References of "Zorzi, Willy"
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See detailA decade of improvements in quantification of gene expression and internal standard selection.
Thellin, Olivier ULg; Elmoualij, Benaïssa ULg; Heinen, Ernst ULg et al

in Biotechnology Advances (2009)

Major improvements have been made in mRNA quantification and internal standard selection over the last decade. Our aim in this paper is to present the main developments that are of interest for practical ... [more ▼]

Major improvements have been made in mRNA quantification and internal standard selection over the last decade. Our aim in this paper is to present the main developments that are of interest for practical laboratory work, contrasting the situation as it is now with the one of ten years ago, and presenting some excellent examples of what can be done today. Specifically, we will mainly discuss Real-Time RT-PCR major improvements that have been performed in the following areas: the most commonly used quantification techniques, the mathematical and software tools created to help researchers in their work on internal standard selection, the availability of detection chemistries and technical information and of commercial tools and services. In addition to mRNA quantification, we will also discuss some aspects of non-coding RNA and protein quantification. In addition to technical improvements, the development of international cooperation and the creation of technical databases are likely to represent a major tool for the future in the standardization of gene expression quantification. [less ▲]

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See detailCreutzfeldt-jakob, Parkinson, lewy body dementia and Alzheimer diseases: from diagnosis to therapy.
Dupiereux-Fettweis, Ingrid ULg; Zorzi, Willy ULg; Quadrio, Isabelle et al

in Central Nervous System Agents in Medicinal Chemistry (2009), 9(1), 2-11

Depositions of proteins in form of amyloid and non-amyloid plaques are common pathogenic signs of more than 20 degenerative diseases affecting the central nervous system or a variety of peripheral tissues ... [more ▼]

Depositions of proteins in form of amyloid and non-amyloid plaques are common pathogenic signs of more than 20 degenerative diseases affecting the central nervous system or a variety of peripheral tissues. Among the neuropathological conditions, Alzheimer's, Parkinson's and the prion diseases, such as Creutzfeldt-Jakob disease (CJD), present ambiguities as regarding their differential diagnosis. At present, their diagnosis must be confirmed by post-mortem examination of the brain. Currently the ante-mortem diagnosis is still based on the integration of multiple data (clinical, paraclinical and biological analyses) because no unique marker exists for such diseases. The detection of specific biomarkers would be useful to develop a differential diagnostic, distinguishing not only different neurodegenerative diseases but also the disease from the non-pathological effects of aging. Several neurodegenerative biomarkers are present at very low levels during the early stages of the disease development and their ultra-low detection is needed for early diagnosis, which should permit more effective therapeutic interventions, before the disease concerned can progress to a stage where considerable damage to the brain has already occurred. In the case of prion diseases, there are concerns regarding not only patient care, but the wider community too, with regard to the risk of transmission of prions, especially during blood transfusion, for which, four cases of variant CJD infection associated with transfusion of non-leukocyte-depleted blood components have been confirmed. Therefore the development of techniques with high sensitivity and specificity represent the major challenge in the field of the protein misfolding diseases. In this paper we review the current analytical and/or biochemical diagnostic technologies used mainly in prion, but also in Alzheimer and Parkinson diseases and emphasizing work on the protein detection as a surrogates and specific biomarker in the body fluid of patients (urine, CSF and blood). This review highlights the urgency of the development of early and sensitive diagnostics in terms of therapeutic challenge. [less ▲]

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See detailImmuno PCR : a new tool for the ultra-sensitive detection of biomolecules
Zorzi, Willy ULg; Elmoualij, Benaïssa ULg

Conference (2008, December 11)

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See detailProtective effect of prion protein via the N-terminal region in mediating a protective effect on paraquat-induced oxidative injury in neuronal cells.
Dupiereux-Fettweis, Ingrid ULg; Falisse-Poirier, Nandini; Zorzi, Willy ULg et al

in Journal of Neuroscience Research (2008), 86(3), 653-9

Transmissible spongiform encephalopathies are a group of neurodegenerative disorders caused by a posttranslational, conformational change in the cellular isoform of the prion protein (PrP(C)) into an ... [more ▼]

