Neurotrophic factor 3-chitosan carrier induces neural stem cells to differentiate into neuronal subtypes and their electrophysiological properties

doi:10.12307/2023.081

Abstract

Abstract: BACKGROUND: Previous studies have confirmed that the neurotrophic factor 3-chitosan carrier can support the survival and proliferation of neural stem cells, and can efficiently induce neural stem cells to differentiate into neurons.
OBJECTIVE: To observe the effects of neurotrophic factor 3-chitosan carrier on neuronal development and its electrophysiological properties at various stages of development and the subtypes of mature neurons.
METHODS: Passage 3 spinal cord neural stem cells of neonatal rats were collected and cultured in four groups. The blank control group was added with neural stem cell culture medium. Chitosan group was added with neural stem cell culture medium containing chitosan. Neurotrophic factor 3 group was added with neural stem cell medium containing neurotrophic factor 3. Neurotrophic factor 3-chitosan group was added with neural stem cell medium containing neurotrophic factor 3-chitosan carrier. Immunofluorescence staining was performed to observe the development of neurons at various stages. The changes in electrophysiological properties during neural stem cell development were evaluated by whole-cell patch-clamp technology. Interneuron subtypes were visualized by immunofluorescence staining 21 days after neural stem cell differentiation.
RESULTS AND CONCLUSION: (1) Immunofluorescence staining for Nestin, DCX, Tuj1 and MAP2 showed that the neurotrophic factor 3-chitosan carrier maintained the homeostasis of the neural stem cell pool and promoted neuronal maturation by accelerating the developmental process of neuroblasts. (2) Whole-cell patch-clamp technology results demonstrated that the neurotrophic factor 3 and neurotrophic factor 3-chitosan promoted cell membrane function and the maturation of ion channels on the cell membrane early in development. However, only neurotrophic factor 3-chitosan can maintain this advantage to the middle and late stages of development, that is, 7-14 days after differentiation. (3) Immunofluorescence staining exhibited that after 21 days of neural stem cell differentiation, mature neurons in the neurotrophic factor 3-chitosan group could express the motor neuron specific marker HB9, V1 type interneuron FOXP1, V2 type interneuron specific marker LHX3, and regulate a specific marker VGLUT3 that regulates mechanical pain sensory interneurons. (4) These results have verified that the neurotrophic factor 3-chitosan carrier can promote the development of neural stem cells to neuroblasts, and the function of the cell membrane and the maturation of ion channels on the cell membrane in the early stage of development to a certain degree, and can induce the diversification of mature neuronal subtypes.

Key words: neurotrophic factor 3, chitosan carrier, neural stem cell, development, neuron, patch clamp, differentiation, ion channel, electrophysiology, interneuron

CLC Number:

Zhang Boya, Duan Hongmei, Bai Tianyu, Hao Fei, Hao Peng, Zhao Wen, Gao Yudan, Li Xiaoguang, Yang Zhaoyang. Neurotrophic factor 3-chitosan carrier induces neural stem cells to differentiate into neuronal subtypes and their electrophysiological properties[J]. Chinese Journal of Tissue Engineering Research, 2023, 27(25): 4020-4027.

Figures/Tables 7

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