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A simple method for large‐scale generation of dopamine neurons from human embryonic stem cells
Author(s) -
Morizane Asuka,
Darsalia Vladimer,
Guloglu M. Oktar,
Hjalt Tord,
Carta Manolo,
Li JiaYi,
Brundin Patrik
Publication year - 2010
Publication title -
journal of neuroscience research
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.72
H-Index - 160
eISSN - 1097-4547
pISSN - 0360-4012
DOI - 10.1002/jnr.22515
Subject(s) - embryonic stem cell , progenitor cell , biology , dopaminergic , transplantation , directed differentiation , basic fibroblast growth factor , stem cell , progenitor , dopamine , stromal cell , microbiology and biotechnology , neuroscience , induced pluripotent stem cell , cancer research , growth factor , medicine , genetics , receptor , gene
Dopamine (DA) neurons derived from human embryonic stem cells (hESCs) are potentially valuable in drug screening and as a possible source of donor tissue for transplantation in Parkinson's disease. However, existing culture protocols that promote the differentiation of DA neurons from hESCs are complex, involving multiple steps and having unreliable results between cultures. Here we report a simple and highly reproducible culture protocol that induces expandable DA neuron progenitors from hESCs in attached cultures. We found that the hESC‐derived neuronal progenitors retain their full capacity to generate DA neurons after repeated passaging in the presence of basic fibroblast growth factor (bFGF) and medium conditioned with PA6 stromal cells. Using immunocytochemistry and RT‐PCR, we found that the differentiated DA neurons exhibit a midbrain phenotype and express, e.g., Aldh1a, Ptx3, Nurr1, and Lmx1a. Using HPLC, we monitored their production of DA. We then demonstrated that the expanded progenitors are possible to cryopreserve without loosing the dopaminergic phenotype. With our protocol, we obtained large and homogeneous populations of dopaminergic progenitors and neurons. We conclude that our protocol can be used to generate human DA neurons suitable for the study of disease mechanisms, toxicology, drug screening, and intracerebral transplantation. © 2010 Wiley‐Liss, Inc.