA levels. HBcrAg concentration provides a reflection of HBV virus load equivalent to HBV-DNA level, and the assay therefore offers a simple method for monitoring hepatitis B patients.

  1. Inhibition of existing denitrification enzyme activity by chloramphenicol

    Science.gov (United States)

    Brooks, M.H.; Smith, R.L.; Macalady, D.L.

    1992-01-01

    Chloramphenicol completely inhibited the activity of existing denitrification enzymes in acetylene-block incubations with (i) sediments from a nitrate-contaminated aquifer and (ii) a continuous culture of denitrifying groundwater bacteria. Control flasks with no antibiotic produced significant amounts of nitrous oxide in the same time period. Amendment with chloramphenicol after nitrous oxide production had begun resulted in a significant decrease in the rate of nitrous oxide production. Chloramphenicol also decreased (>50%) the activity of existing denitrification enzymes in pure cultures of Pseudomonas denitrificans that were harvested during log- phase growth and maintained for 2 weeks in a starvation medium lacking electron donor. Short-term time courses of nitrate consumption and nitrous oxide production in the presence of acetylene with P. denitrificans undergoing carbon starvation were performed under optimal conditions designed to mimic denitrification enzyme activity assays used with soils. Time courses were linear for both chloramphenicol and control flasks, and rate estimates for the two treatments were significantly different at the 95% confidence level. Complete or partial inhibition of existing enzyme activity is not consistent with the current understanding of the mode of action of chloramphenicol or current practice, in which the compound is frequently employed to inhibit de novo protein synthesis during the course of microbial activity assays. The results of this study demonstrate that chloramphenicol amendment can inhibit the activity of existing denitrification enzymes and suggest that caution is needed in the design and interpretation of denitrification activity assays in which chloramphenicol is used to prevent new protein synthesis.

  2. Hepatic Cytochrome P450 Activity, Abundance, and Expression Throughout Human Development.

    Science.gov (United States)

    Sadler, Natalie C; Nandhikonda, Premchendar; Webb-Robertson, Bobbie-Jo; Ansong, Charles; Anderson, Lindsey N; Smith, Jordan N; Corley, Richard A; Wright, Aaron T

    2016-07-01

    Cytochrome P450s are oxidative metabolic enzymes that play critical roles in the biotransformation of endogenous compounds and xenobiotics. The expression and activity of P450 enzymes varies considerably throughout human development; the deficit in our understanding of these dynamics limits our ability to predict environmental and pharmaceutical exposure effects. In an effort to develop a more comprehensive understanding of the ontogeny of P450 enzymes, we employed a multi-omic characterization of P450 transcript expression, protein abundance, and functional activity. Modified mechanism-based inhibitors of P450s were used as chemical probes for isolating active P450 proteoforms in human hepatic microsomes with developmental stages ranging from early gestation to late adult. High-resolution liquid chromatography-mass spectrometry was used to identify and quantify probe-labeled P450s, allowing for a functional profile of P450 ontogeny. Total protein abundance profiles and P450 rRNA was also measured, and our results reveal life-stage-dependent variability in P450 expression, abundance, and activity throughout human development and frequent discordant relationships between expression and activity. We have significantly expanded the knowledge of P450 ontogeny, particularly at the level of individual P450 activity. We anticipate that these results will be useful for enabling predictive therapeutic dosing, and for avoiding potentially adverse and harmful reactions during maturation from both therapeutic drugs and environmental xenobiotics. Copyright © 2016 by The American Society for Pharmacology and Experimental Therapeutics.

  3. Profiling Kinase Activity during Hepatitis C Virus Replication Using a Wortmannin Probe.

    Science.gov (United States)

    Desrochers, Geneviève F; Sherratt, Allison R; Blais, David R; Nasheri, Neda; Ning, Zhibin; Figeys, Daniel; Goto, Natalie K; Pezacki, John Paul

    2015-09-11

    To complete its life cycle, the hepatitis C virus (HCV) induces changes to numerous aspects of its host cell. As kinases act as regulators of many pathways utilized by HCV, they are likely enzyme targets for virally induced inhibition or activation. Herein, we used activity-based protein profiling (ABPP), which allows for the identification of active enzymes in complex protein samples and the quantification of their activity, to identify kinases that displayed differential activity in HCV-expressing cells. We utilized an ABPP probe, wortmannin-yne, based on the kinase inhibitor wortmannin, which contains a pendant alkyne group for bioconjugation using bioorthogonal chemistry. We observed changes in the activity of kinases involved in the mitogen-activated protein kinase pathway, apoptosis pathways, and cell cycle control. These results establish changes to the active kinome, as reported by wortmannin-yne, in the proteome of human hepatoma cells actively replicating HCV. The observed changes include kinase activity that affect viral entry, replication, assembly, and secretion, implying that HCV is regulating the pathways that it uses for its life cycle through modulation of the active kinome.

  4. Dynamic relationships between microbial biomass, respiration, inorganic nutrients and enzyme activities: informing enzyme based decomposition models

    Directory of Open Access Journals (Sweden)

    Daryl L Moorhead

    2013-08-01

    Full Text Available We re-examined data from a recent litter decay study to determine if additional insights could be gained to inform decomposition modeling. Rinkes et al. (2013 conducted 14-day laboratory incubations of sugar maple (Acer saccharum or white oak (Quercus alba leaves, mixed with sand (0.4% organic C content or loam (4.1% organic C. They measured microbial biomass C, carbon dioxide efflux, soil ammonium, nitrate, and phosphate concentrations, and β-glucosidase (BG, β-N-acetyl-glucosaminidase (NAG, and acid phosphatase (AP activities on days 1, 3, and 14. Analyses of relationships among variables yielded different insights than original analyses of individual variables. For example, although respiration rates per g soil were higher for loam than sand, rates per g soil C were actually higher for sand than loam, and rates per g microbial C showed little difference between treatments. Microbial biomass C peaked on day 3 when biomass-specific activities of enzymes were lowest, suggesting uptake of litter C without extracellular hydrolysis. This result refuted a common model assumption that all enzyme production is constitutive and thus proportional to biomass, and/or indicated that part of litter decay is independent of enzyme activity. The length and angle of vectors defined by ratios of enzyme activities (BG/NAG versus BG/AP represent relative microbial investments in C (length, and N and P (angle acquiring enzymes. Shorter lengths on day 3 suggested low C limitation, whereas greater lengths on day 14 suggested an increase in C limitation with decay. The soils and litter in this study generally had stronger P limitation (angles > 45˚. Reductions in vector angles to < 45˚ for sand by day 14 suggested a shift to N limitation. These relational variables inform enzyme-based models, and are usually much less ambiguous when obtained from a single study in which measurements were made on the same samples than when extrapolated from separate studies.

  5. Chimeric enzymes with improved cellulase activities

    Science.gov (United States)

    Xu, Qi; Baker, John O; Himmel, Michael E

    2015-03-31

    Nucleic acid molecules encoding chimeric cellulase polypeptides that exhibit improved cellulase activities are disclosed herein. The chimeric cellulase polypeptides encoded by these nucleic acids and methods to produce the cellulases are also described, along with methods of using chimeric cellulases for the conversion of cellulose to sugars such as glucose.

  6. Enzyme activities in mitochondria isolated from ripening tomato fruit.

    Science.gov (United States)

    Jeffery, D; Goodenough, P W; Weitzman, P D

    1986-09-01

    Mitochondria were isolated from tomato (Lycopersicon esculentum L.) fruit at the mature green, orange-green and red stages and from fruit artificially suspended in their ripening stage. The specific activities of citrate synthase (EC 4.1.3.7), malate dehydrogenase (EC 1.1.1.37), NAD-linked isocitrate dehydrogenase (EC 1.1.1.41) and NAD-linked malic enzyme (EC 1.1.1.38) were determined. The specific activities of all these enzymes fell during ipening, although the mitochondria were fully functional as demonstrated by the uptake of oxygen. The fall in activity of mitochondrial malate dehydrogenase was accompanied by a similar fall in the activity of the cytosolic isoenzyme. Percoll-purified mitochondria isolated from mature green fruit remained intact for more than one week and at least one enzyme, citrate synthase, did not exhibit the fall in specific activity found in normal ripening fruit.

  7. Dynamical changing pattems of glycogen and enzyme histochemical activities in rat liver graft undergoing warm ischemia injury

    Institute of Scientific and Technical Information of China (English)

    Xiao-Shun He; Yi Ma; Lin-Wei Wu; Jin-Lang Wu; Rui-De Hu; Gui-Hua Chen; Jie-Fu Huang

    2005-01-01

    AIM: To investigate the changing patterns of glycogen and enzyme histochemical activities in rat liver graft under a dif ferent warm ischemia time (WIT) and to predict the tolerant time limitation of the liver graft to warm ischemia injury.METHODS: The rats were randomized into five groups, WTT was 0, 15, 30, 45, 60 min, respectively, and histochemical staining of liver graft specimens was observed. The recovery changes of glycogen and enzyme histochemistry activities were measured respectively 6 and 24 h following liver graft implantation.RESULTS: The activities of succinic dehydrogenase, cytochrome oxidase, apyrase (Mg++-ATPase) and content of glycogen were decreased gradually after different WIT in a time-dependent manner. The changes were significant when WIT was over 30 min.CONCLUSION: Hepatic injury is reversible within 30 min of warm ischemia injury. Glycogen and enzyme histochemistry activities of liver grafts and their recovery potency after reperfusion may serve as criteria to evaluate the quality of liver grafts.

  8. Effect of Boswellia serrata supplementation on blood lipid, hepatic enzymes and fructosamine levels in type2 diabetic patients.

    Science.gov (United States)

    Ahangarpour, Akram; Heidari, Hamid; Fatemeh, Ramezani Ali Akbari; Pakmehr, Mostafa; Shahbazian, Hajeye; Ahmadi, Iraj; Mombeini, Zahra; Mehrangiz, Babadi Hajani

    2014-02-04

    Type 2 diabetes is an endocrine disorder that affects a large percentage of patients. High blood glucose causes fatty deposits in the liver which is likely to increase in SGOT and SGPT activities. Significant increase in SGOT/SGPT and low HDL levels is observed in patients with diabetes. Serum fructosamine concentration reflects the degree of blood glucose control in diabetic patients. This study was aimed to investigate the antidiabetic, hypolipidemic and hepatoprotective effects of supplementation of Boswellia serrata in type2 diabetic patients. 60 type 2 diabetic patients from both sexes (30 males and 30 females) were dedicated to the control and intervention groups (30 subjects per group). Boswellia serrata gum resin in amount of 900 mg daily for 6 weeks were orally administered (as three 300 mg doses) in intervention group and the control group did not receive anything. Blood samples were taken at the beginning of the study and after 6 weeks. Blood levels of fructosamine, lipid profiles as well as hepatic enzyme in type 2 diabetic patients were measured. Treatment of diabetic patient with Boswellia serrata was caused to significant increase in blood HDL levels as well as a remarkable decrease in cholesterol, LDL, fructosamine (p < 0.05) SGPT and SGOT levels after 6 weeks (p < 0.01). In spite of reduction of serum triglyceride, VLDL levels in intervention group, we did not detect a significant difference after 6 weeks. This study showed that Boswellia serrata supplementation can be beneficial in controlling blood parameters in patients with type 2 diabetes. Therefore, its use can be useful in patients with medicines.

  9. Catalytically active nanomaterials: a promising candidate for artificial enzymes.

    Science.gov (United States)

    Lin, Youhui; Ren, Jinsong; Qu, Xiaogang

    2014-04-15

    Natural enzymes, exquisite biocatalysts mediating every biological process in living organisms, are able to accelerate the rate of chemical reactions up to 10(19) times for specific substrates and reactions. However, the practical application of enzymes is often hampered by their intrinsic drawbacks, such as low operational stability, sensitivity of catalytic activity to environmental conditions, and high costs in preparation and purification. Therefore, the discovery and development of artificial enzymes is highly desired. Recently, the merging of nanotechnology with biology has ignited extensive research efforts for designing functional nanomaterials that exhibit various properties intrinsic to enzymes. As a promising candidate for artificial enzymes, catalytically active nanomaterials (nanozymes) show several advantages over natural enzymes, such as controlled synthesis in low cost, tunability in catalytic activities, as well as high stability against stringent conditions. In this Account, we focus on our recent progress in exploring and constructing such nanoparticulate artificial enzymes, including graphene oxide, graphene-hemin nanocomposites, carbon nanotubes, carbon nanodots, mesoporous silica-encapsulated gold nanoparticles, gold nanoclusters, and nanoceria. According to their structural characteristics, these enzyme mimics are categorized into three classes: carbon-, metal-, and metal-oxide-based nanomaterials. We aim to highlight the important role of catalytic nanomaterials in the fields of biomimetics. First, we provide a practical introduction to the identification of these nanozymes, the source of the enzyme-like activities, and the enhancement of activities via rational design and engineering. Then we briefly describe new or enhanced applications of certain nanozymes in biomedical diagnosis, environmental monitoring, and therapeutics. For instance, we have successfully used these biomimetic catalysts as colorimetric probes for the detection of

  10. Optimization to Low Temperature Activity in Psychrophilic Enzymes

    Directory of Open Access Journals (Sweden)

    Caroline Struvay

    2012-09-01

    Full Text Available Psychrophiles, i.e., organisms thriving permanently at near-zero temperatures, synthesize cold-active enzymes to sustain their cell cycle. These enzymes are already used in many biotechnological applications requiring high activity at mild temperatures or fast heat-inactivation rate. Most psychrophilic enzymes optimize a high activity at low temperature at the expense of substrate affinity, therefore reducing the free energy barrier of the transition state. Furthermore, a weak temperature dependence of activity ensures moderate reduction of the catalytic activity in the cold. In these naturally evolved enzymes, the optimization to low temperature activity is reached via destabilization of the structures bearing the active site or by destabilization of the whole molecule. This involves a reduction in the number and strength of all types of weak interactions or the disappearance of stability factors, resulting in improved dynamics of active site residues in the cold. Considering the subtle structural adjustments required for low temperature activity, directed evolution appears to be the most suitable methodology to engineer cold activity in biological catalysts.

  11. Influence of Piper betle on hepatic marker enzymes and tissue antioxidant status in ethanol-treated Wistar rats.

    Science.gov (United States)

    Saravanan, R; Prakasam, A; Ramesh, B; Pugalendi, K V

    2002-01-01

    Piper betle L. is a commonly used masticatory in Asia. This study was carried out to investigate the hepatoprotective and antioxidant properties of P. betle, using ethanol intoxication as a model of hepatotoxic and oxidative damage. Ethanol-treated rats exhibited elevation of hepatic marker enzymes and disturbances in antioxidant defense when compared with normal rats. Oral administration of P. betle extract (100, 200, or 300 mg/kg body weight) for 30 days significantly (P betle extract (300 mg/kg body weight) was most effective. The results were comparable with the known hepatoprotective drug, silymarin. These results indicate that P. betle could afford a significant hepatoprotective and antioxidant effect.

  12. Enzyme activity in forest peat soils

    OpenAIRE

    Błońska, Ewa

    2010-01-01

    The aim of the study was to determine the activity of dehydrogenases and urease in forest peat soils of different fertility. There were selected 23 experimental plots localised in central and northern Poland. The research was conducted on forest fens, transition bogs and raised bogs. The biggest differences in soil physical and chemical properties were detected between fen and raised bog soils while raised bog soils and transition bog soils differed the least. Statistically significant dif...

  13. Curcumin attenuates diet-induced hepatic steatosis by activating AMP-activated protein kinase.

    Science.gov (United States)

    Um, Min Young; Hwang, Kwang Hyun; Ahn, Jiyun; Ha, Tae Youl

    2013-09-01

    Curcumin is a well-known component of traditional turmeric (Curcuma longa), which has been reported to prevent obesity and diabetes. However, the effect of curcumin on hepatic lipid metabolism remains unclear. The aim of this study was to examine the effects of curcumin on hepatic steatosis in high-fat/cholesterol diet (HFD)-induced obese mice. Male C57BL/6J mice were fed a normal diet (ND), HFD or HFD with 0.15% curcumin (HFD+C) for 11 weeks. We found that curcumin significantly lowered the body-weight and adipose tissue weight of mice in the HFD+C group compared with the findings for the HFD group (p cholesterol, fasting glucose and insulin in serum were decreased, and HFD-induced impairment of insulin sensitivity was improved by curcumin supplementation (p Curcumin protected against the development of hepatic steatosis by reducing hepatic fat accumulation. Moreover, curcumin activated AMP-activated protein kinase (AMPK) and elevated the gene expression of peroxisome proliferator-activated receptor alpha. By contrast, curcumin suppressed the HFD-mediated increases in sterol regulatory element-binding protein-1, acetyl-CoA carboxylase 1, fatty acid synthase and cluster of differentiation 36 expression. Taken together, these findings indicate that curcumin attenuates HFD-induced hepatic steatosis by regulating hepatic lipid metabolism via AMPK activation, suggesting its use as a therapeutic for hepatic steatosis.

  14. Effect of Commiphora mukul gum resin on hepatic marker enzymes, lipid peroxidation and antioxidants status in pancreas and heart of streptozotocin induced diabetic rats

    Institute of Scientific and Technical Information of China (English)

    Ramesh B; Karuna R; Sreenivasa Reddy S; Haritha K; Sai Mangala D; Sasi Bhusana Rao B; Saralakumari D

    2012-01-01

    Objective: To study the antioxidant efficacy of Commiphora mukul (C. mukul) gum resin ethanolic extract in streptozotocin (STZ) induced diabetic rats. Methods: The male Wistar albino rats were randomly divided into four groups of eight animals each: Control group (C), CM-treated control group (C+CMEE), Diabetic control group (D), CM- treated diabetic group (D+CMEE). Diabetes was induced by intraperitoneal injection of STZ (55 mg/kg/ bwt). After being confirmed the diabetic rats were treated with C. mukul gum resin ethanolic extract (CMEE) for 60 days. The biochemical estimations like antioxidant, oxidative stress marker enzymes and hepatic marker enzymes of tissues were performed. Results: The diabetic rats showed increased level of enzymatic activities aspartate aminotransaminase (AST), alanine aminotransaminase (ALT) in liver and kidney and oxidative markers like lipid peroxidation (LPO) and protein oxidation (PO) in pancreas and heart. Antioxidant enzyme activities were significantly decreased in the pancreas and heart compared to control group. Administration of CMEE (200 mg/kg bw) to diabetic rats for 60 days significantly reversed the above parameters towards normalcy. Conclusions: In conclusion, our data indicate the preventive role of C. mukul against STZ-induced diabetic oxidative stress; hence this plant could be used as an adjuvant therapy for the prevention and/or management of diabetes and aggravated antioxidant status.

  15. Improving Activity of Salt-Lyophilized Enzymes in Organic Media

    Science.gov (United States)

    Borole, Abhijeet P.; Davison, Brian H.

    Lyophilization with salts has been identified as an important method of activating enzymes in organic media. Using salt-activated enzymes to transform molecules tethered to solid surfaces in organic phase requires solubilization of enzymes in the solvents. Methods of improving performance of salt-lyophilized enzymes, further, via chemical modification, and use of surfactants and surfactants to create fine emulsions prior to lyophilization are investigated. The reaction system used is transesterification of N-acetyl phenylalanine ethyl ester with methanol or propanol. Initial rate of formation of amino acid esters by subtilisin Carlsberg (SC) was studied and found to increase two to sevenfold by either chemical modification or addition of surfactants in certain solvents, relative to the salt (only)-lyophilized enzyme. The method to prepare highly dispersed enzymes in a salt-surfactant milieu also improved activity by two to threefold. To test the effect of chemical modification on derivatization of drug molecules, acylation of bergenin was investigated using chemically modified SC.

  16. [Effects of starvation on digestive enzyme activities of Monopterus albus].

    Science.gov (United States)

    Yang, Dai-qin; Chen, Fang; Ruan, Guo-liang; Hu, Cheng-wen; Cao, Sheng-huan

    2007-05-01

    Starvation is a major environmental stress, which has a broad effect on the physiology and ecology of aquatic animals. In this study, Monopterus albus was starved for 30 days at (20 +/- 0.5) degrees C, and the activities of protease, trypsin, amylase and lipase in its digestive organs were measured on the 0, 3rd, 5th, 10th, 15th, 20th, and 30th day of starvation. The results showed that starvation had definite effects on the activities of all test enzymes. With the prolongation of starvation, the activities of test enzymes decreased, which was most significant when the fish was starved for 5-10 days. After 10 days of starvation, the decreasing trend of the enzyme activities became less obvious.

  17. Extraction of Active Enzymes from "Hard-to-Break-Cells"

    DEFF Research Database (Denmark)

    Ottaviani, Alessio; Tesauro, Cinzia; Fjelstrup, S

    We present the utilization of a rolling circle amplification (RCA) based assay to investigate the extraction efficiency of active enzymes from a class of “hard-to-break” cells, yeast Saccaramyces cerevisiae. Current analyses of microorganisms, such as pathogenic bacteria, parasites or particular...... life stages of microorganisms (e.g. spores from bacteria or fungi) is hampered by the lack of efficient lysis protocols that preserve the activity and integrity of the cellular content. Presented herein is a flexible scheme to screen lysis protocols for active enzyme extraction. We also report a gentle...... yet effective approach for extraction of active enzymes by entrapping cells in microdroplets. Combined effort of optimized extraction protocols and effective analytical approaches is expected to generate impact in future disease diagnosis and environmental safety....

  18. Glyphosate on digestive enzymes activity in piava (Leporinus obtusidens

    Directory of Open Access Journals (Sweden)

    Joseânia Salbego

    2014-09-01

    Full Text Available The effects of glyphosate, a nonselective herbicide (1.0 or 5.0mg L-1 on digestive enzymes activity (stomach and intestine were evaluated in juveniles of piava (Leporinus obtusidens after 90 days of exposure. The activity of acid protease, trypsin, chymotrypsin and amylase increased with the increase of glyphosate concentration. These results indicate that glyphosate affects digestive enzyme activities in this species, and may be an indicator of poor nutrient availability when fish survive in herbicide-contaminated water.

  19. Hepatitis

    Institute of Scientific and Technical Information of China (English)

    2005-01-01

    2005226 Characteristics of peripheral blood T lymphocyte subsets in hepatitis B patients. FAN Zhen-ping(范振平),et al. Center Bio Ther, Instit Infect Dis, 302 Hosp Chin PLA, Beijing 100039. World Chin J Digestol, 2005;13(2): 194-197. Objective: To characterize the T-lymphocyte subsets in peripheral blood of patients with acute and chronic hepatitis B, and to explore their relations with the disease state. Methods: Peripheral blood

  20. Lipid peroxidation and antioxidant enzymes activity in avian semen.

    Science.gov (United States)

    Partyka, Agnieszka; Lukaszewicz, Ewa; Niżański, Wojciech

    2012-10-01

    The present study compared the antioxidant system and lipid peroxidation in semen of two avian species: chicken and goose. The experiment was conducted on Greenleg Partridge roosters and White Koluda(®) ganders, each represented by 10 mature males. Malondialdehyde (MDA) concentration, catalase (CAT), glutathione peroxidase (GPx) and superoxide dismutase (SOD) activities were determined in sperm cells and seminal plasma. In gander spermatozoa, the amount of MDA was 10 times greater (Pantioxidant enzymes had greater (Pactivity in goose than chicken sperm. Catalase activity was detected in seminal plasma and spermatozoa from both studied species for the first time. In seminal plasma, the activity of GPx was two times greater (Pactivity was less (Pactivity of antioxidant defense and LPO. The greater amount of lipid peroxidation and greater activity of antioxidant enzymes in goose semen might suggest that spermatozoa were under greater oxidative stress and the enzymes were not utilized for the protection of functionally and structurally impaired cells. In turn, in fresh chicken semen a lesser activity of antioxidant enzymes accompanied with a lesser lipid peroxidation amount and good semen quality could indicate that fowl spermatozoa were under oxidative stress, but the enzymes were employed to protect and maintain sperm quality. Copyright © 2012 Elsevier B.V. All rights reserved.

  1. Serum arylesterase and paraoxonase activity in patients with chronic hepatitis

    Institute of Scientific and Technical Information of China (English)

    Suleyman Sirri Kilic; Suleyman Aydin; Nermin Kilic; Fazilet Erman; Suna Aydin; (I)lhami Celik

    2005-01-01

    AIM: To investigate the relationship between serum paraoxonase (PON1), AST, ALT, GGT, and arylesterase (AE) activity alterations and the degree of liver damage in patients with chronic hepatitis.METHODS: We studied 34 chronic hepatitis patients and 32 control subjects, aged between 35 and 65 years,in the Department of Infection and Clinical Microbiology at the Firat University School of Medicine. Blood samples were collected from subjects between 8:00 and 10:00 a.m. following a 12-h fast. Baseline and salt-stimulated PON1 activities were measured by the hydrolysis of paraoxon. Phenyl acetate was used as the substrate and formed phenol was measured spectrophotometrically at 270 nm after the addition of a 10-fold diluted serum sample in AE activity measurements.RESULTS: The results of this investigation revealed that the levels of AE activity decreased from 132±52 to 94±36 (29%), baseline PON1 activity from 452±112 to 164±67 (64%), salt-stimulated PON1 activity from 746±394 to 294±220 (61%), HDL from 58.4±5.1 to 47.2±5.6(20%), triglyceride from 133±51.2 to 86±34.0 (35%),while a slight increase in the level of LDL (from 163±54.1 to 177.3±56.0; 9%) and significant increases in the levels of AST (from 29±9.3 to 98±44), ALP (from 57.2±13.1 to 91±38.1), ALT (from 27.9±3.32 to 89±19.1), GGT (from 24.3±2.10 to 94±48.2), total bilirubin (from 0.74±0.02 to 1.36±0.06; 84%) and direct bilirubin (from 0.18±0.01 to 0.42±0.04; 133%) were detected.However, the levels of albumin, total protein, cholesterol,and uric acid were almost the same in chronic hepatitis and the control subjects.CONCLUSION: Low PON1 and AE activity may contribute to the increased liver dysfunction in chronic hepatitis patients by reducing the ability of HDL to retard LDL oxidation and might be clinically useful for monitoring the disease of chronic hepatitis.

  2. Extraction of Active Enzymes from "Hard-to-Break-Cells"

    DEFF Research Database (Denmark)

    Ottaviani, Alessio; Tesauro, Cinzia; Fjelstrup, S

    life stages of microorganisms (e.g. spores from bacteria or fungi) is hampered by the lack of efficient lysis protocols that preserve the activity and integrity of the cellular content. Presented herein is a flexible scheme to screen lysis protocols for active enzyme extraction. We also report a gentle...

  3. Soil Enzyme Activities under Agroforestry Systems in Northern Jiangsu Province

    Institute of Scientific and Technical Information of China (English)

    Wan Fuxu; Chen Ping

    2004-01-01

    The authors presented the enzyme characteristics of catalase, sucrase, urease and alkaline phosphatase under agroforestry systems in northern Jiangsu Province. The results show that soil enzyme activities reduce gradually from top to bottom layer of the soil profile, and the fluctuations of catalase and urease are smaller than those of sucrase and alkaline phosphatase. Soil enzyme activities differe significantly in different samples, and the order is arranged as poplar-crop intercropping segment (A, D) > paulownia-crop intercropping segment (B, C) > CK. Furthermore, soil enzyme activities increase with intercropping age. On the other hand, in the same plot, there are closer relationships between enzymes in the soil samples. Catalase, alkaline phosphatase and urease are negatively related, while alkaline phosphatase and urease are positively related (except in samples B and C). In addition, the enzyme activities have a close relationship with the fertilizers. Catalase is positively correlated with the soil pH value (r = 0.854, 0.804, 0.078 and 0.082, respectively), and is negatively correlated with total N (r = -0.201, -0.529, -0.221 and -0.821, respectively), total P (r = -0.143, -0.213, -0.362 and -0.751, respectively) and available P (r = -0.339, -0.351, -0.576, and -0.676, respectively). Sucrase, urease and alkaline phosphatase are negatively correlated with the pH value, while positively correlated with the other fertilizers (r ≈ 1). The authors suggest that enzyme activity will be a great potential as an indicator of soil quality.

  4. Antioxidant enzymes activities in obese Tunisian children

    Directory of Open Access Journals (Sweden)

    Sfar Sonia

    2013-01-01

    Full Text Available Abstract Background The oxidant stress, expected to increase in obese adults, has an important role in the pathogenesis of many diseases. It results when free radical formation is greatly increased or protective antioxidant mechanisms are compromised. The main objective of this study is to evaluate the antioxidant response to obesity-related stress in healthy children. Methods A hundred and six healthy children (54 obese and 52 controls, aged 6–12 years old, participated in this study. The collected data included anthropometric measures, blood pressure, fasting glucose, total cholesterol, triglycerides and enzymatic antioxidants (Superoxide dismutase: SOD, Catalase: CAT and Glutathione peroxidase: GPx. Results The first step antioxidant response, estimated by the SOD activity, was significantly higher in obese children compared with normal-weight controls (p  Conclusions The obesity-related increase of the oxidant stress can be observed even in the childhood period. In addition to the complications of an increased BMI, obesity itself can be considered as an independent risk factor of free radical production resulting in an increased antioxidant response.

  5. Marked inhibition of hepatic cytochrome P450 activity in cholesterol-induced atherosclerosis in rabbits.

    Science.gov (United States)

    Irizar, A; Ioannides, C

    1998-04-03

    The objective of the present study was to investigate the expression of major xenobiotic-metabolising cytochrome P450 proteins, and of other enzyme systems, in hepatic and extrahepatic tissues of rabbits rendered atherosclerotic by the dietary administration of 1% cholesterol diets for 8 weeks. Individual cytochrome P450 proteins were monitored using diagnostic substrates and immunologically in Western blot analysis. The activity of all hepatic isoforms studied was depressed in the atherosclerotic animals; when, however, apoprotein levels were determined immunologically, no major differences were evident between the control and the atherosclerotic rabbits. In vitro studies indicated that neither cholesterol nor palm oil inhibited cytochrome P450 activity. The effects of cholesterol treatment leading to atherosclerosis on kidney, heart and lung cytochrome P450 activities were isoform- and tissue-specific; no change was evident in the heart activities, but in the lung and kidney cytochrome P450 activities were clearly modulated by the treatment with cholesterol. Apoprotein levels did not always parallel the changes in activities. Western blot analysis of aortic cytochromes P450 revealed that administration of cholesterol-rich diets enhanced CYP2B and CYP3A apoprotein levels. Cholesterol feeding to rabbits gave rise to a marked decrease in hepatic glutathione S-transferase activity but did not influence glutathione reductase or total glutathione levels. The same treatment had no effect on catalase, glutathione peroxidase and superoxide dismutase. It is concluded that treatment of rabbits with cholesterol-rich diets leading to atherosclerosis gives rise to profound changes in the expression of cytochrome P450 proteins in the liver and other tissues; possible mechanisms are discussed.

  6. Determination of Intestinal Enzyme Activities During Infancy Period

    Directory of Open Access Journals (Sweden)

    Emel Örün

    2016-08-01

    Full Text Available Introduction: Intestinal enzyme activities are indirect indicators that reflect the existence and metabolic activity of bacteria living in the intestinal flora. The purpose of the study was to measure fecal beta (β-glucuronidase, β-glucosidase and urease enzyme activities and to determine the factors that affect levels in 6 week old and 8 month old babies. Materials and Methods: The study comprised 100 healthy infants at 6 weeks of age. Feces samples were collected from all infants. However, 17 of the feces samples were not included due to the lack of particles in the feces. The same samples were also taken from 35 infants at 8 months of age. Twenty-five of the infants had given feces samples at both 6 weeks and 8 months of age. Urease, β-glucuronidase and β-glucosidase enzyme activities (nmol/min-1/mg-protein-1 were measured. Results: In repeated measures, the levels of β-glucuronidase and urease declined over time and β-glucosidase levels increased. At 8 months of age, higher β-glucuronidase levels were obtained in premature infants. At 6 weeks of age, lower levels of urease were measured in babies who were started breastfeeding at the first hour of life and were bottle-fed. Exclusive breastfeeding had no influence on the intestinal enzyme activities. Conclusions: In early infancy period when microflora is structured, intestinal enzyme activities are important that show indirectly functionality of the microflora. However, it is difficult to highlight what affects the levels of intestinal enzymes because activities vary according to the age.

  7. Chemoproteomic profiling of host and pathogen enzymes active in cholera.

    Science.gov (United States)

    Hatzios, Stavroula K; Abel, Sören; Martell, Julianne; Hubbard, Troy; Sasabe, Jumpei; Munera, Diana; Clark, Lars; Bachovchin, Daniel A; Qadri, Firdausi; Ryan, Edward T; Davis, Brigid M; Weerapana, Eranthie; Waldor, Matthew K

    2016-04-01

    Activity-based protein profiling (ABPP) is a chemoproteomic tool for detecting active enzymes in complex biological systems. We used ABPP to identify secreted bacterial and host serine hydrolases that are active in animals infected with the cholera pathogen Vibrio cholerae. Four V. cholerae proteases were consistently active in infected rabbits, and one, VC0157 (renamed IvaP), was also active in human choleric stool. Inactivation of IvaP influenced the activity of other secreted V. cholerae and rabbit enzymes in vivo, and genetic disruption of all four proteases increased the abundance of intelectin, an intestinal lectin, and its binding to V. cholerae in infected rabbits. Intelectin also bound to other enteric bacterial pathogens, suggesting that it may constitute a previously unrecognized mechanism of bacterial surveillance in the intestine that is inhibited by pathogen-secreted proteases. Our work demonstrates the power of activity-based proteomics to reveal host-pathogen enzymatic dialog in an animal model of infection.

  8. Chemoproteomic profiling of host and pathogen enzymes active in cholera

    Science.gov (United States)

    Hatzios, Stavroula K.; Hubbard, Troy; Sasabe, Jumpei; Munera, Diana; Clark, Lars; Bachovchin, Daniel A.; Qadri, Firdausi; Ryan, Edward T.; Davis, Brigid M.; Weerapana, Eranthie; Waldor, Matthew K.

    2016-01-01

    Activity-based protein profiling (ABPP) is a chemoproteomic tool for detecting active enzymes in complex biological systems. We used ABPP to identify secreted bacterial and host serine hydrolases that are active in animals infected with the cholera pathogen Vibrio cholerae. Four V. cholerae proteases were consistently active in infected rabbits, and one, VC0157 (renamed IvaP), was also active in human cholera stool. Inactivation of IvaP influenced the activity of other secreted V. cholerae and rabbit enzymes in vivo, while genetic disruption of all four proteases increased the abundance and binding of an intestinal lectin—intelectin—to V. cholerae in infected rabbits. Intelectin also bound to other enteric bacterial pathogens, suggesting it may constitute a previously unrecognized mechanism of bacterial surveillance in the intestine that is inhibited by pathogen-secreted proteases. Our work demonstrates the power of activity-based proteomics to reveal host-pathogen enzymatic dialogue in an animal model of infection. PMID:26900865

  9. Evolutionary transitions in enzyme activity of ant fungus gardens.

    Science.gov (United States)

    De Fine Licht, Henrik H; Schiøtt, Morten; Mueller, Ulrich G; Boomsma, Jacobus J

    2010-07-01

    Fungus-growing (attine) ants and their fungal symbionts passed through several evolutionary transitions during their 50 million year old evolutionary history. The basal attine lineages often shifted between two main cultivar clades, whereas the derived higher-attine lineages maintained an association with a monophyletic clade of specialized symbionts. In conjunction with the transition to specialized symbionts, the ants advanced in colony size and social complexity. Here we provide a comparative study of the functional specialization in extracellular enzyme activities in fungus gardens across the attine phylogeny. We show that, relative to sister clades, gardens of higher-attine ants have enhanced activity of protein-digesting enzymes, whereas gardens of leaf-cutting ants also have increased activity of starch-digesting enzymes. However, the enzyme activities of lower-attine fungus gardens are targeted primarily toward partial degradation of plant cell walls, reflecting a plesiomorphic state of nondomesticated fungi. The enzyme profiles of the higher-attine and leaf-cutting gardens appear particularly suited to digest fresh plant materials and to access nutrients from live cells without major breakdown of cell walls. The adaptive significance of the lower-attine symbiont shifts remains unclear. One of these shifts was obligate, but digestive advantages remained ambiguous, whereas the other remained facultative despite providing greater digestive efficiency.

  10. Hydrophobic Core Flexibility Modulates Enzyme Activity in HIV-1 Protease

    Energy Technology Data Exchange (ETDEWEB)

    Mittal, Seema; Cai, Yufeng; Nalam, Madhavi N.L.; Bolon, Daniel N.A.; Schiffer, Celia A. (UMASS, MED)

    2012-09-11

    Human immunodeficiency virus Type-1 (HIV-1) protease is crucial for viral maturation and infectivity. Studies of protease dynamics suggest that the rearrangement of the hydrophobic core is essential for enzyme activity. Many mutations in the hydrophobic core are also associated with drug resistance and may modulate the core flexibility. To test the role of flexibility in protease activity, pairs of cysteines were introduced at the interfaces of flexible regions remote from the active site. Disulfide bond formation was confirmed by crystal structures and by alkylation of free cysteines and mass spectrometry. Oxidized and reduced crystal structures of these variants show the overall structure of the protease is retained. However, cross-linking the cysteines led to drastic loss in enzyme activity, which was regained upon reducing the disulfide cross-links. Molecular dynamics simulations showed that altered dynamics propagated throughout the enzyme from the engineered disulfide. Thus, altered flexibility within the hydrophobic core can modulate HIV-1 protease activity, supporting the hypothesis that drug resistant mutations distal from the active site can alter the balance between substrate turnover and inhibitor binding by modulating enzyme activity.

  11. Does diet influence salivary enzyme activities in elephant species?

    Science.gov (United States)

    Boehlke, Carolin; Pötschke, Sandra; Behringer, Verena; Hannig, Christian; Zierau, Oliver

    2017-01-01

    Asian elephants (Elephas maximus) and African elephants (Loxodonta africana) are herbivore generalists; however, Asian elephants might ingest a higher proportion of grasses than Africans. Although some studies have investigated nutrition-specific morphological adaptations of the two species, broader studies on salivary enzymes in both elephant species are lacking. This study focuses on the comparison of salivary enzymes activity profiles in the two elephant species; these enzymes are relevant for protective and digestive functions in humans. We aimed to determine whether salivary amylase (sAA), lysozyme (sLYS), and peroxidase (sPOD) activities have changed in a species-specific pattern during evolutionary separation of the elephant genera. Saliva samples of 14 Asian and eight African elephants were collected in three German zoos. Results show that sAA and sLYS are salivary components of both elephant species in an active conformation. In contrast, little to no sPOD activity was determined in any elephant sample. Furthermore, sAA activity was significantly higher in Asian compared with African elephants. sLYS and sPOD showed no species-specific differences. The time of food provision until sample collection affected only sAA activity. In summary, the results suggest several possible factors modulating the activity of the mammal-typical enzymes, such as sAA, sLYS, and sPOD, e.g., nutrition and sampling procedure, which have to be considered when analyzing differences in saliva composition of animal species.

  12. A DNA enzyme with N-glycosylase activity

    Science.gov (United States)

    Sheppard, T. L.; Ordoukhanian, P.; Joyce, G. F.

    2000-01-01

    In vitro evolution was used to develop a DNA enzyme that catalyzes the site-specific depurination of DNA with a catalytic rate enhancement of about 10(6)-fold. The reaction involves hydrolysis of the N-glycosidic bond of a particular deoxyguanosine residue, leading to DNA strand scission at the apurinic site. The DNA enzyme contains 93 nucleotides and is structurally complex. It has an absolute requirement for a divalent metal cation and exhibits optimal activity at about pH 5. The mechanism of the reaction was confirmed by analysis of the cleavage products by using HPLC and mass spectrometry. The isolation and characterization of an N-glycosylase DNA enzyme demonstrates that single-stranded DNA, like RNA and proteins, can form a complex tertiary structure and catalyze a difficult biochemical transformation. This DNA enzyme provides a new approach for the site-specific cleavage of DNA molecules.

  13. De novo active sites for resurrected Precambrian enzymes

    Science.gov (United States)

    Risso, Valeria A.; Martinez-Rodriguez, Sergio; Candel, Adela M.; Krüger, Dennis M.; Pantoja-Uceda, David; Ortega-Muñoz, Mariano; Santoyo-Gonzalez, Francisco; Gaucher, Eric A.; Kamerlin, Shina C. L.; Bruix, Marta; Gavira, Jose A.; Sanchez-Ruiz, Jose M.

    2017-07-01

    Protein engineering studies often suggest the emergence of completely new enzyme functionalities to be highly improbable. However, enzymes likely catalysed many different reactions already in the last universal common ancestor. Mechanisms for the emergence of completely new active sites must therefore either plausibly exist or at least have existed at the primordial protein stage. Here, we use resurrected Precambrian proteins as scaffolds for protein engineering and demonstrate that a new active site can be generated through a single hydrophobic-to-ionizable amino acid replacement that generates a partially buried group with perturbed physico-chemical properties. We provide experimental and computational evidence that conformational flexibility can assist the emergence and subsequent evolution of new active sites by improving substrate and transition-state binding, through the sampling of many potentially productive conformations. Our results suggest a mechanism for the emergence of primordial enzymes and highlight the potential of ancestral reconstruction as a tool for protein engineering.

  14. Hepatic glucuronidation of resveratrol: interspecies comparison of enzyme kinetic profiles in human, mouse, rat, and dog.

    Science.gov (United States)

    Maier-Salamon, Alexandra; Böhmdorfer, Michaela; Thalhammer, Theresia; Szekeres, Thomas; Jaeger, Walter

    2011-01-01

    The enzyme kinetic profiles of the formation of resveratrol-3-O-glucuronide (R3G) and resveratrol-4'-O-glucuronide (R4'G) by liver microsomes from humans, dogs, and rodents were investigated. Glucuronidation by human and dog liver microsomes to R3G and R4'G occurred for about 65% of applied resveratrol, and was significantly reduced to 10% when substrate concentration was increased 10-fold. In contrast, rodent microsomes glucuronidated about 90% of applied resveratrol independently of substrate concentration. Furthermore, in mouse and rat liver microsomes, resveratrol was almost exclusively conjugated at position 3, whereas human and dog livers also glucuronidated resveratrol at position 4' (ratio R3G:R4'G = 5:1). Interspecies differences were also found when calculating the enzyme kinetic profiles of both conjugates. Formation of R4'G in human and dog microsomes followed Michaelis-Menten kinetics, while R3G showed substrate inhibition at higher resveratrol concentrations. In mouse and rat microsomes, however, both R3G and R4'G formation exhibited auto-activation kinetics. Formation of R3G and R4'G by recombinant UGT1A1 also showed substrate inhibition kinetics that led to decreased intrinsic clearance values, while UGT1A9-catalyzed glucuronidation demonstrated substrate inhibition kinetics at position 3 and Hill kinetics for the formation of R4'G. In conclusion, resveratrol glucuronidation exhibited species-dependent differences, with the dog as the animal model that most closely represents humans in terms of this process.

  15. Regulation of eNOS enzyme activity by posttranslational modification.

    Science.gov (United States)

    Heiss, Elke H; Dirsch, Verena M

    2014-01-01

    The regulation of endothelial NO synthase (eNOS) employs multiple different cellular control mechanisms impinging on level and activity of the enzyme. This review aims at summarizing the current knowledge on the posttranslational modifications of eNOS, including acylation, nitrosylation, phosphorylation, acetylation, glycosylation and glutathionylation. Sites, mediators and impact on enzyme localization and activity of the single modifications will be discussed. Moreover, interdependence, cooperativity and competition between the different posttranslational modifications will be elaborated with special emphasis on the susceptibility of eNOS to metabolic cues.

  16. Enzyme Activities in Perfluorooctanoic Acid (PFOA)-Polluted Soils

    Institute of Scientific and Technical Information of China (English)

    ZHANG Wei; LIN Kuang-Fei; YANG Sha-Sha; ZHANG Meng

    2013-01-01

    Perfluorooctanoic acid (PFOA) is a popular additive of the chemical industry; its effect on activities of important soil enzymes is not well understood.A laboratory incubation experiment was carried out to analyze the PFOA-induced changes in soil urease,catalase,and phosphatase activities.During the entire incubation period,the activities of the three soil enzymes generally declined with increasing PFOA concentration,following certain dose-response relationships.The values of EC10,the contaminant concentration at which the biological activity is inhibited by 10%,of PFOA for the soil enzyme activity calculated from the modeling equation of the respective dose-response curve suggested a sensitivity order of phosphatase > catalase > urease.The effect of PFOA on soil enzyme activities provided a basic understanding of the eco-toxicological effect of PFOA in the environment.Results of this study supported using soil phosphatase as a convenient biomarker for ecological risk assessment of PFOA-polluted soils.

  17. Influence of long-term fertilization on soil enzyme activities

    Directory of Open Access Journals (Sweden)

    Alina Dora SAMUEL

    2009-05-01

    Full Text Available Soil enzyme activities (actual and potential dehydrogenase, catalase, acid and alkaline phosphatase were determined in the 0–10, 10–20, and 20–30 cm layers of a brown luvic soil submitted to a complex fertilization experiment with different types of green manure. It was found that each activity decreased with increasing sampling depth. It should be emphasized that greenmanuring of maize led to a significant increase in each of the five enzymatic activities determined. The enzymatic indicators of soil quality calculated from the values of enzymatic activities showed the order: lupinus + rape + oat > lupinus > vetch + oat + ryegrass > lupinus + oat + vetch > unfertilized plot. This order means that by determination of enzymatic activities valuable information can be obtained regarding fertility status of soils. There were significant correlations of soil enzyme activities with chemical properties.

  18. Roles of the Lipid Metabolism in Hepatic Stellate Cells Activation

    Institute of Scientific and Technical Information of China (English)

    Xin-yan Jing; Xue-feng Yang; Kai Qing; Yan Ou-Yang

    2013-01-01

    The lipids present in hepatic stellate cells (HSCs) lipid droplets include retinyl ester, triglyceride, cholesteryl ester, cholesterol, phospholipids and free fatty acids. Activation of HSCs is crucial to the development of fibrosis in liver disease. During activation, HSCs transform into myofibroblasts with concomitant loss of their lipid droplets and production of excessive extracellular matrix. Release of lipid droplets containing retinyl esters and triglyceride is a defining feature of activated HSCs. Accumulating evidence supports the proposal that recovering the accumulation of lipids would inhibit the activation of HSCs. In healthy liver, quiescent HSCs store 80%of total liver retinols and release them depending on the extracellular retinol status. However, in injured liver activated HSCs lose their retinols and produce a considerable amount of extracellular matrix, subsequently leading to liver fibrosis. Further findings prove that lipid metabolism of HSCs is closely associated with its activation, yet relationship between activated HSCs and the lipid metabolism has remained mysterious.

