Multi-scale systems analysis of plant development : beyond cellular signaling and tissue morphodynamics

Les plantes sont essentielles pour les sociétés humaines. Notre alimentation quotidienne, les matériaux de constructions et les sources énergétiques dérivent de la biomasse végétale. En revanche, la compréhension des multiples aspects développementaux des plantes est encore peu exploitée et représente un sujet de recherche majeur pour la science. L'émergence des technologies à haut débit pour le séquençage de génome à grande échelle ou l'imagerie de haute résolution permet à présent de produire des quantités énormes d'information. L'analyse informatique est une façon d'intégrer ces données et de réduire la complexité apparente vers une échelle d'abstraction appropriée, dont la finalité est de fournir des perspectives de recherches ciblées. Ceci représente la raison première de cette thèse. En d'autres termes, nous appliquons des méthodes descriptives et prédictives combinées à des simulations numériques afin d'apporter des solutions originales à des problèmes relatifs à la morphogénèse à l'échelle de la cellule et de l'organe. Nous nous sommes fixés parmi les objectifs principaux de cette thèse d'élucider de quelle manière l'interaction croisée des phytohormones auxine et brassinosteroïdes (BRs) détermine la croissance de la cellule dans la racine du méristème apical d'Arabidopsis thaliana, l'organisme modèle de référence pour les études moléculaires en plantes. Pour reconstruire le réseau de signalement cellulaire, nous avons extrait de la littérature les informations pertinentes concernant les relations entre les protéines impliquées dans la transduction des signaux hormonaux. Le réseau a ensuite été modélisé en utilisant un formalisme logique et qualitatif pour pallier l'absence de données quantitatives. Tout d'abord, Les résultats ont permis de confirmer que l'auxine et les BRs agissent en synergie pour contrôler la croissance de la cellule, puis, d'expliquer des observations phénotypiques paradoxales et au final, de mettre à jour une interaction clef entre deux protéines dans la maintenance du méristème de la racine. Une étude ultérieure chez la plante modèle Brachypodium dystachion (Brachypo- dium) a révélé l'ajustement du réseau d'interaction croisée entre auxine et éthylène par rapport à Arabidopsis. Chez ce dernier, interférer avec la biosynthèse de l'auxine mène à la formation d'une racine courte. Néanmoins, nous avons isolé chez Brachypodium un mutant hypomorphique dans la biosynthèse de l'auxine qui affiche une racine plus longue. Nous avons alors conduit une analyse morphométrique qui a confirmé que des cellules plus anisotropique (plus fines et longues) sont à l'origine de ce phénotype racinaire. Des analyses plus approfondies ont démontré que la différence phénotypique entre Brachypodium et Arabidopsis s'explique par une inversion de la fonction régulatrice dans la relation entre le réseau de signalisation par l'éthylène et la biosynthèse de l'auxine. L'analyse morphométrique utilisée dans l'étude précédente exploite le pipeline de traitement d'image de notre méthode d'histologie quantitative. Pendant la croissance secondaire, la symétrie bilatérale de l'hypocotyle est remplacée par une symétrie radiale et une organisation concentrique des tissus constitutifs. Ces tissus sont initialement composés d'une douzaine de cellules mais peuvent aisément atteindre des dizaines de milliers dans les derniers stades du développement. Cette échelle dépasse largement le seuil d'investigation par les moyens dits 'traditionnels' comme l'imagerie directe de tissus en profondeur. L'étude de ce système pendant cette phase de développement ne peut se faire qu'en réalisant des coupes fines de l'organe, ce qui empêche une compréhension des phénomènes cellulaires dynamiques sous-jacents. Nous y avons remédié en proposant une stratégie originale nommée, histologie quantitative. De fait, nous avons extrait l'information contenue dans des images de très haute résolution de sections transverses d'hypocotyles en utilisant un pipeline d'analyse et de segmentation d'image à grande échelle. Nous l'avons ensuite combiné avec un algorithme de reconnaissance automatique des cellules. Cet outil nous a permis de réaliser une description quantitative de la progression de la croissance secondaire révélant des schémas développementales non-apparents avec une inspection visuelle classique. La formation de pôle de phloèmes en structure répétée et espacée entre eux d'une longueur constante illustre les bénéfices de notre approche. Par ailleurs, l'exploitation approfondie de ces résultats a montré un changement de croissance anisotropique des cellules du cambium et du phloème qui semble en phase avec l'expansion du xylème. Combinant des outils génétiques et de la modélisation biomécanique, nous avons démontré que seule la croissance plus rapide des tissus internes peut produire une réorientation de l'axe de croissance anisotropique des tissus