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Method Article
Bronwen Connor
Connor lab, University of Auckland
Corresponding Author
License:
This work is licensed under a CC BY-NC 3.0 License. Read Full License
Direct reprogramming offers a unique approach by which to generate mature neural lineages for the study and treatment of neurological diseases and neurodevelopmental disorders. However, few studies have directly generated neural stem/precursor cells (iNPs) from adult human fibroblasts that are capable of producing a wide range of mature neuronal phenotypes. Further, current reprogramming protocols require the use of viral-mediated gene delivery to induce the generation of iNPs from human fibroblasts. Here we describe a robust and efficient protocol to directly reprogram adult human fibroblasts to expandable iNPs using transient non-viral gene delivery in a feeder-free cell culture system. The two transfection factors required for direct conversion to iNP cells, SOX2 and PAX6, do not generate a pluripotent cell state, reducing the potential risk of tumor formation following transplantation. By following the current protocol, iNPs are observed 6-9 weeks after transfection. Upon differentiation, iNPs generate both astrocytes and a range of phenotype-specific neurons.
Keywords
cell reprogramming, neural precursor cells, human fibroblasts, SOX2, PAX6
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The authors declare no competing financial interests.
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Method Article
Bronwen Connor
Connor lab, University of Auckland
Corresponding Author
Version History
CURRENT STATUS: Posted
Version 1
Posted 14 Apr, 2015
No community comments so far
Metrics
Comments: 0
HTML Views: ...
License:
This work is licensed under a CC BY-NC 3.0 License. Read Full License
Direct reprogramming offers a unique approach by which to generate mature neural lineages for the study and treatment of neurological diseases and neurodevelopmental disorders. However, few studies have directly generated neural stem/precursor cells (iNPs) from adult human fibroblasts that are capable of producing a wide range of mature neuronal phenotypes. Further, current reprogramming protocols require the use of viral-mediated gene delivery to induce the generation of iNPs from human fibroblasts. Here we describe a robust and efficient protocol to directly reprogram adult human fibroblasts to expandable iNPs using transient non-viral gene delivery in a feeder-free cell culture system. The two transfection factors required for direct conversion to iNP cells, SOX2 and PAX6, do not generate a pluripotent cell state, reducing the potential risk of tumor formation following transplantation. By following the current protocol, iNPs are observed 6-9 weeks after transfection. Upon differentiation, iNPs generate both astrocytes and a range of phenotype-specific neurons.
Figure 1
Figure 2
Figure 3
Figure 4
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