Transmissible spongiform encephalopathies are a group of neurodegenerative disorders caused by a posttranslational, conformational change in the cellular isoform of the prion protein (PrP(C)) into an infectious, disease-associated form (PrP(Sc)). Increasing evidence supports a role for PrP(C) in the cellular response to oxidative stress. We investigated the effect of oxidative stress mediated by paraquat exposure on SH-SY5Y neuroblastoma cells. A loss of mitochondrial membrane potential and subsequent reduction in ATP production were demonstrated in untransfected SH-SY5Y cells, an effect that was ameliorated by the expression of PrP(C). Cells expressing either PrP-DeltaOct, which lacks the octapeptide repeats, or PrP-DA, in which the N-terminus is tethered to the membrane, showed increased sensitivity to paraquat compared with cells expressing wild-type PrP(C) as shown by reduced viability, loss of their membrane integrity, and reduced mitochondrial bioenergetic measurements. Exposure of prion-infected mouse SMB15S cells to paraquat resulted in a reduction in viability to levels similar to those seen in the untransfected SH-SY5Y cells. However, "curing" the cells with pentosan sulfate restored the viability to the level observed in the SH-SY5Y cells expressing PrP(C). These data would indicate that the molecular mechanism promoting cellular resistance to oxidative stress had been compromised in the infected SMB15S cells, which could be reinstated upon curing. Our study supports the hypothesis that PrP(C) expression protects cells against paraquat-induced oxidative injury, demonstrates the significance of the N-terminal region of the protein in mediating this protective effect, and also shows that the biochemical consequences of prion infection may be reversed with therapeutic intervention. [less ▲]

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See detailAn immuno-PF2D-MS/MS proteomic approach for bacterial antigenic characterization: To Bacillus and beyond
Ruelle, Virginie ULg; Falisse-Poirier, Nandini; Elmoualij, Benaïssa ULg et al

in Journal of Proteome Research (2007), 6(6), 2168-2175

We are confronted daily to unknown microorganisms that have yet to be characterized, detected, and/ or analyzed. We propose, in this study, a multidimensional strategy using polyclonal antibodies ... [more ▼]

We are confronted daily to unknown microorganisms that have yet to be characterized, detected, and/ or analyzed. We propose, in this study, a multidimensional strategy using polyclonal antibodies, consisting of a novel proteomic tool, the ProteomeLab PF2D, coupled to immunological techniques and mass spectrometry ( i-PF2D-MS/MS). To evaluate this strategy, we have applied it to Bacillus subtilis, considered here as our unknown bacterial model. [less ▲]

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See detailTRANSMISSION ET RÉSISTANCE DES PRIONS : LA PRATIQUE DE LA MÉDECINE DENTAIRE EN SERA T-ELLEAFFECTÉE ?
Elmoualij, Benaïssa ULg; Heinen, Ernst ULg; Zorzi, Willy ULg et al

in Journal de l'Ordre des Dentistes du Québec (2006), 43(9), 461-467

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See detailAdvances in immunoproteomics for serological characterization of microbial antigens
Falisse-Poirier, Nandini; Ruelle, Virginie ULg; Elmoualij, Benaïssa ULg et al

in Journal of Microbiological Methods (2006), 67(3), 593-596

We propose a multi-dimensional strategy, associating immunodetection to a protein fractionating two-dimensional liquid chromatography tool, for serological characterization of microbial antigens. The ... [more ▼]

We propose a multi-dimensional strategy, associating immunodetection to a protein fractionating two-dimensional liquid chromatography tool, for serological characterization of microbial antigens. The originality of such immunoproteomic approaches resides in their application in large-scale studies for rapid serotyping of micro-organisms, evaluation of immunomes and could be proposed in the development and monitoring of vaccines. (c) 2006 Elsevier B.V. All rights reserved. [less ▲]

