Géraldine Lucchi

Centre Hospitalier Universitaire de Dijon, Dijon, Bourgogne, France

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Publications (22)60.79 Total impact

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    ABSTRACT: Abstract Saliva has different functions in the mouth and is involved, for example, in taste perception. Saliva composition can also be modified rapidly by taste stimulation. It remains unclear, however, whether the perceived intensity of a tastant may modulate this response. Based on increasing evidence that fat can be perceived by the taste system and that fat taste perception may be associated with fat intake, the aim of this work was to study if stimulation by a fatty acid (oleic acid) modifies saliva composition differently in subjects highly (sensitive+) or weakly (sensitive-) sensitive to that taste. For that purpose, saliva of two groups of subjects was collected after stimulation by either a control emulsion or an emulsion containing 5.61 mM oleic acid. Saliva was analyzed by 2D electrophoresis and (1)H NMR spectroscopy. The results show that sensitive+ and sensitive- subjects differ in their salivary response in terms of proteome and metabolome composition. Oppositely to sensitive- subjects, sensitive+ subjects responded to oleic acid by increased abundance of polymeric immunoglobulin receptor, rab GDP dissociation inhibitor beta, and organic acids, and decreased abundance of metabolites characteristic of mucins. The results highlight that modification of saliva composition by taste stimulation may be modulated by taste perception.
    Omics : a journal of integrative biology. 10/2014;
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    ABSTRACT: High plasma concentrations of non-esterified fatty acids (NEFAs), transported bound to serum albumin, are associated with type 2 diabetes (T2D). The effects of albumin on platelet function were investigated in vitro. Modifications of albumin, such as those due to glycoxidation, were found in patients with T2D, and the consequences of these modifications on biological mechanisms related to NEFA handling were investigated. Mass spectrometry profiles of albumin from patients with T2D differed from those from healthy controls. Diabetic albumin showed impaired NEFA binding capacity, and both structural and functional alterations could be reproduced in vitro by incubating native albumin with glucose and methylglyoxal. Platelets incubated with albumin isolated from patients with T2D aggregated approximately twice as much as platelets incubated with albumin isolated from healthy controls. Accordingly, platelets incubated with modified albumin produced significantly higher amounts of arachidonate metabolites than did platelets incubated with control albumin. It is concluded that higher amounts of free arachidonate are made available for the generation of active metabolites in platelets when the NEFA binding capacity of albumin is blunted by glycoxidation. This newly described mechanism, in addition to hypoalbuminaemia may contribute to platelet hyperactivity and increased thrombosis, known to occur in patients with T2D.
    Diabetes 08/2014; · 7.90 Impact Factor
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    ABSTRACT: Based on recent studies in adult subjects, saliva composition is increasingly considered as a physiological factor contributing to taste sensitivity or acceptance. In order to evaluate a possible link between salivary protein composition and taste acceptance in infants, 73 infants participated longitudinally in taste acceptance tests and donated saliva at the age at 3 and 6 months. Intake ratios, reflecting acceptance of a taste solution relative to water were calculated for the five basic tastes. Salivary proteins were separated by one-dimensional electrophoresis and bands were semi-quantified by image analysis. Partial least square (PLS) regression analyses were performed for each taste at both ages to explain intake ratios by band intensities. Bitterness acceptance in the younger infants was unique in the sense that salivary protein profiles could partly predict bitter taste acceptance. At that age, infants were on average indifferent to the 0.18-M urea solution, but great variability in acceptance was observed. The six bands considered as the best predictors for bitterness acceptance were identified by MALDI-TOF mass spectrometry. Higher abundance of bands containing secretory component, zinc-α-2-glycoprotein and carbonic anhydrase 6 was associated to a lower bitterness acceptance, while higher abundance of bands containing lactoperoxidase, prolactin-inducible protein and S-type cystatins was associated to a higher bitterness acceptance. In a second stage, S-type cystatin abundance was measured by Western blotting in order to tentatively confirm this particular finding in an independent group of 22 infants. Although not reaching statistical significance, probably due to a relatively small sample size, it was again observed that cystatin abundance was higher in infants accepting more readily the bitter solution over water. Conclusion: saliva protein composition may contribute to bitter taste acceptance in the younger infants.
