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See detailImprovement of a stability-indicating method by Quality-by-Design versus Quality-by-Testing: A case of a learning process
Hubert, Cédric ULg; Lebrun, Pierre ULg; Houari, Sabah ULg et al

in Journal of Pharmaceutical & Biomedical Analysis (2014), 88

The understanding of the method is a major concern when developing a stability-indicating method and even more so when dealing with impurity assays from complex matrices. In the presented case study, a ... [more ▼]

The understanding of the method is a major concern when developing a stability-indicating method and even more so when dealing with impurity assays from complex matrices. In the presented case study, a Quality-by-Design approach was applied in order to optimize a routinely used method. An analytical issue occurring at the last stage of a long-term stability study involving unexpected impurities perturbing the monitoring of characterized impurities needed to be resolved. A compliant Quality-by-Design (QbD) methodology based on a Design of Experiments (DoE) approach was evaluated within the framework of a Liquid Chromatography (LC) method. This approach allows the investigation of Critical Process Parameters (CPPs), which have an impact on Critical Quality Attributes (CQAs) and, consequently, on LC selectivity. Using polynomial regression response modeling as well as Monte Carlo simulations for error propagation, Design Space (DS) was computed in order to determine robust working conditions for the developed stability-indicating method. This QbD compliant development was conducted in two phases allowing the use of the Design Space knowledge acquired during the first phase to define the experimental domain of the second phase, which constitutes a learning process. The selected working condition was then fully validated using accuracy profiles based on statistical tolerance intervals in order to evaluate the reliability of the results generated by this LC/ESI-MS stability-indicating method. A comparison was made between the traditional Quality-by-Testing (QbT) approach and the QbD strategy, highlighting the benefit of this QbD strategy in the case of an unexpected impurities issue. On this basis, the advantages of a systematic use of the QbD methodology were discussed. [less ▲]

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See detailDevelopment of a liquid chromatographic method for thesimultaneous quantification of curcumin, -arteether,tetrahydrocurcumin and dihydroartemisinin. Application to lipid-based formulations
Memvanga Bondo, Patrick; Mbinze Kindenge, Jérémie ULg; Rozet, Eric ULg et al

in Journal of Pharmaceutical & Biomedical Analysis (2014), 88(-), 447-456

A liquid chromatographic method was developed for the simultaneous separation of curcumin, B-arteether, tetrahydrocurcumin and dihydroartemisinin based on the design of experiments and the design space ... [more ▼]

A liquid chromatographic method was developed for the simultaneous separation of curcumin, B-arteether, tetrahydrocurcumin and dihydroartemisinin based on the design of experiments and the design space methodology. The influence of the percentage of organic modifier, flow rate of the mobile phase and column temperature on the analytes separation was investigated. The optimal chromatographic separation was achieved on a C18 column (125 mm × 4 mm, 5 µm) using an isocratic elution with a mobile phase consisting of methanol:ammonium acetate (pH 4; 10 mM) (80/20, v/v) at a flow rate of 0.45 ml/min and a column temperature of 32.5◦C. This method was then validated for simultaneous quantification of curcumin and B -arteether contained in lipid-based formulations taking into account the B -expectation tolerance interval for the total error measurement. Finally, the suitability of the proposed liquid chromatographic method for routine analysis of curcumin and B -arteether loaded in lipid-based formulations has been proven. [less ▲]

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See detailUnion européenne : passé, présent et futur
Dumoulin, André ULg

Conference given outside the academic context (2014)

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See detailFormer et accompagner les professionnels. Diversité dans les soins et l’éducation des jeunes enfants, l’accent étant mis sur les enfants vulnérables et défavorisés
Pirard, Florence ULg; Van Avermaet, Piet

Conference given outside the academic context (2014)

L’accueil et l’éducation de jeunes enfants en dehors du cadre familial (dans les services autour de la naissance, en milieu d’accueil, à l’école maternelle, dans l’accueil extrascolaire…) nécessitent ... [more ▼]

L’accueil et l’éducation de jeunes enfants en dehors du cadre familial (dans les services autour de la naissance, en milieu d’accueil, à l’école maternelle, dans l’accueil extrascolaire…) nécessitent l’acquisition de compétences professionnelles et de conditions propices à leur développement. L’importance accordée aujourd’hui à une prise en compte des diversités dans l’accueil et l’éducation des jeunes enfants, en particulier celles liées aux situations de précarité, voire de pauvreté, conduit inévitablement à s’interroger sur les compétences professionnelles des divers acteurs opérant dans le secteur (puériculteurs/trices, auxiliaires de l’enfance, accueillant-e-s extrascolaires, enseignant-e-s préscolaires, etc.). [less ▲]

