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See detailIn vitro study of coinfection/superinfection parameters which can influence recombination events in noroviruses
Di Felice, Elisabetta; Ceci, Chiara; Toffoli, Barbara et al

Poster (2014, October 17)

Noroviruses (NoVs) are non-enveloped, single-stranded, positive-sense RNA viruses. They are important causes of acute non-bacterial gastroenteritis in humans worldwide but their study is currently yet ... [more ▼]

Noroviruses (NoVs) are non-enveloped, single-stranded, positive-sense RNA viruses. They are important causes of acute non-bacterial gastroenteritis in humans worldwide but their study is currently yet hampered by the lack of a cell culture system. NoVs genetically evolve by both point mutations and recombination and the murine norovirus (MuNoV) is considered as the best model for human NoVs. The aim of this study was to develop an experimental model based on the MuNoV in order to investigate coinfection/superinfection parameters that could impact recombination events. Monolayers of RAW264.7 cells were coinfected or superinfected with two MuNoV strains (CW1 and WU20) using different multiplicity of infection (0.1/1; 1/1 and 10/1 for CW1 and Wu20 , respectively) and time delays (0h; 0.5h; 1h; 2h; 4h; 8h; 12h and 24h) for infection. Supernatants were collected at 24 and 48 hours post-infection. Genomic copies of both viruses were first quantified by RT-QPCR. Then, viruses from the supernatants were plaque purified (36 clones per condition) and their recombinant status was checked by a real-time PCR discriminating method using primers targeting both extremity of the MuNoV genome. Results of quantitative and plaque picking assays are compared. Together, the results confirm that recombination does not frequently occur, at least in vitro and raise the issue on why these events are however so usual with in silico detection methods. The data also showed that superinfection exclusion seems to be triggered from 4h post infection with the first MuNoV. The mechanisms of the later should be still studied. [less ▲]

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See detailIn vitro study of fibroblasts (L929) and osteoblasts (MG-63) within networks differing in fiber density.
Lombart, François ULg; Alexandre, Diane ULg; de Bien, Charlotte ULg et al

Poster (2013, June 18)

Animal cells are typically cultivated in vitro on 2D surfaces therefore in conditions totally differing from their 3D in vivo environments. It is therefore really attractive to generate better in vitro ... [more ▼]

Animal cells are typically cultivated in vitro on 2D surfaces therefore in conditions totally differing from their 3D in vivo environments. It is therefore really attractive to generate better in vitro animal cell models where animal cells can adhere and proliferate within a 3D networks. When facing to a third dimension, the design of the scaffolds tailored to support the organization and communication of cells should favor at least cell adhesion, proliferation and differentiation, while promoting nutrient, gas (O2 / CO2) and waste product diffusion. In order to construct this in vitro model, we have compared the cell reactivity of two model cell lines fibroblaste L 929 and osteoblaste MG-63 within three 3D networks differing in fiber density. This parameter has been altered in order to increase progressively the total surface exposed to the cells, whilst increasing correspondingly the mean pore size and total porosity of the network, whilst keeping the same architecture and surface chemistry of the fibers within the scaffold. [less ▲]

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See detailIn Vitro Study of the Antioxidant Properties of Nimesulide and 4-Oh Nimesulide: Effects on Hrp- and Luminol-Dependent Chemiluminescence Produced by Human Chondrocytes
Zheng, S. X.; Mouithys-Mickalad, Ange ULg; Deby-Dupont, G. P. et al

in Osteoarthritis and Cartilage (2000), 8(6), 419-25

OBJECTIVES: Reactive oxygen species (ROS) are now recognized to play an important role in the pathogenesis of rheumatic diseases and constitute an interesting therapeutic target for drugs. This in vitro ... [more ▼]