    European Journal of Pediatrics 11/2013; · 1.91 Impact Factor
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    ABSTRACT: MOZ and MLL encoding a histone acetyltransferase and a histone methyltransferase, respectively, are targets for recurrent chromosomal translocations found in acute myeloblastic or lymphoblastic leukemia. We have previously shown that MOZ and MLL cooperate to activate HOXA9 gene expression in hematopoietic stem/progenitors cells. To dissect the mechanism of action of this complex, we decided to identify new proteins interacting with MOZ. We found that the scaffold protein Symplekin that supports the assembly of polyadenylation machinery was identified by mass spectrometry. Symplekin interacts and co-localizes with both MOZ and MLL in immature hematopoietic cells. Its inhibition leads to a decrease of HOXA9 protein level but not of Hoxa9 mRNA and to an over-recruitment of MOZ and MLL onto HOXA9 promoter. Altogether, our results highlight the role of Symplekin in transcription repression involving a regulatory network between MOZ, MLL and Symplekin.
    Biochimica et Biophysica Acta 08/2013; · 4.66 Impact Factor
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    ABSTRACT: We report here that modifications of the preanalytical steps of matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) identification of yeasts, with regard to the original protocol provided by the manufacturers, appear to be efficient for the reliable routine identification of clinical yeast isolates in medical laboratories. Indeed, when one colony was sampled instead of five and the protein extraction protocol was modified, the performance of MALDI-TOF MS was superior to that of the API ID 32C method (discrepancies were confirmed by using molecular identification), allowing the correct identification of 94% of the 335 clinical isolates prospectively tested. We then demonstrated that the time for which the primary cultures were preincubated on CHROMagar did not impact the identification of yeasts by MALDI-TOF MS, since 95.1 and 96.2% of the 183 clinical yeast isolates prospectively tested were correctly identified after 48 and 72 h of preincubation, respectively.
    Journal of clinical microbiology 06/2012; 50(9):3066-8. · 4.16 Impact Factor
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    ABSTRACT: Conventional identification (CI) of yeasts is based on morphological, biochemical and/or immunological methods. Matrix-assisted laser desorption/ionization - time of flight (MALDI-TOF or MT-MS) mass spectrometry has been proposed as a new method for the identification of microorganisms. This prospective study compared the performance of MT-MS and CI for the identification of yeasts isolated from clinical samples. Sequencing of the internal transcribed spacer (ITS) regions of ribosomal DNA was used as the reference method in the analysis of a total of 1207 yeast isolates. Concordance between MT-MS and CI was observed for 1105 isolates (91.5%), while 74 isolates (6.1%) were misidentified. Molecular identification revealed that 73 of these 74 isolates were identified correctly by MT-MS and CI correctly identified the last one. Concordance between the two techniques was excellent for the medically-important species (98-100%), including the identification of closely-related species (Candida albicans/C. dubliniensis; C. inconspicua/C. norvegensis; C. parapsilosis/C. metapsilosis/C. orthopsilosis). Only 2.3% of isolates belonging to C. famata, C. lambica and C. magnoliae or to Geotrichum spp. and Trichosporon spp. were not identified by MT-MS. This investigation highlights the potential of MT-MS-based yeast identification as a reliable, time and cost-efficient alternative to CI.
