Présentation graphique des caractères technologiques des principaux bois tropicaux. Tome 8 : Bois du Burundi

Caractéristiques physiques et mécaniques des bois sous forme de tableaux comparatifs pour le Burund

In vivo Bioluminescence Imaging of Ca(2+) Signalling in the Brain of Drosophila

Many different cells' signalling pathways are universally regulated by Ca(2+) concentration [Ca(2+)] rises that have highly variable amplitudes and kinetic properties. Optical imaging can provide the means to characterise both the temporal and spatial aspects of Ca(2+) signals involved in neurophysiological functions. New methods for in vivo imaging of Ca(2+) signalling in the brain of Drosophila are required for probing the different dynamic aspects of this system. In studies here, whole brain Ca(2+) imaging was performed on transgenic flies with targeted expression of the bioluminescent Ca(2+) reporter GFP-aequorin (GA) in different neural structures. A photon counting based technique was used to undertake continuous recordings of cytosolic [Ca(2+)] over hours. Time integrals for reconstructing images and analysis of the data were selected offline according to the signal intensity. This approach allowed a unique Ca(2+) response associated with cholinergic transmission to be identified by whole brain imaging of specific neural structures. Notably, [Ca(2+)] transients in the Mushroom Bodies (MBs) following nicotine stimulation were accompanied by a delayed secondary [Ca(2+)] rise (up to 15 min. later) in the MB lobes. The delayed response was sensitive to thapsigargin, suggesting a role for intra-cellular Ca(2+) stores. Moreover, it was reduced in dunce mutant flies, which are impaired in learning and memory. Bioluminescence imaging is therefore useful for studying Ca(2+) signalling pathways and for functional mapping of neurophysiological processes in the fly brain

Morbillivirus Glycoprotein Expression Induces ER Stress, Alters Ca2+ Homeostasis and Results in the Release of Vasostatin

Although the pathology of Morbillivirus in the central nervous system (CNS) is well described, the molecular basis of neurodegenerative events still remains poorly understood. As a model to explore Morbillivirus-mediated CNS dysfunctions, we used canine distemper virus (CDV) that we inoculated into two different cell systems: a monkey cell line (Vero) and rat primary hippocampal neurons. Importantly, the recombinant CDV used in these studies not only efficiently infects both cell types but recapitulates the uncommon, non-cytolytic cell-to-cell spread mediated by virulent CDVs in brain of dogs. Here, we demonstrated that both CDV surface glycoproteins (F and H) markedly accumulated in the endoplasmic reticulum (ER). This accumulation triggered an ER stress, characterized by increased expression of the ER resident chaperon calnexin and the proapoptotic transcription factor CHOP/GADD 153. The expression of calreticulin (CRT), another ER resident chaperon critically involved in the response to misfolded proteins and in Ca2+ homeostasis, was also upregulated. Transient expression of recombinant CDV F and H surface glycoproteins in Vero cells and primary hippocampal neurons further confirmed a correlation between their accumulation in the ER, CRT upregulation, ER stress and disruption of ER Ca2+ homeostasis. Furthermore, CDV infection induced CRT fragmentation with re-localisation of a CRT amino-terminal fragment, also known as vasostatin, on the surface of infected and neighbouring non-infected cells. Altogether, these results suggest that ER stress, CRT fragmentation and re-localization on the cell surface may contribute to cytotoxic effects and ensuing cell dysfunctions triggered by Morbillivirus, a mechanism that might potentially be relevant for other neurotropic viruses

Non-Invasive In Vivo Imaging of Calcium Signaling in Mice

Rapid and transient elevations of Ca2+ within cellular microdomains play a critical role in the regulation of many signal transduction pathways. Described here is a genetic approach for non-invasive detection of localized Ca2+ concentration ([Ca2+]) rises in live animals using bioluminescence imaging (BLI). Transgenic mice conditionally expressing the Ca2+-sensitive bioluminescent reporter GFP-aequorin targeted to the mitochondrial matrix were studied in several experimental paradigms. Rapid [Ca2+] rises inside the mitochondrial matrix could be readily detected during single-twitch muscle contractions. Whole body patterns of [Ca2+] were monitored in freely moving mice and during epileptic seizures. Furthermore, variations in mitochondrial [Ca2+] correlated to behavioral components of the sleep/wake cycle were observed during prolonged whole body recordings of newborn mice. This non-invasive imaging technique opens new avenues for the analysis of Ca2+ signaling whenever whole body information in freely moving animals is desired, in particular during behavioral and developmental studies

Mortality and pulmonary complications in patients undergoing surgery with perioperative SARS-CoV-2 infection: an international cohort study

