Etienne Harté

Detection of an estrogen derivative in two breast cancer cell lines using a single core multimodal probe for imaging (SCoMPI) imaged by a panel of luminescent and vibrational techniques

3-Methoxy-17α-ethynylestradiol or mestranol is a prodrug for ethynylestradiol and the estrogen component of some oral contraceptive formulations. We demonstrate here that a single core multimodal probe for imaging - SCoMPI - can be efficiently grafted onto mestranol allowing its tracking in two breast cancer cell lines, MDA-MB-231 and MCF-7 fixed cells. Correlative imaging studies based on luminescence (synchrotron UV spectromicroscopy, wide field and confocal fluorescence microscopies) and vibrational (AFMIR, synchrotron FTIR spectromicroscopy, synchrotron-based multiple beam FTIR imaging, confocal Raman microspectroscopy) spectroscopies were consistent with one another and showed a Golgi apparatus distribution of the SCoMPI-mestranol conjugate in both cell lines.

New concepts in molecular imaging: non-invasive MRI spotting of proteolysis using an Overhauser effect switch.

Background Proteolysis, involved in many processes in living organisms, is tightly regulated in space and time under physiological conditions. However deregulation can occur with local persistent proteolytic activities, e.g. in inflammation, cystic fibrosis, tumors, or pancreatitis. Furthermore, little is known about the role of many proteases, hence there is a need of new imaging methods to visualize specifically normal or disease-related proteolysis in intact bodies. Methodology/Principal Findings In this paper, a new concept for non invasive proteolysis imaging is proposed. Overhauser-enhanced Magnetic Resonance Imaging (OMRI) at 0.2 Tesla was used to monitor the enzymatic hydrolysis of a nitroxide-labeled protein. In vitro, image intensity switched from 1 to 25 upon proteolysis due to the associated decrease in the motional correlation time of the substrate. The OMRI experimental device used in this study is consistent with protease imaging in mice at 0.2 T without significant heating. Simulations show that this enzymatic-driven OMRI signal switch can be obtained at lower frequencies suitable for larger animals or humans. Conclusions/Significance The method is highly sensitive and makes possible proteolysis imaging in three dimensions with a good spatial resolution. Any protease could be targeted specifically through the use of taylor-made cleavable macromolecules. At short term OMRI of proteolysis may be applied to basic research as well as to evaluate therapeutic treatments in small animal models of experimental diseases.

Bimetallic cyanido-bridged magnetic materials derived from manganese(III) Schiff-base complexes and pentacyanidonitrosylferrate(II) precursor

Three heterobimetallic MnIIIFeII complexes: [Mn(5-Br-salpn)(H2O)]2[Fe(CN)5NO]·2H2O (1), [{Mn(salen)}2Fe(CN)5NO]·2H2O (2) and [{Mn(saltmen)}4Fe(CN)5NO](ClO4)2·H2O·2CH3OH (3), have been synthesized by the reactions of MnIII/Schiff–base (SB) complexes, [Mn(SB)(H2O)]+, (SB being salen2− = N,N′-ethylenebis(salicylideneiminato) dianion; 5-Br-salpn2− = N,N′-1,3-propylenebis(5-bromosalicylideneiminato) dianion or saltmen2− = N,N′-(1,1,2,2-tetramethylethylene) bis(salicylideneiminato) dianion) with the [Fe(CN)5NO]2− building block. X-Ray diffraction analyses on single crystals reveal that 1 has a trinuclear molecular structure, 2 displays a two-dimensional (2D) network structure, and 3 possesses a new 2D network of MnIII dinuclear motifs. The magnetic measurements show that the interaction between the MnIIIS = 2 spins through the [NC–Fe–CN] bridges is systematically of antiferromagnetic nature: J/kB = −0.95(5) K and J/kB = −1.15(5) K for 1 and 2, respectively, while the direct MnIII–MnIII interaction, in the {Mn2(saltmen)2} dinuclear units present in 3, is ferromagnetic with J/kB = +1.75(5) K. Due to the ST = 4 spin ground state of these dinuclear units and the uniaxial magnetic anisotropy brought by the Mn(III) metal ions, 3 exhibits single-molecule magnet properties. The 400–900 nm optical properties of the three compounds have been also investigated, showing no significant photoactivity unlike in the Na2[Fe(CN)5NO] precursor.

