Universite de Strasbourg

19 articles published in JoVE

• Behavior

  
  Animal Models of Depression - Chronic Despair Model (CDM) Stefan Vestring^1,2, Tsvetan Serchov^3,4, Claus Normann^{1,5 1}Department of Psychiatry and Psychotherapy, Medical Center - University of Freiburg, Faculty of Medicine, University of Freiburg, ²Berta-Ottenstein-Programme for Clinician Scientists, Faculty of Medicine, University of Freiburg, ³Centre National de la Recherche Scientifique (CNRS), Université de Strasbourg, Institut des Neurosciences Cellulaires et Intégratives, ⁴Department of Stereotactic and Functional Neurosurgery, Medical Center - University Freiburg, Faculty of Medicine, University of Freiburg, ⁵Center for Basics in Neuromodulation, Faculty of Medicine, University of Freiburg The chronic despair mouse model (CDM) of depression consists of repetitive forced swim sessions and another delayed swim phase as a read-out. It represents a suitable model for induction of a chronic depressive-like state stable for at least 4 weeks, amendable to evaluate subchronic and acute treatment interventions.

• Biology

  
  In Situ Exploration of Murine Megakaryopoiesis using Transmission Electron Microscopy Cyril Scandola¹, François Lanza¹, Christian Gachet¹, Anita Eckly^{1 1}Université de Strasbourg, INSERM, EFS Grand Est, BPPS UMR-S 1255, FMTS Here, we present a protocol to analyze ultrastructure of the megakaryocytes in situ using transmission electron microscopy (TEM). Murine bone marrows are collected, fixed, embedded in epoxy resin and cut in ultrathin sections. After contrast staining, the bone marrow is observed under a TEM microscope at 120 kV.

• Biology

  
  Megakaryocyte Culture in 3D Methylcellulose-Based Hydrogel to Improve Cell Maturation and Study the Impact of Stiffness and Confinement Julie Boscher¹, Christian Gachet¹, François Lanza¹, Catherine Léon^{1 1}Université de Strasbourg, INSERM, EFS Grand Est, BPPS UMR-S 1255, FMTS It is now acknowledged that the three-dimensional environment of cells can play an important role in their behavior, maturation and/or differentiation. This protocol describes a three-dimensional cell culture model designed to study the impact of physical containment and mechanical constraints on megakaryocytes.

• Biology

  
  In Vivo Two-photon Imaging of Megakaryocytes and Proplatelets in the Mouse Skull Bone Marrow Alicia Bornert¹, Fabien Pertuy¹, François Lanza¹, Christian Gachet¹, Catherine Léon^{1 1}INSERM, EFS Grand Est, Université de Strasbourg We describe here the method for imaging megakaryocytes and proplatelets in the marrow of the skull bone of living mice using two-photon microscopy.

• Biology

  
  Proplatelet Formation Dynamics of Mouse Fresh Bone Marrow Explants Inès Guinard¹, François Lanza¹, Christian Gachet¹, Catherine Léon¹, Anita Eckly^{1 1}Université de Strasbourg, INSERM, EFS Grand Est, BPPS UMR-S 1255, FMTS Here, we detail the bone marrow explant method, from sample preparation to microscopic slide analysis, to evaluate the ability of megakaryocytes which have differentiated in their physiological environment to form proplatelets.

• Biology

  
  Leukodepletion Filters-Derived CD34+ Cells As a Cell Source to Study Megakaryocyte Differentiation and Platelet Formation Anaïs Pongerard¹, Lea Mallo¹, Christian Gachet¹, Henri de La Salle¹, François Lanza¹, Catherine Strassel^{1 1}BPPS UMR-S 1255, FMTS, Université de Strasbourg, INSERM, EFS Grand Est This protocol describes in detail all the steps involved in obtaining leukofilter-derived CD34+ hematopoietic progenitors and their in vitro differentiation and maturation into proplatelet-bearing megakaryocytes that are able to release platelets in the culture medium. This procedure is useful for in-depth analysis of cellular and molecular mechanisms controlling megakaryopoiesis.

• Biology

  
  Isolation of Mouse Megakaryocyte Progenitors Quentin Kimmerlin¹, Manuela Tavian², Christian Gachet¹, François Lanza¹, Nathalie Brouard^{1 1}Université de Strasbourg, INSERM UMR S1255, EFS Grand-EST, ²UMR-S1113 –IRFAC, Université de Strasbourg This method describes the purification by flow cytometry of MEP and MKp from mice femurs, tibias, and pelvic bones.