Background: The impact of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) on postoperative recovery needs to be understood to inform clinical decision making during and after the COVID-19 pandemic. This study reports 30-day mortality and pulmonary complication rates in patients with perioperative SARS-CoV-2 infection. Methods: This international, multicentre, cohort study at 235 hospitals in 24 countries included all patients undergoing surgery who had SARS-CoV-2 infection confirmed within 7 days before or 30 days after surgery. The primary outcome measure was 30-day postoperative mortality and was assessed in all enrolled patients. The main secondary outcome measure was pulmonary complications, defined as pneumonia, acute respiratory distress syndrome, or unexpected postoperative ventilation. Findings: This analysis includes 1128 patients who had surgery between Jan 1 and March 31, 2020, of whom 835 (74·0%) had emergency surgery and 280 (24·8%) had elective surgery. SARS-CoV-2 infection was confirmed preoperatively in 294 (26·1%) patients. 30-day mortality was 23·8% (268 of 1128). Pulmonary complications occurred in 577 (51·2%) of 1128 patients; 30-day mortality in these patients was 38·0% (219 of 577), accounting for 81·7% (219 of 268) of all deaths. In adjusted analyses, 30-day mortality was associated with male sex (odds ratio 1·75 [95% CI 1·28–2·40], p\textless0·0001), age 70 years or older versus younger than 70 years (2·30 [1·65–3·22], p\textless0·0001), American Society of Anesthesiologists grades 3–5 versus grades 1–2 (2·35 [1·57–3·53], p\textless0·0001), malignant versus benign or obstetric diagnosis (1·55 [1·01–2·39], p=0·046), emergency versus elective surgery (1·67 [1·06–2·63], p=0·026), and major versus minor surgery (1·52 [1·01–2·31], p=0·047). Interpretation: Postoperative pulmonary complications occur in half of patients with perioperative SARS-CoV-2 infection and are associated with high mortality. Thresholds for surgery during the COVID-19 pandemic should be higher than during normal practice, particularly in men aged 70 years and older. Consideration should be given for postponing non-urgent procedures and promoting non-operative treatment to delay or avoid the need for surgery. Funding: National Institute for Health Research (NIHR), Association of Coloproctology of Great Britain and Ireland, Bowel and Cancer Research, Bowel Disease Research Foundation, Association of Upper Gastrointestinal Surgeons, British Association of Surgical Oncology, British Gynaecological Cancer Society, European Society of Coloproctology, NIHR Academy, Sarcoma UK, Vascular Society for Great Britain and Ireland, and Yorkshire Cancer Research

The c.429_452 duplication of the ARX gene: a unique developmental-model of limb kinetic apraxia:

BACKGROUND: The c.429_452dup24 of the ARX gene is a rare genetic anomaly, leading to X-Linked Intellectual Disability without brain malformation. While in certain cases c.429_452dup24 has been associated with specific clinical patterns such as Partington syndrome, the consequence of this mutation has been also often classified as "non-specific Intellectual Disability". The present work aims at a more precise description of the clinical features linked to the c.429_452dup24 mutation. METHODS: We clinically reviewed all affected patients identified in France over a five-year period, i.e. 27 patients from 12 different families. Detailed cognitive, behavioural, and motor evaluation, as well as standardized videotaped assessments of oro-lingual and gestural praxis, were performed. In a sub-group of 13 ARX patients, kinematic and MRI studies were further accomplished to better characterize the motor impairment prevalent in the ARX patients group. To ensure that data were specific to the ARX gene mutation and did not result from low-cognitive functioning per se, a group of 27 age- and IQ-matched Down syndrome patients served as control. RESULTS: Neuropsychological and motor assessment indicated that the c.429_452dup24 mutation constitutes a recognizable clinical syndrome: ARX patients exhibiting Intellectual Disability, without primary motor impairment, but with a very specific upper limb distal motor apraxia associated with a pathognomonic hand-grip. Patients affected with the so-called Partington syndrome, which involves major hand dystonia and orolingual apraxia, exhibit the most severe symptoms of the disorder. The particular "reach and grip" impairment which was observed in all ARX patients, but not in Down syndrome patients, was further characterized by the kinematic data: (i) loss of preference for the index finger when gripping an object, (ii) major impairment of fourth finger deftness, and (iii) a lack of pronation movements. This lack of distal movement coordination exhibited by ARX patients is associated with the loss of independent digital dexterity and is similar to the distortion of individual finger movements and posture observed in Limb Kinetic Apraxia. CONCLUSION: These findings suggest that the ARX c.429_452dup24 mutation may be a developmental model for Limb Kinetic Apraxia

Création et caractérisation d'outils génétiques destinés à visualiser les réseaux neuronaux chez la souris

