References of "Fillet, Marianne"
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See detailEspectroscopia aplicada
Ziemons, Eric ULg; Frederich, Michel ULg; Fillet, Marianne ULg et al

Scientific conference (2011, May)

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See detailidentification of new parterns of the NOD2 protein
Lecat, Aurore ULg; Di Valentin, Emmanuel ULg; Fillet, Marianne ULg et al

Poster (2011, January 27)

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See detailEvaluation of chlorine containing cellulose-based chiral stationary phases for the LC enantioseparation of basic pharmaceuticals using polar non-aqueous mobile phases.
Dossou, Katina S. S.; CHIAP, Patrice ULg; Servais, Anne-Catherine ULg et al

in Journal of Separation Science (2011), 34(6), 617-22

The discrimination ability of three cellulose-based chiral stationary phases (CSPs) was evaluated towards the enantiomers of basic drugs, using ACN as the main solvent in polar organic mobile phases. The ... [more ▼]

The discrimination ability of three cellulose-based chiral stationary phases (CSPs) was evaluated towards the enantiomers of basic drugs, using ACN as the main solvent in polar organic mobile phases. The study was focused on CSPs containing cellulose tris(3-chloro-4-methylphenylcarbamate) (3-Cl-4-MePC), cellulose tris(4-chloro-3-methylphenylcarbamate) (4-Cl-3-MePC) or cellulose tris(3,5-dichlorophenylcarbamate) (3,5-diClPC) as the chiral selector. The behaviour of these CSPs was studied systematically in order to investigate the influence of the presence and position of the chlorine substituents on the phenylcarbamate moieties on the retention and resolution of the enantiomers. The evaluation was made with three different generic mobile phases, namely ACN/0.1%DEA/0.1% TFA (DEA, diethylamine), ACN/0.1%DEA/0.2% FA and ACN/0.1%DEA/0.2%AcA, deduced from the previous study. The nature of the acidic additive and of the chiral selector was found to be particularly important for the retention and enantioresolution of these basic compounds. High-resolution values could be obtained for most studied enantiomers with these CSPs, clearly demonstrating the interest of using them in combination with polar organic mobile phases. However, significant differences in enantioresolution between the CSPs have been observed for many compounds, indicating that these phases seem to be quite complementary. [less ▲]

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See detailDevelopment and validation of a LC method for the enantiomeric purity determination of S-ropivacaine in a pharmaceutical formulation using a recently commercialized cellulose-based chiral stationary phase and polar non-aqueous mobile phase.
Dossou, Katina Sourou Sylvestre; CHIAP, Patrice ULg; Servais, Anne-Catherine ULg et al

in Journal of Pharmaceutical & Biomedical Analysis (2011), 54(4), 687-93

Ropivacaine is the first enantiomerically pure long-acting local anaesthetic used for surgical anaesthesia and post-operative pain relief. A liquid chromatographic (LC) method using acetonitrile as the ... [more ▼]

Ropivacaine is the first enantiomerically pure long-acting local anaesthetic used for surgical anaesthesia and post-operative pain relief. A liquid chromatographic (LC) method using acetonitrile as the main solvent and cellulose tris(4-chloro-3-methylphenylcarbamate) coated on silica as chiral stationary phase was successfully developed and applied for the enantiomeric purity determination of S-ropivacaine in a pharmaceutical formulation (Naropin((R))). The key role played by the acidic additive (trifluoroacetic acid or formic acid) in the enantioseparation of basic drugs in these LC systems was demonstrated by the reversal of ropivacaine enantiomers elution order observed when both acids were compared. In order to elute the enantiomeric impurity (R-ropivacaine) before S-ropivacaine, formic acid (FA) was selected. The temperature and the percentages of acidic additive and hexane in the mobile phase were found to significantly influence the retention and resolution of these enantiomers. The optimized mobile phase consisted of ACN/0.1% DEA/0.2% FA/5% hexane (v/v/v/v). The temperature was set at 35 degrees C to avoid the interference from a peak system related to the presence of water in the sample on ropivacaine enantiomers. The LC method was then fully validated applying the strategy based on total measurement error and accuracy profiles. The accuracy profile obtained by linear regression after square root transformation was selected, the acceptance limits being settled at +/-10% for the intended use of this analytical method. The relative bias was lower than 1.5%, while the RSD values for repeatability and intermediate precision were both below 1.0%. The limit of detection (LOD) and the limit of quantification (LOQ) were found to be about 0.2 and 1.0 mug/mL, respectively, corresponding to 0.02 and 0.1% of the enantiomeric impurity in S-ropivacaine. [less ▲]

