Institut de Chimie & Biologie des Membranes & des Nano-objets • Bordeaux

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James Nowick, 15 september 2017, 11am, IECB Emmanuelle Thinon, 28 June 2017, 11 am, IECB Caroline Tokarski, 15 June 2017, 4pm, ENSCBP Christian Salesse, 8 June 2017, 4pm, ENSCBP Patrice Rey, 01 June 2017, 2h30 pm, ENSCBP Christina Sizun, 18 May 2017, 4 pm, ENSCBP Marisela Velez, 5 May 2017, 11am, IECB Iqbal Choudhary, 27 April 2017, 4 pm, ENSCBP Vincent Aucagne, 21 April 2017, 11 am, IECB Sophie Zinn-Justin, 14 April 2017, 11 am, IECB Cécile Feuillie, 16 February 2017, 4 PM, ENSCBP Félix M. Goñi, 8 december 2016, 4 pm, ENSCBP Félix M. Goñi, 17 november 2016, 4 pm, ENSCBP Carl Creutz, 20 october 2016, 4 pm, ENSCBP Diego Romero, 30 september 2016, 11 am, IECB A. Ciaccafava, 8 september 2016, 14h, ENSCBP A. Ramamoorthy, 16 June 2016, 16h, ENSCBP Alexandre de Brevern, 2 june 2016, ENSCBP Isabelle Landrieu, 26 May, 16h, ENSCBP Frances Sepavoric, 12 May, 16h, ENSCBP Thomas Pradeu, 7 April 2016, 9h, ENSCBP Françoise Argoul, 3 march 2016, 16h, ENSCBP Corinne Loutelier-Bourhis, 16/12/2015, 16h Aristotelis XENAKIS, 3 december 2015, 16h Fabian Kiessling, 26 november 2015, 11h Michaël Molinari, 20 november 2015, 11h Dipankar Das Sarma, 28 October 2015, 14h. Olivier Donard, 22 october 2015, 16h, ENSCBP E. Morvan & A.Grelard, 17/12/2015, 16h, ENSCBP Christophe Cullin, 10 september 2015, 14 h Brigitte Lindet, 18 June 2015, 16h, ENSCBP(B) Marion Decossas, 4 June 2015, 16h, ENSCBP(B) Pascale Schellenberger, 21 Mai 2015, 16h F. Leal-Calderon, 7 May 2015, 16h, ENSCBP(B) Ibrahim Abdulhalim, 9 April 2015, 14h, ENSCBP Manon Carré, 26 March 2015, 14h, ENSCBP(B) C. Bure & JM Schmitter, 19 March 2015, 16h T. Ogata & H. Ihara, 17 March 2015, 11h, IECB Banafshe Larijani, 12 March 2015, 16h, ENSCBP
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LIA France-Japan GEM Réseau RMN Aquitain Master AC2QMAPS IPEP master UREKA UMT FOLIES INDIA
Offres d'Emploi Plateau Technique Conseil d'Unité Animation Scientifique WEB Editorial Committee Hygiène & Sécurité Communication Administration Directoire Scientifique Direction
Chimie Biophysique Chimie Biomimétique et Thérapeuti... Biologie et Biotechnologie Administration
James Nowick, 15 september 2017, 11... Emmanuelle Thinon, 28 June 2017, 11... Caroline Tokarski, 15 June 2017, 4p... Christian Salesse, 8 June 2017, 4pm... Patrice Rey, 01 June 2017, 2h30 pm,... Christina Sizun, 18 May 2017, 4 pm,... Marisela Velez, 5 May 2017, 11am, I... Iqbal Choudhary, 27 April 2017, 4 p... Vincent Aucagne, 21 April 2017, 11 ... Sophie Zinn-Justin, 14 April 2017, ... Cécile Feuillie, 16 February 2017,... Félix M. Goñi, 8 december 2016, 4... Félix M. Goñi, 17 november 2016, ... Carl Creutz, 20 october 2016, 4 pm,... Diego Romero, 30 september 2016, 11... A. Ciaccafava, 8 september 2016, 14... A. Ramamoorthy, 16 June 2016, 16h,... Alexandre de Brevern, 2 june 2016, ... Isabelle Landrieu, 26 May, 16h, ENS... Frances Sepavoric, 12 May, 16h, ENS... Thomas Pradeu, 7 April 2016, 9h, EN... Françoise Argoul, 3 march 2016, 1... Corinne Loutelier-Bourhis, 16/12/20... Aristotelis XENAKIS, 3 december 201... Fabian Kiessling, 26 november 2015,... Michaël Molinari, 20 november 2015... Dipankar Das Sarma, 28 October 2015... Olivier Donard, 22 october 2015, 16... E. Morvan & A.Grelard, 17/12/2015, ... Christophe Cullin, 10 september 201... Brigitte Lindet, 18 June 2015, 16h,... Marion Decossas, 4 June 2015, 16h, ... Pascale Schellenberger, 21 Mai 2015... F. Leal-Calderon, 7 May 2015, 16h, ... Ibrahim Abdulhalim, 9 April 2015, 1... Manon Carré, 26 March 2015, 14h, ... C. Bure & JM Schmitter, 19 March 20... T. Ogata & H. Ihara, 17 March 2015,... Banafshe Larijani, 12 March 2015, 1...
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1 […]
Xiang, Shengqi; Kulminskaya, Natalia; Habenstein, Birgit; Biernat, Jacek; Tepper, Katharina; Paulat, Maria; Griesinger, Christian; Becker, Stefan; Lange, Adam; Mandelkow, Eckhard; Linser, Rasmus;
A Two-Component Adhesive: Tau Fibrils Arise from a Combination of a Well-Defined Motif and Conformationally Flexible Interactions
Journal of the American Chemical Society, 2017, 139, 2639-2646
Fibrillar aggregates of A beta and Tau in the brain are the major hallmarks of Alzheimer's disease. Most Tau fibers have a twisted appearance, but the twist can be variable and even absent. This ambiguity, which has also been associated with different phenotypes of tauopathies, has led to controversial assumptions about fibril constitution, and it is unclear to-date what the molecular causes of this polymorphism are. To tackle this question, we used solid-state NMR strategies providing assignments of non-seeded three-repeat-domain Tau(3RD) with an inherent heterogeneity. This is in contrast to the general approach to characterize the most homogeneous preparations by construct truncation or intricate seeding protocols. Here, carbon and nitrogen chemical-shift conservation between fibrils revealed invariable secondary-structure properties, however, with inter-monomer interactions variable among samples. Residues with variable amide shifts are localized mostly to N- and C-terminal regions within the rigid beta structure in the repeat region of Tau(3RD). By contrast, the hexapeptide motif in repeat R3, a crucial motif for fibril formation, shows strikingly low variability of all NMR parameters: Starting as a nucleation site for monomer monomer contacts, this six-residue sequence element also turns into a well-defined structural element upon fibril formation. Given the absence of external causes in vitro, the interplay of structurally differently conserved elements in this protein likely reflects an intrinsic property of Tau fibrils.
2 […]
Wolff, Philippe; Da Veiga, Cyrielle; Ennifar, Eric; Bec, Guillaume; Guichard, Gilles; Burnouf, Dominique; Dumas, Philippe;
Native ESI Mass Spectrometry Can Help to Avoid Wrong Interpretations from Isothermal Titration Calorimetry in Difficult Situations
Journal of the American Society for Mass Spectrometry, 2017, 28, 347-357
We studied by native ESI-MS the binding of various DNA-polymerase-derived peptides onto DNA-polymerase processivity rings from Escherichia coli, Pseudomonas aeruginosa, and Mycobacterium tuberculosis. These homodimeric rings present two equivalent specific binding sites, which leads to successive formation during a titration experiment of singly- and doubly occupied rings. By using the ESI-MS free-ring spectrum as a ruler, we derived by robust linear regression the fractions of the different ring species at each step of a titration experiment. These results led to accurate K-d values (from 0.03 to 0.5 mu M) along with the probability of peptide loss due to gas phase dissociation (GPD). We show that this good quality is due to the increased information content of a titration experiment with a homodimer. Isothermal titration calorimetry (ITC) led with the same binding model to K-d(ITC) values systematically higher than their ESI-MS counterparts and, often, to poor fit of the ITC curves. A processing with two competing modes of binding on the same site requiring determination of two (K-d, Delta H) pairs greatly improved the fits and yielded a second K-d(ITC) close to K-d(ESI-MS). The striking features are: (1) ITC detected a minor binding mode (similar to 20%) of 'low-affinity' that did not appear with ESI-MS; (2) the simplest processing of ITC data with only one (K-d, Delta H) pair led wrongly to the Kd of the low-affinity binding mode but to the Delta H of the high-affinity binding mode. Analogous misleading results might well exist in published data based on ITC experiments.
3 […]
Wang, Xiang; Wicher, Barbara; Ferrand, Yann; Huc, Ivan;
Orchestrating Directional Molecular Motions: Kinetically Controlled Supramolecular Pathways of a Helical Host on Rodlike Guests
Journal of the American Chemical Society, 2017, 139, 9350-9358
An aromatic oligoamide sequence was designed to fold and self-assemble into a double helical host having a cylindrical Cavity complementary to linear oligocarbamate guests. Formation of helical pseudorotaxane complexes, foldaxanes, between the host and guests having binding stations of different affinities was evidenced by NMR and X-ray crystallography. Rodlike guests possessing two or three binding stations, long alkyl or oligoethylene glycol spacers or bulky barriers in-between the binding stations, and a single bulky stopper at one end were synthesized. Kinetic investigations of the threading and translation of the double helix along multistation rods were monitored by H-1 NMR Results show that multiple events may occur upon sliding Of the host from the nonbulky end of the rod to reach the thermodynamically most stable state before unfolding-mediated dissociation has time to take place, including binding on intermediate stations and rapid sliding along nonbinding spacers. Conversely, installing a kinetic barrier that blocks sliding allows for the deliberate integration of a helix dissociation reassociation step in the supramolecular trajectory. Typical sliding processes can be monitored over the course of hours whereas steps involving unwinding rewinding of the helix proceeded over the course of days. These results further demonstrate the interest in foldaxanes to design complex sequences of supramolecular events within networks of equilibria through the adjustment of the kinetics of the individual steps involved.
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Wadeesirisak, Kanthida; Castano, Sabine; Berthelot, Karine; Vaysse, Laurent; Bonfils, Frederic; Peruch, Frederic; Rattanaporn, Kittipong; Liengprayoon, Siriluck; Lecomte, Sophie; Bottier, Celine;
Rubber particle proteins REF1 and SRPP1 interact differently with native lipids extracted from Hevea brasiliensis latex
Biochimica Et Biophysica Acta-Biomembranes, 2017, 1859, 201-210
Rubber particle membranes from the Hevea latex contain predominantly two proteins, REF1 and SRPP1 involved in poly(cis-1,4-isoprene) synthesis or rubber quality. The repartition of both proteins on the small or large rubber particles seems to differ, but their role in the irreversible coagulation of the rubber particle is still unknown. In this study we highlighted the different modes of interactions of both recombinant proteins with different classes of lipids extracted from Hevea brasiliensis latex, and defined as phospholipids (PL), glycolipids (GL) and neutral lipids (NL). We combined two biophysical methods, polarization modulated-infrared reflection adsorption spectroscopy (PM-IRRAS) and ellipsometry to elucidate their interactions with monolayers of each class of lipids. REF1 and SRPP1 interactions with native lipids are clearly different; SRPP1 interacts mostly in surface with PI, GL or NL, without modification of its structure. In contrast REF1 inserts deeply in the lipid monolayers with all lipid classes. With NI, REF1 is even able to switch from alpha-helice conformation to beta-sheet structure, as in its aggregated form (amyloid form). Interaction between REF1 and NL may therefore have a specific role in the irreversible coagulation of rubber particles. (C) 2016 Elsevier B.V. All rights reserved.
5 […]
Vallet-Courbin, Amelie; Lariviere, Melusine; Hocquellet, Agnes; Hemadou, Audrey; Parimala, Sarjapura-Nagaraja; Laroche-Traineau, Jeanny; Santarelli, Xavier; Clofent-Sanchez, Gisele; Jacobin-Valat, Marie-Josee; Noubhani, Abdelmajid;
A Recombinant Human Anti-Platelet scFv Antibody Produced in Pichia pastoris for Atheroma Targeting
PloS one, 2017, 12, e0170305-e0170305
Cells of the innate and adaptive immune system are key factors in the progression of atherosclerotic plaque, leading to plaque instability and rupture, potentially resulting in acute atherothrombotic events such as coronary artery disease, cerebrovascular disease and peripheral arterial disease. Here, we describe the cloning, expression, purification, and immunoreactivity assessment of a recombinant single-chain variable fragment (scFv) derived from a human anti-alphaIIbbeta3 antibody (HuAb) selected to target atheromatous lesions for the presence of platelets. Indeed, platelets within atheroma plaques have been shown to play a role in inflammation, in platelet-leucocyte aggregates and in thrombi formation and might thus be considered relevant biomarkers of atherosclerotic progression. The DNA sequence that encodes the anti-alphaIIbbeta3 TEG4 scFv previously obtained from a phage-display selection on activated platelets, was inserted into the eukaryote vector (pPICZalphaA) in fusion with a tag sequence encoding 2 cysteines useable for specific probes grafting experiments. The recombinant protein was expressed at high yields in Pichia pastoris (30 mg/L culture). The advantage of P. pastoris as an expression system is the production and secretion of recombinant proteins in the supernatant, ruling out the difficulties encountered when scFv are produced in the cytoplasm of bacteria (low yield, low solubility and reduced affinity). The improved conditions allowed for the recovery of highly purified and biologically active scFv fragments ready to be grafted in a site-directed way to nanoparticles for the imaging of atherosclerotic plaques involving inflammatory processes and thus at high risk of instability.
