A method for inducing neural stem cells to differentiate into neurons in vitro

A neural stem cell and directional differentiation technology, applied in the field of inducing neural stem cells directional differentiation into neurons in vitro, can solve the problems of low neural stem cell differentiation ratio, low neuron purity, and no unified standard, and achieves the promotion of pure differentiation and high purity. , the effect of the simple method

Active Publication Date: 2021-04-09
XINYANG NORMAL UNIVERSITY
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Problems solved by technology

[0003] At present, the method of inducing neural stem cells to differentiate into neurons in vitro is still in the exploratory stage, and no unified standard has been formed. The differentiation ratio of neural stem cells into neurons is relatively low, and the purity of neurons obtained is low.

Method used

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  • A method for inducing neural stem cells to differentiate into neurons in vitro
  • A method for inducing neural stem cells to differentiate into neurons in vitro
  • A method for inducing neural stem cells to differentiate into neurons in vitro

Examples

Experimental program
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Embodiment 1

[0028] A method for inducing directional differentiation of neural stem cells into neurons in vitro, comprising the steps of:

[0029] (1) Primary culture of neural stem cells

[0030] Under sterile conditions, 14.5-day-pregnant ICR mice were sacrificed by dislocation of the cervical spine, soaked in 75% ethanol for 5-8 minutes, then transferred to an ultra-clean table for dissection, the fetal mice were taken out, and the brain cortex was separated, and then passed through 1×PBS After rinsing with DMEM / F12 solution, transfer to the growth medium containing 1% N2, 1% B27, 20ng / mL bFGF, 20ng / mL EGF, 97% DMEM / F12, and cut the brain tissue with dissecting scissors, Use a 1mL sterile pipette tip to blow repeatedly into a single-cell suspension. After the single-cell suspension is filtered through a 140-mesh nylon screen, transfer it to a 50ML centrifuge tube. The solution formula is: 1% N2, 1% B27, 20ng / mL bFGF, 20ng / mL EGF, 97% DMEM / F12) to adjust the cell density to 1×10 6 cel...

Embodiment 2

[0038] In step (3), the cell density of plating is 5×10 4 cells / mL, the rest are the same as in Example 1.

Embodiment 3

[0040] In step (3), the cell density of plating is 5×10 6 cells / mL, the rest are the same as in Example 1.

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Abstract

The invention relates to a method for inducing neural stem cells to differentiate into neurons in vitro. The fetal mouse brain tissue of ICR pregnant mice with 14-15 days of pregnancy is used as a raw material to perform primary culture of neural stem cells to obtain neurospheres, which are then centrifuged and removed. Wash the precipitate with PBS and digest it, then add the culture medium to stop, centrifuge to remove the supernatant, add the growth medium to the precipitate, carry out the subculture of neural stem cells, and take the fourth or fifth generation of neurospheres to digest and prepare single cells Suspension, to 5×10 4 -5×10 6 The cell density of cells / mL was plated on a 12-well cell culture plate coated with polyornithine and laminin, cultured overnight, and differentiated culture medium was added the next day to obtain mature neurons. The method of the invention is simple and has good repeatability, can significantly increase the differentiation ratio of neural stem cells to neurons, promote the purity differentiation of neurons, and can obtain neurons with good growth state and high purity.

Description

technical field [0001] The invention relates to a method for inducing directional differentiation of neural stem cells into neurons in vitro, belonging to the technical field of biomedicine. Background technique [0002] Neural stem cells are pluripotent cells with self-renewal and differentiation capabilities in the central nervous system, which can differentiate into neurons, astrocytes and oligodendrocytes. The discovery of neural stem cells is of great significance, providing new research directions for central nervous system development, nerve injury repair, and treatment of neurodegenerative diseases, and has broad prospects for the repair and treatment of nervous system diseases or injuries. Studies have shown that the number of neural stem cells in the adult animal brain is small and most of them are in a resting state. Other studies have shown that most of the exogenous neural stem cells transplanted into the host differentiate into glial cells, and only a small num...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N5/0793C12N5/0797
CPCC12N5/0619C12N2501/11C12N2501/115C12N2506/08C12N2533/32C12N2533/52
Inventor 周棋赢韩月华周玉玲刘磊杨宁宁王仪佳王艳丽梁萍
Owner XINYANG NORMAL UNIVERSITY
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