périphériques. Cette prédiction a été confirmée par le calcul du ratio des taux de croissance du xylème et du phloème au cours de développement secondaire ; des ratios élevés sont effectivement observés et concomitant à l'établissement progressif et tangentiel du cambium. Ces résultats suggèrent un mécanisme d'auto-organisation établi par un gradient de division méristématique qui génèrent une distribution de contraintes mécaniques. Ceci réoriente la croissance anisotropique des tissus périphériques pour supporter la croissance secondaire. - Plants are essential for human society, because our daily food, construction materials and sustainable energy are derived from plant biomass. Yet, despite this importance, the multiple developmental aspects of plants are still poorly understood and represent a major challenge for science. With the emergence of high throughput devices for genome sequencing and high-resolution imaging, data has never been so easy to collect, generating huge amounts of information. Computational analysis is one way to integrate those data and to decrease the apparent complexity towards an appropriate scale of abstraction with the aim to eventually provide new answers and direct further research perspectives. This is the motivation behind this thesis work, i.e. the application of descriptive and predictive analytics combined with computational modeling to answer problems that revolve around morphogenesis at the subcellular and organ scale. One of the goals of this thesis is to elucidate how the auxin-brassinosteroid phytohormone interaction determines the cell growth in the root apical meristem of Arabidopsis thaliana (Arabidopsis), the plant model of reference for molecular studies. The pertinent information about signaling protein relationships was obtained through the literature to reconstruct the entire hormonal crosstalk. Due to a lack of quantitative information, we employed a qualitative modeling formalism. This work permitted to confirm the synergistic effect of the hormonal crosstalk on cell elongation, to explain some of our paradoxical mutant phenotypes and to predict a novel interaction between the BREVIS RADIX (BRX) protein and the transcription factor MONOPTEROS (MP),which turned out to be critical for the maintenance of the root meristem. On the same subcellular scale, another study in the monocot model Brachypodium dystachion (Brachypodium) revealed an alternative wiring of auxin-ethylene crosstalk as compared to Arabidopsis. In the latter, increasing interference with auxin biosynthesis results in progressively shorter roots. By contrast, a hypomorphic Brachypodium mutant isolated in this study in an enzyme of the auxin biosynthesis pathway displayed a dramatically longer seminal root. Our morphometric analysis confirmed that more anisotropic cells (thinner and longer) are principally responsible for the mutant root phenotype. Further characterization pointed towards an inverted regulatory logic in the relation between ethylene signaling and auxin biosynthesis in Brachypodium as compared to Arabidopsis, which explains the phenotypic discrepancy. Finally, the morphometric analysis of hypocotyl secondary growth that we applied in this study was performed with the image-processing pipeline of our quantitative histology method. During its secondary growth, the hypocotyl reorganizes its primary bilateral symmetry to a radial symmetry of highly specialized tissues comprising several thousand cells, starting with a few dozens. However, such a scale only permits observations in thin cross-sections, severely hampering a comprehensive analysis of the morphodynamics involved. Our quantitative histology strategy overcomes this limitation. We acquired hypocotyl cross-sections from tiled high-resolution images and extracted their information content using custom high-throughput image processing and segmentation. Coupled with an automated cell type recognition algorithm, it allows precise quantitative characterization of vascular development and reveals developmental patterns that were not evident from visual inspection, for example the steady interspace distance of the phloem poles. Further analyses indicated a change in growth anisotropy of cambial and phloem cells, which appeared in phase with the expansion of xylem. Combining genetic tools and computational modeling, we showed that the reorientation of growth anisotropy axis of peripheral tissue layers only occurs when the growth rate of central tissue is higher than the peripheral one. This was confirmed by the calculation of the ratio of the growth rate xylem to phloem throughout secondary growth. High ratios are indeed observed and concomitant with the homogenization of cambium anisotropy. These results suggest a self-organization mechanism, promoted by a gradient of division in the cambium that generates a pattern of mechanical constraints. This, in turn, reorients the growth anisotropy of peripheral tissues to sustain the secondary growth.