  19. Effects of dietary inulin, statin, and their co-treatment on hyperlipidemia, hepatic steatosis and changes in drug-metabolizing enzymes in rats fed a high-fat and high-sucrose diet

    Directory of Open Access Journals (Sweden)

    Sugatani Junko

    2012-03-01

    Full Text Available Abstract Background Rats fed a high-fat and high-sucrose (HF diet develop hepatic steatosis and hyperlipidemia. There are several reports that a change in nutritional status affects hepatic levels of drug-metabolizing enzymes. Synthetic inulin is a dietary component that completely evades glucide digestion. Supplementing a HF diet with inulin ameliorates hypertriglycemia and hepatic steatosis, but not hypercholesterolemia. This study aimed at distinguishing the effects of synthetic inulin and 3-hydroxy-3-methylglutaryl coenzyme A reductase inhibitor (statin, which inhibit cholesterol biosynthesis. Methods We examined effects of co-treatment with synthetic inulin (5% and fluvastatin (0, 4, and 8 mg/kg, per os on body weight, epidydimal white adipose tissue weight, serum and hepatic lipid profiles, and hepatic cytochrome P450 (CYP mRNA and protein profiles in rats fed a standard diet or a HF diet for 3 weeks. Results Treatment with the synthetic inulin (5% or fluvastatin at 4 mg/kg (lethal dose in rats fed the HF diet, 8 mg/kg ameliorated the elevation in hepatic triacylglycerol and total cholesterol levels in rats fed the HF diet. Whereas co-treatment with the inulin (5% and fluvastatin (4 mg/kg had a tendency to more strongly suppress the elevation in serum levels of very low density lipoprotein triacylglycerol than either treatment alone, no additive or synergistic effect was found in decrease in hepatic lipid levels. Hepatic levels of CYP1A1/2 and CYP2E1 mRNA and protein and methoxyresorufin O-demethylase and ethoxyresorufin O-deethylase activities were reduced in rats fed the HF diet. The synthetic inulin alleviated the reduction in hepatic levels of CYP1A1/2 and CYP2E1 mRNA and protein more strongly than fluvastatin, and no synergistic effects were observed on co-treatment. Furthermore, hepatic levels of aryl hydrocarbon receptor mRNA were decreased in rats fed the HF diet and recovered to near normal values with the intake of dietary inulin

  20. 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

    Energy Technology Data Exchange (ETDEWEB)

    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.

  1. [Enzyme activity of an actinomycete producer of carotenes and macrotetrolides].

    Science.gov (United States)

    Nefelova, M V; Sverdlova, A N

    1982-01-01

    The activity of pyruvate dehydrogenase and dehydrogenases of the tricarboxylic acid cycle was assayed in the mycelium of Streptomyces chrysomallus var. Carotenoides growing under different conditions of the medium. The activity of the enzymes increased when acetic, citric and succinic acids were added at different periods of the growth. Moreover, addition of the acids increased the time of intensive functioning of the dehydrogenases whose activity abruptly decreased after 60 h of the growth under the control conditions.

  2. Effects of hepatic enzyme inducers on thyroxine (T4) catabolism in primary rat hepatocytes

    Science.gov (United States)

    Nuclear receptor agonists such as phenobarbital (PB), 2,2',4,4',5,5'-hexachlorobiphenyl (PCB 153), and 3-methylcholantrene (3-MC) decrease circulating thyroxine (T4) concentrations in rats. It is suspected that this decrease occurs through the induction of hepatic metabolizing en...

  3. Early feeding to modify digestive enzyme activity in broiler chickens

    Directory of Open Access Journals (Sweden)

    Milagro León T.

    2014-09-01

    Full Text Available Objective. To evaluate the effect on digestive enzyme activity in broiler chickens by providing food in the first 48 hrs. after birth. Materials and methods. After incubating 300 fertile eggs from Hubbard breeding and immediately after hatching, the chicks were randomly assigned to treatments: fasting (from hatching to 48 hrs.; Hydrated Balanced Food (HBF from birth to 48 hrs.; commercial hydrating supplement (CHS from birth to 48 hrs. The diets were provided ad libitum. After 48 hrs. a commercial diet was fed. At birth and at 48 and 72 hrs. of age 30 chicks/treatment were sacrificed to determine the enzyme activity of maltase, sucrase, alkaline phosphatase, phytase, a-amylase, trypsin and lipase in samples of duodenal or pancreatic homogenate. Results. The supply of HBF or CHS during the first 48 hrs. of life increased the activity of maltase, sucrase and phytase in the first 3 days of life, with values between 1.2 and up to 4-fold compared to the control (p<0.05. Chickens that fasted for the first 48 hrs. had higher activity of the pancreatic enzymes a-amylase, trypsin, and lipase at 72 hrs. of life (p<0.05. Conclusions. The food supply in the first 48 hrs. after hatching increases the duodenal enzyme activity in the intestinal brush border during the first 3 days of age in broiler chickens.

  4. Relationship between somatostatin receptors and activation of hepatic stellate cells

    Institute of Scientific and Technical Information of China (English)

    潘勤; 李定国; 陆汉明; 陆良勇; 尤汉宁; 徐芹芳

    2004-01-01

    Background Somafostatin receptors (SSTRs) have been suggested to involve in mediating the effect of somatostatin on hepatic stellate cells (HSCs) in an activation-dependent way. We, therefore, try to investigate the relationship between expression of SSTRs and activation of rat HSCs.Methods HSCs were isolated from rats by in situ perfusion and single-step density gradient centrifugation.SSTR1-5 mRNA levels in the differentiated first passage HSCs were detected by means of a reverse transcription polymerase chain reaction. On the other hand, hepatic fibrosis was induced in adult male Sprague-Dawley rats by carbon tetrachloride intoxication, and the expression of SSTR1-5 in normal as well as fibrotic livers was measured by immunohistochemical staining.Results SSTR mRNA and SSTR could not be found in freshly isolated rat HSCs or normal rat liver. However, SSTR1-3 mRNA appeared as HSCs became wholly activated, and could also be identified on the membrane of activated HSCs in the perisinusoid space, fibrous septa, etc.Conclusion The expression of SSTR1-3 in the rat HSC is closely related to its activation. This may reflect one of the main negative regulation mechanisms in the course of HSC activation.

  5. Identification of glyoxalase 1 polymorphisms associated with enzyme activity.

    Science.gov (United States)

    Peculis, Raitis; Konrade, Ilze; Skapare, Elina; Fridmanis, Davids; Nikitina-Zake, Liene; Lejnieks, Aivars; Pirags, Valdis; Dambrova, Maija; Klovins, Janis

    2013-02-15

    The glyoxalase system and its main enzyme, glyoxalase 1 (GLO1), protect cells from advanced glycation end products (AGEs), such as methylglyoxal (MG) and other reactive dicarbonyls, the formation of which is increased in diabetes patients as a result of excessive glycolysis. MG is partly responsible for harmful protein alterations in living cells, notably in neurons, leading to their dysfunction, and recent studies have shown a negative correlation between GLO1 expression and tissue damage. Neuronal dysfunction is a common diabetes complication due to elevated blood sugar levels, leading to high levels of AGEs. The aim of our study was to determine whether single nucleotide polymorphisms (SNPs) in the GLO1 gene influence activity of the enzyme. In total, 125 healthy controls, 101 type 1 diabetes, and 100 type 2 diabetes patients were genotyped for three common SNPs, rs2736654 (A111E), rs1130534 (G124G), and rs1049346 (5'-UTR), in GLO1. GLO1 activity was determined in whole blood lysates for all participants of the study. Our results showed a significant association between the minor alleles rs1130534 and rs1049346 and decreased enzyme activity (P=0.001 and P=2.61×10(-5), respectively). Increased allelic counts of the risk alleles were strongly associated with decreased GLO1 activity (standardised β=-0.24, P=2.15×10(-5)), indicating independent actions of these variants on GLO1 activity, as supported by the haplotype analysis. We showed for the first time an association between genetic variants with GLO1 enzyme activity in humans. SNPs in GLO1 can be used to predict enzyme activity and detoxifying capabilities, but further studies are needed to link these SNPs with common complications in diabetes.

  6. Evaluation of cytochrome P450 activity in vitro, using dermal and hepatic microsomes from four species and two keratinocyte cell lines in culture.

    Science.gov (United States)

    Rolsted, Kamilla; Kissmeyer, Anne-Marie; Rist, Gerda Marie; Hansen, Steen Honoré

    2008-01-01

    The Cytochrome P450 (CYP450) enzymes are expressed in the skin, and despite a low activity, as compared to the hepatic counterpart, a role during transdermal delivery of a drug cannot be excluded. Additionally, the enzymes may play a role in local toxicity, and further knowledge of dermal CYP450 activity can contribute to elucidate this issue. To achieve this, a cocktail of six selective CYP450 probe substrates were incubated with dermal and hepatic microsomes isolated from mouse, rat, minipig and man. Different species were used to evaluate if a reliable substitute for human tissue was possible. Further, the hepatic microsomes were included in this study, to estimate if the hepatic CYP450 activity is predictive of dermal CYP450 activity. The CYP450 activity was determined in two keratinocyte cell lines as well, as this in vitro model is desirable due to the ease in handling, among other factors. Overall, the metabolism found in the dermal microsomes was very low, and major differences were observed between species. When comparing the activities in dermal and hepatic microsomes, the qualitative pattern was to some extent similar within species, but also a number of differences were observed. The CYP450 metabolic activity in the two keratinocyte cell lines was not comparable to metabolism in the human dermal microsomes.

  7. Activity of Krebs cycle enzymes in mdx mice.

    Science.gov (United States)

    Comim, Clarissa M; Hoepers, Andreza; Ventura, Letícia; Freiberger, Viviane; Dominguini, Diogo; Mina, Francielle; Mendonça, Bruna P; Scaini, Giselli; Vainzof, Mariz; Streck, Emílio L; Quevedo, João

    2016-01-01

    Duchenne muscular dystrophy (DMD) is a degenerative disease of skeletal, respiratory, and cardiac muscles caused by defects in the dystrophin gene. More recently, brain involvement has been verified. Mitochondrial dysfunction and oxidative stress may underlie the pathophysiology of DMD. In this study we evaluate Krebs cycle enzymes activity in the cerebral cortex, diaphragm, and quadriceps muscles of mdx mice. Cortex, diaphragm, and quadriceps tissues from male dystrophic mdx and control mice were used. We observed increased malate dehydrogenase activity in the cortex; increased malate dehydrogenase and succinate dehydrogenase activities in the diaphragm; and increased citrate synthase, isocitrate dehydrogenase, and malate dehydrogenase activities in the quadriceps of mdx mice. This study showed increased activity of Krebs cycle enzymes in cortex, quadriceps, and diaphragm in mdx mice. © 2015 Wiley Periodicals, Inc.

  8. Fluorogenic Substrates for Visualizing Acidic Organelle Enzyme Activities.

    Directory of Open Access Journals (Sweden)

    Fiona Karen Harlan

    Full Text Available Lysosomes are acidic cytoplasmic organelles that are present in all nucleated mammalian cells and are involved in a variety of cellular processes including repair of the plasma membrane, defense against pathogens, cholesterol homeostasis, bone remodeling, metabolism, apoptosis and cell signaling. Defects in lysosomal enzyme activity have been associated with a variety of neurological diseases including Parkinson's Disease, Lysosomal Storage Diseases, Alzheimer's disease and Huntington's disease. Fluorogenic lysosomal staining probes were synthesized for labeling lysosomes and other acidic organelles in a live-cell format and were shown to be capable of monitoring lysosomal metabolic activity. The new targeted substrates were prepared from fluorescent dyes having a low pKa value for optimum fluorescence at the lower physiological pH found in lysosomes. They were modified to contain targeting groups to direct their accumulation in lysosomes as well as enzyme-cleavable functions for monitoring specific enzyme activities using a live-cell staining format. Application to the staining of cells derived from blood and skin samples of patients with Metachromatic Leukodystrophy, Krabbe and Gaucher Diseases as well as healthy human fibroblast and leukocyte control cells exhibited localization to the lysosome when compared with known lysosomal stain LysoTracker® Red DND-99 as well as with anti-LAMP1 Antibody staining. When cell metabolism was inhibited with chloroquine, staining with an esterase substrate was reduced, demonstrating that the substrates can be used to measure cell metabolism. When applied to diseased cells, the intensity of staining was reflective of lysosomal enzyme levels found in diseased cells. Substrates specific to the enzyme deficiencies in Gaucher or Krabbe disease patient cell lines exhibited reduced staining compared to that in non-diseased cells. The new lysosome-targeted fluorogenic substrates should be useful for research

  9. Effects of selenium dietary enhancement on hatchery-reared coho salmon, Oncorhynchus kisutch (Walbaum), when compared with wild coho: hepatic enzymes and seawater adaptation evaluated.

    Science.gov (United States)

    Felton, S.P.; Landolt, M.L.; Grace, R.; Palmisano, A.N.

    1996-01-01

    Hatchery-reared coho salmon, Oncorhynchus kisutch (Walbaum), were fed elevated levels of selenium (as Na2SeO3) to raise eviscerated body burdens to the level measured in wild counterparts. The goal was to find a dietary concentration that would achieve the desired effect without causing damage to growth and normal development. To measure some indices of health, the detoxifying enzymes chosen were hepatic glutathione peroxidase (GSH-Px) and hepatic superoxide dismutase (SOD). Eviscerated body selenium (Se) concentration, GSH-Px and SOD levels were measured during and at the end of the 9 month freshwater feeding trial. Selenium retention and enzyme activity were also measured during 6 months’residence in sea water (SW). Selenium supplements were added to a commercial ration to give final concentrations of 1.1, 8.6, 11.1, 13.6 μg g-1 Se in the four respective diets. The results indicated that a dietary concentration of 8.6 μg g-1selenium was capable of inducing eviscerated body burdens similar to those found in wild fish. The elevated selenium levels persisted throughout the freshwater (FW) rearing phase, but declined when the fish were fed an unsupplemented ration upon SW entry. Superoxide dismutase levels did not increase above control levels. Glutathione peroxidase levels increased in fish fed the supplemented diets. GSH-Px activity declined in the higher supplemented dietary groups when all groups were reduced to the control group level of 1.1 μg g-1. Cumulative mortality in SW was 20% in fish fed either the 1.1 or the 8.6 μg g-1 Se diets. The 8.6 μg g-1 Se supplemented diets did produce healthy coho, comparable to their wild counterparts.

  10. 蛋氨酸钴对珍珠龙胆石斑鱼幼鱼生长、矿物元素沉积及肝脏酶活力的影响%Effects of dietary cobalt methionine on growth performance, mineral deposition, and hepatic enzyme activities in juvenile pearl gentian grouper (Epinephelus lanceolatus♂ ×E. fuscoguttatus♀)

    Institute of Scientific and Technical Information of China (English)

    刘云; 王际英; 李宝山; 乔洪金; 柳旭东; 郝甜甜; 王晓艳

    2016-01-01

    在基础饲料中添加不同梯度的蛋氨酸钴(CoMet),制成钴含量分别为0.30 mg/kg、1.75 mg/kg、3.42 mg/kg、6.73 mg/kg、12.56 mg/kg、25.50 mg/kg的6组实验饲料(记作D1、D2、D3、D4、D5、D6组),饲喂初始体重(60.02±0.42) g的珍珠龙胆石斑鱼(Epinephelus lanceolatu♂×E. fuscoguttatus♀)幼鱼8周,研究蛋氨酸钴对其生长、矿物元素沉积和肝脏酶活力的影响。结果表明,随饲料中蛋氨酸钴含量的增加,实验鱼增重率(WGR)和特定生长率(SGR)均先升后降, D3组显著高于其他组(P0.05);粗蛋白含量呈降低的趋势, D5、D6组显著低于D1~D4组(P0.05);粗脂肪含量先降后升, D4组显著低于其他组(P0.05)。本实验条件下,以SGR为评价指标,经折线回归分析,珍珠龙胆石斑鱼幼鱼对钴的最适需求量为3.25 mg/kg,即53.28 mg(CoMet)/kg(饲料)。%Cobalt (Co) is one of the essential elements for fish, with various biological functions, including in growth promotion, blood production, reproduction, and immunity. In this study, we evaluated the effects of dietary cobalt me-thionine (CoMet) on growth performance, mineral deposition, and hepatic enzyme activities in the juvenile pearl gen-tian grouper (Epinephelus lanceolatus♂ ×E. fuscoguttatus♀). Six extruded isonitrogenous and isoenergetic diets (51% crude protein and 13% crude lipid) were formulated by supplementing the basal diet with CoMet, providing actual Co contents of 0.30, 1.75, 3.42, 6.73, 12.56, or 25.50 mg/kg diet, which were fed to groups designated D1, D2, D3, D4, D5, and D6, respectively. Each diet was randomly assigned to triplicate groups of 20 juveniles (initial weight, 60.02±0.42 g). The experiment was conducted for 8 weeks in plastic cylindrical tanks in an indoor recirculated feeding system. The weight gain rate (WGR) and specific growth rate (SGR) of the juveniles both increased and then decreased with higher CoMet levels, and both were significantly higher in the D3 group than in

  11. Synthesis and study on biological activity of nitrogen-containing heterocyclic compounds – regulators of enzymes of nucleic acid biosynthesis

    Directory of Open Access Journals (Sweden)

    Alexeeva I. V.

    2013-07-01

    Full Text Available Results of investigations on the development of new regulators of functional activity of nucleic acid biosynthesis enzymes based on polycyclic nitrogen-containing heterosystems are summarized. Computer design and molecular docking in the catalytic site of target enzyme (T7pol allowed to perform the directed optimization of basic structures. Several series of compounds were obtained and efficient inhibitors of herpes family (simple herpes virus type 2, Epstein-Barr virus, influenza A and hepatitis C viruses were identified, as well as compounds with potent antitumor, antibacterial and antifungal activity. It was established that the use of model test systems based on enzymes participating in nucleic acids synthesis is a promising approach to the primary screening of potential inhibitors in vitro.

  12. Enzyme activities by indicator of quality in organic soil

    Science.gov (United States)

    Raigon Jiménez, Mo; Fita, Ana Delores; Rodriguez Burruezo, Adrián

    2016-04-01

    The analytical determination of biochemical parameters, as soil enzyme activities and those related to the microbial biomass is growing importance by biological indicator in soil science studies. The metabolic activity in soil is responsible of important processes such as mineralization and humification of organic matter. These biological reactions will affect other key processes involved with elements like carbon, nitrogen and phosphorus , and all transformations related in soil microbial biomass. The determination of biochemical parameters is useful in studies carried out on organic soil where microbial processes that are key to their conservation can be analyzed through parameters of the metabolic activity of these soils. The main objective of this work is to apply analytical methodologies of enzyme activities in soil collections of different physicochemical characteristics. There have been selective sampling of natural soils, organic farming soils, conventional farming soils and urban soils. The soils have been properly identified conserved at 4 ° C until analysis. The enzyme activities determinations have been: catalase, urease, cellulase, dehydrogenase and alkaline phosphatase, which bring together a representative group of biological transformations that occur in the soil environment. The results indicate that for natural and agronomic soil collections, the values of the enzymatic activities are within the ranges established for forestry and agricultural soils. Organic soils are generally higher level of enzymatic, regardless activity of the enzyme involved. Soil near an urban area, levels of activities have been significantly reduced. The vegetation cover applied to organic soils, results in greater enzymatic activity. So the quality of these soils, defined as the ability to maintain their biological productivity is increased with the use of cover crops, whether or spontaneous species. The practice of cover based on legumes could be used as an ideal choice

  13. Hepatitis B antigen in hepatocytes of chronic active liver disease.

    Science.gov (United States)

    Kawanishi, H

    1979-04-01

    To study the morphologic interrelation of hepatocytes with the replication of hepatitis B vius (HBV) and immunocompetent cells in chronic active liver disease(CALD), organ cultures were prepared from liver biopsy specimens. Replication of hepatitis B core antigen (HBcAg) appears to occur in the nucleus of the hepatocyte in close association with intranuclear electron-dense strands and sometimes intranucleolar matrixes (likely HBcAg genomes), and cytoplasmic maturation of the HBcAg takes place in the preautolytic condition of host hepatocytes. Immunocompetent cells became progressively autolyzed in the early period of cultures. No difference in progression of hepatocyte injury in tissues from normal subjects and from hepatitis B surface antigen (HBsAg)-positive and HBsAg-negative patients with CALD may suggest that intracellular synthesis of HBV alone is not cytopathic to host hepatocytes. This model is promising for the study of HBV replication and development, and also for testing the efficacy of new antiviral agents against the virus.

  14. Branched chain amino acid transaminase and branched chain alpha-ketoacid dehydrogenase activity in the brain, liver and skele­tal muscle of acute hepatic failure rats

    Directory of Open Access Journals (Sweden)

    Takei,Nobuyuki

    1985-02-01

    Full Text Available Branched chain amino acid (BCAA transaminase activity increased in both the mitochondrial and supernatant fractions of brain from hepatic failure rats, in which a partial hepatectomy was performed 24h following carbon tetrachloride (CCl4 administration, although the activity of liver and skeletal muscle was the same as in control rats. The elevation of mitochondrial BCAA transaminase activity in liver-injured rats was partly due to increased activity of brain specific Type III isozyme. Branched chain alpha-ketoacid (BCKA dehydrogenase in the brain homogenates was not significantly altered in acute hepatic failure rats, while the liver enzyme activity was markedly diminished. BCKA dehydrogenase activity in the brain homogenates was inhibited by adding ATP to the assay system, and was activated in vitro by preincubating the brain homogenate at 37 degrees C for 15 min. These findings suggest that brain BCAA catabolism is accelerated in acute hepatic failure rats.

  15. RELATIONSHIP BETWEEN SOMATOSTATIN RECEPTORS AND ACTIVATION OF HEPATIC STELLATE CELL

    Institute of Scientific and Technical Information of China (English)

    潘勤; 李定国; 陆汉明; 尤汉宁; 徐芹芳; 陆良勇

    2004-01-01

    Objective To investigate the relationship between expression of somatostatin receptors (SSTRs) and activation of rat hepatic stellate cell (HSC). Methods HSCs were isolated from rats by in situ perfusion and single-step density gradient centrifugation, and then SSTR1 ~5 mRNA levels in the differentiated first passage HSCs were detected by means of reverse transcription polymerase chain reaction. On the other hand, hepatic fibrosis was induced in adult male Sprague-Dawley rats by carbon tetrachloride intoxication, and the expression of SSTR1 ~5 in normal as well as fibrotic liver was measured by immunohistochemical staining. Results SSTR mRNA and SSTR could not be found in freshly isolated rat HSCs and normal rat liver. But SSTR1~3 mRNA appeared as HSCs became wholly activated, and SSTR1 ~3 could also be identified on the membrane of activated HSCs in the perisinusoid space, fibrous septa, etc Conclusion The expression of SSTR1~3 in the rat HSC is closely related to its activation. This may reflect one of the main negative regulation mechanisms in the course of HSC activation.

  16. EVOLUTIONARY TRANSITIONS IN ENZYME ACTIVITY OF ANT FUNGUS GARDENS

    DEFF Research Database (Denmark)

    De Fine Licht, Henrik H; Schiøtt, Morten; Mueller, Ulrich G

    2010-01-01

    Fungus-growing (attine) ants and their fungal symbionts passed through several evolutionary transitions during their 50 million year old evolutionary history. The basal attine lineages often shifted between two main cultivar clades, whereas the derived higher-attine lineages maintained an associa......Fungus-growing (attine) ants and their fungal symbionts passed through several evolutionary transitions during their 50 million year old evolutionary history. The basal attine lineages often shifted between two main cultivar clades, whereas the derived higher-attine lineages maintained...... an association with a monophyletic clade of specialized symbionts. In conjunction with the transition to specialized symbionts, the ants advanced in colony size and social complexity. Here we provide a comparative study of the functional specialization in extracellular enzyme activities in fungus gardens across...... the attine phylogeny. We show that, relative to sister clades, gardens of higher-attine ants have enhanced activity of protein-digesting enzymes, whereas gardens of leaf-cutting ants also have increased activity of starch-digesting enzymes. However, the enzyme activities of lower-attine fungus gardens...

  17. Variation in Soil Enzyme Activities in a Temperate Agroforestry Watershed

    Science.gov (United States)

    Integration of agroforestry and grass buffers into row crop watersheds improves overall environmental quality, including soil quality. The objective of this study was to examine management and landscape effects on soil carbon, soil nitrogen, microbial diversity, enzyme activity, and DNA concentrati...

  18. [Interaction between CYP450 enzymes and metabolism of traditional Chinese medicine as well as enzyme activity assay].

    Science.gov (United States)

    Lu, Tu-lin; Su, Lian-lin; Ji, De; Gu, Wei; Mao, Chun-qin

    2015-09-01

    Drugs are exogenous compounds for human bodies, and will be metabolized by many enzymes after administration. CYP450 enzyme, as a major metabolic enzyme, is an important phase I drug metabolizing enzyme. In human bodies, about 75% of drug metabolism is conducted by CYP450 enzymes, and CYP450 enzymes is the key factor for drug interactions between traditional Chinese medicine( TCM) -TCM, TCM-medicine and other drug combination. In order to make clear the interaction between metabolic enzymes and TCM metabolism, we generally chose the enzymatic activity as an evaluation index. That is to say, the enhancement or reduction of CYP450 enzyme activity was used to infer the inducing or inhibitory effect of active ingredients and extracts of traditional Chinese medicine on enzymes. At present, the common method for measuring metabolic enzyme activity is Cocktail probe drugs, and it is the key to select the suitable probe substrates. This is of great significance for study drug's absorption, distribution, metabolism and excretion (ADME) process in organisms. The study focuses on the interaction between TCMs, active ingredients, herbal extracts, cocktail probe substrates as well as CYP450 enzymes, in order to guide future studies.

  19. Construction of chimeric enzymes out of maize endosperm branching enzymes I and II: activity and properties.

    Science.gov (United States)

    Kuriki, T; Stewart, D C; Preiss, J

    1997-11-14

    Branching enzyme I and II isoforms from maize endosperm (mBE I and mBE II, respectively) have quite different properties, and to elucidate the domain(s) that determines the differences, chimeric genes consisting of part mBE I and part mBE II were constructed. When expressed under the control of the T7 promoter in Escherichia coli, several of the chimeric enzymes were inactive. The only fully active chimeric enzyme was mBE II-I BspHI, in which the carboxyl-terminal part of mBE II was exchanged for that of mBE I at a BspHI restriction site and was purified to homogeneity and characterized. Another chimeric enzyme, mBE I-II HindIII, in which the amino-terminal end of mBE II was replaced with that of mBE I, had very little activity and was only partially characterized. The purified mBE II-I BspHI exhibited higher activity than wild-type mBE I and mBE II when assayed by the phosphorylase a stimulation assay. mBE II-I BspHI had substrate specificity (preference for amylose rather than amylopectin) and catalytic capacity similar to mBE I, despite the fact that only the carboxyl terminus was from mBE I, suggesting that the carboxyl terminus may be involved in determining substrate specificity and catalytic capacity. In chain transfer experiments, mBE II-I BspHI transferred more short chains (with a degree of polymerization of around 6) in a fashion similar to mBE II. In contrast, mBE I-II HindIII transferred more long chains (with a degree of polymerization of around 11-12), similar to mBE I, suggesting that the amino terminus of mBEs may play a role in the size of oligosaccharide chain transferred. This study challenges the notion that the catalytic centers for branching enzymes are exclusively located in the central portion of the enzyme; it suggests instead that the amino and carboxyl termini may also be involved in determining substrate preference, catalytic capacity, and chain length transfer.

  20. Regulation of human hepatic drug transporter activity and expression by diesel exhaust particle extract.

    Directory of Open Access Journals (Sweden)

    Marc Le Vee

    Full Text Available Diesel exhaust particles (DEPs are common environmental air pollutants primarily affecting the lung. DEPs or chemicals adsorbed on DEPs also exert extra-pulmonary effects, including alteration of hepatic drug detoxifying enzyme expression. The present study was designed to determine whether organic DEP extract (DEPe may target hepatic drug transporters that contribute in a major way to drug detoxification. Using primary human hepatocytes and transporter-overexpressing cells, DEPe was first shown to strongly inhibit activities of the sinusoidal solute carrier (SLC uptake transporters organic anion-transporting polypeptides (OATP 1B1, 1B3 and 2B1 and of the canalicular ATP-binding cassette (ABC efflux pump multidrug resistance-associated protein 2, with IC50 values ranging from approximately 1 to 20 μg/mL and relevant to environmental exposure situations. By contrast, 25 μg/mL DEPe failed to alter activities of the SLC transporter organic cation transporter (OCT 1 and of the ABC efflux pumps P-glycoprotein and bile salt export pump (BSEP, whereas it only moderately inhibited those of sodium taurocholate co-transporting polypeptide and of breast cancer resistance protein (BCRP. Treatment by 25 μg/mL DEPe was next demonstrated to induce expression of BCRP at both mRNA and protein level in cultured human hepatic cells, whereas it concomitantly repressed mRNA expression of various transporters, including OATP1B3, OATP2B1, OCT1 and BSEP. Such changes in transporter expression were found to be highly correlated to those caused by 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD, a reference activator of the aryl hydrocarbon receptor (AhR pathway. This suggests that DEPe, which is enriched in known ligands of AhR like polycyclic aromatic hydrocarbons, alters drug transporter expression via activation of the AhR cascade. Taken together, these data established human hepatic transporters as targets of organic chemicals containing in DEPs, which may contribute

  1. Anti-hepatitis B virus active constituents from Swertia chirayita.

    Science.gov (United States)

    Zhou, Ning-Jia; Geng, Chang-An; Huang, Xiao-Yan; Ma, Yun-Bao; Zhang, Xue-Mei; Wang, Ju-Le; Chen, Ji-Jun

    2015-01-01

    Four new compounds swertiachiralatone A (1), swertiachoside A (2), swertiachirdiol A (3) and swertiachoside B (4), together with twenty-six known ones were isolated from the ethanol extract of Swertia chirayita. Their structures were elucidated by extensive spectroscopic analyses (1D- and 2D-NMR, HRESIMS, UV, IR and [α]D). All compounds were evaluated for anti-hepatitis B virus (anti-HBV) activities on HepG 2.2.15 cells line in vitro, of which compounds 14 and 19 showed inhibitory activity on hepatitis B surface antigen (HBsAg) secretion with IC50 values of 0.31 ± 0.045 and 1.49 ± 0.033 mM; compounds 14 and 28 exhibited activity against hepatitis B e antigen (HBeAg) secretion with IC50 values of 0.77 ± 0.076 and 5.92 ± 1.02 mM; and eight compounds (8,9,13,14,24-26,29) possessed activity against HBV DNA replication with IC50 values of 0.07-0.33 mM. In particular (+)-cycloolivil-4'-O-β-d-glucopyranoside (14) exhibited inhibition not only on the secretions of HBsAg and HBeAg with IC50 values of 0.31 ± 0.045 mM (SI=4.29) and 0.77 ± 0.076 mM (SI=1.75), respectively, but also on HBV DNA replication with an IC50 value of 0.29 ± 0.034 mM (SI=4.66).

  2. Potential enzyme activities in cryoturbated organic matter of arctic soils

    Science.gov (United States)

    Schnecker, J.; Wild, B.; Rusalimova, O.; Mikutta, R.; Guggenberger, G.; Richter, A.

    2012-12-01

    An estimated 581 Gt organic carbon is stored in arctic soils that are affected by cryoturbtion, more than in today's atmosphere (450 Gt). The high amount of organic carbon is, amongst other factors, due to topsoil organic matter (OM) that has been subducted by freeze-thaw processes. This cryoturbated OM is usually hundreds to thousands of years old, while the chemical composition remains largely unaltered. It has therefore been suggested, that the retarded decomposition rates cannot be explained by unfavourable abiotic conditions in deeper soil layers alone. Since decomposition of soil organic material is dependent on extracellular enzymes, we measured potential and actual extracellular enzyme activities in organic topsoil, mineral subsoil and cryoturbated material from three different tundra sites, in Zackenberg (Greenland) and Cherskii (North-East Siberia). In addition we analysed the microbial community structure by PLFAs. Hydrolytic enzyme activities, calculated on a per gram dry mass basis, were higher in organic topsoil horizons than in cryoturbated horizons, which in turn were higher than in mineral horizons. When calculated on per gram carbon basis, the activity of the carbon acquiring enzyme exoglucanase was not significantly different between cryoturbated and topsoil organic horizons in any of the three sites. Oxidative enzymes, i.e. phenoloxidase and peroxidase, responsible for degradation of complex organic substances, showed higher activities in topsoil organic and cryoturbated horizons than in mineral horizons, when calculated per gram dry mass. Specific activities (per g C) however were highest in mineral horizons. We also measured actual cellulase activities (by inhibiting microbial uptake of products and without substrate addition): calculated per g C, the activities were up to ten times as high in organic topsoil compared to cryoturbated and mineral horizons, the latter not being significantly different. The total amount of PLFAs, as a proxy for

  3. Hepatitis

    Institute of Scientific and Technical Information of China (English)

    2008-01-01

    2008449 A cross-sectional survey of occult hepatitis B virus infection in HIV-infected patients. MA Jianxin(马建新), et al.Dept Infect Dis, Shanghai Public Health Clin Center, Shanghai 201508. Chin J Intern Med 2008;47(7):574-577. Objective To assess the prevalence of occult HBV infection in HIV-infected patients.

  4. Carotenoid-cleavage activities of crude enzymes from Pandanous amryllifolius.

    Science.gov (United States)

    Ningrum, Andriati; Schreiner, Matthias

    2014-11-01

    Carotenoid degradation products, known as norisoprenoids, are aroma-impact compounds in several plants. Pandan wangi is a common name of the shrub Pandanus amaryllifolius. The genus name 'Pandanus' is derived from the Indonesian name of the tree, pandan. In Indonesia, the leaves from the plant are used for several purposes, e.g., as natural colorants and flavor, and as traditional treatments. The aim of this study was to determine the cleavage of β-carotene and β-apo-8'-carotenal by carotenoid-cleavage enzymes isolated from pandan leaves, to investigate dependencies of the enzymatic activities on temperature and pH, to determine the enzymatic reaction products by using Headspace Solid Phase Microextraction Gas Chromatography/Mass Spectrophotometry (HS-SPME GC/MS), and to investigate the influence of heat treatment and addition of crude enzyme on formation of norisoprenoids. Crude enzymes from pandan leaves showed higher activity against β-carotene than β-apo-8'-carotenal. The optimum temperature of crude enzymes was 70°, while the optimum pH value was 6. We identified β-ionone as the major volatile reaction product from the incubations of two different carotenoid substrates, β-carotene and β-apo-8'-carotenal. Several treatments, e.g., heat treatment and addition of crude enzymes in pandan leaves contributed to the norisoprenoid content. Our findings revealed that the crude enzymes from pandan leaves with carotenoid-cleavage activity might provide a potential application, especially for biocatalysis, in natural-flavor industry.

  5. Descriptive and predictive assessment of enzyme activity and enzyme related processes in biorefinery using IR spectroscopy and chemometrics

    DEFF Research Database (Denmark)

    Baum, Andreas

    the understanding of the structural properties of the extracted pectin. Secondly, enzyme kinetics of biomass converting enzymes was examined in terms of measuring enzyme activity by spectral evolution profiling utilizing FTIR. Chemometric multiway methods were used to analyze the tensor datasets enabling the second......-order calibration advantage (reference Theory of Analytical chemistry). As PAPER 3 illustrates the method is universally applicable without the need of any external standards and was exemplified by performing quantitative enzyme activity determinations for glucose oxidase, pectin lyase and a cellolytic enzyme blend...... (Celluclast 1.5L). In PAPER 4, the concept is extended to quantify enzyme activity of two simultaneously acting enzymes, namely pectin lyase and pectin methyl esterase. By doing so the multiway methods PARAFAC, TUCKER3 and NPLS were compared and evaluated towards accuracy and precision....

  6. A metal-based inhibitor of NEDD8-activating enzyme.

    Directory of Open Access Journals (Sweden)

    Hai-Jing Zhong

    Full Text Available A cyclometallated rhodium(III complex [Rh(ppy(2(dppz](+ (1 (where ppy=2-phenylpyridine and dppz=dipyrido[3,2-a:2',3'-c]phenazine dipyridophenazine has been prepared and identified as an inhibitor of NEDD8-activating enzyme (NAE. The complex inhibited NAE activity in cell-free and cell-based assays, and suppressed the CRL-regulated substrate degradation and NF-κB activation in human cancer cells with potency comparable to known NAE inhibitor MLN4924. Molecular modeling analysis suggested that the overall binding mode of 1 within the binding pocket of the APPBP1/UBA3 heterodimer resembled that for MLN4924. Complex 1 is the first metal complex reported to suppress the NEDDylation pathway via inhibition of the NEDD8-activating enzyme.

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

    Energy Technology Data Exchange (ETDEWEB)

    Gueguen, Y.; Souidi, M.; Baudelin, C.; Dudoignon, N.; Grison, S.; Dublineau, I.; Marquette, C.; Voisin, P.; Gourmelon, P.; Aigueperse, J. [Direction de la RadioProtection de l' Homme, Service de Radiobiologie et d' Epidemiologie. IRSN, Institut de Radioprotection et de Surete Nucleaire, B.P. No. 17, Fontenay-aux-Roses Cedex (France)

    2006-04-15

    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: 7{alpha}-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 PPAR{alpha} 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.)

  8. The Effects of Chloroquine-Resistant and Chloroquine-Sensitive Strains of Berghei on Rodent Hepatic Drug-Metabolizing Enzymes

    Science.gov (United States)

    1993-10-14

    Dipetolllonema M<’I:stomys 42.94 106 58.46 viteolle Schistosollloll mansoni S . I1\\I!lnsoni Fasciola hepatica matalensis Mi ce (SWiss) Mice (Swiss...infected with Fasciola hepatica (Table 4). This may be due to direct residence of the parasites in their host’s liver and severe injury of the liver...vitro drug- metabolizing activity of the hepatic mixed function oxidase system in rats infected experimentally with Fasciola hepatica

  9. Lipoprotein Receptors and Lipid Enzymes in Hepatitis C Virus Entry and Early Steps of Infection

    OpenAIRE

    Eve-Isabelle Pécheur

    2012-01-01

    Viruses are obligate intracellular agents that depend on host cells for successful propagation, hijacking cellular machineries to their own profit. The molecular interplay between host factors and invading viruses is a continuous coevolutionary process that determines viral host range and pathogenesis. The hepatitis C virus (HCV) is a strictly human pathogen, causing chronic liver injuries accompanied by lipid disorders. Upon infection, in addition to protein-protein and protein-RNA interacti...

  10. Enzyme butilcholinesterase activity variation in hormonal contraception users

    OpenAIRE

    Vásquez, Llermén; Dirección Regional de Salud - Puno, EsSALUD, Puno, Perú; Osorio, Jorge; Complejo Hospitalario Clínica San Pablo, Lima, Perú

    2014-01-01

    OBJECTIVE: To evaluate enzyme butilcholinesterase activity in women using two hormonal contraceptives, norgestrel ethinylestradiol and medroxyprogesterone acetate, using alkaline phosphatase as indirect liver marker. MATERIAL AND METHODS: Women attending family planning program at El Agustino "7 de Octubre" health center, continuing users of hormonal contraception for over six months. Fasting blood was obtained and enzymatic activity was measured in serum. RESULTS: Users of both contraceptive...

  11. County-Scale Spatial Distribution of Soil Enzyme Activities and Enzyme Activity Indices in Agricultural Land: Implications for Soil Quality Assessment

    OpenAIRE

    Xiangping Tan; Baoni Xie; Junxing Wang; Wenxiang He; Xudong Wang; Gehong Wei

    2014-01-01

    Here the spatial distribution of soil enzymatic properties in agricultural land was evaluated on a county-wide (567 km2) scale in Changwu, Shaanxi Province, China. The spatial variations in activities of five hydrolytic enzymes were examined using geostatistical methods. The relationships between soil enzyme activities and other soil properties were evaluated using both an integrated total enzyme activity index (TEI) and the geometric mean of enzyme activities (GME). At the county scale, soil...

  12. Inhibitors of Testosterone Biosynthetic and Metabolic Activation Enzymes

    Directory of Open Access Journals (Sweden)

    Leping Ye

    2011-12-01

    Full Text Available The Leydig cells of the testis have the capacity to biosynthesize testosterone from cholesterol. Testosterone and its metabolically activated product dihydrotestosterone are critical for the development of male reproductive system and spermatogenesis. At least four steroidogenic enzymes are involved in testosterone biosynthesis: Cholesterol side chain cleavage enzyme (CYP11A1 for the conversion of cholesterol into pregnenolone within the mitochondria, 3β-hydroxysteroid dehydrogenase (HSD3B, for the conversion of pregnenolone into progesterone, 17α-hydroxylase/17,20-lyase (CYP17A1 for the conversion of progesterone into androstenedione and 17β-hydroxysteroid dehydrogenase (HSD17B3 for the formation of testosterone from androstenedione. Testosterone is also metabolically activated into more potent androgen dihydrotestosterone by two isoforms 5α-reductase 1 (SRD5A1 and 2 (SRD5A2 in Leydig cells and peripheral tissues. Many endocrine disruptors act as antiandrogens via directly inhibiting one or more enzymes for testosterone biosynthesis and metabolic activation. These chemicals include industrial materials (perfluoroalkyl compounds, phthalates, bisphenol A and benzophenone and pesticides/biocides (methoxychlor, organotins, 1,2-dibromo-3-chloropropane and prochloraz and plant constituents (genistein and gossypol. This paper reviews these endocrine disruptors targeting steroidogenic enzymes.

  13. Peroxisome Proliferator-Activated Receptor α Activation Induces Hepatic Steatosis, Suggesting an Adverse Effect

    Science.gov (United States)

    Yan, Fang; Wang, Qi; Xu, Chao; Cao, Mingfeng; Zhou, Xiaoming; Wang, Tingting; Yu, Chunxiao; Jing, Fei; Chen, Wenbin; Gao, Ling; Zhao, Jiajun

    2014-01-01

    Non-alcoholic fatty liver disease (NAFLD) is characterized by hepatic triglyceride accumulation, ranging from steatosis to steatohepatitis and cirrhosis. NAFLD is a risk factor for cardiovascular diseases and is associated with metabolic syndrome. Antihyperlipidemic drugs are recommended as part of the treatment for NAFLD patients. Although fibrates activate peroxisome proliferator-activated receptor α (PPARα), leading to the reduction of serum triglyceride levels, the effects of these drugs on NAFLD remain controversial. Clinical studies have reported that PPARα activation does not improve hepatic steatosis. In the present study, we focused on exploring the effect and mechanism of PPARα activation on hepatic triglyceride accumulation and hepatic steatosis. Male C57BL/6J mice, Pparα-null mice and HepG2 cells were treated with fenofibrate, one of the most commonly used fibrate drugs. Both low and high doses of fenofibrate were administered. Hepatic steatosis was detected through oil red O staining and electron microscopy. Notably, in fenofibrate-treated mice, the serum triglyceride levels were reduced and the hepatic triglyceride content was increased in a dose-dependent manner. Oil red O staining of liver sections demonstrated that fenofibrate-fed mice accumulated abundant neutral lipids. Fenofibrate also increased the intracellular triglyceride content in HepG2 cells. The expression of sterol regulatory element-binding protein 1c (SREBP-1c) and the key genes associated with lipogenesis were increased in fenofibrate-treated mouse livers and HepG2 cells in a dose-dependent manner. However, the effect was strongly impaired in Pparα-null mice treated with fenofibrate. Fenofibrate treatment induced mature SREBP-1c expression via the direct binding of PPARα to the DR1 motif of the SREBP-1c gene. Taken together, these findings indicate the molecular mechanism by which PPARα activation increases liver triglyceride accumulation and suggest an adverse effect of

  14. Cytidine-5'-monophosphate-N-acetylneuraminic acid. Asialoglycoprotein sialic acid transferase activity in liver and serum of patients with juvenile hepatic cirrhosis and alpha-1-antitrypsin deficiency.

    Science.gov (United States)

    Kuhlenschmidt, M S; Peters, S P; Pinkard, O D; Glew, R H; Sharp, H

    1976-04-08

    The molecular basis for the accumulation of a substance which displays the immunological reactivity of alpha-1-antitrypsin within vesicles of liver parenchymal cells of individuals with hepatic cirrhosis and serum alpha-1-antitrypsin deficiency remains unclear. We recently reported that serum from a patient with alpha-1-antitrypsin deficiency and hepatic cirrhosis was substantially deficient in sialyltransferease (EC 2.4.99.1) an enzyme which transfers sialic acid from cytidine 5'-monophosphate-N-acetylneuraminic acid to a variety of asialoglycoprotein acceptors. In the present report we have extended these studies to include serum from five additional patients with alpha-1-antitrypsin deficiency and juvenile hepatic cirrhosis as well as a liver specimen obtained at autopsy of one of these patients. We find the sialytransferase activity in serum from six patients with alpha-1-antitrypsin deficiency and hepatic cirrhosis to be 50% of healthy pediatric control values and 30% of pediatric patients with liver disease. However, serum from family members homozygous for alpha-1-antitrypsin deficiency but without hepatic cirrhosis, and serum from patients with a variety of other kinds of liver disease, failed to exhibit the marked sialytransferase deficiency. Similar assays carried out on a homogenate of a liver sample from one patient with alpha-1-antitrypsin deficiency and hepatic cirrhosis indicated that the deficiency of sialyltransferase activity was not demonstrable in liver. Furthermore, a comparative kinetic analysis of serum and liver sialytransferase in normal and afflicted individuals failed to detect differences in substrate affinities which might account for a decrease in functional sialyltransferase capacity in individuals with alpha-1-antitrypsin deficiency and hepatic cirrhosis. These observations suggest that the serum sialyltransferase deficiency in such patients probably arises after chronic and extensive liver disease involving hepatic accumulation of

  15. Micropollutant degradation via extracted native enzymes from activated sludge.

    Science.gov (United States)

    Krah, Daniel; Ghattas, Ann-Kathrin; Wick, Arne; Bröder, Kathrin; Ternes, Thomas A

    2016-05-15

    A procedure was developed to assess the biodegradation of micropollutants in cell-free lysates produced from activated sludge of a municipal wastewater treatment plant (WWTP). This proof-of-principle provides the basis for further investigations of micropollutant biodegradation via native enzymes in a solution of reduced complexity, facilitating downstream protein analysis. Differently produced lysates, containing a variety of native enzymes, showed significant enzymatic activities of acid phosphatase, β-galactosidase and β-glucuronidase in conventional colorimetric enzyme assays, whereas heat-deactivated controls did not. To determine the enzymatic activity towards micropollutants, 20 compounds were spiked to the cell-free lysates under aerobic conditions and were monitored via LC-ESI-MS/MS. The micropollutants were selected to span a wide range of different biodegradabilities in conventional activated sludge treatment via distinct primary degradation reactions. Of the 20 spiked micropollutants, 18 could be degraded by intact sludge under assay conditions, while six showed reproducible degradation in the lysates compared to the heat-deactivated negative controls: acetaminophen, N-acetyl-sulfamethoxazole (acetyl-SMX), atenolol, bezafibrate, erythromycin and 10,11-dihydro-10-hydroxycarbamazepine (10-OH-CBZ). The primary biotransformation of the first four compounds can be attributed to amide hydrolysis. However, the observed biotransformations in the lysates were differently influenced by experimental parameters such as sludge pre-treatment and the addition of ammonium sulfate or peptidase inhibitors, suggesting that different hydrolase enzymes were involved in the primary degradation, among them possibly peptidases. Furthermore, the transformation of 10-OH-CBZ to 9-CA-ADIN was caused by a biologically-mediated oxidation, which indicates that in addition to hydrolases further enzyme classes (probably oxidoreductases) are present in the native lysates. Although the

  16. ENZYME ACTIVITY OF SEVERAL SOILS OF THE CRIMEA

    Directory of Open Access Journals (Sweden)

    Kazeev K. S.