6 […]
Smulski, Cristian R.; Decossas, Marion; Chekkat, Neila; Beyrath, Julien; Willen, Laure; Guichard, Gilles; Lorenzetti, Raquel; Rizzi, Marta; Eibel, Hermann; Schneider, Pascal; Fournel, Sylvie;
Hetero-oligomerization between the TNF receptor superfamily members CD40, Fas and TRAILR2 modulate CD40 signalling
Cell Death & Disease, 2017, 8,
TNF receptor superfamily members (TNFRSF) such as CD40, Fas and TRAIL receptor 2 (TRAILR2) participate to the adaptive immune response by eliciting survival, proliferation, differentiation and/or cell death signals. The balance between these signals determines the fate of the immune response. It was previously reported that these receptors are able to self-assemble in the absence of ligand through their extracellular regions. However, the role of this oligomerization is not well understood, and none of the proposed hypotheses take into account potential hetero-association of receptors. Using CD40 as bait in a flow cytometry Forster resonance energy transfer assay, TNFRSF members with known functions in B cells were probed for interactions. Both Fas and TRAILR2 associated with CD40. Immunoprecipitation experiments confirmed the interaction of CD40 with Fas at the endogenous levels in a BJAB B-cell lymphoma cell line deficient for TRAILR2. TRAILR2-expressing BJAB cells displayed a robust CD40-TRAILR2 interaction at the expense of the CD40-Fas interaction. The same results were obtained by proximity ligation assay, using TRAILR2-positive and -negative BJAB cells and primary human B cells. Expression of the extracellular domains of Fas or TRAILR2 with a glycolipid membrane anchor specifically reduced the intrinsic signalling pathway of CD40 in 293T cells. Conversely, BJAB cells lacking endogenous Fas or TRAILR2 showed an increased NF-kappa B response to CD40L. Finally, upregulation of TRAILR2 in primary human B cells correlated with reduced NF-kappa B activation and reduced proliferation in response to CD40L. Altogether, these data reveal that selective interactions between different TNFRSF members may modulate ligand-induced responses upstream signalling events.
7 […]
Ryu, Naoya; Okazaki, Yutaka; Pouget, Emilie; Takafuji, Makoto; Nagaoka, Shoji; Ihara, Hirotaka; Oda, Reiko;
Fluorescence emission originated from the H-aggregated cyanine dye with chiral gemini surfactant assemblies having a narrow absorption band and a remarkably large Stokes shift
Chemical Communications, 2017, 53, 8870-8873
We demonstrate that the fluorescence emission that originated from a cyanine dye forming H-aggregates with a narrow absorption band and a remarkably large Stokes shift can be induced by symmetry breaking. The H-aggregate formation was successfully induced using chirally assembled cationic gemini surfactants with enantiomeric tartrate counterions as templates in water.
8 […]
Rombaut, Natacha; Savoire, Raphaelle; Van Hecke, Elisabeth; Thomasset, Brigitte;
Supercritical CO2 extraction of linseed: Optimization by experimental design with regards to oil yield and composition
European Journal of Lipid Science and Technology, 2017, 119,
The aim of the present study is to evaluate the effect of CO2 pressure, temperature, and CO2 flow rate on supercritical fluid extraction of linseed oil using neat CO2. Chosen methodology is based on central composite rotatable design, responses studied are extract yield, oil yield, water-extract ratio, oil -tocopherol content, oil polyphenols content, alpha linolenic acid proportion in oil, and oil acid value. Acid value, tocopherol content, and linolenic acid proportion are independent responses of studied factors. On other responses, only pressure and temperature have an influence. Based on response surface analysis and multiresponses optimization, optimal conditions for oil yield, and oil polyphenol content maximization and water-extract minimization were determined (42.5MPa, 120 degrees C, 12.5kg/h [125g/g solvent:feed ratio]). Optimal conditions were tested and validated on three linseed varieties. A maximal oil yield of approximate to 60% was reached containing up to 100mg FAE/kg oil. Practical applications: Results of this study can be applied for developing a new linseed oil extraction process based on supercritical fluid extraction without co-solvent addition. The optimized process permits an increased extraction of antioxidants. Extraction of antioxidants such as -tocopherol or polyphenols could provide a protection of oil against oxidation. This protection is of great interest for sensitive oils such as the highly polyunsaturated linseed oil. Application to other oleaginous seeds is also possible. Supercritical CO2 extraction of linseed oil is optimized through design of experiment with regards to oil yield and oil quality. Optimization enable to identify conditions (42.5MPa, 120 degrees C) that favors oil yield (approximate to 60%) while enhancing oil polyphenols content (50-100mg FAE/kg). Extraction performances are variety dependent.
9 […]
Reiner, Agnes T.; Tan, Sisareuth; Agreiter, Christiane; Auer, Katharina; Bachmayr-Heyda, Anna; Aust, Stefanie; Pecha, Nina; Mandorfer, Mattias; Pils, Dietmar; Brisson, Alain R.; Zeillinger, Robert; Lim, Sai Kiang;
EV-Associated MMP9 in High-Grade Serous Ovarian Cancer Is Preferentially Localized to Annexin V-Binding EVs
Disease markers, 2017, 2017, 9653194-9653194
High-grade serous ovarian cancer (HGSOC) is the most aggressive type of ovarian cancer and is responsible for most deaths caused by gynecological cancers. Numerous candidate biomarkers were identified for this disease in the last decades, but most were not sensitive or specific enough for clinical applications. Hence, new biomarkers for HGSOC are urgently required. This study aimed to identify new markers by isolating different extracellular vesicle (EV) types from the ascites of ovarian cancer patients according to their affinities for lipid-binding proteins and analyzing their protein cargo. This approach circumvents the low signal-to-noise ratio when using biological fluids for biomarker discovery and the issue of contamination by large non-EV complexes. We isolated and analyzed three distinct EV populations from the ascites of patients with ovarian cancer or cirrhosis and observed that Annexin V-binding EVs have higher levels of matrix metalloproteinase 9 in malignant compared to portal-hypertensive ascites. As this protein was not detected in other EV populations, this study validates our approach of using different EV types for optimal biomarker discovery. Furthermore, MMP9 in Annexin V-binding EVs could be a HGSOC biomarker with enhanced specificity, because its identification requires detection of two distinct components, that is, lipid and protein.
10 […]
Ravaud, Alain; de la Fouchardiere, Christelle; Caron, Philippe; Doussau, Adelaide; Do Cao, Christine; Asselineau, Julien; Rodien, Patrice; Pouessel, Damien; Nicolli-Sire, Patricia; Klein, Marc; Bournaud-Salinas, Claire; Wemeau, Jean-Louis; Gimbert, Anne; Picat, Marie-Quitterie; Pedenon, Delphine; Digue, Laurence; Daste, Amaury; Catargi, Bogdan; Delord, Jean-Pierre;
A multicenter phase II study of sunitinib in patients with locally advanced or metastatic differentiated, anaplastic or medullary thyroid carcinomas: mature data from the THYSU study
European Journal of Cancer, 2017, 76, 110-117
Purpose: Patients with advanced radioactive iodine resistant differentiated (MDTC) or medullary (MMTC) thyroid cancer had an unmet need. Early data showed promising efficacy of vascular endothelial growth factor receptor inhibitors. We investigated sunitinib in this setting. Patients and methods: This phase 2 trial enrolled MDTC, anaplastic (MATC) and MMTC patients in 1st line anti-angiogenic therapy with sunitinib at 50 mg/d, 4/6w. Objective response rate was the primary end-point. Secondary end-points were progression-free survival, overall survival and safety. Results: Seventy-one patients were enrolled from August 2007 to October 2009, 41 MDTC/4 MATC patients and 26 MMTC patients. Patients received a median of 8 and 9 cycles, respectively. In the MDTC/MATC group, 13% of patients and 43% of cycles and in the MMTC group, 23% of the patients and 48.8% of cycles remained at 50 mg/d, respectively. The primary end-point was reached with an objective response rate of 22% (95% Cl: 10.6-37.6) in MDTC patients and in 38.5% (95% CI: 22.6-56.4) in MMTC patients. No objective response was seen in MATC patients. Median progression-free survival and overall survival were 13.1 and 26.4 months in MDTC patients, 16.5 and 29.4 months in MMTC patients. The most frequent side effects were asthenia/fatigue (27.8% > grade 3), mucosal (9.9% > grade 3), cutaneous toxicities, hand-foot syndrome (18.3% > grade 3). Of all, 14.1% had a cardiac event. Nine unexpected side effects were reported, out of which, five induced deaths. Conclusion: Sunitinib is active in MDTC and MMTC patients. Side effects were more severe than with previous reports. If using sunitinib, alternative schedule/dosage should be considered. (C) 2017 Elsevier Ltd. All rights reserved.
11 […]
Petitdemange, Rosine; Garanger, Elisabeth; Bataille, Laure; Dieryck, Wilfrid; Bathany, Katell; Garbay, Bertrand; Deming, Timothy J.; Lecommandoux, Sebastien;
Selective Tuning of Elastin-like Polypeptide Properties via Methionine Oxidation
Biomacromolecules, 2017, 18, 544-550
We have designed and prepared a recombinant elastin-like polypeptide (ELP) containing precisely positioned methionine residues, and performed the selective and complete oxidation of its methionine thioether groups to both sulfoxide and sulfone derivatives. Since these oxidation reactions substantially increase methionine residue polarity, they were found to be a useful means to precisely adjust the temperature responsive behavior of ELPs in aqueous solutions. In particular, lower critical solution temperatures were found to be elevated in oxidized sample solutions, but were not eliminated. These transition temperatures were found to be further tunable by the use of solvents containing different Hofmeister salts. Overall, the ability to selectively and fully oxidize methionine residues in ELPs proved to be a convenient postmodification strategy for tuning their transition temperatures in aqueous media.
12 […]
Petitdemange, Rosine; Garanger, Elisabeth; Bataille, Laure; Bathany, Katell; Garbay, Bertrand; Deming, Timothy J.; Lecommandoux, Sebastien;
Tuning Thermoresponsive Properties of Cationic Elastin-like Polypeptides by Varying Counterions and Side-Chains
Bioconjugate chemistry, 2017, 0,
We report the synthesis of methionine-containing recombinant elastin-like polypeptides (ELPs) of different lengths that contain periodically spaced methionine residues. These ELPs were chemoselectively alkylated at all methionine residues to give polycationic derivatives. Some of these samples were found to possess solubility transitions in water, where the temperature of these transitions varied with ELP concentration, nature of the methionine alkylating group, and nature of the sulfonium counterions. These studies show that introduction and controlled spacing of methionine sulfonium residues into ELPs can be used as a means both to tune their solubility transition temperatures in water using a variety of different parameters and to introduce new side-chain functionality.
13 […]
Neal, Bruce; Perkovic, Vlado; Mahaffey, Kenneth W.; de Zeeuw, Dick; Fulcher, Greg; Erondu, Ngozi; Shaw, Wayne; Law, Gordon; Desai, Mehul; Matthews, David R.; Catargi, B.; Canvas Program Collaborative Grp;
Canagliflozin and Cardiovascular and Renal Events in Type 2 Diabetes
New England Journal of Medicine, 2017, 377, 644-657
BACKGROUND Canagliflozin is a sodium-glucose cotransporter 2 inhibitor that reduces glycemia as well as blood pressure, body weight, and albuminuria in people with diabetes. We report the effects of treatment with canagliflozin on cardiovascular, renal, and safety outcomes. METHODS The CANVAS Program integrated data from two trials involving a total of 10,142 participants with type 2 diabetes and high cardiovascular risk. Participants in each trial were randomly assigned to receive canagliflozin or placebo and were followed for a mean of 188.2 weeks. The primary outcome was a composite of death from cardiovascular causes, nonfatal myocardial infarction, or nonfatal stroke. RESULTS The mean age of the participants was 63.3 years, 35.8% were women, the mean duration of diabetes was 13.5 years, and 65.6% had a history of cardiovascular disease. The rate of the primary outcome was lower with canagliflozin than with placebo (occurring in 26.9 vs. 31.5 participants per 1000 patient-years; hazard ratio, 0.86; 95% confidence interval [CI], 0.75 to 0.97; P< 0.001 for noninferiority; P = 0.02 for superiority). Although on the basis of the prespecified hypothesis testing sequence the renal outcomes are not viewed as statistically significant, the results showed a possible benefit of canagliflozin with respect to the progression of albuminuria (hazard ratio, 0.73; 95% CI, 0.67 to 0.79) and the composite outcome of a sustained 40% reduction in the estimated glomerular filtration rate, the need for renal-replacement therapy, or death from renal causes (hazard ratio, 0.60; 95% CI, 0.47 to 0.77). Adverse reactions were consistent with the previously reported risks associated with canagliflozin except for an increased risk of amputation (6.3 vs. 3.4 participants per 1000 patient-years; hazard ratio, 1.97; 95% CI, 1.41 to 2.75); amputations were primarily at the level of the toe or metatarsal. CONCLUSIONS In two trials involving patients with type 2 diabetes and an elevated risk of cardiovascular disease, patients treated with canagliflozin had a lower risk of cardiovascular events than those who received placebo but a greater risk of amputation, primarily at the level of the toe or metatarsal.