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See detailImmunoquantitative real-time PCR for detection and quantification of Staphylococcus aureus enterotoxin B in foods
Rajkovic, A.; Elmoualij, Benaïssa ULg; Uyttendaele, M. et al

in Applied and Environmental Microbiology (2006), 72(10), 6593-6599

A real-time immunoquantitative PCR (iqPCR) method for detection of Staphylococcus aureus enterotoxin B (SEB) was developed and evaluated using both pure cultures and foods. The assay consisted of ... [more ▼]

A real-time immunoquantitative PCR (iqPCR) method for detection of Staphylococcus aureus enterotoxin B (SEB) was developed and evaluated using both pure cultures and foods. The assay consisted of immunocapture of SEB and real-time PCR amplification of the DNA probe linked to the detection antibody. iqPCR was compared to an in-house enzyme-linked immunosorbent assay (ELISA) using the same couple of capture-detection antibodies and to commercial kits for detection of S. aureus enterotoxins (SE). The iqPCR was approximately 1,000 times more sensitive (< 10 pg ml(-1)) than the in-house ELISA and had a dynamic range of approximately 10 pg ml(-1) to approximately 30,000 pg ml(-1). iqPCR was not inhibited by any of the foods tested and was able to detect SEB present in these foods. No cross-reactivity with SE other than SEB was observed. Application of iqPCR for detection of SEB in cultures of S. aureus revealed the onset of SEB production after 4 It of incubation at 22, 37, and 42 degrees C, which was in the first half of the exponential growth phase. The total amounts of SEB produced by the two strains tested were larger at 42 degrees C than at 37 degrees C and were strain dependent. [less ▲]

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See detailStudy on the toxic mechanism of prion protein peptide 106-126 in neuronal and non neuronal cells
Dupiereux-Fettweis, Ingrid ULg; Zorzi, Willy ULg; Rachidi, W. et al

in Journal of Neuroscience Research (2006), 84(3), 637-646

A synthetic peptide corresponding to the 106-126 amyloidogenic region of the cellular human prion protein (PrPc) is useful for in vitro study of prion-induced neuronal cell death. The aim of the present ... [more ▼]

A synthetic peptide corresponding to the 106-126 amyloidogenic region of the cellular human prion protein (PrPc) is useful for in vitro study of prion-induced neuronal cell death. The aim of the present work was to examine the implication of the cellular prion protein in the toxicity mechanism induced by PrP 106-126. The effect of PrP 106-126 was investigated both on human neuroblastoma SH-SY5Y cells and on SH-SY5Y over-expressing murine cellular prions (wtPrP). We show by metabolic assay tests and ATP assays that PrPc expression does not modulate the toxicity of the prion peptide. Moreover, we investigated the effect of this peptide on an established non neuronal model, rabbit kidney epithelial A74 cells that express a doxycycline-inducible murine PrPc gene. We show for the first time that the prion peptide 106-126 does not exert any toxic effect on this cell line in the presence or absence of doxycycline. Our results show that the PrP 106-126-induced cell alteration is independent of PrPc expression. Rather, it seems to act via an interaction with lipidic components of the plasma membrane as strengthened by our results showing the differential susceptibility of neuronal and non neuronal cell lines that significantly differ by their membrane fatty acid composition. (c) 2006 Wiley-Liss, Inc. [less ▲]

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See detailDevelopment of a serum-free co-culture of human intestinal epithelium cell-lines (Caco-2/HT29-5M21)
Nollevaux, Géraldine; Deville, Christelle ULg; Elmoualij, Benaïssa ULg et al

in BMC Cell Biology (2006), 7

Background: The absorptive and goblet cells are the main cellular types encountered in the intestine epithelium. The cell lineage Caco-2 is a model commonly used to reproduce the features of the bowel ... [more ▼]

