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Title: Human pluripotent stem cell-derived neurons are functionally mature in vitro and integrate into the mouse striatum following transplantation
Author: Comella Bolla, Andrea
Orlandi, Javier G.
Miguez, Andrés
Straccia, Marco
García Bravo, María
Bombau, Georgina
Galofré, Mireia
Sanders, Phil
Carrere, Jordi
Segovia, José Carlos
Blasi Cabús, Joan
Allen, Nicholas D.
Alberch i Vié, Jordi
Soriano i Fradera, Jordi
Canals i Coll, Josep M.
Keywords: Malalties neurodegeneratives
Cèl·lules mare
Neurodegenerative Diseases
Stem cells
Issue Date: 30-Apr-2020
Publisher: Humana Press
Abstract: Human Pluripotent Stem Cells (hPSCs) are a powerful tool for modelling human development. In recent years, hPSCSs have become central in cell-based therapies for neurodegenerative diseases given their potential to replace affected neurons. However, directing hPSCs into specific neuronal types is complex and requires an accurate protocol that mimics endogenous neuronal development. Here we describe step-by-step a novel and fast feeder-free neuronal differentiation protocol to direct hPSCs onto mature forebrain neurons in 37 days in vitro (DIV). The protocol is grounded on a combination of specific morphogens, trophic and growth factors, ions, neurotransmitters and extracellular matrix elements. An induced hPSC line (Ctr-Q33) and an embryonic hPSC line (GEN-Q18) were used to reinforce the potential of the protocol. Neuronal activity was analysed by single-cell calcium imaging. At 8 DIV, we obtained a homogeneous population of hPSCs-derived neuroectodermal progenitors which self-arranged in bi-dimensional neural tube-like structures. At 16 DIV, we generated hPSC-derived neural progenitors (NPCs) with mostly subpallial identity along with a subpopulation of pallial NPCs. Terminal in vitro neuronal differentiation was confirmed by the expression of microtubule associated protein 2b (Map2b) by almost 100% of hPSC-derived neurons and the expression of specific-striatal neuronal markers including GABA, CTIP2 and DARPP-32. HPSC-derived neurons showed mature and functional phenotypes as they expressed synaptic markers, voltage-gated ion channels and neurotransmitter receptors. Neurons displayed diverse spontaneous activity patterns that were classified into three major groups, namely 'high', 'intermediate' and 'low' firing neurons. Finally, transplantation experiments in vivo showed that highly relevant, committed NPCs survived within mouse striatum for at least 3 months. NPCs embodied host environmental cues and differentiated into striatal medium size spiny neurons (MSNs), which successfully integrated into the endogenous circuitry without the appearance of any teratoma symptom. Altogether, present findings demonstrate the potential of this in vitro human neuronal differentiation protocol, which will bring new opportunities for the study of human neurodevelopment and neurodegeneration, and will open new avenues in cell-based therapies, cutting-edge pharmacological studies and toxicology.
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It is part of: Molecular Neurobiology, 2020, vol. 57, p. 2766-2798
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ISSN: 0893-7648
Appears in Collections:Articles publicats en revistes (IDIBAPS: Institut d'investigacions Biomèdiques August Pi i Sunyer)
Articles publicats en revistes (Institut de Recerca en Sistemes Complexos (UBICS))
Articles publicats en revistes (Física de la Matèria Condensada)
Articles publicats en revistes (Institut d'lnvestigació Biomèdica de Bellvitge (IDIBELL))
Articles publicats en revistes (Institut de Neurociències (UBNeuro))
Articles publicats en revistes (Biomedicina)

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