DEVELOPMENTAL NEUROBIOLOGY, Issue 3 2003
Kiran K. Soma
Abstract In many species, territoriality is expressed only during the breeding season, when plasma testosterone (T) is elevated. In contrast, in song sparrows (Melospiza melodia morphna), males are highly territorial during the breeding (spring) and nonbreeding (autumn) seasons, but not during molt (late summer). In autumn, plasma sex steroids are basal, and castration has no effect on aggression. However, inhibition of aromatase reduces nonbreeding aggression, suggesting that neural steroid metabolism may regulate aggressive behavior. In wild male song sparrows, we examined the neural distribution of aromatase mRNA and seasonal changes in the activities of aromatase, 5,-, and 5,-reductase, enzymes that convert T to 17,-estradiol, 5,-dihydrotestosterone (5,-DHT, a potent androgen), or 5,-DHT (an inactive metabolite), respectively. Enzyme activities were measured in the diencephalon, ventromedial telencephalon (vmTEL, which includes avian amygdala), caudomedial neostriatum (NCM), and the hippocampus of birds captured during spring, molt, or autumn. Aromatase and 5,-reductase changed seasonally in a region-specific manner. Aromatase in the diencephalon was higher in spring than in molt and autumn, similar to seasonal changes in male sexual behavior. Aromatase activity in the vmTEL was high in both spring and autumn but significantly reduced at molt, similar to seasonal changes in aggression. 5,-Reductase was not elevated during molt, suggesting that low aggression during molt is not a result of increased inactivation of androgens. These data highlight the relevance of neural steroid metabolism to the expression of natural behaviors by free-living animals. © 2003 Wiley Periodicals, Inc. J Neurobiol 56: 209,221, 2003 [source]

The Role of Axial Ligation in Nitrate Reductase: A Model Study by DFT Calculations on the Mechanism of Nitrate Reduction

EUROPEAN JOURNAL OF INORGANIC CHEMISTRY, Issue 34 2008
Kuntal Pal
Abstract The reactivity differences of the model anionic complexes [Mo(mnt)2(X)(PPh3)], [mnt2, = 1,2-dicyanoethylenedithiolate; X = SPh (1a), SEt (1b), Cl (1c), Br (1b)] towards oxygen atom transfer from nitrate, which is a key step performed by nitrate reductase, has been investigated by density functional theory calculations. Unlike complexes 1a and 1b, complexes 1c and 1d do not react with nitrate. Thermodynamically, all these complexes have a similar ability to generate the pentacoordinate active state [Mo(mnt)2(X)], by dissociation of PPh3, although the inaccessibility of the dxy orbital in 1c,d and the instability of the corresponding nitrate-bound enzyme substrate (ES) type complex contributes to their failure to reduce nitrate. The nature of the ES complex for 1a,b is described. The variation in the experimental data due to the change of axial ligation from SPh to SEt on the catalytic pathway has also been addressed. The gas-phase and solvent-corrected potential energy surface for the reaction of 1a,b with nitrate are established with fully optimized minima and transition states.(© Wiley-VCH Verlag GmbH & Co. KGaA, 69451 Weinheim, Germany, 2008) [source]

Immune Response to a 26-kDa Protein, Alkyl Hydroperoxide Reductase, in Helicobacter pylori-Infected Mongolian Gerbil Model

HELICOBACTER, Issue 4 2001
Jing Yan
ABSTRACT Background. The host immune response is thought to play an important role in the outcome of Helico-bacter pylori infection. The successful development of the H. pylori -infected Mongolian gerbil model that mimics human disease has enabled study of the antibody response against H. pylori antigens. Materials and Methods. Serum samples from ulcer and carcinogenesis models of H. pylori -infected gerbils were used to screen for H. pylori antigens that cause a humoral immune response in the infected hosts. H. pylori alkyl hydroperoxide reductase (AhpC) is one such antigen on which we report here. The tsaA gene encoding AhpC was amplified by PCR from H. pylori ATCC 43504 strain, cloned into pMALTM -c2 expression vector and expressed in Escherichia coli. Maltose-binding protein fusion protein (MBP-AhpC) was purified by a MBP affinity column. Using purified recombinant AhpC protein as an antigen, the antibody response and changes of antibody levels against AhpC in the gerbil models were studied by Western blotting and ELISA. Results. Antibody against AhpC was negative in the early stages of infection, and became positive in the gerbils with the emergence of gastric diseases such as chronic active gastritis, gastric ulcer and gastric cancer. The antibody levels (ELISA) increased gradually over time and were higher in gerbils with gastric ulcer than that in gerbils without ulcers. Conclusions. Use of the gerbil model that mimics human H. pylori infection is likely to provide insights into the role of H. pylori -specific antigens possibly related to the subsequent development of gastric diseases. [source]