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See detailMetallothioneins pattern during ontogeny of coastal dolphin, Pontoporia blainvillei, from Argentina
Polizzi, P.S.; Romero, M.B.; Chiodi Boudet, L.N. et al

in Marine Pollution Bulletin (2014)

Metallothioneins are signals of metal exposure and widely used in biomonitoring. Franciscana dolphin is an endemic cetacean from the Southwestern Atlantic Ocean, classified as Vulnerable A3d by the IUCN ... [more ▼]

Metallothioneins are signals of metal exposure and widely used in biomonitoring. Franciscana dolphin is an endemic cetacean from the Southwestern Atlantic Ocean, classified as Vulnerable A3d by the IUCN. Metallothionein, copper and zinc in Franciscana were assessed in two geographic groups; one inhabits La Plata River estuary, anthropogenically impacted, and the other inhabits marine coastal ecosystems, with negligible pollution. Despite the environment, hepatic and renal MT concentrations were similar, but there was a declining trend from early to later developmental stages. Metallothionein K/L, Cu and Zn levels corresponded to normal reported ranges. MT was not related with Cd. Fetal concentrations were higher than its mother. These results and the health status of dolphins are suggesting that MT correspond to physiological ranges for the species, and they are closely to homeostasis of Zn and Cu, according to its ontogenetic changes. The information constitutes the first MT information on Franciscana dolphin and can be considered as baseline for the species conservation. [less ▲]

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See detailL'âge d'abattage influence-t-il la qualité de la viande?
De Maeseneire, Coraline ULg; Sindic, Marianne ULg; Colot, Catherine et al

Article for general public (2014)

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See detail« Toward a Visual Sociology of Migration »
Martiniello, Marco ULg

Scientific conference (2014, January 24)

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See detailNanobodies as tools to investigate the mechanism of aggregation of chimeric proteins made by the insertion of polyglutamine stretches into the beta-lactamase BlaP
Pain, Coralie ULg

Doctoral thesis (2014)

Among the neurodegenerative amyloidoses, ten disorders, referred to as polyglutamine (polyQ) diseases and including Huntington's disease and several spinocerebellar ataxias, are associated with ten ... [more ▼]

