Université Paris Cité

9 ARTICLES PUBLISHED IN JoVE

Immunology and Infection

Monitoring Influenza Virus Survival Outside the Host Using Real-Time Cell Analysis

Thomas Labadie¹, Quentin Grassin², Christophe Batéjat², Jean-Claude Manuguerra², India Leclercq^2,3

¹Department of Infection Biology, London School of Hygiene and Tropical Medicine, ²Institut Pasteur, Unité Environnement et Risques Infectieux, Cellule d'Intervention Biologique d'Urgence (CIBU), ³Cellule Pasteur, Université de Paris

Reported here is a protocol for the quantification of infectious viral particles using real-time monitoring of electrical impedance of infected cells. A practical application of this method is presented by quantifying influenza A virus decay under different physicochemical parameters mimicking environmental conditions.

Neuroscience

Whole-Brain 3D Activation and Functional Connectivity Mapping in Mice using Transcranial Functional Ultrasound Imaging

Adrien Bertolo^*1, Mohamed Nouhoum^*1, Silvia Cazzanelli^*1, Jeremy Ferrier^*3, Jean-Charles Mariani², Andrea Kliewer^4,2, Benoit Belliard¹, Bruno-Félix Osmanski³, Thomas Deffieux^*1, Sophie Pezet^*1, Zsolt Lenkei^*2, Mickael Tanter^*1

¹Physics for Medicine Paris, ESPCI Paris, INSERM, CNRS, PSL Research University, ²Institute of Psychiatry and Neurosciences of Paris, INSERM U1266, Université de Paris, ³Iconeus, ⁴Department of Pharmacology and Toxicology, Jena University Hospital - Friedrich Schiller University Jena

This protocol describes the quantification of volumetric cerebral hemodynamic variations in the mouse brain using functional ultrasound (fUS). Procedures for 3D functional activation map following sensory stimulation as well as resting-state functional connectivity are provided as illustrative examples, in anesthetized and awake mice.

Immunology and Infection

Partial Heterotopic Hindlimb Transplantation Model in Rats

Marion Goutard^1,2,3, Mark A. Randolph^1,2,3, Corentin B. Taveau^1,2,3,4, Elise Lupon^1,2,3, Laurent Lantieri⁴, Korkut Uygun^1,2,3, Curtis L. Cetrulo Jr.^*1,2,3, Alexandre G. Lellouch^*1,2,3,4

¹Division of Plastic and Reconstructive Surgery, Massachusetts General Hospital, Harvard Medical School, ²Vascularized Composite Allotransplantation Laboratory, Center for Transplantation Sciences, Massachusetts General Hospital, Harvard Medical School, ³Shriners Hospital for Children, ⁴Service de Chirurgie Plastique, Hôpital Européen Georges Pompidou, Assistance Publique-Hôpitaux de Paris (APHP), Université de Paris

This paper presents a partial heterotopic osteomyocutaneous flap transplantation protocol in rats and its potential outcomes in the mid-term follow-up.

Developmental Biology

2D and 3D Human Induced Pluripotent Stem Cell-Based Models to Dissect Primary Cilium Involvement during Neocortical Development

Lucile Boutaud^*1, Marie Michael^*1, Céline Banal², Damelys Calderon¹, Sarah Farcy¹, Julie Pernelle¹, Nicolas Goudin³, Camille Maillard¹, Clémantine Dimartino¹, Cécile Deleschaux¹, Sébastien Dupichaud⁴, Corinne Lebreton¹, Sophie Saunier¹, Tania Attié-Bitach^1,5, Nadia Bahi-Buisson^1,6, Nathalie Lefort², Sophie Thomas¹

¹Imagine Institute, INSERM UMR 1163, Université de Paris, ²Imagine Institute, iPSC Core Facility, INSERM UMR U1163, Université de Paris, ³Necker Bio-image Analysis platform of the SFR Necker, ⁴Imagine Institute, Cell Imaging Platform, INSERM-US24-CNRS UMS 3633 Structure Fédérative de Recherche Necker, INSERM UMR U1163, Université de Paris, ⁵Fédération de Génétique, Hôpital Necker-Enfants Malades, Assistance Publique Hôpitaux de Paris, ⁶Pediatric Neurology, APHP- Necker Enfants Malades Hospital

We present detailed protocols for the generation and characterization of 2D and 3D human induced pluripotent stem cell (hIPSC)-based models of neocortical development as well as complementary methodologies enabling qualitative and quantitative analysis of primary cilium (PC) biogenesis and function.