The Substrate Spectra of Pentaerythritol Tetranitrate Reductase, Morphinone Reductase, N -Ethylmaleimide Reductase and Estrogen-Binding Protein in the Asymmetric Bioreduction of Activated Alkenes

ADVANCED SYNTHESIS & CATALYSIS (PREVIOUSLY: JOURNAL FUER PRAKTISCHE CHEMIE), Issue 2-3 2010
Nicole
Abstract Four flavoproteins from the old yellow enzyme (OYE) family, pentaerythritol tetranitrate (PETNR) reductase, N -ethylmaleimide reductase (NEMR), morphinone reductase (MorR) and estrogen-binding protein (EBP1), exhibited a broad substrate tolerance by accepting conjugated enals, enones, imides, dicarboxylic acids and esters, as well as a nitroalkene and therefore can be employed for the asymmetric bioreduction of carbon-carbon double (CC) bonds. In particular, morphinone reductase and estrogen-binding protein often showed a complementary stereochemical preference in comparison to that of previously investigated OYEs. [source]

Asymmetric Reduction of Activated Alkenes by Pentaerythritol Tetranitrate Reductase: Specificity and Control of Stereochemical Outcome by Reaction Optimisation

ADVANCED SYNTHESIS & CATALYSIS (PREVIOUSLY: JOURNAL FUER PRAKTISCHE CHEMIE), Issue 17 2009
Anna Fryszkowska
Abstract We show that pentaerythritol tetranitrate reductase (PETNR), a member of the ,ene' reductase old yellow enzyme family, catalyses the asymmetric reduction of a variety of industrially relevant activated ,,,-unsaturated alkenes including enones, enals, maleimides and nitroalkenes. We have rationalised the broad substrate specificity and stereochemical outcome of these reductions by reference to molecular models of enzyme-substrate complexes based on the crystal complex of the PETNR with 2-cyclohexenone 4a. The optical purity of products is variable (49,99% ee), depending on the substrate type and nature of substituents. Generally, high enantioselectivity was observed for reaction products with stereogenic centres at C, (>99% ee). However, for the substrates existing in two isomeric forms (e.g., citral 11a or nitroalkenes 18,19a), an enantiodivergent course of the reduction of E/Z -forms may lead to lower enantiopurities of the products. We also demonstrate that the poor optical purity obtained for products with stereogenic centres at C, is due to non-enzymatic racemisation. In reactions with ketoisophorone 3a we show that product racemisation is prevented through reaction optimisation, specifically by shortening reaction time and through control of solution pH. We suggest this as a general strategy for improved recovery of optically pure products with other biocatalytic conversions where there is potential for product racemisation. [source]

Enantioselective Reduction of Diaryl Ketones Catalyzed by a Carbonyl Reductase from Sporobolomyces salmonicolor and its Mutant Enzymes

ADVANCED SYNTHESIS & CATALYSIS (PREVIOUSLY: JOURNAL FUER PRAKTISCHE CHEMIE), Issue 4 2009
Hongmei Li
Abstract The carbonyl reductase from red yeast Sporobolomyces salmonicolor AKU4429 (SSCR) and its mutant enzymes effectively catalyzed the enantioselective reduction of diaryl ketones to give the corresponding chiral alcohols. Both conversion and enantioselectivity were dependent on the co-solvent in the reaction medium. Diaryl ketones with a para -substituent on one of the phenyl groups were reduced with high enantioselectivity (up to 99% ee), which is difficult to achieve using chemical methods such as chiral borane reduction, asymmetric hydrogenation or hydrosilylation. Mutation of SSCR at Q245 resulted in a higher amount of (S)-enantiomer in the products, and in the case of mutant Q245P with para -substituted diaryl ketones as substrate, this effect was so remarkable that the reduction enantiopreference was switched from (R) to (S). The present study provides valuable information about the catalytic properties of the carbonyl reductase SSCR toward the reduction of diaryl ketones, serving as basis for further engineering of this enzyme to develop efficient biocatalysts for highly enantiospecific reduction of diaryl ketones without high electronic dissymmetry or an ortho -substituent on one of the aryl groups. [source]

X-ray crystallographic analysis of the complexes of enoyl acyl carrier protein reductase of Plasmodium falciparum with triclosan variants to elucidate the importance of different functional groups in enzyme inhibition