Among the neurodegenerative amyloidoses, ten disorders, referred to as polyglutamine (polyQ) diseases and including Huntington's disease and several spinocerebellar ataxias, are associated with ten proteins within which a polyQ tract is expanded above a threshold of typically 35-45 glutamine residues. Such expanded polyQ tracts lead to the aggregation of the host protein into amyloid fibrils that accumulate in the nucleus of some populations of neurons; these aggregates or some of their precursors are thought to contribute to neuronal death. So far, no preventive or curative treatment exists for these devastating pathologies. While the expansion of the polyQ tract above the threshold is the determinant factor for aggregation, recent studies suggest that non-polyQ regions of these proteins can play a significant role, either preventative or facilitative, in the aggregation process. The general principles governing the complex interplay between the role of the expanded polyQ tract and the role of the non-polyQ regions in the aggregation process are not well understood yet. In order to develop therapeutic strategies, it is important to better understand this complex interplay. To contribute to this aim, we have engineered chimeric proteins via the insertion of polyQ repeats of various lengths (23, 30, 55 and 79Q) into two sites (197 and 216) of the BlaP beta-lactamase from Bacillus licheniformis 749/C. The properties of these chimeric proteins recapitulate the characteristic features of the disease-associated polyQ proteins, i.e. (i) there is a minimum number of inserted glutamines (threshold) required to trigger the aggregation of the chimeras into amyloid fibrils, and (ii) above the threshold, the longer the polyQ tract, the faster the aggregation. Interestingly, for the same polyQ length, the chimeras with insertions in position 216 have an increased propensity to form amyloid fibrils compared to their counterparts with insertions in position 197. These findings highlight the strong influence of the overall protein context on aggregation triggered by expanded polyQ tracts. This thesis addresses the use of the variable domains of camelid heavy-chain antibodies, referred to as nanobodies or VHHs, as structural and mechanistic probes to better understand the different aggregating properties of the two sets of BlaP-polyQ chimeras (197 and 216). We have also performed limited proteolysis experiments and transglutaminase-mediated reactions on the monomeric form of the BlaP-polyQ chimeras to further investigate the effects of the polyQ insertions on the structure and dynamics of the BlaP moiety, as well as the structure of the polyQ tract itself. From the blood of a llama immunised with BlaP197(Gln)55, we isolated more than 60 VHHs specific to the BlaP-polyQ chimeras. Twenty eight of them were produced, purified and characterised. These VHHs were found to be all specific to the BlaP moiety and could be classified into four different groups recognising distinct epitopes on the surface of BlaP. One representative VHH of each group (i.e. cAb-A3S, cAb-H7S, cAb-F11N and cAb-G10S) was selected as probe to investigate the mechanism of aggregation of the BlaP-polyQ chimeras. The epitope of three of them was determined by X-ray diffraction and/or by NMR spectroscopy. Although they recognise distinct epitopes and exhibit different affinities for BlaP, the binding of the four VHHs significantly slows down the aggregation of all the BlaP-polyQ chimeras investigated (i.e. BlaP197(Gln)55, BlaP197(Gln)79 and BlaP216(Gln)79). The extent of inhibition depends however on the chimera and on the experimental conditions. We show that the inhibition of the aggregation of BlaP197(Gln)55 and BlaP197(Gln)79 upon binding of the four VHHs is correlated with the stabilisation of their native state. In the case of BlaP216(Gln)79, the extent of inhibition could not be only correlated to the stabilisation of its native state; the location of the epitope of the VHH is instead also determinant. This observation demonstrates that the lower thermodynamic stability of BlaP216(Gln)79 is not the unique factor responsible for its increased aggregation propensity. It also further highlights the complexity of the aggregation mechanism of polyQ proteins and the strong influence of the non-polyQ regions on the amyloid fibril formation triggered by the expanded polyQ tract. All together our results suggest that antibodies or antibody fragments raised against the non-polyQ regions of polyQ proteins associated with diseases could constitute a relevant therapeutic strategy. They also further demonstrate the power of nanobodies as probes to get a deeper knowledge of the underlying mechanisms of amyloid fibril formation. The preliminary limited proteolysis and transglutamination experiments obtained suggest that the polyQ tracts are all flexible, except that of 23 glutamines inserted in position 197 of BlaP, which seems to be more rigid than the others. The results obtained confirm that, globally, the structure of BlaP is not significantly modified by the insertions while the 216 chimeras seem more dynamic than the 197 chimeras. [less ▲]

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See detailLosses – Marks&Spencer
Richelle, Isabelle ULg

Conference (2014, January 23)

This conference will explore the history, impact, and potential for further development of the CJEU’s and the EFTA Court’s landmark decisions relating to direct taxation.

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See detailAspetti prosodici e testuali del raccontare: dalla letteratura orale al parlato dei media
Moreno, Paola ULg; Di Salvo, Margherita

Conference (2014, January 23)

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See detailComment on devient camorristes. La transformation d'une société méridionale
De Biase, Marco ULg

Scientific conference (2014, January 23)

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See detailClassification of periodic orbits of two-dimensional homogeneous granular crystals with no pre-compression
Detroux, Thibaut ULg; Starosvetsky, Yuli; Kerschen, Gaëtan ULg et al

in Nonlinear Dynamics (2014), 76(April 2014), 673-696

In the present study we classify the periodic orbits of a squarely packed, uncompressed and undamped, homogeneous granular crystal, assuming that all elastic granules oscillate with the same frequency (i ... [more ▼]

In the present study we classify the periodic orbits of a squarely packed, uncompressed and undamped, homogeneous granular crystal, assuming that all elastic granules oscillate with the same frequency (i.e., under condition of 1:1 resonance); this type of Hamiltonian periodic orbits have been labeled as nonlinear normal modes. To this end we formulate an auxiliary system which consists of a two-dimensional, vibro-impact lattice composed of non-uniform “effective particles” oscillating in an anti-phase fashion. The analysis is based on the idea of balancing linear momentum in both horizontal and vertical directions for separate, groups of particles, whereby each such a group is represented by the single effective particle of the auxiliary system. It is important to emphasize that the auxiliary model can be defined for general finite, squarely packed granular crystals composed of n rows and m columns. The auxiliary model is successful in predicting the total number of such periodic orbits, as well as the amplitude ratios for different periodic regimes including strongly localized ones. In fact this methodology enables one to systematically study the generation of mode localization in these strongly nonlinear, highly degenerate dynamical systems. Good correspondence between the results of the theoretical model and direct numerical simulations is observed. The results presented herein can be further extended to study the intrinsic dynamics of the more complex granular materials, such as heterogeneous two-dimensional and three-dimensional granular crystals and multi-layered structures. [less ▲]

