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Updated: Aug 7, 2026
Protocol for the Differentiation of Human Induced Pluripotent Stem Cells into Mixed Cultures of Neurons and Glia for Neurotoxicity Testing
Published on: June 9, 2017
POU3F4 up-regulates Gli1 expression and promotes neuronal differentiation and synaptic development of hippocampal
Lei Zhang1,2, Jue Wang1, Naijuan Xu1
1Department of Human Anatomy, Co-Innovation Center of Neuroregeneration, Nantong University, No.19 Qixiu Road, Nantong, 226001, Jiangsu, People's Republic of China.
Background:
Neural stem cells (NSCs) are considered to be the most promising cell type for cell replacement therapy in neurodegenerative diseases. However, their low neuronal differentiation ratio impedes their application in such conditions. Elucidating the molecular mechanism of NSC differentiation may provide the necessary experimental basis for expanding their application. Previous studies have indicated that POU3F4 can induce neuronal differentiation of NSCs, this study aims to underly the possible exact mechanism of POU3F4 on the NSC differentiation and development.
Methods:
NSCs were isolated and cultured from the hippocampus of neonatal mice. The frozen hippocampal sections were prepared for immunohistochemical staining. Synaptic development was assessed using electron microscopy. High-throughput sequencing was employed to analyze the gene expression profile following the overexpression of Brn4. Gene expression levels were determined through Western blotting and qRT-PCR. Cell cycle and differentiation were evaluated using flow cytometry and immunofluorescent staining.
Results:
It was found that POU3F4 promoted the neuronal differentiation of hippocampal NSCs and synapse development, and inhibited NSC proliferation. POU3F4-deficient mice exhibited impairments in learning and memory. RNA sequencing and ChIP assays confirmed that Gli1 was downstream of POU3F4. Loss and gain function experiments indicated that Gli1 mediated POU3F4 promoting neuronal differentiation and synapse development. Forced expression of Gli1 in hippocampus improved learning and memory function of animal models.
Conclusions:
The results suggest that POU3F4 and Gli1 promote neuronal differentiation and synaptic development of NSCs, and that Gli1 partially mediates the effects of POU3F4.