OBJECTIVES: Reactive oxygen species (ROS) are now recognized to play an important role in the pathogenesis of rheumatic diseases and constitute an interesting therapeutic target for drugs. This in vitro study was designed to evaluate the antioxidant properties of nimesulide (NIM), a nonsteroidal antiinflammatory drug of the sulfonanilide class, and its main metabolite 4-OH nimesulide (4-OHNIM). METHODS: The scavenging effects of NIM and 4-OH NIM on hydroxyl radical ((.)OH) and superoxide anions (O(minusd)(2)) were investigated by electron spin resonance (ESR), using 5, 5-dimethylpyrroline-N-oxide (DMPO) as the spin trap agent. The quenching properties of these drugs on hypochlorite anion was studied by luminol enhanced chemiluminescence. Finally, the effects of NIM and 4-OHNIM on the reactive oxygen species production by human articular chondrocytes were recorded by HRP and luminol-enhanced chemiluminescence. RESULTS: By this method it has been demonstrated that NIM and 4-OHNIM, at concentrations ranging from 10 to 100 microM, are potent scavengers of(.)OH whereas only 4-OHNIM was capable to scavenge O(minusd)(2). Chemiluminescence generated by HOCl was also significantly and dose-dependently inhibited by both NIM and 4-OHNIM. Nevertheless, at each concentration tested, the inhibitory effect of 4-OHNIM was significantly more marked, even at the highest concentration (100 microM). Furthermore, when chondrocytes were pre-incubated for 48-96 h with NIM or 4-OHNIM, the luminol- and HRP-dependent CL produced by the cells was significantly inhibited in a dose-dependent manner. CONCLUSIONS: NIM and 4-OHNIM may protect cartilage against oxidative stress, not only by scavenging ROS but also by inhibiting their production by chondrocytes. [less ▲]

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See detailIn vitro study of the antioxidant properties of non steroidal anti-inflammatory drugs by chemiluminescence and electron spin resonance (ESR).
Mouithys-Mickalad, Ange ULg; Zheng, S. X.; Deby-Dupont, G. P. et al

in Free Radical Research (2000), 33(5), 607-21

OBJECTIVES: To determine the antioxidant activities of nonsteroidal anti-inflammatory drugs (NSAIDS), we examined by chemiluminescence (CL) and electron spin resonance (ESR) their scavenging properties ... [more ▼]

OBJECTIVES: To determine the antioxidant activities of nonsteroidal anti-inflammatory drugs (NSAIDS), we examined by chemiluminescence (CL) and electron spin resonance (ESR) their scavenging properties towards lipid peroxides, hypochlorous acid and peroxynitrite. METHODS: The antioxidant properties of nimesulide (NIM), 4-hydroxynimesulide (4-HONIM), aceclofenac (ACLO), 4-hydroxyaceclofenac (4-HOA-CLO), diclofenac (DICLO) and indomethacin (INDO) were tested on four different reactive oxygen species (ROS) generating systems: (I) phorbol-myristate acetate (PMA)-activated neutrophils, (II) Fe2+/ascorbate-induced lipid peroxidation, (III) HOCl-induced light emission, (IV) the kinetics of ONOO- decomposition followed by spectrophotometry. ROS production was monitored by luminol-enhanced CL or by ESR using two different spin traps. RESULTS: At 10 microM, ACLO, NIM, 4-HONIM, 4-HOA-CLO, and DICLO decreased luminol-enhanced CL generated by PMA-activated neutrophils. Inversely, INDO increased the luminol enhanced CL. Interestingly, hydroxylated metabolites were more potent antioxidants than the parent drugs. Furthermore, all drugs tested, excepted ACLO, lowered lipid peroxidation induced by Fe2+/ascorbate system. ACLO and DICLO, even at the highest concentration tested (100 microM), did not significantly lower HOCl induced CL, whereas the other drugs were potent scavengers. Finally, all the NSAIDS accelerated decomposition of ONOO-, suggesting a potential capacity of the molecules to scavenge peroxynitrite. CONCLUSION: The NSAIDs possess variable degrees of antioxidant activities, linked to their ability to react with HOCl, lipid peroxides or ONOO-. These antioxidant activities could offer interesting targeted side-effects in the treatment of joint inflammatory diseases. [less ▲]

