Publications by authors named "Amélie Slembrouck-Brec"

6 Publications

  • Page 1 of 1

Reproducing diabetic retinopathy features using newly developed human induced-pluripotent stem cell-derived retinal Müller glial cells.

Glia 2021 Mar 8. Epub 2021 Mar 8.

Institut de la Vision, Sorbonne Université, INSERM, CNRS, Paris, France.

Muller glial cells (MGCs) are responsible for the homeostatic and metabolic support of the retina. Despite the importance of MGCs in retinal disorders, reliable and accessible human cell sources to be used to model MGC-associated diseases are lacking. Although primary human MGCs (pMGCs) can be purified from post-mortem retinal tissues, the donor scarcity limits their use. To overcome this problem, we developed a protocol to generate and bank human induced pluripotent stem cell-derived MGCs (hiMGCs). Using a transcriptome analysis, we showed that the three genetically independent hiMGCs generated were homogeneous and showed phenotypic characteristics and transcriptomic profile of pMGCs. These cells expressed key MGC markers, including Vimentin, CLU, DKK3, SOX9, SOX2, S100A16, ITGB1, and CD44 and could be cultured up to passage 8. Under our culture conditions, hiMGCs and pMGCs expressed low transcript levels of RLPB1, AQP4, KCNJ1, KCJN10, and SLC1A3. Using a disease modeling approach, we showed that hiMGCs could be used to model the features of diabetic retinopathy (DR)-associated dyslipidemia. Indeed, palmitate, a major free fatty acid with elevated plasma levels in diabetic patients, induced the expression of inflammatory cytokines found in the ocular fluid of DR patients such as CXCL8 (IL-8) and ANGPTL4. Moreover, the analysis of palmitate-treated hiMGC secretome showed an upregulation of proangiogenic factors strongly related to DR, including ANG2, Endoglin, IL-1β, CXCL8, MMP-9, PDGF-AA, and VEGF. Thus, hiMGCs could be an alternative to pMGCs and an extremely valuable tool to help to understand and model glial cell involvement in retinal disorders, including DR.
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http://dx.doi.org/10.1002/glia.23983DOI Listing
March 2021

Generation of a Transplantable Population of Human iPSC-Derived Retinal Ganglion Cells.

Front Cell Dev Biol 2020 27;8:585675. Epub 2020 Oct 27.

Institut de la Vision, Sorbonne Université, INSERM, CNRS, Paris, France.

Optic neuropathies are a major cause of visual impairment due to retinal ganglion cell (RGC) degeneration. Human induced-pluripotent stem cells (iPSCs) represent a powerful tool for studying both human RGC development and RGC-related pathological mechanisms. Because RGC loss can be massive before the diagnosis of visual impairment, cell replacement is one of the most encouraging strategies. The present work describes the generation of functional RGCs from iPSCs based on innovative 3D/2D stepwise differentiation protocol. We demonstrate that targeting the cell surface marker THY1 is an effective strategy to select transplantable RGCs. By generating a fluorescent GFP reporter iPSC line to follow transplanted cells, we provide evidence that THY1-positive RGCs injected into the vitreous of mice with optic neuropathy can survive up to 1 month, intermingled with the host RGC layer. These data support the usefulness of iPSC-derived RGC exploration as a potential future therapeutic strategy for optic nerve regeneration.
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http://dx.doi.org/10.3389/fcell.2020.585675DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7652757PMC
October 2020

Generation of human induced pluripotent stem cell lines from a patient with ITM2B-related retinal dystrophy and a non mutated brother.

Stem Cell Res 2019 12 5;41:101625. Epub 2019 Nov 5.

INSERM, CNRS, Institut de la Vision, Sorbonne Université, 17 rue Moreau, Paris, F-75012, France; CHNO des Quinze-Vingts, INSERM-DGOS CIC 1423, 28 rue de Charenton, Paris, F-75012, France. Electronic address:

Human induced pluripotent stem cell (iPSC) lines were generated from fibroblasts of a patient affected with an autosomal dominant retinal dystrophy carrying the mutation c.782A>C, p.Glu261Ala in ITM2B and from an unaffected brother. Three different iPSC lines were generated and characterized from primary dermal fibroblasts of the affected subject and two from the unaffected brother. All iPSC lines expressed the pluripotency markers, were able to differentiate into the three germ layers and presented normal karyotypes. This cellular model will provide a powerful tool to study this retinal dystrophy and better understand the role of ITM2B.
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http://dx.doi.org/10.1016/j.scr.2019.101625DOI Listing
December 2019

Reprogramming of Adult Retinal Müller Glial Cells into Human-Induced Pluripotent Stem Cells as an Efficient Source of Retinal Cells.