Background: The absorptive and goblet cells are the main cellular types encountered in the intestine epithelium. The cell lineage Caco-2 is a model commonly used to reproduce the features of the bowel epithelium. However, there is a strong debate regarding the value of Caco-2 cell culture to mimick in vivo situation. Indeed, some authors report in Caco-2 a low paracellular permeability and an ease of access of highly diffusible small molecules to the microvilli, due to an almost complete lack of mucus. The HT29-5M21 intestinal cell lineage is a mucin-secreting cellular population. A co-culture system carried out in a serum-free medium and comprising both Caco-2 and HT29-5M21 cells was developed. The systematic use of a co-culture system requires the characterization of the monolayer under a given experimental procedure. Results: In this study, we investigated the activity and localization of the alkaline phosphatase and the expression of IAP and MUC5AC genes to determine a correlation between these markers and the cellular composition of a differentiated monolayer obtained from a mixture of Caco-2 and HT29-5M21 cells. We observed that the culture conditions used ( serum-free medium) did not change the phenotype of each cell type, and produced a reproducible model. The alkaline phosphatase expression characterizing Caco-2 cells was influenced by the presence of HT29-5M21 cells. Conclusion: The culture formed by 75% Caco-2 and 25% HT29-5M21 produce a monolayer containing the two main cell types of human intestinal epithelium and characterized by a reduced permeability to macromolecules. [less ▲]

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See detailL'infection par les prions pathogènes modifie l'expression menbranaire de la PrPc par les cellules dendritiques
Dorban, G; Demonceau,C; Levavasseur, E et al

Poster (2005, October)

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See detailImmunoquantitative PCR for prion protein detection in sporadic Creutzfeldt-Jakob disease.
Gofflot, Stéphanie ULg; Deprez, Manuel ULg; Elmoualij, Benaïssa ULg et al

in Clinical Chemistry (2005), 51(9), 1605-11

BACKGROUND: The most common human prion disorder is Creutzfeldt-Jakob disease (CJD); it includes sporadic, familial, iatrogenic, and variant subtypes. Diagnostic tests aim at detection with the highest ... [more ▼]

BACKGROUND: The most common human prion disorder is Creutzfeldt-Jakob disease (CJD); it includes sporadic, familial, iatrogenic, and variant subtypes. Diagnostic tests aim at detection with the highest specificity of very small deposits of abnormal prion protein (PrP). METHODS: We used immunoquantitative PCR (iqPCR) to detect proteinase K-resistant PrP (PrPRes) in tissue from the middle frontal gyrus of 7 patients with sporadic CJD and 7 non-CJD cases. We compared iqPCR with routine optimized ELISA, Western blotting, and immunohistochemical analyses. RESULTS: The 4 methods showed similar 100% sensitivity and specificity for the diagnosis of CJD. Along with high specificity, however, iqPCR had a threshold for PrP(Res) detection at least 10-fold lower than that of the classic ELISA. CONCLUSIONS: iqPCR is a new method for PrPRes detection that combines 100% specificity with a detection threshold at least 10-fold lower than classic techniques. This method may improve the detection of minute PrPRes deposits in tissues and body fluids and thus be useful for diagnostic and sterilization applications. [less ▲]

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See detailInteraction of the 106-126 prion peptide with lipid membranes and potential implication for neurotoxicity.
Dupiereux-Fettweis, Ingrid ULg; Zorzi, Willy ULg; Lins, Laurence ULg et al

in Biochemical and Biophysical Research Communications (2005), 331(4), 894-901

Prion diseases are fatal neurodegenerative disorders characterized by the accumulation in the brain of an abnormally misfolded, protease-resistant, and beta-sheet rich pathogenic isoform (PrP(SC)) of the ... [more ▼]

