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Aix-Marseille Université

11 ARTICLES PUBLISHED IN JoVE

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Medicine

Implanting Glass Spinal Cord Windows in Adult Mice with Experimental Autoimmune Encephalomyelitis
Keith K. Fenrich 1, Pascal Weber 1, Genevieve Rougon 1,2, Franck Debarbieux 1,2
1Developmental Biology Institute of Marseille-Luminy (IBDML), Aix Marseille University, 2Campus de la Timone, European Research Center for Medical Imaging (CERIMED)

We describe a method for implanting and maintaining glass windows over the exposed spinal cords of adult mice for studying experimental autoimmune encephalomyelitis. These windows provide chronic optical access to the spinal cord for monitoring cell dynamics at the subcellular level in live animals using two-photon microscopy.

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Immunology and Infection

Bone Marrow-derived Macrophage Production
Virginie Trouplin 1, Nicolas Boucherit 1, Laurent Gorvel 1, Filippo Conti 1, Giovanna Mottola 1,2, Eric Ghigo 1
1CNRS UMR 7278, IRD198, INSERM U1095, Aix-Marseille Université, 2Department of Molecular Medicine and Medical Biotechnology, University of Naples "Federico II"

Macrophages have long been recognized as a critical component of the innate and adaptive immune responses. The recent explosion of knowledge pertaining to evolutionary, genetic, and biochemical aspects of the interaction between macrophages and microbes has renewed scientific attention to macrophages. This article describes a method to differentiate macrophages from mouse bone marrow.

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Medicine

An Orthotopic Glioblastoma Mouse Model Maintaining Brain Parenchymal Physical Constraints and Suitable for Intravital Two-photon Microscopy
Clément Ricard 1,2, Fabio Stanchi 1,3, Geneviève Rougon 1,2, Franck Debarbieux 1,2
1Developmental Biology Institute of Marseille, Aix Marseille University, 2European Research Center for Medical Imaging, Campus de la Timone, 3Vesalius Research Center, KU Leuven Campus Gasthuisberg

We have established a cortical orthotopic glioblastoma model in mice for intravital two-photon microscopy that recapitulates the biophysical constraints normally at play during the growth of the tumor. A chronic glass window replacing the skull above the tumor enables the follow-up of the tumor progression over time by two-photon microscopy.

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Biology

Setting Up a Simple Light Sheet Microscope for In Toto Imaging of C. elegans Development
Claire Chardès *1, Pauline Mélénec *1, Vincent Bertrand 1, Pierre-François Lenne 1
1Institut de Biologie du Développement de Marseille, UMR7288 CNRS, Aix-Marseille Université

This protocol describes the setup of a light sheet microscope and its implementation for in vivo imaging of C. elegans embryos.

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Biology

High Throughput Quantitative Expression Screening and Purification Applied to Recombinant Disulfide-rich Venom Proteins Produced in E. coli
Natalie J. Saez 1, Hervé Nozach 2, Marilyne Blemont 1, Renaud Vincentelli 1
1Architecture et Fonction des Macromolécules Biologiques (AFMB), Aix-Marseille Université, 2iBiTec-S, Service d'Ingénierie Moléculaire des Protéines (SIMOPRO), Commissariat à l'énergie atomique et aux énergies alternatives (CEA) Saclay, France

A protocol for the quantitative, high throughput expression screening and analytical purification of fusion proteins from small-scale Escherichia coli cultures is described and applied to the expression of disulfide-rich animal venom protein targets.

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Neuroscience

Metabolomic Analysis of Rat Brain by High Resolution Nuclear Magnetic Resonance Spectroscopy of Tissue Extracts
Norbert W. Lutz 1, Evelyne Béraud 2, Patrick J. Cozzone 1
1Centre de Résonance Magnétique Biologique et Médicale, UMR 7339 CNRS, Faculté de Médecine, Aix-Marseille Université, 2Centre de Recherches en Oncologie Biologique et Oncopharmacologie, UMR 911 INSERM, Faculté de Médecine, Aix-Marseille Université

The neurochemistry of mammalian brain is changed in many neurological and systemic diseases. Characteristic profiles of cerebral metabolites can be efficiently obtained based on crude extracts of brain tissue. To this end, high-resolution NMR spectroscopy is employed, enabling detailed quantitative analysis of metabolite concentrations (metabolomics).