IUBMB LIFE, Issue 6 2010
Koustav Maity
Abstract Triclosan, a well-known inhibitor of Enoyl Acyl Carrier Protein Reductase (ENR) from several pathogenic organisms, is a promising lead compound to design effective drugs. We have solved the X-ray crystal structures of Plasmodium falciparum ENR in complex with triclosan variants having different substituted and unsubstituted groups at different key functional locations. The structures revealed that 4 and 2, substituted compounds have more interactions with the protein, cofactor, and solvents when compared with triclosan. New water molecules were found to interact with some of these inhibitors. Substitution at the 2, position of triclosan caused the relocation of a conserved water molecule, leading to an additional hydrogen bond with the inhibitor. This observation can help in conserved water-based inhibitor design. 2, and 4, unsubstituted compounds showed a movement away from the hydrophobic pocket to compensate for the interactions made by the halogen groups of triclosan. This compound also makes additional interactions with the protein and cofactor which compensate for the lost interactions due to the unsubstitution at 2, and 4,. In cell culture, this inhibitor shows less potency, which indicates that the chlorines at 2, and 4, positions increase the ability of the inhibitor to cross multilayered membranes. This knowledge helps us to modify the different functional groups of triclosan to get more potent inhibitors. © 2010 IUBMB IUBMB Life, 467,476, 2010 [source]

Ribonucleotide activation by enzyme ribonucleotide reductase: Understanding the role of the enzyme

JOURNAL OF COMPUTATIONAL CHEMISTRY, Issue 16 2004
Nuno M. F. S. A. Cerqueira
Abstract This article focuses on the first step of the catalytic mechanism for the reduction of ribonucleotides catalyzed by the enzyme Ribonucleotide Reductase (RNR). This corresponds to the activation of the substrate. In this work a large model of the active site region involving 130 atoms was used instead of the minimal gas phase models used in previous works. The ONIOM method was employed to deal with such a large system. The results gave additional information, which previous small models could not provide, allowing a much clearer evaluation of the role of the enzyme in this step. Enzyme,substrate interaction energies, specific transition state stabilization, and substrate steric strain energies were obtained. It was concluded that the transition state is stabilized in 4.0 kcal/mol by specific enzyme,substrate interactions. However, this stabilization is cancelled by the cost in conformational energy for the enzyme to adopt the transition state geometry; the overall result is that the enzyme machinery does not lead to a rate enhancement in this step. It was also found that the substrate binds to the active site with almost no steric strain, emphasizing the complementarity and specificity of the RNR active site for nucleotide binding. The main role of the enzyme at the very beginning of the catalytic cycle was concluded to be to impose stereospecifity upon substrate activation and to protect the enzyme radical from the solvent, rather than to be an reaction rate enhancement. © 2004 Wiley Periodicals, Inc. J Comput Chem 25: 2031,2037, 2004 [source]

Theoretical study of ribonucleotide reductase mechanism-based inhibition by 2,-azido-2,-deoxyribonucleoside 5,-diphosphates

JOURNAL OF COMPUTATIONAL CHEMISTRY, Issue 2 2004
Susana Pereira
Abstract 2,-Azido-2,-deoxyribonucleoside 5,-diphosphates are mechanism-based inhibitors of Ribonucleotide Reductase. Considerable effort has been made to elucidate their mechanism of inhibition, which is still controversial and not fully understood. Previous studies have detected the formation of a radical intermediate when the inhibitors interact with the enzyme, and several authors have proposed possible structures for this radical. We have conducted a theoretical study of the possible reactions involved, which allowed us to identify the structure of the new radical among the several proposals. A new reactional path is also proposed that is the most kinetically favored to yield this radical and ultimately inactivate the enzyme. The energetic involved in this mechanism, both for radical formation and radical decay, as well as the calculated Hyperfine Coupling Constants for the radical intermediate, are in agreement with the correspondent experimental values. This mechanistic alternative is fully coherent with remaining experimental data. © 2003 Wiley Periodicals, Inc. J Comput Chem 25: 227,237, 2004 [source]

Aldose Reductase and AGE,RAGE pathways: central roles in the pathogenesis of vascular dysfunction in aging rats