Prion diseases are fatal neurodegenerative disorders characterized by the accumulation in the brain of an abnormally misfolded, protease-resistant, and beta-sheet rich pathogenic isoform (PrP(SC)) of the cellular prion protein (PrP(C)). In the present work, we were interested to study the mode of prion protein interaction with the membrane using the 106-126 peptide and small unilamellar lipid vesicles as model. As previously demonstrated, we showed by MTS assay that PrP 106-126 induces alterations in the human neuroblastoma SH-SY5Y cell line. We demonstrated for the first time by lipid-mixing assay and by the liposome vesicle leakage test that PrP 106-126, a non-tilted peptide, induces liposome fusion thus a potential cell membrane destabilization, as supported by membrane integrity assay (LDH). By circular dichroism (CD) analysis we showed that the fusogenic property of PrP 106-126 in the presence of liposome is associated with a predominantly beta-sheet structure. These data suggest that the fusogenic property associated with a predominant beta-sheet structure exhibited by the prion peptides contributes to the neurotoxicity of these peptides by destabilizing cellular membranes. The latter might be attached at the membrane surface in a parallel orientation as shown by molecular modeling. [less ▲]

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See detailDetection of biomarkers of pathogenic bacteria by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry
Ruelle, Virginie; El Moualij, B.; Zorzi, Willy ULg et al

in Lichtfouse, Eric; Schwarzbauer, Jan; Robert, Didier (Eds.) Environmental Chemistry : Green Chemistry and Pollutants in Ecosystems (2005)

In recent years, various mass spectrometry procedures has been developed for identifying bacteria. The accuracy and speed with which data can be obtained by Matrix-assisted laser Desorption/Ionization ... [more ▼]

In recent years, various mass spectrometry procedures has been developed for identifying bacteria. The accuracy and speed with which data can be obtained by Matrix-assisted laser Desorption/Ionization Time-of-flight Mass Spectrometry (MALDI-TOF-MS) make this an advantageous technique for environmental monitoring. However, minor variations in the sample preparation can influence the mass spectra significantly. In the present study, we have introduced a procedure to prepare bacteria by microextraction and we have optimized experimental parameters for rapid identification by MALDI-TOF-MS of whole bacterial cells isolated from environmental samples such as wastewater and soil. [less ▲]

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See detailImmuno-quantitative polymerase chain reaction for detection and quantitation of prion protein
Gofflot, Stéphanie ULg; Elmoualij, Benaïssa ULg; Zorzi, Danièle ULg et al

in Journal of Immunoassay & Immunochemistry (2004), 25(3), 241-258

Immuno-polymerase chain reaction (PCR) is an extremely sensitive detection method, combining the specificity of antibody detection and the sensitivity of PCR. We have developed an immuno-quantitative PCR ... [more ▼]

Immuno-polymerase chain reaction (PCR) is an extremely sensitive detection method, combining the specificity of antibody detection and the sensitivity of PCR. We have developed an immuno-quantitative PCR (iqPCR), exploiting real-time PCR technology, in order to improve this immuno-detection method and make it quantitative. To illustrate the advantages of iqPCR, we have compared it with a conventional enzyme linked immuno sorbent assay (ELISA) technique in experiments aimed at detecting the cellular and the resistant form of prion protein in bovine brain extract. The iqPCR technique proved to be more sensitive than ELISA, so it could be a technique of choice for the diagnosis of infected animals both at an ante mortem and post-mortem stage. [less ▲]

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See detailHuman recombinant thiamine triphosphatase: purification, secondary structure and catalytic properties
Lakaye, Bernard ULg; Makarchikov, Alexander F; Wins, Pierre et al

in International Journal of Biochemistry & Cell Biology (2004), 36(7), 1348-1364

Thiamine triphosphate (ThTP) is found in most living organisms and it may act as a phosphate donor for protein phosphorylation. We have recently cloned the cDNA coding for a highly specific mammalian 25 ... [more ▼]