14 […]
Mendoza, Oscar; Houmadi, Said; Aime, Jean-Pierre; Elezgaray, Juan;
Signal replication in a DNA nanostructure
Journal of Chemical Physics, 2017, 146,
Logic circuits based on DNA strand displacement reaction are the basic building blocks of future nanorobotic systems. The circuits tethered to DNA origami platforms present several advantages over solution-phase versions where couplings are always diffusion-limited. Here we consider a possible implementation of one of the basic operations needed in the design of these circuits, namely, signal replication. We showthat with an appropriate preparation of the initial state, signal replication performs in a reproducible way. We also show the existence of side effects concomitant to the high effective concentrations in tethered circuits, such as slow leaky reactions and cross-activation. Published by AIP Publishing.
15 […]
Mendoza, O.; Calmet, P.; Alves, I.; Lecomte, S.; Raoux, M.; Cullin, C.; Elezgaray, J.;
A tensegrity driven DNA nanopore
Nanoscale, 2017, 9, 9762-9769
Control of transport across membranes, whether natural or synthetic, is fundamental in many biotechnology applications, including sensing and drug release. Mutations of naturally existing protein channels, such as hemolysin, have been explored in the past. More recently, DNA channels with conductivities in the nanosiemens range have been designed. Regulating transport across DNA channels in response to external stimuli remains an important challenge. Previous designs relied on steric hindrance to control the inner diameter of the channel, which resulted in unstable electric signatures. In this paper we introduce a new design to control electric channel conductance of a DNA nanopore. The tensegrity driven mechanism inhibits the flux of small analytes while keeping a tightly controlled ionic transport modulated by the addition of specific DNA sequences. Current signals are clearly defined, with no sign of gating, opening new perspectives in single molecule DNA sensing.
16 […]
Mateus, Pedro; Wicher, Barbara; Ferrand, Yann; Huc, Ivan;
Alkali and alkaline earth metal ion binding by a foldamer capsule: selective recognition of magnesium hydrate
Chemical Communications, 2017, 53, 9300-9303
Alkali and alkaline earth metal ion binding by an aromatic oligoamide foldamer was shown to induce its folding into a helical capsule. CD and NMR titrations revealed tight and selective binding of Mg2+. Crystallographic studies demonstrated that, depending on the metal, binding may involve the first or second coordination spheres of the metal hydrates.
17 […]
Martinez, Denis; Langlois d'Estaintot, Beatrice; Granier, Thierry; Tolchard, James; Courreges, Cecile; Prouzet-Mauleon, Valerie; Hugues, Michel; Gallois, Bernard; Doignon, Francois; Odaert, Benoit;
Structural evidence of a phosphoinositide-binding site in the Rgd1-RhoGAP domain
The Biochemical journal, 2017, 474, 3307-3319
Phosphoinositide lipids recruit proteins to the plasma membrane involved in the regulation of cytoskeleton organization and in signalling pathways that control cell polarity and growth. Among those, Rgd1p is a yeast GTPase-activating protein (GAP) specific for Rho3p and Rho4p GTPases, which control actin polymerization and stress signalling pathways. Phosphoinositides not only bind Rgd1p, but also stimulate its GAP activity on the membrane-anchored form of Rho4p. Both F-BAR (F-BAR FCH, and BAR) and RhoGAP domains of Rgd1p are involved in lipid interactions. In the Rgd1p-F-BAR domain, a phosphoinositide-binding site has been recently characterized. We report here the X-ray structure of the Rgd1p-RhoGAP domain, identify by NMR spectroscopy and confirm by docking simulations, a new but cryptic phosphoinositide-binding site, comprising contiguous A1, A1' and B helices. The addition of helix A1', unusual among RhoGAP domains, seems to be crucial for lipid interactions. Such a site was totally unexpected inside a RhoGAP domain, as it was not predicted from either the protein sequence or its three-dimensional structure. Phosphoinositide-binding sites in RhoGAP domains have been reported to correspond to polybasic regions, which are located at the unstructured flexible termini of proteins. Solid-state NMR spectroscopy experiments confirm the membrane interaction of the Rgd1p-RhoGAP domain upon the addition of PtdIns(4,5)P2 and indicate a slight membrane destabilization in the presence of the two partners.
18 […]
Martinez, Denis; Decossas, Marion; Kowal, Julia; Frey, Lukas; Stahlberg, Henning; Dufourc, Erick J.; Riek, Roland; Habenstein, Birgit; Bibow, Stefan; Loquet, Antoine;
Lipid Internal Dynamics Probed in Nanodiscs
Chemphyschem : a European journal of chemical physics and physical chemistry, 2017, 0,
Nanodiscs offer a very promising tool to incorporate membrane proteins into native-like lipid bilayers and an alternative to liposomes to maintain protein functions and protein-lipid interactions in a soluble nanoscale object. The activity of the incorporated membrane protein appears to be correlated to its dynamics in the lipid bilayer and by protein-lipid interactions. These two parameters depend on the lipid internal dynamics surrounded by the lipid-encircling discoidal scaffold protein that might differ from more unrestricted lipid bilayers observed in vesicles or cellular extracts. A solid-state NMR spectroscopy investigation of lipid internal dynamics and thermotropism in nanodiscs is reported. The gel-to-fluid phase transition is almost abolished for nanodiscs, which maintain lipid fluid properties for a large temperature range. The addition of cholesterol allows fine-tuning of the internal bilayer dynamics by increasing chain ordering. Increased site-specific order parameters along the acyl chain reflect a higher internal ordering in nanodiscs compared with liposomes at room temperature; this is induced by the scaffold protein, which restricts lipid diffusion in the nanodisc area.
19 […]
Lozan, Ecaterina; Shinkaruk, Svitlana; Al Abed, Shaam Alice; Lamothe, Valerie; Potier, Mylene; Marighetto, Aline; Schmitter, Jean-Marie; Bennetau-Pelissero, Catherine; Bure, Corinne;
Derivatization-free LC-MS/MS method for estrogen quantification in mouse brain highlights a local metabolic regulation after oral versus subcutaneous administration
Analytical and Bioanalytical Chemistry, 2017, 409, 5279-5289
17 beta-Estradiol (17 beta-E-2) is a steroid with pleiotropic actions. In addition to being a sexual hormone, it is also produced in the brain where it modulates the reproductive axis. It has been shown that 17 beta-E-2 also acts on synaptic plasticity and plays a role in neurological pathways and in neurodegenerative diseases. Assaying this steroid in the brain is thus interesting to improve our knowledge of 17 beta-E-2 effects in the brain. However, 17 beta-E-2 concentration in the central nervous system has been reported to be of a few nanograms per gram wet weight (nanomolar range concentration); therefore, its quantification requires both an efficient extraction process and a sensitive detection method. Herein is presented a derivatization-free procedure based on solid-phase extraction followed by LC-MS/MS analysis, targeted on 17 beta-E-2, its isomer17 alpha-E-2, and its metabolites estrone (E-1) and estriol (E-3). This extraction process allowed reaching 96% 17 beta-E-2 recovery from the mouse brain. Limit of detection (LOD) and limit of quantification (LOQ) values of 0.5 and 2.5 pmol mL(-1), respectively, were reached for both 17 alpha-E-2 and 17 beta-E-2. LOD values for E-1 and E-3 were 0.01 and 0.025 pmol mL(-1), respectively. The variation coefficients for intra- and inter-assays were 6 and 14%, respectively, for both estradiol forms. The method was applied to assess estrogen levels in the mouse brain and hippocampus after 17 beta-E-2 acute (subcutaneous injection) and chronic (drinking water) physiological administration. Total estrogen levels were determined after enzymatic deconjugation and compared to free estrogen levels. While 17 alpha-E-2 was not detected in biological samples, 17 beta-E-2 and metabolite measurements highlight a local biotransformation of estrogens after physiological administration via drinking water.
20 […]
Loquet, Antoine; Saupe, Sven J.;
Diversity of Amyloid Motifs in NLR Signaling in Fungi
Biomolecules, 2017, 7,
Amyloid folds not only represent the underlying cause of a large class of human diseases but also display a variety of functional roles both in prokaryote and eukaryote organisms. Among these roles is a recently-described activity in signal transduction cascades functioning in host defense and programmed cell death and involving Nod-like receptors (NLRs). In different fungal species, prion amyloid folds convey activation signals from a receptor protein to an effector domain by an amyloid templating and propagation mechanism. The discovery of these amyloid signaling motifs derives from the study of [Het-s], a fungal prion of the species Podospora anserina. These signaling pathways are typically composed of two basic components encoded by adjacent genes, the NLR receptor bearing an amyloid motif at the N-terminal end and a cell death execution protein with a HeLo pore-forming domain bearing a C-terminal amyloid motif. Activation of the NLR receptor allows for amyloid folding of the N-terminal amyloid motifs which then template trans-conformation of the homologous motif in the cell death execution protein. A variety of such motifs, which differ by their sequence signature, have been described in fungi. Among them, the PP-motif bears resemblance with the RHIM amyloid motif involved in the necroptosis pathway in mammals suggesting an evolutionary conservation of amyloid signaling from fungi to mammals.
21 […]
Liu, Z. W.; Hu, X. B.; Abramyan, A. M.; Meszaros, A.; Csekei, M.; Kotschy, A.; Huc, I.; Pophristic, V.;
Computational Prediction and Rationalization, and Experimental Validation of Handedness Induction in Helical Aromatic Oligoamide Foldamers
Chemistry-a European Journal, 2017, 23, 3605-3615
Metadynamics simulations were used to describe the conformational energy landscapes of several helically folded aromatic quinoline carboxamide oligomers bearing a single chiral group at either the C or Nterminus. The calculations allowed the prediction of whether a helix handedness bias occurs under the influence of the chiral group and gave insight into the interactions (sterics, electrostatics, hydrogen bonds) responsible for a particular helix sense preference. In the case of camphanyl-based and morpholine-based chiral groups, experimental data confirming the validity of the calculations were already available. New chiral groups with a proline residue were also investigated and were predicted to induce handedness. This prediction was verified experimentally through the synthesis of proline-containing monomers, their incorporation into an oligoamide sequence by solid phase synthesis and the investigation of handedness induction by NMR spectroscopy and circular dichroism.
22 […]
Linares, Romain; Tan, Sisareuth; Gounou, Celine; Brisson, Alain R.;
Imaging and Quantification of Extracellular Vesicles by Transmission Electron Microscopy
Methods in molecular biology (Clifton, N.J.), 2017, 1545, 43-54
Extracellular vesicles (EVs) are cell-derived vesicles that are present in blood and other body fluids. EVs raise major interest for their diverse physiopathological roles and their potential biomedical applications. However, the characterization and quantification of EVs constitute major challenges, mainly due to their small size and the lack of methods adapted for their study. Electron microscopy has made significant contributions to the EV field since their initial discovery. Here, we describe the use of two transmission electron microscopy (TEM) techniques for imaging and quantifying EVs. Cryo-TEM combined with receptor-specific gold labeling is applied to reveal the morphology, size, and phenotype of EVs, while their enumeration is achieved after high-speed sedimentation on EM grids.
23 […]
Legrand, Bernard; Salvetat, Jean-Paul; Walter, Benjamin; Faucher, Marc; Theron, Didier; Aime, Jean-Pierre;
Multi-MHz micro-electro-mechanical sensors for atomic force microscopy
Ultramicroscopy, 2017, 175, 46-57
Silicon ring-shaped micro-electro-mechanical resonators have been fabricated and used as probes for dynamic atomic force microscopy (AFM) experiments. They offer resotnance frequency above 10MHz, which is notably greater than that of usual cantilevers and quartz-based AFM probes. On-chip electrical actuation and readout of the tip oscillation are obtained by means of built-in capacitive transducers. Displacement and force resolutions have been determined from noise analysis at 1.5fm/Hz and 0.4 pN/Hz, respectively. Despite the high effective stiffness of the probes, the tip-surface interaction force is kept below 1 nN by using vibration amplitude significantly below 100pm and setpoint close to the free vibration conditions. Imaging capabilities in amplitude- and frequency-modulation AFM modes have been demonstrated on block copolymer surfaces. Z-spectroscopy experiments revealed that the tip is vibrating in permanent contact with the viscoelastic material, with a pinned contact line. Results are compared to those obtained with commercial AFM cantilevers driven at large amplitudes (>10nm).
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LEGERON, Rachel; XUEREB, Fabien; BREILH, Dominique; GADEAU, Alain Pierrre; BOIRON, Jean-Michel; SCHMITTER, Jean-Marie; DUPUY, Jean-William; CHAIGNEPAIN, Stéphane;
, 2017, 0,
The present invention relates to the use of an Internal Standard compound in a method for quantifying Bevacizumab in a sample by mass spectrometry, wherein the said Internal Standard compound is described in the specification.
