論文

査読有り 国際誌
2020年2月25日

miRNA-Based Rapid Differentiation of Purified Neurons from hPSCs Advancestowards Quick Screening for Neuronal Disease Phenotypes In Vitro.

Cells
  • Mitsuru Ishikawa
  • ,
  • Takeshi Aoyama
  • ,
  • Shoichiro Shibata
  • ,
  • Takefumi Sone
  • ,
  • Hiroyuki Miyoshi
  • ,
  • Hirotaka Watanabe
  • ,
  • Mari Nakamura
  • ,
  • Saori Morota
  • ,
  • Hiroyuki Uchino
  • ,
  • Andrew S Yoo
  • ,
  • Hideyuki Okano

9
3
記述言語
英語
掲載種別
研究論文(学術雑誌)
DOI
10.3390/cells9030532

Obtaining differentiated cells with high physiological functions by an efficient, but simple and rapid differentiation method is crucial for modeling neuronal diseases in vitro using human pluripotent stem cells (hPSCs). Currently, methods involving the transient expression of one or a couple of transcription factors have been established as techniques for inducing neuronal differentiation in a rapid, single step. It has also been reported that microRNAs can function as reprogramming effectors for directly reprogramming human dermal fibroblasts to neurons. In this study, we tested the effect of adding neuronal microRNAs, miRNA-9/9*, and miR-124 (miR-9/9*-124), for the neuronal induction method of hPSCs using Tet-On-driven expression of the Neurogenin2 gene (Ngn2), a proneural factor. While it has been established that Ngn2 can facilitate differentiation from pluripotent stem cells into neurons with high purity due to its neurogenic effect, a long or indefinite time is required for neuronal maturation with Ngn2 misexpression alone. With the present method, the cells maintained a high neuronal differentiation rate while exhibiting increased gene expression of neuronal maturation markers, spontaneous calcium oscillation, and high electrical activity with network bursts as assessed by a multipoint electrode system. Moreover, when applying this method to iPSCs from Alzheimer's disease (AD) patients with presenilin-1 (PS1) or presenilin-2 (PS2) mutations, cellular phenotypes such as increased amount of extracellular secretion of amyloid β42, abnormal oxygen consumption, and increased reactive oxygen species in the cells were observed in a shorter culture period than those previously reported. Therefore, it is strongly anticipated that the induction method combining Ngn2 and miR-9/9*-124 will enable more rapid and simple screening for various types of neuronal disease phenotypes and promote drug discovery.

リンク情報
DOI
https://doi.org/10.3390/cells9030532
PubMed
https://www.ncbi.nlm.nih.gov/pubmed/32106535
PubMed Central
https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7140514

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