Cell culture fluid, application of cell culture fluid and method of inducting DPSCs to differentiate into neuron-like cells

A dental pulp stem cell and cell culture technology, which can be applied to non-embryonic pluripotent stem cells, animal cells, nervous system cells, etc., and can solve the problems of long cycle and low differentiation efficiency.

Active Publication Date: 2015-11-25
GUANGZHOU SALIAI STEMCELL SCI & TECH CO LTD
View PDF1 Cites 3 Cited by
  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

In vitro studies have found that DPSCs are pluripotent stem cells derived from neural crest and me

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
  • Cell culture fluid, application of cell culture fluid and method of inducting DPSCs to differentiate into neuron-like cells
  • Cell culture fluid, application of cell culture fluid and method of inducting DPSCs to differentiate into neuron-like cells
  • Cell culture fluid, application of cell culture fluid and method of inducting DPSCs to differentiate into neuron-like cells

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1～5

[0059] Using 3-5 passages of dental pulp stem cells, according to 2×10 3 piece / cm 2 Seeding Density Seeding was done in six-well plates. After cultivating 4h~6h cell adherence with DMEM / F12, after adding EGF pre-induction respectively, discard the complete medium, wash the cells twice with PBS, add the cell culture fluid (containing GDNF, Shh, EGF, cAMP) provided by the present invention DMEM / F12 serum-free medium), after induction of cultured cells. Observe the differentiation of dental pulp stem cells. The pre-induction conditions and cell culture medium components nuclear induction conditions used in each embodiment are shown in Table 1:

[0060] Table 1, Examples 1-5

[0061]

[0062] After the cells of each example were pre-induced and induced, the cell morphology was observed with an electron microscope. The results showed that the cell morphology began to change after 12 hours, and changed significantly after 24 hours of induction. The refraction of the cells beg...

Embodiment 6

[0069] Extract the total proteins with cells induced in Examples 1-5 and Comparative Examples 1-3, and perform Western blot to detect the changes in the expression of Nestin and MAP2 during the induction process. Specifically, the total proteins obtained by each extraction were polymerized with 12% SDS-PAGE. Acrylamide gel electrophoresis, transfer to PVDF membrane at 80V for 1h. Block with 5% BSA at room temperature for 1 h, add anti-Nestin, MAP2 primary antibody (1:200) or β-tubulin antibody (1:200), overnight at 4°C; wash the membrane, add HRP-labeled secondary antibody (1:2000), Incubate at room temperature for 60 min; ECL emits light. The result is as Figure 5 .

[0070] The results showed that the expressions of Nestin and MAP2 increased after the dental pulp stem cells were induced to differentiate into neural-like cells in each of the examples and comparative examples, which was consistent with the expression of specific proteins in neural-like cells. The expressio...

Embodiment 7

[0072] The cells were induced for 48 hours using the protocols of Comparative Example 1 and Example 1, and the cells were collected at 24 hours, 36 hours, and 48 hours of induction to extract total protein, and the expression levels of Nestin and MAP2 were detected by the method described in Example 6. The result is as Figure 6 . The results showed that after the cells were induced for 24 hours in Example 1, the expression levels of Nestin and MAP2 in the cells were significantly higher than those in the cells induced in Comparative Example 1 for 48 hours.

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 technical field of stem cell culture, in particular to cell culture fluid, application of the cell culture fluid and a method of inducting DPSCs (Dental Pulp Stem Cells) to differentiate into neuron-like cells. The cell culture fluid provided by the invention comprises basic culture fluid, GDNFs (Glial Cell-Derived Neurotrophic Factors), Shh (Sonic hedgehog), EGFs (Epidermal Growth Factors) and cAMP (Cyclic Adenosine monophosphate). The culture fluid provided by the invention can be used for inducting the DPSCs to differentiate into the neuron-like cells. When the culture fluid is used for inducing the DPSCs, the form of the DPSCs starts to change from 12h; after the 24h from the induction, the form of the DPSCs is obviously changed; the cell refractivity starts to be enhanced; cell processes increase and grows and differentiate into the form of the neuron-like cells. Through western bolt detection, after 24h from the induction, the expression amounts of Nestin and MAP2 (Microtubule-Associated Protein 2) in the DPSCs are obviously increased. Therefore the culture fluid has the advantages of short period and high efficiency when being used for inducing the DPSCs to differentiate into the neuron-like cells.

Description

technical field [0001] The invention relates to the technical field of stem cell culture, in particular to a cell culture solution and its application and a method for inducing differentiation of dental pulp stem cells into nerve-like cells. Background technique [0002] Many clinically common chronic neurodegenerative diseases, such as: Parkinson's syndrome, Alzheimer's disease, Huntington's disease and amyotrophic lateral sclerosis. These diseases are caused by the degeneration and death of nerve cells in the central nervous system. In addition, external trauma can also cause neurological damage, leading to severe behavioral dysfunction. [0003] Nerve cells do not regenerate as quickly as epidermis and liver cells. Although stem cells in the central nervous system can also replicate and differentiate, their replication speed is extremely slow, and the ratio of differentiation into nerve cells is also very low. Most of them will differentiate into glial cells, which cann...

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
IPC IPC(8): C12N5/074C12N5/079
Inventor 陈海佳王一飞葛啸虎冯德龙马岩岩
Owner GUANGZHOU SALIAI STEMCELL SCI & TECH CO LTD
Who we serve
  • R&D Engineer
  • R&D Manager
  • IP Professional
Why Eureka
  • Industry Leading Data Capabilities
  • Powerful AI technology
  • Patent DNA Extraction
Social media
Try Eureka
PatSnap group products

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

© 2024 PatSnap. All rights reserved.Legal|Privacy policy|Modern Slavery Act Transparency Statement|Sitemap