    2014-12-01

    Full Text Available The article considers the enzymatic activity and some other ecological and biological properties of zonal soils of the Crimea (cambisol, Chromic cambisol, different subtypes of chernozems. We have revealed significant differences in catalase, dehydrogenase, polyphenol oxidase, peroxidase, invertase for soils of the Crimea, which can not be explained only by the content of soil organic matter. Despite the low humus content of the soil, some have a high level of some enzymes. The level of enzyme activity depends on the reaction medium, the content of carbonate and other soil properties. We have also revealed that the agricultural use of brown soils under vineyards leads to a significant change in their properties and enzymatic activity

  17. Microbial Community Structure and Enzyme Activities in Semiarid Agricultural Soils

    Science.gov (United States)

    Acosta-Martinez, V. A.; Zobeck, T. M.; Gill, T. E.; Kennedy, A. C.

    2002-12-01

    The effect of agricultural management practices on the microbial community structure and enzyme activities of semiarid soils of different textures in the Southern High Plains of Texas were investigated. The soils (sandy clay loam, fine sandy loam and loam) were under continuous cotton (Gossypium hirsutum L.) or in rotations with peanut (Arachis hypogaea L.), sorghum (Sorghum bicolor L.) or wheat (Triticum aestivum L.), and had different water management (irrigated or dryland) and tillage (conservation or conventional). Microbial community structure was investigated using fatty acid methyl ester (FAME) analysis by gas chromatography and enzyme activities, involved in C, N, P and S cycling of soils, were measured (mg product released per kg soil per h). The activities of b-glucosidase, b-glucosaminidase, alkaline phosphatase, and arylsulfatase were significantly (Pconservation tillage in comparison to continuous cotton under conventional tillage. Principal component analysis showed FAME profiles of these soils separated distinctly along PC1 (20 %) and PC2 (13 %) due to their differences in soil texture and management. No significant differences were detected in FAME profiles due to management practices for the same soils in this sampling period. Enzyme activities provide early indications of the benefits in microbial populations and activities and soil organic matter under crop rotations and conservation tillage in comparison to the typical practices in semiarid regions of continuous cotton and conventional tillage.

  18. Effects of long-term tea polyphenols consumption on hepatic microsomal drug-metabolizing enzymes and liver function in Wistar rats

    Institute of Scientific and Technical Information of China (English)

    Tao-Tao Liu; Ning-Sheng Liang; Yan Li; Fan Yang; Yi Lu; Zi-Qing Meng; Li-Sheng Zhang,

    2003-01-01

    AIM: To investigate the effects of long-term tea polyphenols (TPs) consumption on hepatic microsomal drug-metabolizing enzymes and liver function in rats.METHODS: TPs were administered intragastrically to rats at the doses of 833 mg.kg-1.d-1 (n=20) and 83.3 mg.kg-1@d-1 (n=20) respectively for six months. Controlled group (n=20)was given same volume of saline solution. Then the contents of cytochrome P450, bS, enzyme activities of aminopyrine N-demethylase (ADM), glutathione S-trasferase (GST) and the biochemical liver function of serum were determined.RESULTS: The contents of cytochrome P450 and b5 in the livers of male rats in high dose groups (respectively 2.66±0.55,10.43±2.78 nmol.mg MS pro-1) were significantly increased compared with the control group (1.08±1.04, 5.51±2.98nmol.mg MS pro-1; P<0.01, respectively). The enzymatic activities of ADM in the livers of female rats in high dose groups (0.91±0.08 mmol@mg MS pro-1min-1) were increased compared with the control group (0.82±0.08 mmol.mg MS pro-1.min-1; P<0.05). The GST activity was unchanged in all treated groups, and the function of liver was not obviously changed.CONCLUSION: The antidotal capability of rats' livers can be significantly improved after long-term consumption of TPs.There are differences in changes of drug-metabolizing enzymes between the sexes induced by TPs and normal condition.

  19. Functionally diverse biotin-dependent enzymes with oxaloacetate decarboxylase activity.

    Science.gov (United States)

    Lietzan, Adam D; St Maurice, Martin

    2014-02-15

    Biotin-dependent enzymes catalyze carboxylation, decarboxylation and transcarboxylation reactions that participate in the primary metabolism of a wide range of organisms. In all cases, the overall reaction proceeds via two half reactions that take place in physically distinct active sites. In the first half-reaction, a carboxyl group is transferred to the 1-N' of a covalently tethered biotin cofactor. The tethered carboxybiotin intermediate subsequently translocates to a second active site where the carboxyl group is either transferred to an acceptor substrate or, in some bacteria and archaea, is decarboxylated to biotin and CO2 in order to power the export of sodium ions from the cytoplasm. A homologous carboxyltransferase domain is found in three enzymes that catalyze diverse overall reactions: carbon fixation by pyruvate carboxylase, decarboxylation and sodium transport by the biotin-dependent oxaloacetate decarboxylase complex, and transcarboxylation by transcarboxylase from Propionibacterium shermanii. Over the past several years, structural data have emerged which have greatly advanced the mechanistic description of these enzymes. This review assembles a uniform description of the carboxyltransferase domain structure and catalytic mechanism from recent studies of pyruvate carboxylase, oxaloacetate decarboxylase and transcarboxylase, three enzymes that utilize an analogous carboxyltransferase domain to catalyze the biotin-dependent decarboxylation of oxaloacetate.

  20. Hepatitis

    Institute of Scientific and Technical Information of China (English)

    2008-01-01

    2008079 Relationship of HBV genotype and bcp and pc mutations with HBV DNA rebound after lamivudine therapy. SU Minghua(苏明华), et al. Dept Infect Dis Clin Hosp, Guangxi Med Univ, Nanning 530027. World Chin J Digestol 2007;15(33):3507-3513. Objective To investigate the relationship of HBV gene mutations with HBV DNA rebound after lamivudine therapy. Methods Twenty-seven hepatitis B patients with HBV DNA rebound after

  1. Extracellular enzyme activity and biogeochemical cycling in restored prairies

    Science.gov (United States)

    Lynch, L.; Hernandez, D.; Schade, J. D.

    2011-12-01

    Winter microbial activity in mid-latitude prairie ecosystems is thermally sensitive and significantly influenced by snow depth. Snow insulates the soil column facilitating microbial processing of complex organic substrates. Previous studies in forests and tundra ecosystems suggest patterns of substrate utilization and limitation are seasonal; above freezing, soil microbes access fresh litter inputs and sugar exudates from plant roots, while under frozen condition they recycle nutrients incorporated in microbial biomass. In order to liberate nutrients required for carbon degradation, soil microbes invest energy in the production of extracellular enzymes that cleave monomers from polymer bonds. The inverse relationship between relative enzyme abundance and substrate availability makes enzyme assays a useful proxy to assess changes in resources over time. Our objective in this study was to assess patterns in microbial biomass, nutrient availability, and extracellular enzyme activity in four snow exclosure sites over a seven-month period. Over the past three years, we have maintained a snow removal experiment on two restored prairies in central Minnesota. In each prairie, snow was continuously removed annually from two 4 x 4 m plots by shoveling after each snow event. Extractable C, N and P, and microbial C, N and P in soil samples were measured in samples collected from these snow removal plots, as well as in adjacent unmanipulated prairie control plots. Pools of C, N, and P were estimated using standard extraction protocols, and microbial pools were estimated using chloroform fumigation direct extraction (CFDE). We conducted fluorometric extracellular enzyme assays (EEA) to assess how the degradation potential of cellulose (cellobiohydrolase, CBH), protein (leucine aminopeptidase, LAP), and phosphate esters (phosphatase, PHOS) changed seasonally. Microbial C and N declined between October and June, while microbial P declined during the fall and winter, but increased

  2. The Effect of Prolonged Fasting on Total Lipid Synthesis and Enzyme Activities in the Liver of the European Eel (Anguilla anguilla)

    DEFF Research Database (Denmark)

    Abraham, S. A.; Hansen, Heinz Johs. Max; Hansen, F.N.

    1984-01-01

    The extent of fatty acid synthesis from [1-14C]acetate in liver slices was reduced 6-fold when eels were fasted for 1-7 wk and 20-fold when fasted for 39 wk, thereafter hepatic lipogenesis seemed to remain constant for up to 95 wk of fasting. After a 1-3 wk fast some hepatic enzyme activities were...... total lipid synthesis and lipogenic enzyme activity in eel liver was 30.degree. C....... reduced (acetyl-CoA carboxylase decreased 2-fold and fatty acid synthetase declined 5-fold); others remained unchanged (G-6-P dehydrogenase, 6-phosphogluconate dehydrogenase, .alpha.-glycerol phosphate dehydrogenase as well as malic enzyme and ATP-citrate lyase). The optimum temperature for measuring both...

  3. Functional Integrity of the Chimeric (Humanized) Mouse Liver: Enzyme Zonation, Physiologic Spaces, and Hepatic Enzymes and Transporters.

    Science.gov (United States)

    Chow, Edwin C Y; Wang, Jason Z Ya; Quach, Holly P; Tang, Hui; Evans, David C; Li, Albert P; Silva, Jose; Pang, K Sandy

    2016-09-01

    Chimeric mouse liver models are useful in vivo tools for human drug metabolism studies; however, liver integrity and the microcirculation remain largely uninvestigated. Hence, we conducted liver perfusion studies to examine these attributes in FRGN [Fah(-/-), Rag2(-/-), and Il2rg(-/-), NOD strain] livers (control) and chimeric livers repopulated with mouse (mFRGN) or human (hFRGN) hepatocytes. In single-pass perfusion studies (2.5 ml/min), outflow dilution profiles of noneliminated reference indicators ((51)Cr-RBC, (125)I-albumin, (14)C-sucrose, and (3)H-water) revealed preservation of flow-limited distribution and reduced water and albumin spaces in hFRGN livers compared with FRGN livers, a view supported microscopically by tightly packed sinusoids. With prograde and retrograde perfusion of harmol (50 µM) in FRGN livers, an anterior sulfation (Sult1a1) over the posterior distribution of glucuronidation (Ugt1a1) activity was preserved, evidenced by the 42% lower sulfation-to-glucuronidation ratio (HS/HG) and 14% higher harmol extraction ratio (E) upon switching from prograde to retrograde flow. By contrast, zonation was lost in mFRGN and hFRGN livers, with HS/HG and E for both flows remaining unchanged. Remnant mouse genes persisted in hFRGN livers (10%-300% those of FRGN). When hFRGN livers were compared with human liver tissue, higher UGT1A1 and MRP2, lower MRP3, and unchanged SULT1A1 and MRP4 mRNA expression were observed. Total Sult1a1/SULT1A1 protein expression in hFRGN livers was higher than that of FRGN livers, consistent with higher harmol sulfate formation. The composite data on humanized livers suggest a loss of zonation, lack of complete liver humanization, and persistence of murine hepatocyte activities leading to higher sulfation.

  4. Activity of peroxisomal enzymes, and levels of polyamines in LPA-transgenic mice on two different diets

    Directory of Open Access Journals (Sweden)

    Rønning Helle

    2005-10-01

    Full Text Available Abstract Background In man, elevated levels of plasma lipoprotein (a(Lp(a is a cardiovascular risk factor, and oxidized phospholipids are believed to play a role as modulators of inflammatory processes such as atherosclerosis. Polyamines are potent antioxidants and anti-inflammatory agents. It was therefore of interest to examine polyamines and their metabolism in LPA transgenic mice. Concentration of the polyamines putrescine, spermidine and spermine as well as the activity of peroxisomal polyamine oxidase and two other peroxisomal enzymes, acyl-CoA oxidase and catalase were measured. The mice were fed either a standard diet or a diet high in fat and cholesterol (HFHC. Some of the mice in each feeding group were in addition given aminoguanidine (AG, a specific inhibitor of diamine oxidase, which catalyses degradation of putrescine, and also inhibits non-enzymatic glycosylation of protein which is implicated in the aetiology of atherosclerosis in diabetic patients. Non-transgenic mice were used as controls. Results Intestinal peroxisomal polyamine oxidase activity was significantly higher in LPA transgenic mice than in the non-transgenic mice, while intestinal peroxisomal catalase activity was significantly lower. Hepatic β-oxidation increased in Lp(a transgenic mice fed the HFHC diet, but not in those on standard diet. Hepatic spermidine concentration was increased in all mice fed the HFHC diet compared to those fed a standard diet, while spermine concentration was decreased. With exception of the group fed only standard diet, transgenic mice showed a lower degree of hepatic steatosis than non-transgenic mice. AG had no significant effect on hepatic steatosis. Conclusion The present results indicate a connection between peroxisomal enzyme activity and the presence of the human LPA gene in the murine genome. The effect may be a result of changes in oxidative processes in lipid metabolism rather than resulting from a direct effect of the LPA

  5. Effect of Barley and Enzyme on Performance, Carcass, Enzyme Activity and Digestion Parameters of Broilers

    Directory of Open Access Journals (Sweden)

    majid kalantar

    2016-04-01

    Full Text Available Introduction Corn has been recently used for producing ethanol fuel in the major corn-producing countries such as the US and Brazil. Recent diversion of corn for biofuel production along with the increased world's demand for this feedstuff has resulted in unprecedented rise in feed cost for poultry worldwide. Alternative grains such as wheat and barley can be successfully replaced for corn in poultry diets. These cereal grains can locally grow in many parts of the world as they have remarkably lower water requirement than corn. Wheat and barley are generally used as major sources of energy in poultry diets. Though the major components of these grains are starch and proteins, they have considerable content of non-starch polysaccharides (NSPs, derived from the cell walls (Olukosi et al. 2007; Mirzaie et al. 2012. NSPs are generally considered as anti-nutritional factors (Jamroz et al. 2002. The content and structure of NSP polymers vary between different grains, which consequently affect their nutritive value (Olukosi et al. 2007.Wheat and barley are generally used as major sources of energy in poultry diets. The major components of these grains are starch and proteins, they have considerable content of non-starch polysaccharides (NSPs, derived from the cell walls. NSPs are generally considered as anti-nutritional factors. The content and structure of NSP polymers vary between different grains, which consequently affect their nutritive value. The major part of NSPs in barley comprises polymers of (1→3 (1→4-β- glucans which could impede growth factors and consequently carcass quality through lowering the rate and amount of available nutrients in the mucosal surface of the intestinal. Materials and Methods This experiment was conducted to evaluate the effect of corn and barley based diets supplemented with multi-enzyme on growth, carcass, pancreas enzyme activity and physiological characteristics of broilers. A total number of 375 one day old

  6. Extracellular enzyme activity in a willow sewage treatment system.

    Science.gov (United States)

    Brzezinska, Maria Swiontek; Lalke-Porczyk, Elżbieta; Kalwasińska, Agnieszka

    2012-12-01

    This paper presents the results of studies on the activity of extra-cellular enzymes in soil-willow vegetation filter soil which is used in the post-treatment of household sewage in an onsite wastewater treatment system located in central Poland. Wastewater is discharged from the detached house by gravity into the onsite wastewater treatment system. It flows through a connecting pipe into a single-chamber septic tank and is directed by the connecting pipe to a control well to be further channelled in the soil-willow filter by means of a subsurface leaching system. Soil samples for the studies were collected from two depths of 5 cm and 1 m from three plots: close to the wastewater inflow, at mid-length of the plot and close to its terminal part. Soil samples were collected from May to October 2009. The activity of the extra-cellular enzymes was assayed by the fluorometric method using 4-methylumbelliferyl and 7-amido-4-methylcoumarin substrate. The ranking of potential activity of the assayed enzymes was the same at 5 cm and 1 m soil depths, i.e. esterase > phosphmomoesterase > leucine-aminopeptidase > β-glucosidase > α-glucosidase. The highest values of enzymatic activity were recorded in the surface layer of the soil at the wastewater inflow and decreased with increasing distance from that point.

  7. Enzyme-like activities of algal polysaccharide - cerium complexes

    Science.gov (United States)

    Wang, Dongfeng; Sun, Jipeng; Du, Dehong; Ye, Shen; Wang, Changhong; Zhou, Xiaoling; Xue, Changhu

    2005-01-01

    Water-soluble algal polysaccharides (APS) (alginic acid, fucoidan and laminaran) possess many pharmacological activities. The results of this study showed that the APS-Ce4+ complexes have some enzyme-like activities. Fucoidan and its complex with Ce4+ have activities similar to those of SOD. The activities of laminaran, alginic acid and their complexes are not measurable. The APS do not show measurable activities in the digestion of plasmid DNA. In contrast, the APS - Ce4+ complexes show these measurable activities under the comparable condition when APS bind Ce4+ and form homogenous solutions. The laminaran - Ce4+ complex shows the most obvious activity in the digestion of plasmid DNA, pNPP and chloropy-rifos under neutral conditions.

  8. Digestive enzymes activity in larvae of Cameraria ohridella (Lepidoptera: Gracillariidae).

    Science.gov (United States)

    Stygar, Dominika; Dolezych, Bogdan; Nakonieczny, Mirosław; Migula, Pawel; Michalczyk, Katarzyna; Zaak, Maria

    2010-10-01

    This article presents the activity of carbohydratases and proteases in the midgut of Cameraria ohridella larvae--an oligophagous pest whose preferred feeding is horse chestnuts leaves. Optimal media pH of the assayed enzymes were similar to those of other Lepidopterans. Relatively high amylase activity, as well as maltase and sucrase activities, indicates that starch and sucrose are the main digested saccharides. Trehalase activity was similar to that described in other Lepidopterans. Activities of glycosidases were significantly lower than those of disaccharidases what suggests that neither cellulose nor glycosides are important for C. ohridella. Trypsin is the main endoprotease of this pest. Like in other leaf-eaters carboxypeptidase activity was higher than that of aminopeptidase. The activity of the majority of examined enzymes increased in the following successive pest generations, which could be explained by the decreased nutritional value of older leaves. Probably this phenomenon in hydrolases activity in Cameraria is a nonspecific mechanism present at this stage of co-evolution of the horse chestnut and its pest.

  9. Enantioselective effects of metalaxyl on soil enzyme activity.

    Science.gov (United States)

    Yue, Heng; Fang, Song; Zhang, Yizhi; Ning, Yang; Yu, Weisong; Kong, Fanyu; Qiu, Jun

    2016-12-01

    The enantioselective effects of the chiral pesticide metalaxyl on soil enzyme activity were investigated. Incubation experiments were conducted to investigate the effects of metalaxyl enantiomers at different concentrations on the activities of urease, invertase, and catalase as well as the type of activity change (activation vs. inhibition) at different times during incubation. The results indicated that the effects of metalaxyl on the activity of soil enzymes were not only related to the concentration of the enantiomers and soil incubation time, but also to the chiral configuration, suggesting the effects were enantioselective. A pattern of inhibition-recovery-slight stimulation was observed in urease activity of the soil samples treated with metalaxyl enantiomers, but the effects of (-) -R-metalaxyl were stronger than those of (+)-S-metalaxyl at the same concentration. Invertase activity in soil samples treated with metalaxyl enantiomers initially sharply decreased before finally returning to the normal level, and the effects of (+)-S-metalaxyl were stronger than those of (-) -R-metalaxyl at the same concentration. Metalaxyl enantiomers influenced catalase activity in a pattern of slight stimulation-inhibition-recovery, and the effects of (-) -R-metalaxyl were stronger than those of (+)-S-metalaxyl at the same concentration. © 2016 Wiley Periodicals, Inc.

  10. Evidence that the RNAseH activity of the duck hepatitis B virus is unable to act on exogenous substrates

    Directory of Open Access Journals (Sweden)

    Tavis John E

    2001-07-01

    Full Text Available Abstract Background The hepadnaviral reverse transcriptase can synthesize DNA on its native RNA template within viral cores but it is usually unable to synthesize DNA employing exogenous nucleic acids as a template. The mechanism of this template commitment is unknown. Here we provide evidence that the RNAseH activity of duck hepatitis B virus reverse transcriptase may also be unable to act on exogenous substrates. Results RNAseH assays were performed under a wide variety of conditions employing substrate RNAs of Duck Hepatitis B Virus sequence annealed to complementary DNA oligonucleotides and permeabilized intracellular viral core particles. Temperature, pH, cation type, salt concentration, substrate concentration, and the sequences of the cleavage sites were varied, and the effects of ATP and dNTPs on RNAseH activity were examined. duck hepatitis B virus RNAseH activity was not detected under any of these conditions, although E. coli or Avian Myeloblastosis Virus RNAseH activity could be detected under all conditions. Access of the RNA substrate to the enzyme within the viral cores was confirmed. Conclusions These results imply that the RNAseH activity of the DHBV reverse transcriptase may not be able to degrade exogenous RNA:DNA heteroduplexes, although it can degrade heteroduplexes of the same sequence generated during reverse transcription of the endogenous RNA template. Therefore, the RNAseH activity appears to be "substrate committed" in a manner similar to the template commitment observed for the DNA polymerase activity.

  11. Diverticular bile duct lesion in chronic active hepatitis

    DEFF Research Database (Denmark)

    Vyberg, M

    1989-01-01

    Liver needle biopsies from patients with non-A, non-B chronic active hepatitis and so-called abnormal bile duct epithelium were studied with a three-dimensional method. Photographs of bile duct structures in serial sections were transferred to acrylic plates. Five bile duct lesions of a not previ......Liver needle biopsies from patients with non-A, non-B chronic active hepatitis and so-called abnormal bile duct epithelium were studied with a three-dimensional method. Photographs of bile duct structures in serial sections were transferred to acrylic plates. Five bile duct lesions...... cells, but most were larger, with rounded nuclei, prominent nucleoli and abundant eosinophilic cytoplasm, sometimes with periodic acid-Schiff-positive, diastase-resistant granules. The lesions were only partly surrounded by a basement membrane. They were all embedded in a tight mononuclear inflammatory...... infiltrate associated with pronounced periportal piecemeal necrosis. In two cases, a germinal center was adjacent to the epithelium. The pathogenesis of the diverticular bile duct lesion is unknown, but the diverticuli probably represent Hering ducts and groups of periportal liver cells which have escaped...

  12. Extracellular Enzyme Activity assay as indicator of soil microbial functional diversity and activity

    DEFF Research Database (Denmark)

    Hendriksen, Niels Bohse; Winding, Anne

    2012-01-01

    and especially hydrolytic extracellular enzymes are of pivotal importance for decomposition of organic substrates and biogeochemical cycling. Their activity reflects the functional diversity and activity of the microorganisms involved in decomposition processes which are essential processes for soil functioning...... and soil ecosystem services. The soil enzyme activity has been measured by the use of fluorogenic model substrates e.g. methylumbelliferyl (MUF) substrates for a number of enzymes involved in the degradation of polysaccharides as cellulose, hemicellulose and chitin, while degradation of proteins has been...... followed by amino-methyl-coumaric substrates (AMC). The already developed Extracellular Enzyme Activity (EEA) assay was further optimized as a microwell based assay for the activity of enzymes involved in degradation of polysaccharides and proteins. Using specific MUF and AMC substrates on European soils...

  13. Extracellular enzyme activity assay as indicator of soil microbial functional diversity and activity

    DEFF Research Database (Denmark)

    Hendriksen, Niels Bohse; Winding, Anne

    2012-01-01

    of soil microbial functions is still needed. In soil, enzymes originate from a variety of organisms, notably fungi and bacteria and especially hydrolytic extracellular enzymes are of pivotal importance for decomposition of organic substrates and biogeochemical cycling. Their activity will reflect...... the functional diversity and activity of the microorganisms involved in decomposition processes. Their activity has been measured by the use of fluorogenic model substrates e.g. methylumbelliferyl (MUF) substrates for a number of enzymes involved in the degradation of polysacharides as cellulose, hemicellulose...... and chitin, while degradation of proteins has been followed by amino-methyl-coumaric substrates (AMC). Based on these fluorogenic substrates the Extracellular Enzyme Activity assay was optimized as a microwell based standardized assay for the activity of enzymes involved in degradation of polysaccharides...

  14. IL28B polymorphism correlates with active hepatitis in patients with HBeAg-negative chronic hepatitis B.

    Directory of Open Access Journals (Sweden)

    I-Cheng Lee

    Full Text Available BACKGROUND AIMS: The clinical relevance of single nucleotide polymorphisms (SNPs near the IL28B gene is controversial in patients with hepatitis B virus (HBV infection. This study aimed to investigate the role of viral and host factors, including IL28B genotypes, in the natural course of chronic hepatitis B (CHB. METHODS: The study enrolled consecutive 115 treatment-naive CHB patients. HBV viral loads, genotypes, precore and basal core promotor mutations, serum hepatitis B surface antigen (HBsAg and interferon-gamma inducible protein 10 (IP-10 levels as well as four SNPs of IL28B were determined. Serial alanine transaminase (ALT levels in the previous one year before enrollment at an interval of three months were recorded. Factors associated with active hepatitis, defined as persistent ALT >2× upper limit of normal (ULN or a peak ALT level >5× ULN, were evaluated. RESULTS: The prevalence of rs8105790 TT, rs12979860 CC, rs8099917 TT, and rs10853728 CC genotypes were 88.3%, 87.4%, 88.4% and 70.9%, respectively. In HBeAg-positive patients (n = 48, HBV viral load correlated with active hepatitis, while in HBeAg-negative patients (n = 67, rs10853728 CC genotype (p = 0.032 and a trend of higher IP-10 levels (p = 0.092 were associated with active hepatitis. In multivariate analysis, high viral load (HBV DNA >10(8 IU/mL, p = 0.042, odds ratio = 3.946 was significantly associated with HBeAg-positive hepatitis, whereas rs10853728 CC genotype (p = 0.019, odds ratio = 3.927 was the only independent factor associated with active hepatitis in HBeAg-negative population. CONCLUSIONS: HBV viral load and IL28B rs10853728 CC genotype correlated with hepatitis activity in HBeAg-positive and HBeAg-negative CHB, respectively. Both viral and host factors play roles in disease activity during different phases of CHB.

  15. Growth characteristics and enzyme activity in Batrachochytrium dendrobatidis isolates.

    Science.gov (United States)

    Symonds, E Pearl; Trott, Darren J; Bird, Philip S; Mills, Paul

    2008-09-01

    Batrachochytrium dendrobatidis is a member of the phylum Chytridiomycota and the causative organism chytridiomycosis, a disease of amphibians associated with global population declines and mass mortality events. The organism targets keratin-forming epithelium in adult and larval amphibians, which suggests that keratinolytic activity may be required to infect amphibian hosts. To investigate this hypothesis, we tested 10 isolates of B. dendrobatidis for their ability to grow on a range of keratin-supplemented agars and measured keratolytic enzyme activity using a commercially available kit (bioMerieux API ZYM). The most dense and fastest growth of isolates were recorded on tryptone agar, followed by growth on frog skin agar and the slowest growth recorded on feather meal and boiled snake skin agar. Growth patterns were distinctive for each nutrient source. All 10 isolates were strongly positive for a range of proteolytic enzymes which may be keratinolytic, including trypsin and chymotrypsin. These findings support the predilection of B. dendrobatidis for amphibian skin.

  16. A DNA enzyme with Mg(2+)-Dependent RNA Phosphoesterase Activity

    Science.gov (United States)

    Breaker, Ronald R.; Joyce, Gerald F.

    1995-01-01

    Previously we demonstrated that DNA can act as an enzyme in the Pb(2+)-dependent cleavage of an RNA phosphoester. This is a facile reaction, with an uncatalyzed rate for a typical RNA phosphoester of approx. 10(exp -4)/ min in the presence of 1 mM Pb(OAc)2 at pH 7.0 and 23 C. The Mg(2+) - dependent reaction is more difficult, with an uncatalyzed rate of approx. 10(exp -7)/ min under comparable conditions. Mg(2+) - dependent cleavage has special relevance to biology because it is compatible with intracellular conditions. Using in vitro selection, we sought to develop a family of phosphoester-cleaving DNA enzymes that operate in the presence of various divalent metals, focusing particularly on the Mg(2+) - dependent reaction. Results: We generated a population of greater than 10(exp 13) DNAs containing 40 random nucleotides and carried out repeated rounds of selective amplification, enriching for molecules that cleave a target RNA phosphoester in the presence of 1 mM Mg(2+), Mn(2+), Zn(2+) or Pb(2+). Examination of individual clones from the Mg(2+) lineage after the sixth round revealed a catalytic motif comprised of a three-stem junction.This motif was partially randomized and subjected to seven additional rounds of selective amplification, yielding catalysts with a rate of 0.01/ min. The optimized DNA catalyst was divided into separate substrate and enzyme domains and shown to have a similar level of activity under multiple turnover conditions. Conclusions: We have generated a Mg(2+) - dependent DNA enzyme that cleaves a target RNA phosphoester with a catalytic rate approx. 10(exp 5) - fold greater than that of the uncatalyzed reaction. This activity is compatible with intracellular conditions, raising the possibility that DNA enzymes might be made to operate in vivo.

  17. Effect of Commiphora mukul gum resin on hepatic and renal marker enzymes, lipid peroxidation and antioxidants status in pancreas and heart in fructose fed insulin resistant rats

    Directory of Open Access Journals (Sweden)

    B. Ramesh

    2015-12-01

    Full Text Available This work aims to study the antioxidant efficacy of Commiphora mukul (C. mukul gum resin ethanolic extract in high fructose diet (HFD insulin resistant rats. The male Wistar albino rats were randomly divided into four groups of eight animals each; two of these groups (Control group [C] and Control treated with C. mukul [C + CM] were fed with standard pellet diet and the other two groups (Fructose fed rats [F-group] and fructose fed with C. mukul treated group [F + CM] were fed with high fructose diet (HFD (66%. C. mukul gum resin ethanolic extract (200 mg/kg body weight/day was administered orally to group C + CM and group F + CM. At the end of 60-day experimental period biochemical parameters related to antioxidant, oxidative stress marker enzymes and hepatic and renal marker enzymes of tissues were performed. The fructose fed rats showed increased level of enzymatic activities aspartate aminotransminases (AST, alanine aminotransminases (ALT in liver and kidney and oxidative markers like lipid peroxidation (LPO and protein oxidation (PO in pancreas and heart. Antioxidant enzyme activities were significantly decreased in the pancreas and heart compared to control groups. Administration of C. mukul (200 mg/kg bwt to fructose fed insulin resistant rats for 60 days significantly reversed the above parameters toward normal. In conclusion, our data indicate the preventive role of C. mukul against fructose-induced insulin resistance and oxidative stress; hence this plant could be used as an adjuvant therapy for the prevention and/or management of chronic diseases characterized by hyperinsulinemia, insulin resistance and aggravated antioxidant status.

  18. ACTIVITY OF SUPEROXIDE DISMUTASE ENZYME IN YEAST SACCHAROMYCES CEREVISIAE

    OpenAIRE

    2014-01-01

    Reactive oxygen species (ROS) with reactive nitrogen species (RNS) are known to play dual role in biological systems, they can be harmful or beneficial to living systems. ROS can be important mediators of damage to cell structures, including proteins, lipids and nucleic acids termed as oxidative stress. The antioxidant enzymes protect the organism against the oxidative damage caused by active oxygen forms. The role of superoxide dismutase (SOD) is to accelerate the dismutation of the toxic su...

  19. Controlling the enzymatic activity of a restriction enzyme by light.

    Science.gov (United States)

    Schierling, Benno; Noël, Ann-Josée; Wende, Wolfgang; Hien, Le Thi; Volkov, Eugeny; Kubareva, Elena; Oretskaya, Tatiana; Kokkinidis, Michael; Römpp, Andreas; Spengler, Bernhard; Pingoud, Alfred

    2010-01-26

    For many applications it would be desirable to be able to control the activity of proteins by using an external signal. In the present study, we have explored the possibility of modulating the activity of a restriction enzyme with light. By cross-linking two suitably located cysteine residues with a bifunctional azobenzene derivative, which can adopt a cis- or trans-configuration when illuminated by UV or blue light, respectively, enzymatic activity can be controlled in a reversible manner. To determine which residues when cross-linked show the largest "photoswitch effect," i.e., difference in activity when illuminated with UV vs. blue light, > 30 variants of a single-chain version of the restriction endonuclease PvuII were produced, modified with azobenzene, and tested for DNA cleavage activity. In general, introducing single cross-links in the enzyme leads to only small effects, whereas with multiple cross-links and additional mutations larger effects are observed. Some of the modified variants, which carry the cross-links close to the catalytic center, can be modulated in their DNA cleavage activity by a factor of up to 16 by illumination with UV (azobenzene in cis) and blue light (azobenzene in trans), respectively. The change in activity is achieved in seconds, is fully reversible, and, in the case analyzed, is due to a change in V(max) rather than K(m).

  20. Extracellular enzyme activity assay as indicator of soil microbial functional diversity and activity

    DEFF Research Database (Denmark)

    Hendriksen, Niels Bohse; Winding, Anne

    2012-01-01

    Extracellular enzyme activity assay as indicator of soil microbial functional diversity and activity Niels Bohse Hendriksen, Anne Winding. Department of Environmental Science, Aarhus University, 4000 Roskilde, Denmark Soils provide numerous essential ecosystem services such as carbon cycling...... the functional diversity and activity of the microorganisms involved in decomposition processes. Their activity has been measured by the use of fluorogenic model substrates e.g. methylumbelliferyl (MUF) substrates for a number of enzymes involved in the degradation of polysacharides as cellulose, hemicellulose...

  1. A lard-rich high-fat diet increases hepatic peroxisome proliferator-activated receptors in endotoxemic rats.

    Science.gov (United States)

    Kai, Motoki; Miyoshi, Makoto; Fujiwara, Mayu; Nishiyama, Yuya; Inoue, Taketo; Maeshige, Noriaki; Hamada, Yasuhiro; Usami, Makoto

    2017-05-15

    Diets high in saturated fatty acids activate chronic inflammation. We previously reported that, in even acute inflammation caused by lipopolysaccharide (LPS), liver injury was exacerbated in rats fed a lard-rich diet. Peroxisome proliferator-activated receptors (PPARs) are related to inflammation and are also key regulators of lipid metabolism. In this study, we examined effects of high-fat diet on liver injury and hepatic lipid metabolism during endotoxemia, measuring hepatic PPARs and other markers. Male Wistar rats were fed a high-fat diet (HFD, 60 kcal% fat) or control diet (CD, 10 kcal% fat) for 4 or 12 wk, injected with LPS and sacrificed at 0, 1.5, or 6 h. Analyses included plasma aspartate transaminase (AST) and alanine transaminase (ALT) levels, messenger RNA (mRNA) and protein levels of hepatic PPARα and PPARγ, and mRNA levels of enzymes related to fatty acid oxidation and synthesis. Endotoxemic rats on HFD for 12 wk, but not 4 wk, had higher mRNA and protein levels for hepatic PPARs, than did those on CD (P PPARα and PPARγ expression in endotoxemic rats. Copyright © 2016 Elsevier Inc. All rights reserved.

  2. Evaluation of pancreatin stability through enzyme activity determination

    Directory of Open Access Journals (Sweden)

    Terra Gleysson De Paula

    2016-09-01

    Full Text Available Pancreatin is a biotechnological product containing an enzyme complex, obtained from porcine pancreas, that is employed in treating pancreatic diseases. Experiments regarding the stability of the pharmaceutical formulation containing pancreatin were performed using standard binary mixtures with 6 excipients in a 1:1 ratio (m/m and a commercial formulation. To accomplish these goals, samples were stored for 1, 3 and 6 months at 40 ± 1 °C and 75 ± 5 % relative humidity (RH and 40 ± 1 °C and 0 % RH. Stress testing was also performed. All samples were analyzed to evaluate the α-amylase, lipase and protease activities through UV/Vis spectrophotometry. The results revealed that the excipient proprieties and the storage conditions affected enzyme stability. Humidity was a strong influencing factor in the reduction of α-amylase and protease activities. Stress testing indicated that pH 9.0 and UV light did not induce substantial alterations in enzyme activity.

  3. In vivo enzyme activity in inborn errors of metabolism

    Energy Technology Data Exchange (ETDEWEB)

    Thompson, G.N.; Walter, J.H.; Leonard, J.V.; Halliday, D. (Clinical Research Centre, Harrow (England))

    1990-08-01

    Low-dose continuous infusions of (2H5)phenylalanine, (1-13C)propionate, and (1-13C)leucine were used to quantitate phenylalanine hydroxylation in phenylketonuria (PKU, four subjects), propionate oxidation in methylmalonic acidaemia (MMA, four subjects), and propionic acidaemia (PA, four subjects) and leucine oxidation in maple syrup urine disease (MSUD, four subjects). In vivo enzyme activity in PKU, MMA, and PA subjects was similar to or in excess of that in adult controls (range of phenylalanine hydroxylation in PKU, 3.7 to 6.5 mumol/kg/h, control 3.2 to 7.9, n = 7; propionate oxidation in MMA, 15.2 to 64.8 mumol/kg/h, and in PA, 11.1 to 36.0, control 5.1 to 19.0, n = 5). By contrast, in vivo leucine oxidation was undetectable in three of the four MSUD subjects (less than 0.5 mumol/kg/h) and negligible in the remaining subject (2 mumol/kg/h, control 10.4 to 15.7, n = 6). These results suggest that significant substrate removal can be achieved in some inborn metabolic errors either through stimulation of residual enzyme activity in defective enzyme systems or by activation of alternate metabolic pathways. Both possibilities almost certainly depend on gross elevation of substrate concentrations. By contrast, only minimal in vivo oxidation of leucine appears possible in MSUD.

  4. [Mass spectrometry assay for denitrifying enzyme activity measurement].

    Science.gov (United States)

    Zhang, Li-li; Wu, Zhi-jie; Song, Yu-chao

    2010-07-01

    Nitrification inhibitors exert inhibition function in soil nitrification process (NH4(+)-N to NO3(-)-N) and are widely applied in order to improve N fertilizer use efficiency. Before the new nitrification inhibitor is used, its effects on denitrification process must be investigated and denitrifying enzyme activity (DEA) is an effective indicator to show this process. In the present paper, a mass spectrometery (MS) method was taken to measure the denitrifying enzyme activity in the new nitrification inhibitor 3, 4-dimethylpyrazole phosphate (DMPP) incubation system. The results showed that the method measured the concentration of N2O in the incubation system accurately and the concentration measured by MS had marked correlation with that measured by gas chromatogram (GC) (MS(N2O) = -0.45 + 1.03GC(N2O, R2 = 0.995). At the same time, enrichment of 15 N2O and 15 N2 was measured to discriminate the nitrate reductase and N2O reductase. Differed with traditional method, the enzymatic inhibitor-acetylene was not appended. The results showed that DMPP had no influence on the denitrifying enzyme activity and indicated that the denitrification process was not influenced by DMPP.

  5. Microsomal enzyme activity, glutathione S-transferase-placental form expression, cell proliferation, and vitamin A stores in livers of rats consuming Great Lakes salmon.

    Science.gov (United States)

    Iverson, F; Mehta, R; Hierlihy, L; Gurofsky, S; Lok, E; Mueller, R; Bourbonnais, D H; Spear, P A

    1998-02-01

    Male and female Sprague-Dawley rats were fed diets incorporating lyophilized chinook salmon obtained from Lake Ontario and Lake Huron. After 70 days, females were bred and the progeny (F1) were reared on the same fish-based diets as the adults (F0). After 78-133 days on the diets, males and females of both generations were sacrificed and hepatic microsomal enzyme activities determined, along with glutathione S-transferase-placental form (GSTP) expression and hepatic cellular proliferation. Hepatic P450 enzyme activities (MROD, EROD, PROD, BROD, and aminopyrine) were increased significantly by fish diets from both sources. Increases in hepatic enzyme activity were greatest for fish caught from Lake Ontario and reflected the total levels of organochlorine contaminants in the fish. GSTP and cell proliferation rates did not show any diet-related or dose-related changes. Vitamin A stores were analyzed as the concentration of liver retinyl palmitate. In rats receiving the highest TEQ dose (i.e., 20% Lake Ontario fish diet), vitamin A stores were significantly lower in F0 adults, F1 weanlings, and F1 adult females.

  6. Glial fibrillary acidic protein as an early marker of hepatic stellate cell activation in chronic and posttransplant recurrent hepatitis C.

    Science.gov (United States)

    Carotti, Simone; Morini, Sergio; Corradini, Stefano Ginanni; Burza, Maria Antonella; Molinaro, Antonio; Carpino, Guido; Merli, Manuela; De Santis, Adriano; Muda, Andrea Onetti; Rossi, Massimo; Attili, Adolfo Francesco; Gaudio, Eugenio

    2008-06-01

    Activated alpha-smooth muscle actin (alpha-SMA)-positive hepatic stellate cells (HSCs) are pericytes responsible for fibrosis in chronic liver injury. The glial fibrillary acidic protein (GFAP), commonly expressed by astrocytes in the central nervous system, is expressed in vivo in the liver in a subpopulation of quiescent stellate cells. In the rat, increased GFAP expression in the acute response to injury and down-regulation in the chronic response have been observed, whereas reports concerning GFAP expression in human liver are still conflicting. We investigated the utility of GFAP compared to alpha-SMA as an immunohistochemical marker of early activated HSCs in chronic and posttransplant recurrent hepatitis C and correlated GFAP expression with vascular remodeling and fibrosis progression. With immunohistochemistry and a semiquantitative scoring system, the expression of GFAP and alpha-SMA in HSCs and the microvessel density were analyzed in biopsies from normal livers obtained from cadaveric donors [donor liver (DL); n = 21] and from livers from posttransplant hepatitis C virus recurrent hepatitis (HCV-PTR) patients (n = 19), hepatitis C virus chronic hepatitis (HCV-CH) patients, (n = 12), and hepatitis C virus cirrhosis (HCV-C) patients (n = 16). The percentage of alpha-SMA-positive HSCs was significantly higher in the HCV-PTR, HCV-CH, and HCV-C groups compared to the DL group (P HCV-PTR group compared to the DL, HCV-C (P HCV-CH (P HCV-CH group compared to the DL group (P HCV-PTR group, the percentage of GFAP-positive HSCs correlated with fibrosis progression (P HCV-CH and seems to predict fibrosis progression in HCV-PTR.

  7. Engineered N-acetylhexosamine-active enzymes in glycoscience.

    Science.gov (United States)

    Slámová, Kristýna; Bojarová, Pavla

    2017-08-01

    In recent years, enzymes modifying N-acetylhexosamine substrates have emerged in numerous theoretical studies as well as practical applications from biology, biomedicine, and biotechnology. Advanced enzyme engineering techniques converted them into potent synthetic instruments affording a variety of valuable glycosides. This review presents the diversity of engineered enzymes active with N-acetylhexosamine carbohydrates: from popular glycoside hydrolases and glycosyltransferases to less known oxidases, epimerases, kinases, sulfotransferases, and acetylases. Though hydrolases in natura, engineered chitinases, β-N-acetylhexosaminidases, and endo-β-N-acetylglucosaminidases were successfully employed in the synthesis of defined natural and derivatized chitooligomers and in the remodeling of N-glycosylation patterns of therapeutic antibodies. The genes of various N-acetylhexosaminyltransferases were cloned into metabolically engineered microorganisms for producing human milk oligosaccharides, Lewis X structures, and human-like glycoproteins. Moreover, mutant N-acetylhexosamine-active glycosyltransferases were applied, e.g., in the construction of glycomimetics and complex glycostructures, industrial production of low-lactose milk, and metabolic labeling of glycans. In the synthesis of biotechnologically important compounds, several innovative glycoengineered systems are presented for an efficient bioproduction of GlcNAc, UDP-GlcNAc, N-acetylneuraminic acid, and of defined glycosaminoglycans. The above examples demonstrate that engineering of N-acetylhexosamine-active enzymes was able to solve complex issues such as synthesis of tailored human-like glycoproteins or industrial-scale production of desired oligosaccharides. Due to the specific catalytic mechanism, mutagenesis of these catalysts was often realized through rational solutions. Specific N-acetylhexosamine glycosylation is crucial in biological, biomedical and biotechnological applications and a good

  8. Endoplasmic reticulum localization and activity of maize auxin biosynthetic enzymes.

    Science.gov (United States)

    Kriechbaumer, Verena; Seo, Hyesu; Park, Woong June; Hawes, Chris

    2015-09-01

    Auxin is a major growth hormone in plants and the first plant hormone to be discovered and studied. Active research over >60 years has shed light on many of the molecular mechanisms of its action including transport, perception, signal transduction, and a variety of biosynthetic pathways in various species, tissues, and developmental stages. The complexity and redundancy of the auxin biosynthetic network and enzymes involved raises the question of how such a system, producing such a potent agent as auxin, can be appropriately controlled at all. Here it is shown that maize auxin biosynthesis takes place in microsomal as well as cytosolic cellular fractions from maize seedlings. Most interestingly, a set of enzymes shown to be involved in auxin biosynthesis via their activity and/or mutant phenotypes and catalysing adjacent steps in YUCCA-dependent biosynthesis are localized to the endoplasmic reticulum (ER). Positioning of auxin biosynthetic enzymes at the ER could be necessary to bring auxin biosynthesis in closer proximity to ER-localized factors for transport, conjugation, and signalling, and allow for an additional level of regulation by subcellular compartmentation of auxin action. Furthermore, it might provide a link to ethylene action and be a factor in hormonal cross-talk as all five ethylene receptors are ER localized.

  9. Trace elements as an activator of antioxidant enzymes

    Directory of Open Access Journals (Sweden)

    Marta Wołonciej

    2016-12-01

    Full Text Available Oxidative stress is a state of impaired balance between the formation of free radicals and antioxidant capacity of the body. It causes many defects of the body, e.g. lipid peroxidation, DNA and protein damage. In order to prevent the effects of oxidative stress, the organism has developed defence mechanisms. These mechanisms capture and inhibit the formation of free radicals and also chelate ion metals that catalyse free radical reactions. Trace elements are components of antioxidant enzymes involved in antioxidant mechanisms. Selenium, as a selenocysteine, is a component of the active site of glutathione peroxidase (GPx. The main function of GPx is neutralization of hydrogen peroxide (H2O2 and organic peroxide (LOOH. Furthermore, selenium is a structural part of a large group of selenoproteins that are necessary for proper functioning of the body. Manganese, copper and zinc are a part of the group of superoxide dismutase enzymes (MnSOD, Cu/ZnSOD, which catalyse the superoxide anion dismutation into hydrogen peroxide and oxygen. Formed hydrogen peroxide is decomposed into water and oxygen by catalase or glutathione peroxidase. An integral component of catalase (CAT is iron ions. The concentration of these trace elements has a significant influence on the activity of antioxidant enzymes, and thus on defence against oxidative stress. Even a small change in the level of trace elements in the tissue causes a disturbance in their metabolism, leading to the occurrence of many diseases.