25 […]
Leger, Antoine; Hocquellet, Agnes; Dieryck, Wilfrid; Moine, Virginie; Marchal, Axel; Marullo, Philippe; Josseaume, Annabelle; Cabanne, Charlotte;
Production and Purification of the Native Saccharomyces cerevisiae Hsp12 in Escherichia coli
Journal of agricultural and food chemistry, 2017, 65, 8154-8161
Hsp12 is a small heat shock protein produced in many organisms, including the yeast Saccharomyces cerevisiae. It has been described as an indicator of yeast stress rate and has also been linked to the sweetness sensation of wine. To obtain a sufficient amount of protein, we produced and purified Hsp12 without tag in Escherichia coli. A simple fast two-step process was developed using a microplate approach and a design of experiments. A capture step on an anion-exchange salt-tolerant resin was followed by size exclusion chromatography for polishing, leading to a purity of 97%. Thereafter, specific anti-Hsp12 antibodies were obtained by rabbit immunization. An ELISA was developed to quantify Hsp12 in various strains of Saccharomyces cerevisiae. The antibodies showed high specificity and allowed the quantitation of Hsp12 in the yeast. The quantities of Hsp12 measured in the strains differed in direct proportion to the level of expression found in previous studies.
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Lefevre, M.; Racedo, S. M.; Denayrolles, M.; Ripert, G.; Desfougeres, T.; Lobach, A. R.; Simon, R.; Pelerin, F.; Justen, P.; Urdaci, M. C.;
Safety assessment of Bacillus subtilis CU1 for use as a probiotic in humans
Regulatory Toxicology and Pharmacology, 2017, 83, 54-65
Bacillus subtilis CU1 is a recently described probiotic strain with beneficial effects on immune health in elderly subjects. The following work describes a series of studies supporting the safety of the strain for use as an ingredient in food and supplement preparations. Using a combination of 16S rDNA and gyrB nucleotide analyses, the species was identified as a member of the Bacillus subtilis complex (B. subtilis subsp. spizizenii). Further characterization of the organism at the strain level was achieved using random amplified polymorphic DNA polymerase chain reaction (RAPD PCR) and pulsed field gel electrophoresis (PFGE) analyses. B. subtilis CU1 did not demonstrate antibiotic resistance greater than existing regulatory cutoffs against clinically important antibiotics, did not induce hemolysis or produce surfactant factors, and was absent of toxigenic activity in vitro. Use of B. subtilis CU1 as a probiotic has recently been evaluated in a 16-week randomized, double-blind, placebo-controlled, parallel-arm study, in which 2 x 10(9) spores per day of B. subtilis CU1 were administered for a total 40 days to healthy elderly subjects (4 consumption periods of 10 days separated by 18-day washouts). This work describes safety related endpoints not previously reported. B. subtilis CU1 was safe and well-tolerated in the clinical subjects without undesirable physiological effects on markers of liver and kidney function, complete blood counts, hemodynamic parameters, and vital signs. (C) 2016 The Authors. Published by Elsevier Inc.
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Larrieu, Isabelle; Tolchard, James; Sanchez, Corinne; Kone, Edmond Yazo; Barras, Alexandre; Stines-Chaumeil, Claire; Odaert, Benoit; Giraud, Marie-France;
Cell-Free Expression for the Study of Hydrophobic Proteins: The Example of Yeast ATP-Synthase Subunits
Methods in molecular biology (Clifton, N.J.), 2017, 1635, 57-90
Small hydrophobic membrane proteins or proteins with hydrophobic domains are often difficult to produce in bacteria. The cell-free expression system was found to be a very good alternative for the expression of small hydrophobic subunits of the yeast ATP-synthase, such as subunits e, g, k, i, f and the membrane domain of subunit 4, proteins that are suspected to play a role in the stability of ATP-synthase dimers. All of these proteins could be produced in milligrams amounts using the cell-free "precipitate mode" and were successfully solubilized in the presence of lysolipid 1-myristoyl-2-hydroxy-sn-glycero-3-phospho-1'-rac-glycerol. Purified proteins were also found suitable for structural investigations. An example is given with the NMR backbone assignment of the isotopically labeled subunit g. Protocols are also described for raising specific polyclonal antibodies against overexpressed cell-free proteins.
28 […]
Koutsouras, Dimitrios A.; Perrier, Romain; Marquez, Ariana Villarroel; Pirog, Antoine; Pedraza, Eileen; Cloutet, Eric; Renaud, Sylvie; Raoux, Matthieu; Malliaras, George G.; Lang, Jochen;
Simultaneous monitoring of single cell and of micro-organ activity by PEDOT:PSS covered multi-electrode arrays
Materials Science & Engineering C-Materials for Biological Applications, 2017, 81, 84-89
Continuous and long-term monitoring of cellular and micro-organ activity is required for new insights into physiology and novel technologies such as Organs-on-Chip. Moreover, recent advances in stem cell technology and especially in the field of diabetes call for non-invasive approaches in quality testing of the large quantities of surrogate pancreatic islets to be generated. Electrical activity of such a micro-organ results in single cell action potentials (APs) of high frequency and in low frequency changes in local field potentials (slow potentials or SPs), reflecting coupled cell activity and overall organ physiology. Each of them is indicative of different physiological stages in islet activation. Action potentials in islets are of small amplitude and very difficult to detect. The use of PEDOT:PSS to coat metal electrodes is expected to reduce noise and results in a frequency-dependent change in impedance, as shown here. Whereas detection of high-frequency APs improves, low frequency SPs are less well detected which is, however, an acceptable trade off in view of the strong amplitude of SPs. Using a dedicated software, recorded APs and SPs can be automatically diagnosed and analyzed. Concomitant capture of the two signals will considerably increase the diagnostic power of monitoring islets and islet surrogates in fundamental research as well as drug screening or the use of islets as biosensors.
29 […]
Kamgar-Parsi, Kian; Tolchard, James; Habenstein, Birgit; Loquet, Antoine; Naito, Akira; Ramamoorthy, Ayyalusamy;
Structural Biology of Calcitonin: From Aqueous Therapeutic Properties to Amyloid Aggregation
Israel Journal of Chemistry, 2017, 57, 634-650
Under appropriate conditions, peptides and proteins can assemble from their native state into prefibrillar oligomers and then mature into fibrillar aggregates. This transition forms the molecular basis of several pathologies, often related to the deposition of these amyloid fibrils. Several hormone peptides involved in fundamental biological processes have the tendency to self-assemble into amyloid fibrils, resulting in a loss of their native functions, and more importantly, entailing devastating consequences, such as the formation of amyloid depositions. Calcitonin is a 32 amino-acid hormone peptide that can be considered a molecular paradigm for the central events associated with hormone misfolding. Calcitonin in its native form is involved in various physiological functions, including mediating calcium homeostasis and maintaining bone structure. It is the latter function that has motivated the use of calcitonin as an aqueous therapeutic agent for the treatment of bone-related pathologies such as osteoporosis and Paget's disease. Despite some success as a therapeutic, calcitonin's ability to control these diseases is limited by its aggregation along the canonical amyloid aggregation pathway, compromising its long-term stability as a therapeutic agent. A better understanding of the misfolding process would not only provide the structural basis to improve calcitonin's long-term stability and activity as a therapeutic, but also provide valuable insights into pathological aggregation of other amyloids. In this work, we review the physiological roles of calcitonin, its structure, and aggregation process, and consider the effects of calcitonin's structure on its role as a therapeutic.
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Joly, P.; Vignaud, H.; Di Martino, J.; Ruiz, M.; Garin, R.; Restier, L.; Belmalih, A.; Marchal, C.; Cullin, C.; Arveiler, B.; Fergelot, P.; Gitler, A. D.; Lachaux, A.; Couthouis, J.; Bouchecareilh, M.;
y ERAD defects and the HFE-H63D variant are associated with increased risk of liver damages in Alpha 1-Antitrypsin Deficiency
Plos One, 2017, 12,
Background The most common and severe disease causing allele of Alpha 1-Antitrypsin Deficiency (1ATD) is Z-1AT. This protein aggregates in the endoplasmic reticulum, which is the main cause of liver disease in childhood. Based on recent evidences and on the frequency of liver disease occurrence in Z-1AT patients, it seems that liver disease progression is linked to still unknown genetic factors. Methods We used an innovative approach combining yeast genetic screens with next generation exome sequencing to identify and functionally characterize the genes involved in 1ATD associated liver disease. Results Using yeast genetic screens, we identified HRD1, an Endoplasmic Reticulum Associated Degradation (ERAD) associated protein, as an inducer of Z-mediated toxicity. Whole exome sequencing of 1ATD patients resulted in the identification of two variants associated with liver damages in Z-1AT homozygous cases: HFE H63D and HERPUD1 R50H. Functional characterization in Z-1AT model cell lines demonstrated that impairment of the ERAD machinery combined with the HFE H63D variant expression decreased both cell proliferation and cell viability, while Unfolded Protein Response (UPR)-mediated cell death was hyperstimulated. Conclusion This powerful experimental pipeline allowed us to identify and functionally validate two genes involved in Z-1AT-mediated severe liver toxicity. This pilot study moves forward our understanding on genetic modifiers involved in 1ATD and highlights the UPR pathway as a target for the treatment of liver diseases associated with 1ATD. Finally, these findings support a larger scale screening for HERPUD1 R50H and HFE H63D variants in the sub-group of 1ATD patients developing significant chronic hepatic injuries (hepatomegaly, chronic cholestasis, elevated liver enzymes) and at risk developing liver cirrhosis.
31 […]
Jiao, C. Y.; Sachon, E.; Alves, I. D.; Chassaing, G.; Bolbach, G.; Sagan, S.;
Exploiting Benzophenone Photoreactivity To Probe the Phospholipid Environment and Insertion Depth of the Cell-Penetrating Peptide Penetratin in Model Membranes
Angew Chem Int Ed Engl, 2017, 0,
Penetratin (RQIKIWFQNRRMKWKK) enters cells by different mechanisms, including membrane translocation, thus implying that the peptide interacts with the lipid bilayer. Penetratin also crosses the membrane of artificial vesicles, depending on their phospholipid content. To evaluate the phospholipid preference of penetratin, as the first step of translocation, we exploited the benzophenone triplet kinetics of hydrogen abstraction, which is slower for secondary than for allylic hydrogen atoms. By using multilamellar vesicles of varying phospholipid content, we identified and characterized the cross-linked products by MALDI-TOF mass spectrometry. Penetratin showed a preference for negatively charged (vs. zwitterionic) polar heads, and for unsaturated (vs. saturated) and short (vs. long) saturated phospholipids. Our study highlights the potential of using benzophenone to probe the environment and insertion depth of membranotropic peptides in membranes.
32 […]
Jewginski, Michal; Granier, Thierry; Langlois d'Estaintot, Beatrice; Fischer, Lucile; Mackereth, Cameron D.; Huc, Ivan;
Self-Assembled Protein-Aromatic Foldamer Complexes with 2:3 and 2:2:1 Stoichiometries
Journal of the American Chemical Society, 2017, 139, 2928-2931
The promotion of protein dimerization using the aggregation properties of a protein ligand was explored and shown to produce complexes with unusual stoichiometries. Helical foldamer 2 was synthesized and bound to human carbonic anhydrase (HCA) using a nanomolar active site ligand. Crystal structures show that the hydrophobicity of 2 and interactions of its side chains lead to the formation of an HCA2-23 complex in which three helices of 2 are stacked, two of them being linked to an HCA molecule. The middle foldamer in the stack can be replaced by alternate sequences 3 or 5. Solution studies by CD and NMR confirm left-handedness of the helical foldamers as well as HCA dimerization.
33 […]
Hu, X. B.; Dawson, S. J.; Mandal, P. K.; de Hatten, X.; Baptiste, B.; Huc, I.;
Optimizing side chains for crystal growth from water: a case study of aromatic amide foldamers
Chemical Science, 2017, 8, 3741-3749
The growth of crystals of aromatic compounds from water much depends on the nature of the water solubilizing functions that they carry. Rationalizing crystallization from water, and structure elucidation, of aromatic molecular and supramolecular systems is of general value across various fields of chemistry. Taking helical aromatic foldamers as a test case, we have validated several short polar side chains as efficient substituents to provide both solubility in, and crystal growth ability from, water. New 8-amino-2-quinolinecarboxylic acids bearing charged or neutral aminomethyl, carboxymethyl, sulfonic acid, or bis(hydroxymethyl)-methoxy side chains in position 4 or 5, were prepared on a multi gram scale. Fmoc protection of the main chain amine and suitable protections of the side chains ensured compatibility with solid phase synthesis. One tetrameric and five octameric oligoamides displaying these side chains were synthesized and shown to be soluble in water. In all cases but one, crystals were obtained using the hanging drop method, thus validating the initial design principle to combine polarity and rigidity. The only case that resisted crystallization appeared to be due to exceedingly high water solubility endowed by eight sulfonic acid functions. The neutral side chain did provide crystal growth ability from water but contributed poorly to solubility.