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See detailIn vitro study of the influence of temperature, pH and Aw on the growth rate of trichoderma aspergillum
Begoude, B. A. D.; Lahlali, R.; Friel, D. et al

in Bulletin OILB/SROP = IOBC/WPRS Bulletin (2007), 30

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See detailIn vitro study of the interactions between bovine herpesvirus 4 and the bovine host cells
Vanderplasschen, Alain ULg

in Bulletin et Mémoires de l'Académie Royale de Médecine de Belgique (1999), 154

This work was devoted to the study of the interactions between bovine herpesvirus 4 (BHV-4) and bovine cells in vitro. It led to the discovery of two interesting properties of BVH-4 replication cycle ... [more ▼]

This work was devoted to the study of the interactions between bovine herpesvirus 4 (BHV-4) and bovine cells in vitro. It led to the discovery of two interesting properties of BVH-4 replication cycle: first, the cellular receptor heparan sulfate was proven to mediate BVH-4 binding to target cells. This is the first description of the implication of heparan sulfate in the binding process of a gammaherpesvirus. Second, using synchronised cells, the replication of BVH-4 DNA was proven to be dependent on the S phase of the cell cycle. This dependence could explain some properties of BVH-4 infection in vitro and could play an important role in the biology of the infection in vivo. Finally, in order to produce monoclonal antibodies against BVH-4 IE1 and IE2 proteins, the genes coding for these proteins were cloned and expressed in prokaryotic cells. [less ▲]

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See detailIn vitro study on the effects of aceclofenac on proteoglycan and type II collagen production by human chondrocytes
Henrotin, Yves ULg; Labasse, A; Degroote, D et al

in Osteoarthritis and Cartilage (1997), 5

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See detailIn vitro study toward the endocrine activity and the genotoxic potential of migration products from plastic baby bottles
Simon, Coraline ULg; Onghena, M.; Covaci, A. et al

in Organohalogen Compounds (2014), 76

Bisphenol A (BPA) is documented in several studies to be a well-known source of food contamination. Recently, the European Union took a series of measures, including a ban for the import and sale of ... [more ▼]

Bisphenol A (BPA) is documented in several studies to be a well-known source of food contamination. Recently, the European Union took a series of measures, including a ban for the import and sale of polycarbonate (PC) baby bottles to reduce BPA exposure of infants. Plastic alternatives to PC, which have massively appeared on the Belgian market, include polypropylene (PP), silicone, polyamide (PA) and polyethersulfone (PES). In a recent opinion (No. 8697, 11.03.2010), the Belgian Superior Health Council issued its concern regarding the alternatives to PC currently used. Furthermore, they asked to investigate the possible risks associated with the use of these alternatives. In this study, a screening towards the endocrine activity of chemicals migrating from PC alternatives, identified by the group of Simoneau, was performed by using different reporter gene assays. Furthermore, the genotoxic potential of these compounds was also assessed with the Vitotox assay, an indicator test for DNA damage. The aim of the screening was to select the substances that may present a risk for human health and thus require further characterization. [less ▲]

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See detailIn vitro study toward the endocrine activity and the genotoxic potential of migration products from plastic baby bottles
Simon, Coraline ULg; Onghena, M.; Covaci, A. et al

Poster (2014, November 20)

Bisphenol A (BPA) is used since 1960 as a primary raw material for the production of polycarbonate (PC) plastic and epoxy resin, widely used in a variety of common products including digital media (e.g ... [more ▼]

Bisphenol A (BPA) is used since 1960 as a primary raw material for the production of polycarbonate (PC) plastic and epoxy resin, widely used in a variety of common products including digital media (e.g., CDs, DVDs), electrical and electronic equipment, automobiles, sports safety equipment, reusable food and drink containers , as well as baby bottle. During the last decades, in several studies, the migration of BPA is documented to be a well-known source of food contamination. Some studies have shown that BPA, which is able to mimic the action of and hormone and to disrupt normal endocrine function, may be associated to several health problems and diseases. Recently, the European Union took a series of measures, including a ban for the import and sale PC baby bottles to reduce BPA exposure to infants. Plastic alternatives to polycarbonate have massively appeared on Belgium market. Although there are several studies on BPA migration from polycarbonate into foodstuff under a variety of conditions, there is a small amount of information about consequences on human health of the potential exposure to chemicals migrating from PC alternatives, including bottles commonly labelled “free BPA”. In a recent opinion (No. 8697, 11.03.2010), the Belgium Superior Health Council's issued its concern regarding the currently used alternatives to PC. Furthermore, they asked to investigate the possible risks associated with the use of these alternatives. In order to evaluate the safety of these alternatives, the genotoxicity and the activity on several receptors of chemical compounds migrating from PC alternatives, identified by Simoneau & al, 2012 , were evaluated using reporter gene assays. Receptor agonistic and antagonistic activities of 39 pure compounds were measured. After the first screening, certain substances clearly showed an activity on several receptors such as BPA, and Bisphenol S, while only a few substances showed no reaction on the different receptor. None of the 39 components was genotoxic as identified in the Vitotox test. However, further experiments will be performed to characterize their activity and confirm the result for the genotoxicity. [less ▲]