AGING CELL, Issue 5 2010
Kellie McCormick Hallam
Summary Aging is inevitably accompanied by gradual and irreversible innate endothelial dysfunction. In this study, we tested the hypothesis that accentuation of glucose metabolism via the aldose reductase (AR) pathway contributes to age-related vascular dysfunction. AR protein and activity levels were significantly increased in aged vs. young aortic homogenates from Fischer 344 rats. Immunostaining revealed that the principal site of increased AR protein was the aortic endothelium as well as smooth muscle cells. Studies revealed that endothelial-dependent relaxation (EDR) in response to acetylcholine was impaired in aged rats compared to young rats and that treatment with the AR inhibitor (ARI) zopolrestat significantly improved EDR in aged rats. Methylglyoxal (MG), a key precursor of advanced glycation endproducts (AGEs), was significantly increased in the aortas of aged rats vs. young rats. Consistent with central roles for AR in generation of MG in aging, ARI treatment significantly reduced MG levels in aged rat aorta to those in young rats. Treatment of aged rats with soluble(s) RAGE, a soluble form of the chief signal transduction receptor for AGEs, RAGE, significantly improved EDR in aged rats, thus establishing the contribution of age-related increases in AGEs to endothelial dysfunction. These findings reveal that significant increases in AR expression and activity in aged rat vasculature linked to endothelial dysfunction may be mitigated, at least in part, via ARI and that aging-linked increased flux via AR generates AGEs; species which transduce endothelial injury consequent to their interaction with RAGE. These data demonstrate for the first time that AR mediates aging-related vascular dysfunction, at least in part, via RAGE. [source]

Localization of Sepiapterin Reductase in Pigment Cells of Oryzias latipes

PIGMENT CELL & MELANOMA RESEARCH, Issue 5 2003
Sumiko Negishi
Body colors of poikilothermal vertebrates are derived from three distinct types of pigment cells, melanophores, erythro/xanthophores and irido/leucophores. It is well known that melanin in melanophores is synthesized by tyrosinase within a specific organelle termed the melanosome. Although sepiapterin reductase (SPR) is an important enzyme involved in metabolizing biopterin and sepiapterin (a conspicuous pteridine as a coloring pigment in xanthophores) the distribution of SPR has not been shown in pigment cells. An antibody raised in rabbits against rat SPR was used to demonstrate the presence of SPR in pigment cells of Oryzias latipes. This study, which used immunohistochemistry with fluorescence or peroxidase/diaminobenzidine as markers, revealed that SPR could be detected readily in xanthophores, but only faintly in melanophores. These results suggest that sepiapterin is metabolized within xanthophores. Moreover, these experiments show that a protein sharing immunological cross-reactivity with rat SPR is located in teleost O. latipes xanthophores, which is significant considering the relationship of pteridine metabolism between poikilothermal vertebrates and mammals. Further progress in investigations of the roles of pteridines in vertebrates will be promoted by using these fish which can be bred in mass rather easily in the laboratory. [source]

Activity of NADPH-Cytochrome P-450 Reductase of the Human Heart, Liver and Lungs in the Presence of (-)-Epigallocatechin Gallate, Quercetin and Resveratrol: An in vitro Study

BASIC AND CLINICAL PHARMACOLOGY & TOXICOLOGY, Issue 2 2005
Jaroslaw Dudka
The enzyme is also involved in the toxicity of some clinically important antitumour drugs (doxorubicin) and pesticides (paraquat). P-450 reductase activates them to their more toxic metabolites via one electron reduction which triggers free radical cascade. In some cases however, such transformation is essential to produce therapeutic effect in anticancer drugs. The main purpose of the paper was to evaluate the effect of three natural compounds found in human diet: (-)-epigallocatechin gallate (EGCG), quercetin and resveratrol on P-450 reductase activity. The activity of the enzyme was determined spectrophotometrically by measurement of the rate of cytochrome c reduction at 550 nm, in vitro, using human heart, liver and lung microsomes. It was found that quercetin increased the P-450 reductase activity in human organs at all tested doses. The activity of microcosms in all organs was enhanced according to the concentrations of quercetin, which increased the activity in the order lung>heart>liver. Addition of EGCG to the reaction mixture enhanced the P-450 reductase activity in the following order: liver>heart>lung. However, no significant effect of resveratrol on P-450 reductase activity was observed. It seems that the presence of quercetin and EGCG in the diet may increase P-450 reductase activity during doxorubicin therapy with subsequent increased risk of toxicity. A beneficial effect may be obtained in anticancer therapy with bioreductive agents like tirapazamine. [source]

Design of a Functional Nitric Oxide Reductase within a Myoglobin Scaffold

CHEMBIOCHEM, Issue 8 2010
Valentin Köhler Dr.
One site fits all: Yi Lu and co-workers have reported the conversion of sperm-whale myoglobin into a functional nitric oxide reductase. For this purpose, they designed a second metal binding site in the wild-type holo-protein and demonstrated NO reduction with the structurally characterized model, making thereby a significant contribution to the rapidly developing field of artificial metalloenzymes. [source]