Anterior inferior cerebellar artery aneurysms mimicking vestibular schwannomas.

BACKGROUND AND PURPOSE: Unruptured anterior inferior cerebellar artery (AICA) aneurysms are rare but potentially lethal cerebellopontine angle (CPA) lesions that may be misdiagnosed as vestibular schwannomas when they present with vestibulo-cochlear symptoms. METHODS: We report two cases of unruptured but symptomatic AICA aneurysms initially referred to us as atypical vestibular schwannomas requiring surgery. Two discriminant MR features are described. RESULTS: One patient refused treatment. The other was successfully treated by coil occlusion. CONCLUSIONS: Caution is advised before suspecting a CPA mass to be a purely extra-canalicular schwannoma, given its extreme rarity. Deafness and cerebellar ischemia may be prevented if AICA aneurysms are correctly identified preoperatively. In the absence of specific arterial imaging, two MR features may distinguish them from vestibular schwannomas: (1) the absence of internal auditory canal enlargement and (2) the "blurry dot sign," representing blood flow artefacts on pre- and postcontrast studies.

Statin treatment is associated with improved prognosis in patients with AF-related stroke.

BACKGROUND/OBJECTIVES: The most recent ACC/AHA guidelines recommend high-intensity statin therapy in ischemic stroke patients of presumably atherosclerotic origin. On the contrary, there is no specific recommendation for the use of statin in patients with non-atherosclerotic stroke, e.g. strokes related to atrial fibrillation (AF). We investigated whether statin treatment in patients with AF-related stroke is associated with improved survival and reduced risk for stroke recurrence and future cardiovascular events. METHODS: All consecutive patients registered in the Athens Stroke Registry with AF-related stroke and no history of coronary artery disease nor clinically manifest peripheral artery disease were included in the analysis and categorized in two groups depending on whether statin was prescribed at discharge. The primary outcome was overall mortality; the secondary outcomes were stroke recurrence and a composite cardiovascular endpoint comprising of recurrent stroke, myocardial infarction, aortic aneurysm rupture or sudden cardiac death during the 5-year follow-up. RESULTS: Among 1602 stroke patients, 404 (25.2%) with AF-related stroke were included in the analysis, of whom 102 (25.2%) were discharged on statin. On multivariate Cox-proportional-hazards model, statin treatment was independently associated with a lower mortality (hazard-ratio (HR): 0.49, 95%CI:0.26-0.92) and lower risk for the composite cardiovascular endpoint during the median 22months follow-up (HR: 0.44, 95%CI:0.22-0.88), but not with stroke recurrence (HR: 0.47, 95%CI:0.22-1.01, p: 0.053). CONCLUSIONS: In this long-term registry of patients with AF-related stroke, statin treatment was associated with improved survival and reduced risk for future cardiovascular events.

Associations of baroreflex sensitivity, heart rate variability, and initial orthostatic hypotension with prenatal and recent postnatal methylmercury exposure in the Seychelles Child Development Study at age 19 years.