  10. Effects of Different Substrates on Lignocellulosic Enzyme Expression, Enzyme Activity, Substrate Utilization and Biological Efficiency of Pleurotus Eryngii

    Directory of Open Access Journals (Sweden)

    Chunliang Xie

    2016-09-01

    Full Text Available Background/Aims: Pleurotus eryngii is one of the most valued and delicious mushrooms which are commercially cultivated on various agro-wastes. How different substrates affect lignocellulosic biomass degradation, lignocellulosic enzyme production and biological efficiency in Pleurotus eryngii was unclear. Methods and Results: In this report, Pleurotus eryngii was cultivated in substrates including ramie stalks, kenaf stalks, cottonseed hulls and bulrush stalks. The results showed that ramie stalks and kenaf stalks were found to best suitable to cultivate Pleurotus eryngii with the biological efficiency achieved at 55% and 57%, respectively. In order to establish correlations between different substrates and lignocellulosic enzymes expression, the extracellular proteins from four substrates were profiled with high throughput TMT-based quantitative proteomic approach. 241 non-redundant proteins were identified and 74 high confidence lignocellulosic enzymes were quantified. Most of the cellulases, hemicellulases and lignin depolymerization enzymes were highly up-regulated when ramie stalks and kenaf stalks were used as carbon sources. The enzyme activities results suggested cellulases, hemicellulases and lignin depolymerization enzymes were significantly induced by ramie stalks and kenaf stalks. Conclusion: The lignocelluloses degradation, most of the lignocellulosic enzymes expressions and activities of Pleurotus eryngii had positive correlation with the biological efficiency, which depend on the nature of lignocellulosic substrates. In addition, the lignocellulosic enzymes expression profiles during Pleurotus eryngii growth in different substrates were obtained. The present study suggested that most of the lignocellulosic enzymes expressions and activities can be used as tools for selecting better performing substrates for commercial mushroom cultivation.

  11. Cysteine 904 is required for maximal insulin degrading enzyme activity and polyanion activation.

    Directory of Open Access Journals (Sweden)

    Eun Suk Song

    Full Text Available Cysteine residues in insulin degrading enzyme have been reported as non-critical for its activity. We found that converting the twelve cysteine residues in rat insulin degrading enzyme (IDE to serines resulted in a cysteine-free form of the enzyme with reduced activity and decreased activation by polyanions. Mutation of each cysteine residue individually revealed cysteine 904 as the key residue required for maximal activity and polyanion activation, although other cysteines affect polyanion binding to a lesser extent. Based on the structure of IDE, Asn 575 was identified as a potential hydrogen bond partner for Cys904 and mutation of this residue also reduced activity and decreased polyanion activation. The oligomerization state of IDE did not correlate with its activity, with the dimer being the predominant form in all the samples examined. These data suggest that there are several conformational states of the dimer that affect activity and polyanion activation.

  12. Preliminary hepatoprotective activity of Jigarine Cl on carbon tetra chloride induced hepatic damage in rats

    Directory of Open Access Journals (Sweden)

    Muhammad Ahmed

    2014-02-01

    Full Text Available Jigarine CL  is an herbal preparation of Hamdard laboratories waqf Pakistan that has been used as hepatoprotective agent. In this study, the protective effects of Jigarine CL  against liver damage were evaluated in carbon tetrachloride (CCl4-induced chronic hepatotoxicity in rats. Sprague-Dawley (SD rats were divided in to four groups (I, II, III, IV. Groups I received olive oil 10ml/kg p.o served as control and group II  and III were orally fed with Jigarine CL (20ml/kg for 14 consecutive days while Groups IV were orally fed with saline. After the last dose of jigarine CL and saline group III and IV received a single dose of CCl4 (0.3 ml/kg body weight in a 20% olive oil was injected intraperitoneally 30 min after the last dose of Barrisal and the animals were starved for 24 h. The degree of hepatic-protection was measured using biochemical parameters (serum glutamate oxaloacetate transaminase (SGOT and serum glutamate pyruvate transaminase (SGPT and Bilirubin and histopathology. Data obtained from results were compared using student’s t-test. The results showed that the treatment of Jigarine CL  significantly lowered the CCl4-induced serum levels of hepatic enzyme markers (GOT, GPT and total bilirubin indicated hepatoprotective effect of the Jigarine CL. Histopathological examination of liver sections confirmed that, pre-treatment with Jigarine CL  decreased the degree of hepatic damage induced by CCl4. The  significant reversal of the biochemical and histological changes induced by CCl4treatment in rats, indicating promising hepatoprotective activity of Jigarine CL in rats.

  13. St. John's Wort increases brain serotonin synthesis by inhibiting hepatic tryptophan 2, 3 dioxygenase activity and its gene expression in stressed rats.

    Science.gov (United States)

    Bano, Samina; Ara, Iffat; Saboohi, Kausar; Moattar, Tariq; Chaoudhry, Bushra

    2014-09-01

    We aimed to investigate the effects of herbal St. John's Wort (SJW) on transcriptional regulation of hepatic tryptophan 2, 3 - dioxygenase (TDO) enzyme activity and brain regional serotonin (5-HT) levels in rats exposed to forced swim test (FST). TDO mRNA expression was quantified using real-time reverse transcription polymerase chain (RT-PCR) reaction and brain regional indoleamines were determined by high performance liquid chromatography coupled to fluorescence detector. Behavioral analysis shows significant reduction in immobility time in SJW (500mg/kg/ml) administered rats. It was found that pretreatment of SJW to rats did not prevent stress-induced elevation in plasma corticosterone levels however it increases serotonin synthesis by virtue of inhibiting hepatic TDO enzyme activity and its gene expression, ascertaining the notion that there exists an inverse relationship between hepatic TDO enzyme activity and brain 5-HT. The drug also decreases serotonin turnover in all the brain areas (hypothalamus, hippocampus amygdala) in stressed rats endorsing its monoamine oxidase inhibition property. Inhibition of TDO enzyme activity and its gene expression by the drug provides new insights for the development of therapeutic interventions for stress related mental illnesses.

  14. Impact of hyperlipidemia on plasma protein binding and hepatic drug transporter and metabolic enzyme regulation in a rat model of gestational diabetes.

    Science.gov (United States)

    Anger, Gregory J; Piquette-Miller, Micheline

    2010-07-01

    It is currently unknown whether gestational diabetes mellitus (GDM), a prevalent obstetrical complication, compounds the changes in drug disposition that occur naturally in pregnancy. Hyperlipidemia occurs in GDM. Using a rat model of GDM, we determined whether excess lipids compete with drugs for plasma protein binding. Because lipids activate nuclear receptors that regulate drug transporters and metabolic enzymes, we used proteome analysis to determine whether hyperlipidemia indirectly leads to the dysregulation of these proteins in the liver. GDM was induced on gestational day 6 (GD6) via streptozotocin injection. Controls received either vehicle alone or streptozotocin with subsequent insulin treatment. Liver and plasma were collected on GD20. Glyburide and saquinavir protein binding was determined by ultrafiltration, and an established solvent method was used for plasma delipidation. Proteomics analysis was performed by using isobaric tags for relative and absolute quantitation methodology with membrane-enriched hepatic protein samples. Relative to controls, GDM rat plasma contained more cholesterol and triglycerides. Plasma protein binding of glyburide and saquinavir was decreased in GDM. Delipidation normalized protein binding in GDM plasma. Proteins linked to lipid metabolism were strongly affected in the GDM proteomics data set, with prohyperlipidemic and antihyperlipidemic changes observed, and formed networks that implicated several nuclear receptors. Up-regulation of drug transporters and metabolic enzymes was observed (e.g., multidrug resistance 1/2, CYP2A1, CYP2B9, and CYP2D3). In this study, GDM-induced hyperlipidemia decreased protein binding and was associated with drug transporter and metabolic enzyme up-regulation in the liver. Both of these findings could change drug disposition in affected pregnancies, compounding changes associated with pregnancy itself.

  15. ANTIOXIDANT ENZYME ACTIVITY AND FRESH-CUT ARRACACHA QUALITY

    Directory of Open Access Journals (Sweden)

    Hêmina Carla Vilela

    2015-06-01

    Full Text Available The arracacha is an alternative of fresh-cut product; however it can be easily degraded after the processing techniques. The objective of this work was to evaluate the useful life of fresh-cut arracacha submitted to two types of cuts and storage, as well as to evaluate the activity of antioxidant enzymes. The roots were selected, sanitized and submitted to two cut types: cubed and grated. Then they were evaluated at 3 times: 0, 3 and 7 days. The cutting in cubes provided higher quality and lower SOD, CAT and APX activity. However, the grated product presented higher PG activity and lower PPO activity. The microbiological safety and the nutritional value were maintained in both cuts during the whole storage period. The useful life, regarding the physicochemical, nutritional and microbiological aspects, can be established at 7 days under refrigeration for fresh-cut arracacha.

  16. Relationship between Estradiol and Antioxidant Enzymes Activity of Ischemic Stroke

    Directory of Open Access Journals (Sweden)

    Nasrin Sheikh

    2009-01-01

    Full Text Available Some evidence suggests the neuroprotection of estrogen provided by the antioxidant activity of this compound. The main objective of this study was to determine the level of estradiol and its correlation with the activity of antioxidant enzymes, total antioxidant status and ferritin from ischemic stroke subjects. The study population consisted of 30 patients with acute ischemic stroke and 30 controls. There was no significant difference between estradiol in stroke and control group. The activity of superoxide dismutase and level of ferritin was higher in stroke compared with control group (<.05, <.001, resp.. There was no significant correlation between estradiol and glutathione peroxidase, glutathione reductase, catalase, total antioxidant status, and ferritin in stroke and control groups. We observed inverse correlation between estradiol with superoxide dismutase in males of stroke patients (=−0.54, =.029. Our results supported that endogenous estradiol of elderly men and women of stroke or control group has no antioxidant activity.

  17. Protoplast fusion enhances lignocellulolytic enzyme activities in Trichoderma reesei.

    Science.gov (United States)

    Cui, Yu-xiao; Liu, Jia-jing; Liu, Yan; Cheng, Qi-yue; Yu, Qun; Chen, Xin; Ren, Xiao-dong

    2014-12-01

    Protoplast fusion was used to obtain a higher production of lignocellulolytic enzymes with protoplast fusion in Trichoderma reesei. The fusant strain T. reesei JL6 was obtained from protoplast fusion from T. reesei strains QM9414, MCG77, and Rut C-30. Filter paper activity of T. reesei JL6 increased by 18% compared with that of Rut C-30. β-Glucosidase, hemicellulase and pectinase activities of T. reesei JL6 were also higher. The former activity was 0.39 Uml(-1), while those of QM9414, MCG77, and Rut C-30 were 0.13, 0.11, and 0.16 Uml(-1), respectively. Pectinase and hemicellulase activities of JL6 were 5.4 and 15.6 Uml(-1), respectively, which were slightly higher than those of the parents. The effects of corn stover and wheat bran carbon sources on the cellulase production and growth curve of T. reesei JL6 were also investigated.

  18. ACTIVITY OF SUPEROXIDE DISMUTASE ENZYME IN YEAST SACCHAROMYCES CEREVISIAE

    Directory of Open Access Journals (Sweden)

    Blažena Lavová

    2014-02-01

    Full Text Available Reactive oxygen species (ROS with reactive nitrogen species (RNS are known to play dual role in biological systems, they can be harmful or beneficial to living systems. ROS can be important mediators of damage to cell structures, including proteins, lipids and nucleic acids termed as oxidative stress. The antioxidant enzymes protect the organism against the oxidative damage caused by active oxygen forms. The role of superoxide dismutase (SOD is to accelerate the dismutation of the toxic superoxide radical, produced during oxidative energy processes, to hydrogen peroxide and molecular oxygen. In this study, SOD activity of three yeast strains Saccharomyces cerevisiae was determined. It was found that SOD activity was the highest (23.7 U.mg-1 protein in strain 612 after 28 hours of cultivation. The lowest SOD activity from all tested strains was found after 56 hours of cultivation of strain Gyöng (0.7 U.mg-1 protein.

  19. Understanding drivers of peatland extracellular enzyme activity in the PEATcosm experiment: mixed evidence for enzymic latch hypothesis

    Science.gov (United States)

    Karl J. Romanowicz; Evan S. Kane; Lynette R. Potvin; Aleta L. Daniels; Randy Kolka; Erik A. Lilleskov

    2015-01-01

    Aims. Our objective was to assess the impacts of water table position and plant functional groups on peatland extracellular enzyme activity (EEA) framed within the context of the enzymic latch hypothesis. Methods. We utilized a full factorial experiment with 2 water table (WT) treatments (high and low) and 3 plant functional...

  20. An Extended Polyanion Activation Surface in Insulin Degrading Enzyme.

    Directory of Open Access Journals (Sweden)

    Eun Suk Song

    Full Text Available Insulin degrading enzyme (IDE is believed to be the major enzyme that metabolizes insulin and has been implicated in the degradation of a number of other bioactive peptides, including amyloid beta peptide (Aβ, glucagon, amylin, and atrial natriuretic peptide. IDE is activated toward some substrates by both peptides and polyanions/anions, possibly representing an important control mechanism and a potential therapeutic target. A binding site for the polyanion ATP has previously been defined crystallographically, but mutagenesis studies suggest that other polyanion binding modes likely exist on the same extended surface that forms one wall of the substrate-binding chamber. Here we use a computational approach to define three potential ATP binding sites and mutagenesis and kinetic studies to confirm the relevance of these sites. Mutations were made at four positively charged residues (Arg 429, Arg 431, Arg 847, Lys 898 within the polyanion-binding region, converting them to polar or hydrophobic residues. We find that mutations in all three ATP binding sites strongly decrease the degree of activation by ATP and can lower basal activity and cooperativity. Computational analysis suggests conformational changes that result from polyanion binding as well as from mutating residues involved in polyanion binding. These findings indicate the presence of multiple polyanion binding modes and suggest the anion-binding surface plays an important conformational role in controlling IDE activity.

  1. A Computational Methodology to Screen Activities of Enzyme Variants

    CERN Document Server

    Hediger, Martin R; Svendsen, Allan; Besenmatter, Werner; Jensen, Jan H

    2012-01-01

    We present a fast computational method to efficiently screen enzyme activity. In the presented method, the effect of mutations on the barrier height of an enzyme-catalysed reaction can be computed within 24 hours on roughly 10 processors. The methodology is based on the PM6 and MOZYME methods as implemented in MOPAC2009, and is tested on the first step of the amide hydrolysis reaction catalyzed by Candida Antarctica lipase B (CalB) enzyme. The barrier heights are estimated using adiabatic mapping and are shown to give barrier heights to within 3kcal/mol of B3LYP/6-31G(d)//RHF/3-21G results for a small model system. Relatively strict convergence criteria (0.5kcal/(mol{\\AA})), long NDDO cutoff distances within the MOZYME method (15{\\AA}) and single point evaluations using conventional PM6 are needed for reliable results. The generation of mutant structure and subsequent setup of the semiempirical calculations are automated so that the effect on barrier heights can be estimated for hundreds of mutants in a matte...

  2. Tissue enzyme activities in the loggerhead sea turtle (Caretta caretta).

    Science.gov (United States)

    Anderson, Eric T; Socha, Victoria L; Gardner, Jennifer; Byrd, Lynne; Manire, Charles A

    2013-03-01

    The loggerhead sea turtle, Caretta caretta, one of the seven species of threatened or endangered sea turtles worldwide, is one of the most commonly encountered marine turtles off the eastern coast of the United States and Gulf of Mexico. Although biochemical reference ranges have been evaluated for several species of sea turtles, tissue specificity of the commonly used plasma enzymes is lacking. This study evaluated the tissue specificity of eight enzymes, including amylase, lipase, creatine kinase (CK), gamma-glutamyl transferase (GGT), alkaline phosphatase (ALP), lactate dehydrogenase (LDH), aspartate aminotransferase (AST), and alanine aminotransferase (ALT), in 30 tissues from five stranded loggerhead sea turtles with no evidence of infectious disease. Amylase and lipase showed the greatest tissue specificity, with activity found only in pancreatic samples. Creatine kinase had high levels present in skeletal and cardiac muscle, and moderate levels in central nervous system and gastrointestinal samples. Gamma-glutamyl transferase was found in kidney samples, but only in very low levels. Creatine kinase, ALP, AST, and LDH were found in all tissues evaluated and ALT was found in most, indicating low tissue specificity for these enzymes in the loggerhead.

  3. A computational methodology to screen activities of enzyme variants.

    Directory of Open Access Journals (Sweden)

    Martin R Hediger

    Full Text Available We present a fast computational method to efficiently screen enzyme activity. In the presented method, the effect of mutations on the barrier height of an enzyme-catalysed reaction can be computed within 24 hours on roughly 10 processors. The methodology is based on the PM6 and MOZYME methods as implemented in MOPAC2009, and is tested on the first step of the amide hydrolysis reaction catalyzed by the Candida Antarctica lipase B (CalB enzyme. The barrier heights are estimated using adiabatic mapping and shown to give barrier heights to within 3 kcal/mol of B3LYP/6-31G(d//RHF/3-21G results for a small model system. Relatively strict convergence criteria (0.5 kcal/(molÅ, long NDDO cutoff distances within the MOZYME method (15 Å and single point evaluations using conventional PM6 are needed for reliable results. The generation of mutant structures and subsequent setup of the semiempirical calculations are automated so that the effect on barrier heights can be estimated for hundreds of mutants in a matter of weeks using high performance computing.

  4. Activation of Variants of Hypoxanthine-Guanine Phosphoribosyl Transferase by the Normal Enzyme

    Science.gov (United States)

    Bakay, Bohdan; Nyhan, William L.

    1972-01-01

    Deficient hypoxanthine-guanine phosphoribosyl transferase (HGPRT; EC 2.4.2.8) enzymes from erythrocytes of patients with hyperuricemia and with the Lesch-Nyhan syndrome migrate 15% faster in polyacrylamide gel disc electrophoresis than the normal enzyme. A half-sister of two males with partial deficiency, who had 34% of normal HGPRT activity in her erythrocytes, yielded profiles containing two distinct zones of activity; one corresponded to the enzyme found in normal individuals and one to the variant of her half-brothers. However, in her profile her variant enzyme showed notably greater activity than that observed in her half-brothers. This increase was due to an activation of the variant by normal enzyme. Electrophoresis of mixtures of normal enzyme with partially deficient enzymes from patients with hyperuricemia and with the Lesch-Nyhan syndrome also led to activation of deficient HGPRT variants by normal enzymes. Deficient variants were also activated by normal enzyme on filtration through Sephadex G-25. Experiments in which deficient variant enzymes were activated with purified normal enzyme labeled with 125I indicated that deficient enzymes incorporate components of the normal enzyme. No such activation of deficient enzymes was ever obtained when mixtures of deficient and normal enzymes were put together in a test tube. Images PMID:4341698

  5. The slowed brain: cortical oscillatory activity in hepatic encephalopathy.

    Science.gov (United States)

    Butz, Markus; May, Elisabeth S; Häussinger, Dieter; Schnitzler, Alfons

    2013-08-15

    Oscillatory activity of the human brain has received growing interest as a key mechanism of large-scale integration across different brain regions. Besides a crucial role of oscillatory activity in the emergence of other neurological and psychiatric diseases, recent evidence indicates a key role in the pathophysiology of hepatic encephalopathy (HE). This review summarizes the current knowledge on pathological alterations of oscillatory brain activity in association with liver dysfunction and HE in the context of spontaneous brain activity, motor symptoms, sensory processing, and attention. The existing literature demonstrates a prominent slowing of the frequency of oscillatory activity as shown for spontaneous brain activity at rest, with respect to deficits of motor behavior and motor symptoms, and in the context of visual attention processes. The observed slowing extends across different subsystems of the brain and has been confirmed across different frequency bands, providing evidence for ubiquitous changes of oscillatory activity in HE. For example, the frequency of cortico-muscular coherence in HE patients appears at the frequency of the mini-asterixis (⩽12Hz), while cirrhotics without overt signs of HE show coherence similar to healthy subjects, i.e. at 13-30Hz. Interestingly, the so-called critical flicker frequency (CFF) as a measure of the processing of an oscillating visual stimulus has emerged as a useful tool to quantify HE disease severity, correlating with behavioral and neurophysiological alterations. Moreover, the CFF reliably distinguishes patients with manifest HE from cirrhotics without any signs of HE and healthy controls using a cut-off frequency of 39Hz. In conclusion, oscillatory activity is globally slowed in HE in close association with HE symptoms and disease severity. Although the underlying causal mechanisms are not yet understood, these results indicate that pathological changes of oscillatory activity play an important role in the

  6. ANTIMICROBIAL ACTIVITY AND BIODEGRADING ENZYMES OF ENDOPHYTIC FUNGI FROM EUCALYPTUS

    Directory of Open Access Journals (Sweden)

    K. Ananda et al

    2012-08-01

    Full Text Available A total of thirty endophytic fungi were isolated from leaves and twigs of Eucalyptus globulus and Eucalyptus citriodora. Among thirty endophytic fungal isolates, four (P3MT1, P3MT2, OP4MT2 and P7ML2 are consistently producing compounds which are inhibiting Pseudomonas aeroginosa, Mycobacterium smegmatis and Candida albicans even after 10 generations tested under dual culture, well diffusion and disc diffusion methods. P3MT1 and OP4MT2 are inhibiting even a filamentous fungi Penicillium chrysogenum. The fungal isolate OP4MT2 showed highest zone of inhibition (20 mm against Penicillium chrysogenum among two test fungi. The crude ethyl acetate extract of P3MT1 isolate showed highest zone of inhibition against Candida albicans (19 mm by both well and disc diffusion method when compared to other fungal isolates. Another four fungal isolates (P3ML1, P6MT1, P5MT1 and P2MT1 from the same set of thirty isolates showed positive for the secretion of amylase, protease and laccase enzymes in agar plate method. Two endophytic fungal isolates (P6MT1 & P2MT1 among thirty are able to oxidize guaiacol indicating the presence of Lignin degrading enzymes. Four fungal isolates indicated presence of laccase enzymes by qualitative test were able to decolorize both methylene blue and aniline blue (synthetic dyes in solid and liquid media. The quantitative estimation of percent decolorization of synthetic dyes by spectrophotometric method confirmed more than 90 % reduction in color is made possible by the endophytic fungi. All these fungal strains with good bioactivity are of worth studying in detail for the purification and characterization of the active compounds and enzymes.

  7. Global Profiling of Carbohydrate Active Enzymes in Human Gut Microbiome.

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    Tanudeep Bhattacharya

    Full Text Available Carbohydrate Active enzyme (CAZyme families, encoded by human gut microflora, play a crucial role in breakdown of complex dietary carbohydrates into components that can be absorbed by our intestinal epithelium. Since nutritional wellbeing of an individual is dependent on the nutrient harvesting capability of the gut microbiome, it is important to understand how CAZyme repertoire in the gut is influenced by factors like age, geography and food habits.This study reports a comprehensive in-silico analysis of CAZyme profiles in the gut microbiomes of 448 individuals belonging to different geographies, using similarity searches of the corresponding gut metagenomic contigs against the carbohydrate active enzymes database. The study identifies a core group of 89 CAZyme families that are present across 85% of the gut microbiomes. The study detects several geography/age-specific trends in gut CAZyme repertoires of the individuals. Notably, a group of CAZymes having a positive correlation with BMI has been identified. Further this group of BMI-associated CAZymes is observed to be specifically abundant in the Firmicutes phyla. One of the major findings from this study is identification of three distinct groups of individuals, referred to as 'CAZotypes', having similar CAZyme profiles. Distinct taxonomic drivers for these CAZotypes as well as the probable dietary basis for such trends have also been elucidated. The results of this study provide a global view of CAZyme profiles across individuals of various geographies and age-groups. These results reiterate the need of a more precise understanding of the role of carbohydrate active enzymes in human nutrition.

  8. Activated hepatic stellate cells in liver cirrhosis. A morphologic and morphometrical study.

    Science.gov (United States)

    Carpino, Guido; Franchitto, Antonio; Morini, Sergio; Corradini, Stefano Ginanni; Merli, Manuela; Gaudio, Eugenio

    2004-01-01

    Hepatic stellate cells have been considered the most important cell-type involved in hepatic fibrogenesis. Proliferation and differentiation of hepatic stellate cells into myofibroblast-like cells has been related to the development of liver fibrosis. The alpha-actin expressed by hepatic stellate cells was considered a marker of their activation to myofibroblast-like cell. The aim of this study is to evaluate the changes in morphology, distribution, percentage and alpha-smooth muscle actin expression of hepatic stellate cells in normal and cirrhotic livers, and to correlate activated hepatic stellate cells with the progression of fibrosis. Human liver biopsies (n=121) were divided in five groups: 1) normal livers (controls); 2) cirrhosis post-HCV hepatitis; 3) cirrhosis post-HBV hepatitis; 4) non viral related cirrhosis; 5) recurrent HCV hepatitis after orthotopic liver transplantation. Samples immunostained with anti alpha-smooth muscle actin antibody by immunoperoxidase method were semi-quantitatively evaluated. Liver fibrosis was quantified by computer image analysis on specimens stained with Masson's trichrome. In normal adult livers stellate cells were very rarely stained for alpha-smooth muscle actin. In cirrhotic livers, a strongly enhanced percentage of stellate cells expressing alpha-smooth muscle actin was detected in cirrhotic fragments with respect to the control group, with a significant correlation between alpha-smooth muscle actin positive stellate cells and the volume fraction of fibrosis. Moreover, liver biopsies of recurrent hepatitis revealed an increased number of activated stellate cells compared to normal livers, and intermediate volume fraction of fibrosis. These results confirmed that a direct correlation existed between activated stellate cells and the progression of fibrosis. Alpha-smooth muscle actin confirmed to be a reliable marker of hepatic stellate cells activation also in precocious stages of the disease.

  9. Virtual Biochemistry – pH effect on enzyme activity

    Directory of Open Access Journals (Sweden)

    D.N. Heidrich

    2011-04-01

    Full Text Available Protocols of laboratory experiments, followed by teacher's explanation, not always clearly translate to the student the dynamics to beadopted for the implementation of the proposed practice. One of these cases is related to the study of the effect of pH on enzyme activity. For better help the understanding of the technical procedure, a hypermedia was built based on a protocol adopted at the Department of Biochemistry, UFSC. The hypermedia shows how theeffect of variations in pH can be observed  in vitro. Taking as example salivary amylase and the consumption of starch (substrate by means of iodine staining, a set of pH buffers was tested to identify the best pH for this enzyme  activity. This hypermedia as introductory tool for such practice was tested on aNutrition course classroom. Students agree that the hypermedia provided a better understanding of the proposed activities. Teachers also notice a smallerreagents consumption and reduction of the time spent by the students in the achievement of the experiment.

  10. Protein stability and enzyme activity at extreme biological temperatures

    Energy Technology Data Exchange (ETDEWEB)

    Feller, Georges, E-mail: gfeller@ulg.ac.b [Laboratory of Biochemistry, Centre for Protein Engineering, Institute of Chemistry B6a, University of Liege, B-4000 Liege (Belgium)

    2010-08-18

    Psychrophilic microorganisms thrive in permanently cold environments, even at subzero temperatures. To maintain metabolic rates compatible with sustained life, they have improved the dynamics of their protein structures, thereby enabling appropriate molecular motions required for biological activity at low temperatures. As a consequence of this structural flexibility, psychrophilic proteins are unstable and heat-labile. In the upper range of biological temperatures, thermophiles and hyperthermophiles grow at temperatures > 100 {sup 0}C and synthesize ultra-stable proteins. However, thermophilic enzymes are nearly inactive at room temperature as a result of their compactness and rigidity. At the molecular level, both types of extremophilic proteins have adapted the same structural factors, but in opposite directions, to address either activity at low temperatures or stability in hot environments. A model based on folding funnels is proposed accounting for the stability-activity relationships in extremophilic proteins. (topical review)

  11. Extracellular Enzyme Activity assay as indicator of soil microbial functional diversity and activity

    DEFF Research Database (Denmark)

    Hendriksen, Niels Bohse; Winding, Anne

    2012-01-01

    Extracellular Enzyme Activity assay as indicator of soil microbial functional diversity and activity Niels Bohse Hendriksen, Anne Winding. Department of Environmental Science, Aarhus University, 4000 Roskilde, Denmark Soil enzymes originate from a variety of organisms, notably fungi and bacteria...... and especially hydrolytic extracellular enzymes are of pivotal importance for decomposition of organic substrates and biogeochemical cycling. Their activity reflects the functional diversity and activity of the microorganisms involved in decomposition processes which are essential processes for soil functioning......, experimental conditions of extraction of enzymes from soils, buffer and pH, substrate concentration, temperature and the necessary controls were optimized and standardized. This has resulted in an optimized standard operating procedure of EEA, which are being tested as an indicator of soil functional diversity...

  12. Saturated fatty acids activate ERK signaling to downregulate hepatic sortilin 1 in obese and diabetic mice.

    Science.gov (United States)

    Bi, Lipeng; Chiang, John Y L; Ding, Wen-Xing; Dunn, Winston; Roberts, Benjamin; Li, Tiangang

    2013-10-01

    Hepatic VLDL overproduction is a characteristic feature of diabetes and an important contributor to diabetic dyslipidemia. Hepatic sortilin 1 (Sort1), a cellular trafficking receptor, is a novel regulator of plasma lipid metabolism and reduces plasma cholesterol and triglycerides by inhibiting hepatic apolipoprotein B production. Elevated circulating free fatty acids play key roles in hepatic VLDL overproduction and the development of dyslipidemia. This study investigated the regulation of hepatic Sort1 in obesity and diabetes and the potential implications in diabetic dyslipidemia. Results showed that hepatic Sort1 protein was markedly decreased in mouse models of type I and type II diabetes and in human individuals with obesity and liver steatosis, whereas increasing hepatic Sort1 expression reduced plasma cholesterol and triglycerides in mice. Mechanistic studies showed that the saturated fatty acid palmitate activated extracellular signal-regulated kinase (ERK) and inhibited Sort1 protein by mechanisms involving Sort1 protein ubiquitination and degradation. Consistently, hepatic ERK signaling was activated in diabetic mice, whereas blocking ERK signaling by an ERK inhibitor increased hepatic Sort1 protein in mice. These results suggest that increased saturated fatty acids downregulate liver Sort1 protein, which may contribute to the development of dyslipidemia in obesity and diabetes.

  13. Repressor and activator protein accelerates hepatic ischemia reperfusion injury by promoting neutrophil inflammatory response.

    Science.gov (United States)

    Li, Chang Xian; Lo, Chung Mau; Lian, Qizhou; Ng, Kevin Tak-Pan; Liu, Xiao Bing; Ma, Yuen Yuen; Qi, Xiang; Yeung, Oscar Wai Ho; Tergaonkar, Vinay; Yang, Xin Xiang; Liu, Hui; Liu, Jiang; Shao, Yan; Man, Kwan

    2016-05-10

    Repressor and activator protein (Rap1) directly regulates nuclear factor-κB (NF-κB) dependent signaling, which contributes to hepatic IRI. We here intended to investigate the effect of Rap1 in hepatic ischemia reperfusion injury (IRI) and to explore the underlying mechanisms. The association of Rap1 expression with hepatic inflammatory response were investigated in both human and rat liver transplantation. The effect of Rap1 in hepatic IRI was studied in Rap1 knockout mice IRI model in vivo and primary cells in vitro. Our results showed that over expression of Rap1 was associated with severe liver graft inflammatory response, especially in living donor liver transplantation. The results were also validated in rat liver transplantation model. In mice hepatic IRI model, the knockout of Rap1 reduced hepatic damage and hepatic inflammatory response. In primary cells, the knockout of Rap1 suppressed neutrophils migration activity and adhesion in response to liver sinusoidal endothelial cells through down-regulating neutrophils F-Actin expression and CXCL2/CXCR2 pathway. In addition, the knockout of Rap1 also decreased production of pro-inflammatory cytokines/chemokines in primary neutrophils and neutrophils-induced hepatocyte damage. In conclusion, Rap1 may induce hepatic IRI through promoting neutrophils inflammatory response. Rap1 may be the potential therapeutic target of attenuating hepatic IRI.

  14. The ameliorating effects of vitamin E on hepatic antioxidant system and xenobiotic-metabolizing enzymes in fenvalerate-exposed iodine-deficient rats.

    Science.gov (United States)

    Kocer-Gumusel, Belma; Erkekoglu, Pinar; Caglayan, Aydan; Hincal, Filiz

    2016-01-01

    This study investigated the effects of vitamin E (VE) on hepatic antioxidant system and drug-metabolizing enzymes in fenvalerate (FEN)-exposed iodine-deficient (ID) Wistar rats. ID was produced by perchlorate containing drinking water. VE was introduced by a loading dose of 100 mg/kg/d, i.g. for the first three days in the last week of feeding period; then with a single maintenance dose of 40 mg/kg on the 4th day. During last week, FEN groups (F) received 100 mg/kg/d, i.p. FEN. VE alone did not significantly affect thyroid hormones and antioxidant parameters; however, significantly increased total cytochrome P450 (38%) and cytochrome b5 levels (36%). In all ID groups, plasma thyroid-stimulating hormone (TSH) levels increased markedly, but remained at control level in vitamin E plus FEN receiving iodine-deficient group (IDVF) group. Glutathione peroxidase activity showed marked increases in F (19%) and FEN-exposed iodine-deficient group (IDF, 48%) groups. FEN treatment significantly increased total cytochrome P450 (28%) and thiobarbituric acid reactive substance levels (36%), as well as 7-ethoxyresorufin O-deethylase (120%), 7-penthoxyresorufin O-deethylase (139%) and glutathione S-transferase (15%) activities and decreased total glutathione concentrations (28%) versus control. Overall results suggest that vitamin E has ameliorating effects on the measured parameters in ID and/or FEN exposure.

  15. Antiviral activity of glycyrrhizin against hepatitis C virus in vitro.

    Directory of Open Access Journals (Sweden)

    Yoshihiro Matsumoto

    Full Text Available Glycyrrhizin (GL has been used in Japan to treat patients with chronic viral hepatitis, as an anti-inflammatory drug to reduce serum alanine aminotransferase levels. GL is also known to exhibit various biological activities, including anti-viral effects, but the anti-hepatitis C virus (HCV effect of GL remains to be clarified. In this study, we demonstrated that GL treatment of HCV-infected Huh7 cells caused a reduction of infectious HCV production using cell culture-produced HCV (HCVcc. To determine the target step in the HCV lifecycle of GL, we used HCV pseudoparticles (HCVpp, replicon, and HCVcc systems. Significant suppressions of viral entry and replication steps were not observed. Interestingly, extracellular infectivity was decreased, and intracellular infectivity was increased. By immunofluorescence and electron microscopic analysis of GL treated cells, HCV core antigens and electron-dense particles had accumulated on endoplasmic reticulum attached to lipid droplet (LD, respectively, which is thought to act as platforms for HCV assembly. Furthermore, the amount of HCV core antigen in LD fraction increased. Taken together, these results suggest that GL inhibits release of infectious HCV particles. GL is known to have an inhibitory effect on phospholipase A2 (PLA2. We found that group 1B PLA2 (PLA2G1B inhibitor also decreased HCV release, suggesting that suppression of virus release by GL treatment may be due to its inhibitory effect on PLA2G1B. Finally, we demonstrated that combination treatment with GL augmented IFN-induced reduction of virus in the HCVcc system. GL is identified as a novel anti-HCV agent that targets infectious virus particle release.

  16. Peroxisome Proliferator-Activated Receptor α Activation Induces Hepatic Steatosis, Suggesting an Adverse Effect

    OpenAIRE

    Fang Yan; Qi Wang; Chao Xu; Mingfeng Cao; Xiaoming Zhou; Tingting Wang; Chunxiao Yu; Fei Jing; Wenbin Chen; Ling Gao; Jiajun Zhao

    2014-01-01

    Non-alcoholic fatty liver disease (NAFLD) is characterized by hepatic triglyceride accumulation, ranging from steatosis to steatohepatitis and cirrhosis. NAFLD is a risk factor for cardiovascular diseases and is associated with metabolic syndrome. Antihyperlipidemic drugs are recommended as part of the treatment for NAFLD patients. Although fibrates activate peroxisome proliferator-activated receptor α (PPARα), leading to the reduction of serum triglyceride levels, the effects of these drugs ...

  17. Effect of Pesticides on soil microbial and enzyme activity

    Institute of Scientific and Technical Information of China (English)

    WANG Lan; LI Xiao-hui

    2008-01-01

    Objective Pesticides has gain an increasing awareness because of it is becoming a serious environmental problem and come to threaten the health of humanbeing. The effect of five pesticides (zineb, copforce, the mixture of earbendazim and mancozeb, hymexazol) on soil bacteria, fungi, actinomyces, and Five specific enzymes were chosen for investigation (urease, dehydrogenase, invertase, acid phosphates and protease). Methods The enumeration of the soil micro flora was done by the dilution plate method; The enzyme activity was determined by traditional methods. Shannon-Wiener index as well as 16S rRNA-PCR amplification and DGGE fingerprinting was used for detection of shift in microbial community diversity in pesticides contaminated agricultural soil. Results The outcome showed that the microbial diversity was significantly changed after the application of pesticides, the effect of pesticides on microbe had a order from top to bottom:bacteria-actinomyces-fungi. Conclusions Our results indicate that the use of the pesticides hymexazol resulted in an altered soil community structure, in particular for the actinomyces. Invertase was markedly inhibited by hymexazol, zineb, carbendazim and mancozeb and the inhibiting rates were varied between 30.30 % and 21.21%;Urease activity was also inhibited significantly by hymexazol, the inhibiting rate was 37.67%;Protease activity was markedly inhibited by zineb and hymexazol, the inhibiting rates were 27.27 % and 18.18 % respectively; Phosphates activity was inhibited significantly by hymexazol, zineb, earbendazim and mancozeb, the inhibiting rates were range from 22.12 %-3.54 %; Dehydrogenase activity was not significantly affected by pesticides. Meanwhile, the correlation of all indexes were analyzed, the data suggested that all indexes existed certain correlation.

  18. Hepatoprotective Activity of Cassia fistula root against Carbon tetrachloride-Induced Hepatic Injury in rats (Wistar

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    SAGAR DAWADA

    2012-04-01

    Full Text Available The protective effects of the alcoholic extract of Cassia fistula root; against CCl4 induced hepatic failure in male albino rats (wistar strain was investigated. For acute and massive invasion of hepatopathy, CCl4 (s.c injection of CCl4+Olive Oil in 1:1 ratio; 2ml/kg was used and the insidious intoxication was evidenced bysignificant turmoil of various biochemical parameters followed by significant (p<0.001 weight loss in toxic control group. The administration of alcoholic root extract (200mg/kg and 100mg/kg of body weight for 7 days, elicited protective action since the elevated levels of marker enzymes (SGOT, SGPT, ALP of liver functionswere found to be decreasing progressively in a dose dependent manner. The final body weight was also significantly (p<0.001 increased when compared with the toxic control group. The serum total protein and theserum albumin were also approaching normal values. The results found in alcoholic extract 200mg/kg treated rat were quite promising and were comparable with a standard drug Silymarin. In the alcoholic extract 200mg/kg treated rat group all the marker enzymes were analyzed to be decreasing significantly. The statistically processed results support the conclusion, that the alcoholic root extract of Cassia fistula root (200mg/kg and 100mg/kg possesses dose dependent, significant protective activity against CCl4 induced hepatotoxicity.

  19. Redox regulation of apurinic/apyrimidinic endonuclease 1 activity in Long-Evans Cinnamon rats during spontaneous hepatitis.

    Science.gov (United States)

    Karmahapatra, Soumendra Krishna; Saha, Tapas; Adhikari, Sanjay; Woodrick, Jordan; Roy, Rabindra

    2014-03-01

    The Long-Evans Cinnamon (LEC) rat is an animal model for Wilson's disease. This animal is genetically predisposed to copper accumulation in the liver, increased oxidative stress, accumulation of DNA damage, and the spontaneous development of hepatocellular carcinoma. Thus, this animal model is useful for studying the relationship of endogenous DNA damage to spontaneous carcinogenesis. In this study, we have investigated the apurinic/apyrimidinic endonuclease 1 (APE1)-mediated excision repair of endogenous DNA damage, apurinic/apyrimidinic (AP)-sites, which is highly mutagenic and implicated in human cancer. We found that the activity was reduced in the liver extracts from the acute hepatitis period of LEC rats as compared with extracts from the age-matched Long-Evans Agouti rats. The acute hepatitis period had also a heightened oxidative stress condition as assessed by an increase in oxidized glutathione level and loss of enzyme activity of glyceraldehyde 3-phosphate dehydrogenase, a key redox-sensitive protein in cells. Interestingly, the activity reduction was not due to changes in protein expression but apparently by reversible protein oxidation as the addition of reducing agents to extracts of the liver from acute hepatitis period reactivated APE1 activity and thus, confirmed the oxidation-mediated loss of APE1 activity under increased oxidative stress. These findings show for the first time in an animal model that the repair mechanism of AP-sites is impaired by increased oxidative stress in acute hepatitis via redox regulation which contributed to the increased accumulation of mutagenic AP-sites in liver DNA.

  20. Acute Interstitial Nephritis Proteinuria and Herpes Simplex Virus Hepatitis in Pregnancy Mimic HELLP Syndrome (Hemolysis, Elevated Liver Enzymes, Low Platelets

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    Wendy M. White

    2011-12-01

    Full Text Available Elevated transaminases, hemolysis, and thrombocytopenia in pregnancy are most often caused by a preeclampsia variant—HELLP syndrome (hemolysis, elevated liver enzymes, low platelets. In atypical cases, it is important to consider other causes, such as herpes simplex virus (HSV hepatitis. Acute interstitial nephritis (AIN-induced proteinuria can make distinguishing HELLP from its mimics more difficult. A 43-year-old G4P3 gestational carrier at 28 weeks had abnormal laboratory findings consistent with HELLP, including proteinuria. However, she was normotensive and febrile, prompting an investigation into other possible causes of her signs and symptoms. She ultimately was diagnosed with disseminated HSV infection, started on definitive therapy, and allowed to continue her pregnancy to term. The proteinuria was attributed to AIN. AIN can cause proteinuria in the critically ill pregnant patient. When mimics of HELLP syndrome, such as disseminated HSV infection, are the cause of critical illness, the presence of AIN-induced proteinuria may falsely implicate a hypertensive disorder of pregnancy, resulting in iatrogenic premature delivery of the fetus and failure to initiate definitive potential lifesaving treatment.

  1. Enzymic synthesis and cofactor activity of 3'-pyrophosphocoenzyme A.

    Science.gov (United States)

    Mukai, J I; Sy, J; Lipmann, F

    1983-01-01

    The 3'-pyrophosphate derivative of CoA was synthesized by using the excreted 5'-to-3' pyrophosphoryl-transferring enzyme from Streptomyces adephospholyticus and ATP as donor and dephospho-CoA as acceptor. Cofactor activity of this new coenzyme A derivative was tested with Clostridium kluyveri phosphotransacetylase and hog heart succinic thiokinase. With the phosphotransacetylase, 3'-pyrophospho-CoA was found to be twice as active as CoA whereas dephospho-CoA was inactive. However, succinic thiokinase utilized all three types of CoA equally well. Adenosine 5'-monophosphate 3'-pyrophosphate also was synthesized and used as an analog of adenosine 5'-monophosphate 3'-monophosphate in the dog liver's sulfotransferase-catalyzed sulfate transfer from p-nitrophenyl sulfate to phenol. In contrast to the pyrophospho derivative of coenzyme A, adenosine 5'-monophosphate 3'-pyrophosphate was inactive as a cofactor. PMID:6574458

  2. Two azole fungicides (carcinogenic triadimefon and non-carcinogenic myclobutanil) exhibit different hepatic cytochrome P450 activities in medaka fish.

    Science.gov (United States)

    Lin, Chun-Hung; Chou, Pei-Hsin; Chen, Pei-Jen

    2014-07-30

    Conazoles are a class of imidazole- or triazole-containing drugs commonly used as fungicides in agriculture and medicine. The broad application of azole drugs has led to the contamination of surface aquifers receiving the effluent of municipal or hospital wastewater or agricultural runoff. Several triazoles are rodent carcinogens; azole pollution is a concern to environmental safety and human health. However, the carcinogenic mechanisms associated with cytochrome P450 enzymes (CYPs) of conazoles remain unclear. We exposed adult medaka fish (Oryzias latipes) to continuous aqueous solutions of carcinogenic triadimefon and non-carcinogenic myclobutanil for 7 to 20 days at sub-lethal or environmentally relevant concentrations and assessed hepatic CYP activity and gene expression associated with CYP-mediated toxicity. Both triadimefon and myclobutanil induced hepatic CYP3A activity, but only triadimefon enhanced CYP1A activity. The gene expression of cyp3a38, cyp3a40, pregnane x receptor (pxr), cyp26b, retinoid acid receptor γ1 (rarγ1) and p53 was higher with triadimefon than myclobutanil. As well, yeast-based reporter gene assay revealed that 4 tested conazoles were weak agonists of aryl hydrocarbon receptor (AhR). We reveal differential CYP gene expression with carcinogenic and non-carcinogenic conazoles in a lower vertebrate, medaka fish. Liver CYP-enzyme induction may be a key event in conazole-induced tumorigenesis. This information is essential to evaluate the potential threat of conazoles to human health and fish populations in the aquatic environment.

  3. Overexpression of hepatic plasminogen activator inhibitor type 1 mRNA in rabbits with fatty liver

    Institute of Scientific and Technical Information of China (English)

    Jian-Gao Fan; Liang-Hua Chen; Zheng-Jie Xu; Min-De Zeng

    2001-01-01

    @@ INTRODUCTION Plasminogen activator inhibitor type 1 ( PAI-I ), an approximately Mr 50000 glycoprotein, is the major physiological inhibitor of plasminogen activators. It is not only the priming factor for atherosclerosis and coronary thrombosis[1-3] , but also participates in the genesis of chronic hepatitis and liver fibrosis[4-11] . However, there has been no available report yet about the research of hepatic PAl-1 gene expression in hyperlipidemia and fatty liver. The present study aimed to explore the change of hepatic PAl-1 mRNA and its plasma activity by means of animal model.

  4. Energy Landscape Topography Reveals the Underlying Link Between Binding Specificity and Activity of Enzymes

    Science.gov (United States)

    Chu, Wen-Ting; Wang, Jin

    2016-06-01

    Enzyme activity (often quantified by kcat/Km) is the main function of enzyme when it is active against the specific substrate. Higher or lower activities are highly desired for the design of novel enzyme and drug resistance. However, it is difficult to measure the activities of all possible variants and find the “hot-spot” within the limit of experimental time. In this study, we explore the underlying energy landscape of enzyme-substrate interactions and introduce the intrinsic specificity ratio (ISR), which reflects the landscape topography. By studying two concrete systems, we uncover the statistical correlation between the intrinsic specificity and the enzyme activity kcat/Km. This physics-based concept and method show that the energy landscape topography is valuable for understanding the relationship between enzyme specificity and activity. In addition, it can reveal the underlying mechanism of enzyme-substrate actions and has potential applications on enzyme design.