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See detailDiscovery and biochemical characterisation of four novel biomarkers for osteoarthritis.
DE SENY, Dominique ULg; Sharif, Mohammed; Fillet, Marianne ULg et al

in Annals of the Rheumatic Diseases (2011), 70(6), 1144-52

OBJECTIVE: Knee osteoarthritis (OA) is a heterogeneous, complex joint pathology of unknown aetiology. Biomarkers have been widely used to investigate OA but currently available biomarkers lack specificity ... [more ▼]

OBJECTIVE: Knee osteoarthritis (OA) is a heterogeneous, complex joint pathology of unknown aetiology. Biomarkers have been widely used to investigate OA but currently available biomarkers lack specificity and sensitivity. Therefore, novel biomarkers are needed to better understand the pathophysiological processes of OA initiation and progression. METHODS: Surface enhanced laser desorption/ionisation-time of flight-mass spectrometry proteomic technique was used to analyse protein expression levels in 284 serum samples from patients with knee OA classified according to Kellgren and Lawrence (K&L) score (0-4). OA serum samples were also compared to serum samples provided by healthy individuals (negative control subjects; NC; n=36) and rheumatoid arthritis (RA) patients (n=25). Proteins that gave similar signal in all K&L groups of OA patients were ignored, whereas proteins with increased or decreased levels of expression were selected for further studies. RESULTS: Two proteins were found to be expressed at higher levels in sera of OA patients at all four K&L scores compared to NC and RA, and were identified as V65 vitronectin fragment and C3fpeptide. Of the two remaining proteins, one showed increased expression (unknown protein at m/z of 3762) and the other (identified as connective tissue-activating peptide III protein) was decreased in K&L scores >2 subsets compared to NC, RA and K&L scores 0 or 1 subsets. CONCLUSION: The authors detected four unexpected biomarkers (V65 vitronectin fragment, C3f peptide, CTAP-III and m/z 3762 protein) that could be relevant in the pathophysiological process of OA as having significant correlation with parameters reflecting local inflammation and bone remodelling, as well as decrease in cartilage turnover. [less ▲]

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See detailPotential Therapeutic Target Discovery by 2D-DIGE Proteomic Analysis in Mouse Models of Asthma
QUESADA CALVO, Florence ULg; Fillet, Marianne ULg; Renaut, Jenny et al

in Journal of Proteome Research (2011), 10(9), 4291-4301

As asthma physiopathology is complex and not fully understood to date; it is expected that new key mediators are still to be unveiled in this disease. The main objective of this study was to discover ... [more ▼]