The Role of Arginine 28 in Catalysis by Dihydrofolate Reductase from the Hyperthermophile Thermotoga maritima

CHEMBIOCHEM, Issue 16 2009
E. Joel Loveridge Dr.
Get a grip: Dihydrofolate reductase from Thermotoga maritima (TmDHFR) is unusual in that it has an arginine residue within its active site (ringed residue). Here, we address the role of this residue in catalysis. We find no evidence that Arg28 compromises catalysis in TmDHFR by preventing protonation of the substrate or that it acts as an acid to protonate the substrate. Instead, it appears that this residue plays an important role in binding the substrate tightly to ensure its thermal stability. [source]

Enantioselective Enzymatic Reductions of Sterically Bulky Aryl Alkyl Ketones Catalyzed by a NADPH-Dependent Carbonyl Reductase.

CHEMINFORM, Issue 16 2007
Dunming Zhu
Abstract ChemInform is a weekly Abstracting Service, delivering concise information at a glance that was extracted from about 200 leading journals. To access a ChemInform Abstract, please click on HTML or PDF. [source]

Asymmetric Reduction of a Variety of Ketones with a Recombinant Carbonyl Reductase: Identification of the Gene Encoding a Versatile Biocatalyst.

CHEMINFORM, Issue 31 2005
Tadashi Ema
No abstract is available for this article. [source]

High Throughput Screening Identifies Novel Inhibitors of Escherichia coli Dihydrofolate Reductase that Are Competitive with Dihydrofolate.

CHEMINFORM, Issue 43 2003
Michela Zolli-Juran
Abstract For Abstract see ChemInform Abstract in Full Text. [source]

Cyclohex-1-ene Carboxylic Acids: Synthesis and Biological Evaluation of Novel Inhibitors of Human 5, Reductase.

CHEMINFORM, Issue 28 2003
Eckhard Baston
Abstract For Abstract see ChemInform Abstract in Full Text. [source]

ChemInform Abstract: 4-(Benzoylindolizinyl)butyric Acids; Novel Nonsteroidal Inhibitors of Steroid 5,-Reductase.

CHEMINFORM, Issue 48 2001
Part 3.
Abstract ChemInform is a weekly Abstracting Service, delivering concise information at a glance that was extracted from about 100 leading journals. To access a ChemInform Abstract of an article which was published elsewhere, please select a "Full Text" option. The original article is trackable via the "References" option. [source]

ChemInform Abstract: 1,4-Disubstituted Imidazoles Are Potential Antibacterial Agents Functioning as Inhibitors of Enoyl Acyl Carrier Protein Reductase (FabI).

CHEMINFORM, Issue 45 2001
Dirk A. Heerding
Abstract ChemInform is a weekly Abstracting Service, delivering concise information at a glance that was extracted from about 100 leading journals. To access a ChemInform Abstract of an article which was published elsewhere, please select a "Full Text" option. The original article is trackable via the "References" option. [source]

Synthesis and Evaluation of 1-(1-(Benzo[b]thiophen-2-yl)cyclohexyl)piperidine (BTCP) Analogues as Inhibitors of Trypanothione Reductase

CHEMMEDCHEM, Issue 8 2009
Stephen Patterson Dr.
Abstract Thirty two analogues of phencyclidine were synthesised and tested as inhibitors of trypanothione reductase (TryR), a potential drug target in trypanosome and leishmania parasites. The lead compound BTCP (1, 1-(1-benzo[b]thiophen-2-yl-cyclohexyl) piperidine) was found to be a competitive inhibitor of the enzyme (Ki=1,,M) and biologically active against bloodstream T.,brucei (EC50=10,,M), but with poor selectivity against mammalian MRC5 cells (EC50=29,,M). Analogues with improved enzymatic and biological activity were obtained. The structure,activity relationships of this novel series are discussed. [source]

Analysis of proliferating cell fraction determined by monoclonal antibody to M1-subunit ribonucleotide reductase and Ki-67 in relation to p53 protein expression in fine-needle aspirates from non-Hodgkin's lymphomas