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See detailTailoring the Properties of Metallic Clusters by Ligand Coatings
Fresch, Barbara ULg

Conference (2014, January 23)

Tuning the properties of metallic clusters using different protecting ligand shells is an important step toward the application-orientated design of nanoparticles for nano-electronics and catalysis. An ... [more ▼]

Tuning the properties of metallic clusters using different protecting ligand shells is an important step toward the application-orientated design of nanoparticles for nano-electronics and catalysis. An attractive property of these materials is the ability to engineer ligand shells composed of different molecules that influence the electronic structure of the system due to their chemical interaction with the metal core. Sometimes properties are not simply additive, and cooperative effects emerge so that the presence of more than one ligand type in the ligand shell imparts novel properties to the nanoparticles. We theoretically investigate at the DFT level palladium and gold metal cores interacting with only-thiol, only-phosphine and mixed phosphine-thiol ligand shells. In this talk, we present some examples of how these capping systems affect structural, optical, magnetic and charging properties of metallic clusters. For Au11 and Au13, we show that ligation stabilizes structural isomers characterized by a flake geometry of the metal core, and they show different optical properties than isomers with a compact gold core. The magnetic ground state of small bare Pd13 clusters is a high spin state, nonet or septet depending on the structural isomer. We demonstrate that interaction with thiols and phosphines differently affect structure and magnetic ground state in these systems. The mixed ligand species Pd13(SCH3)6(PH3)6 shows multiple energetically accessible spin states and consequently an unusual thermal behaviour of the average magnetic moment. The charge transfer character of the metal-ligand interaction is studied in small Pd13 and truly nano-sized Pd55 ligand protected metal cores. We discuss cooperative effects in the ligand induced charge redistribution and demonstrate that the electron donor or acceptor character of the ligand molecules affects the energetics of charging processes [less ▲]

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See detailEpigenetics and local adaptation
Hanikenne, Marc ULg

Conference (2014, January 22)

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See detailIntervention à la table ronde "Politics of the Debt", autour du travail d'Etienne Balibar
Pieron, Julien ULg

Scientific conference (2014, January 22)

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See detailContribution à l'étude de la réponse immune au cours d'une infection à Microsporum canis et établissement d'un modèle murin de dermatophytose
Cambier, Ludivine ULg

Doctoral thesis (2014)

Les dermatophytoses (ou « teignes ») sont des mycoses cutanées superficielles provoquées par des champignons filamenteux, appelés dermatophytes, affectant les animaux et l’homme. Microsporum canis est un ... [more ▼]