As asthma physiopathology is complex and not fully understood to date; it is expected that new key mediators are still to be unveiled in this disease. The main objective of this study was to discover potential new target proteins with a molecular weight >20 kDa by using two-dimensional differential in-gel electrophoresis (2D-DIGE) on lung parenchyma extracts from control or allergen-exposed mice (ovalbumin). Two different mouse models leading to the development of acute airway inflammation (5 days allergen exposure) and airway remodeling (10 weeks allergen exposure) were used. This experimental setting allowed the discrimination of 33 protein spots in the acute inflammation model and 31 spots in the remodeling model displaying a differential expression. Several proteins were then identified by MALDI-TOF/TOF MS. Among those differentially expressed proteins, PDIA6, GRP78, Annexin A6, hnRPA3, and Enolase display an increased expression in lung parenchyma from mice exposed to allergen for 5 days. Conversely, Apolipoprotein A1 was shown to be decreased after allergen exposure in the same model. Analysis on lung parenchyma of mice exposed to allergens for 10 weeks showed decreased calreticulin levels. Changes in the levels of those different mediators were confirmed by Western blot and immunohistochemical analysis. Interestingly, alveolar macrophages isolated from lungs in the acute inflammation model displayed enhanced levels of GRP78. Moreover, intratracheal instillation of anti-GRP78 siRNA in allergen-exposed animals led to a decrease in eosinophilic inflammation and bronchial hyperresponsiveness. This study unveils new mediators of potential importance that are up- and down-regulated in asthma. Among up-regulated mediators, GRP-78 appears as a potential new therapeutic target worthy of further investigations. [less ▲]

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See detailDetermination of enantiomeric purity of S-amlodipine by chiral LC with emphasis on reversal of enantiomer elution order.
Dossou, Katina ULg; Edorh, P. A.; Chiap, Patrice ULg et al

in Journal of Separation Science (2011), 34

An LC method was developed and prevalidated for the enantiomeric purity determination of S-amlodipine in polar organic solvent chromatography using a chlorine-containing cellulose-based chiral stationary ... [more ▼]

An LC method was developed and prevalidated for the enantiomeric purity determination of S-amlodipine in polar organic solvent chromatography using a chlorine-containing cellulose-based chiral stationary phase (CSP). The concentration of formic acid (FA) (0.01-0.2%) in the mobile phase containing acetonitrile as the main solvent was found to influence the elution order of amlodipine enantiomers as well as the enantioresolution. A reversal of the enantiomer elution order of amlodipine was only observed with chiral stationary phases with both electron-withdrawing (chloro) and electron-donating groups (methyl) on the phenyl moieties of the chiral selector, namely cellulose tris(3-chloro-4-methylphenylcarbamate) and cellulose tris(4-chloro-3-methylphenylcarbamate). The highest enantioresolution (R(s) : 4.1) value was obtained at the lowest FA concentration (0.01%) using cellulose tris(4-chloro-3-methylphenylcarbamate) as the chiral selector and the enantiomeric impurity, R-amlodipine, eluted first under these conditions. Therefore, the mobile phase selected for the prevalidation of the method consisted of ACN/0.1% DEA/0.01% FA and the temperature was set at 25 degrees C. The method was prevalidated by means of the strategy based on the total measurement error and the accuracy profile. The method was found to be selective and the limit of quantification was found to be about 0.05% for R-amlodipine, while the limit of detection was close to 0.02%. [less ▲]

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See detailPre-study and in-study validation of a SPE-LC-MS-MS method for the determination of 5-S-cysteinyldopa, a melanoma biomarker, in human plasma.
Martin, Gaelle; Mansion, François ULg; Houbart, Virginie ULg et al

in Talanta (2011), 84(2), 280-6

The incidence of malignant melanoma has increased over the past decades, particularly in Caucasian population. This disease presents defavourable prognosis in terms of survey, especially when detection ... [more ▼]

The incidence of malignant melanoma has increased over the past decades, particularly in Caucasian population. This disease presents defavourable prognosis in terms of survey, especially when detection occurs at the metastatic phase. Reliable analytical methods for biomarker determination are thus an interesting tool in pathology detection and follow-up. In this context, a method using SPE-LC-ESI-MS-MS for the determination of 5-S-cysteinyldopa (5-SCD) in human plasma was optimized. The presence of matrix effect was investigated in details while 5-SCD stability was studied according to FDA requirements for the validation of bioanalytical methods. Pre-study and in-study validations of the entire method were then successfully performed by applying the approach based on total measurement error and accuracy profiles over a concentration ranges from 1.6 to 200 ng/ml. Good results with respect to accuracy, trueness and precision were obtained. The maximum risk of observing future measurements falling outside the acceptance limits during routine analysis was also estimated. [less ▲]