Stem Cells Int 2019 15;2019:7858796. Epub 2019 Jul 15.

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

The reprogramming of human somatic cells to induced pluripotent stem cells (iPSCs) has broad applications in regenerative medicine. The generation of self-organized retinal structures from these iPSCs offers the opportunity to study retinal development and model-specific retinal disease with patient-specific iPSCs and provides the basis for cell replacement strategies. In this study, we demonstrated that the major type of glial cells of the human retina, Müller cells, can be reprogrammed into iPSCs that acquire classical signature of pluripotent stem cells. These Müller glial cell-derived iPSCs were able to differentiate toward retinal fate and generate concomitantly retinal pigmented epithelial cells and self-forming retinal organoid structures containing retinal progenitor cells. Retinal organoids recapitulated retinal neurogenesis with differentiation of retinal progenitor cells into all retinal cell types in a sequential overlapping order. With a modified retinal maturation protocol characterized by the presence of serum and high glucose levels, our study revealed that the retinal organoids contained pseudolaminated neural retina with important features reminiscent of mature photoreceptors, both rod and cone subtypes. This advanced maturation of photoreceptors not only supports the possibility to use 3D retinal organoids for studying photoreceptor development but also offers a novel opportunity for disease modeling, particularly for inherited retinal diseases.
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http://dx.doi.org/10.1155/2019/7858796DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6664555PMC
July 2019

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

J Vis Exp 2018 09 6(139). Epub 2018 Sep 6.

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

The production of specialized cells from pluripotent stem cells provides a powerful tool to develop new approaches for regenerative medicine. The use of human-induced pluripotent stem cells (iPSCs) is particularly attractive for neurodegenerative disease studies, including retinal dystrophies, where iPSC-derived retinal cell models mark a major step forward to understand and fight blindness. In this paper, we describe a simple and scalable protocol to generate, mature, and cryopreserve retinal organoids. Based on medium changing, the main advantage of this method is to avoid multiple and time-consuming steps commonly required in a guided differentiation of iPSCs. Mimicking the early phases of retinal development by successive changes of defined media on adherent human iPSC cultures, this protocol allows the simultaneous generation of self-forming neuroretinal structures and retinal pigmented epithelial (RPE) cells in a reproducible and efficient manner in 4 weeks. These structures containing retinal progenitor cells (RPCs) can be easily isolated for further maturation in a floating culture condition enabling the differentiation of RPCs into the seven retinal cell types present in the adult human retina. Additionally, we describe quick methods for the cryopreservation of retinal organoids and RPE cells for long-term storage. Combined together, the methods described here will be useful to produce and bank human iPSC-derived retinal cells or tissues for both basic and clinical research.
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http://dx.doi.org/10.3791/57795DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6235103PMC
September 2018

Characterization and Transplantation of CD73-Positive Photoreceptors Isolated from Human iPSC-Derived Retinal Organoids.

Stem Cell Reports 2018 09 9;11(3):665-680. Epub 2018 Aug 9.

Institut de la Vision, Sorbonne Université, INSERM, CNRS, 17, Rue Moreau, Paris 75012, France. Electronic address:

Photoreceptor degenerative diseases are a major cause of blindness for which cell replacement is one of the most encouraging strategies. For stem cell-based therapy using human induced pluripotent stem cells (hiPSCs), it is crucial to obtain a homogenous photoreceptor cell population. We confirmed that the cell surface antigen CD73 is exclusively expressed in hiPSC-derived photoreceptors by generating a fluorescent cone rod homeobox (Crx) reporter hiPSC line using CRISPR/Cas9 genome editing. We demonstrated that CD73 targeting by magnetic-activated cell sorting (MACS) is an effective strategy to separate a safe population of transplantable photoreceptors. CD73+ photoreceptor precursors can be isolated in large numbers and transplanted into rat eyes, showing capacity to survive and mature in close proximity to host inner retina of a model of photoreceptor degeneration. These data demonstrate that CD73+ photoreceptor precursors hold great promise for a future safe clinical translation.
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http://dx.doi.org/10.1016/j.stemcr.2018.07.005DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6135113PMC
September 2018