Precision-Cut Liver Slices of Salmo salar as a tool to investigate the oxidative impact of CYP1A-mediated PCB 126 and 3-methylcholanthrene metabolism

a b s t r a c t Fish isolated cell systems have long been used to predict in vivo toxicity of man-made chemicals. In present study, we tested the suitability of Precision-Cut Liver Slices (PCLS) as an alternative to these models that allows the evaluation of a global tissue response to toxicants, to investigate oxidative stress response to cytochrome P450 1A (CYP1A) induction in fish liver. PCLS of Salmo salar were exposed for 21 h to increasing doses of 3-methylcholanthrene (3-MC) and Polychlorobiphenyl 126 (PCB 126). 3-MC (25 lM) strongly induced CYP1A transcription. In dose-response analysis (25-100 lM), EROD activity was strongly increased at intermediate 3-MC concentrations. We found the counter-intuitive decline of EROD at the highest 3-MC doses to result from reversible competition with ethoxyresorufin. No increases of H 2 O 2 production, antioxidant enzymes activities or oxidative damage to lipids were found with 3-MC treatments. PCLS subjected to PCB 126 (2-200 nM) showed increased contamination levels and a parallel increased CYP1A mRNA synthesis and EROD activity. H 2 O 2 production tended to increase but no oxidative damage to lipids was found. As antioxidant enzymes activities declined at the highest PCB 126 dose, it is suggested that longer incubation periods could be required to generate oxidative stress in PCLS

PRIMA subretinal wireless photovoltaic microchip implantation in non-human primate and feline models

Purpose To evaluate the surgical technique for subretinal implantation of two sizes of PRIMA photovoltaic wireless microchip in two animal models, and refine these surgical procedures for human trials. Methods Cats and Macaca fascicularis primates with healthy retina underwent vitrectomy surgery and were implanted with subretinal wireless photovoltaic microchip at the macula/central retina. The 1.5mm PRIMA chip was initially studied in feline eyes. PRIMA implant (2mm,1.5mm sizes) arrays were studied in primates. Feasibility of subretinal chip implantation was evaluated with a newly-developed surgical technique, with surgical complications and adverse events recorded. Results The 1.5mm implant was placed in the central retina of 11 feline eyes, with implantation duration 43-106 days. The 1.5mm implant was correctly positioned into central macula of 11 primate eyes, with follow-up periods of minimum 6 weeks (n = 11), 2 years (n = 2), and one eye for 3 years. One primate eye underwent multi-chip 1.5mm implantation using two 1.5mm chips. The 2mm implant was delivered to 4 primate eyes. Optical coherence tomography confirmed correct surgical placement of photovoltaic arrays in the subretinal space in all 26 eyes. Intraoperative complications in primate eyes included retinal tear, macular hole, retinal detachment, and vitreous hemorrhage that resolved spontaneously. Postoperatively, there was no case of significant ocular inflammation in the 1.5mm implant group. Conclusions We report subretinal implantation of 1.5mm and 2mm photovoltaic arrays in the central retina of feline and central macula of primate eyes with a low rate of device-related complications. The in vivo PRIMA implantation technique has been developed and refined for use for a 2mm PRIMA implant in ongoing human trials

Restoring Specific Lactobacilli Levels Decreases Inflammation and Muscle Atrophy Markers in an Acute Leukemia Mouse Model

