Rapid generation of regionally specified CNS neurons by sequential patterning and conversion of human induced pluripotent stem cells

Muwan Chen, Muyesier Maimaitili, Mette Habekost, Katherine P. Gill, Noëmie Mermet-Joret, Sadegh Nabavi, Fabia Febbraro, Mark Denham*

*Corresponding author for this work

Research output: Contribution to journal/Conference contribution in journal/Contribution to newspaperJournal articleResearchpeer-review

14 Citations (Scopus)

Abstract

The differentiation of patient-specific induced pluripotent stem cells (iPSCs) into specific neuronal subtypes has been exploited as an approach for modeling a variety of neurological disorders. However, achieving a highly pure population of neurons is challenging when using directed differentiation methods, especially for neuronal subtypes generated by complex and protracted protocols. In this study, we efficiently produced highly pure populations of regionally specified CNS neurons by using a modified NGN2-Puromycin direct conversion protocol. The protocol is amenable across a range of iPSC lines, with more than 95% of cells at day 21 positive for the neuronal marker MAP2. We found that conversion from pluripotent stem cells resulted in neurons from the central and peripheral nervous system; however, by incorporating a short CNS patterning step, we eliminated these peripheral neurons. Furthermore, we used the patterning step to control the rostral-caudal identity. This approach of sequential patterning and conversion produced pure populations of forebrain neurons, when patterned with SMAD inhibitors. Additionally, when SMAD inhibitors and WNT agonists were applied, the approach produced anterior hindbrain excitatory neurons and resulted in a neuronal population containing VSX2/SHOX2 V2a interneurons. Overall, this sequential patterning and conversion protocol can be used for the production of a variety of CNS excitatory neurons from patient-derived iPSCs, and is a highly versatile system for investigating early disease events for a range of neurological disorders including Alzheimer's disease, motor neurons disease and spinal cord injury.

Original languageEnglish
Article number101945
JournalStem Cell Research
Volume48
ISSN1873-5061
DOIs
Publication statusPublished - Oct 2020

Keywords

  • Cortical neurons
  • Direct conversion
  • Induced pluripotent stem cells
  • Patterning
  • V2a interneuron

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