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Horeau, Maxime; Lautrette, Guillaume; Wicher, Barbara; Blot, Virginie; Lebreton, Jacques; Pipelier, Muriel; Dubreuil, Didier; Ferrand, Yann; Huc, Ivan;
Metal-Coordination-Assisted Folding and Guest Binding in Helical Aromatic Oligoamide Molecular Capsules
Angewandte Chemie-International Edition, 2017, 56, 6823-6827
The development of foldamer-based receptors is driven by the design of monomers with specific properties. Herein, we introduce a pyridazine-pyridine-pyridazine diacid monomer and its incorporation into helical aromatic oligoamide foldamer containers. This monomer codes for a wide helix diameter and can sequester metal ions on the inner wall of the helix cavity. Crystallographic studies and NMR titrations show that part of the metal coordination sphere remains available and may then promote the binding of a guest within the cavity. In addition to metal coordination, binding of the guest is assisted by cooperative interactions with the helix host, thereby resulting in significant enhancements depending on the foldamer sequence, and in slow guest capture and release on the NMR time scale. In the absence of metal ions, the pyridazine-pyridine-pyridazine monomer promotes an extended conformation of the foldamer that results in aggregation, including the formation of an intertwined duplex.
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Hastoy, B.; Scotti, P. A.; Milochau, A.; Fezoua-Boubegtiten, Z.; Rodas, J.; Megret, R.; Desbat, B.; Laguerre, M.; Castano, S.; Perrais, D.; Rorsman, P.; Oda, R.; Lang, J.;
A Central Small Amino Acid in the VAMP2 Transmembrane Domain Regulates the Fusion Pore in Exocytosis
Sci Rep, 2017, 7, 2835
Exocytosis depends on cytosolic domains of SNARE proteins but the function of the transmembrane domains (TMDs) in membrane fusion remains controversial. The TMD of the SNARE protein synaptobrevin2/VAMP2 contains two highly conserved small amino acids, G100 and C103, in its central portion. Substituting G100 and/or C103 with the beta-branched amino acid valine impairs the structural flexibility of the TMD in terms of alpha-helix/beta-sheet transitions in model membranes (measured by infrared reflection-absorption or evanescent wave spectroscopy) during increase in protein/lipid ratios, a parameter expected to be altered by recruitment of SNAREs at fusion sites. This structural change is accompanied by reduced membrane fluidity (measured by infrared ellipsometry). The G100V/C103V mutation nearly abolishes depolarization-evoked exocytosis (measured by membrane capacitance) and hormone secretion (measured biochemically). Single-vesicle optical (by TIRF microscopy) and biophysical measurements of ATP release indicate that G100V/C103V retards initial fusion-pore opening, hinders its expansion and leads to premature closure in most instances. We conclude that the TMD of VAMP2 plays a critical role in membrane fusion and that the structural mobility provided by the central small amino acids is crucial for exocytosis by influencing the molecular re-arrangements of the lipid membrane that are necessary for fusion pore opening and expansion.
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Hano, C.; Corbin, C.; Drouet, S.; Quero, A.; Rombaut, N.; Savoire, R.; Molinie, R.; Thomasset, B.; Mesnard, F.; Laine, E.;
The lignan (+)-secoisolariciresinol extracted from flax hulls is an effective protectant of linseed oil and its emulsion against oxidative damage
European Journal of Lipid Science and Technology, 2017, 119,
Secoisolariciresinol (SECO) is a natural antioxidant lignan accumulated in large amounts in the seedcoat of flax and retained in the flaxseed cake residue during linseed oil pressing. Here SECO was extracted and purified from flaxseed cake and assayed for its ability to prevent oxidation of linseed oil and an o/w emulsion containing linseed oil. For this purpose, an accelerated storage (Schaal oven) test was performed and SECO effectiveness was compared to that of two antioxidants commonly used in food and cosmetic products: -tocopherol (-TOCO) and butylated hydroxyanisole (BHA). In our hands, SECO addition, ranging from 50 to 500mole per kg oil, significantly decreased the production of both primary (conjugated dienes, CD) and secondary (thiobarbituric acid-reactive substances, TBARS) oxidation products. This study evidenced that SECO is an effective stabilizer of linseed oil and its o/w emulsion and this protective effect outperformed both the natural -TOCO and the synthetic BHA antioxidants. In particular, SECO was the most effective in the protection of the o/w emulsion against secondary oxidation products, which makes it a potential alternative preservative for oily products in foods and cosmetics. Practical applications: The present study could lead to applications in the food and cosmetic industries for the stabilization of o/w emulsions. Addition of SECO to linseed oil, already rich in -3, could also result in a new functional food with synergistic beneficial actions on human health.
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Gyorgy, Bence; Sage, Cyrille; Indzhykulian, Artur A.; Scheffer, Deborah I.; Brisson, Alain R.; Tan, Sisareuth; Wu, Xudong; Volak, Adrienn; Mu, Dakai; Tamvakologos, Panos I.; Li, Yaqiao; Fitzpatrick, Zachary; Ericsson, Maria; Breakefield, Xandra O.; Corey, David P.; Maguire, Casey A.;
Rescue of Hearing by Gene Delivery to Inner-Ear Hair Cells Using Exosome-Associated AAV
Molecular therapy : the journal of the American Society of Gene Therapy, 2017, 25, 379-391
Adeno-associated virus (AAV) is a safe and effective vector for gene therapy for retinal disorders. Gene therapy for hearing disorders is not as advanced, in part because gene delivery to sensory hair cells of the inner ear is inefficient. Although AAV transduces the inner hair cells of the mouse cochlea, outer hair cells remain refractory to transduction. Here, we demonstrate that a vector, exosome-associated AAV (exo-AAV), is a potent carrier of transgenes to all inner ear hair cells. Exo-AAV1-GFP is more efficient than conventional AAV1-GFP, both in mouse cochlear explants invitro and with direct cochlear injection invivo. Exo-AAV shows no toxicity invivo, as assayed by tests of auditory and vestibular function. Finally, exo-AAV1 gene therapy partially rescues hearing in a mouse model of hereditary deafness (lipoma HMGIC fusion partner-like 5/tetraspan membrane protein of hair cell stereocilia [Lhfpl5/Tmhs-/-]). Exo-AAV is a powerful gene delivery system for hair cell research and may be useful for gene therapy for deafness.
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GUICHARD, Gilles; (FR).; COLLIE, Gavin; (GB).; PULKA-ZIACH, Karolina; (PL).; LOMBARDO, Caterina; (IT).; FREMAUX, Juliette; (FR) ;
, 2017, 0,
The present description provides compositions and methods for producing therapeutic oligomeric compounds. In another aspect the description provides methods for administering the oligomeric compounds for the treatment and prevention of disease in a mammal. In particular, the disclosure relates to medicaments comprising various novel oligomeric compounds and pharmaceutically acceptable salts thereof. The compounds of the disclosure may optionally be administered with at least one of a pharmaceutically acceptable excipient, additional pharmacologically active agent or a combination thereof.
39 […]
GUICHARD, Gilles; COLLIE, Gavin; PULKA-ZIACH, Karolina; LOMBARDO, Caterina; FREMAUX, Juliette;
, 2017, 0,
The present description provides compositions and methods for producing therapeutic oligomeric compounds. In another aspect the description provides methods for administering the oligomeric compounds for the treatment and prevention of disease in a mammal. In particular, the disclosure relates to medicaments comprising various novel oligomeric compounds and pharmaceutically acceptable salts thereof. The compounds of the disclosure may optionally be administered with at least one of a pharmaceutically acceptable excipient, additional pharmacologically active agent or a combination thereof.
40 […]
Gronnier, Julien; Crowet, Jean-Marc; Habenstein, Birgit; Nasir, Mehmet Nail; Bayle, Vincent; Hosy, Eric; Platre, Matthieu Pierre; Gouguet, Paul; Raffaele, Sylvain; Martinez, Denis; Grelard, Axelle; Loquet, Antoine; Simon-Plas, Francoise; Gerbeau-Pissot, Patricia; Der, Christophe; Bayer, Emmanuelle M.; Jaillais, Yvon; Deleu, Magali; Germain, Veronique; Lins, Laurence; Mongrand, Sebastien;
Structural basis for plant plasma membrane protein dynamics and organization into functional nanodomains
Elife, 2017, 6,
Plasma Membrane is the primary structure for adjusting to ever changing conditions. PM sub-compartmentalization in domains is thought to orchestrate signaling. Yet, mechanisms governing membrane organization are mostly uncharacterized. The plant-specific REMORINs are proteins regulating hormonal crosstalk and host invasion. REMs are the best-characterized nanodomain markers via an uncharacterized moiety called REMORIN C-terminal Anchor. By coupling biophysical methods, super-resolution microscopy and physiology, we decipher an original mechanism regulating the dynamic and organization of nanodomains. We showed that targeting of REMORIN is independent of the COP-II-dependent secretory pathway and mediated by PI4P and sterol. REM-CA is an unconventional lipid-binding motif that confers nanodomain organization. Analyses of REM-CA mutants by single particle tracking demonstrate that mobility and supramolecular organization are critical for immunity. This study provides a unique mechanistic insight into how the tight control of spatial segregation is critical in the definition of PM domain necessary to support biological function.
41 […]
Graf, Michael; Mardirossian, Mario; Nguyen, Fabian; Seefeldt, A. Carolin; Guichard, Gilles; Scocchi, Marco; Innis, C. Axel; Wilson, Daniel N.;
Proline-rich antimicrobial peptides targeting protein synthesis
Natural Product Reports, 2017, 34, 702-711
The innate immune system employs a broad array of antimicrobial peptides (AMPs) to attack invading microorganisms. While most AMPs act by permeabilizing the bacterial membrane, specific subclasses of AMPs have been identified that pass through membranes and inhibit bacterial growth by targeting fundamental intracellular processes. One such subclass is the proline-rich antimicrobial peptides (PrAMPs) that bind to the ribosome and interfere with the process of protein synthesis. A diverse range of PrAMPs have been identified in insects, such as bees, wasps and beetles, and crustaceans, such as crabs, as well as in mammals, such as cows, sheep, goats and pigs. Mechanistically, the best-characterized PrAMPs are the insect oncocins, such as Onc112, and bovine bactenecins, such as Bac7. Biochemical and structural studies have revealed that these PrAMPs bind within the ribosomal exit tunnel with a reverse orientation compared to a nascent polypeptide chain. The PrAMPs allow initiation but prevent the transition into the elongation phase of translation. Insight into the interactions of PrAMPs with their ribosomal target provides the opportunity to further develop these peptides as novel antimicrobial agents.
42 […]
Goto, Taisei; Okazaki, Yutaka; Ueki, Masahiro; Kuwahara, Yutaka; Takafuji, Makoto; Oda, Reiko; Ihara, Hirotaka;
Induction of Strong and Tunable Circularly Polarized Luminescence of Nonchiral, Nonmetal, Low-Molecular-Weight Fluorophores Using Chiral Nanotemplates
Angewandte Chemie-International Edition, 2017, 56, 2989-2993
A new strategy is described for generating strong circularly polarized luminescence with highly tunable emission bands through chiral induction in nonchiral, totally organic, low-molecular-weight fluorescent dyes by chiral nanotemplate systems. Our approach allows the first systematic investigation to clarify the correlation between the circular dichroism and circularly polarized luminescence intensities. As a result, a dilute solution system with the highest circularly polarized luminescence intensity achieved to date and a dissymmetry factor of over 0.1 was identified.
43 […]
Goibier, Lucie; Lecomte, Sophie; Leal-Calderon, Fernando; Faure, Chrystel;
The effect of surfactant crystallization on partial coalescence in O/W emulsions
Journal of colloid and interface science, 2017, 500, 304-314
Partial coalescence is a ubiquitous instability in emulsions whose dispersed phase is partially crystallized. When emulsions are stabilized with proteins, interfacial stiffness and long-range repulsive surface forces hinder this type of instability. The addition of low molecular weight surfactants modifies the interfacial properties and surface forces, generally promoting partial coalescence. In the present work, various surfactants (Tween 80, palmitic acid and monoglycerides) differing in their crystallization temperature were probed for their ability to induce partial coalescence in model O/W emulsions stabilized by sodium caseinate. The initially fluid emulsions were submitted to a tempering cycle leading to the gelation of the system. The extent of partial coalescence was evaluated by measuring the bulk storage modulus. DSC was used to determine the melting range of the oil phase and surfactants, while polarized microscopy, Raman imaging, and surface rheology measurements were performed to characterize the oil/water interface. The experimental conditions in terms of droplet size, surfactant-to-protein molar ratio and tempering history favoring partial coalescence were first explored in presence of Tween 80. We show that partial coalescence is rather marginal when crystallizable surfactants are added, and pronounced with liquid surfactants. The phenomena underlying this result, especially interfacial crystallization of surfactants, are evidenced and discussed.