Thiamine triphosphate (ThTP) is found in most living organisms and it may act as a phosphate donor for protein phosphorylation. We have recently cloned the cDNA coding for a highly specific mammalian 25 kDa thiamine triphosphatase (ThTPase; EC 3.6.1.28). As the enzyme has a high catalytic efficiency and no sequence homology with known phosphohydrolases, it was worth investigating its structure and catalytic properties. For this purpose, we expressed the untagged recombinant human ThTPase (hThTPase) in E. coli, produced the protein on a large scale and purified it to homogeneity. Its kinetic properties were similar to those of the genuine human enzyme, indicating that the recombinant hThTPase is completely functional. Mg2+ ions were required for activity and Ca2+ inhibited the enzyme by competition with Mg2+. With ATP as substrate, the catalytic efficiency was 10(-4)-fold lower than with ThTP, confirming the nearly absolute specificity of the 25 kDa ThTPase for ThTP. The activity was maximum at pH 8.5 and very low at pH 6.0. Zn2+ ions were inhibitory at micromolar concentrations at pH 8.0 but activated at pH 6.0. Kinetic analysis suggests an activator site for Mg2+ and a separate regulatory site for Zn2+. The effects of group-specific reagents such as Woodward's reagent K and diethylpyrocarbonate suggest that at least one carboxyl group in the active site is essential for catalysis, while a positively charged amino group may be involved in substrate binding. The secondary structure of the enzyme, as determined by Fourier-transform infrared spectroscopy, was predominantly beta-sheet and alpha-helix. [less ▲]

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See detailHuman immune cells express ppMCH mRNA and functional MCHR1 receptor
Verlaet, Myriam ULg; Adamantidis, Antoine ULg; Coumans, Bernard ULg et al

in FEBS Letters (2002), 527(1-3), 205-210

Melanin-concentrating hormone (MCH) is highly expressed in the brain and modulates feeding behavior. It is also expressed in some peripheral tissues where its role remains unknown. We have investigated ... [more ▼]

Melanin-concentrating hormone (MCH) is highly expressed in the brain and modulates feeding behavior. It is also expressed in some peripheral tissues where its role remains unknown. We have investigated MCH function in human and mouse immune cells. RT-PCR analysis revealed a low expression of prepro-MCH and MCH receptor 1 (MCHR1) but not of MCHR2 transcript in tissular and peripheral blood immune cells. FACS and in vitro assay studies demonstrated that MCHR1 receptor expression on most cell types can trigger, in the presence of MCH, cAMP synthesis and calcium mobilization in peripheral blood mononuclear cells (PBMCs). Moreover, MCH treatment decreases the CD3-stimulated PBMC proliferation in vitro. Accordingly, our data indicate for the first time that MCH and MCHR1 may exert immunomodulatory functions. (C) 2002 Federation of European Biochemical Societies. Published by Elsevier Science B.V. All rights reserved. [less ▲]

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See detailMolecular characterization of a specific thiamine triphosphatase widely expressed in mammalian tissues
Lakaye, Bernard ULg; Makarchikov, Alexander F; Antunes, Adelio F et al

in Journal of Biological Chemistry (2002), 277(16), 13771-13777

Thiamine triphosphate (ThTP) is found at low concentrations in most animal tissues, and recent data suggest that it may act as a phosphate donor for the phosphorylation of some proteins. In the mammalian ... [more ▼]

Thiamine triphosphate (ThTP) is found at low concentrations in most animal tissues, and recent data suggest that it may act as a phosphate donor for the phosphorylation of some proteins. In the mammalian brain, ThTP synthesis is rapid, but its steady-state concentration remains low, presumably because of rapid hydrolysis. In this report we purified a soluble thiamine triphosphatase (ThTPase; EC 3.6.1.28) from calf brain. The bovine ThTPase is a 24-kDa monomer, hydrolyzing ThTP with virtually absolute specificity. Partial sequence data obtained from the purified bovine enzyme by tandem mass spectrometry were used to search the GenBank(TM) data base. A significant identity was found with only one human sequence, the hypothetical 230-amino acid protein MGC2652. The coding regions from human and bovine brain mRNA were amplified by reverse transcription-PCR, cloned in Escherichia coli, and sequenced. The human open reading frame was expressed in E. coli as a GST fusion protein. Transformed bacteria had a high isopropyl-beta-D-thiogalactopyranoside-inducible ThTPase activity. The recombinant ThTPase had properties similar to those of human brain ThTPase, and it was specific for ThTP. The mRNA was expressed in most human tissues but at relatively low levels. This is the first report of a molecular characterization of a specific ThTPase. [less ▲]

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