Enhanced differentiation of human induced pluripotent stem cells toward the midbrain dopaminergic neuron lineage through GLYPICAN-4 downregulation

Serena Corti, Remi Bonjean, Thomas Legier, Diane Rattier, Christophe Melon, Pascal Salin, Erik A Toso, Michael Kyba, Lydia Kerkerian-Le Goff, Flavio Maina, Rosanna Dono

Research output: Contribution to journalArticlepeer-review

Abstract

Enhancing the differentiation potential of human induced pluripotent stem cells (hiPSC) into disease-relevant cell types is instrumental for their widespread application in medicine. Here, we show that hiPSCs downregulated for the signaling modulator GLYPICAN-4 (GPC4) acquire a new biological state characterized by increased hiPSC differentiation capabilities toward ventral midbrain dopaminergic (VMDA) neuron progenitors. This biological trait emerges both in vitro, upon exposing cells to VMDA neuronal differentiation signals, and in vivo, even when transplanting hiPSCs at the extreme conditions of floor-plate stage in rat brains. Moreover, it is compatible with the overall neuronal maturation process toward acquisition of substantia nigra neuron identity. HiPSCs with downregulated GPC4 also retain self-renewal and pluripotency in stemness conditions, in vitro, while losing tumorigenesis in vivo as assessed by flank xenografts. In conclusion, our results highlight GPC4 downregulation as a powerful approach to enhance generation of VMDA neurons. Outcomes may contribute to establish hiPSC lines suitable for translational applications.

Original languageEnglish (US)
Pages (from-to)725-742
Number of pages18
JournalStem Cells Translational Medicine
Volume10
Issue number5
DOIs
StatePublished - May 2021

Bibliographical note

Funding Information:
We thank: all members of our labs for helpful discussions and comments; A. Fico for critical reading of the manuscript; F. Shi and M. Sorce for contribution during the initial analysis of GPC4sh hiPSCs, and during the initial analysis of cell transplanted rat brain, respectively; A. Gagna for helping us to develop a mathematical method to calculate differentiation efficiency of different experiments; S. Lev and L. Khera for producing lentivirus particles carrying shRNA and for advises on use of lentivirus. Microscopy was performed at the imaging platform of the IBDM, supported by the ANR through the “Investments for the Future” program (France‐BioImaging, ANR‐10‐INSB‐04‐01). The authors thank members of the IBDM core facility for microscopy and mouse husbandry for their technical support. This project is a part of the research program of the Centre of Excellence DHUNE, which is supported by the French National Plan on Neurodegenerative Diseases funded by the French Ministry of national education, higher education and research and the “Investissements d'Avenir” French Government program. The IBDM is affiliated with NeuroMarseille (AMX‐19‐IET‐004), the AMU neuroscience network, and with NeuroSchool, the AMU graduate school in neuroscience supported by the A*MIDEX foundation and the “Investissements d'Avenir” program (nEURo*AMU, ANR‐17‐EURE‐0029 grant). This work was funded by Association France Parkinson (Convention 096038), Fondation de France (2013_00043173), SATT Sud‐Est “Technology Transfer accelerator”, Programme de pré‐maturation du Centre National de la Recherche Scientifique to R.D.,Network of Centres of Excellence in Neurodegeneration (COEN Pathfinder III 4014) to R.D. and L.K.L.G., and was supported by Centre National de la Recherche Scientifique and Aix‐Marseille Université. F.M. received support from FRM (Fondation Recherche Médicale; DEQ20141231766). S.C. was supported by the BIOTRAIL PH.D. Program in Life Science, Marseille, France and by Association France Parkinson 4th year PH.D. fellowships. R.B. was supported by a grant from the Programme de pré‐maturation du Centre National de la Recherche Scientifique to R.D. T.L. and D.R. were supported by the French Ministry of Higher Education, Innovation and Research fellowship. M.K. and E.A.T. were supported by the National Institute of Arthritis and Musculoskeletal and Skin Diseases; R01 AR075413. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.

Funding Information:
We thank: all members of our labs for helpful discussions and comments; A. Fico for critical reading of the manuscript; F. Shi and M. Sorce for contribution during the initial analysis of GPC4sh hiPSCs, and during the initial analysis of cell transplanted rat brain, respectively; A. Gagna for helping us to develop a mathematical method to calculate differentiation efficiency of different experiments; S. Lev and L. Khera for producing lentivirus particles carrying shRNA and for advises on use of lentivirus. Microscopy was performed at the imaging platform of the IBDM, supported by the ANR through the ?Investments for the Future? program (France-BioImaging, ANR-10-INSB-04-01). The authors thank members of the IBDM core facility for microscopy and mouse husbandry for their technical support. This project is a part of the research program of the Centre of Excellence DHUNE, which is supported by the French National Plan on Neurodegenerative Diseases funded by the French Ministry of national education, higher education and research and the ?Investissements d'Avenir? French Government program. The IBDM is affiliated with NeuroMarseille (AMX-19-IET-004), the AMU neuroscience network, and with NeuroSchool, the AMU graduate school in neuroscience supported by the A*MIDEX foundation and the ?Investissements d'Avenir? program (nEURo*AMU, ANR-17-EURE-0029 grant). This work was funded by Association France Parkinson (Convention 096038), Fondation de France (2013_00043173), SATT Sud-Est ?Technology Transfer accelerator?, Programme de pr?-maturation du Centre National de la Recherche Scientifique to R.D.,Network of Centres of Excellence in Neurodegeneration (COEN Pathfinder III 4014) to R.D. and L.K.L.G., and was supported by Centre National de la Recherche Scientifique and Aix-Marseille Universit?. F.M. received support from FRM (Fondation Recherche M?dicale; DEQ20141231766). S.C. was supported by the BIOTRAIL PH.D. Program in Life Science, Marseille, France and by Association France Parkinson 4th year PH.D. fellowships. R.B. was supported by a grant from the Programme de pr?-maturation du Centre National de la Recherche Scientifique to R.D. T.L. and D.R. were supported by the French Ministry of Higher Education, Innovation and Research fellowship. M.K. and E.A.T. were supported by the National Institute of Arthritis and Musculoskeletal and Skin Diseases; R01 AR075413. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.

Publisher Copyright:
© 2021 The Authors. STEM CELLS TRANSLATIONAL MEDICINE published by Wiley Periodicals LLC on behalf of AlphaMed Press

Keywords

  • GLYPICAN-4
  • hiPSCs
  • hPSC-derived dopaminergic neurons
  • intrastriatal transplantation
  • self-renewal and differentiation

PubMed: MeSH publication types

  • Journal Article

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