BACKGROUND: A few studies have suggested an association between prenatal exposure to methylmercury and decreased heart rate variability (HRV) related to autonomic heart function, but no study has examined this association using baroreflex sensitivity (BRS). In this study we assessed the distribution of BRS and immediate orthostatic hypotension (IOH) in young Seychellois adults and their associations with exposure to prenatal and recent postnatal methylmercury. METHODS: Subjects in the Seychelles Child Development Study (SCDS) main cohort were evaluated at age 19 years. Non-invasive beat-to-beat blood pressure (BP) monitoring (Finapres, Ohmeda) was performed at rest and during active standing in 95 consecutive subjects. Recent postnatal mercury exposure was measured in subjects' hair at the age of 19 years and prenatal exposure in maternal hair grown during pregnancy. BRS was estimated by sequence analysis to identify spontaneous ascending and descending BP ramps. HRV was estimated by the following markers: PNN50 (relative numbers of normal-to-normal intervals which are shorter by more than 50 ms than the immediately following normal-to-normal intervals); rMSSD (root mean of the squared sum of successive interval differences); LF/HF (low frequency/high frequency component ratio); ratio of the mean expiratory/inspiratory RR intervals (EI ratio); and the ratio between the longest RR interval 30 s after active standing and the shortest RR interval at 15 s (Max30/Min15). IOH was estimated by the deepest BP fall within the first 15 s after active standing up. RESULTS: Prenatal MeHg exposures were similar in boys and girls (6.7±4.3, 6.7±3.8 ng/g) but recent postnatal mercury levels were higher in males than females (11.2±5.8 vs 7.9±4.3 ng/g, p=0.003). Markers of autonomic heart rate control were within the normal range (BRS: 24.8±7 ms/mm Hg, PNN50: 24.9±6.8%, rMSSD: 68±22, LF/HF: 0.61±0.28) in both sexes. After standing, 51.4% of subjects had a transient systolic BP drop>40 mm Hg, but only 5.3% reported dizziness or had syncope. Prenatal and recent postnatal MeHg levels, overall, were not associated with BRS, E/I ratio, PNN50, rMSSD, LF/HF ratio, Max30/Min15 ratio, and IOH. CONCLUSIONS: This study provides no support for the hypothesis that prenatal or recent postnatal MeHg exposure from fish consumption is associated with impaired autonomic heart rate control.

Intravoxel incoherent motion diffusion-weighted imaging in the liver: comparison of mono-, bi- and tri-exponential modelling at 3.0-T.

PURPOSE: To determine whether a mono-, bi- or tri-exponential model best fits the intravoxel incoherent motion (IVIM) diffusion-weighted imaging (DWI) signal of normal livers. MATERIALS AND METHODS: The pilot and validation studies were conducted in 38 and 36 patients with normal livers, respectively. The DWI sequence was performed using single-shot echoplanar imaging with 11 (pilot study) and 16 (validation study) b values. In each study, data from all patients were used to model the IVIM signal of normal liver. Diffusion coefficients (Di ± standard deviations) and their fractions (fi ± standard deviations) were determined from each model. The models were compared using the extra sum-of-squares test and information criteria. RESULTS: The tri-exponential model provided a better fit than both the bi- and mono-exponential models. The tri-exponential IVIM model determined three diffusion compartments: a slow (D1 = 1.35 ± 0.03 × 10(-3) mm(2)/s; f1 = 72.7 ± 0.9 %), a fast (D2 = 26.50 ± 2.49 × 10(-3) mm(2)/s; f2 = 13.7 ± 0.6 %) and a very fast (D3 = 404.00 ± 43.7 × 10(-3) mm(2)/s; f3 = 13.5 ± 0.8 %) diffusion compartment [results from the validation study]. The very fast compartment contributed to the IVIM signal only for b values ≤15 s/mm(2) CONCLUSION: The tri-exponential model provided the best fit for IVIM signal decay in the liver over the 0-800 s/mm(2) range. In IVIM analysis of normal liver, a third very fast (pseudo)diffusion component might be relevant. KEY POINTS: ? For normal liver, tri-exponential IVIM model might be superior to bi-exponential ? A very fast compartment (D = 404.00 ± 43.7 × 10 (-3)  mm (2) /s; f = 13.5 ± 0.8 %) is determined from the tri-exponential model ? The compartment contributes to the IVIM signal only for b ≤ 15 s/mm (2.)

