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Zhang Qunwei; Salnikow, Konstantin; Kluz, Thomas; Chen, L.C.; Su, W.C.; Costa, Max, E-mail: costam@env.med.nyu.edu
arXiv e-print [ PDF ]2003
arXiv e-print [ PDF ]2003
AbstractAbstract
[en] The carcinogenic process initiated by nongenotoxic carcinogens involves modulation of gene expression. Nickel compounds have low mutagenic activity, but are highly carcinogenic. In vitro both mouse and human cells can be efficiently transformed by soluble and insoluble nickel compounds to anchorage-independent growth. Because previous studies have shown that carcinogenic nickel compounds silence genes by inhibiting histone acetylation and enhancing DNA methylation, we investigated the effect of enhancing histone acetylation on cell transformation. The exposure of nickel-transformed cells to the histone deacetylase inhibitor trichostatin A (TSA) resulted in the appearance of significant number of revertants measured by their inability to grow in soft agar. Using the Affymetrix GeneChip we found that the level of expression of a significant number of genes was changed (suppressed or upregulated) in nickel-transformed clones but returned to a normal level in revertants obtained following TSA treatment. Moreover, we found that treatment of cells with TSA inhibited the ability of nickel to transform mouse PW cells to anchorage-independent growth. Treatment with TSA also inhibited the ability of nickel to transform human HOS cells, although to a lesser extent. In contrast, treatment with TSA was not able to revert established cancer cell lines as readily as the nickel-transformed cells. These data indicated that modulation of gene expression is important for nickel-induced transformation
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S0041008X03002801; Copyright (c) 2003 Elsevier Science B.V., Amsterdam, The Netherlands, All rights reserved.; Country of input: International Atomic Energy Agency (IAEA)
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[en] The role of metal ions as enzyme effectors is considered. Data on inhibitory and activating effects of metal ions are summarised. The dual character of action of the effectors depending on their concentration and the nature of the enzyme is highlighted. The analytical applications of these effects are discussed. The bibliography includes 66 references.
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Available from http://dx.doi.org/10.1070/RC1998v067n03ABEH000228; Country of input: International Atomic Energy Agency (IAEA)
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Russian Chemical Reviews (Print); ISSN 0036-021X;
; v. 67(3); p. 225-232

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[en] We report on an electrochemical biosensor for the determination of the activity of dipeptidyl peptidase-IV (DPP-IV), and on a method for screening the effect of its inhibitors. An enzyme substrate (Fc-peptide) was immobilized on the surface of a gold electrode, and double signal amplification was accomplished via an additional layer consisting of phenyl rings and gold nanoparticles. The activity of DPP-IV was determined at levels as low as 39 nU·mL−1 and over a linear detection range as wide as from 0.5 μU·mL−1 to 2.5 mU·mL−1. The inhibitory effects of diprotin A and the His-Leu dipeptide on the activity of DPP-IV also were tested and gave IC50 values of 93.5 and 95.5 μM, respectively. The assay is rapid, precise and selective. It may be extended to other peptidases and, possibly, proteases and their inhibitors. (author)
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Microchimica Acta (Online); ISSN 1436-5073;
; v. 182(1-2); p. 281-288

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[en] The immobilization of biological molecules onto polymeric membranes to produce biofunctional membranes is used for selective catalysis, separation, analysis, and artificial organs. Normally, random immobilization of enzymes onto polymeric membranes leads to dramatic reduction in activity due to chemical reactions involved in enzyme immobilization, multiple-point binding, etc., and the extent of activity reduction is a function of membrane hydrophilicity (e.g. activity in cellulosic membrane >> polysulfone membrane). We have used molecular biology to effect site-specific immobilization of enzymes in a manner that orients the active site away from the polymeric membrane surface, thus resulting in higher enzyme activity that approaches that in solution and in increased stability of the enzyme relative to the enzyme in solution. A prediction of this site-specific method of enzyme immobilization, which in this study with subtilisin and organophosphorus hydrolase consists of a fusion tag genetically added to these enzymes and subsequent immobilization via the anti-tag antibody and membrane-bound protein A, is that the active site conformation will more closely resemble that of the enzyme in solution than is the case for random immobilization. This hypothesis was confirmed using a new electron paramagnetic resonance (EPR) spin label active site titration method that determines the amount of spin label bound to the active site of the immobilized enzyme. This value nearly perfectly matched the enzyme activity, and the results suggested: (a) a spectroscopic method for measuring activity and thus the extent of active enzyme immobilization in membrane, which may have advantages in cases where optical methods can not be used due to light scattering interference; (b) higher spin label incorporation (and hence activity) in enzymes that had been site-specifically immobilized versus random immobilization; (c) higher spin label incorporation in enzymes immobilized onto hydrophilic bacterial cellulose membranes versus hydrophobic modified poly(ether)sulfone membranes. These results are discussed with reference to analysis and utilization of biofunctional membranes
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S0003267002005366; Copyright (c) 2002 Elsevier Science B.V., Amsterdam, The Netherlands, All rights reserved.; Country of input: International Atomic Energy Agency (IAEA)
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[en] Bromodomain-containing protein 4 (Brd4) is known to play a key role in tumorigenesis. It binds acetylated histones to regulate the expression of numerous genes. Because of the importance of brd4 in tumorigenesis, much research has been undertaken to develop brd4 inhibitors with therapeutic potential. As a result, various scaffolds for bromodomain inhibitors have been identified. To discover new scaffolds, we performed mid-throughput screening using two different enzyme assays, alpha-screen and ELISA. We found a novel bromodomain inhibitor with a unique scaffold, aristoyagonine. This natural compound showed inhibitory activity in vitro and tumor growth inhibition in a Ty82-xenograft mouse model. In addition, we tested Brd4 inhibitors in gastric cancer cell lines, and found that aristoyagonine exerted cytotoxicity not only in I-BET-762-sensitive cancer cells, but also in I-BET-762-resistant cancer cells. This is the first paper to describe a natural compound as a Brd4 bromodomain inhibitor.
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S0006291X18314037; Available from http://dx.doi.org/10.1016/j.bbrc.2018.06.091; Copyright (c) 2018 Elsevier Inc. All rights reserved.; Country of input: International Atomic Energy Agency (IAEA)
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Biochemical and Biophysical Research Communications; ISSN 0006-291X;
; CODEN BBRCA9; v. 503(2); p. 882-887