Le fonctionnement du système nerveux central (SNC) des mammifères met en jeu la communication entre les neurones reliés entre eux par des connexions synaptiques et formant des réseaux neuronaux. Afin d étudier la formation, le maintien et le fonctionnement des réseaux neuronaux, nous avons développé et amélioré de nouveaux outils génétiques codant pour deux types de protéines, fluorescentes ou bioluminescentes : La protéine hybride fluorescente GFP-TTC qui conserve les propriétés de transport rétrograde et trans-synaptique de la toxine tétanique tout en étant dépourvue de toxicité. Cette protéine est ainsi un marqueur anatomique des réseaux de neurones actifs connectés entre eux. Différentes constructions moléculaires gfp-ttc contenant une séquence IRES et des sites loxP, ont été fabriquées dans le but de construire des souris rapporteurs universels pouvant exprimer ces constructions de manière ubiquitaire dans tous les neurones du SNC. L idée, à plus long terme, mais qui ne fait pas partie de ce travail, est de croiser ces souris rapporteurs avec des souris exprimant la recombinase Cre dans des sous-populations de neurones très spécifiques afin d étudier les réseaux neuronaux associés à ces populations neuronales. Deux souris transgéniques, chacune contenant une version différente du transgène gfp-ttc, ont été obtenues à l issue de ce travail de thèse. Le premier animal, obtenu par transgenèse classique, ne présentait malheureusement pas une expression ubiquitaire du transgène et ne pourra donc pas être utilisé comme souris rapporteur universel . Néanmoins, le transgène est exprimé dans des régions spécifiques du SNC et notamment dans la rétine au cours de l embryogenèse jusque chez l adulte. Par ailleurs, son expression est sélective des cellules bipolaires et ganglionnaires de la rétine, faisant de la protéine fluorescente GFP-TTC un marqueur fluorescent spécifique de ces types cellulaires. En conséquence, cette lignée de souris pourrait représenter un outil de choix pour suivre anatomiquement le développement, la migration et l architecture des ces neurones rétiniens au cours des stades embryonnaires et adultes. Suite à cet échec, nous avons généré un second animal transgénique par recombinaison homologue au locus HPRT afin d optimiser l expression ubiquitaire du transgène. Cet animal, obtenu très récemment, est en cours de caractérisation et nous espérons vivement qu il exprimera gfp-ttc de manière ubiquitaire. Ce travail de transgenèse a été parallèlement complété par une étude de caractérisation des propriétés moléculaires et cellulaires des facteurs agissant sur l internalisation neuronale de la sonde GFP-TTC à la synapse neuromusculaire. L idée était d identifier la voie de transport intracellulaire utilisée par la protéine GFP-TTC dans le but de pouvoir ensuite améliorer le transport intracellulaire et trans-synaptique de cette sonde. Ainsi, des expériences de coinjection de la protéine GFP-TTC et des neurotrophines BDNF ou NT-4 dans le muscle, nous ont permis de mettre en évidence une augmentation de l endocytose de la protéine GFP-TTC en présence de ces neurotrophines. Ces résultats suggèrent la présence d un mécanisme d internalisation similaire entre ces neurotrophines et la sonde GFP-TTC. La protéine hybride bioluminescente GFP-aequorine (GA) qui est sensible au Ca2+. Sa liaison au Ca2+ entraîne l émission de lumière verte ( max=509nm) et permet de suivre les variations de concentrations calciques associées à l activité cellulaire. Afin d étudier l activité de réseaux neuronaux, provenant des régions profondes du cerveau de la souris adulte, nous avons dû construire de nouvelles sondes bioluminescentes en se basant sur le transfert d énergie chimioluminescente, retrouvé dans GA. Deux nouvelles sondes génétiquement codées ont ainsi été obtenues : Venus-aequorine (VA) et mRFP1-aequorine (RA) dont les signaux luminescents ont été déplacés vers les longueurs d ondes rouges par rapport à GA. Nous avons observé que l intensité bioluminescente de la sonde VA était plus importante que celle de RA. Toutefois, seul le signal émis par RA (dans un rouge plus lointain que le signal émis par VA) et provenant de régions profondes du cerveau de la souris adulte, s est avéré être détectable. Cette amélioration, par déplacement de la bioluminescence vers des longueurs d ondes lointaines, représente une avancée technique dans le développement des nouvelles technologies d imageries du petit animal. Ces deux nouvelles sondes sont en cours d être brevetées par l Institut Pasteur. En conclusion, l ensemble de ce travail de thèse a essentiellement consisté en (1) la création, l amélioration et la caractérisation de nouvelles sondes génétiques (destinées à l analyse de l activité des réseaux neuronaux) et (2) la génération de souris transgéniques exprimant certaines de ces sondes. Le développement de tels outils biotechnologiques se place en amont d études à plus long terme du développement et de l activité des réseaux neuronaux chez l animal vivant.PARIS-BIUSJ-Thèses (751052125) / SudocPARIS-BIUSJ-Physique recherche (751052113) / SudocSudocFranceF