    Medical mycology: official publication of the International Society for Human and Animal Mycology 06/2012; · 2.13 Impact Factor
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    ABSTRACT: Protein biomarker discovery and validation are crucial for diagnosis, prognosis, and theranostics of human pathologies; "omics" approaches bring new insights in this field. In particular, the combination of immuno-sensors in array format with mass spectrometry efficiently extends the classical immunoassay format and includes molecular characterization. Here, we coupled surface plasmon resonance imaging (SPRi) with MALDI-TOF mass spectrometry in a hyphenated technique which enables multiplexed quantification of binding by SPRi and molecular characterization of interacting partners by subsequent MS analysis. This adds specificity, because MS enables differentiation of molecules that are difficult to distinguish by use of antibodies, for example truncation variants or protein isoforms. Proof of concept was established for detection, identification, and characterization of a potential breast cancer marker, the LAG3 protein, at ~1 μg mL(-1), added to human plasma. The analytical performance of this new method, dubbed "SUPRA-MS", was established, particularly its specificity (S/N > 10) and reliability (100 % LAG3 identification with high significant mascot score >87.9). The adjusted format for rapid, collective, and automated on-chip MALDI-MS analysis is robust at the femtomole level and has numerous potential applications in proteomics.
    Analytical and Bioanalytical Chemistry 06/2012; 404(2):423-32. · 3.66 Impact Factor
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    ABSTRACT: In order to describe developmental changes in human salivary peptidome, whole saliva was obtained from 98 infants followed longitudinally at 3 and 6months of age. Data on teeth eruption and diet at the age of 6months were also recorded. Salivary peptide extracts were characterised by label-free MALDI-MS. Peptides differentially expressed between the two ages, and those significantly affected by teeth eruption or introduction of solid foods were identified by MALDI TOF-TOF and LC ESI MS-MS. Out of 81 peaks retained for statistical analysis, 26 were overexpressed at the age of 6months. Exposure to solid foods had a more pronounced effect on profiles (overexpression of nine peaks) than teeth eruption (overexpression of one peak). Differential peaks corresponded to fragments of acidic and basic PRPs, statherin and histatin. Comparison with existing knowledge on adult saliva peptidome revealed that proteolytic processing of salivary proteins is qualitatively quite comparable in infants and in adults. However, age and diet are modulators of salivary peptidome in human infants.
    Journal of proteomics 04/2012; 75(12):3665-73. · 5.07 Impact Factor
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    ABSTRACT: Immuno-SPR-MS is the combination of immuno-sensors in biochip format with mass spectrometry. This association of instrumentation allows the detection and the quantification of proteins of interest by SPR and their molecular characterization by additional MS analysis. However, two major bottlenecks must be overcome for a wide diffusion of the SPR-MS analytical platform: (i) To warrant all the potentialities of MS, an enzymatic digestion step must be developed taking into account the spot formats on the biochip and (ii) the biological relevancy of such an analytical solution requires that biosensing must be performed in complex media. In this study, we developed a procedure for the detection and the characterization at ~1 µg/mL of the LAG3 protein spiked in human plasma. The analytical performances of this new method was established, particularly its specificity (S/N > 9) and sensitivity (100% of LAG3 identification with high significant mascot score >68 at the femtomole level). The collective and automated on-chip MALDI-MS imaging and analysis based on peptidic fragments opens numerous applications in the fields of proteomics and diagnosis.
    Sensors 01/2012; 12(11):15119-32. · 2.05 Impact Factor
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    ABSTRACT: GABARAPL1 belongs to the small family of GABARAP proteins (including GABARAP, GABARAPL1 and GABARAPL2/GATE-16), one of the two subfamilies of the yeast Atg8 orthologue. GABARAPL1 is involved in the intracellular transport of receptors, via an interaction with tubulin and GABA(A) or kappa opioid receptors, and also participates in autophagy and cell proliferation. In the present study, we identify the HSP90 protein as a novel interaction partner for GABARAPL1 using GST pull-down, mass spectrometry and coimmunoprecipitation experiments. GABARAPL1 and HSP90 partially colocalize in MCF-7 breast cancer cells overexpressed Dsred-GABARAPL1 and in rat brain. Moreover, treatment of MCF-7 cells overexpressed FLAG-GABARAPL1-6HIS with the HSP90 inhibitor 17-AAG promotes the GABARAPL1 degradation, a process that is blocked by proteasome inhibitors such as MG132, bortezomib and lactacystin. Accordingly, we demonstrate that HSP90 interacts and protects GABARAPL1 from its degradation by the proteasome.