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See detailGeneric systems for the enantioseparation of basic drugs in NACE using single-isomer anionic CDs
Rousseau, Anne ULg; Gillotin, Florian; Chiap, Patrice ULg et al

in Journal of Pharmaceutical & Biomedical Analysis (2011), 54

The enantioseparation of ten basic drugs was evaluated in NACE systems using heptakis(2-O-methyl-3-O-acetyl-6-O-sulfo)-β-CD (HMAS-β-CD). For this purpose, a D-optimal design with 21 experimental points ... [more ▼]

The enantioseparation of ten basic drugs was evaluated in NACE systems using heptakis(2-O-methyl-3-O-acetyl-6-O-sulfo)-β-CD (HMAS-β-CD). For this purpose, a D-optimal design with 21 experimental points was applied. Four antifungal agents (econazole, isoconazole, miconazole, sulconazole), three local anesthetics (bupivacaine, mepivacaine and prilocaine), two sympathomimetics (salbutamol and terbutaline) and one β-blocker (carvedilol) were selected as basic model analytes. The influence on the enantiomeric resolution of anionic CD and BGE anion concentrations as well as the BGE anion nature was investigated. For all studied analytes, the enantiomeric resolution was shown to be significantly influenced by the CD concentration. Based on the observed results, a generic NACE system was recommended, namely 20 mM HMAS-β-CD and 10 mM ammonium camphorSO3- in methanol acidified with 0.75 M formic acid. Moreover, this NACE system was compared to previous conditions with heptakis(2,3-di-O-methyl-6-O-sulfo)-β-CD (HDMS-β-CD) or heptakis(2,3-di-O-acetyl-6-O-sulfo)-β-CD (HDAS-β-CD). Finally, two generic systems using either HDAS-β-CD or HMAS-β-CD were proposed and evaluated for the enantioseparation of ketamine and norketamine after incubation of ketamine in phenobarbital-induced male rat liver microsomes systems. [less ▲]

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See detailA DESIGN SPACE APPROACH TO DEVELOP A GENERIC CE METHOD FOR THE SEPARATION OF 19 ANTIMALARIAL DRUGS
Lamalle, Caroline ULg; Marini Djang'Eing'A, Roland ULg; Debrus, Benjamin ULg et al

Poster (2010, September)

This project consists in analysing different molecules used against malaria by capillary electrophoresis (CE). As qualitative and quantitative counterfeit is largely present in Africa, it is important to ... [more ▼]

This project consists in analysing different molecules used against malaria by capillary electrophoresis (CE). As qualitative and quantitative counterfeit is largely present in Africa, it is important to develop a simple method which can control the conformity of medicines from African market. For the moment no CE method has been developed to analyse simultaneously the most common antipaludics; but it can be very useful for the control of unknown tablets. The method development was performed on 4 preservatives (methylparaben, propylparaben, butylhydroxyanisol and butylhydroxytoluen) and 16 antipaludics (artesunate, artemether, amodiaquine hydrochloride, chloroquine diphosphate, piperaquine, primaquine diphosphate, quinine hydrochloride, cinchonine, mefloquine hydrochloride, lumefantrine, halofantrine, sulfadoxine, sulfalen, atovaquone, proguanil hydrochloride, pyrimethamine). Micellar electrokinetic chromatography (MEKC) was chosen because of the presence of neutral and charged compounds in the studied mixture. The first step of method development was to screen CE experimental conditions to select the most crucial factors. Several conditions were tested with antipaludics diluted in 100% methanol and in 70:30 (v/v) methanol/water in which resolution was better. Four parameters as well as their investigation domain were then chosen: pH (5-10), SDS concentration (20-90nM), acetonitrile proportion (10-40%) and temperature (20-35°C). Then, in order to predict the best condition for the method, an experimental design methodology using a face-centered central composite design (CCD) was realised. Twenty five experiments were defined by CCD. Molecules were separated in four groups and each molecule could be found in two groups. Four samples containing 10 molecules were therefore injected to reduce the number of runs. All the results were analysed and allowed the prediction of optimal conditions in terms of analyte separation. Finally, the condition giving the best separation was tested to verify the prediction. [less ▲]

