Search results for "Luciferase"
showing 10 items of 67 documents
MicroRNA hsa-miR-4717-5p regulates RGS2 and may be a risk factor for anxiety-related traits
2015
Regulator of G-protein Signaling 2 (RGS2) is a key regulator of G-protein-coupled signaling pathways involved in fear and anxiety. Data from rodent models and genetic analysis of anxiety-related traits and disorders in humans suggest down-regulation of RGS2 expression to be a risk factor for anxiety. Here we investigated, whether genetic variation in microRNAs mediating posttranscriptional down-regulation of RGS2 may be a risk factor for anxiety as well. 75 microRNAs predicted to regulate RGS2 were identified by four bioinformatic algorithms and validated experimentally by luciferase reporter gene assays. Specificity was confirmed for six microRNAs (hsa-miR-1271-5p, hsa-miR-22-3p, hsa-miR-3…
Surfactant effect on the physicochemical characteristics of cationic solid lipid nanoparticles
2016
Solid lipid nanoparticles (SLNs) may be considered as a new approach for therapeutics for many diseases. In addition to drug delivery, their use as non-viral vectors for gene delivery can be obtained by including cationic lipids, which provide a positive surface potential that favors binding to the nucleic acids as DNA, siRNA, miRNA, etc. In fact, the addition of cationic surfactants is indispensable for obtaining nanoparticles with surface positive charge. In this study, three different cationic lipids (dioctadecyl dimethyl ammonium bromide, cetyltrimethyl ammonium bromide, cetylpyridinium chloride) and Brij 76 as nonionic surfactant were employed to formulate Precirol ATO 5 based cSLN usi…
Discovery of 5-benzyl-3-phenyl-4,5-dihydroisoxazoles and 5-benzyl-3-phenyl-1,4,2-dioxazoles as potent firefly luciferase inhibitors.
2013
Luciferase reporter assays are commonly used in high-throughput screening methods. Here, we report new firefly luciferase (FLuc) inhibitors based on 5-benzyl-3-phenyl-4,5-dihydroisoxazoles and 5-benzyl-3-phenyl-1,4,2-dioxazoles, which showed up as "false positives" in a luciferase reporter gene-based assay for nuclear receptor antagonists. The inhibition was shown to be noncompetitive for both natural enzyme substrates (d-luciferin and ATP) and selective to FLuc and proven to arise from a direct interaction between the enzyme and the inhibitor. Of the 63 evaluated compounds, 28 showed significantly better inhibition potency than the well-known inhibitor resveratrol (IC(50) = 59 nM), with fi…
Polyhydroxyethylaspartamide-spermine copolymers: Efficient vectors for gene delivery
2008
Abstract Aim of this paper was that to prepare biocompatible, polyaspartamide based copolymers containing spermine or spermine/hydrophobic side chains able to condense nucleic acids and to transfect mammalian cells. Copolymers were prepared starting from α,β-poly-(N-2-hydroxyethyl)- d , l -aspartamide (PHEA) and exploiting the reactive hydroxyl groups in the polymeric side chains by subsequent activation reactions to obtain PHEA-Spermine (PHEA-Spm) and PHEA-Spermine-Butyramide (PHEA-Spm-C4). Molecular, physico-chemical and biological characterization of copolymers and interpolyelectrolyte complexes with plasmid DNA was performed. Experimental results evidenced that these copolymers are able…
Bis(hydroxyphenyl)methane-bisphenol F-metabolism by the HepG2 human hepatoma cell line and cryopreserved human hepatocytes
2011
author cannot archive publisher's version/PDF; International audience; Bisphenol F (BPF) is present in the environment and as a contaminant of food. Humans may, therefore, be exposed to BPF, and an assessment of this risk is required. BPF has been shown to have genotoxic and endocrine-disruptor properties in a human hepatoma cell line (HepG2), which is a model system for studies of xenobiotic toxicity. In this study, we investigated the ability of HepG2 cells to biotransform BPF, because metabolism may affect the observed effects of BPF, and we compared this metabolic capacity with that of human hepatocytes. Cells were incubated for 24 hours with [(3)H]-BPF. The culture medium was then conc…