  5. Feed-drug interaction of orally applied butyrate and phenobarbital on hepatic cytochrome P450 activity in chickens.

    Science.gov (United States)

    Mátis, G; Kulcsár, A; Petrilla, J; Hermándy-Berencz, K; Neogrády, Zs

    2016-08-01

    The expression of hepatic drug-metabolizing cytochrome P450 (CYP) enzymes may be affected by several nutrition-derived compounds, such as by the commonly applied feed additive butyrate, possibly leading to feed-drug interactions. The aim of this study was to provide some evidence if butyrate can alter the activity of hepatic CYPs in chickens exposed to CYP-inducing xenobiotics, monitoring for the first time the possibility of such interaction. Ross 308 chickens in the grower phase were treated with daily intracoelomal phenobarbital (PB) injection (80 mg/kg BW), applied as a non-specific CYP-inducer, simultaneously with two different doses of intra-ingluvial sodium butyrate boluses (0.25 and 1.25 g/kg BW) for 5 days. Activity of CYP2H and CYP3A subfamilies was assessed by specific enzyme assays from isolated liver microsomes. According to our results, the lower dose of orally administered butyrate significantly attenuated the PB-triggered elevation of both hepatic CYP2H and CYP3A activities, which might be in association with the partly common signalling pathways of butyrate and CYP-inducing drugs, such as that of PB. Based on these data, butyrate may take part in pharmacoepigenetic interactions with simultaneously applied drugs or other CYP-inducing xenobiotics, with possible consequences for food safety and pharmacotherapy. Butyrate was found to be capable to maintain physiological CYP activity by attenuating CYP induction, underlining the safety of butyrate application in poultry nutrition. Journal of Animal Physiology and Animal Nutrition © 2015 Blackwell Verlag GmbH.

  6. Metabolic enzyme activities and drug excretion in the small intestine and in the liver in the rat.

    Science.gov (United States)

    Almási, A; Bojcsev, Sz; Fischer, T; Simon, H; Perjési, P; Fischer, Emil

    2013-12-01

    The aim of these experiments was the investigation of the correlation between the metabolic enzyme activities and the intestinal and hepatic excretion of p-nitrophenol (PNP) and its metabolites (PNP-glucuronide: PNP-G and PNP-sulfate: PNP-S) in the same group of rats (n = 10). A jejunal loop was perfused with isotonic medium containing PNP in a concentration of 500 μM. The samples were obtained from the luminal perfusion medium and from the bile. For enzyme assays tissue samples were obtained from the liver and jejunum at the end of experiments. Significant differences were calculated by the Student's t-test. The activity of UDP-glucuronyltransferase and sulfotransferase was about three times higher in the liver than in the small intestine. The activity of the ß-glucuronidase was about six times higher, the activity of the arylsulfatase was approximately seven times greater in the liver than in the jejunum. No significant difference was found between the luminal appearance and the biliary excretion of PNP-G. Contrary to these findings, the biliary excretion of PNP-S was significantly higher than the luminal appearance of PNP-sulfate. It can be concluded that no direct correlation exists between the activity of metabolic enzymes and the excretion rate of PNP-metabolites in the liver and in the jejunal segment of the small intestine.

  7. Acute hypoxia and cytochrome P450-mediated hepatic drug metabolism in humans

    DEFF Research Database (Denmark)

    Jürgens, Gesche; Christensen, Hanne Rolighed; Brøsen, Kim;

    2002-01-01

    Our objective was to investigate the effect of acute hypoxia on the activity of hepatic cytochrome P450 (CYP) enzymes.......Our objective was to investigate the effect of acute hypoxia on the activity of hepatic cytochrome P450 (CYP) enzymes....

  8. The effects of hydroalcoholic extract of Portulaca Oleracea on the serum concentreation of Hepatic enzymes in Rats

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    Ali Zarei

    2014-11-01

    Full Text Available Background Hyperlipidemia can be cause a variety of diseases such as atherosclerosis, diabetes and fatty liver and subsequent liver enzyme increases. The Portulaca Oleracea plant has hypoglycemic and hypolipidemic properties. Therefore, in this study the effect of Portulca Oleracea herb extract on serum liver enzymes including aspartate aminotransferase (SGOT or AST, alanine aminotransferase (SGPT or ALT and alkaline phosphatase (ALP in rats were studied. Materials and Methods: In this experimental study, 60 Wistar rats were divided into 6 groups (n=10. Control group with normal diet, fat diet group and other groups, the experimental group received the same diet plus fat Portulaca oleracea extract maximum dose (800, the mean dose of (400, and a minimum dose of (200 mg / kg or intraperitoneally injection (ip and sort of Atorvastatin (10 mg kg. After the end of this period (21 days, blood sampling was performed and collected data were analyzed using the t and Tukey test, and SPSS software version 11.5. Results Comparison of statistical results indicated that alanine aminotransferase (ALT and Alkaline phosphatase (ALP increase in the control group that received only fatty foods, while the experimental groups received extract of Portulaca Oleracea, and groups receiving Atorvastatin had reduced levels of liver enzymes. Conclusion: Regarding hypoglycemic and hypolipidemic antioxidant activity of the extract and its effect on reducing liver enzymes, plant extracts can be recommended to improve liver function.

  9. County-Scale Spatial Distribution of Soil Enzyme Activities and Enzyme Activity Indices in Agricultural Land: Implications for Soil Quality Assessment

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    Xiangping Tan

    2014-01-01

    Full Text Available Here the spatial distribution of soil enzymatic properties in agricultural land was evaluated on a county-wide (567 km2 scale in Changwu, Shaanxi Province, China. The spatial variations in activities of five hydrolytic enzymes were examined using geostatistical methods. The relationships between soil enzyme activities and other soil properties were evaluated using both an integrated total enzyme activity index (TEI and the geometric mean of enzyme activities (GME. At the county scale, soil invertase, phosphatase, and catalase activities were moderately spatially correlated, whereas urease and dehydrogenase activities were weakly spatially correlated. Correlation analysis showed that both TEI and GME were better correlated with selected soil physicochemical properties than single enzyme activities. Multivariate regression analysis showed that soil OM content had the strongest positive effect while soil pH had a negative effect on the two enzyme activity indices. In addition, total phosphorous content had a positive effect on TEI and GME in orchard soils, whereas alkali-hydrolyzable nitrogen and available potassium contents, respectively, had negative and positive effects on these two enzyme indices in cropland soils. The results indicate that land use changes strongly affect soil enzyme activities in agricultural land, where TEI provides a sensitive biological indicator for soil quality.

  10. RNA editing of hepatitis B virus transcripts by activation-induced cytidine deaminase.

    Science.gov (United States)

    Liang, Guoxin; Kitamura, Kouichi; Wang, Zhe; Liu, Guangyan; Chowdhury, Sajeda; Fu, Weixin; Koura, Miki; Wakae, Kousho; Honjo, Tasuku; Muramatsu, Masamichi

    2013-02-01

    Activation-induced cytidine deaminase (AID) is essential for the somatic hypermutation (SHM) and class-switch recombination (CSR) of Ig genes. The mechanism by which AID triggers SHM and CSR has been explained by two distinct models. In the DNA deamination model, AID converts cytidine bases in DNA into uridine. The uridine is recognized by the DNA repair system, which produces DNA strand breakages and point mutations. In the alternative model, RNA edited by AID is responsible for triggering CSR and SHM. However, RNA deamination by AID has not been demonstrated. Here we found that C-to-T and G-to-A mutations accumulated in hepatitis B virus (HBV) nucleocapsid DNA when AID was expressed in HBV-replicating hepatic cell lines. AID expression caused C-to-T mutations in the nucleocapsid DNA of RNase H-defective HBV, which does not produce plus-strand viral DNA. Furthermore, the RT-PCR products of nucleocapsid viral RNA from AID-expressing cells exhibited significant C-to-T mutations, whereas viral RNAs outside the nucleocapsid did not accumulate C-to-U mutations. Moreover, AID was packaged within the nucleocapsid by forming a ribonucleoprotein complex with HBV RNA and the HBV polymerase protein. The encapsidation of the AID protein with viral RNA and DNA provides an efficient environment for evaluating AID's RNA and DNA deamination activities. A bona fide RNA-editing enzyme, apolipoprotein B mRNA editing catalytic polypeptide 1, induced a similar level of C-to-U mutations in nucleocapsid RNA as AID. Taken together, the results indicate that AID can deaminate the nucleocapsid RNA of HBV.

  11. Neferine inhibits cultured hepatic stellate cell activation and facilitates apoptosis: A possible molecular mechanism.

    Science.gov (United States)

    Ding, Hui; Shi, Jinghong; Wang, Ying; Guo, Jia; Zhao, Juhui; Dong, Lei

    2011-01-10

    Neferine is a major alkaloid component of "Lian Zi Xin", embryos of the seeds of Nelumbo nucifera Gaertner, Nymphaeaceae. Previous studies have shown that neferine has an inhibitory effect on pulmonary fibrosis through its anti-inflammatory and anti-oxidative activities and inhibition of cytokines and NF-κB. However, it is unknown whether neferine also has an inhibitory effect on liver fibrosis through inhibition of TGF-β1 and collagen I and facilitation of apoptosis of hepatic stellate cells. This study examined the effects of neferine on cultured hepatic stellate (HSC-T6) cells and explored its possible action mechanisms by means of MTT assay, enzyme-linked immunosorbent assay, flow-cytometric annexin V-PI assay and Hoechst 33258 staining, as well as real-time PCR and western blotting. The results showed that neferine administration (2, 4, 6, 8 and 10μmol/l) significantly decreased the TGF-β1 and collagen I produced in HSC-T6 cells, and increased the HSC-T6 cell apoptosis in a dose-dependent manner. Neferine treatment for 48h at concentrations of 6 and 10μmol/l significantly increased Bax and caspase 3 mRNAs and proteins, and reduced Bcl2 and alpha-smooth muscle actin (α-SMA) mRNAs and proteins. Our data indicate that neferine efficiently inhibits cultured HSC-T6 cell activation and induces apoptosis by increasing Bax and caspase 3 expression via the mitochondrial pathway.

  12. Phlorotannins from Alaskan Seaweed Inhibit Carbolytic Enzyme Activity

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    Joshua Kellogg

    2014-10-01

    Full Text Available Global incidence of type 2 diabetes has escalated over the past few decades, necessitating a continued search for natural sources of enzyme inhibitors to offset postprandial hyperglycemia. The objective of this study was to evaluate coastal Alaskan seaweed inhibition of α-glucosidase and α-amylase, two carbolytic enzymes involved in serum glucose regulation. Of the six species initially screened, the brown seaweeds Fucus distichus and Alaria marginata possessed the strongest inhibitory effects. F. distichus fractions were potent mixed-mode inhibitors of α-glucosidase and α-amylase, with IC50 values of 0.89 and 13.9 μg/mL, respectively; significantly more efficacious than the pharmaceutical acarbose (IC50 of 112.0 and 137.8 μg/mL, respectively. The activity of F. distichus fractions was associated with phlorotannin oligomers. Normal-phase liquid chromatography-mass spectrometry (NPLC-MS was employed to characterize individual oligomers. Accurate masses and fragmentation patterns confirmed the presence of fucophloroethol structures with degrees of polymerization from 3 to 18 monomer units. These findings suggest that coastal Alaskan seaweeds are sources of α-glucosidase and α-amylase inhibitory phlorotannins, and thus have potential to limit the release of sugar from carbohydrates and thus alleviate postprandial hyperglycemia.

  13. Effect of hepatitis C virus infection on the mRNA expression of drug transporters and cytochrome p450 enzymes in chimeric mice with humanized liver.

    Science.gov (United States)

    Kikuchi, Ryota; McCown, Matthew; Olson, Pamela; Tateno, Chise; Morikawa, Yoshio; Katoh, Yumiko; Bourdet, David L; Monshouwer, Mario; Fretland, Adrian J

    2010-11-01

    The expression of drug transporters and metabolizing enzymes is a primary determinant of drug disposition. Chimeric mice with humanized liver, including PXB mice, are an available model that is permissive to the in vivo infection of hepatitis C virus (HCV), thus being a promising tool for investigational studies in development of new antiviral molecules. To investigate the potential of HCV infection to alter the pharmacokinetics of small molecule antiviral therapeutic agents in PXB mice, we have comprehensively determined the mRNA expression profiles of human ATP-binding cassette (ABC) transporters, solute carrier (SLC) transporters, and cytochrome P450 (P450) enzymes in the livers of these mice under noninfected and HCV-infected conditions. Infection of PXB mice with HCV resulted in an increase in the mRNA expression levels of a series of interferon-stimulated genes in the liver. For the majority of genes involved in drug disposition, minor differences in the mRNA expression of ABC and SLC transporters as well as P450s between the noninfected and HCV-infected groups were observed. The exceptions were statistically significantly higher expression of multidrug resistance-associated protein 4 and organic anion-transporting polypeptide 2B1 and lower expression of organic cation transporter 1 and CYP2D6 in HCV-infected mice. Furthermore, the enzymatic activities of the major human P450s were, in general, comparable in the two experimental groups. These data suggest that the pharmacokinetic properties of small molecule antiviral therapies in HCV-infected PXB mice are likely to be similar to those in noninfected PXB mice. However, caution is needed in the translation of this relationship to HCV-infected patients as the PXB mouse model does not accurately reflect the pathology of patients with chronic HCV infection.

  14. Chronic viral hepatitis C in pediatric age group; assessment of viral activity and hepatic fibrosis by 1H magnetic resonance spectroscopy and diffusion weighted imaging in asymptomatic

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    Shereen Mansour Galal

    2016-09-01

    Conclusion: Early diagnosis of asymptomatic chronic hepatitis C is essential to prevent or delay end stage chronic parenchymal liver disease. 1H MRS may be a potential noninvasive helpful diagnostic tool in the assessment of staging and fibrosis of asymptomatic chronic hepatitis C. The increase in metabolites were correlated with histopathological changes. DW-MRI can be considered as an effective predictor in the assessment of activity in chronic hepatitis C.

  15. Hepatic xenobiotic metabolizing enzyme and transporter gene expression through the life stages of the mouse.

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    Janice S Lee

    Full Text Available BACKGROUND: Differences in responses to environmental chemicals and drugs between life stages are likely due in part to differences in the expression of xenobiotic metabolizing enzymes and transporters (XMETs. No comprehensive analysis of the mRNA expression of XMETs has been carried out through life stages in any species. RESULTS: Using full-genome arrays, the mRNA expression of all XMETs and their regulatory proteins was examined during fetal (gestation day (GD 19, neonatal (postnatal day (PND 7, prepubescent (PND32, middle age (12 months, and old age (18 and 24 months in the C57BL/6J (C57 mouse liver and compared to adults. Fetal and neonatal life stages exhibited dramatic differences in XMET mRNA expression compared to the relatively minor effects of old age. The total number of XMET probe sets that differed from adults was 636, 500, 84, 5, 43, and 102 for GD19, PND7, PND32, 12 months, 18 months and 24 months, respectively. At all life stages except PND32, under-expressed genes outnumbered over-expressed genes. The altered XMETs included those in all of the major metabolic and transport phases including introduction of reactive or polar groups (Phase I, conjugation (Phase II and excretion (Phase III. In the fetus and neonate, parallel increases in expression were noted in the dioxin receptor, Nrf2 components and their regulated genes while nuclear receptors and regulated genes were generally down-regulated. Suppression of male-specific XMETs was observed at early (GD19, PND7 and to a lesser extent, later life stages (18 and 24 months. A number of female-specific XMETs exhibited a spike in expression centered at PND7. CONCLUSIONS: The analysis revealed dramatic differences in the expression of the XMETs, especially in the fetus and neonate that are partially dependent on gender-dependent factors. XMET expression can be used to predict life stage-specific responses to environmental chemicals and drugs.

  16. Effects of Fertilization on Tomato Growth and Soil Enzyme Activity

    Science.gov (United States)

    Mu, Zhen; Hu, Xue-Feng; Cheng, Chang; Luo, Zhi-qing

    2015-04-01

    To study the effects of different fertilizer applications on soil enzyme activity, tomato plant growth and tomato yield and quality, a field experiment on tomato cultivation was carried out in the suburb of Shanghai. Three fertilizer treatments, chemical fertilizer (CF) (N, 260 g/kg; P, 25.71g/kg; K, 83.00g/kg), rapeseed cake manure (CM) (N, 37.4 g/kg; P, 9.0 g/kg; K, 8.46 g/kg), crop-leaf fermenting manure (FM) (N, 23.67 g/kg; P, 6.39 g/kg; K 44.32 g/kg), and a control without using any fertilizers (CK), were designed. The total amounts of fertilizer application to each plot for the CF, CM, FM and CK were 0.6 kg, 1.35 kg, 3.75 kg and 0 kg, respectively, 50% of which were applied as base fertilizer, and another 50% were applied after the first fruit picking as top dressing. Each experimental plot was 9 m2 (1 m × 9 m) in area. Each treatment was replicated for three times. No any pesticides and herbicides were applied during the entire period of tomato growth to prevent their disturbance to soil microbial activities. Soil enzyme activities at each plot were constantly tested during the growing period; the tomato fruit quality was also constantly analyzed and the tomato yield was calculated after the final harvesting. The results were as follows: (1) Urease activity in the soils treated with the CF, CM and FM increased quickly after applying base fertilizer. That with the CF reached the highest level. Sucrase activity was inhibited by the CF and CM to some extent, which was 32.4% and 11.2% lower than that with the CK, respectively; while that with the FM was 15.7% higher than that with the CK. Likewise, catalase activity with the CF increased by 12.3% - 28.6%; that with the CM increased by 87.8% - 95.1%; that with the FM increased by 86.4% - 93.0%. Phosphatase activity with the CF increased rapidly and reached a maximum 44 days after base fertilizer application, and then declined quickly. In comparison, that with the CM and FM increased slowly and reached a maximum

  17. Effect of thymol and carvacrol feed supplementation on performance, antioxidant enzyme activities, fatty acid composition, digestive enzyme activities, and immune response in broiler chickens

    NARCIS (Netherlands)

    Hashemipour, H.; Kermanshahi, H.; Golian, A.; Veldkamp, T.

    2013-01-01

    This trial was conducted to evaluate the effects of dietary supplementation of phytogenic product containing an equal mixture of thymol and carvacrol at 4 levels (0, 60, 100, and 200 mg/kg of diet) on performance, antioxidant enzyme activities, fatty acid composition, digestive enzyme activities, an

  18. Effect of thymol and carvacrol feed supplementation on performance, antioxidant enzyme activities, fatty acid composition, digestive enzyme activities, and immune response in broiler chickens

    NARCIS (Netherlands)

    Hashemipour, H.; Kermanshahi, H.; Golian, A.; Veldkamp, T.

    2013-01-01

    This trial was conducted to evaluate the effects of dietary supplementation of phytogenic product containing an equal mixture of thymol and carvacrol at 4 levels (0, 60, 100, and 200 mg/kg of diet) on performance, antioxidant enzyme activities, fatty acid composition, digestive enzyme activities,

  19. Effect of thymol and carvacrol feed supplementation on performance, antioxidant enzyme activities, fatty acid composition, digestive enzyme activities, and immune response in broiler chickens

    NARCIS (Netherlands)

    Hashemipour, H.; Kermanshahi, H.; Golian, A.; Veldkamp, T.

    2013-01-01

    This trial was conducted to evaluate the effects of dietary supplementation of phytogenic product containing an equal mixture of thymol and carvacrol at 4 levels (0, 60, 100, and 200 mg/kg of diet) on performance, antioxidant enzyme activities, fatty acid composition, digestive enzyme activities, an

  20. Structure-based rational design of peptide hydroxamic acid inhibitors to target tumor necrosis factor-α converting enzyme as potential therapeutics for hepatitis.

    Science.gov (United States)

    Wu, Dan; Gu, Qiuhong; Zhao, Ning; Xia, Fei; Li, Zhiwei

    2015-12-01

    The human tumor necrosis factor-α converting enzyme (TACE) has recently been raised as a new and promising therapeutic target of hepatitis and other inflammatory diseases. Here, we reported a successful application of the solved crystal structure of TACE complex with a peptide-like ligand INN for rational design of novel peptide hydroxamic acid inhibitors with high potency and selectivity to target and inhibit TACE. First, the intermolecular interactions between TACE catalytic domain and INN were characterized through an integrated bioinformatics approach, with which the key substructures of INN that dominate ligand binding were identified. Subsequently, the INN molecular structure was simplified to a chemical sketch of peptide hydroxamic acid compound, which can be regarded as a linear tripeptide capped by a N-terminal carboxybenzyl group (chemically protective group) and a C-terminal hydroxamate moiety (coordinated to the Zn(2+) at TACE active site). Based on the sketch, a virtual combinatorial library containing 180 peptide hydroxamic acids was generated, from which seven samples were identified as promising candidates by using a knowledge-based protein-peptide affinity predictor and were then tested in vitro with a standard TACE activity assay protocol. Consequently, three designed peptide hydroxamic acids, i.e. Cbz-Pro-Ile-Gln-hydroxamic acid, Cbz-Leu-Ile-Val-hydroxamic acid and Cbz-Phe-Val-Met-hydroxamic acid, exhibited moderate or high inhibitory activity against TACE, with inhibition constants Ki of 36 ± 5, 510 ± 46 and 320 ± 26 nM, respectively. We also examined the structural basis and non-bonded profile of TACE interaction with a designed peptide hydroxamic acid inhibitor, and found that the inhibitor ligand is tightly buried in the active pocket of TACE, forming a number of hydrogen bonds, hydrophobic forces and van der Waals contacts at the interaction interface, conferring both stability and specificity for TACE-inhibitor complex

  1. 2-octynoic acid inhibits hepatitis C virus infection through activation of AMP-activated protein kinase.

    Directory of Open Access Journals (Sweden)

    Darong Yang

    Full Text Available Many chronic hepatitis C virus (HCV-infected patients with current therapy do not clear the virus. It is necessary to find novel treatments. The effect of 2-octynoic acid (2-OA on HCV infection in human hepatocytes was examined. The mechanism of 2-OA antiviral activity was explored. Our data showed that 2-OA abrogated lipid accumulation in HCV replicon cells and virus-infected hepatocytes. It suppressed HCV RNA replication and infectious virus production with no cytotoxicity to the host cells. 2-OA did not affect hepatitis B virus replication in HepG2.2.15 cells derived from HepG2 cells transfected with full genome of HBV. Further study demonstrated that 2-OA activated AMP-activated protein kinase (AMPK and inhibited acetyl-CoA carboxylase in viral-infected cells. Compound C, a specific inhibitor of AMPK, inhibited AMPK activity and reversed the reduction of intracellular lipid accumulation and the antiviral effect of 2-OA. Knockdown of AMPK expression by RNA interference abolished the activation of AMPK by 2-OA and blocked 2-OA antiviral activity. Interestingly, 2-OA induced interferon-stimulated genes (ISGs and inhibited microRNA-122 (miR-122 expression in virus-infected hepatocytes. MiR-122 overexpression reversed the antiviral effect of 2-OA. Furthermore, knockdown of AMPK expression reversed both the induction of ISGs and suppression of miR-122 by 2-OA, implying that activated AMPK induces the intracellular innate response through the induction of ISGs and inhibiting miR-122 expression. 2-OA inhibits HCV infection through regulation of innate immune response by activated AMPK. These findings reveal a novel mechanism by which active AMPK inhibits HCV infection. 2-OA and its derivatives hold promise for novel drug development for chronic hepatitis C.

  2. Surface binding sites in carbohydrate active enzymes: An emerging picture of structural and functional diversity

    DEFF Research Database (Denmark)

    Svensson, Birte; Cockburn, Darrell

    2013-01-01

    Carbohydrate active enzymes, particularly those that are active on polysaccharides, are often found associated with carbohydrate binding modules (CBMs), which can play several roles in supporting enzyme function, such as localizing the enzyme to the substrate. However, the presence of CBMs...

  3. 不同蛋白质水平下葡萄糖添加水平对大黄鱼生长性能、糖酵解和糖异生关键酶活性的影响%Effects of Glucose Supplemental Level at Different Protein Levels on Growth Performance, Hepatic Glycolysis and Gluconeogenic Key Enzyme Activities of Large Yellow Croaker (Larmichthys crocea Richardson)

    Institute of Scientific and Technical Information of China (English)

    王猛强; 周飘苹; 黄文文; 周歧存

    2015-01-01

    本试验旨在研究不同蛋白质水平下葡萄糖添加水平对大黄鱼生长性能、血清指标、肝脏糖酵解和糖异生关键酶活性、糖原含量以及消化酶活性的影响。采用2×3双因素试验设计,其中蛋白质水平设42%、48%2个水平,葡萄糖添加水平设10%、20%、30%3个水平,共配制6种试验饲料。每种饲料设3个重复,每个重复放养平均体重为(14.89±0.11) g的大黄鱼幼鱼50尾。试验期为8周。结果表明:饲料蛋白质水平和葡萄糖添加水平的交互作用对大黄鱼的增重率(WGR)和特定生长率(SGR)有显著影响(P0.05)。饲料蛋白质水平和葡萄糖添加水平的交互作用对大黄鱼肝脏葡萄糖激酶( GK)、6-磷酸果糖激酶( PFK)、丙酮酸激酶( PK)、葡萄糖-6-磷酸酶( G6Pase)、果糖-1,6-二磷酸酶( FBPase)及磷酸烯醇式丙酮酸羧激酶( PEPCK)活性有显著影响(P0.05)。由此得出,当蛋白质水平为42%时,随着饲料葡萄糖添加水平的升高,大黄鱼能够通过调节糖酵解和糖异生关键酶活性及肝糖原含量来维持血糖含量的平衡,改善对葡萄糖的利用能力;而当蛋白质水平为48%时,随着饲料葡萄糖添加水平的升高,大黄鱼对葡萄糖的利用能力降低。%An 8-week feeding trial was conducted to evaluate the effects of glucose supplemental level at differ-ent protein levels on growth performance, serum indices, hepatic glycolysis and gluconeogenic key enzyme ac-tivities, glycogen content and digestive enzyme activities of large yellow croaker ( Larmichthys crocea Richard-son) . Six experimental diets were formulated to contain two protein levels ( 42% and 48%, respectively) and three glucose supplemental levels ( 10%, 20% and 30%, respectively) . Each diet was randomly assigned to 3 replicates, and each replicate had 50 juvenile large yellow croaker with the average body weight of (14.89± 0.11) g. The results showed that weight gain rate ( WGR) and specific growth rate

  4. Genetic abolishment of hepatocyte proliferation activates hepatic stem cells.

    Directory of Open Access Journals (Sweden)

    Yoko Endo

    Full Text Available Quiescent hepatic stem cells (HSCs can be activated when hepatocyte proliferation is compromised. Chemical injury rodent models have been widely used to study the localization, biomarkers, and signaling pathways in HSCs, but these models usually exhibit severe promiscuous toxicity and fail to distinguish damaged and non-damaged cells. Our goal is to establish new animal models to overcome these limitations, thereby providing new insights into HSC biology and application. We generated mutant mice with constitutive or inducible deletion of Damaged DNA Binding protein 1 (DDB1, an E3 ubiquitin ligase, in hepatocytes. We characterized the molecular mechanism underlying the compensatory activation and the properties of oval cells (OCs by methods of mouse genetics, immuno-staining, cell transplantation and gene expression profiling. We show that deletion of DDB1 abolishes self-renewal capacity of mouse hepatocytes in vivo, leading to compensatory activation and proliferation of DDB1-expressing OCs. Partially restoring proliferation of DDB1-deficient hepatocytes by ablation of p21, a substrate of DDB1 E3 ligase, alleviates OC proliferation. Purified OCs express both hepatocyte and cholangiocyte markers, form colonies in vitro, and differentiate to hepatocytes after transplantation. Importantly, the DDB1 mutant mice exhibit very minor liver damage, compared to a chemical injury model. Microarray analysis reveals several previously unrecognized markers, including Reelin, enriched in oval cells. Here we report a genetic model in which irreversible inhibition of hepatocyte duplication results in HSC-driven liver regeneration. The DDB1 mutant mice can be broadly applied to studies of HSC differentiation, HSC niche and HSCs as origin of liver cancer.

  5. Peroxisome proliferator-activated receptor alpha controls hepatic heme biosynthesis through ALAS1.

    Science.gov (United States)

    Degenhardt, Tatjana; Väisänen, Sami; Rakhshandehroo, Maryam; Kersten, Sander; Carlberg, Carsten

    2009-05-01

    Heme is an essential prosthetic group of proteins involved in oxygen transport, energy metabolism and nitric oxide production. ALAS1 (5-aminolevulinate synthase) is the rate-limiting enzyme in heme synthesis in the liver and is highly regulated to adapt to the metabolic demand of the hepatocyte. In the present study, we describe human hepatic ALAS1 as a new direct target for the nuclear receptor peroxisome proliferator-activated receptor alpha (PPARalpha). In primary human hepatocytes and in HepG2 cells, PPARalpha agonists induced an increase in ALAS1 mRNA levels, which was abolished by PPARalpha silencing. These effects are mediated by two functional PPAR binding sites at positions -9 and -2.3 kb relative to the ALAS1 transcription start site. PPARalpha ligand treatment also up-regulated the mRNA levels of the genes ALAD (5-aminolevulinate dehydratase), UROS (uroporphyrinogen III synthase), UROD (uroporphyrinogen decarboxylase), CPOX (coproporphyrinogen oxidase) and PPOX (protoporphyrinogen oxidase) encoding for enzymes controlling further steps in heme biosynthesis. In HepG2 cells treated with PPARalpha agonists and in mouse liver upon fasting, the association of PPARalpha, its partner retinoid X receptor, PPARgamma co-activator 1alpha and activated RNA polymerase II with the transcription start site region of all six genes was increased, leading to higher levels of the metabolite heme. In conclusion, these data strongly support a role of PPARalpha in the regulation of human ALAS1 and of five additional genes of the pathway, consequently leading to increased heme synthesis.

  6. Tailor-Made Enzyme Carriers: Preparation and Use of Adsorbents Specifically Designed to Immobilize Allosteric Enzymes in Activated Conformation

    Directory of Open Access Journals (Sweden)

    Zahra Salemi

    2010-01-01

    Full Text Available Problem statement: The enzyme immobilization has experienced substantial growth in the recent past and an ever increasing amount of study has been reported on various aspects of immobilized enzymes. In most of these investigations, catalytic activities are found to be diminished as compared to the enzyme free in solution. Approach: Hydrophobic adsorbents were prepared containing L-leucine or citric acid, two positive allosteric effectors, for bovine liver Glutamate Dehydrogenase (GDH, EC 1.4.1.3 and heart mitochondrial Malate Dehydrogenase (MDH, EC 1.1.1.37 , respectively. Results: Immobilized preparations of these well-defined allosteric enzymes indicated improved catalytic activities as compared with those involving use of the adsorbents without these activators. Conclusion/Recommendations: It is concluded that the regulatory proteins are Furthermore; they retain their natural capacity for undergoing the conformational transitions needed for enhanced catalytic activities. Adsorptive immobilization of these two allosteric proteins in activated conformation may serve as useful models in relation to design strategies for preparation of tailor-made enzyme carriers.

  7. Angiotensin-I converting enzyme inhibitory and antioxidant activities of egg protein hydrolysates produced with gastrointestinal and nongastrointestinal enzymes.

    Science.gov (United States)

    You, Sun-Jong; Wu, Jianping

    2011-08-01

    Egg is a well-known rich source of bioactive peptides. In this study, egg protein (egg white and egg yolk proteins) hydrolysates were produced with gastrointestinal enzymes (pepsin and pancreatin) or nongastrointestinal enzymes (thermolysin and alcalase), and fractionated by ultrafiltration and cation exchange chromatography. Angiotensin-I converting enzyme (ACE) inhibitory and antioxidant activities, amino acid composition and molecular weight distribution were studied, and the physicochemical properties were related with the bioactivities. Our results showed that egg protein hydrolysates produced with non-GI enzymes (thermolysin and alcalase) showed significantly higher ACE inhibitory activity, whereas similar or even lower antioxidative activities, than those of hydrolysates produced with GI enzymes. ACE-inhibitory activity significantly correlated with the amino acid composition, especially the proportion of positively charged amino acid, whereas antioxidant activities correlated with the proportion of low molecular weight peptides under 500 Da. Understanding the relationship between the bioactivities and physicochemical properties of the hydrolysates/fractions is important to facilitate the development technologies for preparing fractions with improved bioactivities. © 2011 Institute of Food Technologists®

  8. Development of radiometric assays for quantification of enzyme activities of the key enzymes of thyroid hormones metabolism.

    Science.gov (United States)

    Pavelka, S

    2014-01-01

    We newly elaborated and adapted several radiometric enzyme assays for the determination of activities of the key enzymes engaged in the biosynthesis (thyroid peroxidase, TPO) and metabolic transformations (conjugating enzymes and iodothyronine deiodinases, IDs) of thyroid hormones (THs) in the thyroid gland and in peripheral tissues, especially in white adipose tissue (WAT). We also elaborated novel, reliable radiometric methods for extremely sensitive determination of enzyme activities of IDs of types 1, 2 and 3 in microsomal fractions of different rat and human tissues, as well as in homogenates of cultured mammalian cells. The use of optimized TLC separation of radioactive products from the unconsumed substrates and film-less autoradiography of radiochromatograms, taking advantage of storage phosphor screens, enabled us to determine IDs enzyme activities as low as 10(-18) katals. In studies of the interaction of fluoxetine (Fluox) with the metabolism of THs, we applied adapted radiometric enzyme assays for iodothyronine sulfotransferases (ST) and uridine 5'-diphospho-glucuronyltransferase (UDP-GT). Fluox is the most frequently used representative of a new group of non-tricyclic antidepressant drugs--selective serotonin re-uptake inhibitors. We used the elaborated assays for quantification the effects of Fluox and for the assessment of the degree of potential induction of rat liver ST and/or UDP-GT enzyme activities by Fluox alone or in combination with T(3). Furthermore, we studied possible changes in IDs activities in murine adipose tissue under the conditions that promoted either tissue hypertrophy (obesogenic treatment) or involution (caloric restriction), and in response to leptin, using our newly developed radiometric enzyme assays for IDs. Our results suggest that deiodinase D1 has a functional role in WAT, with D1 possibly being involved in the control of adipose tissue metabolism and/or accumulation of the tissue. Significant positive correlation between

  9. Gene Cloning, Expression and Enzyme Activity of Vitis vinifera Vacuolar Processing Enzymes (VvVPEs).

    Science.gov (United States)

    Tang, Yujin; Wang, Ruipu; Gong, Peijie; Li, Shuxiu; Wang, Yuejin; Zhang, Chaohong

    2016-01-01

    Vacuolar processing enzymes (VPEs) have received considerable attention due to their caspase-1-like activity and ability to regulate programmed cell death (PCD), which plays an essential role in the development of stenospermocarpic seedless grapes ovules. To characterize VPEs and the relationship between stenospermocarpic grapes and the VPE gene family, we identified 3 Vitis vinifera VPE genes (VvβVPE, VvγVPE, and VvδVPE) from the PN40024 grape genome and cloned the full-length complementary DNAs (cDNAs) from the 'Vitis vinifera cv. Pinot Noir' and 'Vitis vinifera cv. Thompson Seedless' varietals. Each of the VPEs contained a typical catalytic dyad [His (177), Cys (219)] and substrate binding pocket [Arg (112), Arg (389), Ser (395)], except that Ser (395) in the VvγVPE protein sequence was replaced with alanine. Phylogenetic analysis of 4 Arabidopsis thaliana and 6 Vitis vinifera VPEs revealed that the 10 VPEs form 3 major branches. Furthermore, the 6 grapevine VPEs share a similar gene structure, with 9 exons and 8 introns. The 6 grapevine VPEs are located on 3 different chromosomes. We also tested the enzymatic activity of recombinant VPEs expressed in the Pichia Pastoris expression system and found that the VvVPEs exhibit cysteine peptidase activity. Tissue-specific expression analysis showed that VvδVPE is only expressed in flowers, buds and ovules, that VvγVPE is expressed in various tissues, and that VvβVPE was expressed in roots, flowers, buds and ovules. The results of quantitative real-time PCR (qRT-PCR) suggested that VvβVPE in seeded grapes increased significantly at 30 days after full-bloom (DAF), close to the timing of endosperm abortion at 32 DAF. These results suggested that VvβVPE is related to ovule abortion in seedless grapes. Our experiments provide a new perspective for understanding the mechanism of stenospermocarpic seedlessness and represent a useful reference for the further study of VPEs.

  10. Two azole fungicides (carcinogenic triadimefon and non-carcinogenic myclobutanil) exhibit different hepatic cytochrome P450 activities in medaka fish

    Energy Technology Data Exchange (ETDEWEB)

    Lin, Chun-Hung [Department of Agricultural Chemistry, National Taiwan University, Taipei, Taiwan (China); Chou, Pei-Hsin [Department of Environmental Engineering, National Cheng-Kung University, Tainan, Taiwan (China); Chen, Pei-Jen, E-mail: chenpj@ntu.edu.tw [Department of Agricultural Chemistry, National Taiwan University, Taipei, Taiwan (China)

    2014-07-30

    Highlights: • We assess ecotoxicological impact of azole fungicides in the aquatic environment. • Carcinogenic and non-carcinogenic azoles show different CYP activities in medaka. • We compare azole-induced CYP expression and carcinogenesis between fish and rodents. • Liver CYP-enzyme induction is a key event in conazole-induced tumorigenesis. • We suggest toxicity evaluation methods for azole fungicides using medaka fish. - Abstract: Conazoles are a class of imidazole- or triazole-containing drugs commonly used as fungicides in agriculture and medicine. The broad application of azole drugs has led to the contamination of surface aquifers receiving the effluent of municipal or hospital wastewater or agricultural runoff. Several triazoles are rodent carcinogens; azole pollution is a concern to environmental safety and human health. However, the carcinogenic mechanisms associated with cytochrome P450 enzymes (CYPs) of conazoles remain unclear. We exposed adult medaka fish (Oryzias latipes) to continuous aqueous solutions of carcinogenic triadimefon and non-carcinogenic myclobutanil for 7 to 20 days at sub-lethal or environmentally relevant concentrations and assessed hepatic CYP activity and gene expression associated with CYP-mediated toxicity. Both triadimefon and myclobutanil induced hepatic CYP3A activity, but only triadimefon enhanced CYP1A activity. The gene expression of cyp3a38, cyp3a40, pregnane x receptor (pxr), cyp26b, retinoid acid receptor γ1 (rarγ1) and p53 was higher with triadimefon than myclobutanil. As well, yeast-based reporter gene assay revealed that 4 tested conazoles were weak agonists of aryl hydrocarbon receptor (AhR). We reveal differential CYP gene expression with carcinogenic and non-carcinogenic conazoles in a lower vertebrate, medaka fish. Liver CYP-enzyme induction may be a key event in conazole-induced tumorigenesis. This information is essential to evaluate the potential threat of conazoles to human health and fish

  11. A Review on the Effects of Supercritical Carbon Dioxide on Enzyme Activity

    Directory of Open Access Journals (Sweden)

    Marie Zarevúcka

    2010-01-01

    Full Text Available Different types of enzymes such as lipases, several phosphatases, dehydrogenases, oxidases, amylases and others are well suited for the reactions in SC-CO2. The stability and the activity of enzymes exposed to carbon dioxide under high pressure depend on enzyme species, water content in the solution and on the pressure and temperature of the reaction system. The three-dimensional structure of enzymes may be significantly altered under extreme conditions, causing their denaturation and consequent loss of activity. If the conditions are less adverse, the protein structure may be largely retained. Minor structural changes may induce an alternative active protein state with altered enzyme activity, specificity and stability.

  12. Chronic active hepatitis in transgenic mice expressing interferon-gamma in the liver.

    OpenAIRE

    1994-01-01

    Interferon-gamma may play an important role in the immune response and in inflammatory diseases, including chronic active hepatitis. To understand the role of interferon-gamma in the regulation of inflammation and to establish a mouse model of chronic active hepatitis, we produced transgenic mice in which the mouse interferon-gamma gene was regulated by a liver-specific promoter, the serum amyloid P component gene promoter. Four transgenic mouse lines were generated, and two of these lines ex...

  13. Fisetin improves glucose homeostasis through the inhibition of gluconeogenic enzymes in hepatic tissues of streptozotocin induced diabetic rats.

    Science.gov (United States)

    Prasath, Gopalan Sriram; Pillai, Subramanian Iyyam; Subramanian, Sorimuthu Pillai

    2014-10-05

    Liver plays a vital role in blood glucose homeostasis. Recent studies have provided considerable evidence that hepatic glucose production (HGP) plays an important role in the development of fasting hyperglycemia in diabetes. From this perspective, diminution of HGP has certainly been considered for the treatment of diabetes. In the present study, we have analyzed the modulatory effects of fisetin, a flavonoid of strawberries, on the expression of key enzymes of carbohydrate metabolism in STZ induced experimental diabetic rats. The physiological criterions such as food and fluid intake were regularly monitored. The levels of blood glucose, plasma insulin, hemoglobin and glycosylated hemoglobin were analyzed. The mRNA and protein expression levels of gluconeogenic genes such as phosphoenolpyruvate carboxykinase (PEPCK) and glucose-6-phosphatase (G6Pase) were determined by immunoblot as well as PCR analysis. Diabetic group of rats showed significant increase in food and water intake when compared with control group of rats. Upon oral administration of fisetin as well as gliclazide to diabetic group of rats, the levels were found to be decreased. Oral administration of fisetin (10 mg/kg body weight) to diabetic rats for 30 days established a significant decline in blood glucose and glycosylated hemoglobin levels and a significant increase in plasma insulin level. The mRNA and protein expression levels of gluconeogenic genes, such as phosphoenolpyruvate carboxykinase (PEPCK) and glucose-6-phosphatase (G6Pase), were decreased in liver tissues upon treatment with fisetin. The results of the present study suggest that fisetin improves glucose homeostasis by direct inhibition of gluconeogenesis in liver. Copyright © 2014 Elsevier B.V. All rights reserved.

  14. Diurnal variations of mouse plasma and hepatic bile acid concentrations as well as expression of biosynthetic enzymes and transporters.

    Directory of Open Access Journals (Sweden)

    Yu-Kun Jennifer Zhang

    Full Text Available BACKGROUND: Diurnal fluctuation of bile acid (BA concentrations in the enterohepatic system of mammals has been known for a long time. Recently, BAs have been recognized as signaling molecules beyond their well-established roles in dietary lipid absorption and cholesterol homeostasis. METHODS AND RESULTS: The current study depicted diurnal variations of individual BAs detected by ultra-performance liquid chromatography/mass spectrometry (UPLC/MS in serum and livers collected from C57BL/6 mice fed a regular chow or a chow containing cholestyramine (resin. Circadian rhythms of mRNA of vital BA-related nuclear receptors, enzymes, and transporters in livers and ilea were determined in control- and resin-fed mice, as well as in farnesoid X receptor (FXR null mice. The circadian profiles of BAs showed enhanced bacterial dehydroxylation during the fasting phase and efficient hepatic reconjugation of BAs in the fed phase. The resin removed more than 90% of BAs with β-hydroxy groups, such as muricholic acids and ursodeoxycholic acid, from serum and livers, but did not exert as significant influence on CA and CDCA in both compartments. Both resin-fed and FXR-null mouse models indicate that BAs regulate their own biosynthesis through the FXR-regulated ileal fibroblast growth factor 15. BA flux also influences the daily mRNA levels of multiple BA transporters. CONCLUSION: BA concentration and composition exhibit circadian variations in mouse liver and serum, which influences the circadian rhythms of BA metabolizing genes in liver and ileum. The diurnal variations of BAs appear to serve as a signal that coordinates daily nutrient metabolism in mammals.

  15. Effect of age and diet composition on activity of pancreatic enzymes in birds.

    Science.gov (United States)

    Brzęk, Paweł; Ciminari, M Eugenia; Kohl, Kevin D; Lessner, Krista; Karasov, William H; Caviedes-Vidal, Enrique

    2013-07-01

    Digestive enzymes produced by the pancreas and intestinal epithelium cooperate closely during food hydrolysis. Therefore, activities of pancreatic and intestinal enzymes processing the same substrate can be hypothesized to change together in unison, as well as to be adjusted to the concentration of their substrate in the diet. However, our knowledge of ontogenetic and diet-related changes in the digestive enzymes of birds is limited mainly to intestinal enzymes; it is largely unknown whether they are accompanied by changes in activities of enzymes produced by the pancreas. Here, we analyzed age- and diet-related changes in activities of pancreatic enzymes in five passerine and galloanserine species, and compared them with simultaneous changes in activities of intestinal enzymes. Mass-specific activity of pancreatic amylase increased with age in young house sparrows but not in zebra finches, in agreement with changes in typical dietary starch content and activity of intestinal maltase. However, we found little evidence for the presence of adaptive, diet-related modulation of pancreatic enzymes in both passerine and galloanserine species, even though in several cases the same birds adaptively modulated activities of their intestinal enzymes. In general, diet-related changes in mass-specific activities of pancreatic and intestinal enzymes were not correlated. We conclude that activity of pancreatic enzymes in birds is under strong genetic control, which enables evolutionary adjustment to typical diet composition but is less adept for short term, diet-related flexibility.

  16. Hydrolytic enzymes conjugated to quantum dots mostly retain whole catalytic activity.

    Science.gov (United States)

    Iyer, Aditya; Chandra, Anil; Swaminathan, Rajaram

    2014-09-01

    Tagging a luminescent quantum dot (QD) with a biological like enzyme (Enz) creates value-added entities like quantum dot-enzyme bioconjugates (QDEnzBio) that find utility as sensors to detect glucose or beacons to track enzymes in vivo. For such applications, it is imperative that the enzyme remains catalytically active while the quantum dot is luminescent in the bioconjugate. A critical feature that dictates this is the quantum dot-enzyme linkage chemistry. Previously such linkages have put constraints on polypeptide chain dynamics or hindered substrate diffusion to active site, seriously undermining enzyme catalytic activity. In this work we address this issue using avidin-biotin linkage chemistry together with a flexible spacer to conjugate enzyme to quantum dot. The catalytic activity of three biotinylated hydrolytic enzymes, namely, hen egg white lysozyme (HEWL), alkaline phosphatase (ALP) and acetylcholinesterase (AChE) was investigated post-conjugation to streptavidin linked quantum dot for multiple substrate concentrations and varying degrees of biotinylation. We demonstrate that all enzymes retain full catalytic activity in the quantum dot-enzyme bioconjugates in comparison to biotinylated enzyme alone. However, unlike alkaline phosphatase and acetylcholinesterase, the catalytic activity of hen egg white lysozyme was observed to be increasingly susceptible to ionic strength of medium with rising level of biotinylation. This susceptibility was attributed to arise from depletion of positive charge from lysine amino groups after biotinylation. We reasoned that avidin-biotin linkage in the presence of a flexible seven atom spacer between biotin and enzyme poses no constraints to enzyme structure/dynamics enabling retention of full enzyme activity. Overall our results demonstrate for the first time that streptavidin-biotin chemistry can yield quantum dot enzyme bioconjugates that retain full catalytic activity as native enzyme. Copyright © 2014 Elsevier B

  17. Hepatic ethoxy-, methoxy- and pentoxyresorufin O-dealkylase activities in Landrace and Duroc pigs stimulated with HCG.