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See detailThe repressing function of the oncoprotein BCL-3 requires CtBP while its polyubiquitination and degradation involve the E3 ligase TBLR1
Keutgens, Aurore ULg; Shostak, Kateryna ULg; Close, Pierre ULg et al

in Molecular & Cellular Biology (2010), 30

The nuclear and oncogenic BCL-3 protein activates or represses gene transcription when bound to NF-kB proteins p50 and p52, yet the molecules that specifically interact with BCL-3 and drive BCL-3-mediated ... [more ▼]

The nuclear and oncogenic BCL-3 protein activates or represses gene transcription when bound to NF-kB proteins p50 and p52, yet the molecules that specifically interact with BCL-3 and drive BCL-3-mediated effects on gene expression remain largely uncharacterized. Moreover, GSK3-mediated phosphorylation of BCL-3 triggers its degradation through the proteasome, but the proteins involved in this degradative pathway are poorly characterized. Biochemical purification of interacting partners of BCL-3 led to the identification of CtBP as a molecule required for the ability of BCL-3 to repress gene transcription. CtBP is also required for the oncogenic potential of BCL-3 and for its ability to inhibit UV-mediated cell apoptosis in keratinocytes. We also defined the E3 ligase TBLR1 as a protein involved in BCL-3 degradation through a GSK3-independent pathway. Thus, our data demonstrate that the LSD1/CtBP complex is required for the repressing abilities of an oncogenic IkB protein, and they establish a functional link between the E3 ligase TBLR1 and NF-kB. [less ▲]

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See detailAssessment of high sensitive troponin T and I immunoassays in patients with acute chest
Le Goff, Caroline ULg; Garweg, Christophe ULg; Laurent, Terry et al

in Clinical Chemistry (2010, July), 56(S6), 127

Introduction: Cardiac troponin I and T are specific markers of myocardial injury that are widely used for the diagnosis of acute coronary syndrome (ACS). In acute chest pain without ST-segment elevation ... [more ▼]

Introduction: Cardiac troponin I and T are specific markers of myocardial injury that are widely used for the diagnosis of acute coronary syndrome (ACS). In acute chest pain without ST-segment elevation, they are used to differentiate unstable angina from non ST-segment elevation myocardial infarction (NSTEMI). Recently, troponin assays with higher analytical sensitivities became available to enable the detection of minor myocardial damage and identify individuals at higher risk for ACS. As a result of its high tissue-specificity, cardiac troponin T and I are cardio-specific, highly sensitive markers for myocardial damage. The aim of this study was to evaluate the new higher sensitive troponin (T and I) in patients with stable angina and acute chest pain without ST-segment elevation. Methods: Sixty subjects (mean age : 65.5± 11 years), were included: 20 healthy controls, 20 patients with stable angina, 9 with unstable angina (troponin-) and 18 patients with NSTEMI myocardial infarction (troponin+). The protocol was approved by the ethic committee of the University of Liège (Belgium). High sensitive troponin T (hsTnT) determination was realized on heparin plasma by electrochemiluminescence immunoassay on Modular E (Roche Diagnostic). Troponin I II (TnI II) is a chemiluminescent microparticle immunoassay for the quantitative determination of cardiac troponin-I in heparine plasma on the ARCHITECT i System (Abbott Diagnostic). The lower detection limit of these assays was 0.005μg/L for hsTnT and 0.01μg/L for TnI II. Stastistical analysis was performed using t test. P value <0.05 was considered significant. Results: HsTNT levels were 0.003(0.003, 0.004) [median baseline (1st, 3rd quartile)]ng/ml in controls, 0.0075 (0.00475, 0.014) ng/ml in stable angina, 0.011(0.006, 0.012) ng/ml in unstable angina and 0.3715 (0.1795, 1.00725) ng/ml in NSTEMI ACS. TnI II levels were 0 (0, 0.001) ng/ml in controls and in patients with stable angina, 0.07 (0.005, 0.014) ng/ml in unstable angina and 1.4475 (0.0407, 2.656) ng/ml in NSTEMI. HsTNT and TnI II levels were significantly increased in NSTEMI as compared to control subjects, patients with stable and unstable angina. TnI II levels were also increased in unstable angina as compared to controls. Conclusion: In our population, TnI II was more sensitive than hsTNT to detect minor myocardial damage in patients with unstable angina as compared to controls. Therefore, future studies will have to determine whether TnI II might contribute to better risk stratification and treatment strategy in this group of patients. [less ▲]

