NQO2 is a reactive oxygen species generating off-target for acetaminophen

[Image: see text] The analgesic and antipyretic compound acetaminophen (paracetamol) is one of the most used drugs worldwide. Acetaminophen overdose is also the most common cause for acute liver toxicity. Here we show that acetaminophen and many structurally related compounds bind quinone reductase 2 (NQO2) in vitro and in live cells, establishing NQO2 as a novel off-target. NQO2 modulates the levels of acetaminophen derived reactive oxygen species, more specifically superoxide anions, in cultured cells. In humans, NQO2 is highly expressed in liver and kidney, the main sites of acetaminophen toxicity. We suggest that NQO2 mediated superoxide production may function as a novel mechanism augmenting acetaminophen toxicity

A microfluidic device for the quantization of subpopulations of cells

International audienc

LUCS (Light-Up Cell System), a universal high throughput assay for homeostasis evaluation in live cells

Observations of fluorescent cyanine dye behavior under illumination at 500 nm lead to a novel concept in cell biology allowing the development of a new live cell assay called LUCS, for Light-Up Cell System, measuring homeostasis in live cells. Optimization of the LUCS process resulted in a standardized, straightforward and high throughput assay with applications in toxicity assessment. The mechanisms of the LUCS process were investigated. Electron Paramagnetic Resonance experiments showed that the singlet oxygen and hydroxyl radical are involved downstream of the light effect, presumably leading to deleterious oxidative stress that massively opens access of the dye to its intracellular target. Reversible modulation of LUCS by both verapamil and proton availability indicated that plasma membrane proton/cation antiporters, possibly of the MATE drug efflux transport family, are involved. A mechanistic model is presented. Our data show that intracellular oxidation can be controlled by tuning light energy, opening applications in regulatory purposes, anti-oxidant research, chemotherapy efficacy and dynamic phototherapy strategies

Strategy of red blood cells immobilisation onto a gold electrode: Characterization by electrochemical impedance spectroscopy and quartz crystal microbalance

International audienceThis study describes the grafting of red blood cells (RBC) onto a gold electrode. The erythrocytes were immobilised using antigen/antibody crosslinking based on the bonding of anti-D with the corresponding antigen of the RBC membrane that is shared by all erythrocytes from the positive rhesus group (O+). To optimise the reproducibility of the modified electrode and to avoid nonspecific interactions, the anti-D layer was deposited onto a protein G layer. The bridge between the protein G and the gold transducer was formed using mixed thiol-based self-assembled monolayer (SAM) (a mixture of 11-mercaptoundecanoic acid and 6-mercapto-1-hexanol in a 1/10 ratio). Each layer deposited was characterized, firstly, with a quartz crystal microbalance to obtain the deposited mass and the corresponding number of moles per square centimetres and secondly, by electrochemical impedance spectroscopy (EIS) using a redox couple Fe(CN)63−/4− (1:1) as an EIS probe. Subsequent modelling with appropriate circuitry enabled the values for each component representing the interface (electrode/film/solution) to be calculated at each step of the grafting process. From these results the surface coverage has been calculated to range from 95 to 98%.RésuméCe travail présente une stratégie innovante permettant le greffage de globules rouges à la surface d’électrodes en or. L’immobilisation du globule rouge repose sur l’interaction de type antigène/anticorps se produisant entre un anticorps (anti-D) et l’antigène correspondant présent dans la membrane du globule rouge de groupe O+. Afin d’optimiser la reproductibilité et d’éviter les interactions non spécifiques, l’anti-D a été déposé sur un film de protéine G. Le lien entre l’or et la protéine G a été réalisé grâce à une monocouche de thiols mixtes autoassemblée (SAM) (mélange d’acide 11-mercaptoundécanoïque et de 6-mercapto-1-hexanol en proportion 1/10). Chaque couche a été caractérisée, d’une part, par microbalance à quartz afin d’obtenir la masse du film déposé ainsi que le nombre de moles de greffons par centimètre carré correspondant et, d’autre part, par spectroscopie d’impédance électrochimique en utilisant le couple redox Fe(CN)63−/4− (1:1) comme sonde. Les mesures impédancemétriques ainsi que les modélisations utilisant un modèle adéquat ont permis de calculer la valeur des composants du circuit électrique équivalent ainsi que le taux de recouvrement de chacune des couches déposées qui sont de l’ordre de 95 à 98 %

Ability of certain plant extracts traditionally used to treat ciguatera fish poisoning to inhibit nitric oxide production in RAW 264.7 macrophages