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See detailIn vitro survival of vitrified goat embryos: comparison of two vitrification methods
Guignot, F.; Baril, G.; Pougnard, J. L. et al

in Proceedings: 17e Réunion A.E.T.E. (2001)

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See detailIn vitro susceptibility of Clostridium perfringens isolated from farm animals to growth-enhancing antibiotics.
Devriese, L. A.; Daube, Georges ULg; Hommez, J. et al

in Journal of Applied Bacteriology (1993), 75

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See detailIn Vitro Susceptibility Testing of Aspergillus fumigatus against Posaconazole: Comparison of NCCLS M38-P and E-Test Methods
Hayette, Marie-Pierre ULg; Amadore, Agatha; Seidel, Laurence ULg et al

Poster (2002, September)

Posaconazole is a second-generation triazole and structural analogue of itraconazole. This drug has fungicidal activity against yeasts and filamentous fungi. The aim of our study was to evaluate E-test ... [more ▼]

Posaconazole is a second-generation triazole and structural analogue of itraconazole. This drug has fungicidal activity against yeasts and filamentous fungi. The aim of our study was to evaluate E-test method for in vitro susceptibility testing of Aspergillus fumigatus isolates against posaconazole. METHODS: A total of 121 isolates of A. fumigatus were selected as follows: 106 clinical strains from colonized patients, 18 from patients with invasive aspergillosis and 7 environmental isolates. Their in vitro susceptibility was evaluated by E-test (Abbiodisk, Sweden) and compared with NCCLS microdilution reference method (M38-P). Both tests were performed with RPMI 1640 medium at 35 degrees C. MIC values were read after 24h (MIC-24h) and 48h (MIC-48h) incubation time by E-test method. Two MIC endpoints were determined by NCCLS method: 1.no visible reduction of growth (MIC-0); 2. 50% reduction (or more) of growth (MIC-2). Three A. fumigatus reference strains (IHEM 5734, 6149 and 13935) were included as control. RESULTS: Geometric mean MICs (microg/ml) were respectively 0.02 for E-test at 24h and 0.029 at 48h. MIC-0 and MIC-2 values were respectively 0.19 and 0.018 microg/ml. One correlation between both methods was observed for MICs-24h and MICs-2s (p<.05). However, there was no significant difference according to origin of isolates (p<.05). CONCLUSIONS: 1. This study assessed the potent role of posaconazole against A. fumigatus isolates with very low MICs. 2. MIC values were not predictive of pathogenicity. 3. E-test method by reading after 24h-incubation time could easily replace the time-consuming NCCLS M38-P reference method. [less ▲]

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See detailIn Vitro Synergy of Polymyxins and Carbapenems: Systematic Review and Meta Analysis.
Zusman, Oren; Avni, Tomer; Leibovici, Leonard et al

in Antimicrobial Agents and Chemotherapy (2013), 57(10), 5104-51011

ObjectivesTo examine the evidence on in-vitro synergy of polymyxin-carbapenem combination therapy against Gram-negative bacteria (GNB)MethodsSystematic review and meta-analysis. All studies examining in ... [more ▼]

