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Method for differentiating neural stem cells into oligodendroglia cells, culture medium and application

A technology for oligodendrocytes and neural stem cells, applied in the field of culture medium and serum substitutes, can solve the problems of low induction efficiency, hindering the deep research of oligodendrocytes, failing to meet the standards of clinical research, etc. Low cost and the effect of improving the induction rate

Pending Publication Date: 2022-07-19
INNER MONGOLIA UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

At present, there are some methods for inducing neural stem cells into oligodendrocytes, but these methods have the defect of low induction efficiency and cannot meet the standards of clinical research, which hinders people's in-depth research on oligodendrocytes

Method used

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  • Method for differentiating neural stem cells into oligodendroglia cells, culture medium and application
  • Method for differentiating neural stem cells into oligodendroglia cells, culture medium and application
  • Method for differentiating neural stem cells into oligodendroglia cells, culture medium and application

Examples

Experimental program
Comparison scheme
Effect test

preparation example 1

[0095] (1) The 14.5-day pregnant ICR mice were killed by cervical dislocation, the abdomen was dissected, and the uterus and fetus were taken out.

[0096] (2) After washing the uterus and the fetal rat for 2-3 times with ice PBS buffer, the fetal rat is separated from other tissues. The head of the fetal rat was separated with forceps, and the fetal rat brain was dissected out after washing with ice-cold PBS buffer.

[0097] (3) The striatum was isolated from the fetal rat brain, and the striatum was minced in ice PBS buffer, then digested with Accutase cell separation solution, and centrifuged to collect and separate the neural stem cell single cell suspension.

[0098] (4) The single-cell suspension of isolated neural stem cells was divided into 1x10 5 Cells / mL were seeded in 1 wt% gelatin-coated dishes and incubated at 37 °C and 5% CO. 2 Cultured under the same conditions, the neural stem cell culture medium was replaced every two days; when cultured to the 4th to 5th da...

Embodiment 1

[0101] The neural stem cells to be differentiated obtained in Preparation Example 1 were cultured in the first induction medium, and the first induction medium was replaced every two days on days 1 to 4; 50% of the supernatant was removed every day from days 5 to 7 and replaced with the second Inducer; second inducer was replaced every two days after day 7.

[0102] The first inducing group is composed of the first mixed inducing solution, platelet-derived growth factor PDGF-aa and epidermal growth factor EGF. The first mixed induction solution consisted of 87 vol% DMEM:F12 (1:1) liquid medium medium, 10 vol% KnockOut serum replacement, 2 vol% B27 cell culture supplement and 1 vol% penicillin-streptomycin mixture. Based on the first mixed induction solution, the concentration of platelet-derived growth factor PDGF-aa was 10 ng / mL, and the concentration of epidermal growth factor EGF was 20 ng / mL.

[0103]The second inducer is composed of the second mixed inducer and sonic hed...

experiment example

[0109] 1. mRNA expression test of cell marker genes

[0110] Identification was performed using RT-PCR (Reverse Transcript-Polymer Chain Reaction) technology. Therefore, during the induction process, samples were taken on day 0, day 3, day 5, day 7, day 9, day 11, and day 13, respectively. Extraction of total RNA was carried out according to the instructions, followed by mRNA concentration detection, reverse transcription reaction, PCR amplification reaction and gel electrophoresis, and finally exposure detection and analysis were carried out according to standard experimental techniques.

[0111] 2. Protein expression test of cell marker genes

[0112] Identification was performed using Western Blot technology. Therefore, during the induction process, samples were taken on day 0, day 3, day 5, day 7, day 9, day 11, and day 13, respectively. According to the instructions, the total protein was centrifuged, followed by protein concentration detection, protein SDS-PAGE gel el...

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Abstract

The invention discloses a method for differentiating neural stem cells into oligodendroglia cells, a culture medium and application. The method comprises the following steps: culturing neural stem cells to be differentiated in a first induction medium to form oligodendrocyte precursor cells; culturing the oligodendroglia precursor cells in a second induction medium to form oligodendroglia cells; wherein the first induction medium contains a first mixed induction liquid, a platelet-derived growth factor PDGF-aa and an epidermal growth factor EGF, and the first mixed induction liquid contains a KnockOut serum substitute; the second induction medium contains a second mixed induction liquid and a sound hedgehog Shh, and the second mixed induction liquid contains a KnockOut serum substitute. According to the method, the inductivity is improved in a low-cost manner. The invention further discloses application of the culture medium and the serum substitute.

Description

technical field [0001] The present invention relates to a method for differentiating neural stem cells into oligodendrocytes, and also relates to the use of culture medium and serum substitutes. Background technique [0002] Oligodendrocytes are distributed in the central nervous system, and their main functions are to wrap axons in the central nervous system, form an insulating myelin sheath, assist in the efficient transmission of bioelectrical signals, and maintain and protect the normal functioning of neurons. Function. Abnormal oligodendrocytes can not only lead to demyelination of the central nervous system, but also cause neuronal damage or neurological diseases, and even brain tumors. It can be seen that oligodendrocytes have a major influence in the brain and are the objects that people urgently need to study. [0003] CN107810268A discloses a method for inducing oligodendrocyte precursor cells from human cells by direct reprogramming, comprising: culturing human ...

Claims

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Application Information

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IPC IPC(8): C12N5/079
CPCC12N5/0622C12N2500/90C12N2501/11C12N2501/135
Inventor 贺喜白乙玲玲那木拉巴特尔
Owner INNER MONGOLIA UNIVERSITY
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