    Science.gov (United States)

    Zamaratskaia, G; Zlabek, V; Ropstad, E; Tajet, H; Andresen, Ø

    2010-12-01

    The effect of human chorionic gonadotropin (hCG) stimulation on the activities of ethoxyresorufin O-deethylase (EROD), methoxyresorufin O-demethylase (MROD) and pentoxyresorufin O-depentylase (PROD) was studied in intact male pigs of purebred Landrace and Duroc breeds. Pigs were divided into four groups: two control groups of each breed, without hCG stimulation (n = 20 for each breed), and two experimental groups (n = 18 for each breed), with hCG stimulation (Pregnyl(®); N.V. Organon, Oss, The Netherlands, 30 IU/kg live weight). Pigs were slaughtered 3 days after hCG stimulation and enzyme activities were measured in hepatic microsomes using two approaches. First, only one substrate concentration was used for the analysis of each enzyme activity. We found that EROD activity was suppressed by hCG-stimulation in Landrace (p = 0.004), but not Duroc pigs (p > 0.05). Generally, EROD activity was higher in Duroc pigs compared with Landrace (p = 0.017). Methoxyresorufin O-demethylase and PROD activities did not differ between groups (p > 0.05). To further characterize EROD, MROD and PROD, enzyme kinetic studies were performed. V(max) values for EROD and MROD in both breeds were lower after hCG stimulation (p Landrace and p Landrace and Duroc pigs being higher in Duroc pigs (p < 0.05). We concluded that both hCG stimulation and breed differences may be important in the regulation of EROD and MROD activities. This study provides the first data on the effect of hCG stimulation and thus high testicular steroids, on EROD, MROD and PROD activities. Further studies are needed to investigate individual CYP450 enzymes and their regulation in porcine tissues.

  18. The hepatoselective glucokinase activator PF-04991532 ameliorates hyperglycemia without causing hepatic steatosis in diabetic rats.

    Directory of Open Access Journals (Sweden)

    Derek M Erion

    Full Text Available Hyperglycemia resulting from type 2 diabetes mellitus (T2DM is the main cause of diabetic complications such as retinopathy and neuropathy. A reduction in hyperglycemia has been shown to prevent these associated complications supporting the importance of glucose control. Glucokinase converts glucose to glucose-6-phosphate and determines glucose flux into the β-cells and hepatocytes. Since activation of glucokinase in β-cells is associated with increased risk of hypoglycemia, we hypothesized that selectively activating hepatic glucokinase would reduce fasting and postprandial glucose with minimal risk of hypoglycemia. Previous studies have shown that hepatic glucokinase overexpression is able to restore glucose homeostasis in diabetic models; however, these overexpression experiments have also revealed that excessive increases in hepatic glucokinase activity may also cause hepatosteatosis. Herein we sought to evaluate whether liver specific pharmacological activation of hepatic glucokinase is an effective strategy to reduce hyperglycemia without causing adverse hepatic lipids changes. To test this hypothesis, we evaluated a hepatoselective glucokinase activator, PF-04991532, in Goto-Kakizaki rats. In these studies, PF-04991532 reduced plasma glucose concentrations independent of changes in insulin concentrations in a dose-dependent manner both acutely and after 28 days of sub-chronic treatment. During a hyperglycemic clamp in Goto-Kakizaki rats, the glucose infusion rate was increased approximately 5-fold with PF-04991532. This increase in glucose infusion can be partially attributed to the 60% reduction in endogenous glucose production. While PF-04991532 induced dose-dependent increases in plasma triglyceride concentrations it had no effect on hepatic triglyceride concentrations in Goto-Kakizaki rats. Interestingly, PF-04991532 decreased intracellular AMP concentrations and increased hepatic futile cycling. These data suggest that

  19. The hepatoselective glucokinase activator PF-04991532 ameliorates hyperglycemia without causing hepatic steatosis in diabetic rats.

    Science.gov (United States)

    Erion, Derek M; Lapworth, Amanda; Amor, Paul A; Bai, Guoyun; Vera, Nicholas B; Clark, Ronald W; Yan, Qingyun; Zhu, Yimin; Ross, Trenton T; Purkal, Julie; Gorgoglione, Matthew; Zhang, Guodong; Bonato, Vinicius; Baker, Levenia; Barucci, Nicole; D'Aquila, Theresa; Robertson, Alan; Aiello, Robert J; Yan, Jiangli; Trimmer, Jeff; Rolph, Timothy P; Pfefferkorn, Jeffrey A

    2014-01-01

    Hyperglycemia resulting from type 2 diabetes mellitus (T2DM) is the main cause of diabetic complications such as retinopathy and neuropathy. A reduction in hyperglycemia has been shown to prevent these associated complications supporting the importance of glucose control. Glucokinase converts glucose to glucose-6-phosphate and determines glucose flux into the β-cells and hepatocytes. Since activation of glucokinase in β-cells is associated with increased risk of hypoglycemia, we hypothesized that selectively activating hepatic glucokinase would reduce fasting and postprandial glucose with minimal risk of hypoglycemia. Previous studies have shown that hepatic glucokinase overexpression is able to restore glucose homeostasis in diabetic models; however, these overexpression experiments have also revealed that excessive increases in hepatic glucokinase activity may also cause hepatosteatosis. Herein we sought to evaluate whether liver specific pharmacological activation of hepatic glucokinase is an effective strategy to reduce hyperglycemia without causing adverse hepatic lipids changes. To test this hypothesis, we evaluated a hepatoselective glucokinase activator, PF-04991532, in Goto-Kakizaki rats. In these studies, PF-04991532 reduced plasma glucose concentrations independent of changes in insulin concentrations in a dose-dependent manner both acutely and after 28 days of sub-chronic treatment. During a hyperglycemic clamp in Goto-Kakizaki rats, the glucose infusion rate was increased approximately 5-fold with PF-04991532. This increase in glucose infusion can be partially attributed to the 60% reduction in endogenous glucose production. While PF-04991532 induced dose-dependent increases in plasma triglyceride concentrations it had no effect on hepatic triglyceride concentrations in Goto-Kakizaki rats. Interestingly, PF-04991532 decreased intracellular AMP concentrations and increased hepatic futile cycling. These data suggest that hepatoselective glucokinase

  20. IL-25 or IL-17E Protects against High-Fat Diet-Induced Hepatic Steatosis in Mice Dependent upon IL-13 Activation of STAT6.

    Science.gov (United States)

    Wang, An-Jiang; Yang, Zhonghan; Grinchuk, Viktoriya; Smith, Allen; Qin, Bolin; Lu, Nonghua; Wang, Duan; Wang, Hongbing; Ramalingam, Thirumalai R; Wynn, Thomas A; Urban, Joseph F; Shea-Donohue, Terez; Zhao, Aiping

    2015-11-15

    IL-25 or IL-17E is a member of IL-17 cytokine family and has immune-modulating activities. The role of IL-25 in maintaining lipid metabolic homeostasis remains unknown. We investigated the effects of exogenous IL-25 or deficiency of IL-25 on hepatic lipid accumulation. IL-25 expression was examined in paraffin-embedded tissue sections of liver from patients or in the livers from mice. Mouse model of steatosis was induced by feeding a high-fat diet (HFD). Extent of steatosis as well as expression of cytokines, key enzymes for lipid metabolic pathways, markers for Kupffer cells/macrophages, and lipid droplet (LD) proteins, were analyzed. Our results show that hepatic steatosis in mice was accompanied by increased LD proteins, but decreased IL-25 in the liver. Decreased hepatic IL-25 was also observed in patients with fatty liver. Administration of IL-25 to HFD-fed wild-type mice led to a significant improvement in hepatic steatosis. This effect was associated with increased expression of IL-13, development of alternatively activated Kupffer cells/macrophages, and decreased expression of LD proteins in the liver. In contrast, administration of IL-25 to HFD-fed mice deficient in STAT6 or IL-13 had no effects. In addition, stimulation of primary hepatocytes with IL-13, but not IL-25, resulted in downregulation of LD proteins. Finally, mice deficient in IL-25 had exacerbated hepatic lipid accumulation when fed the HFD. These data demonstrate that dysregulated IL-25 expression contributes to lipid accumulation, whereas exogenous IL-25 protects against hepatic steatosis through IL-13 activation of STAT6. IL-25 and IL-13 are potential therapeutic agents for hepatic steatosis and associated pathologies.

  1. Amphiregulin activates human hepatic stellate cells and is upregulated in non alcoholic steatohepatitis

    Science.gov (United States)

    McKee, Chad; Sigala, Barbara; Soeda, Junpei; Mouralidarane, Angelina; Morgan, Maelle; Mazzoccoli, Gianluigi; Rappa, Francesca; Cappello, Francesco; Cabibi, Daniela; Pazienza, Valerio; Selden, Claire; Roskams, Tania; Vinciguerra, Manlio; Oben, Jude A.

    2015-01-01

    Amphiregulin (AR) involvement in liver fibrogenesis and hepatic stellate cells (HSC) regulation is under study. Non-alcoholic fatty liver disease (NAFLD) and its more severe form non-alcoholic steatohepatitis (NASH) may progress to cirrhosis and hepatocellular cancer (HCC). Our aim was to investigate ex vivo the effect of AR on human primary HSC (hHSC) and verify in vivo the relevance of AR in NAFLD fibrogenesis. hHSC isolated from healthy liver segments were analyzed for expression of AR and its activator, TNF-α converting enzyme (TACE). AR induction of hHSC proliferation and matrix production was estimated in the presence of antagonists. AR involvement in fibrogenesis was also assessed in a mouse model of NASH and in humans with NASH. hHSC time dependently expressed AR and TACE. AR increased hHSC proliferation through several mitogenic signaling pathways such as EGFR, PI3K and p38. AR also induced marked upregulation of hHSC fibrogenic markers and reduced hHSC death. AR expression was enhanced in the HSC of a murine model of NASH and of severe human NASH. In conclusion, AR induces hHSC fibrogenic activity via multiple mitogenic signaling pathways, and is upregulated in murine and human NASH, suggesting that AR antagonists may be clinically useful anti-fibrotics in NAFLD. PMID:25744849

  2. Inhibition of cytochrome P450 enzymes by saturated and unsaturated fatty acids in human liver microsomes, characterization of enzyme kinetics in the presence of bovine serum albumin (0.1 and 1.0% w/v) and in vitro - in vivo extrapolation of hepatic clearance.

    Science.gov (United States)

    Palacharla, Raghava Choudary; Uthukam, Venkatesham; Manoharan, Arunkumar; Ponnamaneni, Ranjith Kumar; Padala, Nagasurya Prakash; Boggavarapu, Rajesh Kumar; Bhyrapuneni, Gopinadh; Ajjala, Devender Reddy; Nirogi, Ramakrishna

    2017-04-01

    The objective of the study was to determine the effect of fatty acids on CYP enzymes and the effect of BSA on intrinsic clearance of probe substrates. The inhibitory effect of thirteen fatty acids including saturated, mono-unsaturated and polyunsaturated fatty acids on CYP enzymes, kinetic parameters and intrinsic clearance values of nine CYP marker probe substrate reactions in the absence and presence of BSA (0.1 and 1.0% w/v) were characterized in human liver microsomes. The results demonstrate that most of the unsaturated fatty acids showed marked inhibition towards CYP2C8 mediated amodiaquine N-deethylation followed by inhibition of CYP2C9 and CYP2B6 mediated activities. The addition of 0.1% BSA in the incubation markedly improved the unbound intrinsic clearance values of probe substrates by reducing the Km values with little or no effect on maximal velocity. The addition of BSA (0.1 and 1.0% w/v) did not influence the unbound intrinsic clearance of marker reactions for CYP2A6, and CYP3A4 enzymes. The addition of 0.1% w/v BSA is sufficient to determine the intrinsic clearance of marker probe reactions by metabolite formation approach. The predicted hepatic clearance values for the substrates using the well-stirred model, in the presence of BSA (0.1% BSA), are comparable to the in vivo hepatic clearance values.

  3. Quantification of viral DNA and liver enzymes in plasma improves early diagnosis and management of herpes simplex virus hepatitis

    NARCIS (Netherlands)

    M.F.C. Beersma (Thijs); G.M.G.M. Verjans (George); H.J. Metselaar (Herold); A.D.M.E. Osterhaus (Albert); W.R. Berrington; G.J.J. van Doornum (Gerard)

    2011-01-01

    textabstractHerpes simplex virus (HSV) hepatitis is a rare and potential life-threatening disease. The diagnosis of HSV hepatitis is hampered by its indifferent clinical presentation, which necessitates confirmatory laboratory data to identify HSV in the affected liver. However, liver biopsies are o

  4. Quantification of viral DNA and liver enzymes in plasma improves early diagnosis and management of herpes simplex virus hepatitis

    NARCIS (Netherlands)

    M.F.C. Beersma (Thijs); G.M.G.M. Verjans (George); H.J. Metselaar (Herold); A.D.M.E. Osterhaus (Albert); W.R. Berrington; G.J.J. van Doornum (Gerard)

    2011-01-01

    textabstractHerpes simplex virus (HSV) hepatitis is a rare and potential life-threatening disease. The diagnosis of HSV hepatitis is hampered by its indifferent clinical presentation, which necessitates confirmatory laboratory data to identify HSV in the affected liver. However, liver biopsies are

  5. Phosphate-Modified Nucleotides for Monitoring Enzyme Activity.

    Science.gov (United States)

    Ermert, Susanne; Marx, Andreas; Hacker, Stephan M

    2017-04-01

    Nucleotides modified at the terminal phosphate position have been proven to be interesting entities to study the activity of a variety of different protein classes. In this chapter, we present various types of modifications that were attached as reporter molecules to the phosphate chain of nucleotides and briefly describe the chemical reactions that are frequently used to synthesize them. Furthermore, we discuss a variety of applications of these molecules. Kinase activity, for instance, was studied by transfer of a phosphate modified with a reporter group to the target proteins. This allows not only studying the activity of kinases, but also identifying their target proteins. Moreover, kinases can also be directly labeled with a reporter at a conserved lysine using acyl-phosphate probes. Another important application for phosphate-modified nucleotides is the study of RNA and DNA polymerases. In this context, single-molecule sequencing is made possible using detection in zero-mode waveguides, nanopores or by a Förster resonance energy transfer (FRET)-based mechanism between the polymerase and a fluorophore-labeled nucleotide. Additionally, fluorogenic nucleotides that utilize an intramolecular interaction between a fluorophore and the nucleobase or an intramolecular FRET effect have been successfully developed to study a variety of different enzymes. Finally, also some novel techniques applying electron paramagnetic resonance (EPR)-based detection of nucleotide cleavage or the detection of the cleavage of fluorophosphates are discussed. Taken together, nucleotides modified at the terminal phosphate position have been applied to study the activity of a large diversity of proteins and are valuable tools to enhance the knowledge of biological systems.

  6. Anti-hepatitis B virus activities of cinobufacini and its active components bufalin and cinobufagin in HepG2.2.15 cells.

    Science.gov (United States)

    Cui, Xiaoyan; Inagaki, Yoshinori; Xu, Huanli; Wang, Dongliang; Qi, Fanghua; Kokudo, Norihiro; Fang, Dingzhi; Tang, Wei

    2010-01-01

    Cinobufacini (Huachansu) is a Chinese medicine prepared from the skin of Bufo bufo gargarizans Cantor (Bufonidae), which has long been used in traditional Chinese medicine (TCM). The aim of present study was to examine the anti-hepatitis B virus (HBV) activities of cinobufacini and its active components bufalin and cinobufagin in the human HBV-transfected cell line HepG2.2.15. The hepatitis B surface antigen (HBsAg), hepatitis B e antigen (HBeAg), and hepatitis B core-related antigen (HBcrAg) concentrations in cell culture medium were determined by chemiluminescent enzyme immunoassay after HepG2.2.15 cells were respectively treated with different concentrations of cinobufacini, bufalin, and cinobufagin for 3 or 6 d. HBV DNA and mRNA were determined using transcription-mediated amplification and real-time polymerase chain reaction (PCR), respectively. On d 3, cinobufacini at a concentration of 1 µg/ml had no activity against HBV virological markers. However, on d 6, cinobufacini at 1 µg/ml effectively inhibited the secretion of HBsAg, HBeAg, and HBcrAg by 29.58, 32.87, and 42.52%. It was more potent than the positive control lamivudine (100 µg/ml). Bufalin and cinobufagin slightly inhibited HBV antigen secretion. Treatment with cinobufacini, bufalin, or cinobufagin had no anti-HBV effect on DNA in cell culture medium. Consistent with the HBV antigen reduction, HBV mRNA expression was markedly inhibited in comparison to the control when HepG2.2.15 cells were treated with cinobufacini, bufalin, or cinobufagin. Results suggested that cinobufacini had more potent activity against HBV antigen secretion than its components bufalin and cinobufagin and this inhibitory role was attributed to the specific inhibition of HBV mRNA expression.

  7. Ultrasonic Monitoring of Enzyme Catalysis; Enzyme Activity in Formulations for Lactose-Intolerant Infants.

    Science.gov (United States)

    Altas, Margarida C; Kudryashov, Evgeny; Buckin, Vitaly

    2016-05-03

    The paper introduces ultrasonic technology for real-time, nondestructive, precision monitoring of enzyme-catalyzed reactions in solutions and in complex opaque media. The capabilities of the technology are examined in a comprehensive analysis of the effects of a variety of diverse factors on the performance of enzyme β-galactosidase in formulations for reduction of levels of lactose in infant milks. These formulations are added to infant's milk bottles prior to feeding to overcome the frequently observed intolerance to lactose (a milk sugar), a serious issue in healthy development of infants. The results highlight important impediments in the development of these formulations and also illustrate the capability of the described ultrasonic tools in the assessment of the performance of enzymes in complex reaction media and in various environmental conditions.

  8. Hepatic Insulin Resistance Following Chronic Activation of the CREB Coactivator CRTC2*

    Science.gov (United States)

    Hogan, Meghan F.; Ravnskjaer, Kim; Matsumura, Shigenobu; Huising, Mark O.; Hull, Rebecca L.; Kahn, Steven E.; Montminy, Marc

    2015-01-01

    Under fasting conditions, increases in circulating concentrations of glucagon maintain glucose homeostasis via the induction of hepatic gluconeogenesis. Triggering of the cAMP pathway in hepatocytes stimulates the gluconeogenic program via the PKA-mediated phosphorylation of CREB and dephosphorylation of the cAMP-regulated CREB coactivators CRTC2 and CRTC3. In parallel, decreases in circulating insulin also increase gluconeogenic gene expression via the de-phosphorylation and activation of the forkhead transcription factor FOXO1. Hepatic gluconeogenesis is increased in insulin resistance where it contributes to the attendant hyperglycemia. Whether selective activation of the hepatic CREB/CRTC pathway is sufficient to trigger metabolic changes in other tissues is unclear, however. Modest hepatic expression of a phosphorylation-defective and therefore constitutively active CRTC2S171,275A protein increased gluconeogenic gene expression under fasting as well as feeding conditions. Circulating glucose concentrations were constitutively elevated in CRTC2S171,275A-expressing mice, leading to compensatory increases in circulating insulin concentrations that enhance FOXO1 phosphorylation. Despite accompanying decreases in FOXO1 activity, hepatic gluconeogenic gene expression remained elevated in CRTC2S171,275A mice, demonstrating that chronic increases in CRTC2 activity in the liver are indeed sufficient to promote hepatic insulin resistance and to disrupt glucose homeostasis. PMID:26342077

  9. Hepatic Insulin Resistance Following Chronic Activation of the CREB Coactivator CRTC2.

    Science.gov (United States)

    Hogan, Meghan F; Ravnskjaer, Kim; Matsumura, Shigenobu; Huising, Mark O; Hull, Rebecca L; Kahn, Steven E; Montminy, Marc

    2015-10-23

    Under fasting conditions, increases in circulating concentrations of glucagon maintain glucose homeostasis via the induction of hepatic gluconeogenesis. Triggering of the cAMP pathway in hepatocytes stimulates the gluconeogenic program via the PKA-mediated phosphorylation of CREB and dephosphorylation of the cAMP-regulated CREB coactivators CRTC2 and CRTC3. In parallel, decreases in circulating insulin also increase gluconeogenic gene expression via the de-phosphorylation and activation of the forkhead transcription factor FOXO1. Hepatic gluconeogenesis is increased in insulin resistance where it contributes to the attendant hyperglycemia. Whether selective activation of the hepatic CREB/CRTC pathway is sufficient to trigger metabolic changes in other tissues is unclear, however. Modest hepatic expression of a phosphorylation-defective and therefore constitutively active CRTC2S171,275A protein increased gluconeogenic gene expression under fasting as well as feeding conditions. Circulating glucose concentrations were constitutively elevated in CRTC2S171,275A-expressing mice, leading to compensatory increases in circulating insulin concentrations that enhance FOXO1 phosphorylation. Despite accompanying decreases in FOXO1 activity, hepatic gluconeogenic gene expression remained elevated in CRTC2S171,275A mice, demonstrating that chronic increases in CRTC2 activity in the liver are indeed sufficient to promote hepatic insulin resistance and to disrupt glucose homeostasis.

  10. Correlation Among Soil Enzyme Activities, Root Enzyme Activities, and Contaminant Removal in Two-Stage In Situ Constructed Wetlands Purifying Domestic Wastewater.

    Science.gov (United States)

    Ni, Lixiao; Xu, Jiajun; Chu, Xianglin; Li, Shiyin; Wang, Peifang; Li, Yiping; Li, Yong; Zhu, Liang; Wang, Chao

    2016-07-01

    Two-stage in situ wetlands (two vertical flow constructed wetlands in parallel and a horizontal flow constructed wetland) were constructed for studying domestic wastewater purification and the correlations between contaminant removal and plant and soil enzyme activities. Results indicated the removal efficiency of NH4 (+) and NO3 (-) were significantly correlated with both urease and protease activity, and the removal of total phosphorus was significantly correlated with phosphatase activity. Chemical oxygen demand removal was not correlated with enzyme activity in constructed wetlands. Plant root enzyme (urease, phosphatase, protease and cellulose) activity correlation was apparent with all contaminant removal in the two vertical flow constructed wetlands. However, the correlation between the plant root enzyme activity and contaminant removal was poor in horizontal flow constructed wetlands. Results indicated that plant roots clearly played a role in the removal of contaminants.

  11. Characterization and profiling of hepatic cytochromes P450 and phase II xenobiotic-metabolizing enzymes in beluga whales (Delphinapterus leucas) from the St. Lawrence River Estuary and the Canadian Arctic.

    Science.gov (United States)

    McKinney, Melissa A; Arukwe, Augustine; De Guise, Sylvain; Martineau, Daniel; Béland, Pierre; Dallaire, André; Lair, Stéphane; Lebeuf, Michel; Letcher, Robert J

    2004-07-30

    Cytochromes P450 (CYP, phase I) and conjugating (phase II) enzymes can be induced by and influence the toxicokinetics (metabolism) and toxicity of xenobiotic contaminants in exposed organisms. Beluga whale (Delphinapterus leucas) from the endangered St. Lawrence (SL) River Estuary population exhibit deleterious health effects and various severe pathologies that have been associated with contaminant exposure. In contrast, such effects (e.g. reproductive and immunological impairment) are generally less frequent in less exposed populations in the Canadian Arctic (CA). In the present study, opportunistic sampling resulted in the collection immediately after death of liver tissue from a single female neonate SL beluga (SL6) and male and female CA beluga (n=10) from the Arviat region of western Hudson Bay, in addition to sampling of stranded carcasses of male and female SL beluga (n=5) at least 12 h postmortem. We immunologically characterized cross-reactive proteins of hepatic microsomal CYP1A, CYP2B, CYP3A, CYP2E, epoxide hydrolase (EH) and uridine diphosphoglucuronosyl transferase (UDPGT) isozymes. Cross-reactive proteins were found in all SL and CA beluga using anti-rat CYP1A1, anti-rainbow trout CYP3A, anti-human CYP2E1, anti-rabbit EH and anti-human UDPGT1A1 polyclonal antibodies (Abs), whereas faintly cross-reactive CYP2B proteins were only found in SL6 and the CA samples using an anti-rabbit CYP2B1 Ab. In corresponding catalytic activity assessments, only SL6 and all CA beluga microsomal samples exhibited CYP1A-mediated 7-ethoxyresorufin O-deethylase (EROD) activity (51-260 pmol/mg/min), CYP3A-mediated activity (113-899 pmol/mg/min) based on the formation of 6beta-hydroxytestosterone using a testosterone hydroxylase assay, and UDPGT activity (830-4956 pmol/mg/min) based on 1-naphthylglucuronide formation. The marginal cross-reactivity with the anti-CYP2B1 Ab and lack of catalytically measurable hydroxytestosterone isomers associated with CYP2B-type activity in

  12. Silica Exposure and Serum Angiotensin Converting Enzyme Activity

    Directory of Open Access Journals (Sweden)

    YK Sharma

    2010-01-01

    Full Text Available Background: Silicosis is known in industrial workers for centuries. Till recently, the mainstay of its diagnosis and progress was clinical examination of the respiratory system, pulmonary function test and chest radiography. Several biomarkers such as serum angiotensin converting enzyme (ACE activity have been examined to determine the extent of silicosis. Objective: To elucidate the effect of age, gender, duration of exposure to silica dust, smoking habit, and pulmonary function status on the serum ACE activity among quartz stone workers without disease.Methods: A cross-sectional study was carried out on 134 (111 men and 14 women workers of quartz stone crushing units were studied. Standard diagnostic criteria were used for diagnosing silicosis and tuberculosis. Pulmonary functions of the participants were also assessed.Results: The mean±SD age for participants was 26.1±6.3 years (26.6±6.3 for men and 21.9±4.3 for women. The mean±SD duration of exposure was 1.1±1.9 years. In the present study, only one case of silicosis and eight cases of tuberculosis were found. The mean±SD serum ACE levels for those with and without respiratory disease were 68.44±11.61, and 66.9±14.4 IU/L, respectively (p>0.05.Conclusion: We could not observe any association between serum ACE activity and age, gender, duration of exposure, smoking habits and pulmonary function status. However, elevated levels of serum ACE was found in a solitary case of silicosis.

  13. Angiotensin-Converting Enzyme Inhibitors and Active Tuberculosis

    Science.gov (United States)

    Wu, Jiunn-Yih; Lee, Meng-Tse Gabriel; Lee, Si-Huei; Lee, Shih-Hao; Tsai, Yi-Wen; Hsu, Shou-Chien; Chang, Shy-Shin; Lee, Chien-Chang

    2016-01-01

    Abstract Numerous epidemiological data suggest that the use of angiotensin-converting enzyme inhibitors (ACEis) can improve the clinical outcomes of pneumonia. Tuberculosis (TB) is an airborne bacteria like pneumonia, and we aimed to find out whether the use of ACEis can decrease the risk of active TB. We conducted a nested case–control analysis by using a 1 million longitudinally followed cohort, from Taiwan national health insurance research database. The rate ratios (RRs) for TB were estimated by conditional logistic regression, and adjusted using a TB-specific disease risk score (DRS) with 71 TB-related covariates. From January, 1997 to December, 2011, a total of 75,536 users of ACEis, and 7720 cases of new active TB were identified. Current use (DRS adjusted RR, 0.87 [95% CI, 0.78–0.97]), but not recent and past use of ACEis, was associated with a decrease in risk of active TB. Interestingly, it was found that chronic use (>90 days) of ACEis was associated with a further decrease in the risk of TB (aRR, 0.74, [95% CI, 0.66–0.83]). There was also a duration response effect, correlating decrease in TB risk with longer duration of ACEis use. The decrease in TB risk was also consistent across all patient subgroups (age, sex, heart failure, cerebrovascular diseases, myocardial infraction, renal diseases, and diabetes) and patients receiving other cardiovascular medicine. In this large population-based study, we found that subjects with recent and chronic use of ACEis were associated with decrease in TB risk. PMID:27175655

  14. Controlled exogenous enzyme imbibition and activation in whole chickpea seed enzyme reactor (SER).

    Science.gov (United States)

    Kliger, Eynav; Fischer, Lutz; Lutz-Wahl, Sabine; Saguy, I Sam

    2011-05-01

    Chickpeas are of excellent quality (protein, vitamins, minerals, unsaturated fatty acids) and very low in phytoestrogen, making them a potentially promising source for vegetarian-based infant formula (VBIF). However, their high starch and fiber concentration could hinder their utilization for infants. To overcome this natural shortcoming, a solid-state "enzymation" (SSE) process was developed in which imbibition of exogenous enzyme facilitates hydrolysis within the intact chickpea seed. The process was termed seed enzyme reactor (SER). Liquid imbibition data of dry chickpeas during soaking were fitted with the Weibull distribution model. The derived Weibull shape parameter, β, value (0.77 ± 0.11) indicated that the imbibition mechanism followed Fickian diffusion. Imbibition occurred through the coat and external layers. The process was tested using green fluorescent protein (GFP) as an exogenous marker, and involved soaking, thermal treatment, peeling, microwave partial drying, rehydration in enzyme solution, and SSE at an adjusted pH, time, and temperature. Amylases, or a combination of amylases and cellulases, resulted in significant carbohydrate hydrolysis (23% and 47% of the available starch, respectively). In addition, chickpea initial raffinose and stachyose concentration was significantly reduced (91% and 92%, respectively). The process could serve as a proof of concept, requiring additional development and optimization to become a full industrial application.

  15. Nanocaged enzymes with enhanced catalytic activity and increased stability against protease digestion.

    Science.gov (United States)

    Zhao, Zhao; Fu, Jinglin; Dhakal, Soma; Johnson-Buck, Alexander; Liu, Minghui; Zhang, Ting; Woodbury, Neal W; Liu, Yan; Walter, Nils G; Yan, Hao

    2016-02-10

    Cells routinely compartmentalize enzymes for enhanced efficiency of their metabolic pathways. Here we report a general approach to construct DNA nanocaged enzymes for enhancing catalytic activity and stability. Nanocaged enzymes are realized by self-assembly into DNA nanocages with well-controlled stoichiometry and architecture that enabled a systematic study of the impact of both encapsulation and proximal polyanionic surfaces on a set of common metabolic enzymes. Activity assays at both bulk and single-molecule levels demonstrate increased substrate turnover numbers for DNA nanocage-encapsulated enzymes. Unexpectedly, we observe a significant inverse correlation between the size of a protein and its activity enhancement. This effect is consistent with a model wherein distal polyanionic surfaces of the nanocage enhance the stability of active enzyme conformations through the action of a strongly bound hydration layer. We further show that DNA nanocages protect encapsulated enzymes against proteases, demonstrating their practical utility in functional biomaterials and biotechnology.

  16. Nanocaged enzymes with enhanced catalytic activity and increased stability against protease digestion

    Science.gov (United States)

    Zhao, Zhao; Fu, Jinglin; Dhakal, Soma; Johnson-Buck, Alexander; Liu, Minghui; Zhang, Ting; Woodbury, Neal W.; Liu, Yan; Walter, Nils G.; Yan, Hao

    2016-01-01

    Cells routinely compartmentalize enzymes for enhanced efficiency of their metabolic pathways. Here we report a general approach to construct DNA nanocaged enzymes for enhancing catalytic activity and stability. Nanocaged enzymes are realized by self-assembly into DNA nanocages with well-controlled stoichiometry and architecture that enabled a systematic study of the impact of both encapsulation and proximal polyanionic surfaces on a set of common metabolic enzymes. Activity assays at both bulk and single-molecule levels demonstrate increased substrate turnover numbers for DNA nanocage-encapsulated enzymes. Unexpectedly, we observe a significant inverse correlation between the size of a protein and its activity enhancement. This effect is consistent with a model wherein distal polyanionic surfaces of the nanocage enhance the stability of active enzyme conformations through the action of a strongly bound hydration layer. We further show that DNA nanocages protect encapsulated enzymes against proteases, demonstrating their practical utility in functional biomaterials and biotechnology. PMID:26861509

  17. Enzymes in Action: An Interactive Activity Designed to Highlight Positive Attributes of Extracellular Enzymes Synthesized by Microbes

    Directory of Open Access Journals (Sweden)

    Rachel M.C. Gillespie

    2014-07-01

    Full Text Available Microbial activities are widely exploited in the manufacture of valuable products. However, the many beneficial uses of microorganisms are often overshadowed by negative associations with disease and decay. This article describes an interactive activity aimed at school-aged children and members of the public, which introduces the concept of microbial enzymes and ultimately illustrates how the industrial uses of microbes have a positive impact on everyday life. Participants are guided through a simple chemical assay which allows them to use a hands-on approach to reveal bacterial enzymes at work. This activity is safe and economical to run and is suitable for use in both the classroom and external learning environments. Also included are supplemental educational resources to support the demonstration and suggestions for extensions to the activity described, which enable further exploration of the topic. This activity has been tested by more than 2000 people at public engagement events and has received much positive feedback.

  18. Study on the Model for Regulation of the Allosteric Enzyme Activity

    Institute of Scientific and Technical Information of China (English)

    LI,Qian-Zhong(李前忠); LUO,Liao-Fu(罗辽复); ZHANG,Li-Rong(张利绒)

    2002-01-01

    The effects of activator molecule and repressive molecule on binding process between allosteric enzyme and substrate are disused by considering the heterotropic effect of the regulating molecule that binds to allosteric enzyme. A model of allosteric enzyme with heterotropic effect is presented. The cooperativity and anticooperativity in the regulation process are studied.

  19. Concurrent subacute exposure to arsenic through drinking water and malathion via diet in male rats: effects on hepatic drug-metabolizing enzymes

    Energy Technology Data Exchange (ETDEWEB)

    Naraharisetti, Suresh Babu [Indian Veterinary Research Institute, Division of Pharmacology and Toxicology, Izatnagar, Uttar Pradesh (India); University of Washington, Department of Medicinal Chemistry, Seattle, WA (United States); Aggarwal, Manoj [Indian Veterinary Research Institute, Division of Pharmacology and Toxicology, Izatnagar, Uttar Pradesh (India); Institut fuer Arbeitsphysiologie an der Universitaet Dortmund, Dortmund (Germany); Sarkar, S.N.; Malik, J.K. [Indian Veterinary Research Institute, Division of Pharmacology and Toxicology, Izatnagar, Uttar Pradesh (India)

    2008-08-15

    Arsenic is a known global groundwater contaminant, while malathion is one of the most widely used pesticides in agriculture and public health practices in the world. Here, we investigated whether repeated exposure to arsenic at the groundwater contamination levels and to malathion at sublethal levels exerts adverse effects on the hepatic drug-metabolizing system in rats, and whether concurrent exposure is more hazardous than the single agent. Male Wistar rats were exposed daily to 4 or 40 ppm of arsenic via drinking water, 50 or 500 ppm of malathion-mixed feed and in a similar fashion co-exposed to 4 ppm of arsenic and 50 ppm of malathion or 40 ppm of arsenic and 500 ppm of malathion for 28 days. At term, toxicity was assessed by evaluating changes in body weight, liver weight, levels of cytochrome P{sub 450} (CYP), cytochrome b{sub 5} and microsomal and cytosolic proteins, and activities of aminopyrine-N-demethylase (ANDM), aniline-P-hydroxylase (APH), glutathione-S-transferase (GST) and uridine diphosphate glucuronosyltransferase (UGT) in liver. Arsenic and malathion alone did not alter body weight and liver weight, but these were significantly decreased in both the co-exposed groups. These treatments decreased the activities of ANDM and APH and the levels of liver microsomal and cytosolic proteins, increased GST activity and had no effect on UGT activity. The effects of exposure to low-dose and high-dose combinations on the activities of either phase I or phase II drug-metabolizing enzymes and protein content were mostly similar to that produced by the respective low and high dose of either arsenic or malathion, except APH activity. The effect of arsenic (40 ppm) on APH activity was partially, but significantly, inhibited by malathion (500 ppm). Results indicate that the body or liver weights and the biochemical parameters were differentially affected in male rats following concurrent subacute exposure to arsenic and malathion, with the co-exposure appearing

  20. Cross-linked enzyme aggregates (CLEAs) of Pencilluim notatum lipase enzyme with improved activity, stability and reusability characteristics.

    Science.gov (United States)

    Rehman, Saima; Bhatti, Haq Nawaz; Bilal, Muhammad; Asgher, Muhammad

    2016-10-01

    Cross-linked enzyme aggregates (CLEAs) are considered as an effective tool for the immobilization of enzyme. In this study, Pencillium notatum lipase (PNL) was immobilized as carrier free cross-linked enzyme aggregates using glutaraldehyde (GLA) and Ethylene glycol-bis [succinic acid N-hydroxysuccinimide] (EG-NHS) as cross-linking agents. The optimal conditions for the synthesis of an efficient lipase CLEAs such as precipitant type, the nature and amount of cross-linking reagent, and cross-linking time were optimized. The recovered activities of CLEAs were considerably dependent on the concentration of GLA; however, the activity recovery was not severely affected by EG-NHS as a mild cross-linker. The EG-NHS aggregates displayed superior hydrolytic (52.08±2.52%) and esterification (64.42%) activities as compared to GLA aggregates which showed 23.8±1.86 and 34.54% of hydrolytic and esterification activity, respectively. Morphological analysis by fluorescence and scanning electron microscope revealed that EG-NHS aggregates were smaller in size with larger surface area compared to GLA aggregates. The pH optima of both types of CLEAs were displaced to slightly alkaline region and higher temperature as compared to native enzyme. Highest enzyme activity of CLEAs was achieved at the pH of 9.0 and 42°C temperature. Moreover, a significant improvement in the thermal resistance was also recorded after immobilization. After ten reusability cycles in aqueous medium, GLA and EG-NHS cross-linked lipase CLEAs preserved 63.62% and 70.9% of their original activities, respectively. The results suggest that this novel CLEA-lipase is potentially usable in many industrial applications.

  1. Chromium-Insulin Reduces Insulin Clearance and Enhances Insulin Signaling by Suppressing Hepatic Insulin-Degrading Enzyme and Proteasome Protein Expression in KKAy Mice.

    Science.gov (United States)

    Wang, Zhong Q; Yu, Yongmei; Zhang, Xian H; Komorowski, James

    2014-01-01

    JDS-chromium-insulin (CRI)-003 is a novel form of insulin that has been directly conjugated with chromium (Cr) instead of zinc. Our hypothesis was that CRI enhances insulin's effects by altering insulin-degrading enzyme (IDE) and proteasome enzymes. To test this hypothesis, we measured hepatic IDE content and proteasome parameters in a diabetic animal model. Male KKAy mice were randomly divided into three groups (n = 8/group); Sham (saline), human regular insulin (Reg-In), and chromium conjugated human insulin (CRI), respectively. Interventions were initiated at doses of 2 U insulin/kg body weight daily for 8-weeks. Plasma glucose and insulin were measured. Hepatic IDE, proteasome, and insulin signaling proteins were determined by western blotting. Insulin tolerance tests at week 7 showed that both insulin treatments significantly reduced glucose concentrations and increased insulin levels compared with the Sham group, CRI significantly reduced glucose at 4 and 6 h relative to Reg-In (P < 0.05), suggesting the effects of CRI on reducing glucose last longer than Reg-In. CRI treatment significantly increased hepatic IRS-1 and Akt1 and reduced IDE, 20S as well as 19S protein abundance (P < 0.01, P < 0.05, and P < 0.001, respectively), but Reg-In only significantly increased Akt1 (P < 0.05). Similar results were also observed in Reg-In- and CRI-treated HepG2 cells. This study, for the first time, demonstrates that CRI reduces plasma insulin clearance by inhibition of hepatic IDE protein expression and enhances insulin signaling as well as prevents degradation of IRS-1 and IRS-2 by suppressing ubiquitin-proteasome pathway in diabetic mice.

  2. Chromium-insulin reduces insulin clearance and enhances insulin signaling by suppressing hepatic insulin-degrading enzyme and proteasome protein expression in KKAy mice

    Directory of Open Access Journals (Sweden)

    Zhong Q Wang

    2014-07-01

    Full Text Available JDS-CRI-003 (CRI is a novel form of insulin that has been directly conjugated with chromium (Cr instead of zinc. Our hypothesis was that CRI enhances insulin’s effects by altering insulin degrading enzyme (IDE and proteasome enzymes. To test this hypothesis, we measured hepatic IDE content and proteasome parameters in a diabetic animal model. Male KKAy mice were randomly divided into three groups (n=8/group; Sham (saline, human insulin (Reg-In and chromium conjugated human insulin (CRI, respectively. Interventions were initiated at doses of 2 U insulin/kg body weight daily for eight-weeks. Plasma glucose and insulin were measured. Hepatic IDE, proteasome and insulin signaling proteins were determined by western blotting. Insulin tolerance tests at week 7 showed that both insulin treatments significantly reduced glucose concentrations and increased insulin levels compared with the Sham group, CRI significantly reduced glucose at 4 and 6 hours relative to Reg-In (P<0.05, suggesting the effects of CRI on reducing glucose last longer than Reg-In. CRI treatment significantly increased hepatic IRS-1 and Akt1 and reduced IDE, 20S as well as 19S protein abundance (P<0.01, P<0.05, and P<0.001, respectively, but Reg-In only significantly increased Akt1 (P<0.05. Similar results were also observed in Reg-In and CRI treated HepG2 cells. This study, for the first time, demonstrates that CRI reduces plasma insulin clearance by inhibition of hepatic IDE protein expression and enhances insulin signaling as well as prevents degradation of IRS-1 and IRS-2 by suppressing ubiquitin-proteasome pathway in diabetic mice.

  3. The Effect of Compaction on Urease Enzyme Activity, Carbon Dioxide Evaluation and Nitrogen Mineralisation

    OpenAIRE

    Ayten KARACA; Abdullah BARAN; KAKTANIR, Koray

    2000-01-01

    The effects of compaction on urease enzyme activity, carbon dioxide evaluation and nitrogen mineralisation of urea-treated and untreated soils were investigated. Soils were compacted at compaction levels of O kgcm -2 , 2 kgcm -2 and 4 kgcm -2 and incubated for 28 days. The changes in urease enzyme activity, CO 2 evaluation and nitrogen mineralization were determined during incubation periods. Urease enzyme activity was decreased significantly (P

  4. Detection of Sulfatase Enzyme Activity with a CatalyCEST MRI Contrast Agent.

    Science.gov (United States)

    Sinharay, Sanhita; Fernández-Cuervo, Gabriela; Acfalle, Jasmine P; Pagel, Mark D

    2016-05-01

    A chemical exchange saturation transfer (CEST) MRI contrast agent has been developed that detects sulfatase enzyme activity. The agent produces a CEST signal at δ=5.0 ppm before enzyme activity, and a second CEST signal appears at δ=9.0 ppm after the enzyme cleaves a sulfate group from the agent. The comparison of the two signals improved detection of sulfatase activity.

  5. Variability of human hepatic UDP-glucuronosyltransferase activity

    NARCIS (Netherlands)

    Little, JM; Lester, R; Kuipers, F; Vonk, R; Mackenzie, PI; Drake, RR; Frame, L; Radominska-Pandya, A

    1999-01-01

    The availability of a unique series of liver samples from human subjects, both control patients (9) and those with liver disease (6; biliary atresia (2), retransplant, chronic tyrosinemia type I, tyrosinemia, Wilson's disease) allowed us to characterize human hepatic UDP-glucuronosyltransferases usi

  6. Effect of graded Nrf2 activation on phase-I and -II drug metabolizing enzymes and transporters in mouse liver.

    Directory of Open Access Journals (Sweden)

    Kai Connie Wu

    Full Text Available Nuclear factor erythroid 2-related factor 2 (Nrf2 is a transcription factor that induces a battery of cytoprotective genes in response to oxidative/electrophilic stress. Kelch-like ECH associating protein 1 (Keap1 sequesters Nrf2 in the cytosol. The purpose of this study was to investigate the role of Nrf2 in regulating the mRNA of genes encoding drug metabolizing enzymes and xenobiotic transporters. Microarray analysis was performed in livers of Nrf2-null, wild-type, Keap1-knockdown mice with increased Nrf2 activation, and Keap1-hepatocyte knockout mice with maximum Nrf2 activation. In general, Nrf2 did not have a marked effect on uptake transporters, but the mRNAs of organic anion transporting polypeptide 1a1, sodium taurocholate cotransporting polypeptide, and organic anion transporter 2 were decreased with Nrf2 activation. The effect of Nrf2 on cytochrome P450 (Cyp genes was minimal, with only Cyp2a5, Cyp2c50, Cyp2c54, and Cyp2g1 increased, and Cyp2u1 decreased with enhanced Nrf2 activation. However, Nrf2 increased mRNA of many other phase-I enzymes, such as aldo-keto reductases, carbonyl reductases, and aldehyde dehydrogenase 1. Many genes involved in phase-II drug metabolism were induced by Nrf2, including glutathione S-transferases, UDP- glucuronosyltransferases, and UDP-glucuronic acid synthesis enzymes. Efflux transporters, such as multidrug resistance-associated proteins, breast cancer resistant protein, as well as ATP-binding cassette g5 and g8 were induced by Nrf2. In conclusion, Nrf2 markedly alters hepatic mRNA of a large number of drug metabolizing enzymes and xenobiotic transporters, and thus Nrf2 plays a central role in xenobiotic metabolism and detoxification.

  7. PPARδ activation induces hepatic long-chain acyl-CoA synthetase 4 expression in vivo and in vitro.

    Science.gov (United States)

    Kan, Chin Fung Kelvin; Singh, Amar Bahadur; Dong, Bin; Shende, Vikram Ravindra; Liu, Jingwen

    2015-05-01

    The arachidonic acid preferred long-chain acyl-CoA synthetase 4 (ACSL4) is a key enzyme for fatty acid metabolism in various metabolic tissues. In this study, we utilized hamsters fed a normal chow diet, a high-fat diet or a high cholesterol and high fat diet (HCHFD) as animal models to explore novel transcriptional regulatory mechanisms for ACSL4 expression under hyperlipidemic conditions. Through cloning hamster ACSL4 homolog and tissue profiling ACSL4 mRNA and protein expressions we observed a selective upregulation of ACSL4 in testis and liver of HCHFD fed animals. Examination of transcriptional activators of the ACSL family revealed an increased hepatic expression of PPARδ but not PPARα in HCHFD fed hamsters. To explore a role of PPARδ in dietary cholesterol-mediated upregulation of ACSL4, we administered a PPARδ specific agonist L165041 to normolipidemic and dyslipidemic hamsters. We observed significant increases of hepatic ACSL4 mRNA and protein levels in all L165041-treated hamsters as compared to control animals. The induction of ACSL4 expression by L165041 in liver tissue in vivo was recapitulated in human primary hepatocytes and hepatocytes isolated from hamster and mouse. Moreover, employing the approach of adenovirus-mediated gene knockdown, we showed that depletion of PPARδ in hamster hepatocytes specifically reduced ACSL4 expression. Finally, utilizing HepG2 as a model system, we demonstrate that PPARδ activation leads to increased ACSL4 promoter activity, mRNA and protein expression, and consequently higher arachidonoyl-CoA synthetase activity. Taken together, we have discovered a novel PPARδ-mediated regulatory mechanism for ACSL4 expression in liver tissue and cultured hepatic cells.