    Biochimie 11/2011; 94(3):748-58. · 3.14 Impact Factor
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    ABSTRACT: The interindividual variation in the sensitivity to bitterness is attributed in part to genetic polymorphism at the taste receptor level, but other factors, such as saliva composition, might be involved. In order to investigate this, 2 groups of subjects (hyposensitive, hypersensitive) were selected from 29 healthy male volunteers based on their detection thresholds for caffeine, and their salivary proteome composition was compared. Abundance of 26 of the 255 spots detected on saliva electrophoretic patterns was significantly different between hypo- and hypersensitive subjects. Saliva of hypersensitive subjects contained higher levels of amylase fragments, immunoglobulins, and serum albumin and/or serum albumin fragments. It also contained lower levels of cystatin SN, an inhibitor of protease. The results suggest that proteolysis occurring within the oral cavity is an important perireceptor factor associated to the sensitivity to the bitter taste of caffeine.
    Chemical Senses 08/2011; 37(1):87-95. · 3.22 Impact Factor
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    ABSTRACT: Peroxisome proliferators have been extensively studied in rodents and are known to induce liver tumors, whereas the effects of these compounds are not very clearly identified in humans when they are widely exposed to herbicides, plasticizers, solvents or drugs such as the lipid-lowering fibrate bezafibrate (BEZA). We assessed the effect of BEZA on human hepatocyte proteome. Hepatocyte proteins, including those membrane-associated, were successfully extracted and separated using 2D-liquid chromatography (PF2D, Beckman coulter). Proteins that were regulated by ≥ 1.5 fold compared to controls were identified by mass spectrometry (MALDI-TOF, Bruker Daltonics) and SwissProt bank search. BEZA modified the expression of proteins involved in various metabolic pathways as well as in cell homeostasis. No marker of peroxisome proliferation was obtained but surprisingly the expression of proteins involved in liver carcinogenicity was modulated. The co-treatment of cultures with N-acetylcysteine modified the set of proteins regulated by BEZA, either by a potentiation or an inhibition of the effects. Our study points out that the hepatocellular redox environment has to be taken into account when using fibrates in therapeutics.
    Toxicology Letters 03/2011; 201(2):123-9. · 3.15 Impact Factor
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    ABSTRACT: The objective of this study was to describe the changes in salivary protein profiles in infants between the ages of 3 and 6 months, and to evaluate the impact of teeth eruption and introduction of solid foods on such profiles. 73 infants were followed longitudinally at 3 and 6 months of age. Their whole saliva proteins were separated by SDS-PAGE electrophoresis and semi-quantified by image analysis. Amylase activity was also measured on a sub-sample of the population (n=42 infants). Bands which abundance was significantly different between the two ages according to paired comparisons were identified by mass spectrometry techniques. Out of 21 bands, 13 were significantly different between 3 and 6 months of age. Two short variants of amylase increased in abundance with age, as did amylase activity. Other changes possibly translated developmental physiological events, for example maturation of the adaptive immune system. The balance between S-type cystatins and cystatins A and B was modified, in favour of S-type cystatins at 6 months of age. Teeth eruption resulted in an increase in albumin abundance, whilst introduction of solid foods was associated with higher levels of β-2 microglobulin and S-type cystatins. Salivary profiles were modified substantially between the ages of 3 and 6 months. Both teeth eruption and diet had an impact on abundance changes for some proteins, revealing dynamic interactions between saliva proteome, oral physiology and diet.