Neuroscience

Magnetic Isolation of Microglial Cells from Neonate Mouse for Primary Cell Cultures

Cindy Bokobza^*1, Alice Jacquens^*1,2, Manuela Zinni¹, Valérie Faivre¹, Jennifer Hua¹, David Guenoun¹, Caroline Userovici¹, Shyamala Mani¹, Vincent Degos^1,2, Pierre Gressens¹, Juliette Van Steenwinckel¹

¹NeuroDiderot, Inserm UMR-1141, Hôpital Robert Debré 48, Université de Paris, ²Department of anesthesia and critical care, APHP-Sorbonne university

Primary microglia cultures are commonly used to evaluate new anti-inflammatory molecules. The present protocol describes a reproducible and relevant method to magnetically isolate microglia from neonate pups.

Biology

Primary Human Nasal Epithelial Cells: Biobanking in the Context of Precision Medicine

Mairead Kelly^1,2, Elise Dreano^1,2, Aurelie Hatton^1,2, Agathe Lepissier^1,2, Anita Golec^1,2, Isabelle Sermet-Gaudelus^1,2,3, Iwona Pranke^1,2,3

¹Institut Necker Enfants Malades, ²Université de Paris, ³Centre de Référence Maladies Rares Mucoviscidose et Maladies apparentées, Assistance Publique Hôpitaux de Paris

Here we describe the isolation, amplification, and differentiation of primary human nasal epithelial (HNE) cells at the air-liquid interface and a biobanking protocol allowing to successfully freeze and then thaw amplified HNE. The protocol analyzes electrophysiological properties of differentiated HNE cells and CFTR-related chloride secretion correction upon different modulator treatments.

Biology

Bottom-Up In Vitro Methods to Assay the Ultrastructural Organization, Membrane Reshaping, and Curvature Sensitivity Behavior of Septins

Brieuc Chauvin^*1, Koyomi Nakazawa^*1, Alexandre Beber^1,7, Aurélie Di Cicco¹, Bassam Hajj¹, François Iv², Manos Mavrakis², Gijsje H. Koenderink³, João T. Cabral⁴, Michaël Trichet⁵, Stéphanie Mangenot^*6, Aurélie Bertin^*1

¹Laboratoire Physico Chimie Curie, Institut Curie, PSL Research University, Sorbonne Université, ²Institut Fresnel, CNRS UMR7249, Aix Marseille Univ, Centrale Marseille, ³Department of Bionanoscience, Kavli Institute of Nanoscience Delft, Delft University of Technology, ⁴Department of Chemical Engineering, Imperial College London, ⁵Sorbonne Université, CNRS, Institut de Biologie Paris-Seine (IBPS), Service de microscopie électronique (IBPS-SME), ⁶Laboratoire Matière et Systèmes Complexes (MSC), Université Paris Cité, ⁷Institute of Biotechnology, Czech Academy of Sciences, BIOCEV

Septins are cytoskeletal proteins. They interact with lipid membranes and can sense but also generate membrane curvature at the micron scale. We describe in this protocol bottom-up in vitro methodologies for analyzing membrane deformations, curvature-sensitive septin binding, and septin filament ultrastructure.

Medicine

Robot-Assisted Transcanal Endoscopic Ear Surgery for Congenital Cholesteatoma

François Simon^1,2, Yann Nguyen^3,4, Natalie Loundon², Françoise Denoyelle^1,2

¹Université Paris Cité, ²Pediatric Otolaryngology, Head and neck surgery department, AP-HP, Hôpital Necker-Enfants Malades, ³Sorbonne Université, ⁴Otolaryngology - head and neck surgery department AP-HP, Hôpital Pitié-Salpêtrière

This protocol describes the removal of congenital cholesteatoma using minimally invasive transcanal endoscopic ear surgery with a two-handed approach and a robotic endoscope holder.

Medicine

Assessing Intracardiac Vortices with High Frame-Rate Echocardiography-Derived Blood Speckle Imaging in Newborns

Edward Crendal¹, Koert De Waal², Damien Vitiello³

¹John Hunter Hospital, Department of Cardiology, University of Newcastle, ²John Hunter Children’s Hospital, Department of Neonatology, University of Newcastle, ³Institute of Sport and Health Sciences of Paris (IS3P - URP 3625), Université Paris Cité

The present protocol uses echocardiography-derived blood speckle imaging technology to visualize intracardiac hemodynamics in newborns. The clinical utility of this technology is explored, the rotational body of fluid within the left ventricle (known as a vortex) is accessed, and its significance in understanding diastology is determined.