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Use of protocatechuic acid in accelerating nerve stem cell multiplication in vitro and inducement differentiation

A technology of neural stem cells and protocatechuic acid, applied in medical preparations containing active ingredients, nervous system diseases, biochemical equipment and methods, etc.

Inactive Publication Date: 2009-11-18
DALIAN UNIV OF TECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] However, there have been no domestic and foreign research reports on protocatechuic acid in promoting the proliferation and inducing differentiation of neural stem cells in vitro.

Method used

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  • Use of protocatechuic acid in accelerating nerve stem cell multiplication in vitro and inducement differentiation
  • Use of protocatechuic acid in accelerating nerve stem cell multiplication in vitro and inducement differentiation
  • Use of protocatechuic acid in accelerating nerve stem cell multiplication in vitro and inducement differentiation

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0024]Example 1 Isolation, cultivation and identification of neural stem cells in vitro

[0025] (1) Isolation and culture of neural stem cells derived from midbrain

[0026] E14d mice were killed by necking, the embryos were taken out by aseptic operation, the midbrain tissue of the embryos was separated, digested into single cells, counted with trypan blue, and counted by 2×10 5 cells / mL inoculated in a T25 culture flask at 37°C, 5% CO 2 For suspension culture, the stem cell culture medium is DMEM / F12 (1:1), 2% B27, bFGF (10 ng / mL), EGF (20 ng / mL). Change the medium in half every 3 to 4 days, and observe the number and size of the growing suspended cell spheres after one week of culture. After collecting the neurospheres and mechanically blowing them into a single-cell suspension, they were placed under the same conditions to continue culturing, and whether to pass passage was determined depending on the growth status.

[0027] (2) Continuous passage and induced different...

Embodiment 2

[0032] Example 2 Proliferation-promoting effect of protocatechuic acid on neural stem cells

[0033] (1) Experimental design and drug treatment

[0034] The experiment was divided into five groups, namely the control group, 6×10 -6 mol / L protocatechuic acid group, 6×10 -5 mol / L protocatechuic acid group, 6×10 -4 mol / L protocatechuic acid group and 1.2×10 -3 mol / L protocatechuic acid group. Collect the second-generation neural stem cells from the primary culture, and use 2×10 5 Cells / mL were inoculated in 24-well plates, 500 μL per well, and 6 replicate wells were set up for each treatment group. The number of neurospheres and the survival rate of cells were observed after 4 days and 7 days of culture respectively.

[0035] (2) Detection method of neural stem cell proliferation

[0036] ① Neurosphere counting method: after the primary cultured second-generation neural stem cells were cultured under the action of different concentrations of protocatechuic acid for 4 days a...

Embodiment 3

[0040] Example 3 Ability of Protocatechuic Acid to Induce Neural Stem Cells to Differentiate into Functional Neurons

[0041] (1) Protocatechuic acid induced differentiation of neural stem cells

[0042] The second-generation neural stem cell spheres cultured at the primary stage were mixed with 6×10 -5 The neural stem cell spheres were collected after being treated with mol / L protocatechuic acid for 7 days, and blown into a single-cell suspension in the differentiation medium (DMEM / F12+10% fetal bovine serum) respectively. 5 Inoculate the poly-L-lysine-treated slides in a 24-well plate at a concentration of 1 / ml, 500 μL per well, and set six replicate wells for each treatment group. After 24 hours, it can be seen that the cells are all attached to the wall, and different concentrations of protocatechuic acid (6 × 10 -5 , 6×10 -4 , 1.2×10 -3 mol / L) after treatment, the culture plate was placed in saturated humidity, 5% CO 2 , 37 ° C incubator to continue culturing for 7 d...

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Abstract

The invention belonging to the cell biology and pharmacology technical field relates to the purpose of a Chinese traditional medicine monomeric compound, in particularly to the new purpose of a protocatechuic acid in the aspect of promoting the external proliferation and induction-differentiation of a nerve stem cell. The invention is characterized in the application of producing medicine to treat the neurodegenerative diseases by nerve stem cell transplantation, in the application of producing neurenergen and cell differential agent for restoring damaged nerve tissues and reconstructing nerve functions, and in the application of constructing high efficient medical screening model to initially screen and appraise the efficacy of medicine. The invention confirms the new phamacoloic activity of the protocatechuic acid, providing material basis with regard to prevention effect of the Chinese traditional medicine, and providing reference for the development of new medicine with independent intellectual property rights.

Description

technical field [0001] The invention belongs to the technical field of cell biology and pharmacology, and relates to the use of a traditional Chinese medicine monomer compound, in particular to the new use of protocatechuic acid in promoting the proliferation and inducing differentiation of neural stem cells in vitro. Background technique [0002] Alpinia oxyphylla MIQUEL is a plant of the genus Alpinia Roxb. in the Zingiberaceae family. It has been a commonly used medicinal material in Chinese medicine since ancient times. Modern studies have shown that nootropic contains special chemical components and significant biological activity, and has a wide range of pharmacological activities such as cardiotonic, anticancer, antiallergic and neuroprotective. The inventor screened and proved for the first time that protocatechuic acid is the main neuroactive component in Yizhi through the established in vitro nerve cell injury model (An LJ et al., Food Chem Toxicol, 2006, 44(3 ): ...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N5/08A61K35/30A61P25/00
Inventor 关水刘天庆李香琴马学虎崔占峰
Owner DALIAN UNIV OF TECH
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