CYTOPATHOLOGY, Issue 5 2000
V. Sviatoha
Analysis of proliferating cell fraction determined by monoclonal antibody to M1-subunit ribonucleotide reductase and Ki-67 in relation to p53 protein expression in fine-needle aspirates from non Hodgkin's lymphomas The purpose of this study was to analyse the proliferative fraction with the monoclonal antibody M1-R-R to M1-subunit ribonucleotide reductase and with MIB-1 to Ki-67 antigen in relation to p53 protein expression in fine needle aspirates from B-cell non-Hodgkin's lymphomas. One hundred and thirty-seven cases, previously diagnosed and sub-typed according to the Kiel classification and characterized by immunophenotyping, were included in the study. The M-1 subunit ribonucleotide reductase (M1 -R-R), Ki-67 and p53 antigens were detected using monoclonal antibodies on stored cytospin preparations. There was a good correlation (r = 0.72) between Ki-67 and M1 -R-R positive cell fraction in both high and low grade lymphomas. High-grade lymphomas had a median percentage of M1 -R-R/MIB-1 positive cells of 53.0/73.0 for lymphoblastic, 61.0/52.0 for immunoblastic and 33.5/41.0 for centroblastic lymphomas, respectively. In low grade lymphomas figures of median percentage of M1 -R-R/MIB-1 were 9.0/15.0 for centroblastic/centrocytic, 11.0/9.5 for chronic lymphocytic leukaemia, 16.0/27.0 for centrocytic and 12.0/9.0 for immunocytomas, respectively. The median percentages of M1 -R-R/MIB-1 for high and low grade lymphomas were 37.0/50.5 and 11.0/12.0, respectively. In the p53 positive cases the proliferation rate as measured by staining for M1 -R-R and MIB-1 was higher than in p53 negative cases, but the difference was not statistically significant. The results show that cytospin material obtained by fine needle aspiration and stored at ,70 °C for years can be used reliably for both peroxidase-avidin-biotin and three-step alkaline phosphatase immunocytochemical staining. In addition, proliferation fraction determined by M1 -R-R monoclonal antibody staining correlates well with that measured by an established marker for cell proliferation, the Ki-67 antibody. However, the proliferation fraction as measured by the two antibodies differs in the various subtypes of non-Hodgkin's lymphoma which indicates that they may contribute different prognostic information. [source]

Reduced metabolites mediate neuroprotective effects of progesterone in the adult rat hippocampus.

DEVELOPMENTAL NEUROBIOLOGY, Issue 9 2006
The synthetic progestin medroxyprogesterone acetate (Provera) is not neuroprotective
Abstract The ovarian hormone progesterone is neuroprotective in different experimental models of neurodegeneration. In the nervous system, progesterone is metabolized to 5,-dihydroprogesterone (DHP) by the enzyme 5,-reductase. DHP is subsequently reduced to 3,,5,-tetrahydroprogesterone (THP) by a reversible reaction catalyzed by the enzyme 3,-hydroxysteroid dehydrogenase. In this study we have analyzed whether progesterone metabolism is involved in the neuroprotective effect of the hormone in the hilus of the hippocampus of ovariectomized rats injected with kainic acid, an experimental model of excitotoxic cell death. Progesterone increased the levels of DHP and THP in plasma and hippocampus and prevented kainic-acid-induced neuronal loss. In contrast to progesterone, the synthetic progestin medroxyprogesterone acetate (MPA, Provera) did not increase DHP and THP levels and did not prevent kainic-acid-induced neuronal loss. The administration of the 5,-reductase inhibitor finasteride prevented the increase in the levels of DHP and THP in plasma and hippocampus as a result of progesterone administration and abolished the neuroprotective effect of progesterone. Both DHP and THP were neuroprotective against kainic acid. However, the administration of indomethacin, a 3,-hydroxysteroid dehydrogenase inhibitor, blocked the neuroprotective effect of both DHP and THP, suggesting that both metabolites are necessary for the neuroprotective effect of progesterone. In conclusion, our findings indicate that progesterone is neuroprotective against kainic acid excitotoxicity in vivo while the synthetic progestin MPA is not and suggest that progesterone metabolism to its reduced derivatives DHP and THP is necessary for the neuroprotective effect of the hormone. © 2006 Wiley Periodicals, Inc. J Neurobiol, 2006 [source]

Brain aromatase, 5,-reductase, and 5,-reductase change seasonally in wild male song sparrows: Relationship to aggressive and sexual behavior

