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Sorbonne Université, INSERM, CNRS, F-75012 Paris, France

6 ARTICLES PUBLISHED IN JoVE

Medicine

Transcriptomic Analysis of Human Retinal Surgical Specimens Using jouRNAl

Marie-Noëlle Delyfer^1,2,3,4, Najate Aït-Ali^1,2,3, Hawa Camara^1,2,3, Emmanuelle Clérin^1,2,3, Jean-François Korobelnik⁴, José-Alain Sahel^1,2,3, Thierry Léveillard^1,2,3

¹U968, Institut National de la Santé et de la Recherche Médicale, ²UMR S 968, Université Pierre et Marie Curie, ³UMR 7210, Centre National de la Recherche Scientifique, ⁴Départment d'Ophtalmologie, Centre Hospitalier Universitaire de Bordeaux

We used retinal samples from retinectomy for a transcriptomic analysis of retinal detachment. We developed a procedure that allows RNA conservation between the surgical blocks and the laboratory. We standardized a protocol to purify RNA by cesium chloride ultracentrifugation to assure that the purified RNAs are suitable for microarray analysis.

Neuroscience

Vibratome Sectioning Mouse Retina to Prepare Photoreceptor Cultures

Emmanuelle Clérin^1,4,5,6, Ying Yang^1,4,5,6, Valérie Forster^2,4,5,6, Valérie Fontaine^3,4,5,6, José-Alain Sahel^4,5,6, Thierry Léveillard^1,4,5,6

¹Department of Genetics, UMR_S 968, Institut de la Vision, ²Department of Visual Information, UMR_S 968, Institut de la Vision, ³Exploratory Team, UMR_S 968, Institut de la Vision, ⁴Sorbonne Universités, Paris 06, UMR_S 968, Institut de la Vision, ⁵INSERM, U968, Institut de la Vision, ⁶CNRS, UMR_7210, Institut de la Vision

Neural retina of a mouse aged 8 days is on top of a 4% gelatin block. After isolation of the photoreceptor layer (200 µm) by vibratome, the photoreceptors are seeded after mechanical and enzymatic dissociation for culture. The photoreceptor layer can be used for molecular, biochemical analyses or transplantation.

Medicine

Using Adeno-associated Virus as a Tool to Study Retinal Barriers in Disease

Ophélie Vacca^1,2,3, Brahim El Mathari^1,2,3, Marie Darche^1,2,3, José-Alain Sahel^1,2,3, Alvaro Rendon^1,2,3, Deniz Dalkara^1,2,3

¹Department of Therapeutics, Institut de la Vision, Sorbonne Universtés, UPMC Univ Paris 06, UMR_S 968, ²INSERM, U968, ³CNRS, UMR_7210

To investigate the blood-retinal barrier permeability and the inner limiting membrane integrity in animal models of retinal disease, we used several adeno-associated virus (AAV) variants as tools to label retinal neurons and glia. Virus mediated reporter gene expression is then used as an indicator of retinal barrier permeability.

Developmental Biology

Defined Xeno-free and Feeder-free Culture Conditions for the Generation of Human iPSC-derived Retinal Cell Models

Amélie Slembrouck-Brec^*1, Céline Nanteau^*1, José-Alain Sahel¹, Olivier Goureau¹, Sacha Reichman¹

¹Institut de la Vision, Sorbonne Université, INSERM, CNRS, F-75012 Paris, France

The production of specialized retinal cells from pluripotent stem cells is a turning point in the development of stem cell-based therapy for retinal diseases. The present paper describes a simple method for an efficient generation of retinal organoids and retinal pigmented epithelium for basic, translational, and clinical research.

Developmental Biology

Engineering Transplantation-suitable Retinal Pigment Epithelium Tissue Derived from Human Embryonic Stem Cells

Karim Ben M'Barek^1,2,3, Walter Habeler^1,2,3, Alexandra Plancheron^1,2,3, Mohamed Jarraya⁴, Olivier Goureau⁵, Christelle Monville^1,2

¹U861, I-Stem, Association Française contre les Myopathies (AFM), Institut National de la Santé et de la Recherche Médicale (INSERM), ²U861, I-Stem, Association Française contre les Myopathies (AFM), Université Evry Val-d'Essonne (UEVE), ³I-Stem, Association Française contre les Myopathies (AFM), Centre pour L’Etude des Cellules Souches (CECS), ⁴Banque de tissus humain, Hôpital Saint Louis, Assistance Publique - Hôpitaux de Paris (AP-HP), ⁵Sorbonne Université, INSERM, CNRS, Institut de la Vision, F-75012

We describe a method to engineer a retinal tissue composed of retinal pigment epithelial cells derived from human pluripotent stem cells cultured on top of human amniotic membranes and its preparation for grafting in animal models.

Neuroscience

Cone-Enriched Cultures from the Retina of Chicken Embryos to Study Rod to Cone Cellular Interactions

Géraldine Millet-Puel¹, Myriam Pinault¹, Marie Cordonnier¹, Valérie Fontaine¹, José-Alain Sahel¹, Thierry Léveillard¹

¹Department of Genetics - Sorbonne Université, INSERM, CNRS, Institut de la Vision

We describe a method to obtain primary cultures of cone photoreceptors from the retina of chicken embryos and its use for high content screening.