The multi-facets of sustainable nanotechnology : lessons from a nanosafety symposium

An international symposium for nanosafety was held recently at the Nanyang Technological University in Singapore. Topics relating to understanding nanomaterial properties, tools, and infrastructure required for predicting hazardous outcomes, measuring nanomaterial exposure levels, systems approach for risk assessment and public's perception of nanotechnology were covered. The need for a multidisciplinary approach, across both natural and social sciences, for developing sustainable nanotechnology solutions was heavily emphasized. This commentary highlights the major issues discussed and the commitment of the nanosafety research community in Singapore to contribute collectively to realise the vision of sustainable nanotechnology.

Predicting drug metabolism: experiment and/or computation?

Drug metabolism can produce metabolites with physicochemical and pharmacological properties that differ substantially from those of the parent drug, and consequently has important implications for both drug safety and efficacy. To reduce the risk of costly clinical-stage attrition due to the metabolic characteristics of drug candidates, there is a need for efficient and reliable ways to predict drug metabolism in vitro, in silico and in vivo. In this Perspective, we provide an overview of the state of the art of experimental and computational approaches for investigating drug metabolism. We highlight the scope and limitations of these methods, and indicate strategies to harvest the synergies that result from combining measurement and prediction of drug metabolism.

Cardiogene: an innovative multidisciplinary consultation for genetic arrhythmias

Introduction: Over the past decade clinically relevant progress has been made regarding the genetic origin of sudden cardiac death due to arrhythmic syndromes such as congenital long QT syndrome (LQTS), Brugada syndrome (BrS), catecholinergic polymorphic ventricular tachycardia (CPVT) and short QT (SQTS). An increased number of patients are diagnosed and their offspring sent for screening. In order to optimize care of these families we have set up a multidisciplinary consultation, "Cardiogene", consisting of a pediatric and an adult cardiologist and a clinical geneticist. All families are seen at a common consult in order to take the family history, genetic background and to explain the disease to patients and their families. Appropriate cardiac investigations and genetic testing are then performed and the families seen again in a multidisciplinary fashion for the results. We have reviewed all our cases over the past 5 years. Methods: retrospective review of all cases seen at Cardiogene Clinic for suspicion of arrhythmic syndromes since 2007. Results: 23 families were seen at the Cardiogene Clinic with a total of 41 children. The suspected arrhythmic syndrome was LQTS in 14 families (26 children), BrS in 7 families (14 children), SQTS in1 family (2 children) and CPVT in 1 family (3 children). Of the 41 children 17 were genetically positive for an arrhythmic syndrome: 14 were for LQTS, 3 for BrS. 24 children were genetically negative however 4 of those were phenotypically positive: 2 LQTS, 1 BrS and 1 CPVT. In 3 families the diagnosis was initially made in a child and then found in the parent. In 2 families the diagnosis was made after a sudden death of one of their children, 1 LQTS (3 week old child), 1 BrS (20 year old). Discussion: Genetic testing is an essential part of diagnosis and permits an improved targeting of patients needing follow-up and treatment. In our series, a mutation has been found in most families with LQTS. In all other genetic arrhythmias, the yield of genetic testing is less but nevertheless helpful for medical care of these pts. Conclusion: A multidisciplinary approach to genetic arrhythmias permits a better and more efficient screening and therapy in affected families. It helps families to better understand their disease and improves follow-up in the affected individuals.

Overspeed HIIT in Lower-Body Positive Pressure Treadmill Improves Running Performance.

