Looking for breakthrough ideas for innovation challenges? Try Patsnap Eureka!

In vitro amplification method of human marrow pluripotent stem cells and method for directionally differentiating same into dopaminergic neuron

A technology for pluripotent stem cells and in vitro expansion, which is applied to the in vitro expansion of human bone marrow pluripotent stem cells. Human bone marrow pluripotent stem cells can be directional, efficiently and safely differentiated into dopaminergic neuron-like cells, which can solve the problem of low self-renewal ability, The limitations of clinical cell therapy and the general ability of MSC to proliferate in vitro have achieved the effect of increasing self-stability and reducing the effect of contact inhibition

Inactive Publication Date: 2011-01-05
SECOND MILITARY MEDICAL UNIV OF THE PEOPLES LIBERATION ARMY
View PDF0 Cites 3 Cited by
  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, MSC is a mesenchymal stem cell derived from bone marrow. Its cell composition is complex and its self-renewal ability is low. At the same time, MSC's ability to proliferate in vitro is average, and its subsequent application in clinical cell therapy is also limited.

Method used

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
View more

Image

Smart Image Click on the blue labels to locate them in the text.
Viewing Examples
Smart Image
  • In vitro amplification method of human marrow pluripotent stem cells and method for directionally differentiating same into dopaminergic neuron
  • In vitro amplification method of human marrow pluripotent stem cells and method for directionally differentiating same into dopaminergic neuron
  • In vitro amplification method of human marrow pluripotent stem cells and method for directionally differentiating same into dopaminergic neuron

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0023] Example 1: In vitro expansion of human bone marrow pluripotent stem cells

[0024] 1. Primary culture of bone marrow pluripotent stem cells

[0025] Using the bone marrow from clinical patients, wash it twice with PBS, use 4°C pre-cooled 0.16M Tris-NH4Cl to break the red treatment for 5 minutes, centrifuge at 1000 rpm for 5 minutes, remove the lysed red blood cells, and wash away excess Tris with PBS -NH 4 Cl, after adding a small amount of expansion medium to disperse evenly, the remaining mononuclear cells were placed at the bottom of the cell culture dish pretreated with 100ng / ml FN fibronectin. CO 2 After culturing in an incubator (saturated humidity; 95% air-5% CO2; 37°C) for 3 days, change the medium in full, wash with PBS to remove suspended unattached cells, then change the medium once every 3 days, and reach 100% confluence in about 10 days Afterwards, 0.25% Trypsin-EDTA was digested into a single-cell suspension, subcultured at 1:2, and the same medium was ...

Embodiment 2

[0047] Example 2: Directional induction of bone marrow pluripotent stem cells to differentiate into dopaminergic neurons

[0048] The bone marrow pluripotent stem cells after steps 1-2 in Example 1 were induced to differentiate into dopaminergic neurons by stage induction method. Divided into 2 phases:

[0049] The first stage: take the cells in good growth state, wash them three times with PBS, digest the cells with 0.05% Trypsin / EDTA, pipette the medium after the digestion is terminated, centrifuge the cell suspension at 1000 rpm for 5mins, remove the medium liquid, and then wash with PBS for 2 times, and pipette repeatedly with a wash tube to obtain a single-cell suspension. The cells were inoculated onto the 6-well slides treated with FN at a seeding density of 3000 cells / em 2 , basal medium + bFGF (100ng / ml) + RA (10nM) induced for 7 days. The composition of the basal medium is the same as that shown in the previous table.

[0050] The second phase: the first phase in...

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to View More

PUM

No PUM Login to View More

Abstract

The invention relates to the stem cell and cell directional induced differentiation field. Parkinson's disease (PD) can be treated using a cell transplantation therapy, and treatment difficulty comprises a proper stem cell source and a condition for stably inducing cell differentiation. Marrow is an important source of autologous stem cells; and hybrid cells in the marrow are subject to separation and purification culture to obtain pluripotent stem cells from the marrow, and the condition for differentiating the pluripotent stem cells into a dopaminergic neuron is searched, which is of great significance to clinical treatment of PD. The invention provides an in vitro amplification method of the human marrow pluripotent stem cells, wherein, various culture components are taken as culture media for cloning culture to obtain the pluripotent stem cells from the human marrow and realize in vitro stable proliferation and express a relevant pluripotent marker. The invention further provides a method for directionally, efficiently and safely differentiating the human marrow pluripotent stem cells into the dopaminergic neuron-like cells, wherein, after two induced differentiation cycles by a RA+bFGF and SHH factor staging induction method, the pluripotent stem cells from the marrow can be differentiated into the dopamine (DA) neuron-like cells in vitro.

Description

technical field [0001] The invention belongs to the technical field of biomedicine, specifically relates to the field of stem cells and cell directional induction differentiation, and relates to an in vitro expansion method of human bone marrow pluripotent stem cells, and the directional, efficient and safe differentiation of human bone marrow pluripotent stem cells into dopaminergic neuron-like cells Methods. Background technique [0002] Parkinson's disease Parkinson's Disease (PD), discovered and named by Dr. James Parkinson in 1817, is a chronic degenerative neurological disease, mostly in the elderly. Currently, in the world's population, more than 1% of the elderly over the age of 55 Suffering from Parkinson's disease (Betarbet, R., T.B. Sherer, and J.T.Grenamyre, Animal models of Parkinson's disease. Bioessays, 2002.24 (4): p.308-18. Zhen-Xin Zhang, Gwendolyn E P Zahner, Parkinson's disease in China: prevalence in Beijing, Xian, and Shanghai. Lancet 2005; 365: 595-97...

Claims

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to View More

Application Information

Patent Timeline
no application Login to View More
Patent Type & Authority Applications(China)
IPC IPC(8): C12N5/0775
Inventor 刘厚奇范力星
Owner SECOND MILITARY MEDICAL UNIV OF THE PEOPLES LIBERATION ARMY
Who we serve
  • R&D Engineer
  • R&D Manager
  • IP Professional
Why Patsnap Eureka
  • Industry Leading Data Capabilities
  • Powerful AI technology
  • Patent DNA Extraction
Social media
Patsnap Eureka Blog
Learn More
PatSnap group products

Browse by: Latest US Patents, China's latest patents, Technical Efficacy Thesaurus, Application Domain, Technology Topic, Popular Technical Reports.

© 2024 PatSnap. All rights reserved.Legal|Privacy policy|Modern Slavery Act Transparency Statement|Sitemap|About US| Contact US: help@patsnap.com