ObjectivesTo examine the evidence on in-vitro synergy of polymyxin-carbapenem combination therapy against Gram-negative bacteria (GNB)MethodsSystematic review and meta-analysis. All studies examining in-vitro interactions of antibiotic combinations consisting of any carbapenem with colistin or polymyxin B against any GNB. A broad search was conducted with no language, date or publication status restrictions. Synergy rates, defined as fractional inhibitory concentration index </=0.5 or >2log colony forming unit reduction, were pooled separately for time-kill, checkerboard, and E-test in a mixed-effects meta-analysis of rates. We examined whether synergy rate depended on testing method, type of antibiotic, bacteria and resistance to carbapenems. Pooled rates with 95% confidence intervals are shown.Results39 published studies and 15 conference proceeding were included, reporting on 246 different tests on 1054 bacterial isolates. In time-kill studies, combination therapy showed synergy rates of 77% (95% CI 64-87) for Acinetobacter baumannii, 44% (95% CI 30-59%) for Klebsiella pneumoniae and 50% (95% CI 30-69%) for Pseudomonas aeruginosa with low antagonism rates for all. Doripenem showed high synergy rates for all three bacteria. For A. baumannii, meropenem was more synergistic than imipenem, whereas for P. aeruginosa the opposite was true. Checkerboard and Etest studies generally reported lower synergy rates than time-kill. Use of combination therapy led to less resistance development in-vitro.ConclusionsThe combination of a carbapenem with a polymyxin against GNB, especially A. baumannii, is supported in-vitro by high synergy rates, with low antagonism and less resistance development. These findings should be examined in clinical studies. [less ▲]

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See detailIn vitro tests for evaluation of the hatchability of the eggs of Psoroptes mites following exposure to acaricidal compounds.
Lekimme, Mireille ULg; Mignon, Bernard ULg; Leclipteux, Thierry ULg et al

in Medical and Veterinary Entomology (2006), 20(1), 102-105

Three in vitro assays for the evaluation of the hatchability of the eggs of the mite Psoroptes ovis (Hering) (Acari: Psoroptidae) are described. Preliminary trials showed that hatching occurs at very high ... [more ▼]

Three in vitro assays for the evaluation of the hatchability of the eggs of the mite Psoroptes ovis (Hering) (Acari: Psoroptidae) are described. Preliminary trials showed that hatching occurs at very high rates when eggs are incubated under conditions of high humidity, on a liquid medium and in agarose dishes. These three protocols were compared, taking into account the ease of preparation, follow-up and accuracy of counting. The best protocol was found to be the use of agarose dishes. It was accurate, easy to carry out and reproducible for further evaluation of existing or potentially new compounds against both adults and eggs of Psoroptes spp. The acaricidal properties of phoxim and amitraz were then evaluated against eggs and adults using the three protocols. Results showed that for both drugs, in vitro adulticidal activity was complete, whereas the in vitro ovicidal activity was only partial. Nevertheless, efficacy of amitraz against both adults and eggs was shown to be higher than that of phoxim. [less ▲]

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See detailIn vitro titration of Theileria parva tick derived stabilates
Marcotty, T.; Speybroeck, N.; Berkvens, D. et al

in Parasitology (2004), 128(Part 2), 131-137

Immunization agairist the protozoan Theileria parva by infection and treatment has proved to be very efficient for the Control Of East Coast fever, an acute and often-fatal lymphoproliferative tick-borlic ... [more ▼]

Immunization agairist the protozoan Theileria parva by infection and treatment has proved to be very efficient for the Control Of East Coast fever, an acute and often-fatal lymphoproliferative tick-borlic disease of cattle in Eastern, Central and Southern Africa. The immunizing dose of live T. Parva sporozites used in this method is usually determined by in vitro titration. An alternative in vivo method of quantitification of sporozoites ill whole tick-derived stabilites is proposed. The method consists of incubating serially diluted T. Parva stabilities with boville peripheral blood lymphocytes, the host cell that is infected naturally. Allowing the cultures to incubate undisturbed for the full cultivation period (10 days) reduced the variability amoung replicate titrations. fungal contaminations were avoided by centrifuging stabilates at 400 g prior to the incubation, which did not precipiate sporozoites significantly. Fungistics, Nysatin and Flucytosine did not appear to interfere with the in vitro development of 2 stabilates but their effect on fungal growth was limited. In vitro titration data were compared to in vivo infection data for 2. In vivo titration of T. parva sporozoites should allow more ethicl and efficient research on the preparation and storage of T. Parva tick-derived stabilates. [less ▲]

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