Cataracts in experimentally diabetic mouse: morphological and apoptotic changes

DIABETES OBESITY & METABOLISM, Issue 2 2005
K. R. Hegde
Aim:, The objective of these investigations was to extend our earlier study on the induction of cataracts in diabetic mice, a low aldose reductase (AR) animal model at morphological level. Previous studies were done primarily at biochemical level. Methods:, Diabetes was induced by intraperitoneal administration of streptozotocin. The lenses isolated after the establishment of diabetes were then subjected to histologic and electron microscopic studies. Results:, Morphological alterations were characterized by shrinkage, elongation and lobulization of the nuclei of the epithelial cells. This was associated with chromatin condensation and its margination. Similar structural aberrations were also observed in a significant number of the subepithelial fibre cells representing defect in fibre maturation. More interestingly, unlike that in other common animal models of diabetic cataract, such abnormally nucleated cells were also found to be prevalent in the posterior subcapsular region, a finding common in human diabetics also. Conclusion:, The present studies further affirm the suitability of the mouse model for a study of cataractogenesis induced by diabetes. Because of the findings reported herein, as well as the known biochemical similarity between the lenses of the mice and humans in respect of AR deficiency, contrary to the rat model where it is very high, use of this species is considered more useful towards understanding the basic aetiology as well as for evaluating the efficacy of various referred nutritional and metabolic antioxidants against such cataracts. [source]

The MTHFR C677T polymorphism confers a high risk for stroke in both homozygous and heterozygous T allele carriers with Type 2 diabetes

DIABETIC MEDICINE, Issue 5 2006
M. P. Hermans
Abstract Objective Individuals with Type 2 diabetes are at increased risk of stroke. Plasma homocysteine (tHcy) is an independent risk factor for cardiovascular (CV) disease. The methylene,tetrahydrofolate reductase (MTHFR) gene polymorphism (thermolabile variant C677T) is associated with CV risk, partly as a result of increased Hcy, especially in homozygous subjects. Aim To relate the occurrence of the MTHFR polymorphism with stroke prevalence by examining allelic frequency and genotype distribution in 165 subjects with Type 2 diabetes studied for the presence of thermolabile C677T MTHFR mutation. Results Mean age was 67.7 years, and tHcy 18.2 µmol/l. T allele frequency was 38.5%. MTHFR genotypes were: normal (CC) 40%; heterozygous (CT) 43%; homozygous (TT) 17%. Serum levels of folic acid and B12 vitamin were within normal limits. Stroke prevalence was 14%. Sixty-four per cent of stroke-free subjects had the normal C allele vs. 46% in stroke subjects. The frequencies of genotypes (CC-CT-TT) were (%): 44-41-15 in stroke-free vs. 17-57-26 in stroke patients. Coronary (CAD) and peripheral artery disease (PAD) were common in all groups, with no differences according to genotypes. Stroke prevalence was markedly higher in genotypes CT and TT (18 and 21%) compared with CC (6%). Mean tHcy levels were higher in TT subjects. Conclusion The allelic frequency of C677T MTHFR mutation in Type 2 diabetes subjects with stroke is markedly different from that of subjects without stroke. Genotypic characteristics suggest that C677T MTHFR mutation confers a higher risk for stroke to both homozygous and heterozygous T allele carriers that cannot be ascribed solely to raised tHcy and/or lower folate status in CT subjects, nor to phenotypic expression of conventional risk factors for stroke. The impact of the MTHFR polymorphism on stroke may result from T allele-linked deleterious effects, or C allele-linked protection. Confirmatory studies are warranted, as this cohort was not randomly selected, and a type 1 error cannot be ruled out. [source]

Changes in antioxidant defense status in response to cisplatin and 5-FU in esophageal carcinoma

DISEASES OF THE ESOPHAGUS, Issue 2 2008
T. Kaur
SUMMARY., The ability of reactive oxygen species to induce cellular damage and to cause cell death opens the possibility of exploiting this property in the treatment of esophageal cancer through a free radical mediated mechanism. The present study was carried out with the aim of evaluating the changes in the antioxidant defense status in esophageal cancer patients treated without and with neoadjuvant therapy (NAT). Forty surgically resected tissue specimens from tumors, tissue adjoining the tumors and paired macroscopically normal mucosa were obtained from esophageal cancer patients treated with or without chemo-radiotherapy. An evaluation of antioxidant defense system in the normal, adjoining and tumor esophageal tissues in response to NAT revealed decreased catalase activity in tumor and adjoining tissues as compared to their respective normal tissue levels. Similarly, decreased superoxide dismutase activity was observed in tumor tissue in response to NAT. In both the treatment groups (with and without NAT), no significant change was observed in the enzyme activity of glutathione reductase in the normal, adjoining and tumor tissues. Enhanced glutathione peroxidase activity was found in tumor tissue, as compared to the adjoining and paired normal tissue of patients after NAT. Estimation of reduced glutathione (GSH) levels showed a significant decline in GSH levels in esophageal tumors after NAT. Depletion of GSH, an endogenous antioxidant, would elevate drug sensitivity and might predispose neoplastic cells to apoptosis in response to NAT. The antioxidant enzymes in the esophageal carcinoma thus may play an important role in influencing the final outcome upon NAT course. [source]

