Université de paris

8 ARTICLES PUBLISHED IN JoVE

Medicine

Standardized Measurement of Nasal Membrane Transepithelial Potential Difference (NPD)

George M. Solomon¹, Inez Bronsveld², Kathryn Hayes³, Michael Wilschanski⁴, Paola Melotti⁵, Steven M. Rowe¹, Isabelle Sermet-Gaudelus^6,7

¹Department of Medicine and the Gregory Fleming James Cystic Fibrosis Center, University of Alabama at Birmingham, ²Department of Pulmonology and Tuberculosis, University Medical Center Utrecht, ³Center for Experimental Medicine, Queens University, Northern Ireland, ⁴Hadassah Hebrew University Medical Center, Jerusalem, ⁵Centro Fibrosi Cistica, Azienda Ospedaliera Universitaria Integrata, ⁶Service de Pneumologie et Allergologie Pédiatriques and Center de Ressources et de Compétence de la Mucoviscidose, Hôpital Necker Enfants Malades, ⁷INSERM U 1151, Institut Necker Enfants Malades

Here, we present a standardized protocol to measure the nasal potential difference (NPD). Cystic fibrosis transmembrane conductance regulator (CFTR) and epithelial sodium channel (ENaC) function are evaluated by the change in the voltage across the nasal epithelium after superfusion of solutions that modify ion channel activity, providing an outcome measure.

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.

Bioengineering

A Reliable Porcine Fascio-Cutaneous Flap Model for Vascularized Composite Allografts Bioengineering Studies

Victor Pozzo^1,2,4, Golda Romano^1,2,4, Marion Goutard^1,2,4, Elise Lupon^1,2,4, Pierre Tawa^1,2,4, Aylin Acun^3,4,5, Alec R. Andrews², Corentin B. Taveau^1,2,4,6, Basak E. Uygun^1,2,3,4, Mark A. Randolph^1,2,4, Curtis L. Cetrulo^1,2,4, Alexandre G. Lellouch^1,2,4,6

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, ³Center for Engineering in Medicine and Surgery, Massachusetts General Hospital, Harvard Medical School, ⁴Shriners Hospital for Children, ⁵Department of Biomedical Engineering, Widener University, ⁶Service de Chirurgie Plastique, Hôpital Européen Georges Pompidou, Assistance Publique-Hôpitaux de Paris (APHP), Université Paris Descartes

The present protocol describes the porcine fascio-cutaneous flap model and its potential use in vascularized composite tissue research.