PURPOSE: Optimal high-intensity interval training (HIIT) regimens for running performance are unknown, although most protocols result in some benefit to key performance factors (running economy (RE), anaerobic threshold (AT), or maximal oxygen uptake (V˙O2max)). Lower-body positive pressure (LBPP) treadmills offer the unique possibility to partially unload runners and reach supramaximal speeds. We studied the use of LBPP to test an overspeed HIIT protocol in trained runners. METHODS: Eleven trained runners (35 ± 8 yr, V˙O2max, 55.7 ± 6.4 mL·kg·min) were randomized to an LBPP (n = 6) or a regular treadmill (CON, n = 5), eight sessions over 4 wk of HIIT program. Four to five intervals were run at 100% of velocity at V˙O2max (vV˙O2max) during 60% of time to exhaustion at vV˙O2max (Tlim) with a 1:1 work:recovery ratio. Performance outcomes were 2-mile track time trial, V˙O2max, vV˙O2max, vAT, Tlim, and RE. LBPP sessions were carried out at 90% body weight. RESULTS: Group-time effects were present for vV˙O2max (CON, 17.5 vs. 18.3, P = 0.03; LBPP, 19.7 vs. 22.3 km·h; P < 0.001) and Tlim (CON, 307.0 vs. 404.4 s, P = 0.28; LBPP, 444.5 vs. 855.5, P < 0.001). Simple main effects for time were present for field performance (CON, -18; LBPP, -25 s; P = 0.002), V˙O2max (CON, 57.6 vs. 59.6; LBPP, 54.1 vs. 55.1 mL·kg·min; P = 0.04) and submaximal HR (157.7 vs. 154.3 and 151.4 vs. 148.5 bpm; P = 0.002). RE was unchanged. CONCLUSIONS: A 4-wk HIIT protocol at 100% vV˙O2max improves field performance, vV˙O2max, V˙O2max and submaximal HR in trained runners. Improvements are similar if intervals are run on a regular treadmill or at higher speeds on a LPBB treadmill with 10% body weight reduction. LBPP could provide an alternative for taxing HIIT sessions.

Role of Munc18c in SNARE-mediated exocytosis

Background: The SNARE (Soluble N-ethylmaleimide-sensitive factor Attachment protein Receptors) and SM (Sec1/Munc18) family of proteins form the core machinery that drives the fusion of vesicles in different membrane trafficking steps. They are highly conserved, implying a similar mode of binding and function. In vertebrates, Munc18a is essential for neuronal exocytosis. It binds to its partner syntaxin1a (Syx1a) at both its N-peptide and closed conformation, and thereby inhibits SNARE complex formation in vitro. By contrast, its close homolog Munc18c is thought to interact with only the N-peptide of its partner Syx4. Moreover, different effects of Munc18c on SNARE complex formation have been reported, suggesting that the two Munc18/Syx pairs act differently. Objective: The aim of the present study was to investigate whether the mechanism of action of Munc18c indeed deviates from that of Munc18a by using sensitive biochemical and biophysical methods. Results: I found that Munc18c does have a similar binding mode as Munc18a and interacts tightly with Syx4 at both the N-peptide and closed conformation. Moreover, I established, through a novel assay, that Munc18c inhibits SNARE complex assembly, with both the binding sites contributing to inhibition, similar to Munc18a. However, there were several subtle differences between the two Munc18/Syx pairs. Munc18a exerted stronger inhibition than Munc18c. Also their respective Syx partners were found to differ in the rate of binding to SNAP25, suggesting that the equilibrium of their open and closed conformations is different. Moreover, Munc18a was found to interact with Syx 1, 2, 3 but not 4, while Munc18c bound to Syx 2, 4 and 1 but not 3. By comparing the kinetics of interaction of Syx with either Munc18 or SNAP25, I found that the block of SNARE complex assembly by Munc18 is effective on a shorter time scale, but SNAP25 eventually binds to Syx resulting in SNARE complex formation. Nevertheless, these findings do not explain how Syx can escape the tight grip of Munc18, suggesting that other proteins or mechanisms are needed for this step. I also discovered that Munc18 is able to bind on the surface of the SNARE core complex; however, this observation needs to be tested more rigorously. Conclusion: Munc18c was found to be similar to Munc18a in its mode of binding to Syx and inhibition of SNARE complex assembly. However, differences in kinetics and interaction specificities were observed between the different Munc18/Syx pairs. -- Contexte : Les familles des protéines SNARE (Soluble N-ethylmaleimide-sensitive factor At- tachment protein Receptors) et SM (Sec1/Munc18) forment le coeur de la machinerie chargée de la fusion vésiculaire au cours des différentes étapes du trafic intracellulaire. Elles sont très conservées, suggérant un mode d'interaction et des fonctions semblables. Chez les Verté- brés, Munc18a est essentielle à l'exocytose neuronale. Elle se lie à sa partenaire d'interaction syntaxin1a (Syx1a) à la fois via un peptide N-terminal et la conformation fermée de celle-ci, inhibant ainsi la formation du complexe SNARE in vitro. Son homologue proche Munc18c au contraire, est supposée interagir seulement avec le peptide N-terminal de sa partenaire Syx4. En outre, différents effets de Munc18c sur la formation du complexe SNARE ont été décrits, suggérant que les deux paires Munc18/Syx fonctionnent différemment. Objectif : Le but de cette étude est de tester si les mécanismes de fonctionnement de Munc18c diffèrent vraiment de ceux de Munc18a par le biais de méthodes biochimiques et biophysiques très précises. Résultats : J'ai pu démontrer que Munc18c se comporte en effet de façon semblable à Munc18a, et interagit étroitement avec Syx4 à ses deux sites de liaison. J'ai pu de surcroît montrer par une nouvelle méthode que Munc18c inhibe l'assemblage du complexe SNARE en impliquant ces deux sites de liaison, comme le fait Munc18a. il existe cependant de subtiles différences entre les deux paires Munc18/Syx : Munc18a exerce une inhibition plus forte que Munc18c ; leurs Syx partenaires diffèrent également dans leur degré de liaison à SNAP25, ce qui suggère un équilibre different de leurs conformations ouverte et fermée. De plus, Munc18a interagit avec Syx 1, 2 et 3 mais pas Syx 4, alors que Munc18c se lie à Syx 2, 4 et 1 mais pas Syx 3. En comparant les cinétiques d'interaction de Syx avec Munc18 ou SNAP25, j'ai découvert que le blocage par Munc18 de l'assemblage du complexe SNARE est effectif de façon brève, bien que SNAP25 finisse par se lier à Syx et aboutir ainsi à la formation du complexe SNARE. Ces découvertes n'expliquent cependant pas comment Syx parvient à échapper à la solide emprise de Munc18, et suggèrent ainsi l'intervention nécessaire d'autres protéines ou mécanismes à cette étape. J'ai également découvert que Munc18 peut se lier à la surface de la partie centrale du complexe SNARE - cette observation reste à être testée de façon plus stringente. Conclusion : Il a pu être établi que Munc18c est semblable à Munc18a quant à son mode de liaison à Syx et d'inhibition de l'assemblage du complexe SNARE. Des différences de cinétique et de spécificité d'interaction entre les diverses paires Munc18/Syx ont cependant été identifiées.