Les dermatophytoses (ou « teignes ») sont des mycoses cutanées superficielles provoquées par des champignons filamenteux, appelés dermatophytes, affectant les animaux et l’homme. Microsporum canis est un dermatophyte zoophile dont l’hôte naturel est le chat. Il touche principalement les carnivores domestiques et est un agent fréquent de zoonose. Les dermatophytes envahissant les structures kératinisées de l’épiderme, de nombreuses études ont porté sur la caractérisation des protéases sécrétées comme facteurs de virulence potentiels. Cependant, peu de recherches ont été consacrées aux mécanismes impliqués dans la mise en place de la réponse immune anti-dermatophytes. Il est couramment admis que la réponse immune protectrice lors d’une dermatophytose est une réponse à médiation cellulaire impliquant les lymphocytes de type Th1 et caractérisée par une réaction cutanée d’hypersensibilité de type retardé (Delayed Type Hypersentivity, DTH). Un constituant sécrété de M. canis correspondant au surnageant de culture du champignon cultivé sur milieu de Sabouraud liquide, appelé surnageant non induit (SNI), provoque des DTH chez le cobaye immunisé après une primo-infection, ce qui suggère que le SNI contient des antigènes potentiellement intéressants pour le développement d’un vaccin. Le cobaye est le modèle animal le plus utilisé pour étudier l’immunologie des dermatophytoses. Cependant, la quasi absence d’outils génétiques et immunologiques dans cette espèce ne permet pas d’étudier la réponse immune induite lors de dermatophytoses de manière satisfaisante. La mise au point d’un modèle murin de dermatophytose permettrait de pallier tous ces inconvénients. Au cours d’une dermatophytose à M. canis, les premières cellules rencontrées par le champignon lors de l’infection sont les kératinocytes qui, suite à l’activation par des composants fongiques, peuvent attirer les polymorphonucléaires neutrophiles (PMN) sur le site d’infection. Ces leucocytes sont, avec les macrophages, responsables in fine de l’élimination du dermatophyte. Cependant, le rôle des PMN dans les mécanismes liés à l’établissement de l’immunité anti-dermatophytes demeure inconnu. L’objectif général de ce travail était de contribuer à l’étude de la réponse immune au cours d’une dermatophytose à M. canis et de mettre au point un nouveau modèle animal de dermatophytose. Pour l’atteindre, trois études ont été réalisées : (1) l’évaluation du rôle des PMN félins dans la mise en place d’une réponse immune protectrice au cours d’une dermatophytose à M. canis, (2) l’évaluation des propriétés protectrices du SNI de M. canis dans un essai vaccinal en utilisant le cobaye comme modèle expérimental et (3) la mise au point d’un modèle murin de dermatophytose pour l’étude de l’immunité anti-dermatophytes. Résumé 2 Les résultats obtenus dans la première étude montrent que les PMN félins produisent des cytokines pro-inflammatoires telles le tumor necrosis factor (TNF)-α, l’interleukin (IL)-1β et l’IL-8 lorsqu’ils sont stimulés par les arthrospores vivantes du champignon ainsi que par des constituants structuraux, représentés par des arthrospores tuées par la chaleur, et sécrétés comme le SNI. De plus, après stimulation pendant 2 ou 4 heures par les arthrospores vivantes et tuées ainsi que par les protéases sécrétées Sub3 et Sub6, disponibles sous forme recombinante dans notre laboratoire, les taux d’ARNm du toll like receptor (TLR)-2 et du TLR-4 étaient augmentés dans les PMN. Ces résultats suggèrent que les PMN félins jouent un rôle dans l’établissement de la réponse immune anti-M. canis en produisant des cytokines pro-inflammatoires et que les mécanismes responsables de cette activation feraient intervenir les récepteurs TLR-2 et TLR-4. Dans la deuxième étude, les résultats indiquent que l’adjuvant monophosphoryl lipid-A (MPLA), dérivé non toxique du lipopolysaccharide et agoniste du TLR-4 a, à lui seul, un effet protecteur partiel chez les cobayes soumis à une épreuve d’infection par M. canis. En effet, les symptômes cutanés observés chez les cobayes ayant préalablement reçu l’adjuvant, étaient moins importants que ceux développés par les cobayes infectés n’ayant pas reçu l’adjuvant. En revanche, le SNI ne semble pas protéger les cobayes d’une infection expérimentale dans cet essai vaccinal. Les résultats obtenus dans la troisième étude démontrent que notre modèle murin de dermatophytose est valide et reproductible, en utilisant Arthroderma benhamiae et Arthroderma vanbreuseghemii. Ce n’est pas le cas avec M. canis. En effet, 100 % et 80 % des souris infectées respectivement par A. vanbreuseghemii et A. benhamiae ont développé des symptômes cutanés typiques de dermatophytose, à savoir de l’érythème, des squames, des croûtes et de l’alopécie alors que seulement 40 % des souris infectées par M. canis ont développé des signes cliniques, par ailleurs très discrets. Le recrutement important de PMN dans la peau des souris infectées par A. benhamiae et A. vanbreuseghemii et le profil cytokinique (transforming growth factor (TGF)-β, l’IL-1β, l’IL-6 et l’IL-22) généré par l’infection suggèrent l’implication de la voie Th17 dans la mise en place de la réponse immune anti-dermatophytes. En conclusion, l’ensemble de ce travail a contribué à élargir nos connaissances sur la réponse immune au cours d’une dermatophytose à M. canis. De plus, un nouveau modèle murin de dermatophytose à A. benhamiae et A. vanbreuseghemii a été mis au point et est approprié pour l’étude de la réponse immune anti-dermatophytes. [less ▲]

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