44 […]
Gan, Quan; Wang, Xiang; Kauffmann, Brice; Rosu, Frederic; Ferrand, Yann; Huc, Ivan;
Translation of rod-like template sequences into homochiral assemblies of stacked helical oligomers
Nature nanotechnology, 2017, 0,
At the molecular level, translation refers to the production of a new entity according to a template that has a different chemical composition. In this way, chemical information may be translated from one molecule to another. The process is useful to synthesize structures and thus functions that might be difficult to create otherwise, and it reaches exquisite levels of efficiency in biological systems, as illustrated by protein expression from mRNA templates or by the assembly of the tobacco mosaic virus capsid protein according to the length of its RNA. In synthetic systems, examples of template-directed syntheses are numerous, but general and versatile schemes in which a non-natural sequence actually encodes the information necessary to produce a different sequence are few and far from being optimized. Here we show a high-fidelity enzyme-free translation of long rod-like alkylcarbamate oligomers into well-defined sequences of stacked helical aromatic oligoamides. The features present in the rods, which include the number and distance between carbamate functions and stereogenic centres, template the self-assembly of complementary stacks of helices that each have a defined right (P) or left (M) handedness, length and single or double helicity. This process enables the production of very large (>20 kDa) abiotic artificial folded architectures (foldamers) that may, for example, serve as scaffolds to organize appended functional features at positions in space defined with atomic precision across nanometric distances.
45 […]
Dufour, Florent; Rattier, Thibault; Shirley, Sarah; Picarda, Gaelle; Constantinescu, Andrei Alexandru; Morle, Aymeric; Zakaria, Al Batoul; Marcion, Guillaume; Causse, Sebastien; Szegezdi, Eva; Zajonc, Dirk Michael; Seigneuric, Renaud; Guichard, Gilles; Gharbi, Tijani; Picaud, Fabien; Herlem, Guillaume; Garrido, Carmen; Schneider, Pascal; Benedict, Chris Alan; Micheau, Olivier;
N-glycosylation of mouse TRAIL-R and human TRAIL-R1 enhances TRAIL-induced death
Cell Death and Differentiation, 2017, 24, 500-510
APO2L/TRAIL (TNF-related apoptosis-inducing ligand) induces death of tumor cells through two agonist receptors, TRAIL-R1 and TRAIL-R2. We demonstrate here that N-linked glycosylation (N-glyc) plays also an important regulatory role for TRAIL-R1-mediated and mouse TRAIL receptor (mTRAIL-R)-mediated apoptosis, but not for TRAIL-R2, which is devoid of N-glycans. Cells expressing N-glyc-defective mutants of TRAIL-R1 and mouse TRAIL-R were less sensitive to TRAIL than their wild-type counterparts. Defective apoptotic signaling by N-glyc-deficient TRAIL receptors was associated with lower TRAIL receptor aggregation and reduced DISC formation, but not with reduced TRAIL-binding affinity. Our results also indicate that TRAIL receptor N-glyc impacts immune evasion strategies. The cytomegalovirus (CMV) UL141 protein, which restricts cell-surface expression of human TRAIL death receptors, binds with significant higher affinity TRAIL-R1 lacking N-glyc, suggesting that this sugar modification may have evolved as a counterstrategy to prevent receptor inhibition by UL141. Altogether our findings demonstrate that N-glyc of TRAIL-R1 promotes TRAIL signaling and restricts virus-mediated inhibition.
46 […]
Dufour, Florent; Rattier, Thibault; Constantinescu, Andrei Alexandru; Zischler, Luciana; Morle, Aymeric; Ben Mabrouk, Hazem; Humblin, Etienne; Jacquemin, Guillaume; Szegezdi, Eva; Delacote, Fabien; Marrakchi, Naziha; Guichard, Gilles; Pellat-Deceunynck, Catherine; Vacher, Pierre; Legembre, Patrick; Garrido, Carmen; Micheau, Olivier;
TRAIL receptor gene editing unveils TRAIL-R1 as a master player of apoptosis induced by TRAIL and ER stress
Oncotarget, 2017, 8, 9974-9985
TRAIL induces selective tumor cell death through TRAIL-R1 and TRAIL-R2. Despite the fact that these receptors share high structural homologies, induction of apoptosis upon ER stress, cell autonomous motility and invasion have solely been described to occur through TRAIL-R2. Using the TALEN gene-editing approach, we show that TRAIL-R1 can also induce apoptosis during unresolved unfolded protein response (UPR). Likewise, TRAIL-R1 was found to co-immunoprecipitate with FADD and caspase-8 during ER stress. Its deficiency conferred resistance to apoptosis induced by thaspigargin, tunicamycin or brefeldin A. Our data also demonstrate that tumor cell motility and invasion-induced by TRAIL-R2 is not cell autonomous but induced in a TRAIL-dependant manner. TRAIL-R1, on the other hand, is unable to trigger cell migration owing to its inability to induce an increase in calcium flux. Importantly, all the isogenic cell lines generated in this study revealed that apoptosis induced TRAIL is preferentially induced by TRAIL-R1. Taken together, our results provide novel insights into the physiological functions of TRAIL-R1 and TRAIL-R2 and suggest that targeting TRAIL-R1 for anticancer therapy is likely to be more appropriate owing to its lack of pro-motile signaling capability.
47 […]
Dridi, Wafa; Toutain, Jean; Sommier, Alain; Essafi, Wafa; Leal-Calderon, Fernando; Cansell, Maud;
Direct technique for monitoring lipid oxidation in water-in-oil emulsions based on micro-calorimetry
Food chemistry, 2017, 230, 563-566
An experimental device based on the measurement of the heat flux dissipated during chemical reactions, previously validated for monitoring lipid oxidation in plant oils, was extended to follow lipid oxidation in water-in-oil emulsions. Firstly, validation of the approach was performed by correlating conjugated diene concentrations measured by spectrophotometry and the heat flux dissipated by oxidation reactions and measured directly in water-in-oil emulsions, in isothermal conditions at 60°C. Secondly, several emulsions based on plant oils differing in their n-3 fatty acid content were compared. The oxidability parameter derived from the enthalpy curves reflected the alpha-linolenic acid proportion in the oils. On the whole, the micro-calorimetry technique provides a sensitive method to assess lipid oxidation in water-in-oil emulsions without requiring any phase extraction.
48 […]
Daury, Laetitia; Taveau, Jean-Christophe; Salvador, Dimitri; Glavier, Marie; Lambert, Olivier;
Reconstitution of Membrane Proteins into Nanodiscs for Single-Particle Electron Microscopy
Methods in molecular biology (Clifton, N.J.), 2017, 1635, 317-327
The structure determination of integral membrane protein (IMP) in lipid environment is particularly challenging. Among emerging methods for exchanging detergent required for IMP purification by original compounds, the use of lipid nanodisc preserves a lipid environment. Compared with the classical method of proteoliposome formation, the nanodisc technology provides a better control of IMP molecules inserted in lipid membrane, therefore giving access to structural methodologies developed for soluble proteins. Here, we present the reconstitution of OprM membrane protein into nanodisc associated with a step of size-exclusion chromatography, an approach applicable to prepare IMPs for subsequent visualization by single-particle electron microscopy.
49 […]
Crouzet, M.; Claverol, S.; Lomenech, A. M.; Le Senechal, C.; Costaglioli, P.; Barthe, C.; Garbay, B.; Bonneu, M.; Vilain, S.;
Pseudomonas aeruginosa cells attached to a surface display a typical proteome early as 20 minutes of incubation
Plos One, 2017, 12,
Biofilms are present in all environments and often result in negative effects due to properties of the biofilm lifestyle and especially antibiotics resistance. Biofilms are associated with chronic infections. Controlling bacterial attachment, the first step of biofilm formation, is crucial for fighting against biofilm and subsequently preventing the persistence of infection. Thus deciphering the underlying molecular mechanisms involved in attachment could allow discovering molecular targets from it would be possible to develop inhibitors against bacterial colonization and potentiate antibiotherapy. To identify the key components and pathways that aid the opportunistic pathogen Pseudomonas aeruginosa in attachment we performed for the first time a proteomic analysis as early as after 20 minutes of incubation using glass wool fibers as a surface. We compared the protein contents of the attached and unattached bacteria. Using mass spectrometry, 3043 proteins were identified. Our results showed that, as of 20 minutes of incubation, using stringent quantification criteria 616 proteins presented a modification of their abundance in the attached cells compared to their unattached counterparts. The attached cells presented an overall reduced gene expression and characteristics of slow-growing cells. The over-accumulation of outer membrane proteins, periplasmic folding proteins and O-antigen chain length regulators was also observed, indicating a profound modification of the cell envelope. Consistently the sigma factor AlgU required for cell envelope homeostasis was highly over-accumulated in attached cells. In addition our data suggested a role of alarmone (p) ppGpp and polyphosphate during the early attachment phase. Furthermore, almost 150 proteins of unknown function were differentially accumulated in the attached cells. Our proteomic analysis revealed the existence of distinctive biological features in attached cells as early as 20 minutes of incubation. Analysis of some mutants demonstrated the interest of this proteomic approach in identifying genes involved in the early phase of adhesion to a surface.
50 […]
Coumans, F. A. W.; Brisson, A. R.; Buzas, E. I.; Dignat-George, F.; Drees, E. E. E.; El-Andaloussi, S.; Emanueli, C.; Gasecka, A.; Hendrix, A.; Hill, A. F.; Lacroix, R.; Lee, Y.; van Leeuwen, T. G.; Mackman, N.; Mager, I.; Nolan, J. P.; van der Pol, E.; Pegtel, D. M.; Sahoo, S.; Siljander, P. R. M.; Sturk, G.; de Wever, O.; Nieuwland, R.;
Methodological Guidelines to Study Extracellular Vesicles
Circulation Research, 2017, 120, 1632-1648
Owing to the relationship between extracellular vesicles (EVs) and physiological and pathological conditions, the interest in EVs is exponentially growing. EVs hold high hopes for novel diagnostic and translational discoveries. This review provides an expert-based update of recent advances in the methods to study EVs and summarizes currently accepted considerations and recommendations from sample collection to isolation, detection, and characterization of EVs. Common misconceptions and methodological pitfalls are highlighted. Although EVs are found in all body fluids, in this review, we will focus on EVs from human blood, not only our most complex but also the most interesting body fluid for cardiovascular research.
51 […]
Colombani, Thibault; Peuziat, Pauline; Dallet, Laurence; Haudebourg, Thomas; Mevel, Mathieu; Berchel, Mathieu; Lambert, Olivier; Habrant, Damien; Pitard, Bruno;
Self-assembling complexes between binary mixtures of lipids with different linkers and nucleic acids promote universal mRNA, DNA and siRNA delivery
Journal of controlled release : official journal of the Controlled Release Society, 2017, 249, 131-142
Protein expression and RNA interference require efficient delivery of DNA or mRNA and small double stranded RNA into cells, respectively. Although cationic lipids are the most commonly used synthetic delivery vectors, a clear need still exists for a better delivery of various types of nucleic acids molecules to improve their biological activity. To optimize the transfection efficiency, a molecular approach consisting in modifying the chemical structure of a given cationic lipid is usually performed, but an alternative strategy could rely on modulating the supramolecular assembly of lipidic lamellar phases sandwiching the nucleic acids molecules. To validate this new concept, we synthesized on one hand two paromomycin-based cationic lipids, with either an amide or a phosphoramide linker, and on the other hand two imidazole-based neutral lipids, having as well either an amide or a phosphoramide function as linker. Combinations of cationic and helper lipids containing the same amide or phosphoramide linkers led to the formation of homogeneous lamellar phases, while hybrid lamellar phases were obtained when the linkers on the cationic and helper lipids were different. Cryo-transmission electron microscopy and fluorescence experiments showed that liposomes/nucleic acids complexes resulting from the association of nucleic acids with hybrid lamellar phases led to complexes that were more stable in the extracellular compartment compared to those obtained with homogeneous systems. In addition, we observed that the most active supramolecular assemblies for the delivery of DNA, mRNA and siRNA were obtained when the cationic and helper lipids possess linkers of different natures. The results clearly show that this supramolecular strategy modulating the property of the lipidic lamellar phase constitutes a new approach for increasing the delivery of various types of nucleic acid molecules.
52 […]
Collie, Gavin W.; Bailly, Remy; Pulka-Ziach, Karolina; Lombardo, Caterina M.; Mauran, Laura; Taib-Maamar, Nada; Dessolin, Jean; Mackereth, Cameron D.; Guichard, Gilles;
Molecular Recognition within the Cavity of a Foldamer Helix Bundle: Encapsulation of Primary Alcohols in Aqueous Conditions
Journal of the American Chemical Society, 2017, 0,
Artificial synthetic molecules able to adopt well-defined stable secondary structures comparable to those found in nature ("foldamers") have considerable potential for use in a range of applications such as biomaterials, biorecognition, nanomachines and as therapeutic agents. The development of foldamers with the ability to bind and encapsulate "guest" molecules is of particular interest; as such an ability is a key step toward the development of artificial sensors, receptors and drug-delivery vectors. Although significant progress has been reported within this context, foldamer capsules reported thus far are largely restricted to organic solvent systems, and it is likely that the move to aqueous conditions will prove challenging. Toward this end, we report here structural studies into the ability of a recently reported water-soluble self-assembled foldamer helix bundle to encapsulate simple guest molecules within an internal cavity. Seven high-resolution aqueous crystal structures are reported, accompanied by molecular dynamics and high-field NMR solution data, showing for the first time that encapsulation of guests by a complex self-assembled foldamer in aqueous conditions is possible. The findings also provide ample insight for the future functional development of this system.