  8. Genistein decreases basal hepatic cytochrome P450 1A1 protein expression and activity in Swiss Webster mice.

    Science.gov (United States)

    Froyen, Erik B; Steinberg, Francene M

    2016-05-01

    Soy consumption has been associated with risk reduction for chronic diseases such as cancer. One proposed mechanism for cancer prevention by soy is through decreasing cytochrome P450 1A1 (Cyp1a1) activity. However, it is not known with certainty which soy components modulate Cyp1a1, or the characteristics or mechanisms involved in the responses after short-term (<20 days) dietary treatment without concomitant carcinogen-mediated induction. Therefore, the objective was to test the hypothesis that physiologic concentrations of dietary genistein and/or daidzein will decrease basal hepatic Cyp1a1 protein expression and activity in male and female Swiss Webster mice via inhibiting the bindings of aryl hydrocarbon receptor (AhR)-AhR nuclear translocator (ARNT) and estrogen receptor-α to the Cyp1a1 promoter region xenobiotic response element. The mice were fed the AIN-93G diet supplemented with 1500 mg/kg of genistein or daidzein for up to 1 week. Genistein, but not daidzein, significantly decreased basal hepatic microsomal Cyp1a1 protein expression and activity. AhR protein expression was not altered. Molecular mechanisms were investigated in Hepa-1c1c7 cells treated with 5 μmol/L purified aglycones genistein, daidzein, or equol. Cells treated with genistein exhibited inhibitions in ARNT and estrogen receptor-α bindings to the Cyp1a1 promoter region. This study demonstrated that genistein consumption reduced constitutive hepatic Cyp1a1 protein expression and activity, thereby contributing to the understanding of how soy isoflavone aglycones modulate cytochrome P450 biotransformation enzymes.

  9. Activation of recombinant human neutrophil procollagenase in the presence of doxycycline results in fragmentation of the enzyme and loss of enzyme activity.

    Science.gov (United States)

    Smith, G N; Brandt, K D; Hasty, K A

    1996-02-01

    To determine if reduction of collagenase activity in vitro by doxycycline (doxy) is related to activation of the proenzyme, and to determine how exogenous Ca++ and Zn++ affect the reduction. Recombinant human neutrophil procollagenase was activated with trypsin or APMA. Activity was assayed on a small peptolide substrate or on 14C-acetylated collagen fibers. The molecular weight of the proenzyme, active enzyme, and enzyme fragments was determined by Western blotting, using a polyclonal antiserum raised against the recombinant proenzyme. Similar experiments were performed in the presence of EDTA, EGTA, 1,10-phenanthroline, or doxy. The effects of exogenous Ca++ and Zn++ were also tested. Doxy inhibited activity of the enzyme against both substrates. If the drug was present during activation, the yield of activity was lower than when it was added after activation of the proenzyme. Western blotting showed that activation in the presence of doxy resulted in the appearance of lower molecular weight fragments and accumulation of less active enzyme. APMA generated prominent 28- and 26-kd fragments while trypsin cleavage yield 40- and 30-kd fragments. Fragmentation of the enzyme also occurred in the presence of EDTA or EGTA, but not 1,10-phenanthroline. It was prevented by Ca++ concentrations greater than 50 mM, but was not altered by addition of Zn++ in concentrations as high as 500 microM. Inhibition of collagenase activity by doxy could be overcome by 100 mM Ca++, but addition of Zn++ had no effect. These data suggest that doxy alters the conformation of procollagenase or collagenase by binding enzyme-associated Ca++, rendering the proteins more susceptible to proteolysis and resulting in irreversible loss of enzyme protein.

  10. Activities of five enzymes following soil disturbance and weed control in a Missouri forest

    Science.gov (United States)

    Felix, Jr. Ponder; Frieda Eivazi

    2008-01-01

    Forest disturbances associated with harvesting activities can affect soil properties including enzyme activity and overall soil quality. The activities of five enzymes (acid and alkaline phosphatases, betaglucosidase, aryl-sulfatase, and beta-glucosominidase) were measured after 8 years in soil from clearcut and uncut control plots of a Missouri oak-hickory (...

  11. Inhibition of hepatic cytochrome P450 enzymes and sodium/bile acid cotransporter exacerbates leflunomide-induced hepatotoxicity

    OpenAIRE

    Ma, Lei-Lei; Wu, Zhi-tao; Wang, Le; Zhang, Xue-Feng; Wang, Jing; Chen, Chen; Ni, Xuan; Lin, Yun-fei; Cao, Yi-yi; Luan, Yang; Pan, Guo-yu

    2016-01-01

    Aim: Leflunomide is an immunosuppressive agent marketed as a disease-modifying antirheumatic drug. But it causes severe side effects, including fatal hepatitis and liver failure. In this study we investigated the contributions of hepatic metabolism and transport of leflunomide and its major metabolite teriflunomide to leflunomide induced hepatotoxicity in vitro and in vivo. Methods: The metabolism and toxicity of leflunomide and teriflunomide were evaluated in primary rat hepatocytes in vitro...

  12. Measuring Solution Viscosity and its Effect on Enzyme Activity

    Directory of Open Access Journals (Sweden)

    Uribe Salvador

    2003-01-01

    Full Text Available In proteins, some processes require conformational changes involving structural domain diffusion. Among these processes are protein folding, unfolding and enzyme catalysis. During catalysis some enzymes undergo large conformational changes as they progress through the catalytic cycle. According to Kramers theory, solvent viscosity results in friction against proteins in solution, and this should result in decreased motion, inhibiting catalysis in motile enzymes. Solution viscosity was increased by adding increasing concentrations of glycerol, sucrose and trehalose, resulting in a decrease in the reaction rate of the H+-ATPase from the plasma membrane of Kluyveromyces lactis. A direct correlation was found between viscosity (&eegr; and the inhibition of the maximum rate of catalysis (V max. The protocol used to measure viscosity by means of a falling ball type viscometer is described, together with the determination of enzyme kinetics and the application of Kramers’ equation to evaluate the effect of viscosity on the rate of ATP hydrolysis by the H+-ATPase.

  13. AMP-activated protein kinase and coordination of hepatic fatty acid metabolism of starved/carbohydrate-refed rats.

    Science.gov (United States)

    Assifi, Murwarid M; Suchankova, Gabriela; Constant, Scarlet; Prentki, Marc; Saha, Asish K; Ruderman, Neil B

    2005-11-01

    Acute increases in the concentration of malonyl-CoA play a pivotal role in mediating the decrease in fatty acid oxidation that occurs in many tissues during refeeding after a fast. In this study, we assess whether such increases in malonyl-CoA in liver could be mediated by malonyl-CoA decarboxylase (MCD), as well as acetyl-CoA carboxylase (ACC). In addition, we examine how changes in the activity of ACC, MCD, and other enzymes that govern fatty acid and glycerolipid synthesis relate temporally to alterations in the activities of the fuel-sensing enzyme AMP-activated protein kinase (AMPK). Rats starved for 48 h and refed a carbohydrate chow diet for 1, 3, 12, and 24 h were studied. Refeeding caused a 40% decrease in the activity of the alpha1-isoform of AMPK within 1 h, with additional decreases in AMPKalpha1 activity and a decrease in AMPKalpha2 occurring between 1 and 24 h. At 1 h, the decrease in AMPK activity was associated with an eightfold increase in the activity of the alpha1-isoform of ACC and a 30% decrease in the activity of MCD, two enzymes thought to be regulated by AMPK. Also, the concentration of malonyl-CoA was increased by 50%. Between 1 and 3 h of refeeding, additional increases in the activity of ACC and decreases in MCD were observed, as was a further twofold increase in malonyl-CoA. Increases in the activity (60%) and abundance (12-fold) of fatty acid synthase occurred predominantly between 3 and 24 h and increases in the activity of mitochondrial sn-glycerol-3-phosphate acyltransferase (GPAT) and acyl-CoA:diaclyglycerol acyltransferase (DGAT) at 12 and 24 h. The results strongly suggest that early changes in the activity of MCD, as well as ACC, contribute to the increase in hepatic malonyl-CoA in the starved-refed rat. They also suggest that the changes in these enzymes, and later occurring increases in enzymes regulating fatty acid and glycerolipid synthesis, could be coordinated by AMPK.

  14. Plasmodium Infection Is Associated with Impaired Hepatic Dimethylarginine Dimethylaminohydrolase Activity and Disruption of Nitric Oxide Synthase Inhibitor/Substrate Homeostasis.

    Directory of Open Access Journals (Sweden)

    Jessica H Chertow

    2015-09-01

    Full Text Available Inhibition of nitric oxide (NO signaling may contribute to pathological activation of the vascular endothelium during severe malaria infection. Dimethylarginine dimethylaminohydrolase (DDAH regulates endothelial NO synthesis by maintaining homeostasis between asymmetric dimethylarginine (ADMA, an endogenous NO synthase (NOS inhibitor, and arginine, the NOS substrate. We carried out a community-based case-control study of Gambian children to determine whether ADMA and arginine homeostasis is disrupted during severe or uncomplicated malaria infections. Circulating plasma levels of ADMA and arginine were determined at initial presentation and 28 days later. Plasma ADMA/arginine ratios were elevated in children with acute severe malaria compared to 28-day follow-up values and compared to children with uncomplicated malaria or healthy children (p<0.0001 for each comparison. To test the hypothesis that DDAH1 is inactivated during Plasmodium infection, we examined DDAH1 in a mouse model of severe malaria. Plasmodium berghei ANKA infection inactivated hepatic DDAH1 via a post-transcriptional mechanism as evidenced by stable mRNA transcript number, decreased DDAH1 protein concentration, decreased enzyme activity, elevated tissue ADMA, elevated ADMA/arginine ratio in plasma, and decreased whole blood nitrite concentration. Loss of hepatic DDAH1 activity and disruption of ADMA/arginine homeostasis may contribute to severe malaria pathogenesis by inhibiting NO synthesis.

  15. Application of a Newly Developed High-Sensitivity HBsAg Chemiluminescent Enzyme Immunoassay for Hepatitis B Patients with HBsAg Seroclearance

    OpenAIRE

    2013-01-01

    We modified and automated a highly sensitive chemiluminescent enzyme immunoassay (CLEIA) for surface antigen (HBsAg) detection using a combination of monoclonal antibodies, each for a specific epitope of HBsAg, and by improving an earlier conjugation technique. Of 471 hepatitis B virus (HBV) carriers seen in our hospital between 2009 and 2012, 26 were HBsAg seronegative as determined by the Abbott Architect assay. The Lumipulse HBsAg-HQ assay was used to recheck those 26 patients who demonstr...

  16. Finding Biomass Degrading Enzymes Through an Activity-Correlated Quantitative Proteomics Platform (ACPP)

    Science.gov (United States)

    Ma, Hongyan; Delafield, Daniel G.; Wang, Zhe; You, Jianlan; Wu, Si

    2017-04-01

    The microbial secretome, known as a pool of biomass (i.e., plant-based materials) degrading enzymes, can be utilized to discover industrial enzyme candidates for biofuel production. Proteomics approaches have been applied to discover novel enzyme candidates through comparing protein expression profiles with enzyme activity of the whole secretome under different growth conditions. However, the activity measurement of each enzyme candidate is needed for confident "active" enzyme assignments, which remains to be elucidated. To address this challenge, we have developed an Activity-Correlated Quantitative Proteomics Platform (ACPP) that systematically correlates protein-level enzymatic activity patterns and protein elution profiles using a label-free quantitative proteomics approach. The ACPP optimized a high performance anion exchange separation for efficiently fractionating complex protein samples while preserving enzymatic activities. The detected enzymatic activity patterns in sequential fractions using microplate-based assays were cross-correlated with protein elution profiles using a customized pattern-matching algorithm with a correlation R-score. The ACPP has been successfully applied to the identification of two types of "active" biomass-degrading enzymes (i.e., starch hydrolysis enzymes and cellulose hydrolysis enzymes) from Aspergillus niger secretome in a multiplexed fashion. By determining protein elution profiles of 156 proteins in A. niger secretome, we confidently identified the 1,4-α-glucosidase as the major "active" starch hydrolysis enzyme (R = 0.96) and the endoglucanase as the major "active" cellulose hydrolysis enzyme (R = 0.97). The results demonstrated that the ACPP facilitated the discovery of bioactive enzymes from complex protein samples in a high-throughput, multiplexing, and untargeted fashion.

  17. Finding Biomass Degrading Enzymes Through an Activity-Correlated Quantitative Proteomics Platform (ACPP)

    Science.gov (United States)

    Ma, Hongyan; Delafield, Daniel G.; Wang, Zhe; You, Jianlan; Wu, Si

    2017-01-01

    The microbial secretome, known as a pool of biomass (i.e., plant-based materials) degrading enzymes, can be utilized to discover industrial enzyme candidates for biofuel production. Proteomics approaches have been applied to discover novel enzyme candidates through comparing protein expression profiles with enzyme activity of the whole secretome under different growth conditions. However, the activity measurement of each enzyme candidate is needed for confident "active" enzyme assignments, which remains to be elucidated. To address this challenge, we have developed an Activity-Correlated Quantitative Proteomics Platform (ACPP) that systematically correlates protein-level enzymatic activity patterns and protein elution profiles using a label-free quantitative proteomics approach. The ACPP optimized a high performance anion exchange separation for efficiently fractionating complex protein samples while preserving enzymatic activities. The detected enzymatic activity patterns in sequential fractions using microplate-based assays were cross-correlated with protein elution profiles using a customized pattern-matching algorithm with a correlation R-score. The ACPP has been successfully applied to the identification of two types of "active" biomass-degrading enzymes (i.e., starch hydrolysis enzymes and cellulose hydrolysis enzymes) from Aspergillus niger secretome in a multiplexed fashion. By determining protein elution profiles of 156 proteins in A. niger secretome, we confidently identified the 1,4-α-glucosidase as the major "active" starch hydrolysis enzyme (R = 0.96) and the endoglucanase as the major "active" cellulose hydrolysis enzyme (R = 0.97). The results demonstrated that the ACPP facilitated the discovery of bioactive enzymes from complex protein samples in a high-throughput, multiplexing, and untargeted fashion.

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

    Directory of Open Access Journals (Sweden)

    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

  19. Effect of curcumin supplementation on blood glucose, plasma insulin, and glucose homeostasis related enzyme activities in diabetic db/db mice.

    Science.gov (United States)

    Seo, Kwon-Il; Choi, Myung-Sook; Jung, Un Ju; Kim, Hye-Jin; Yeo, Jiyoung; Jeon, Seon-Min; Lee, Mi-Kyung

    2008-09-01

    We investigated the effect of curcumin on insulin resistance and glucose homeostasis in male C57BL/KsJ-db/db mice and their age-matched lean non-diabetic db/+ mice. Both db/+ and db/db mice were fed with or without curcumin (0.02%, wt/wt) for 6 wks. Curcumin significantly lowered blood glucose and HbA 1c levels, and it suppressed body weight loss in db/db mice. Curcumin improved homeostasis model assessment of insulin resistance and glucose tolerance, and elevated the plasma insulin level in db/db mice. Hepatic glucokinase activity was significantly higher in the curcumin-supplemented db/db group than in the db/db group, whereas glucose-6-phosphatase and phosphoenolpyruvate carboxykinase activities were significantly lower. In db/db mice, curcumin significantly lowered the hepatic activities of fatty acid synthase, beta-oxidation, 3-hydroxy-3-methylglutaryl coenzyme reductase, and acyl-CoA: cholesterol acyltransferase. Curcumin significantly lowered plasma free fatty acid, cholesterol, and triglyceride concentrations and increased the hepatic glycogen and skeletal muscle lipoprotein lipase in db/db mice. Curcumin normalized erythrocyte and hepatic antioxidant enzyme activities (superoxide dismutase, catalase, gluthathione peroxidase) in db/db mice that resulted in a significant reduction in lipid peroxidation. However, curcumin showed no effect on the blood glucose, plasma insulin, and glucose regulating enzyme activities in db/+ mice. These results suggest that curcumin seemed to be a potential glucose-lowering agent and antioxidant in type 2 diabetic db/db mice, but had no affect in non-diabetic db/+ mice.

  20. Correlation of biological serum markers with the degree of hepatic fibrosis and necroinflammatory activity in hepatitis C and schistosomiasis patients

    Directory of Open Access Journals (Sweden)

    Clarice Neuenschwander Lins de Morais

    2010-07-01

    Full Text Available Liver biopsy is the gold-standard method to stage fibrosis; however, it is an invasive procedure and is potentially dangerous. The main objective of this study was to evaluate biological markers, such as cytokines IL-13, IFN-γ, TNF-α and TGF-β, platelets, bilirubins (Bil, alanine aminotransferase (ALT and aspartate aminotransferase (AST, total proteins, γ-glutamil transferase (γ-GT and alkaline phosphatase (AP, that could be used to predict the severity of hepatic fibrosis in schistosomiasis and hepatitis C (HC as isolated diseases or co-infections. The following patient groups were selected: HC (n = 39, HC/hepatosplenic schistosomiasis (HSS (n = 19, HSS (n = 22 and a control group (n = 13. ANOVA and ROC curves were used for statistical analysis. P < 0.05 was considered significant. With HC patients we showed that TNF-α (p = 0.020 and AP (p = 0.005 could differentiate mild and severe fibrosis. With regard to necroinflammatory activity, AST (p = 0.002, γ-GT (p = 0.034 and AP (p = 0.001 were the best markers to differentiate mild and severe activity. In HC + HSS patients, total Bil (p = 0.008 was capable of differentiating between mild and severe fibrosis. In conclusion, our study was able to suggest biological markers that are non-invasive candidates to evaluate fibrosis and necroinflammatory activity in HC and HC + HSS.

  1. Guanidinylated neomycin mediates heparan sulfate-dependent transport of active enzymes to lysosomes.

    Science.gov (United States)

    Sarrazin, Stéphane; Wilson, Beth; Sly, William S; Tor, Yitzhak; Esko, Jeffrey D

    2010-07-01

    Guanidinylated neomycin (GNeo) can transport bioactive, high molecular weight cargo into the interior of cells in a process that depends on cell surface heparan sulfate proteoglycans. In this report, we show that GNeo-modified quantum dots bind to cell surface heparan sulfate, undergo endocytosis and eventually reach the lysosomal compartment. An N-hydroxysuccinimide activated ester of GNeo (GNeo-NHS) was prepared and conjugated to two lysosomal enzymes, beta-D-glucuronidase (GUS) and alpha-L-iduronidase. Conjugation did not interfere with enzyme activity and enabled binding of the enzymes to heparin-Sepharose and heparan sulfate on primary human fibroblasts. Cells lacking the corresponding lysosomal enzyme took up sufficient amounts of the conjugated enzymes to restore normal turnover of glycosaminoglycans. The high capacity of proteoglycan-mediated uptake suggests that this method of delivery might be used for enzyme replacement or introduction of foreign enzymes into cells.

  2. Activation of peroxisome proliferator-activated receptor gamma by rosiglitazone increases sirt6 expression and ameliorates hepatic steatosis in rats.

    Directory of Open Access Journals (Sweden)

    Soo Jin Yang

    Full Text Available BACKGROUND: Sirt6 has been implicated in the regulation of hepatic lipid metabolism and the development of hepatic steatosis. The aim of this study was to address the potential role of Sirt6 in the protective effects of rosiglitazone (RGZ on hepatic steatosis. METHODS: To investigate the effect of RGZ on hepatic steatosis, rats were treated with RGZ (4 mg·kg⁻¹·day⁻¹ by stomach gavage for 6 weeks. The involvement of Sirt6 in the RGZ's regulation was evaluated by Sirt6 knockdown in AML12 mouse hepatocytes. RESULTS: RGZ treatment ameliorated hepatic lipid accumulation and increased expression of Sirt6, peroxisome proliferator-activated receptor gamma coactivtor-1-α (Ppargc1a/PGC1-α and Forkhead box O1 (Foxo1 in rat livers. AMP-activated protein kinase (AMPK phosphorylation was also increased by RGZ, accompanied by alterations in phosphorylation of LKB1. Interestingly, in free fatty acid-treated cells, Sirt6 knockdown increased hepatocyte lipid accumulation measured as increased triglyceride contents (p = 0.035, suggesting that Sirt6 may be beneficial in reducing hepatic fat accumulation. In addition, Sirt6 knockdown abolished the effects of RGZ on hepatocyte fat accumulation, mRNA and protein expression of Ppargc1a/PGC1-α and Foxo1, and phosphorylation levels of LKB1 and AMPK, suggesting that Sirt6 is involved in RGZ-mediated metabolic effects. CONCLUSION: Our results demonstrate that RGZ significantly decreased hepatic lipid accumulation, and that this process appeared to be mediated by the activation of the Sirt6-AMPK pathway. We propose Sirt6 as a possible therapeutic target for hepatic steatosis.

  3. EFFECT OF GREEN TEA AND ITS COMPONENTS ON BODY WEIGHT, FAT ACCUMULATION AND ACTIVITY OF HEPATIC FAT METABOLISM RELATED ENZYMES IN MICE%绿茶及其成分对小鼠体重、脂肪沉积和肝脏脂肪代谢酶活性的影响

    Institute of Scientific and Technical Information of China (English)

    郑国栋; 黎冬明; 张清峰

    2012-01-01

    Objective To investigate the anti-obesity mechanism of green tea and its components in mice. Method Fifty female Kunming mice were randomly assigned to 5 groups and fed on commercial powder diets containing none (control), 2% green tea, 0.05% caffeine, 0.3% catechins and 0.05% caffeine + 0.3% catechins for 8 w respectively. Liver and intraperitoneal adipose tissues (IPAT) were weighed at the end of experiment. Serum levels of glucose, triglycerides (TG), total cholesterols, leptin and insulin, and lipids in liver were measured. Fatty acid synthese (FAS), carnitine acyltransferase (CAT) and acyl-CoA oxidase (ACO) in liver were determined. Results The body weight gain and IPAT weight were significantly reduced by green tea, caffeine and caffeine + catechins. Serum TG and leptin concentrations were significantly decreased by caffeine + catechins. Green tea, catechins and caffeine + catechins reduced TG level in liver. Caffeine + catechins significantly reduced FAS activity, and remarkably increased CAT and ACO activities in liver. Conclusion Caffeine + catechins can inhibit fat accumulation and body weight gain in mice through regulating fat metabolism related enzymes activities. Moreover, caffeine and catechins possess synergistic anti-obesity effect.%目的 研究绿茶及其成分的减肥作用机制.方法 50只雌性昆明小鼠随机分成5组,分别为对照组、2%绿茶组、0.05%咖啡碱组、0.3%儿茶素组及0.05%咖啡碱+0.3%儿茶素组.混合饲料投喂8w后,摘取肝脏和腹腔脂肪(IPAT)并称重.分析血清中血糖、甘油三酯(TG)、总胆固醇、瘦素及胰岛素浓度和肝脏中脂类含量.测定肝脏中脂肪酸合成酶(FAS)、肉毒碱脂酰转移酶(CAT)和酰基辅酶A氧化酶(ACO)的活性.结果 绿茶、咖啡碱、咖啡碱+儿茶素组体重增加和IPAT重量比对照组明显减少,特别是咖啡碱+儿茶素效果极显著.咖啡碱+儿茶素显著降低血中TG和瘦素浓度.绿茶、儿茶素、咖啡

  4. Human amniotic epithelial cell transplantation induces markers of alternative macrophage activation and reduces established hepatic fibrosis.

    Directory of Open Access Journals (Sweden)

    Ursula Manuelpillai

    Full Text Available Chronic hepatic inflammation from multiple etiologies leads to a fibrogenic response that can progress to cirrhosis and liver failure. Transplantation of human amniotic epithelial cells (hAEC from term delivered placenta has been shown to decrease mild to moderate hepatic fibrosis in a murine model. To model advanced human liver disease and assess the efficacy of hAEC therapy, we transplanted hAEC in mice with advanced hepatic fibrosis. Immunocompetent C57BL/6 mice were administered carbon tetrachloride (CCl(4 twice weekly resulting in bridging fibrosis by 12 weeks. hAEC (2 × 10(6 were infused via the tail vein at week 8 or weeks 8 and 10 (single and double dose, respectively. Human cells were detected in mouse liver four weeks after transplantation showing hAEC engraftment. CCl(4 treated mice receiving single or double hAEC doses showed a significant but similar decrease in liver fibrosis area associated with decreased activation of collagen-producing hepatic stellate cells and decreased hepatic protein levels of the pro-fibrogenic cytokine, transforming growth factor-beta1. CCl(4 administration caused hepatic T cell infiltration that decreased significantly following hAEC transplantation. Hepatic macrophages play a crucial role in both fibrogenesis and fibrosis resolution. Mice exposed to CCl(4 demonstrated increased numbers of hepatic macrophages compared to normal mice; the number of macrophages decreased significantly in CCl(4 treated mice given hAEC. These mice had significantly lower hepatic protein levels of the chemokine monocyte chemoattractant protein-1 than mice given CCl(4 alone. Alternatively activated M2 macrophages are associated with fibrosis resolution. CCl(4 treated mice given hAEC showed increased expression of genes associated with M2 macrophages including YM-1, IL-10 and CD206. We provide novel data showing that hAEC transplantation induces a wound healing M2 macrophage phenotype associated with reduction of established

  5. Antioxidative capacity and enzyme activity in Haematococcus pluvialis cells exposed to superoxide free radicals

    Institute of Scientific and Technical Information of China (English)

    刘建国; 张晓丽; 孙延红; 林伟

    2010-01-01

    The antioxidative capacity of astaxanthin and enzyme activity of reactive oxygen eliminating enzymes such as superoxide dismutase (SOD),peroxidase (POD),catalase (CAT) and ascorbate peroxidase (APX) were studied in three cell types of Haematococcus pluvialis exposed to high concentrations of a superoxide anion radical (O2ˉ).The results show that defensive enzymes and astaxanthin-related mechanisms were both active in H.pluvialis during exposure to reactive oxygen species (ROS) such as Oˉ2.Astaxanthin reacte...

  6. Effect of Cereal Type and Enzyme Addition on Performance, Pancreatic Enzyme Activity, Intestinal Microflora and Gut Morphology of Broilers

    Directory of Open Access Journals (Sweden)

    Kalantar M

    2016-06-01

    Full Text Available The effects of grain and carbohydrase enzyme supplementation were investigated on digestive physiology of chickens. A total of 625 one-day-old chicks (Ross 308 were randomly assigned to five treatments in a completely randomized design. Treatments included two different types of grains (wheat, and barley with or without a multi-carbohydrase supplement. A corn-based diet was also considered to serve as a control. Feeding barley-based diet with multi-carbohydrase led to higher feed intake (P < 0.01 than those fed corn- and wheat-based diets. Birds fed on barley and wheat diets had lower weight gain despite a higher feed conversion ratio (P < 0.01. Total count and number of different type of bacteria including Gram-negative, E. coli, and Clostridia increased after feeding wheat and barley but the number of Lactobacilli and Bifidobacteria decreased (P < 0.01. Feeding barley and wheat diets reduced villus height in different parts of the small intestine when compared to those fed on a corn diet. However, enzyme supplementation of barley and wheat diets improved weight gain and feed conversion ratio and resulted in reduced number of E. coli and Clostridia and increased number of Lactobacilli and Bifidobacteria, and also restored the negative effects on intestinal villi height (P < 0.01. The activities of pancreatic α-amylase and lipase were (P < 0.01 increased in chickens fed wheat and barley diets when compared to the control fed on a corn diet. Enzyme supplementation reduced the activities of pancreatic α-amylase and lipase (P < 0.01. In conclusion, various dietary non-starch polysaccharides without enzyme supplementation have an adverse effect on digesta viscosity, ileal microflora, villi morphology, and pancreatic enzyme activity.

  7. Decreased peripheral natural killer cells activity in the immune activated stage of chronic hepatitis B.

    Directory of Open Access Journals (Sweden)

    Yuan Li

    Full Text Available BACKGROUND & AIMS: The natural course of chronic hepatitis B virus (HBV infection is characterized by different immune responses, ranging from immune tolerant (IT to immune activated (IA stages. In our study, we investigated the natural killer (NK cells activity in patients at different immunological stages of chronic HBV infection. METHODS: Blood samples obtained from 57 HBeAg positive patients with chronic hepatitis B (CHB, including 15 patients in the immune tolerant (IT stage, 42 patients in the immune activated (IA stage, and 18 healthy individuals (HI. The analyses included flow cytometry to detect NK cells, the determination of cytokine levels as well as of surface receptor expression and cytotoxicity. RESULTS: NK cells in peripheral blood were significantly lower in patients in the IA stage of CHB compared to HI (p<0.05. Patients in the IA stage of CHB had lower levels of NK cells activating receptor NKp30 and NKG2D expression, cytokine interferon-γ (IFN-γ and tumor necrosis factor-α (TNF-α production, as compared to patients in the IT stage and HI, respectively (p<0.05. Cytotoxicity of NK cells was lower in patients in the IA stage of CHB compared to patients in the IT stage and HI, respectively (p<0.05. The level of IFN-γ but not level of TNF-α and cytotoxicity of NK cells was inversely correlated with serum HBV load in patients with CHB. Peripheral NK cells activity did not correlate with ALT level. CONCLUSION: NK cells activity was lower in CHB patients, especially in those in the IA stage.

  8. The effect of the P1 side chain on the binding of optimized carboxylate and activated carbonyl inhibitors of the hepatitis C virus NS3 protease.

    Science.gov (United States)

    Kawai, Stephen H; LaPlante, Steven R; Llinàs-Brunet, Montse; Hucke, Oliver

    2010-07-01

    Peptidyl inhibitors of the hepatitis C virus NS3 protease hold much promise as direct-acting antiviral agents against hepatitis C infection. The optimization of N-terminal cleavage products, found to exhibit activity (product inhibition) against the enzyme, has led to potent tripeptide inhibitors that bear free C-terminal carboxylate groups. An analogous activated carbonyl compound (pentafluoroethyl ketone) bearing a P1 norvaline (Nva) was found to possess comparable activity against hepatitis C virus protease. However, an analogue bearing an aminocyclopropylcarboxylic acid (Acca) P1 residue exhibited very poor activity. (19)F-NMR studies indicate that the propensity of the Acca-derived activated carbonyl to form hemiketals is only slightly reduced compared with that of a P1 Nva equivalent. These results, as well as molecular modeling studies, argue against steric hindrance of the nucleophilic attack of Ser-139 accounting for the poor mechanism-based inhibition by the former. We hypothesize that the conformational properties of the respective C-termini in the context of an adaptable active site account for the divergent P1 structure-activity relationships.

  9. Changes in the spectrum and rates of extracellular enzyme activities in seawater following aggregate formation

    Science.gov (United States)

    Ziervogel, K.; Steen, A. D.; Arnosti, C.

    2010-03-01

    Marine snow aggregates are heavily colonized by heterotrophic microorganisms that express high levels of hydrolytic activities, making aggregates hotspots for carbon remineralization in the ocean. To assess how aggregate formation influences the ability of seawater microbial communities to access organic carbon, we compared hydrolysis rates of six polysaccharides in coastal seawater after aggregates had been formed (via incubation on a roller table) with hydrolysis rates in seawater from the same site that had not incubated on a roller table (referred to as whole seawater). Hydrolysis rates in the aggregates themselves were up to three orders of magnitude higher on a volume basis than in whole seawater. The enhancement of enzyme activity in aggregates relative to whole seawater differed by substrate, suggesting that the enhancement was under cellular control, rather than due to factors such as lysis or grazing. A comparison of hydrolysis rates in whole seawater with those in aggregate-free seawater, i.e. the fraction of water from the roller bottles that did not contain aggregates, demonstrated a nuanced microbial response to aggregate formation. Activities of laminarinase and xylanase enzymes in aggregate-free seawater were higher than in whole seawater, while activities of chondroitin, fucoidan, and arabinogalactan hydrolyzing enzymes were lower than in whole seawater. These data suggest that aggregate formation enhanced production of laminarinase and xylanase enzymes, and the enhancement also affected the surrounding seawater. Decreased activities of chondroitin, fucoidan, and arabinoglactan-hydrolyzing enzymes in aggregate-free seawaters relative to whole seawater are likely due to shifts in enzyme production by the aggregate-associated community, coupled with the effects of enzyme degradation. Enhanced activities of laminarin- and xylan-hydrolyzing enzymes in aggregate-free seawater were due at least in part to cell-free enzymes. Measurements of enzyme

  10. Protective Immunity to Hepatitis B and Streptococcus Pneumoniae in Active Duty Women Versus Men: Prevalence and Responses to Preventive Immunization

    Science.gov (United States)

    1996-04-01

    Protective Immunity to Hepatitis B and Streptococcus Pneumoniae in Active Duty Women Versus Men: Prevalence and Responses to Preventive Immunization...April 1996 I Final (1 Dec 94 - 31 Dec 95) 4. TITLE AND SUBTITLE Prot~ecti•ve Inmnunity¥ to Hepat~it~is B and 6. FUNDING NUMBERS Streptococcus Pneumoniae in...pneumococcal vaccine is not included in the standard vaccinations for active duty military. The prevalence of immunity to pathogenic Streptococcus pneumoniae in

  11. Hepatic Insulin Resistance Following Chronic Activation of the CREB Coactivator CRTC2

    DEFF Research Database (Denmark)

    Hogan, Meghan F; Ravnskjaer, Kim; Matsumura, Shigenobu

    2015-01-01

    and dephosphorylation of the cAMP regulated CREB coactivators CRTC2 and CRTC3. In parallel, decreases in circulating insulin also increase gluconeogenic gene expression via the de-phosphorylation and activation of the forkhead transcription factor FOXO1. Hepatic gluconeogenesis is increased in insulin resistance where...... accompanying decreases in FOXO1 activity, hepatic gluconeogenic gene expression remained elevated in CRTC2S171,275A mice demonstrating that chronic increases in CRTC2 activity in the liver are indeed sufficient to promote hepatic insulin resistance and to disrupt glucose homeostasis....... increased gluconeogenic gene expression under fasting as well as feeding conditions. Circulating glucose concentrations were constitutively elevated in CRTC2S171,275A expressing mice, leading to compensatory increases in circulating insulin concentrations that enhance FOXO1 phosphorylation. Despite...

  12. Studies on the function of hepatic lipase in the cat after immunological blockade of the enzyme in vivo.

    Science.gov (United States)

    Demacker, P N; Hijmans, A G; Stalenhoef, A F; van 't Laar, A

    1988-02-01

    In order to investigate the in vivo function of hepatic lipase, cats were injected with anti-cat hepatic lipase antibodies which produced a complete and specific inhibition of heparin-releasable hepatic lipase. The cat was chosen as an animal model because it displays, like man, a relative deficiency of lipoprotein lipase compared to hepatic lipase and because the possession of two subfractions of high density lipoproteins, HDL2 and HDL3. In fasted cats no changes were observed in plasma triglycerides or phospholipids. In fed animals triglycerides increased considerably, indicating that hepatic lipase may have a function in the postprandial phase. In fat-loaded cats (6 g of fat/kg) triglycerides in the d less than 1.019 g/ml fraction increased from 4 h after the blockade due to accumulation of lipoproteins with pre-beta-mobility containing the apoproteins, apo B-100, apo E and apo A-I. Apo B-48 did not accumulate consistently. Phospholipids in the HDL2-fraction and those in the HDL3-fraction of the fat-loaded cats tended to increase and decrease from 6 and 9 h after the blockade, respectively. The absolute change in HDL2 phospholipids approximated that of HDL3-phospholipids. Overall, the density of HDL particles decreased, apparently secondary to the accumulation of apo A-I in the d less than 1.019 g/ml fraction. Our findings suggest that hepatic lipase is involved in the hydrolysis of a special class of apo A-I containing triglyceride-rich lipoproteins synthesised in the postprandial phase.

  13. Characterization of E-NTPDase (EC 3.6.1.5) activity in hepatic lymphocytes: A different activity profile from peripheral lymphocytes.

    Science.gov (United States)

    Doleski, Pedro H; Adefegha, Stephen A; Cabral, Fernanda L; Leal, Daniela B R

    2017-03-01

    The activity of ectonucleoside triphosphate diphosphohydrolase (E-NTPDase; EC 3.6.1.5) was characterized in hepatic lymphocytes (HL) of rats. For this purpose, a specific method for the isolation of lymphocytes from hepatic tissue was developed. Subsequently, E-NTPDase activity of rat HL was compared with that of rat peripheral lymphocytes. The HL showed high cell count and viability. Also, the characterization test revealed that the optimal E-NTPDase activities were attained at 37°C and pH 8.0 in the presence of Ca(2+) . In addition, in the presence of specific E-NTPDase inhibitors (20mM sodium azide and 0.3mM suramin), there were significant inhibitions in nucleotide hydrolysis. However, there was no significant change in adenosine triphosphate (ATP) or adenosine diphosphate (ADP) hydrolysis in the presence of inhibitors of other E-ATPase (0.1mM Ouabain, 0.5mM orthovanadate, and 1mM, 5mM, and 10mM sodium azide). Furthermore, the kinetic behavior of the enzyme in HL showed apparent Km of 134.90 ± 0.03μM and 214.40 ± 0.06μM as well as Vmax of 345.0 ± 28.32 and 242.0 ± 27.55 ƞmol Pi/min/mg of protein for ATP and ADP, respectively. The Chevillard plot revealed that ATP and ADP were hydrolyzed at the same active site of the enzyme. Our results suggest that the degradation of extracellular nucleotides in HL may have been primarily accomplished by E-NTPDase. The higher E-NTPDase activity observed in HL may be attributed to the important physiological functions of ATP and ADP in HL.

  14. Effects of Straw Biomass Charcoal on Enzyme Activity in Cd Contaminated Soil

    Directory of Open Access Journals (Sweden)

    SHANG Yi-jie

    2015-02-01

    Full Text Available Selected straw biomass charcoal as raw material for the simulation, this study mainly explore the effect of different amount of biomass charcoal applied on different categories of Cd contaminated soil enzyme activity. The results showed that when the soil Cd content was 5 mg·kg-1 and soil carbon cycle enzymes was with different amount of biomass carbon, the FDA hydrolase and protease were more sensi-tive to Cd contaminated soil; and under soil oxidoreductase in the Cd polluted soil condition with different amount of biomass carbon into soil, urease enzyme and phosphatase enzyme were more sensitivitive. Phosphatase was more significant which changed 79.40% compared soil without Cd. At the same time, calculated the geometric mean of carbon cycle enzymes, oxidoreductase enzymes and the two types of the over-all enzyme were as a measure of its synthesis enzymes activity. Among them, the comprehensive index of the soil carbon cycle enzymes activi-ty lay between 0.071~0.235, and when biomass carbon intake was 2.5%, the value was 0.174, which was higher 7.4%and 19.5%respectively than the enzyme without biomass and biomass carbon dosage was 5%; the comprehensive index of the oxidoreductase enzyme lay between 0.093~0.202, and when biomass carbon intake was 2.5%, the value was 0.131, which was higher 18.50% and 28.90% respectively than the enzyme without biomass and biomass carbon dosage was 5%; the comprehensive index of the soil composite enzyme index lay between 0.077~0.167, and when biomass carbon intake was 2.5%, the value was 0.108, which was higher 16.26% and 28.57% respectively than the enzyme without biomass and biomass carbon dosage was 5%.

  15. Physicochemical Properties and Enzymes Activity Studies in a ...

    African Journals Online (AJOL)

    JTEkanem

    2009-11-04

    Nov 4, 2009 ... Soil Physicochemical properties and enzyme concentration were evaluated in soil from a refined-oil ... the effects of oil pollution on soil, aquatic life, ground and ..... to anthropogenic origin increase in the metal contents in the ...

  16. Mining anaerobic digester consortia metagenomes for secreted carbohydrate active enzymes

    DEFF Research Database (Denmark)

    Wilkens, Casper; Busk, Peter Kamp; Pilgaard, Bo

    was done with the Peptide Pattern Recognition (PPR) program (Busk and Lange, 2013), which is a novel non-alignment based approach that can predict function of e.g. CAZymes. PPR identifies a set of short conserved sequences, which can be used as a finger print when mining genomes for novel enzymes. In both...

  17. [Glycolytic activity of enzyme preparation from the red king crab (Paralithodes camtschaticus) hepatopancreas].

    Science.gov (United States)

    Rysakova, K S; Novikov, V Iu; Mukhin, V A; Serafimchik, E M

    2008-01-01

    Enzyme preparation exhibiting glycolytic activity yielding chitooligosaccharides along with N-acetyl-D-glucosamine was obtained from the red king crab (Paralithodes camtschaticus) hepatopancreas. The results of the analysis confirmed the presence of endo- and exochitinase activities in the preparation. HPLC showed that the hydrolysis products of chitin and chitosan did not contain D(+)-glucosamine, which is indicative of the absence of deacetylase and, apparently, exochitosanase activities. A comparison of the dependence of the enzyme preparation activity on temperature and pH of the incubation medium suggests that chitinase and protease activities are exhibited by different enzymes.

  18. Immunoliposomes as enzyme-carriers (immuno-enzymosomes) for antibody-directed enzyme prodrug therapy (ADEPT) : Optimization of prodrug activating capacity

    NARCIS (Netherlands)

    Vingerhoeds, MH; Haisma, HJ; Belliot, SO; Smit, RHP; Crommelin, DJA; Storm, G

    Purpose. Immuno-enzymosomes are tumor-specific immunoliposomes bearing enzymes on their surface. These enzymes are capable of converting relatively nontoxic prodrugs into active cytostatic agents. The enzyme beta-glucuronidase (GUS)(4) was coupled to the external surface of immunoliposomes directed

  19. Liposome-mediated targeting of enzymes to cancer cells for site-specific activation of prodrugs : Comparison with the corresponding antibody-enzyme conjugate

    NARCIS (Netherlands)

    Fonseca, Maria José; Jagtenberg, Joycelyn C.; Haisma, Hidde J.; Storm, Gert

    2003-01-01

    Purpose. Immunoenzymosomes are tumor-targeted immunoliposomes bearing enzymes on their surface. These enzymes are capable of converting relatively nontoxic prodrugs into active cytostatic agents. The aims of this study were to compare the enzyme delivery capability of immunoenzymosomes with that of

  20. Liposome-mediated targeting of enzymes to cancer cells for site-specific activation of prodrugs : Comparison with the corresponding antibody-enzyme conjugate

    NARCIS (Netherlands)

    Fonseca, Maria José; Jagtenberg, Joycelyn C.; Haisma, Hidde J.; Storm, Gert

    Purpose. Immunoenzymosomes are tumor-targeted immunoliposomes bearing enzymes on their surface. These enzymes are capable of converting relatively nontoxic prodrugs into active cytostatic agents. The aims of this study were to compare the enzyme delivery capability of immunoenzymosomes with that of

  1. Saline soil enzyme activities of four plant communities in Sangong River basin of Xinjiang, China

    Institute of Scientific and Technical Information of China (English)

    ZhengJun GUAN; Qian LUO; Xi CHEN; XianWei FENG; ZhiXi TANG; Wei WEI; YuanRun ZHENG

    2014-01-01

    Soil enzyme activity plays an important role in the conversion of soil organic carbon into inorganic carbon, which is significant for the global carbon cycle. In this study, we investigated the soil enzyme activities of two ligninolytic enzymes (peroxidase and polyphenol oxidase) and five non-ligninolytic enzymes (α-1,4-glucosidase (AG); β-1,4-gluco-sidase (BG); N-acetyl-β-glucosaminidase (NAG); β-D-cellobiosidase (CBH); and β-xylosidase (BXYL)) in four plant communities of the Sangong River basin in Fukang, North Xinjiang, China. The four typical plant communities were dominated by Haloxylon ammodendron, Reaumuria soongonica, Salsola passerina, and Tamarix rarmosissima, respec-tively, with saline soils of varied alkalinity. The results showed that the soil peroxidase activity decreased seasonally. The activities of the five non-ligninolytic enzymes decreased with increasing soil depths, while those of the two ligninolytic enzymes did not show such a trend. In the four plant communities, BG had the highest activity among the five non-ligninolytic enzymes, and the activities of the two ligninolytic enzymes were higher than those of the four non-ligninolytic ones (AG, NAG, CBH, and BXYL). The community of H. ammodendron displayed the highest activity with respect to the two ligninolytic enzymes in most cases, but no significant differences were found among the four plant communities. The geometric mean of soil enzyme activities of the four plant communities was validated through an inde-pendently performed principal component analysis (PCA), which indicated that different plant communities had different soil enzyme activities. The correlation analysis showed that soil polyphenol oxidase activity was significantly positively correlated with the activities of the five non-ligninolytic enzymes. The soil pH value was positively correlated with the ac-tivities of al soil enzymes except peroxidase. Soil microbial carbon content also showed a significant positive

  2. Clinical evaluation of a new enzyme immunoassay for hepatitis B virus core-related antigen; a marker distinct from viral DNA for monitoring lamivudine treatment.

    Science.gov (United States)

    Rokuhara, A; Tanaka, E; Matsumoto, A; Kimura, T; Yamaura, T; Orii, K; Sun, X; Yagi, S; Maki, N; Kiyosawa, K

    2003-07-01

    We aimed to assess the clinical performance of a newly developed chemiluminescence enzyme immunoassay (CLEIA) for the detection of hepatitis B virus (HBV) core-related antigen (HBcrAg) in patients with chronic HBV infection. A total of 82 patients with chronic HBV infection and 167 HBV-negative controls were studied. HBcrAg was measured by CLEIA with monoclonal antibodies to hepatitis B e antigen (HBeAg) and hepatitis B core antigen (HBcAg), and HBV DNA was measured by transcription-mediated amplification assay (TMA) and in-house real-time detection polymerase chain reaction (RTD-PCR). The HBcrAg assay detected viremia in 189 of 216 samples (88%) collected from 72 patients whilst the TMA assay detected viremia in 178 of the 216 samples (82%) (P = 0.019). The HBcrAg concentration correlated linearly with the HBV DNA concentration (P HBcrAg assay was not affected by the absence of hepatitis B e antigen from the serum or the presence of precore mutations in the HBV genome. In patients without anti-viral drugs, changes in their serum HBcrAg concentration over time corresponded to their HBV DNA concentration. In six additional patients who were later treated with lamivudine, HBV DNA concentration declined more rapidly than their HBcrAg concentration. Three months after treatment commenced, the ratio of HBcrAg: HBV DNA had increased in all six patients (P = 0.031). The HBcrAg assay is a sensitive and useful test for the assessment of a patient's HBV load. When monitoring the anti-viral effect of lamivudine, HBcrAg provides a viral marker which is independent of HBV DNA.

  3. Production of Uricase Enzyme from Aspergillus niger and Determination of Some Factors Affecting the Activity

    OpenAIRE

    ERTAN, Figen

    1999-01-01

    This study was undertaken to obtain uricase enzyme from Aspergillus niger and also to determine the production conditions and the effects of some factors on enzyme activity. The results of the experiments, in consideration of the production of the uricase enzyme and the factors affecting the production, demonstrated that enzyme activity was maximum when the production time was 3 days at a temperature of 30ºC with the initial pH at 6.0 and inductor concentration at 0.1%. It was determined th...