• Cancer Research

  
  A Three-dimensional Model of Spheroids to Study Colon Cancer Stem Cells Maria Virginia Giolito^1,2, Léo Claret^1,2, Carla Frau¹, Michelina Plateroti^{1,2 1}Département de la recherche, Centre de Recherche en Cancérologie de Lyon, INSERM U1052, CNRS UMR5286, Université de Lyon, Université Lyon 1, Centre Léon Bérard, ²UMR-S1113 - IRFAC INSERM, Université de Strasbourg This protocol presents a novel, robust, and reproducible culture system to generate and grow three-dimensional spheroids from Caco2 colon adenocarcinoma cells. The results provide the first proof-of-concept for the appropriateness of this approach to study cancer stem cell biology, including the response to chemotherapy.

• Biology

  
  FLIM-FRET Measurements of Protein-Protein Interactions in Live Bacteria. Hanna Manko¹, Vincent Normant^2,3, Quentin Perraud^2,3, Tania Steffan¹, Véronique Gasser^2,3, Emmanuel Boutant¹, Éléonore Réal¹, Isabelle J. Schalk^2,3, Yves Mély¹, Julien Godet^{1,4 1}Université de Strasbourg, Laboratoire de Bioimagerie et Pathologies, UMR CNRS 7021, ²Université de Strasbourg, UMR 7242, ESBS, ³CNRS, UMR 7242, ESBS, ⁴Groupe Méthode Recherche Clinique, Hôpitaux Universitaires de Strasbourg We describe here a protocol to characterize protein-protein interactions between two highly-differently expressed proteins in live Pseudomonas aeruginosa using FLIM-FRET measurements. The protocol includes bacteria strain constructions, bacteria immobilization, imaging and post-imaging data analysis routines.

• Environment

  
  Investigation of Xenobiotics Metabolism In Salix alba Leaves via Mass Spectrometry Imaging Claire Villette¹, Loïc Maurer^1,2, Dimitri Heintz^{1 1}Plant Imaging and Mass Spectrometry (PIMS), Institut de biologie moléculaire des plantes, CNRS, Université de Strasbourg, ² This method uses mass spectrometry imaging (MSI) to understand metabolic processes in S. alba leaves when exposed to xenobiotics. The method allows the spatial localization of compounds of interest and their predicted metabolites within specific, intact tissues.

• Bioengineering

  
  Intra-Omental Islet Transplantation Using h-Omental Matrix Islet filliNG (hOMING) Anaïs Schaschkow¹, Carole Mura¹, Michel Pinget¹, Karim Bouzakri¹, Elisa Maillard^{1 1}Centre Européen d'Etude du Diabète, Université de Strasbourg Here, we present a protocol for in vivo validation of hydrogel-based cell therapy, illustrated by the example of islet transplantation. h-Omental Matrix Islet filliNG (hOMING) implantation allows implantation of a cell-hydrogel mixture between the omental layers, near to blood vessels, to maximize engraftment in a proper metabolic environment.

• Chemistry

  
  Photogeneration of N-Heterocyclic Carbenes: Application in Photoinduced Ring-Opening Metathesis Polymerization Julien Pinaud¹, Emeline Placet¹, Patrick Lacroix-Desmazes¹, Thi Kim Hoang Trinh^2,3, Jean Pierre Malval^2,3, Abraham Chemtob^2,3, Loïc Pichavant⁴, Valérie Héroguez^{4 1}ICGM, Université de Montpellier, CNRS, ENSCM, ²Institut de Science des Matériaux de Mulhouse (IS2M UMR 7361 CNRS), Université de Haute-Alsace, ³Université de Strasbourg, ⁴Laboratoire de Chimie des Polymères Organiques (LCPO UMR 5629 ENSCBP), Université de Bordeaux We describe a protocol to photogenerate N-heterocyclic carbenes (NHCs) by UV irradiation of a 2-isopropylthioxanthone/imidazolium tetraphenylborate salt system. Methods to characterize the photoreleased NHC and elucidate the photochemical mechanism are proposed. The protocols for ring-opening metathesis photopolymerization in solution and miniemulsion illustrate the potential of this 2-component NHC photogenerating system.

• Genetics

  
  Saccharomyces cerevisiae Metabolic Labeling with 4-thiouracil and the Quantification of Newly Synthesized mRNA As a Proxy for RNA Polymerase II Activity Tiago Baptista^1,2,3,4, Didier Devys^{1,2,3,4 1}Institut de Génétique et de Biologie Moléculaire et Cellulaire, Illkirch, France, ²Centre National de la Recherche Scientifique, Illkirch, France, ³Institut National de la Santé et de la Recherche Médicale, Illkirch, France, ⁴Université de Strasbourg The protocol described here is based on the genome-wide quantification of newly synthesized mRNA purified from yeast cells labeled with 4-thiouracil. This method allows to measure mRNA synthesis uncoupled from mRNA decay and, thus, provides an accurate measurement of RNA polymerase II transcription.