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[en] A direct radio-immunoassay (RIA) for renin substrate (RS) was compared to enzymatic (indirect) assay. In normal subjects, a significant, albeit weak, correlation between the methods was seen. In hypertensive patients with different levels of plasma renin activity (PRA), RS concentration measured by both assays increased with increasing PRA, and for patients with PRA > 10 μg AI/l/h, direct assay gave significantly higher RS values (55%), compared to the enzymatic assay, indicating consumption of RS by increasing plasma renin and production rate of RS with increasing PRA. In 11 patients with renovascular hypertension, treatment with angiotensin-converting enzyme (ACE) inhibitor, lisinopril, resulted in a significant increase in PRA, accompanied by a decrease in RS measured by enzymatic assay. No change in RS measured by direct RIA was noticed. The results suggest that ACE inhibition may not have an effect upon RS production and that its effect on plasma RS is limited to a reduction of intact RS measured by the enzymatic assay. (author)
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Scandinavian Journal of Clinical and Laboratory Investigation; ISSN 0036-5513;
; CODEN SJCLA; v. 48(2); p. 131-136

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[en] Enzyme-linked immunoassays based on an antibody-antigen interaction are widely used in biological and medical sciences. However, the conjugation of an enzyme to antibodies needs an additional chemical process, usually resulting in randomly cross-linked molecules and a loss of the binding affinity and enzyme activity. Herein, we present the development of an alkaline phosphatase-fused repebody as a new format of immuno-reagent for immunoassays. A repebody specifically binding to human TNF-α (hTNF-α) was selected through a phage display, and its binding affinity was increased up to 49 nM using a modular engineering approach. A monomeric alkaline phosphatase (mAP), which was previously isolated from a metagenome library, was genetically fused to the repebody as a signal generator, and the resulting repebody-mAP fusion protein was used for direct and sandwich immunoassays of hTNF-α. We demonstrate the utility and potential of the repebody-mAP fusion protein as an immuno-reagent by showing the sensitivity of 216 pg mL"−"1 for hTNF-α in a sandwich immunoassay. Furthermore, this repebody-mAP fusion protein enabled the detection of hTNF-α spiked in a serum-supplemented medium with high accuracy and reproducibility. It is thus expected that a mAP-fused repebody can be broadly used as an immuno-reagent in immunoassays. - Highlights: • A human TNF-α (hTNF-α)-specific repebody was selected using a phage display. • A monomeric alkaline phosphatase (mAP) was genetically fused to the repebody. • mAP-fused repebody enabled detection of hTNF-α with high sensitivity and accuracy. • mAP-fused repebody can be widely used as a new immuno-reagent in immunoassays.
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S0003-2670(16)31298-3; Available from http://dx.doi.org/10.1016/j.aca.2016.11.013; Copyright (c) 2016 Elsevier Science B.V., Amsterdam, The Netherlands, All rights reserved.; Country of input: International Atomic Energy Agency (IAEA)
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[en] Introduction: There are many factors causing endothelial dysfunction. The aim was to observe chosen markers of endothelial function in patients with subclinical and overt hyperthyroidism. Material and methods: We studied 97 patients with hyperthyroidism: 51 with subclinical (44 F/7 M; mean age 49.3 ± 15.9 y) and 46 patients with overt (39 F/7 M, mean age 50.4 ± 13.2 y). The control comprised of 39 healthy volunteers (26 F/13 M, mean age 47.5 ± 11.8 y). Concentration of TSH, FT3, FT4 were measured by MEIA, TPO Ab, TG Ab, E-selectin, interleukin 6, VCAM-1, ICAM-1 by ELISA. Results: The goiter was found in 71 persons 63F/8M, mean age 49.9 ± 15.3 y, (42-subclinical, 29-overt). Morbus Graves--Basedow was diagnosed in 26 persons, 20 F/6 M, mean age 49.5 ± 12.8 y (9-subclinical, 17-overt). There were no significant differences serum concentration of E-selectin, IL-6, ICAM-1 in patients with subclinical and overt hyperthyroidism compared to the control. Statistically significant differences were shown between concentration of IL-6 in patients with Graves-Basedow compared with the control (p < 0.05). Significance of VCAM-1 values were found in the patients with subclinical and overt hyperthyroidism compared to the control (p < 0.001; p < 0.001, respectively). Conclusions: Among persons with overt and subclinical hyperthyroidism occurs endothelial dysfunction which doesn't depends on exciting cause of thyrotoxicosis but on degree of hyperthyroidism. Elevated concentrations of endothelial markers may confirm that persons with thyroid disorders are extremely exposed to the occurrence of the cardiovascular diseases. (author)
Original Title
Wybrane wskazniki uszkodzenia srodblonka wsrod pacjentow z subkliniczna i jawna nadczynnoscia tarczycy
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29 refs., 5 tabs.
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Endokrynologia Polska; ISSN 0423-104X;
; v. 57(3); p. 202-209