53 […]
Chomette, C.; Treguer-Delapierre, M.; Schade, N. B.; Manoharan, V. N.; Lambert, O.; Taveau, J. C.; Ravaine, S.; Duguet, E.;
Colloidal Alchemy: Conversion of Polystyrene Nanoclusters into Gold
Chemnanomat, 2017, 3, 160-163
Isotropic plasmonic clusters consisting of a controlled number of gold satellites around a silica core are fabricated from silica/polystyrene tetrapod, hexapod, and dodecapod templates. The synthetic pathway includes stages of site-specific seed adsorption, seed-mediated growth, and iterative etching/regrowth to reshape the satellites into spheroids. Transmission electron microscopy and electron tomography provide evidence of the symmetry of the clusters. This work paves the way for a comprehensive study of their optical properties.
54 […]
Cheng, Jiaji; Le Saux, Guillaume; Gao, Jie; Buffeteau, Thierry; Battie, Yann; Barois, Philippe; Ponsinet, Virginie; Delville, Marie-Helene; Ersen, Ovidiu; Pouget, Emilie; Oda, Reiko;
GoldHelix: Gold Nanoparticles Forming 3D Helical Superstructures with Controlled Morphology and Strong Chiroptical Property
ACS nano, 2017, 0,
Plasmonic nanoparticles, particularly gold nanoparticles (GNPs) hold a great potential as structural and functional building blocks for three-dimensional (3D) nanoarchitectures with specific optical applications. However, a rational control of their assembly into nanoscale superstructures with defined positioning and overall arrangement still remains challenging. Herein, we propose a solution to this challenge by using as building blocks: (1) nanometric silica helices with tunable handedness and sizes as a matrix and (2) GNPs with diameter varying from 4 to 10 nm to prepare a collection of helical GNPs superstructures (called Goldhelices hereafter). These nanomaterials exhibit well-defined arrangement of GNPs following the helicity of the silica template. Strong chiroptical activity is evidenced by circular dichroism (CD) spectroscopy at the wavelength of the surface plasmon resonance (SPR) of the GNPs with a anisotropy factor (g-factor) of the order of 1 * 10-4, i.e., 10-fold larger than what is typically reported in the literature. Such CD signals were simulated using a coupled dipole method which fit very well the experimental data. The measured signals are 1-2 orders of magnitude lower than the simulated signals, which is explained by the disordered GNPs grafting, the polydispersity of the GNPs, and the dimension of the nanohelices. These Goldhelices based on inorganic templates are much more robust than previously reported organic-based chiroptical nanostructures, making them good candidates for complex hierarchical organization, providing a promising approach for light management and benefits in applications such as circular polarizers, chiral metamaterials, or chiral sensing in the visible range.
55 […]
Chardon, Edith; Dahm, Georges; Guichard, Gilles; Bellemin-Laponnaz, Stephane;
Synthesis and structural characterization of alkyne-functionalized N-heterocyclic carbene complexes of ruthenium, palladium and rhodium
Inorganica Chimica Acta, 2017, 467, 33-38
A series of novel ruthenium(II), palladium(II) and rhodium(I) complexes bearing N-heterocyclic carbene ligand functionalized by an alkyne-protected functional group (-C equivalent to C-SiMe3) were synthesized by transmetalation from the silver-NHC intermediate or by deprotonation of the azolium salt using an external base. The complexes were fully characterized and X-ray diffraction was used for the structure determination. The donating properties of the NHC ligand were evaluated through the synthesis of the rhodium carbonyl complex. The reactivity of the functional group has been studied: deprotection of the alkyne on the ruthenium and rhodium species gave instable products whereas deprotection of the palladium compound was quantitative. However the resulting alkyne-functionalized species was incompatible with conditions for further post-functionalization such as metal-catalyzed 1,3-dipolar cycloaddition. (C) 2017 Elsevier B.V. All rights reserved.
56 […]
Cartier, Flora; Indersie, Emilie; Lesjean, Sarah; Charpentier, Justine; Hooks, Katarzyna B.; Ghousein, Amani; Desplat, Angelique; Dugot-Senant, Nathalie; Trezeguet, Veronique; Sagliocco, Francis; Hagedorn, Martin; Grosset, Christophe F.;
New tumor suppressor microRNAs target glypican-3 in human liver cancer
Oncotarget, 2017, 0,
Glypican-3 (GPC3) is an oncogene, frequently upregulated in liver malignancies such as hepatocellular carcinoma (HCC) and hepatoblastoma and constitutes a potential molecular target for therapy in liver cancer. Using a functional screening system, we identified 10 new microRNAs controlling GPC3 expression in malignant liver cells, five of them e.g. miR-4510, miR-203a-3p, miR-548aa, miR-376b-3p and miR-548v reduce GPC3 expression. These 5 microRNAs were significantly downregulated in tumoral compared to non-tumoral liver and inhibited tumor cell proliferation. Interestingly, miR-4510 inversely correlated with GPC3 mRNA and protein in HCC samples. This microRNA also induced apoptosis of hepatoma cells and blocked tumor growth in vivo in the chick chorioallantoic membrane model. We further show that the tumor suppressive effect of miR-4510 is mediated through direct targeting of GPC3 mRNA and inactivation of Wnt/beta-catenin transcriptional activity and signaling pathway. Moreover, miR-4510 up-regulated the expression of several tumor suppressor genes while reducing the expression of other pro-oncogenes. In summary, we uncovered several new microRNAs targeting the oncogenic functions of GPC3. We provided strong molecular, cellular and in vivo evidences for the tumor suppressive activities of miR-4510 bringing to the fore the potential value of this microRNA in HCC therapy.
57 […]
Carmeille, Romain; Croissant, Coralie; Bouvet, Flora; Bouter, Anthony;
Membrane Repair Assay for Human Skeletal Muscle Cells
Methods in molecular biology (Clifton, N.J.), 2017, 1668, 195-207
The characterization of the membrane repair machinery in human skeletal muscle has become crucial, since it has been shown that some muscular dystrophies result from a defect of this fundamental physiological process. Deciphering membrane repair mechanism requires the development of methodologies allowing studying the response of skeletal muscle cells to sarcolemma damage and identifying candidate proteins playing a role in the membrane repair machinery. Here, we describe a protocol that is based on the creation of cell membrane disruption by infrared laser irradiation in human myotubes. Membrane disruption and repair are assayed by monitoring the incorporation into myotubes of the membrane probe FM1-43. This methodology has recently enabled us to show that Annexin-A5 is required for membrane repair in human skeletal muscle cells (Carmeille et al., Biochim Biophys Acta 1863:2267-2279, 2016).
58 […]
Carasi, Paula; Racedo, Silvia Maria; Jacquot, Claudine; Elie, Anne Marie; de los Angeles Serradell, Maria; Urdaci, Maria C.;
Enterococcus durans EP1 a Promising Anti-inflammatory Probiotic Able to Stimulate slgA and to Increase Faecalibacterium prausnitzii Abundance
Frontiers in Immunology, 2017, 8,
Enterococcus species, principally Enterococcus faecium are used as probiotics since a long time with preference in animal applications but safety considerations were updated and also new uses as probiotics can be envisaged. Fifteen Enterococcus strains isolated from different foods were identified and analyzed for virulence factors and antibiotic resistance. Three Enterococcus durans strains were selected to study their immunomodulatory properties on PBMC and Caco2 cells. Two strains presented a profile toward a mild inflammatory Th1 response considering TNF-alpha/IL-10 and IL-1 beta/IL-10 cytokines ratios. The third strain EP1, presented an anti-inflammatory potential and was selected for in vivo studies. In mice, the strain was well tolerated and did not cause any adverse effects. EP1 administration increased the amount of IgA+ cells in mesenteric lymph node (MLN) after 7 days of administration. In fecal samples, the IgA content increased gradually and significantly from day 7 to day 21 in treated group. Additionally, IL-17, IL-6, IL-1 beta, IFN-gamma, and CXCL1 gene expression significantly decreased on day 21 in Peyer's patches and IL-17 decreased in MLN. Mice treated with the probiotic showed significant lower mRNA levels of pro-inflammatory cytokines and mucins in the ileum at day 7 while their expression was normalized at day 21. Colonic expression of il-1 beta, il6, and mucins remain diminished at day 21. Ileum and colon explants from treated mice stimulated in vitro with LPS showed a significant reduction in IL-6 and an increase in IL-10 secretion suggesting an in vivo protective effect of the probiotic treatment against a proinflammatory stimulus. Interestingly, analysis of feces microbiota demonstrated that EP1 administration increase the amount of Faecalibacterium prausnitzii, a butyrate-producing bacteria, which is known for its anti-inflammatory effects. In conclusion, we demonstrated that EP1 strain is a strong sIgA inducer and possess mucosal anti-inflammatory properties. This strain also modulates gut microbiota increasing Faecalibacterium prausnitzii, a functionally important bacterium. Thus, E. durans EP1 is not only a good candidate to increases F. prausnitzii in some cases of dysbiosis but can also be interesting in gut inflammatory disorders therapy.
59 […]
Cansell, Maud; Bardeau, Tiphaine; Morvan, Estelle; Grelard, Axelle; Bure, Corinne; Subra-Paternault, Pascale;
Phospholipid Profiles of Oleaginous Pressed Cakes Using NMR and Gas Chromatography
Journal of the American Oil Chemists Society, 2017, 94, 1219-1223
Camelina, flaxseed, hemp, sesame, and walnut cakes were analyzed for their phospholipid (PL) content and composition using P-31 and H-1 nuclear magnetic resonance and gas chromatography. The data evidenced variations between the sources in terms of (1) total lipid content and PL concentration, camelina cake being the richest source of PLs, (2) PL composition, phosphatidylcholine being the most abundant phospholipid in sesame and hemp cakes, whereas phosphatidylinositol represented about 25% of the total PLs in most cakes, and (3) fatty acid composition of the PLs, camelina cake being the richest source of omega 3 polyunsaturated fatty acids. These data may be useful to diversify the PL sources available and to provide PL fractions with specific nutritional or functional properties.
60 […]
Brisson, Alain R.; Tan, Sisareuth; Linares, Romain; Gounou, Celine; Arraud, Nicolas;
Extracellular vesicles from activated platelets: a semiquantitative cryo-electron microscopy and immuno-gold labeling study
Platelets, 2017, 0, 1-9
Cells release membrane vesicles in their surrounding medium either constitutively or in response to activating signals. Two main types of extracellular vesicles (EVs) are commonly distinguished based on their mechanism of formation, membrane composition and size. According to the current model, EVs shed from the plasma membrane, often called microvesicles, expose phosphatidylserine (PS) and range in size from 100 nm to 1 m, while EVs originating from endosomal multi-vesicular bodies, called exosomes, contain tetraspanin proteins, including CD63, and range in size from 50 to 100 nm. Heijnen et al. [1] have shown that activated platelets release EVs corresponding to these two types of vesicles, using negative staining electron microscopy (EM) and immuno-gold labeling. Here, we apply cryo-EM and immuno-gold labeling to provide a quantitative analysis of EVs released by platelets activated by thrombin, TRAP and CRP-XL, as well as EVs from serum. We show that EVs activated by these three agonists present a similar size distribution, the majority of them forming a broad peak extending from 50 nm to 1 m, about 50% of them ranging from 50 to 400 nm. We show also that 60% of the EVs from TRAP or CRP-XL activation expose CD41, a majority of them exposing also PS. To explain the presence of large EVs CD41-negative or PS-negative, several alternative mechanisms of EV formation are proposed. We find also that the majority of EVs in activated platelet samples expose CD63, and distinguish two populations of CD63-positive EVs, namely large EVs with low labeling density and small EVs with high labeling density.
61 […]
Briffa, M.; Ghio, S.; Neuner, J.; Gauci, A. J.; Cacciottolo, R.; Marchal, C.; Caruana, M.; Cullin, C.; Vassallo, N.; Cauchi, R. J.;
Extracts from two ubiquitous Mediterranean plants ameliorate cellular and animal models of neurodegenerative proteinopathies
Neuroscience Letters, 2017, 638, 12-20
A signature feature of age-related neurodegenerative proteinopathies is the misfolding and aggregation of proteins, typically amyloid-beta (A beta) in Alzheimer's disease (AD) and alpha-synuclein (alpha-syn) in Parkinson's disease (PD), into soluble oligomeric structures that are highly neurotoxic. Cellular and animal models that faithfully replicate the hallmark features of these disorders are being increasing exploited to identify disease-modifying compounds. Natural compounds have been identified as a useful source of bioactive molecules with promising neuroprotective capabilities. In the present report, we investigated whether extracts derived from two ubiquitous Mediterranean plants namely, the prickly pear Opuntia ficus-indica (EOFI) and the brown alga Padina pavonica (EPP) alleviate neurodegenerative phenotypes in yeast (Saccharomyces cerevisiae) and fly (Drosophila melanogaster) models of AD and PD. Pre-treatment with EPP or EOFI in the culture medium significantly improved the viability of yeast expressing the Arctic A beta 42 (E22G) mutant. Supplementing food with EOFI or EPP dramatically ameliorated lifespan and behavioural signs of flies with brain-specific expression of wild-type A beta 42 (model of late-onset AD) or the Arctic A beta 42 variant (model of early-onset AD). Additionally, we show that either extract prolonged the survival of a PD fly model based on transgenic expression of the human alpha-syn A53T mutant. Taken together, our findings suggest that the plant-derived extracts interfere with shared mechanisms of neurodegeneration in AD and PD. This notion is strengthened by evidence demonstrating that EOFI and to a greater extent EPP, while strongly inhibiting the fibrillogenesis of both A beta 42 and alpha-syn, accumulate remodelled oligomeric aggregates that are less effective at disrupting lipid membrane integrity. Our work therefore opens new avenues for developing therapeutic applications of these natural plant extracts in the treatment of amyloidogenic neurodegenerative disorders. (C) 2016 Elsevier Ireland Ltd. All rights reserved.