Determination of dissociation constants of folic acid, methotrexate, and other photolabile pteridines by pressure-assisted capillary electrophoresis

ELECTROPHORESIS, Issue 17 2006
Zoltán Szakács
Abstract Pressure-assisted CE (PACE) was applied to determine the previously inaccessible complete set of pK values for folic acid and eight related multiprotic compounds. PACE allowed the determination of all acidity macroconstants at low (,0.1,mM) concentration without interferences of selfassociation or photodegradation throughout the pH range. The accuracy of the constants was verified by NMR-pH, UV-pH, and potentiometric titrations and the data could be converted into physiological ionic strength. It was shown that even three overlapping pK values can be determined by CE with good precision (<0.06) and accuracy if an appropriately low sample throughput is used. Experimental aspects of PACE for the quantitation of acid,base properties are analyzed. The site-specific basicity data obtained for folic acid and methotrexate (MTX) reveal that apparently slight constitutional differences between folic acid and MTX carry highly different proton-binding propensities at analogous moieties, especially at the pteridine N1,locus, providing straightforward explanation for the distinctive binding to dihydrofolate reductase at the molecular level. [source]

Focused proteomics: Monoclonal antibody-based isolation of the oxidative phosphorylation machinery and detection of phosphoproteins using a fluorescent phosphoprotein gel stain

ELECTROPHORESIS, Issue 15 2004
James Murray
Abstract We have raised monoclonal antibodies capable of immunocapturing all five complexes involved in oxidative phosphorylation for evaluating their post-translational modifications. Complex I (NADH dehydrogenase), complex II (succinate dehydrogenase), complex III (cytochrome c reductase), complex IV (cytochrome c oxidase), and complex V (F1F0 ATP synthase) from bovine heart mitochondria were obtained in good yield from small amounts of tissue in more than 90% purity in one step. The composition and purity of the complexes was evaluated by Western blotting using monoclonal antibodies against individual subunits of the five complexes. In this first study, the phosphorylation state of the proteins without inducing phosphorylation or dephosphorylation was identified by using the novel Pro-Q Diamond phosphoprotein gel stain. The major phosphorylated components were the same as described before in sucrose gradient enriched complexes. In addition a few additional potential phosphoproteins were observed. Since the described monoclonal antibodies show cross reactivity to human proteins, this procedure will be a fast and efficient way of studying post-translational modifications in control and patient samples using only small amounts of tissue. [source]

Tissue-specific metabolic activation and mutagenicity of 3-nitrobenzanthrone in MutaÔMouse,

ENVIRONMENTAL AND MOLECULAR MUTAGENESIS, Issue 8 2008
Guosheng Chen
Abstract 3-Nitrobenzanthrone (3-NBA) is a mutagen and suspected human carcinogen detected in diesel exhaust, airborne particulate matter, and urban soil. We investigated the tissue specific mutagenicity of 3-NBA at the lacZ locus of transgenic MutaÔMouse following acute single dose or 28-day repeated-dose oral administration. In the acute high dose (50 mg/kg) exposure, increased lacZ mutant frequency was observed in bone marrow and colonic epithelium, but not in liver and bladder. In the repeated-dose study, a dose-dependent increase in lacZ mutant frequency was observed in bone marrow and liver (2- and 4-fold increase above control), but not in lung or intestinal epithelium. In addition, a concentration-dependent increase in mutant frequency (8.5-fold above control) was observed for MutaÔMouse FE1 lung epithelial cells exposed in vitro. 1-Nitropyrene reductase, 3-NBA reductase, and acetyltransferase activities were measured in a variety of MutaÔMouse specimens in an effort to link metabolic activation and mutagenicity. High 3-NBA nitroreductase activities were observed in lung, liver, colon and bladder, and detectable N -acetyltransferase activities were found in all tissues except bone marrow. The relatively high 3-NBA nitroreductase activity in MutaÔMouse tissues, as compared with those in Salmonella TA98 and TA100, suggests that 3-NBA is readily reduced and activated in vivo. High 3-NBA nitroreductase levels in liver and colon are consistent with the elevated lacZ mutant frequency values, and previously noted inductions of hepatic DNA adducts. Despite an absence of induced lacZ mutations, the highest 3-NBA reductase activity was detected in lung. Further studies are warranted, especially following inhalation or intratracheal exposures. Environ. Mol. Mutagen., 2008. Published 2008 Wiley-Liss, Inc. [source]