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[en] Selenium is an essential component of selenoproteins, enzymes with extensive regulatory and protective effect in organism. Immunological effects of Se are documented and are distinct even above concentrations necessary for maximal activity of selenoenzymes. Therefore, we investigated effect of supplementation by 100 μg of yeast-bound Se on concentrations of thyroid autoantibodies TPOAb and TgAb in the group of 253 seniors living in the Asylum Houses of South Bohemia. Increase of serum selenium from 59 to 150 μg Se/L serum in supplemented group and from 59 to 72 μg Se/L serum in group with placebo were detected by Instrumental Neutron Activation Analysis (INAA) and proved increased Se intake during the trial. Autoantibodies were analyzed by ELISA at the beginning of the trial and after 1 year. Statistical evaluation of results in whole groups (regardless of increased autoantibodies) by ANOVA manifested significant decrease of TPOAb and TgAb in non-supplemented group while supplementation did not effect serum autoantibodies concentrations. Evaluation of groups of seniors created from those with increased autoantibodies, ANOVA demonstrated decrease of TPOAb in both groups but Se supplementation did not affect the decrease. In opposite, TgAb increased significantly and Se supplementation led to higher increase of TgAb. Recent results of possibility to decrease serum concentration of TPOAb proved this effect only for high TPOAb concentrations and for higher Se supplements. From this point of view, it is necessary to conduct subsequent trials with the patients with autoimmune thyreoiditis with different levels of autoantibodies and detect also serum Se levels. (author)
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18 refs.
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Journal of Radioanalytical and Nuclear Chemistry; ISSN 0236-5731;
; CODEN JRNCDM; v. 280(2); p. 275-279

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[en] A 96-well microtiter enzyme-linked immunosorbent assay (ELISA) for protein tyrosine kinases has been developed. This assay uses one of several substrates that are phosphorylated by tyrosine kinase, an antibody to phosphotyrosine, and a peroxidase-linked second antibody. Color development is monitored by a change in absorbance at 450 nm and is dependent upon time, enzyme, ATP, and substrate concentrations. Specificity of the ELISA for phosphotyrosine was shown by inhibition of binding of the anti-phosphotyrosine antibody with phenyl phosphate. Results obtained in the ELISA compared favorably with those obtained by direct phosphorylation of the substrate with [32P]ATP. Staurosporine and K252a, protein kinase inhibitors, showed titratable inhibition of tyrosine kinase activity. This assay is a rapid, nonradioactive alternative to traditional methodology and is also amenable to automation
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AMINO ACIDS, AROMATICS, BETA DECAY RADIOISOTOPES, BETA-MINUS DECAY RADIOISOTOPES, CARBOXYLIC ACIDS, CHEMICAL REACTIONS, DAYS LIVING RADIOISOTOPES, ENZYMES, HYDROXY ACIDS, IMMUNOASSAY, ISOTOPE APPLICATIONS, ISOTOPES, LIGHT NUCLEI, NUCLEI, ODD-ODD NUCLEI, ORGANIC ACIDS, ORGANIC COMPOUNDS, PHOSPHORUS ISOTOPES, PHOSPHORUS-GROUP TRANSFERASES, RADIOISOTOPES, TRANSFERASES
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