Interaction of Aβ1–42 Amyloids with Lipids Promotes “Off-Pathway” Oligomerization and Membrane Damage

The toxicity of amyloids, as Aβ(1-42) involved in Alzheimer disease, is a subject under intense scrutiny. Many studies link their toxicity to the existence of various intermediate structures prior to fiber formation and/or their specific interaction with membranes. In this study we focused on the interaction between membrane models and Aβ(1-42) peptides and variants (L34T, mG37C) produced in E. coli and purified in monomeric form. We evaluated the interaction of a toxic stable oligomeric form (oG37C) with membranes as comparison. Using various biophysical techniques as fluorescence and plasmon waveguide resonance, we clearly established that the oG37C interacts strongly with membranes leading to its disruption. All the studied peptides destabilized liposomes and accumulated slowly on the membrane (rate constant 0.02 min(-1)). Only the oG37C exhibited a particular pattern of interaction, comprising two steps: the initial binding followed by membrane reorganization. Cryo-TEM was used to visualize the peptide effect on liposome morphologies. Both oG37C and mG37C lead to PG membrane fragmentation. The PG membrane promotes peptide oligomerization, implicated in membrane disruption. WT (Aβ(1-42)) also perturbs liposome organization with membrane deformation rather than disruption. For all the peptides studied, their interaction with the membranes changes their fibrillization process, with less fibers and more small aggregates being formed. These studies allowed to establish, a correlation between toxicity, fiber formation, and membrane disruption.

Probing the kinetics of lipid membrane formation and the interaction of a nontoxic and a toxic amyloid with plasmon waveguide resonance

The kinetics of formation of solid-supported lipid model membranes were investigated using a home-made plasmon waveguide resonance (PWR) sensor possessing enhanced properties relative to classic surface plasmon resonance sensors. Additionally, the kinetics of interaction of two amyloid peptides with zwitterionic and anionic membranes and their effect on lipid organization were followed.

Real time monitoring of membrane GPCR reconstitution by plasmon waveguide resonance: on the role of lipids

G-protein coupled receptors (GPCRs) are important therapeutic targets since more than 40% of the drugs on the market exert their action through these proteins. To decipher the molecular mechanisms of activation and signaling, GPCRs often need to be isolated and reconstituted from a detergent-solubilized state into a well-defined and controllable lipid model system. Several methods exist to reconstitute membrane proteins in lipid systems but usually the reconstitution success is tested at the end of the experiment and often by an additional and indirect method. Irrespective of the method used, the reconstitution process is often an intractable and time-consuming trial-and-error procedure. Herein, we present a method that allows directly monitoring the reconstitution of GPCRs in model planar lipid membranes. Plasmon waveguide resonance (PWR) allows following GPCR lipid reconstitution process without any labeling and with high sensitivity. Additionally, the method is ideal to probe the lipid effect on receptor ligand binding as demonstrated by antagonist binding to the chemokine CCR5 receptor.

Specific Anti-Leukemic Activity of the Peptide Warnericin RK and Analogues and Visualization of Their Effect on Cancer Cells by Chemical Raman Imaging

Antimicrobial peptides can be used as therapeutic agents against cancer cells. Warnericin RK and derivatives (WarnG20D and WarnF14V) were tested on various, solid tumor or leukemia, cancer cells. These peptides appeared to be cytotoxic on all the cell types tested, cancerous as well healthy, but very interestingly displayed no deleterious effect on healthy mononuclear cells. The mode of action of the peptide was proposed to be membranolytic, using chemical Raman imaging. Addition of peptide induced a large disorganization of the membrane leading to the loss of the content of inner compartments of Jurkat cell, whereas no effect was observed on the healthy mononuclear cells. The less hemolytic peptides WarnG20D and WarnF14V could be good candidates for the leukemia treatment.