• Chemistry

  
  Microfluidic Preparation of Liquid Crystalline Elastomer Actuators Tristan Hessberger*¹, Lukas B. Braun*¹, Christophe A. Serra², Rudolf Zentel^{1 1}Department of Organic Chemistry, Johannes Gutenberg University, ²CNRS, ICS UPR 22, Université de Strasbourg This article describes the microfluidic process and parameters to prepare actuating particles from liquid crystalline elastomers. This process allows the preparation of actuating particles and the variation of their size and shape (from oblate to strongly prolate, core-shell, and Janus morphologies) as well as the magnitude of actuation.

• Neuroscience

  
  Assessment of Morphine-induced Hyperalgesia and Analgesic Tolerance in Mice Using Thermal and Mechanical Nociceptive Modalities Khadija Elhabazi¹, Safia Ayachi¹, Brigitte Ilien¹, Frédéric Simonin^{1 1}Biotechnology and Cellular Signalling, UMR 7242 CNRS, Université de Strasbourg We describe a protocol to examine the development of opioid-induced hyperalgesia and tolerance in mice. Based on the measurement of thermal and mechanical nociceptive responses of naïve and morphine-treated animals, it allows to quantify the increase in pain sensitivity (hyperalgesia) and decrease in analgesia (tolerance) associated with chronic opiate administration.

• Medicine

  
  The Sciatic Nerve Cuffing Model of Neuropathic Pain in Mice Ipek Yalcin¹, Salim Megat^1,2, Florent Barthas^1,2, Elisabeth Waltisperger¹, Mélanie Kremer^1,2, Eric Salvat^1,2,3, Michel Barrot^{1 1}Institut des Neurosciences Cellulaires et Intégratives UPR3212, Centre National de la Recherche Scientifique, ²Université de Strasbourg, ³Hôpitaux Universitaires de Strasbourg Neuropathic pain is a consequence of a lesion or disease affecting the somatosensory system. The “cuff model” of neuropathic pain in mice consists of the implantation of a polyethylene cuff around the main branch of the sciatic nerve. Mechanical allodynia is tested using von Frey filaments.

• Medicine

  
  In vivo Macrophage Imaging Using MR Targeted Contrast Agent for Longitudinal Evaluation of Septic Arthritis Guillaume Bierry^1,2, Sophie Lefevre^2,3, Jean-Louis Dietemann¹, François Jehl^{2,3 1}Department of Radiology, University Hospital of Strasbourg, ²EA 3432, University of Strasbourg, ³Department of Bacteriology, University Hospital of Strasbourg We demonstrate how to perform macrophage MR imaging using ultrasmall superparamagnetic contrast agent (USPIO) in septic arthritis, allowing an initial and longitudinal in vivo non-invasive evaluation of macrophages infiltration and an assessment of therapy efficacy.

• Immunology and Infection

  
  Use of In vivo Imaging to Monitor the Progression of Experimental Mouse Cytomegalovirus Infection in Neonates Eleonore Ostermann¹, Cécile Macquin¹, Seiamak Bahram¹, Philippe Georgel^{1 1}ImmunoRhumatologie Moléculaire, INSERM UMR_S 1109, Centre de Recherche d'Immunologie et d'Hématologie, Université de Strasbourg Human Cytomegalovirus (HCMV) infection of neonates represents an important cause of mental retardation, yet the molecular events leading to virus-induced pathogenesis are still poorly understood. To investigate the dynamics of brain infection, we adapted whole-animal in vivo imaging to perform time-course analysis of neonates infected with a luciferase-recombinant virus.

• Bioengineering

  
  Multi-Scale Modification of Metallic Implants With Pore Gradients, Polyelectrolytes and Their Indirect Monitoring In vivo Nihal E. Vrana¹, Agnes Dupret-Bories^1,2, Christophe Chaubaroux¹, Elisabeth Rieger^1,2, Christian Debry^1,2, Dominique Vautier^1,3, Marie-Helene Metz-Boutigue^1,3, Philippe Lavalle^{1,3 1}Biomatériaux et Bioingénieriee, INSERM, ²Service Oto-Rhino-Laryngologie, Hôpitaux Universitaires de Strasbourg, ³Faculté de Chirurgie Dentaire, Université de Strasbourg In this video, we will demonstrate modification techniques for porous metallic implants to improve their functionality and to control cell migration. Techniques include development of pore gradients to control cell movement in 3D and production of basement membrane mimics to control cell movement in 2-D. Also, a HPLC-based method for monitoring implant integration in-vivo via analysis of blood proteins is described.