The gut microbiota has recently been proposed as a novel component in the regulation of host homeostasis and immunity. We have assessed for the first time the role of the gut microbiota in a mouse model of leukemia (transplantation of BaF3 cells containing ectopic expression of Bcr-Abl), characterized at the final stage by a loss of fat mass, muscle atrophy, anorexia and inflammation. The gut microbial 16S rDNA analysis, using PCR-Denaturating Gradient Gel Electrophoresis and quantitative PCR, reveals a dysbiosis and a selective modulation of Lactobacillus spp. (decrease of L. reuteri and L. johnsonii/gasseri in favor of L. murinus/animalis) in the BaF3 mice compared to the controls. The restoration of Lactobacillus species by oral supplementation with L. reuteri 100-23 and L. gasseri 311476 reduced the expression of atrophy markers (Atrogin-1, MuRF1, LC3, Cathepsin L) in the gastrocnemius and in the tibialis, a phenomenon correlated with a decrease of inflammatory cytokines (interleukin-6, monocyte chemoattractant protein-1, interleukin-4, granulocyte colony-stimulating factor, quantified by multiplex immuno-assay). These positive effects are strain- and/or species-specific since L. acidophilus NCFM supplementation does not impact on muscle atrophy markers and systemic inflammation. Altogether, these results suggest that the gut microbiota could constitute a novel therapeutic target in the management of leukemia-associated inflammation and related disorders in the muscle

N° 32. — Interactions moléculaires dans les mélanges ternaires alcool polyvinylique-eau-sel minéral

L’étude de la résonance du proton dans les solutions ternaires butanediol-1,3, eau, sel minéral met en évidence l’interaction particulière qui s’exerce entre le composé organique et certains électrolytes comme les thiocyanates. Les résultats obtenus peuvent être rapprochés du comportement physicochimique de l’alcool polyvinylique dans les mêmes milieux

Gene Regulatory Network Inference using ensembles of Local Multiple Kernel Models

International audienceReconstructing gene regulatory network from high-throughput data has many potential applications, from understanding a biological organism to identifying potential drug targets. It is also a notoriously difficult problem, tackled by many scientists with various methods. In this paper, we formulate GRN inference as a sparse regression problem. We decompose the prediction of a p-genes system in p different regression problems. For each gene (target gene), we train a kernel-based regression with feature selection, predicting the expression pattern of the target gene using all the other genes (input genes). The regression will give the importance of each input gene in the prediction of the target gene. We take this importance as an indication of a putative regulatory link. Putative links are then aggregated over all genes to provide a ranking of interactions, from which we infer the GRN. Furthermore, biological data are heterogeneous. The method we propose can learn from both steady-state and time-series data, using an ensemble method that can be applied to other regression model. Finally, we compare our method, called LocKING, to state-of-the-art methods on real and realistic datasets, which are widely spread in the GRN inference community. We show that our method is competitive against individual methods. Nevertheless, best results are obtained by integrating multiple methods. We show that using LocKING among other methods significantly enhances the accuracy of the network inferred

Free radical modulation by N-substituted dehydroalanines, a new way to improve therapeutic activity of anticancer drugs.

Journal ArticleSCOPUS: ar.jinfo:eu-repo/semantics/publishe

High hydrostatic pressure influences the in vitro response toxenobiotics in Dicentrarchus labrax liver

Hydrostatic pressure (HP) increases by about 1 atmosphere (0.1 MPa) for each ten-meter depth increasein the water column. This thermodynamical parameter could well influence the response to and effects ofxenobiotics in the deep-sea biota, but this possibility remains largely overlooked. To grasp the extent of HPadaptation in deep-sea fish, comparative studies with living cells of surface species exposed to chemicalsat high HP are required. We initially conducted experiments with precision-cut liver slices of a deep-sea fish (Coryphaenoides rupestris), co-exposed for 15 h to the aryl hydrocarbon receptor (AhR) agonist3-methylcholanthrene at HP levels representative of the surface (0.1 MPa) and deep-sea (5–15 MPa; i.e.,500–1500 m depth) environments. The transcript levels of a suite of stress-responsive genes, such asthe AhR battery CYP1A, were subsequently measured (Lemaire et al., 2012; Environ. Sci. Technol. 46,10310–10316). Strikingly, the AhR agonist-mediated increase of CYP1A mRNA content was pressure-dependently reduced in C. rupestris. Here, the same co-exposure scenario was applied for 6 or 15 h toliver slices of a surface fish, Dicentrarchus labrax, a coastal species presumably not adapted to high HP.Precision-cut liver slices of D. labrax were also used in 1 h co-exposure studies with the pro-oxidant tert-butylhydroperoxide (tBHP) as to investigate the pressure-dependence of the oxidative stress response(i.e., reactive oxygen production, glutathione and lipid peroxidation status). Liver cells remained viable inall experiments (adenosine triphosphate content). High HP precluded the AhR agonist-mediated increaseof CYP1A mRNA expression in D. labrax, as well as that of glutathione peroxidase, and significantly reducedthat of heat shock protein 70. High HP (1 h) also tended per se to increase the level of oxidative stress inliver cells of the surface fish. Trends to an increased resistance to tBHP were also noted. Whether thelatter observation truly reflects a protective response to oxidative stress will be addressed in future co-exposure studies with both surface and deep-sea fish liver cells, using additional pro-oxidant chemicals.Altogether, data on CYP1A inducibility with D. labrax and C. rupestris support the view that high HPrepresses AhR signaling in marine fishes, and that only species adapted to thrive in the deep-sea haveevolved the molecular adaptations necessary to counteract to some extent this inhibition

