Eric Hitti

Spectrin-like repeats 11-15 of human dystrophin show adaptations to a lipidic environment.

Dystrophin is essential to skeletal muscle function and confers resistance to the sarcolemma by interacting with cytoskeleton and membrane. In the present work, we characterized the behavior of dystrophin 11–15 (DYS R11–15), five spectrin-like repeats from the central domain of human dystrophin, with lipids. DYS R11–15 displays an amphiphilic character at the liquid/air interface while maintaining its secondary α-helical structure. The interaction of DYS R11–15 with small unilamellar vesicles (SUVs) depends on the lipid nature, which is not the case with large unilamellar vesicles (LUVs). In addition, switching from anionic SUVs to anionic LUVs suggests the lipid packing as a crucial factor for the interaction of protein and lipid. The monolayer model and the modulation of surface pressure aim to mimic the muscle at work (i.e. dynamic changes of muscle membrane during contraction and relaxation) (high and low surface pressure). Strikingly, the lateral pressure modifies the protein organization. Increasing the lateral pressure leads the proteins to be organized in a regular network. Nevertheless, a different protein conformation after its binding to monolayer is revealed by trypsin proteolysis. Label-free quantification by nano-LC/MS/MS allowed identification of the helices in repeats 12 and 13 involved in the interaction with anionic SUVs. These results, combined with our previous studies, indicate that DYS R11–15 constitutes the only part of dystrophin that interacts with anionic as well as zwitterionic lipids and adapts its interaction and organization depending on lipid packing and lipid nature. We provide strong experimental evidence for a physiological role of the central domain of dystrophin in sarcolemma scaffolding through modulation of lipid-protein interactions.

Qualitative and Spatial Metabolite Profiling of Lichens by a LC–MS Approach Combined With Optimised Extraction

INTRODUCTION Lichens are self-sustaining partnerships comprising fungi as shape-forming partners for their enclosed symbiotic algae. They produce a tremendous diversity of metabolites (1050 metabolites described so far). OBJECTIVES A comparison of metabolic profiles in nine lichen species belonging to three genera (Lichina, Collema and Roccella) by using an optimised extraction protocol, determination of the fragmentation pathway and the in situ localisation for major compounds in Roccella species. METHODS Chemical analysis was performed using a complementary study combining a Taguchi experimental design with qualitative analysis by high-performance liquid chromatography coupled with mass spectrometry techniques. RESULTS Optimal conditions to obtain the best total extraction yield were determined as follows: mortar grinding to a fine powder, two successive extractions, solid:liquid ratio (2:60) and 700 rpm stirring. Qualitative analysis of the metabolite profiling of these nine species extracted with the optimised method was corroborated using MS and MS/MS approaches. Nine main compounds were identified: 1 β-orcinol, 2 orsellinic acid, 3 putative choline sulphate, 4 roccellic acid, 5 montagnetol, 6 lecanoric acid, 7 erythrin, 8 lepraric acid and 9 acetylportentol, and several other compounds were reported. Identification was performed using the m/z ratio, fragmentation pathway and/or after isolation by NMR analysis. The variation of the metabolite profile in differently organised parts of two Roccella species suggests a specific role of major compounds in developmental stages of this symbiotic association. CONCLUSION Metabolic profiles represent specific chemical species and depend on the extraction conditions, the kind of the photobiont partner and the in situ localisation of major compounds.

MRI quantification of diffusion and perfusion in bone marrow by intravoxel incoherent motion (IVIM) and non-negative least square (NNLS) analysis

OBJECT To assess the feasibility of measuring diffusion and perfusion fraction in vertebral bone marrow using the intravoxel incoherent motion (IVIM) approach and to compare two fitting methods, i.e., the non-negative least squares (NNLS) algorithm and the more commonly used Levenberg-Marquardt (LM) non-linear least squares algorithm, for the analysis of IVIM data. MATERIALS AND METHODS MRI experiments were performed on fifteen healthy volunteers, with a diffusion-weighted echo-planar imaging (EPI) sequence at five different b-values (0, 50, 100, 200, 600 s/mm2), in combination with an STIR module to suppress the lipid signal. Diffusion signal decays in the first lumbar vertebra (L1) were fitted to a bi-exponential function using the LM algorithm and further analyzed with the NNLS algorithm to calculate the values of the apparent diffusion coefficient (ADC), pseudo-diffusion coefficient (D*) and perfusion fraction. RESULTS The NNLS analysis revealed two diffusion components only in seven out of fifteen volunteers, with ADC=0.60±0.09 (10(-3) mm(2)/s), D*=28±9 (10(-3) mm2/s) and perfusion fraction=14%±6%. The values obtained by the LM bi-exponential fit were: ADC=0.45±0.27 (10(-3) mm2/s), D*=63±145 (10(-3) mm2/s) and perfusion fraction=27%±17%. Furthermore, the LM algorithm yielded values of perfusion fraction in cases where the decay was not bi-exponential, as assessed by NNLS analysis. CONCLUSION The IVIM approach allows for measuring diffusion and perfusion fraction in vertebral bone marrow; its reliability can be improved by using the NNLS, which identifies the diffusion decays that display a bi-exponential behavior.

Wavelet optimization for classification

This paper addresses supervised signal classification using discrete time-scale representations. Given a set of learning signals and a class of discrete wavelet-basis, we propose to select the mother wavelet which yields the best classification results. This corresponds to determining the filter (used for the decomposition) that is ideally adapted to the specific classification problem at hand. It is realized by optimizing the filter coefficients according to a contrast criterion calculated on the learning set. Simulations show the efficiency of this approach.

