New publication: Improved method to generate neurons using CRISPRa technology and novel biomaterial
Updated: Feb 16, 2022
In collaboration with scientists from the Chinese Academy of Science, the team has reported an improved method to generate neurons from human iPS cells using CRISPRa technology and novel biomaterials composed of binary colloidal crystals (BCCs).
Combinatorial approach of binary colloidal crystals (BCCs) and CRISPR activation to improve induced pluripotent stem cell differentiation into neurons
Daniel Urrutia-Cabrera, Roxanne Hsiang-Chi Liou, Jiao Lin, Yue Shi, Kun Liu, Sandy Shen-Chi Hung, Alex W. Hewitt, Peng-Yuan Wang, Raymond Ching-Bong Wong
ACS Applied Materials & Interfaces, accepted 2022.01.28 (IF = 9.23)
Conventional methods of neuronal differentiation for human induced pluripotent stem cells (iPSCs) are tedious and complicated, involving multi-stage protocols with complex cocktails of growth factors and small molecules. Artificial extracellular matrices with defined surface topography and chemistry represent a promising venue to improve the neuronal differentiation in vitro. In the present study, we test the impact of a type of colloidal self-assembled patterns called binary colloidal crystals (BCCs) on neuronal differentiation. We develop a CRISPR activation (CRISPRa) iPSC platform that constitutively expresses the dCas9-VPR system, which allows robust activation of the proneural transcription factor NEUROD1 to rapidly induce neuronal differentiation within seven days. We show that the combinatorial use of BCCs can further improve this neuronal differentiation system. In particular, our results indicate that fine tuning of silica and polystyrene size is critical to generate specific topographies to improve neuronal differentiation and branching. BCCs with 5 μm silica and 100 nm carboxylated polystyrene have the most prominent effect on increasing neurite outgrowth and more complex ramification, while BCCs with 2 μm silica and 65 nm carboxylated polystyrene are better in promoting neuronal enrichment. These results indicate that biophysical cues can support rapid differentiation and improve neuronal maturation. In summary, our combinatorial approach of CRISPRa and BCCs provides a robust and rapid pipeline for in vitro production of human neurons. Specific BCCs can be adapted to late stages of neuronal differentiation protocols to improve neuronal maturation, which has important implications in tissue engineering, in vitro biological studies and disease modeling.
Read the publication here.