    Archives of oral biology 03/2011; 56(7):634-42. · 1.65 Impact Factor
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    ABSTRACT: In order to document inter-individual variability in salivary protein patterns, unstimulated whole saliva was obtained from 12 subjects at 10 am and 3 pm of the same day. Saliva proteins were separated using two-dimensional gel electrophoresis, and semi-quantified using image analysis. One-way ANOVA was used to test the effects "time of sampling" and "subject". Data were further explored by multivariate analyses (PCA, hierarchical clustering). Spots of interest were identified by mass spectrometry (MALDI-TOF MS/MS and nanoLC ESI-IT MS/MS). A dataset of 509 spots matched in all gels was obtained. There was no diurnal statistical effect on salivary patterns while inter-individual variability was high with 47 spots differentially expressed between subjects (p<1%). Clustering of these spots revealed that subjects could be discriminated first based on several proteins participating to the non-specific immune response (cystatins, lipocalin 1, parotid-secretory protein and prolactin-induced protein). Independently, subjects were also differentiated by their level of proteins originating from serum and involved in the immune system (complement C3, transferrin, IgG2), as well as the relative abundance of enzymes involved in carbohydrates metabolism (amylase and glycolytic enzymes). Inter-individual variability should be accounted for when searching for salivary biomarkers or when studying in-mouth biochemical mechanisms.
    Journal of proteomics 05/2009; 72(5):822-30. · 5.07 Impact Factor
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    ABSTRACT: The objective of the study was to determine optimal conditions for sampling, sample processing and mass-spectrometry based analysis of infants’ salivary peptidome. Saliva was sampled in 3- and 6-month-old infants and peptide extracts were prepared. Various sample pretreatments before profiling by MALDI-ToF were evaluated and peptide identification was undertaken by MALDI-ToF/ToF or nanoLC-ESI-IT tandem MS. A fast and simple protocol (cut-off filtration at 5 kDa) was satisfactory to produce extracts where no proteolysis was detected even when no protease inhibitor was added. Optimal MALDI spectra were generated after purification on C18 tips. Variability of spectra between two samples exceeded that of the technical replicates, validating that the method is suitable to conduct differential studies. Salivary peptides, identified by means of the two complementary mass spectrometry techniques, were fragments of proline-rich proteins and histatins. The fragments originated mainly from the C-terminal protein extremities. Indications on the proteolytic systems involved and the anatomic location where they intervene are proposed.
    International Journal of Peptide Research and Therapeutics 01/2009; 15(3):177-185. · 1.28 Impact Factor
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    ABSTRACT: We present the results of a study in which biomolecular interaction analysis (BIA, Biacoretrade mark 2000) was combined with mass spectrometry (MS) using entire "on-a-chip" procedure. Most BIA-MS studies included an elution step of the analyte prior MS analysis. Here, we report a low-cost approach combining Biacore analysis with homemade chips and MS in situ identification onto the chips without elution step. First experiments have been made with rat serum albumin to determine the sensitivity and validation of the concept has been obtained with an antibody/antigen couple. Our "on-a-chip" procedure allowed complete analysis by MS/MS(2) of the biochip leading to protein identifications at low femtomole to sub-femtomole levels. Using this technique, identification of protein complexes were routinely obtained giving the opportunity to the "on-a-chip" processing to complete the BIA-MS approach in the discovery and analysis of protein complexes.
    Biosensors & bioelectronics 08/2008; 24(5):1121-7. · 5.43 Impact Factor
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    ABSTRACT: Proteins identified in biological fluids of cancer patients could be helpful for both diagnosis and prognosis. However, clinical proteomics based on analysis of protein profiles in biological fluids has demonstrated various flaws, most of them related to the difficulties met in reproducibility. These difficulties could be partly overcome by accurate standardisation of pre-analytical and analytical steps of these studies. The size of the patient cohort is one of the parameters that determine the powerfulness of the study. Recruitment of a cohort with a sufficient size often implies multicentric studies in which analysis of the reproducibility between centres and standardisation of pre-analytical and analytical steps are essential. Such a standardisation requires the use of calibrated samples as common references.