Transcranial Direct Current Stimulation for the Treatment of Chronic Tinnitus: A Randomized Controlled Study.

BACKGROUND: Tinnitus is an often disabling condition for which there is no effective therapy. Current research suggests that tinnitus may develop due to maladaptive plastic changes and altered activity in the auditory and prefrontal cortex. Transcranial direct current stimulation (tDCS) modulates brain activity and has been shown to transiently suppress tinnitus in trials. OBJECTIVE: To investigate the efficacy and safety of tDCS in the treatment of chronic subjective tinnitus. METHODS: In a randomized, parallel, double-blind, sham-controlled study, the efficacy and safety of cathodal tDCS to the auditory cortex with anode over the prefrontal cortex was investigated in five sessions over five consecutive days. Tinnitus was assessed after the last session on day 5, and at follow-up visits 1 and 3 months post stimulation using the Tinnitus Handicap Inventory (THI, primary outcome measure), Subjective Tinnitus Severity Scale, Hospital Anxiety and Depression scale, Visual Analogue Scale, and Clinical Global Impression scale. RESULTS: 42 patients were investigated, 21 received tDCS and 21 sham stimulation. There were no beneficial effects of tDCS on tinnitus as assessed by primary and secondary outcome measures. Effect size assessed with Cohen's d amounted to 0.08 (95% CI: -0.52 to 0.69) at 1 month and 0.18 (95% CI: -0.43 to 0.78) at 3 months for the THI. CONCLUSION: tDCS of the auditory and prefrontal cortices is safe, but does not improve tinnitus. Different tDCS protocols might be beneficial.