Fat ViP MRI: Virtual Phantom Magnetic Resonance Imaging of water-fat systems.

OBJECT Virtual Phantom Magnetic Resonance Imaging (ViP MRI) is a method to generate reference signals on MR images, using external radiofrequency (RF) signals. The aim of this study was to assess the feasibility of ViP MRI to generate complex-data images of phantoms mimicking water-fat systems. MATERIALS AND METHODS Various numerical phantoms with a given fat fraction, T2* and field map were designed. The k-space of numerical phantoms was converted into RF signals to generate virtual phantoms. MRI experiments were performed at 4.7T using a multi-gradient-echo sequence on virtual and physical phantoms. The data acquisition of virtual and physical phantoms was simultaneous. Decomposition of the water and fat signals was performed using a complex-based water-fat separation algorithm. RESULTS Overall, a good agreement was observed between the fat fraction, T2* and phase map values of the virtual and numerical phantoms. In particular, fat fractions of 10.5±0.1 (vs 10% of the numerical phantom), 20.3±0.1 (vs 20%) and 30.4±0.1 (vs 30%) were obtained in virtual phantoms. CONCLUSION The ViP MRI method allows for generating imaging phantoms that i) mimic water-fat systems and ii) can be analyzed with water-fat separation algorithms based on complex data.

Preliminary MRI Quality Assessment and Device Acceptance Guidelines for a Multicenter Bioclinical Study: The GO Glioblastoma Project

It is a major challenge to guarantee homogeneous acquisition during a prospective multicenter magnetic resonance imaging (MRI) study that makes use of different devices. The goal of the multicenter Grand Ouest Glioblastoma Project (GOGP) was to correlate MRI quantitative parameters with biological markers extracted from image‐guided biopsies. Therefore, it was essential to ensure spatial coherence of the parameters as well as the signal intensity and homogeneity. The project included the same MRI protocol implemented on six devices from different manufacturers. The key point was the initial acceptance of the imaging devices and protocol sequences. For this purpose, and to allow comparison of quantitative patient data, we propose a specific method for quality assessment. A common quality control based on 10 parameters was established. Three pulse sequences of the clinical project protocol were applied using three test‐objects. A fourth test‐object was used to assess T1 accuracy. Although geometry‐related parameters, signal‐to‐noise ratio, uniformity, and T1 measurements varied slightly depending on the different devices, they nevertheless remained within the recommendations and expectations of the multicenter project. This kind of quality control procedure should be undertaken as a prerequisite to any multicenter clinical project involving quantitative MRI and comparison of data acquisitions with quantitative biological image‐guided biopsies.

MRI quantification of splenic iron concentration in mouse

To quantify hepatic and splenic iron load, which is a critical issue for iron overload disease diagnosis. MRI is useful to noninvasively determine liver iron concentration, but not proven to be adequate for robust evaluation of splenic iron load. We evaluated the usefulness of MRI‐derived parameters to determine splenic iron concentration in mice.

Abrupt changes detection in the time-scale and in the time-frequency planes: a comparative study

We present in this paper a comparison between two segmentation approaches based on time-scale (t-s) and time-frequency (t-f) analysis. The purpose is to detect abrupt changes in the spectral characteristics of multicomponent nonstationary signals. After exposing the two procedures, we carry out different synthetic simulations in order to test and compare the performances of the methods for strictly and not strictly stepwise stationary signals. Finally, we apply the algorithms to musical signals which are not exactly piecewise stationary. This study demonstrates the potential segmentation capability of the proposed methods.

Eliminating the blood-flow confounding effect in intravoxel incoherent motion (IVIM) using the non-negative least square analysis in liver

Tissue perfusion measurements using intravoxel incoherent motion (IVIM) diffusion‐MRI are of interest for investigations of liver pathologies. A confounding factor in the perfusion quantification is the partial volume between liver tissue and large blood vessels. The aim of this study was to assess and correct for this partial volume effect in the estimation of the perfusion fraction.

Human Dystrophin Rod 11-15 Sub-Domain: A Membrane Interacting Zone Modulated by Lipid Packing

Dystrophin is essential for skeletal muscle function and confers resistance to the sarcolemma by interacting with cytoskeletal and membrane partners. We investigated here protein-lipid interaction of a five repeat from the central domain of dystrophin (DYS R11-15) also referred as an actin binding domain. In a first step, we demonstrated that DYS R11-15 interacts more strongly with anionic than with zwitterionic small unilamellar vesicles. Using large unilamellar vesicles with different radius and trypsin accessibility assays, we showed that the protein presents different conformation depending on vesicle curvature and lipid nature. Using label-free quantification mass spectroscometry, a protein domain protected from proteolysis in presence of anionic vesicles was observed while a protein domain more accessible to trypsin in the presence of either anionic or zwitterionic vesicles was identified. In a second step, we studied the adsorption behavior of the protein at the air-liquid and lipid-liquid interface in a Langmuir trough. DYS R11-15 displayed a surface activity while maintaining its α-helical secondary structure as shown by PM-IRRAS. At 16mN/m lateral pressure of monolayer lipid film, few protein clusters were observed by AFM, while at 20 and 30mN/m, a striking protein network was formed with both negative and zwitterionic phospholipids. However, image analysis and behaviour of the networks towards trypsination in the trough as revealed by AFM showed that trypsin accessibility to the protein network depends on the surface pressure as well as on the nature of the phospholipid. These results indicate that DYS R11-15 constitutes part of the dystrophin protein for which anchoring and interaction with membrane depend on the packing and the nature of lipids. Such behaviour provides a strong experimental support for a physiological role of dystrophin central domain in contraction-relaxation cycles and dynamics of muscle cells.