Activation of Poly(ADP-Ribose)Polymerase in rat hepatocytes does not contribute to their cell death by oxidative stress.

Oxidative stress induced by tert-butyl hydroperoxide (tBOOH) in freshly isolated rat hepatocytes caused DNA damage and loss of membrane integrity. Such DNA lesions are likely to be single strand breaks since neither caryolysis nor chromatine condensation was seen in electron micrographs from tBOOH-treated cells. In addition, pulsed field gel electrophoresis of genomic DNA from both control and tBOOH-treated hepatocytes showed similar profiles, indicating the absence of internucleosomal DNA cleavage, a classical reflection of apoptotic endonuclease activity. The activation of the repair enzyme poly(ADP-ribose)polymerase (PARP) following DNA damage by tBOOH induced a dramatic drop in both NAD(+) and ATP. The inhibition of PARP by 3-aminobenzamide enhanced DNA damage by tBOOH, restored NAD(+) and ATP levels, but did not result in better survival against cell killing by tBOOH. The lack of the protective effect of PARP inhibitor, therefore, does not implicate PARP in the mechanism of tBOOH-induced cytotoxicity. Electron micrographs also show no mitochondrial swelling in cells under oxidative stress, but such organelles were mainly located around the nucleus, a picture already observed in autoschizis, a new suggested kind of cell death which shows both apoptotic and necrotic morphological characteristics

Protein S-thiolation can mediate the inhibition of protein synthesis induced by tert-butyl hydroperoxide in isolated rat hepatocytes.

A rapid inhibition of protein synthesis is observed when isolated rat hepatocytes are incubated in the presence of 0.25-0.5 mM of tert-butyl hydroperoxide (tBOOH). Such an inhibition occurs in the absence of a cytolytic effect by tBOOH. Iron chelators (o-phenanthroline and desferrioxiamine), protected against oxidative cell death, but they did not modify the inhibition of protein synthesis caused by tBOOH (0.5 mM), suggesting that free radicals are less implicated in such an impairment. Electron micrographs of hepatocytes under oxidative stress show disaggregation of polyribosomes but not oxidative alterations, such as blebs or mitochondrial swelling. Protein synthesis inhibition is accompanied by a decrease in reduced glutathione (GSH) and an increase in glutathione disulfide (GSSG) and the level of protein S-thiolation (protein mixed disulfides formation). Such an increase of GSSG appears as a critical event since diethylmaleate (DEM) at 0.2 mM reduced GSH content by more than 50% but did not affect either GSSG content or protein synthesis. The addition of exogenous GSH and N-acetylcysteine (NAC) to tBOOH-treated hepatocytes significantly reduced the formation of protein mixed disulfides and restored the depressed protein synthesis either completely or partially. We suggest that S-thiolation of some key proteins may be involved in protein synthesis inhibition by tBOOH