Exclusive transduction of human CD4+ T Cells upon systemic delivery of CD4-targeted lentiviral vectors
2015
Abstract Playing a central role in both innate and adaptive immunity, CD4+ T cells are a key target for genetic modifications in basic research and immunotherapy. In this article, we describe novel lentiviral vectors (CD4-LV) that have been rendered selective for human or simian CD4+ cells by surface engineering. When applied to PBMCs, CD4-LV transduced CD4+ but not CD4− cells. Notably, also unstimulated T cells were stably genetically modified. Upon systemic or intrasplenic administration into mice reconstituted with human PBMCs or hematopoietic stem cells, reporter gene expression was predominantly detected in lymphoid organs. Evaluation of GFP expression in organ-derived cells and blood …
Geographical mapping of metabolites in biological tissue with quantitative bioluminescence and single photon imaging
1993
This article features a novel technique for measuring the spatial distribution of metabolites, such as ATP, glucose, and lactate, in rapidly frozen tissue. Concentration values are obtained in absolute terms and with a spatial resolution of single-cell dimension. The method is based on enzymatic reactions that link the metabolite of interest to luciferase with subsequent light emission. Using a specific array, cryosections are brought into contact with the enzymes in a well-defined, reproducible way inducing a distribution of light across the section with an intensity that is proportional to the metabolite concentration. The emitted light can be visualized through a microscope and an imagin…
Differential Promotion of Glutamate Transporter Expression and Function by Glucocorticoids in Astrocytes from Various Brain Regions
2005
Steroids that activate glucocorticoid receptors (GRs) and mineralocorticoid receptors have important regulatory effects on neural development, plasticity, and the body's stress response. Here, we investigated the role of corticosteroids in regulating the expression of the glial glutamate transporters glial glutamate transporter-1 (GLT-1) and glutamate-aspartate transporter (GLAST) in rat primary astrocytes. The synthetic glucocorticoid dexamethasone provoked a marked increase of GLT-1 transcription and protein levels in cortical astrocytes, whereas GLAST expression remained unaffected. Up-regulation of GLT-1 expression was accompanied by an enhanced glutamate uptake, which could be blocked …
Sequence similarity of mammalian epoxide hydrolases to the bacterial haloalkane dehalogenase and other related proteins Implication for the potential…
1994
Direct comparison of the amino acid sequences of microsomal and soluble epoxide hydrolase superficially indicates that these enzymes are unrelated. Both proteins, however, share significant sequence similarity to a bacterial haloalkane dehalogenase that has earlier been shown to belong to the alpha/beta hydrolase fold family of enzymes. The catalytic mechanism for the dehalogenase has been elucidated in detail [Verschueren et al. (1993) Nature 363, 693-698] and proceeds via an ester intermediate where the substrate is covalently bound to the enzyme. From these observations we conclude (i) that microsomal and soluble epoxide hydrolase are distantly related enzymes that have evolved from a co…
Comparison of a Novel Homogeneous Cyclic Amp Assay and a Luciferase Assay for Measuring Stimulating Thyrotropin-Receptor Autoantibodies.
2019
Objective: Stimulating thyrotropin-receptor antibodies (TSAb) cause Graves’ disease (GD). We tested a novel homogeneous fluorescent 3′,5′ cyclic adenine monophosphate (cAMP) assay for the detection of TSAb in a bioassay. Methods: Chinese hamster ovary (CHO) cell lines expressing either a chimeric (MC4) or wild-type (WT) TSH-R were incubated with the adenyl cyclase activator forskolin, a human TSAb monoclonal antibody (M22), and with sera from GD patients. Intracellular cAMP levels were measured using a Bridge-It® cAMP assay, and the results were compared with a luciferase-based bioassay. Results: Both cell lines were stimulated with forskolin concentrations (0.006–200 µM) in a dose-dependen…