62 […]
Boscardin, E.; Perrier, R.; Sergi, C.; Maillard, M.; Loffing, J.; Loffing-Cueni, D.; Koesters, R.; Rossier, B. C.; Hummler, E.;
Severe hyperkalemia is rescued by low-potassium diet in renal beta ENaC-deficient mice
Pflugers Archiv-European Journal of Physiology, 2017, 469, 1387-1399
In adulthood, an induced nephron-specific deficiency of alpha ENaC (Scnn1a) resulted in pseudohypoaldosteronism type 1 (PHA-1) with sodium loss, hyperkalemia, and metabolic acidosis that is rescued through high-sodium/low-potassium (HNa+/LK+) diet. In the present study, we addressed whether renal beta ENaC expression is required for sodium and potassium balance or can be compensated by remaining (alpha and gamma) ENaC subunits using adult nephron-specific knockout (Scnn1b(Pax8/LC1)) mice. Upon induction, these mice present a severe PHA-1 phenotype with weight loss, hyperkalemia, and dehydration, but unlike the Scnn1a(Pax8/LC1) mice without persistent salt wasting. This is followed by a marked downregulation of STE20/SPS1-related proline-alanine-rich protein kinase (SPAK) and Na+/Cl- co-transporter (NCC) protein expression and activity. Most of the experimental Scnn1b(Pax8/LC1) mice survived with a HNa+/LK+ diet that partly normalized NCC phosphorylation, but not total NCC expression. Since salt loss was minor, we applied a standard-sodium/LK+ diet that efficiently rescued these mice resulting in normokalemia and normalization of NCC phosphorylation, but not total NCC expression. A further switch to LNa+/standard-K+ diet induced again a severe PHA-1-like phenotype, but with only transient salt wasting indicating that low-K+ intake is critical to decrease hyperkalemia in a NCC-dependent manner. In conclusion, while the beta ENaC subunit plays only a minor role in sodium balance, severe hyperkalemia results in downregulation of NCC expression and activity. Our data demonstrate the importance to primarily correct the hyperkalemia with a low-potassium diet that normalizes NCC activity.
63 […]
Bonhommeau, S.; Talaga, D.; Hunel, J.; Cullin, C.; Lecomte, S.;
Tip-Enhanced Raman Spectroscopy to Distinguish Toxic Oligomers from Abeta1-42 Fibrils at the Nanometer Scale
Angew Chem Int Ed Engl, 2017, 56, 1771-1774
For the first time, natural Abeta1-42 fibrils (WT) implicated in Alzheimer's disease, as well as two synthetic mutants forming less toxic amyloid fibrils (L34T) and highly toxic oligomers (oG37C), are chemically characterized at the scale of a single structure using tip-enhanced Raman spectroscopy (TERS). While the proportion of TERS features associated with amino acid residues is similar for the three peptides, a careful examination of amide I and amide III bands allows us to clearly distinguish WT and L34T fibers organized in parallel beta-sheets from the small and more toxic oG37C oligomers organized in anti-parallel beta-sheets.
64 […]
Bobo, C.; Chaignepain, S.; Henry, S.; Vignaud, H.; Ameadan, A.; Marchal, C.; Prado, E.; Doutch, J.; Schmitter, J. M.; Nardin, C.; Lecomte, S.; Cullin, C.;
Synthetic toxic Abeta1-42 oligomers can assemble in different morphologies
Biochim Biophys Acta, 2017, 1861, 1168-1176
BACKGROUND: Alzheimer's disease is the most common neurodegenerative disease associated with aggregation of Abeta peptides. Abeta toxicity is mostly related to the capacity of intermediate oligomers to disrupt membrane integrity. We previously expressed Abeta1-42 in a eukaryotic cellular system and selected synthetic variants on their sole toxicity. The most toxic mutant G37C forms stable oligomers. METHODS: Different biophysical methods (Fluorescence spectroscopy, cross-linking, mass spectrometry (MS), Small Angle X-ray Scattering (SAXS), Atomic Force Microscopy (AFM), Transmission Electron Microscopy (TEM), calcein leakage) were used. RESULTS: The oligomers are mostly populated by a 14mers resulting from the packing of homodimers. These homodimers come from the formation of a disulfide bridge between two monomers. This link stabilizes the multimers and prevents the assembly into amyloid fibrils. These oligomers affect the membrane integrity. The reduction of disulfide bonds leads to a rearrangement and redirects assembly of Abeta amyloid fibrils. CONCLUSION: The toxic synthetic AbetaG37C mutant can assemble into an amyloid of unusual morphology through the formation of anti-parallel beta-sheets. This pathway involves the formation of oligomers resulting from the arrangement of Abeta dimers linked by covalent di-sulfide link, being these oligomers harmful for the membranes. GENERAL SIGNIFICANCE: The capacity to produce large amount of stable oligomers without additional detergents or extrinsic cross-linkers allow further structural and biophysical studies to understand their capacity to assemble and disrupt the membranes, a key event in Alzheimer's disease.
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Beztsinna, Nataliia; Tsvetkova, Yoanna; Jose, Jithin; Rhourri-Frih, Boutayna; Al Rawashdeh, Wa'el; Lammers, Twan; Kiessling, Fabian; Bestel, Isabelle;
Photoacoustic imaging of tumor targeting with riboflavin-functionalized theranostic nanocarriers
International Journal of Nanomedicine, 2017, 12, 3813-3825
Photoacoustic imaging is an emerging method in the molecular imaging field, providing high spatiotemporal resolution and sufficient imaging depths for many clinical applications. Therefore, the aim of this study was to use photoacoustic imaging as a tool to evaluate a riboflavin (RF)-based targeted nanoplatform. RF is internalized by the cells through a specific pathway, and its derivatives were recently shown as promising tumor-targeting vectors for the drug delivery systems. Here, the RF amphiphile synthesized from a PEGylated phospholipid was successfully inserted into a long-circulating liposome formulation labeled with the clinically approved photoacoustic contrast agent - indocyanine green (ICG). The obtained liposomes had a diameter of 124 nm (polydispersity index = 0.17) and had a negative zeta potential of -26 mV. Studies in biological phantoms indicated a stable and concentration-dependent photoacoustic signal (Vevo (R) LAZR) of the ICG-containing RF-functionalized liposomes. In A431 cells, a high uptake of RF-functionalized liposomes was found and could be blocked competitively. First, studies in mice revealed similar to 3 times higher photoacoustic signal in subcutaneous A431 tumor xenografts (P<0.05) after injection of RF-functionalized liposomes compared to control particles. In this context, the application of a spectral unmixing protocol confirmed the initial quantitative data and improved the localization of liposomes in the tumor. In conclusion, the synthesized RF amphiphile leads to efficient liposomal tumor targeting and can be favorably detected by photoacoustic imaging with a perspective of theranostic applications.
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Berger, Emmanuelle; Nassra, Merian; Atgie, Claude; Plaisancie, Pascale; Geloen, Alain;
Oleic Acid Uptake Reveals the Rescued Enterocyte Phenotype of Colon Cancer Caco-2 by HT29-MTX Cells in Co-Culture Mode
International journal of molecular sciences, 2017, 18,
Gastrointestinal epithelium is the unique route for nutrients and for many pharmaceuticals to enter the body. The present study aimed to analyze precisely whether co-culture of two colon cancer cell lines, mucus-producing cells HT29-MTX and enterocyte-like Caco-2 cells, ameliorate differentiation into an in vitro intestinal barrier model and the signaling pathways involved. Differentiated Caco-2 cells gene datasets were compared first to intestinal or cancer phenotypes and second to signaling pathway gene datasets. Experimental validations were performed in real-time experiments, immunochemistry, and gene expression analyses on Caco-2 versus co-cultures of Caco-2 and HT29-MTX (10%) cells. Partial maintenance of cancer-cell phenotype in differentiated Caco-2 cells was confirmed and fatty acids merged as potential regulators of cancer signaling pathways. HT29-MTX cells induced morphological changes in Caco-2 cells, slightly increased their proliferation rate and profoundly modified gene transcription of phenotype markers, fatty acid receptors, intracellular transporters, and lipid droplet components as well as functional responses to oleic acid. In vitro, enterocyte phenotype was rescued partially by co-culture of cancer cells with goblet cells and completed through oleic acid interaction with signaling pathways dysregulated in cancer cells.
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Becart, Diane; Diemer, Vincent; Salaun, Arnaud; Oiarbide, Mikel; Nelli, Yella Reddy; Kauffmann, Brice; Fischer, Lucile; Palomo, Claudio; Guichard, Gilles;
Helical Oligourea Foldamers as Powerful Hydrogen Bonding Catalysts for Enantioselective C-C Bond-Forming Reactions
Journal of the American Chemical Society, 2017, 139, 12524-12532
Substantial progress has been made toward the development of metal-free catalysts of enantioselective transformations, yet the discovery of organic catalysts effective at low catalyst loadings remains a major challenge. Here we report a novel synergistic catalyst combination system consisting of a peptide-inspired chiral helical (thio)urea oligomer and a simple tertiary amine that is able to promote the Michael reaction between enolizable carbonyl compounds and nitroolefins with excellent enantioselectivities at exceptionally low (1/10 000) chiral catalyst/substrate molar ratios. In addition to high selectivity, which correlates strongly with helix folding, the system we report here is also highly amenable to optimization, as each of its components can be fine-tuned separately to increase reaction rates and/or selectivities. The predictability of the foldamer secondary structure coupled to the high level of control over the primary sequence results in a system with significant potential for future catalyst design.
68 […]
Bazin, D.; Faure, C.;
Superhydrophobic, highly adhesive arrays of copper hollow spheres produced by electro-colloidal lithography
Soft Matter, 2017, 13, 5500-5505
We report the patterning of copper surfaces which display both superhydrophobicity and high adhesion thanks to a new feature geometry, and without resorting to chemical modification. Polystyrene beads organized in 2D crystals under an AC electric field act as a template for the growth of copper deposited via cupric ion-loaded multi-lamellar vesicles. After the removal of the beads, hexagonal arrays of supported hollow spheres or copper bowls are generated, depending on the amount of deposited copper. While the bowl-covered surfaces display a predictable decreasing wettability (Cassie model) as their wall height increases, the hollow sphere-covered surfaces exhibit both high adhesion and superhydrophobicity (Cassie-Baxter state).
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Bayard, Mathilde; Leal-Calderon, Fernando; Cansell, Maud;
Free fatty acids and their esters modulate isothermal crystallization of anhydrous milk fat
Food Chemistry, 2017, 218, 22-29
The effect of free fatty acids with different chain lengths or unsaturation degree on anhydrous milk fat (AMF) crystallization was evaluated. The impact of esterification was also studied using three triglycerides. Melted blends containing the additives at concentrations lower than 12 wt.% were quenched at 25 degrees C and isothermal crystallization was monitored by pulsed low-resolution nuclear magnetic resonance. In parallel, polarized light microscopy was used to observe the microstructure. Compounds based on long chain saturated fatty acids, i.e. palmitic, stearic, eicosanoic acids, tripalmitin and tristearin accelerated crystallization. Conversely, propanoic, hexanoic and oleic acids slowed down the process, while triacetin had no impact. Interestingly, above a critical concentration, the addition of palmitic, stearic or eicosanoic acids caused a transition from a one-step to two-step process. Gompertz model was used to fit the experimental data and to assess the influence of the molecular properties of the additives on the kinetic parameters. (C) 2016 Elsevier Ltd. All rights reserved.
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Antunes, Stephanie; Corre, Jean-Philippe; Mikaty, Guillain; Douat, Celine; Goossens, Pierre L.; Guichard, Gilles;
Effect of replacing main-chain ureas with thiourea and guanidinium surrogates on the bactericidal activity of membrane active oligourea foldamers
Bioorganic & Medicinal Chemistry, 2017, 25, 4245-4252
Membrane-active foldamers have recently emerged as potential mimics of antimicrobial peptides (AMPs). Amphiphilic cationic helical N,N'-linked oligoureas are one such class of AMP mimics with activities in vitro against a broad range of bacteria including Bacillus anthracis, a Gram-positive sporulating bacillus and causing agent of anthrax. Here we have used site-selective chemical modifications of the oligourea backbone to gain additional insight into the relationship between structure and function and modulate anthracidal activity. A series of analogues in which urea linkages at selected positions are replaced by thiourea and guanidium surrogates have been prepared on solid support and tested against different bacterial forms of B. anthracis (germinated spores and encapsulated bacilli). Urea ? thiourea and urea? guanidinium replacements close to the negative end of the helix dipole led to analogues with increased potency and selectivity for B. anthracis versus mammalian cells. (C) 2017 Elsevier Ltd. All rights reserved.
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