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See detailDoes echocardiographic stress test induced release of hsTnT and TnI II?
Le Goff, Caroline ULg; Laurent, Terry; Garweg, Christophe ULg et al

in Clinical Chemistry (2010, July), 56(S6), 128

Background: Cardiac troponins (cTn) are considered as the best biomarkers for detection of myocardial cell injury. In this study, cTnT and cTnI were measured by new commercially available high-sensitive ... [more ▼]

Background: Cardiac troponins (cTn) are considered as the best biomarkers for detection of myocardial cell injury. In this study, cTnT and cTnI were measured by new commercially available high-sensitive methods in patients undergoing brief exercise- or pharmacologicinduced stress. Our aim was to compare cTnT and cTnI levels before and after the stress tests, in the patients with or without reversible ischemia. Materials and Methods: Fifty patients (28 men and 22 women) underwent an echographic stress test (ST) for suspected ischemic heart disease. Of these 50 patients, 28 received pharmacological ST (dobutamine injection) and 22 dynamic ST (bicycle exercise). The patients were subdivided into two groups according to the presence or absence of documented transient reversible ischemia: 14 with reversible ischemia ( mean age: 67.71±9.66 y) and 36 without ischemia ( mean age: 63.17±11.72 y). In all patients, cTnT and cTnI concentrations were measured by high sensitive methods (hsTnT, Roche Diagnostics and TnI II, Abbott Diagnostics) on heparin plasma immediately before (T0) and after ST (T1).The lower detection limit of these assays was 0.005μg/L for hsTnT and 0.01μg/L for TnI II. The protocol was approved by the ethics committee of the University of Liège (Belgium). All patients gave informed consent. All statistical analyses were performed using Medcalc version 8.1 for Windows. P value <0.05 was regarded as statistically significant. Results: There was no significant difference between hsTnT concentrations at T0 and T1, neither in the whole patient group, nor in the subgroups of subjects who received pharmacological ST or dynamic ST. The same was true for TnI II. Although there was no change in hsTnT levels during test in ischemic and in non ischemic patients, the latter tend to demonstrate higher median T0 levels (25th, 75th percentiles) than the others [0.011 (0.007, 0.029) vs 0.007 (0.0047, 0.1125) ng/ml, p=0.09]. They also showed higher median T1 levels [0.014 (0.065, 0.03) vs 0.007 (0.003, 0.0102) ng/ml, p=0.08]. Higher TnI II levels were also recorded in ischemic patients as compared to non ischemic patients at T0[ 0.014 (0.0072; 0.0265) vs 0.005 (0.003; 0.01) ng/ml, p=0.08] and T1[ 0.013 (0.0085- 0.03) vs 0.006 (0.0035-0.008) ng/ml, p=0.08]. Also, TnI II levels did not change during test in both subgroups. Conclusions: Measurement of cardiac troponins by high sensitive methods did not allow to detect significant release of biomarkers from the heart during exercise-or pharmacologic-induced ST, even in patients who demonstrated reversible myocardial ischemia. The type of test – pharmacological or dynamic - was without effect. The patients with induced transient ischemia had however higher troponin T and I levels at baseline, this difference remaining during test. [less ▲]

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