Aim of the study: Ciguatera fish poisoning (CFP) is an intertropical ichthyosarcotoxism that manifests in complex assortment of symptoms in humans. Ciguatoxins (CTXs), issued from Gambierdicus spp., are causative agents of this intoxication. We have recently demonstrated that a Pacific CTX (P-CTX-1 B) strongly modulated iNOS expression, leading to overproduction of nitric oxide (NO) in RAW 264.7 murine macrophage cells. NO produced in large amounts is involved in a wide range of pathophysiological processes. Many traditional remedies are commonly used in the Pacific against CFP. In this context, bioassay-guided screening was carried out to study NO inhibiting capacity of 28 selected plant extracts. Materials and methods: We prepared aqueous extracts of plants used in New Caledonia in the treatment of CFP and screened their NO inhibitory activity in lipopolysaccharide (LPS)-activated RAW 264.7 macrophages. Results: Among 28 plants tested, Euphorbia hirta (Euphorbiaceae), Syzygium malaccense (Myrtaceae), Schinus terebenthifolius (Anacardiaceae), Punica granatum (Punicaceae), Cerbera manghas (Apocynaceae), Vitex trifolia (Labiateae) and Ximenia americana (Olacaceae) showed inhibitory activity, validating their use as traditional remedies in CFP, and the potential for use in the treatment of conditions accompanied by NO overproduction. Conclusion: These plants are promising candidates for further screening of their active compounds through activity-guided fractionation

Effect of ion doping in silica-based nanoparticles on the hemolytic and oxidative activity in contact with human erythrocytes

Aim: The aim of this study was the synthesis of ion doped silica-based nanoparticles and the evaluation of their toxic effect on erythrocytes. Materials & methods: Their synthesis was performed using the sol-gel method, by the progressive addition of calcium, magnesium and copper ions on pure silica nanoparticles. The toxicity evaluation was based on hemolysis, lipid peroxidation, ROS, H2O2 species and antioxidant enzyme production. Results: The addition of Mg and Cu in the SNs presented better hemocompatibility by protecting erythrocytes from oxidative stress. Conclusion: Ion doping with magnesium in the investigated calcium silicate system induces a protective effect in erythrocyte membrane in compare with pure silica nanoparticles

Antileishmanial compounds isolated from Psidium guajava L. using a metabolomic approach

With an estimated annual incidence of one million cases, leishmaniasis is one of the top five vector-borne diseases. Currently available medical treatments involve side effects, including toxicity, non-specific targeting, and resistance development. Thus, new antileishmanial chemical entities are of the utmost interest to fight against this disease. The aim of this study was to obtain potential antileishmanial natural products from Psidium guajava leaves using a metabolomic workflow. Several crude extracts from P. guajava leaves harvested from different locations in the Lao People's Democratic Republic (Lao PDR) were profiled by liquid chromatography coupled to high-resolution mass spectrometry, and subsequently evaluated for their antileishmanial activities. The putative active compounds were highlighted by multivariate correlation analysis between the antileishmanial response and chromatographic profiles of P. guajava mixtures. The results showed that the pooled apolar fractions from P. guajava were the most active (IC50 = 1.96 +/- 0.47 mu g/mL). Multivariate data analysis of the apolar fractions highlighted a family of triterpenoid compounds, including jacoumaric acid (IC50 = 1.318 +/- 0.59 mu g/mL) and corosolic acid (IC50 = 1.01 +/- 0.06 mu g/mL). Our approach allowed the identification of antileishmanial compounds from the crude extracts in only a small number of steps and can be easily adapted for use in the discovery workflows of several other natural products

Electrochemical behavior of indolone-N-oxides : relationship to structure and antiplasmodial activity

Indolone-N-oxides exert high parasiticidal activity at the nanomolar level in vitro against Plasmodium falciparum, the parasite responsible for malaria. The bioreductive character of these molecules was investigated using cyclic voltammetry and EPR spectroelectrochemistry to examine the relationship between electrochemical behavior and antimalarial activity and to understand their mechanisms of action. For all the compounds (37 compounds) studied, the voltammograms recorded in acetonitrile showed a well-defined and reversible redox couple followed by a second complicated electron transfer. The first reduction (-0.88 V < E-1/2 < -0.50 V vs. SCE) was attributed to the reduction of the N-oxide function to form a radical nitroxide anion. The second reduction (-1.65 V <E-1/2 < -1.14 V vs. SCE) was assigned to the reduction of the ketone function. By coupling electrochemistry with EPR spectroscopy, the EPR spectra confirmed the formation of the nitroxide anion radical. Moreover, the experiments demonstrated that a slow protonation occurs at the carbon of the nitrone function and not at the NO function. A relationship between electrochemical behavior and indolone-N-oxide structure can be established for compounds with R-1 = -OCH3, R-2 = H, and electron-withdrawing substituents on the phenyl group at R-3. The results help in the design of new molecules with more potent in vivo antimalarial activity