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Biology

An Easy Method for Plant Polysome Profiling
Cécile Lecampion 1,2,3, Maïna Floris 1,2,3,4, Jean Raphaël Fantino 5,6, Christophe Robaglia 1,2,3, Christophe Laloi 1,2,3
1Laboratoire de Génétique et Biophysique des Plantes, Aix-Marseille Université, 2UMR 7265 Biologie Végétale & Microbiologie Environnementales, CNRS, 3BIAM, CEA, 4Department of Biology, Biocenter, University of Copenhagen, 5Laboratoire de Chimie Bactérienne, 6CNRS, LCB UMR 7283, Aix Marseille Université

This protocol describes an easy method to extract and fractionate transcripts from plant tissues on the basis of the number of bound ribosomes. It allows a global estimate of translation activity and the determination of the translational status of specific mRNAs.

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Genetics

RNA Pull-down Procedure to Identify RNA Targets of a Long Non-coding RNA
Manon Torres 1, Denis Becquet 1, Séverine Guillen 1, Bénédicte Boyer 1, Mathias Moreno 1, Marie-Pierre Blanchard 2, Jean-Louis Franc 1, Anne-Marie François-Bellan 1
1CNRS, CRN2M-UMR7286, Faculté de Médecine Nord, Aix-Marseille Université, 2Plate-forme d'imagerie MRI, UMS3426, CNRS 141, Institut de Génétique Humaine

This RNA pull-down method allows identifying the RNA targets of a long non-coding RNA (lncRNA). Based on the hybridization of home-made, designed anti-sense DNA oligonucleotide probes specific to this lncRNA in an appropriately fixed tissue or cell line, it efficiently allows the capture of all RNA targets of the lncRNA.

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Developmental Biology

Probing Cell Mechanics with Bead-Free Optical Tweezers in the Drosophila Embryo
Claire Chardès *1, Raphael Clement *1, Olivier Blanc 1, Pierre-François Lenne 1
1CNRS, Institut de Biologie du Développement de Marseille, Aix-Marseille Université

We present a setup of optical tweezers coupled to a light sheet microscope, and its implementation to probe cell mechanics without beads in the Drosophila embryo.

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Genetics

Identification of RNAs Engaged in Direct RNA-RNA Interaction with a Long Non-Coding RNA
Audrey Jacq 1, Denis Becquet 1, Séverine Guillen 1, Bénédicte Boyer 1, Maria-Montserrat Bello-Goutierrez 1, Jean-Louis Franc *1, Anne-Marie François-Bellan *1
1Faculté des Sciences Médicales et Paramédicales, Aix Marseille Univ, CNRS, INP

An easy-to-use RNA pull-down protocol is designed for the identification of RNAs engaged in direct RNA/RNA interaction with a long non-coding RNA. The protocol uses psoralen as a fixative to cross-link only RNA/RNA interactions and provides the whole direct RNA interactome of a long non-coding RNA when coupled with RNA sequencing.

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Cancer Research

Flexible Organic Electronic Devices for Pulsed Electric Field Therapy of Glioblastoma
Marie C. Lefevre 1, Attila Kaszas 1, Gerwin Dijk 1,2, Martin Baca 1, Olivier Baudino 2, Bastien Marchiori 2, Loig Kergoat 2, David Moreau 1, Franck Debarbieux 3,4,5, Rodney P. O'Connor 1
1Centre CMP, Bioelectronics Department, Mines Saint-Etienne, 2Panaxium SAS, 3Institut Universitaire de France, 4Institut des Neurosciences de la Timone (UMR7289), CNRS, Aix-Marseille Université, 5Centre Européen de Recherche en Imagerie Médicale, Aix-Marseille Université

This work describes the development of flexible interdigitated electrodes for implementation in 3D brain tumor models, namely, in vitro culture, in ovo model, and in vivo murine model. The proposed method can be used to evaluate the effects of pulsed electric fields on tumors at different levels of complexity.

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