  4. Spatial distribution of enzyme activities along the root and in the rhizosphere of different plants

    Science.gov (United States)

    Razavi, Bahar S.; Zarebanadkouki, Mohsen; Blagodatskaya, Evgenia; Kuzyakov, Yakov

    2015-04-01

    Extracellular enzymes are important for decomposition of many biological macromolecules abundant in soil such as cellulose, hemicelluloses and proteins. Activities of enzymes produced by both plant roots and microbes are the primary biological drivers of organic matter decomposition and nutrient cycling. So far acquisition of in situ data about local activity of different enzymes in soil has been challenged. That is why there is an urgent need in spatially explicit methods such as 2-D zymography to determine the variation of enzymes along the roots in different plants. Here, we developed further the zymography technique in order to quantitatively visualize the enzyme activities (Spohn and Kuzyakov, 2013), with a better spatial resolution We grew Maize (Zea mays L.) and Lentil (Lens culinaris) in rhizoboxes under optimum conditions for 21 days to study spatial distribution of enzyme activity in soil and along roots. We visualized the 2D distribution of the activity of three enzymes:β-glucosidase, leucine amino peptidase and phosphatase, using fluorogenically labelled substrates. Spatial resolution of fluorescent images was improved by direct application of a substrate saturated membrane to the soil-root system. The newly-developed direct zymography shows different pattern of spatial distribution of enzyme activity along roots and soil of different plants. We observed a uniform distribution of enzyme activities along the root system of Lentil. However, root system of Maize demonstrated inhomogeneity of enzyme activities. The apical part of an individual root (root tip) in maize showed the highest activity. The activity of all enzymes was the highest at vicinity of the roots and it decreased towards the bulk soil. Spatial patterns of enzyme activities as a function of distance from the root surface were enzyme specific, with highest extension for phosphatase. We conclude that improved zymography is promising in situ technique to analyze, visualize and quantify

  5. A new versatile microarray-based method for high throughput screening of carbohydrate-active enzymes.

    Science.gov (United States)

    Vidal-Melgosa, Silvia; Pedersen, Henriette L; Schückel, Julia; Arnal, Grégory; Dumon, Claire; Amby, Daniel B; Monrad, Rune Nygaard; Westereng, Bjørge; Willats, William G T

    2015-04-03

    Carbohydrate-active enzymes have multiple biological roles and industrial applications. Advances in genome and transcriptome sequencing together with associated bioinformatics tools have identified vast numbers of putative carbohydrate-degrading and -modifying enzymes including glycoside hydrolases and lytic polysaccharide monooxygenases. However, there is a paucity of methods for rapidly screening the activities of these enzymes. By combining the multiplexing capacity of carbohydrate microarrays with the specificity of molecular probes, we have developed a sensitive, high throughput, and versatile semiquantitative enzyme screening technique that requires low amounts of enzyme and substrate. The method can be used to assess the activities of single enzymes, enzyme mixtures, and crude culture broths against single substrates, substrate mixtures, and biomass samples. Moreover, we show that the technique can be used to analyze both endo-acting and exo-acting glycoside hydrolases, polysaccharide lyases, carbohydrate esterases, and lytic polysaccharide monooxygenases. We demonstrate the potential of the technique by identifying the substrate specificities of purified uncharacterized enzymes and by screening enzyme activities from fungal culture broths.

  6. Pectic enzymes

    NARCIS (Netherlands)

    Benen, J.A.E.; Voragen, A.G.J.; Visser, J.

    2003-01-01

    The pectic enzymes comprise a diverse group of enzymes. They consist of main-chain depolymerases and esterases active on methyl- and acetylesters of galacturonosyl uronic acid residues. The depolymerizing enzymes comprise hydrolases as wel as lyases

  7. Pectic enzymes

    NARCIS (Netherlands)

    Benen, J.A.E.; Voragen, A.G.J.; Visser, J.

    2003-01-01

    The pectic enzymes comprise a diverse group of enzymes. They consist of main-chain depolymerases and esterases active on methyl- and acetylesters of galacturonosyl uronic acid residues. The depolymerizing enzymes comprise hydrolases as wel as lyases

  8. Activity, life time and effect of hydrolytic enzymes for enhanced biogas production from sludge anaerobic digestion.

    Science.gov (United States)

    Odnell, Anna; Recktenwald, Michael; Stensén, Katarina; Jonsson, Bengt-Harald; Karlsson, Martin

    2016-10-15

    As an alternative to energy intensive physical methods, enzymatic treatment of sludge produced at wastewater treatment plants for increased hydrolysis and biogas production was investigated. Several hydrolytic enzymes were assessed with a focus on how enzyme activity and life time was influenced by sludge environments. It could be concluded that the activity life time of added enzymes was limited (biogas in situ experiments, subtilisin at a 1% mixture on basis of volatile solids, was the only enzyme providing a significantly increased biomethane production of 37%. However, even at this high concentration, subtilisin could not hydrolyze all available substrate within the life time of the enzyme. Thus, for large scale implementation, enzymes better suited to the sludge environments are needed. Copyright © 2016 Elsevier Ltd. All rights reserved.

  9. Replacement of Retinyl Esters by Polyunsaturated Triacylglycerol Species in Lipid Droplets of Hepatic Stellate Cells during Activation

    NARCIS (Netherlands)

    Testerink, N.; Ajat, M.A.; Houweling, M.; Brouwers, J.F.; Pully, V.V.; Manen, van H.J.; Otto, C.; Helms, J.B.; Vaandrager, A.B.

    2012-01-01

    Activation of hepatic stellate cells has been recognized as one of the first steps in liver injury and repair. During activation, hepatic stellate cells transform into myofibroblasts with concomitant loss of their lipid droplets (LDs) and production of excessive extracellular matrix. Here we aimed t

  10. Lipid accumulation in hepatocytes induces fibrogenic activation of hepatic stellate cells

    Institute of Scientific and Technical Information of China (English)

    Hella Wobser; Christoph Dorn; Thomas S Weiss; Thomas Amann; Cornelius Bollheimer; Roland Büttner; Jürgen Sc(o)lmerich; Claus Hellerbrand

    2009-01-01

    Despite the initial belief that non-alcoholic fatty liver disease is a benign disorder, it is now recognized that fbrosis progression occurs in a significant number of patients. Furthermore, hepatic steatosis has been identified as a risk factor for the progression of hepatic fibrosis in a wide range of other liver diseases. Here, we established an in vitro model to study the effect of hepatic lipid accumulation on hepatic stellate cells (HSCs), the central mediators of liver fibrogenesis. Primary human hepatocytes were incubated with the saturated fatty acid palmitate to induce intracellular lipid accumulation. Subsequently, human HSCs were incubated with conditioned media (CM) from steatotic or control hepatocytes. Lipid accumulation in hepatocytes induced the release of factors that accelerated the activation and proliferation of HSC, and enhanced their resistance to apoptosis, largely mediated via activation of the PI-3-kinase pathway. Furthermore, CM from steatotic hepatocytes induced the expression of the profibrogenic genes TGF-β, tissue inhibitor of metallo-proteinase-1 (TIMP-1), TIMP-2 and matrix-metallo-proteinase-2, as well as nuclear-factor Κb-dependent MCP-1 expression in HSC. In summary, our in vitro data indicate a potential mechanism for the pathophysiological link between hepatic steatosis and fibrogenesis in vivo. Herewith, this study provides an attractive in vitro model to study the molecular mechanisms of steatosis-induced fibrogenesis, and to identify and test novel targets for antifibrotic therapies in fatty liver disease.

  11. Changes in the spectrum and rates of extracellular enzyme activities in seawater following aggregate formation

    Directory of Open Access Journals (Sweden)

    K. Ziervogel

    2009-12-01

    Full Text Available Marine snow aggregates are heavily colonized by heterotrophic microorganisms that express high levels of hydrolytic activities, making aggregates hotspots for carbon remineralization in the ocean. To assess how aggregate formation influences the ability of seawater microbial communities to access organic carbon, we compared hydrolysis rates of six polysaccharides in coastal seawater after aggregates had been formed (via incubation on a roller table with hydrolysis rates in seawater from the same site that had not incubated on a roller table (referred to as whole seawater. Hydrolysis rates in the aggregates themselves were up to three orders of magnitude higher on a volume basis than in whole seawater. The enhancement of enzyme activity in aggregates relative to whole seawater differed by substrate, suggesting that the enhancement was under cellular control, rather than due to factors such as lysis or grazing. A comparison of hydrolysis rates in whole seawater with those in aggregate-free seawater, i.e. the fraction of water from the roller bottles that did not contain aggregates, demonstrated a nuanced microbial response to aggregate formation. Activities of laminarinase and xylanase enzymes in aggregate-free seawater were higher than in whole seawater, while activities of chondroitin, fucoidan, and arabinogalactan hydrolyzing enzymes were lower than in whole seawater. These data suggest that aggregate formation enhanced production of laminarinase and xylanase enzymes, and the enhancement also affected the surrounding seawater. Decreased activities of chondroitin, fucoidan, and arabinoglactan-hydrolyzing enzymes in aggregate-free seawater relative to whole seawater are likely due to shifts in enzyme production by the aggregate-associated community, coupled with the effects of enzyme degradation. Enhanced activities of laminarin- and xylan-hydrolyzing enzymes in aggregate-free seawater were due at least in part to cell-free enzymes. Measurements

  12. Endogenous n-3 polyunsaturated fatty acids attenuate T cell-mediated hepatitis via autophagy activation

    Directory of Open Access Journals (Sweden)

    Yanli Li

    2016-09-01

    Full Text Available Omega-3 polyunsaturated fatty acids (n-3 PUFAs exert anti-inflammatory effects in several liver disorders, including cirrhosis, acute liver failure, and fatty liver disease. To date, little is known about their role in immune-mediated liver diseases. In this study, we used fat-1 transgenic mice rich in endogenous n-3 PUFAs to examine the role of n-3 PUFAs in immune-mediated liver injury. Concanavalin A (Con A was administered intravenously to wild-type (WT and fat-1 transgenic mice to induce T cell-mediated hepatitis. Reduced liver damage was shown in Con A-administrated fat-1 transgenic mice, as evidenced by decreased mortality, attenuated hepatic necrosis, lessened serum alanine aminotransferase (ALT activity, and inhibited production of pro-inflammatory cytokines (e.g. TNF-α, IL-6, IL-17A and IFN-γ. In vivo and in vitro studies demonstrated that n-3 PUFAs significantly inhibited the activation of hepatic T cells and the differentiation of Th1 cells after Con A challenge. Further studies showed that n-3 PUFAs markedly increased autophagy level in Con A-treated fat-1 T cells compared with the WT counterparts. Blocking hepatic autophagy activity with chloroquine diminished the differences in T cell activation and liver injury between Con A-injected WT and fat-1 transgenic mice. We conclude that n-3 PUFAs limit Con A-induced hepatitis via an autophagy-dependent mechanism, and could be exploited as a new therapeutic approach for autoimmune hepatitis.

  13. Endogenous n-3 Polyunsaturated Fatty Acids Attenuate T Cell-Mediated Hepatitis via Autophagy Activation

    Science.gov (United States)

    Li, Yanli; Tang, Yuan; Wang, Shoujie; Zhou, Jing; Zhou, Jia; Lu, Xiao; Bai, Xiaochun; Wang, Xiang-Yang; Chen, Zhengliang; Zuo, Daming

    2016-01-01

    Omega-3 polyunsaturated fatty acids (n-3 PUFAs) exert anti-inflammatory effects in several liver disorders, including cirrhosis, acute liver failure, and fatty liver disease. To date, little is known about their role in immune-mediated liver diseases. In this study, we used fat-1 transgenic mice rich in endogenous n-3 PUFAs to examine the role of n-3 PUFAs in immune-mediated liver injury. Concanavalin A (Con A) was administered intravenously to wild-type (WT) and fat-1 transgenic mice to induce T cell-mediated hepatitis. Reduced liver damage was shown in Con A-administrated fat-1 transgenic mice, as evidenced by decreased mortality, attenuated hepatic necrosis, lessened serum alanine aminotransferase activity, and inhibited production of pro-inflammatory cytokines (e.g., TNF-α, IL-6, IL-17A, and IFN-γ). In vivo and in vitro studies demonstrated that n-3 PUFAs significantly inhibited the activation of hepatic T cells and the differentiation of Th1 cells after Con A challenge. Further studies showed that n-3 PUFAs markedly increased autophagy level in Con A-treated fat-1 T cells compared with the WT counterparts. Blocking hepatic autophagy activity with chloroquine diminished the differences in T cell activation and liver injury between Con A-injected WT and fat-1 transgenic mice. We conclude that n-3 PUFAs limit Con A-induced hepatitis via an autophagy-dependent mechanism and could be exploited as a new therapeutic approach for autoimmune hepatitis. PMID:27679638

  14. The vagal nerve stimulates activation of the hepatic progenitor cell compartment via muscarinic acetylcholine receptor type 3.

    Science.gov (United States)

    Cassiman, David; Libbrecht, Louis; Sinelli, Nicoletta; Desmet, Valeer; Denef, Carl; Roskams, Tania

    2002-08-01

    In the rat the hepatic branch of the nervus vagus stimulates proliferation of hepatocytes after partial hepatectomy and growth of bile duct epithelial cells after bile duct ligation. We studied the effect of hepatic vagotomy on the activation of the hepatic progenitor cell compartment in human and rat liver. The number of hepatic progenitor cells and atypical reactive ductular cells in transplanted (denervated) human livers with hepatitis was significantly lower than in innervated matched control livers and the number of oval cells in vagotomized rat livers with galactosamine hepatitis was significantly lower than in livers of sham-operated rats with galactosamine hepatitis. The expression of muscarinic acetylcholine receptors (M1-M5 receptor) was studied by immunohistochemistry and reverse transcriptase-polymerase chain reaction. In human liver, immunoreactivity for M3 receptor was observed in hepatic progenitor cells, atypical reactive ductules, intermediate hepatocyte-like cells, and bile duct epithelial cells. mRNA for the M1-M3 and the M5 receptor, but not the M4 receptor, was detected in human liver homogenates. In conclusion, the hepatic vagus branch stimulates activation of the hepatic progenitor cell compartment in diseased liver, most likely through binding of acetylcholine to the M3 receptor expressed on these cells. These findings may be of clinical importance for patients with a transplant liver.

  15. 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.)

  16. An efficient antiviral strategy for targeting hepatitis B virus genome using transcription activator-like effector nucleases.

    Science.gov (United States)

    Chen, Jieliang; Zhang, Wen; Lin, Junyu; Wang, Fan; Wu, Min; Chen, Cuncun; Zheng, Ye; Peng, Xiuhua; Li, Jianhua; Yuan, Zhenghong

    2014-02-01

    The hepatitis B virus (HBV) is a DNA virus that can cause chronic hepatitis B (CHB) in humans. Current therapies for CHB infection are limited in efficacy and do not target the pre-existing viral genomic DNA, which are present in the nucleus as a covalently closed circular DNA (cccDNA) form. The transcription activator-like (TAL) effector nucleases (TALENs) are newly developed enzymes that can cleave sequence-specific DNA targets. Here, TALENs targeting the conserved regions of the viral genomic DNA among different HBV genotypes were constructed. The expression of TALENs in Huh7 cells transfected with monomeric linear full-length HBV DNA significantly reduced the viral production of HBeAg, HBsAg, HBcAg, and pgRNA, resulted in a decreased cccDNA level and misrepaired cccDNAs without apparent cytotoxic effects. The anti-HBV effect of TALENs was further demonstrated in a hydrodynamic injection-based mouse model. In addition, an enhanced antiviral effect with combinations of TALENs and interferon-α (IFN-α) treatment was observed and expression of TALENs restored HBV suppressed IFN-stimulated response element-directed transcription. Taken together, these data indicate that TALENs can specifically target and successfully inactivate the HBV genome and are potently synergistic with IFN-α, thus providing a potential therapeutic strategy for treating CHB infection.

  17. Heating of vegetable oils influences the activity of enzymes participating in arachidonic acid formation in Wistar rats.

    Science.gov (United States)

    Stawarska, Agnieszka; Białek, Agnieszka; Tokarz, Andrzej

    2015-10-01

    Dietary intake of lipids and their fatty acids profile influence many aspects of health. Thermal processing changes the properties of edible oils and can also modify their metabolism, for example, eicosanoids formation. The aim of our study was to verify whether the activity of desaturases can be modified by lipids intake, especially by the fatty acids content. The experimental diets contained rapeseed oil, sunflower oil, and olive oil, both unheated and heated (for 10 minutes at 200 °C each time before administration), and influenced the fatty acids composition in serum and the activity of enzymes participating in arachidonic acid (AA) formation. The activity of desaturases was determined by measuring the amounts of AA formed in vitro derived from linoleic acid as determined in liver microsomes of Wistar rats. In addition, the indices of ∆(6)-desaturase (D6D) and ∆(5)-desaturase (D5D) have been determined. To realize this aim, the method of high-performance liquid chromatography has been used with ultraviolet-visible spectrophotometry detection. Diet supplementation with the oils rich in polyunsaturated fatty acids affects the fatty acids profile in blood serum and the activity of D6D and ∆(5)-desaturase in rat liver microsomes, the above activities being dependent on the kind of oil applied. Diet supplementation with heated oils has been found to increase the amount of AA produced in hepatic microsomes; and in the case of rapeseed oil and sunflower oil, it has also increased D6D activity.

  18. Enzyme Stability and Activity in Non-Aqueous Reaction Systems: A Mini Review

    Directory of Open Access Journals (Sweden)

    Shihui Wang

    2016-02-01

    Full Text Available Enormous interest in biocatalysis in non-aqueous phase has recently been triggered due to the merits of good enantioselectivity, reverse thermodynamic equilibrium, and no water-dependent side reactions. It has been demonstrated that enzyme has high activity and stability in non-aqueous media, and the variation of enzyme activity is attributed to its conformational modifications. This review comprehensively addresses the stability and activity of the intact enzymes in various non-aqueous systems, such as organic solvents, ionic liquids, sub-/super-critical fluids and their combined mixtures. It has been revealed that critical factors such as Log P, functional groups and the molecular structures of the solvents define the microenvironment surrounding the enzyme molecule and affect enzyme tertiary and secondary structure, influencing enzyme catalytic properties. Therefore, it is of high importance for biocatalysis in non-aqueous media to elucidate the links between the microenvironment surrounding enzyme surface and its stability and activity. In fact, a better understanding of the correlation between different non-aqueous environments and enzyme structure, stability and activity can contribute to identifying the most suitable reaction medium for a given biotransformation.

  19. Comparative azo reductase activity of red azo dyes through caecal and hepatic microsomal fraction in rats.

    Science.gov (United States)

    Singh, S; Das, M; Khanna, S K

    1997-09-01

    In order to study the rate of formation of toxic aromatic amines, anaerobic reduction of four red azo dyes viz. amaranth, carmoisine, fast Red E and ponceau 4R was investigated by incubating caecal content and hepatic microsomal fraction of rats with 37.5 microM concentration of dyes in sodium phosphate buffer pH 7.4 using NADPH generating system, glucose oxidase system and nitrogen as the gaseous phase. Caecal suspension exhibited higher azo reductase activity than that of hepatic microsomal fraction using any of the 4 azo dyes. Caecal microbes showed maximal azo reductase activity when ponceau 4R was used as a substrate followed by fast Red E and carmoisine, while with amaranth the activity was minimum. Similarly ponceau 4 R exhibited maximum hepatic microsomal azo reductase activity followed by fast Red E and carmoisine whereas, amaranth had minimum activity. Caecal flora possessed almost 17 fold higher degradative capability of ponceau 4 R and fast Red E colourants than the hepatic microsomal fraction. The higher reductive ability through caecal flora for ponceau 4R and fast Red E signifies the formation of more aromatic amines which may be re-absorbed through the intestine to be either eliminated through urine as conjugates or retained in the target tissues to elicit toxic effects.

  20. Macromolecular (pro)drugs with concurrent direct activity against the hepatitis C virus and inflammation.

    Science.gov (United States)

    Wohl, Benjamin M; Smith, Anton A A; Jensen, Bettina E B; Zelikin, Alexander N

    2014-12-28

    Macromolecular prodrugs (MPs) are a powerful tool to alleviate side-effects and improve the efficacy of the broad-spectrum antiviral agent ribavirin. In this work, we sought an understanding of what makes an optimal formulation within the macromolecular parameter space--nature of the polymer carrier, average molar mass, drug loading, or a good combination thereof. A panel of MPs based on biocompatible synthetic vinylic and (meth)acrylic polymers was tested in an anti-inflammatory assay with relevance to alleviating inflammation in the liver during hepatitis C infection. Pristine polymer carriers proved to have a pronounced anti-inflammatory activity, a notion which may prove significant in developing MPs for antiviral and anticancer treatments. With conjugated ribavirin, MPs revealed enhanced activity but also higher toxicity. Therapeutic windows and therapeutic indices were determined and discussed to reveal the most potent formulation and those with optimized safety. Polymers were also tested as inhibitors of replication of the hepatitis C viral RNA using a subgenomic viral replicon system. For the first time, negatively charged polymers are revealed to have an intracellular activity against hepatitis C virus replication. Concerted activity of the polymer and ribavirin afforded MPs which significantly increased the therapeutic index of ribavirin-based treatment. Taken together, the systematic investigation of the macromolecular space identified lead candidates with high efficacy and concurrent direct activity against the hepatitis C virus and inflammation.

  1. Activity-based protein profiling of the hepatitis C virus replication in Huh-7 hepatoma cells using a non-directed active site probe

    Directory of Open Access Journals (Sweden)

    McKay Craig S

    2010-02-01

    Full Text Available Abstract Background Hepatitis C virus (HCV poses a growing threat to global health as it often leads to serious liver diseases and is one of the primary causes for liver transplantation. Currently, no vaccines are available to prevent HCV infection and clinical treatments have limited success. Since HCV has a small proteome, it relies on many host cell proteins to complete its life cycle. In this study, we used a non-directed phenyl sulfonate ester probe (PS4≡ to selectively target a broad range of enzyme families that show differential activity during HCV replication in Huh-7 cells. Results The PS4≡ probe successfully targeted 19 active proteins in nine distinct protein families, some that were predominantly labeled in situ compared to the in vitro labeled cell homogenate. Nine proteins revealed altered activity levels during HCV replication. Some candidates identified, such as heat shock 70 kDa protein 8 (or HSP70 cognate, have been shown to influence viral release and abundance of cellular lipid droplets. Other differentially active PS4≡ targets, such as electron transfer flavoprotein alpha, protein disulfide isomerase A5, and nuclear distribution gene C homolog, constitute novel proteins that potentially mediate HCV propagation. Conclusions These findings demonstrate the practicality and versatility of non-directed activity-based protein profiling (ABPP to complement directed methods and accelerate the discovery of altered protein activities associated with pathological states such as HCV replication. Collectively, these results highlight the ability of in situ ABPP approaches to facilitate the identification of enzymes that are either predominantly or exclusively labeled in living cells. Several of these differentially active enzymes represent possible HCV-host interactions that could be targeted for diagnostic or therapeutic purposes.

  2. Altered hyperlipidemia, hepatic steatosis, and hepatic peroxisome proliferator-activated receptors in rats with intake of tart cherry.

    Science.gov (United States)

    Seymour, E Mitchell; Singer, Andrew A M; Kirakosyan, Ara; Urcuyo-Llanes, Daniel E; Kaufman, Peter B; Bolling, Steven F

    2008-06-01

    Elevated plasma lipids, glucose, insulin, and fatty liver are among components of metabolic syndrome, a phenotypic pattern that typically precedes the development of Type 2 diabetes. Animal studies show that intake of anthocyanins reduces hyperlipidemia, obesity, and atherosclerosis and that anthocyanin-rich extracts may exert these effects in association with altered activity of tissue peroxisome proliferator-activated receptors (PPARs). However, studies are lacking to test this correlation using physiologically relevant, whole food sources of anthocyanins. Tart cherries are a rich source of anthocyanins, and whole cherry fruit intake may also affect hyperlipidemia and/or affect tissue PPARs. This hypothesis was tested in the Dahl Salt-Sensitive rat having insulin resistance and hyperlipidemia. For 90 days, Dahl rats were pair-fed AIN-76a-based diets supplemented with either 1% (wt:wt) freeze-dried whole tart cherry or with 0.85% additional carbohydrate to match macronutrient and calorie provision. After 90 days, the cherry-enriched diet was associated with reduced fasting blood glucose, hyperlipidemia, hyperinsulinemia, and reduced fatty liver. The cherry diet was also associated with significantly enhanced hepatic PPAR-alpha mRNA, enhanced hepatic PPAR-alpha target acyl-coenzyme A oxidase mRNA and activity, and increased plasma antioxidant capacity. In conclusion, physiologically relevant tart cherry consumption reduced several phenotypic risk factors that are associated with risk for metabolic syndrome and Type 2 diabetes. Tart cherries may represent a whole food research model of the health effects of anthocyanin-rich foods and may possess nutraceutical value against risk factors for metabolic syndrome and its clinical sequelae.

  3. Differences in forage-acquisition and fungal enzyme activity contribute to niche segregation in Panamanian leaf-cutting ants

    DEFF Research Database (Denmark)

    Kooij, Pepijn Wilhelmus; Liberti, Joanito; Giampoudakis, Konstantinos;

    2014-01-01

    activities of twelve fungus garden decomposition enzymes, belonging to the amylases, cellulases, hemicellulases, pectinases and proteinases, and show that average enzyme activity per unit of fungal mass in Atta gardens is lower than in Acromyrmex gardens. Expression profiles of fungal enzymes in Atta also...... for decomposition enzymes....

  4. Inhibitory effects of prostaglandin E1 on activation of hepatic stellate cells in rabbits with schistosomiasis

    Institute of Scientific and Technical Information of China (English)

    Wei-Long Zou; Zhen Yang; Yun-Jin Zang; Dong-Jian Li; Zhi-Peng Liang; Zhong-Yang Shen

    2007-01-01

    BACKGROUND:Liver ifbrosis is the result of an imbalance between synthesis and degradation of extracellular matrix proteins of the liver. At the cellular and molecular levels, this progressive process is mainly characterized by activation of hepatic stellate cells (HSCs). Schistosoma japonica is one of the most prevalent causes of liver ifbrosis in China. It is characterized by hepatocyte damage, inlfammation, and chronic parasite egg-induced granuloma formation leading to ifbrosis. This study aimed to investigate the inhibitory effects of prostaglandin E1 (PGE1) on activation of HSCs and the alteration of type Ⅰ and Ⅲ collagen in rabbits with schistosomiasis. The study may promote the clinical application of praziquantel and PGE1 as a combined therapy to reverse hepatic ifbrosis caused by schistosomiasis. METHODS: Rabbits were percutaneously infected with cercaria of S. japonicum. Seven rabbits were subjected to intravenous injections of PGE1 (2.5 μg/kg daily) from days 60 to 120 after infection. The ultrastructural changes in activated HSCs were observed under transmission electron microscopy. The expression ofα-smooth muscle actin (α-SMA) was detected by immunohistochemistry. Fibril-forming collagens were detected by picrosirius staining. RESULTS: Activation of HSCs was a characteristic alteration in schistosome-induced hepatic ifbrosis. The expression of contraction-related α-SMA and the content of collagens were increased. Exogenous PGE1 markedly inhibited the activation of HSCs and reduced the expression of α-SMA around the hepatic sinusoids (P CONCLUSIONS:Activation of HSCs may play a key role in the progress of schistosome-induced hepatic ifbrosis. PGE1 effectively protects rabbit liver from ifbrosis, at least in part by inhibiting the activation of HSCs.

  5. Activated effects of parathyroid hormone-related protein on human hepatic stellate cells.

    Directory of Open Access Journals (Sweden)

    Fen-Fen Liang

    Full Text Available BACKGROUND & AIMS: After years of experiments and clinical studies, parathyroid hormone-related protein(PTHrP has been shown to be a bone formation promoter that elicits rapid effects with limited adverse reaction. Recently, PTHrP was reported to promote fibrosis in rat kidney in conjunction with transforming growth factor-beta1 (TGF-β1, which is also a fibrosis promoter in liver. However, the effect of PTHrP in liver has not been determined. In this study, the promoting actions of PTHrP were first investigated in human normal hepatic stellate cells (HSC and LX-2 cell lines. METHODS: TGF-β1, alpha-smooth muscle actin (α-SMA, matrix metalloproteinase 2 (MMP-2, and collagen I mRNA were quantified by real-time polymerase chain reaction (PCR after HSCs or LX-2 cells were treated with PTHrP(1-36 or TGF-β1. Protein levels were also assessed by western-blot analysis. Alpha-SMA were also detected by immunofluorescence, and TGF-β1 secretion was measured with enzyme-linked immunosorbent assay (ELISA of HSC cell culture media. RESULTS: In cultured human HSCs, mRNA and protein levels of α-SMA, collagen I, MMP-2, and TGF-β1 were increased by PTHrP treatment. A similar increasing pattern was also observed in LX-2 cells. Moreover, PTHrP significantly increased TGF-β1 secretion in cultured media from HSCs. CONCLUSIONS: PTHrP activated HSCs and promoted the fibrosis process in LX-2 cells. These procedures were probably mediated via TGF-β1, highlighting the potential effects of PTHrP in the liver.

  6. Activated effects of parathyroid hormone-related protein on human hepatic stellate cells.

    Science.gov (United States)

    Liang, Fen-Fen; Liu, Cui-Ping; Li, Li-Xuan; Xue, Min-Min; Xie, Fang; Guo, Yu; Bai, Lan

    2013-01-01

    After years of experiments and clinical studies, parathyroid hormone-related protein(PTHrP) has been shown to be a bone formation promoter that elicits rapid effects with limited adverse reaction. Recently, PTHrP was reported to promote fibrosis in rat kidney in conjunction with transforming growth factor-beta1 (TGF-β1), which is also a fibrosis promoter in liver. However, the effect of PTHrP in liver has not been determined. In this study, the promoting actions of PTHrP were first investigated in human normal hepatic stellate cells (HSC) and LX-2 cell lines. TGF-β1, alpha-smooth muscle actin (α-SMA), matrix metalloproteinase 2 (MMP-2), and collagen I mRNA were quantified by real-time polymerase chain reaction (PCR) after HSCs or LX-2 cells were treated with PTHrP(1-36) or TGF-β1. Protein levels were also assessed by western-blot analysis. Alpha-SMA were also detected by immunofluorescence, and TGF-β1 secretion was measured with enzyme-linked immunosorbent assay (ELISA) of HSC cell culture media. In cultured human HSCs, mRNA and protein levels of α-SMA, collagen I, MMP-2, and TGF-β1 were increased by PTHrP treatment. A similar increasing pattern was also observed in LX-2 cells. Moreover, PTHrP significantly increased TGF-β1 secretion in cultured media from HSCs. PTHrP activated HSCs and promoted the fibrosis process in LX-2 cells. These procedures were probably mediated via TGF-β1, highlighting the potential effects of PTHrP in the liver.

  7. Circadian pancreatic enzyme pattern and relationship between secretory and motor activity in fasting humans.

    Science.gov (United States)

    Keller, Jutta; Layer, Peter

    2002-08-01

    It is unknown whether nonparallel pancreatic enzyme output occurs under basal conditions in humans. We aimed to determine whether the circadian or wake-sleep cycle influences the relationship among pancreatic enzymes or between pancreatic secretory and jejunal motor activity. Using orojejunal multilumen intubation, we measured enzyme outputs and proximal jejunal motility index during consecutive daytime and nighttime periods in each of seven fasting, healthy volunteers. Enzyme outputs were correlated tightly during daytime phases of wakefulness and nighttime phases of sleep (r > 0.72, P activity was directly correlated with jejunal motility index (r > 0.50, P enzymes dominates throughout the circadian cycle. Nonparallel secretion during nocturnal phases of wakefulness may be due to merely circadian effects or to the coupling of the wake-sleep and the circadian cycle. The association between fluctuations of secretory and motor activity appears to be particularly tight during the night.

  8. [Relationship among soil enzyme activities, vegetation state, and soil chemical properties of coal cinder yard].

    Science.gov (United States)

    Wang, Youbao; Zhang, Li; Liu, Dengyi

    2003-01-01

    From field investigation and laboratory analysis, the relationships among soil enzyme activities, vegetation state and soil chemical properties of coal cinder yard in thermal power station were studied. The results showed that vegetation on coal cinder yard was distributed in scattered patch mainly with single species of plant, and herbs were the dominant species. At the same time, the activity of three soil enzymes had a stronger relativity to environment conditions, such as vegetation state and soil chemical properties. The sensitivity of three soil enzymes to environmental stress was in order of urease > sucrase > catalase. The relativity of three soil enzymes to environmental factor was in order of sucrase > urease > catalase. Because of urease being the most susceptible enzyme to environmental conditions, and it was marked or utmost marked interrelated with vegetation state and soil chemical properties, urease activity could be used as an indicator for the reclamation of wasteland.

  9. Inhibitory activity of Plantago major L. on angiotensin I-converting enzyme.

    Science.gov (United States)

    Nhiem, Nguyen Xuan; Tai, Bui Huu; Van Kiem, Phan; Van Minh, Chau; Cuong, Nguyen Xuan; Tung, Nguyen Huu; Thu, Vu Kim; Trung, Trinh Nam; Anh, Hoang Le Tuan; Jo, Sung-Hoon; Jang, Hae-Dong; Kwon, Young-In; Kim, Young Ho

    2011-03-01

    Eight compounds were isolated from methanol extract of Plantago major L. leaves and investigated for their ability to inhibit angiotensin I-converting enzyme activity. Among them, compound 1 showed the most potent inhibition with rate of 28.06 ± 0.21% at a concentration of 100 μM. Compounds 2 and 8 exhibited weak activities. These results suggest that compound 1 might contribute to the ability of P. major to inhibit the activity of angiotensin I- converting enzyme.

  10. Increased hepatic peroxisome proliferator-activated receptor coactivator-1α expression precedes the development of insulin resistance in offspring of rats from severe hyperglycemic mothers

    Institute of Scientific and Technical Information of China (English)

    MA Jing-mei; ZENG Chan-juan; ZHANG Li; SHOU Chong; YANG Hui-xia

    2012-01-01

    Background Prenatal hyperglycaemia may increase metabolic syndrome susceptibility of the offspring.An underlying component of the development of these morbidities is hepatic gluconeogenic molecular dysfunction.We hypothesized that maternal hyperglycaemia will influence her offsprings hepatic peroxisome proliferator-activated receptor coactivator-1α (PGC-1α) expression,a key regulator of glucose production in hepatocytes.@@Method We established maternal hyperglycaemia by streptozotocin injection to induce the maternal hyperglycaemic Wistar rat model.Offspring from the severe hyperglycemia group (SDO) and control group (CO) were monitored until 180 days after birth.Blood pressure,lipid metabolism indicators and insulin resistance (IR) were measured.Hepatic PGC-1α expression was analyzed by reverse transcription polymerase chain reaction and Western blotting.mRNA expression of two key enzymes in gluconeogenesis,glucose-6-phospha-tase (G-6-Pase) and phosphoenolpyruvate carboxykinase (PEPCK),were analyzed and compared.@@Results In the SDO group,PGC-1α expression at protein and mRNA levels were increased,so were expression of G-6-Pase and PEPCK (P<0.05).The above effects were seen prior to the onset of IR.@@Conclusion The hepatic gluconeogenic molecular dysfunction may contribute to the metabolic morbidities experienced by this population.

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

    Directory of Open Access Journals (Sweden)

    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

  12. Exo-inulinase of Aspergillus niger N402 : A hydrolytic enzyme with significant transfructosylating activity

    NARCIS (Netherlands)

    Goosen, C.; Van der Maarel, M. J. E. C.; Dijkhuizen, L.

    2008-01-01

    The purified exo-inulinase enzyme of Aspergillus niger N402 (AngInuE; heterologously expressed in Escherichia coli) displayed a sucrose:inulin (S/I) hydrolysis ratio of 2.3, characteristic for a typical exo-inulinase. The enzyme also had significant transfructosylating activity with increasing

  13. Mini Review: Basic Physiology and Factors Influencing Exogenous Enzymes Activity in the Porcine Gastrointestinal Tract

    DEFF Research Database (Denmark)

    Strube, Mikael Lenz; Meyer, Anne S.; Boye, Mette

    2013-01-01

    activity during intestinal transit are few, it is known that the enzymes, being protein molecules, can be negatively affected by the gastrointestinal proteolytic enzymes and the low pH in the stomach ventricle. In this review, the pH-values, endogenous proteases and other factors native to the digestive...

  14. Enzyme activity in terrestrial soil in relation to exploration of the Martian surface

    Science.gov (United States)

    Mclaren, A. D.

    1974-01-01

    Sensitive tests for the detection of extracellular enzyme activity in Martian soil was investigated using simulated Martian soil. Enzyme action at solid-liquid water interfaces and at low humidity were studied, and a kinetic scheme was devised and tested based on the growth of microorganisms and the oxidation of ammonium nitrite.

  15. Illustrating the Effect of pH on Enzyme Activity Using Gibbs Energy Profiles

    Science.gov (United States)

    Bearne, Stephen L.

    2014-01-01

    Gibbs energy profiles provide students with a visual representation of the energy changes that occur during enzyme catalysis, making such profiles useful as teaching and learning tools. Traditional kinetic topics, such as the effect of pH on enzyme activity, are often not discussed in terms of Gibbs energy profiles. Herein, the symbolism of Gibbs…

  16. Measuring potential denitrification enzyme activity rates using the membrane inlet mass spectrometer

    Science.gov (United States)

    The denitrification enzyme activity (DEA) assay, provides a quantitative assessment of the multi enzyme, biological process of reactive nitrogen removal via the reduction of N03 to N2. Measured in soil, usually under non limiting carbon and nitrate concentrations, this short ter...

  17. Crystallization of Hevamine, an Enzyme with Lysozyme/Chitinase Activity from Hevea brasiliensis Latex

    NARCIS (Netherlands)

    ROZEBOOM, HJ; BUDIANI, A; BEINTEMA, JJ

    1990-01-01

    Hevamine, an enzyme with both lysozyme and chitinase activity, was isolated and purified from Hevea brasiliensis (rubber tree) latex. The enzyme (molecular weight 29,000) is homologous to certain “pathogenesis-related” proteins from plants, but not to hen egg-white or phage T4 lysozyme. To

  18. Crystallization of Hevamine, an Enzyme with Lysozyme/Chitinase Activity from Hevea brasiliensis Latex

    NARCIS (Netherlands)

    ROZEBOOM, HJ; BUDIANI, A; BEINTEMA, JJ

    1990-01-01

    Hevamine, an enzyme with both lysozyme and chitinase activity, was isolated and purified from Hevea brasiliensis (rubber tree) latex. The enzyme (molecular weight 29,000) is homologous to certain “pathogenesis-related” proteins from plants, but not to hen egg-white or phage T4 lysozyme. To investiga

  19. Exo-inulinase of Aspergillus niger N402: A hydrolytic enzyme with significant transfructosylating activity

    NARCIS (Netherlands)

    Goosen, C.; Maarel, M.J. E.C. van der; Dijkhuizen, L.

    2008-01-01

    The purified exo-inulinase enzyme of Aspergillus niger N402 (AngInuE; heterologously expressed in Escherichia coli) displayed a sucrose:inulin (S/I) hydrolysis ratio of 2.3, characteristic for a typical exo-inulinase. The enzyme also had significant transfructosylating activity with increasing

  20. Illustrating the Effect of pH on Enzyme Activity Using Gibbs Energy Profiles

    Science.gov (United States)

    Bearne, Stephen L.

    2014-01-01

    Gibbs energy profiles provide students with a visual representation of the energy changes that occur during enzyme catalysis, making such profiles useful as teaching and learning tools. Traditional kinetic topics, such as the effect of pH on enzyme activity, are often not discussed in terms of Gibbs energy profiles. Herein, the symbolism of Gibbs…

  1. Exo-inulinase of Aspergillus niger N402: A hydrolytic enzyme with significant transfructosylating activity

    NARCIS (Netherlands)

    Goosen, C.; Maarel, M.J. E.C. van der; Dijkhuizen, L.

    2008-01-01

    The purified exo-inulinase enzyme of Aspergillus niger N402 (AngInuE; heterologously expressed in Escherichia coli) displayed a sucrose:inulin (S/I) hydrolysis ratio of 2.3, characteristic for a typical exo-inulinase. The enzyme also had significant transfructosylating activity with increasing sucro

  2. Homology to peptide pattern for annotation of carbohydrate-active enzymes and prediction of function

    DEFF Research Database (Denmark)

    Busk, Peter Kamp; Pilgaard, Bo; Lezyk, Mateusz Jakub

    2017-01-01

    Background: Carbohydrate-active enzymes are found in all organisms and participate in key biological processes.These enzymes are classified in 274 families in the CAZy database but the sequence diversity within each family makes it a major task to identify new family members and to provide basis ...

  3. Enhanced Wnt/ß-catenin and Notch signalling in the activated canine hepatic progenitor cell niche

    NARCIS (Netherlands)

    Schotanus, Baukje A|info:eu-repo/dai/nl/304836524; Kruitwagen, Hedwig S|info:eu-repo/dai/nl/34102046X; van den Ingh, Ted; van Wolferen, Monique E|info:eu-repo/dai/nl/119342561; Rothuizen, Jan|info:eu-repo/dai/nl/071276033; Penning, Louis C|info:eu-repo/dai/nl/110369181; Spee, Bart|info:eu-repo/dai/nl/304830925

    2014-01-01

    BackgroundThe liver has a large regenerative capacity. Hepatocytes can replicate and regenerate a diseased liver. However, as is the case in severe liver diseases, this replication may become insufficient or exhausted and hepatic progenitor cells (HPCs) can be activated in an attempt to restore

  4. Secretion and apparent activation of human hepatic lipase requires proper oligosaccharide processing in the endoplasmic reticulum

    NARCIS (Netherlands)

    A.J.M. Verhoeven (Adrie); B.P. Neve (Bernadette); H. Jansen (Hans)

    1999-01-01

    textabstractHuman hepatic lipase (HL) is a glycoprotein with four N-linked oligosaccharide side chains. The importance of glycosylation for the secretion of catalytically active HL was studied in HepG2 cells by using inhibitors of intracellular trafficking, N-glycosylat

  5. Melatonin suppresses activation of hepatic stellate cells through ROR alpha-mediated inhibition of 5-lipoxygenase

    NARCIS (Netherlands)

    Shajari, Shiva; Laliena, Almudena; Heegsma, Janette; Jesus Tunon, Maria; Moshage, Han; Faber, Klaas Nico

    2015-01-01

    Liver fibrosis is scar tissue resulting from an uncontrolled wound-healing process in response to chronic liver injury. Liver damage generates an inflammatory reaction that activates hepatic stellate cells (HSC) that transdifferentiate from quiescent cells that control retinol metabolism to prolifer

  6. An application of multilayer neural network on hepatitis disease diagnosis using approximations of sigmoid activation function

    Directory of Open Access Journals (Sweden)

    Onursal Çetin

    2015-06-01

    Full Text Available Objective: Implementation of multilayer neural network (MLNN with sigmoid activation function for the diagnosis of hepatitis disease. Methods: Artificial neural networks (ANNs are efficient tools currently in common use for medical diagnosis. In hardware based architectures activation functions play an important role in ANN behavior. Sigmoid function is the most frequently used activation function because of its smooth response. Thus, sigmoid function and its close approximations were implemented as activation function. The dataset is taken from the UCI machine learning database. Results: For the diagnosis of hepatitis disease, MLNN structure was implemented and Levenberg Morquardt (LM algorithm was used for learning. Our method of classifying hepatitis disease produced an accuracy of 91.9% to 93.8% via 10 fold cross validation. Conclusion: When compared to previous work that diagnosed hepatitis disease using artificial neural networks and the identical data set, our results are promising in order to reduce the size and cost of neural network based hardware. Thus, hardware based diagnosis systems can be developed effectively by using approximations of sigmoid function.

  7. [Effect of melatonin on antioxidant enzyme activities in blood erythrocytes of rats during acute emotional stress].

    Science.gov (United States)

    Pertsov, S S; Kalinichenko, L S; Koplik, E V; Nagler, L G; Alinkina, E S; Kozachenko, A I

    2015-01-01

    The effect of the epiphyseal hormone melatonin on the activity of antioxidant enzymes, glutathione peroxidase (GPx), glutathione reductase (GR), and Cu/Zn-superoxide dismutase (Cu/Zn-SOD) was studied in peripheral blood erythrocytes of behaviorally passive and active Wistar rats. Acute emotional stress was modeled by immobilization of animals for1 h with simultaneous electrocutaneous stimulation. Basal activity of antioxidant glutathione enzymes in erythrocytes of behaviorally passive rats was higher than that in active animals. Administration of melatonin (2 mg/kg, intraperitoneally) was accompanied by a decrease in the activity of GPx and GR in erythrocytes from non-stressed passive animals. After experimental stress, passive rats demonstrated a significant increase in the activity of Cu/Zn-SOD and GPx in peripheral blood erythrocytes. The absence of stress-induced changes in functional activity of antioxidant defense enzymes in the blood of behaviorally active animals suggests a relatively constant oxidative status of tissues in these animals under stress conditions. Melatonin administration had little effect on stress-induced changes in functional activity of the erythrocyte antioxidant system in passive rats. Active specimens pretreated with melatonin before stress exposure were characterized by activation of study antioxidant enzymes. Quantitative parameters of the erythrocyte antioxidant defense enzymes did not differ in behaviorally active and passive rats subjected to experimental stress after melatonin injection. Thus, exogenous melatonin abolishes differences in the activity of study antioxidant enzymes in erythrocytes of animals with different behavioral parameters under basal conditions and after experimental stress. In passive rats melatonin mainly reduced the initial tension of oxidative processes. By contrast, administration of this hormone to active specimens is followed by an increase in functional activity of the antioxidant enzyme system under

  8. Activity of an enzyme immobilized on superparamagnetic particles in a rotational magnetic field

    Energy Technology Data Exchange (ETDEWEB)

    Mizuki, Toru; Watanabe, Noriyuki; Nagaoka, Yutaka [Bio-Nano Electronics Research Centre, Toyo University, Saitama 350-8585 (Japan); Fukushima, Tadamasa [Shimadzu GLC Ltd., Phenomenex Support Centre, Tokyo 110-0016 (Japan); Morimoto, Hisao; Usami, Ron [Bio-Nano Electronics Research Centre, Toyo University, Saitama 350-8585 (Japan); Maekawa, Toru, E-mail: maekawa@toyonet.toyo.ac.jp [Bio-Nano Electronics Research Centre, Toyo University, Saitama 350-8585 (Japan)

    2010-03-19

    We immobilize {alpha}-amylase extracted from Bacillus Iicheniformis on the surfaces of superparamagnetic particles and investigate the effect of a rotational magnetic field on the enzyme's activity. We find that the activity of the enzyme molecules immobilized on superparamagnetic particles increases in the rotational magnetic field and reaches maximum at a certain frequency. We clarify the effect of the cluster structures formed by the superparamagnetic particles on the activity. Enzyme reactions are enhanced even in a tiny volume of solution using the present method, which is very important for the development of efficient micro reactors and micro total analysis systems ({mu}-TAS).

  9. Angiotensin converting enzyme in Alzheimer's disease increased activity in caudate nucleus and cortical areas.

    Science.gov (United States)

    Arregui, A; Perry, E K; Rossor, M; Tomlinson, B E

    1982-05-01

    The activity of the dipeptidyl carboxypeptidase, angiotensin converting enzyme, was assayed in several brain regions of patients dying with Alzheimer's disease and compared to that of appropriately age-matched controls. Enzyme activity was found to be elevated by 44% and 41% in the medial hippocampus and parahippocampal gyrus, respectively, and by 27% and 29% in the frontal cortex (area 10 of Brodman) and caudate nucleus, respectively, in Alzheimer's disease patients. Converting enzyme activity did not differ from controls in the nucleus accumbens, substantia nigra, temporal cortex, anterior or posterior hippocampus, amydgala, and septal nuclei.

  10. An appraisal of the enzyme stability-activity trade-off.

    Science.gov (United States)

    Miller, Scott R

    2017-07-01

    A longstanding id