    Medecine sciences: M/S 04/2007; 23 Spec No 1:19-22. · 0.56 Impact Factor
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    ABSTRACT: A great challenge in biosensors and diagnosis devices relies on the way to reconstitute relevant biological mechanisms on surface of the biochips and which analytical tools are convenient to provide accurate and rapid information on the structures and function of molecules attached to this surface. A better control in the realization of biochips can be obtained in combining different complementary approaches while always keeping in mind the biological key point. Researches in CLIPP are focused towards this objective. Conception, realization and characterization of protein and cell chips are presented. We detail different strategies of materials engineering1,2,3, chemical functionalizations and biomolecular graftings4, molecular and cellular characterization in physiological conditions5,6, which lead to the optimization of “biorecognitions events” at the surface of the chip. We present herein an original interdisciplinary approach, consisting to carry out in parallel a micro-scale analysis (SPR, fluorescence microscopy) and nano-scale characterizations (AFM, XPS, TOF-SIMS). Concerning protein interfaces, we demonstrated in particular that the molecular orientation in a protein monolayer can be determined based on the specific fragment ions from the protein in TOF-SIMS spectra7. These developments have also contributed to the establishment of a new biomolecular interaction analysis/mass spectrometry (BIA-MS) combination based on an entire “on-a-chip” procedure8. We report a low-cost approach combining Biacore 2000 analysis with homemade chips and MS and MS/MS identification directly onto the chips without elution step. Using this technique, identification of protein complexes were routinely obtained giving the opportunity to the “on-a-chip” processing to complete the BIA-MS approach in the discovery and analysis of protein complexes in biological fluids. Our interest is also focused on cell/surface interaction. The cell biochips we are developing consist either of circulating or adherent cells, that we characterized in terms of cell capture on biofunctionalized surface or growth with substrate dependency respectively. Parameters such as cell spreading, growth, morphology, and topography are particularly investigated and controlled by atomic force microscopy in physiological conditions6. With the aim to increase the throughput of analysis, we are currently working on cell and protein micro-arrays. Our expertise in cell and protein biochip preparation, and competences in micro- to nanoscale characterization in liquid conditions, represents precious assets enabling a relevant clinical proteomic research, thanks to deeply controlled steps of biosensor development and use.
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    ABSTRACT: L’identification conventionnelle (IC) des levures est fondée sur l’utilisation des tests morphologiques, immunologiques et biochimiques (galerie API® 32 C, bioMérieux). La spectrométrie de masse MALDI-TOF (SM) a été proposée récemment comme nouvelle approche pour l’identification des microorganismes. L’objectif de notre étude était de comparer prospectivement les performances analytiques de la SM et de l’IC pour l’identification des isolats cliniques de levures. En cas de discordance d’identification, le séquençage des régions ITS de l’ADN ribosomal était utilisé comme méthode d’identification de référence. Un total de 1 207 souches a été analysé. Une concordance entre SM et IC a été observée pour 1 105 souches (91,5 %) alors que la proportion de souches différemment identifiées par IC et SM n’était que de 6,1 % (74 souches). Parmi ces 74 identifications discordantes entre SM, l’identification par biologie moléculaire confirmait l’identification obtenue par SM pour 73 isolats (6 %) et celle obtenue par IC pour 1 isolat (0,1 %). Pour les principales espèces d’intérêt médical, les concordances d’identification entre les deux techniques étaient excellentes (entre 98 et 100 %) y compris pour les espèces phylogénétiquement proches (Candida albicans/Candida dubliniensis; Candida inconspicua/Candida norvegensis; Candida parapsilosis/Candida metapsilosis/Candida orthopsilosis). La SM n’a été mise en défaut que pour 2,3 % des souches appar tenant aux espèces Candida famata, Candida lambica, Candida magnoliae et aux genres Geotrichum sp. et Trichosporon sp. Nos investigations soulignent le potentiel de la SM pour l’identification des levures et comme alternative fiable, rapide et de moindre coût par rapport aux méthodes conventionnelles.
    Bio Tribune Magazine 40(1).