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Human mesenchymal stem cell cartilage-formation induced differentiation culture medium and preparation method

A technology of mesenchymal stem cells and induced differentiation, applied in the field of culture medium, can solve the problems of chondrogenic induction efficiency and specificity, long induction time, etc., and achieve the effects of high degree of induced differentiation, simple preparation method, and convenient use

Inactive Publication Date: 2018-09-14
安徽瑞杰赛尔生物科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] In order to solve the problems in the prior art that the efficiency and specificity of chondrogenic induction are not ideal and the induction time is long, the present invention provides a medium capable of efficiently and stably inducing chondrogenic differentiation of mesenchymal stem cells derived from various human tissues and its preparation method

Method used

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  • Human mesenchymal stem cell cartilage-formation induced differentiation culture medium and preparation method
  • Human mesenchymal stem cell cartilage-formation induced differentiation culture medium and preparation method
  • Human mesenchymal stem cell cartilage-formation induced differentiation culture medium and preparation method

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0043] Example 1: Formula screening of human mesenchymal stem cell chondrogenic differentiation medium

[0044] The inventors of the present invention screened the components of the human mesenchymal stem cell ru-induced differentiation medium through a large number of experiments. Human mesenchymal stem cells were cultured and expanded in low-sugar DMEM medium containing 10% FBS. Trypsinize the cells, resuspend the cells with the chondrogenic differentiation medium described in the patent claims, the concentration is 2X10 7 cells / mL; add 10 μL cell suspension dropwise to a six-well plate, 5 drops per well; place the cells in the incubator for 2 hours, and add differentiation-inducing medium 1, differentiation-inducing medium 2, and differentiation-inducing medium to each well. Medium 3, and induction differentiation medium 4, the medium was changed every 3 days. At 2 and 3 weeks of induction, the expression levels of chondrocyte-related genes CollagenII and Aggrecan were de...

Embodiment 2

[0045] Example 2: Preparation of human mesenchymal stem cell chondrogenic differentiation medium

[0046] Human mesenchymal stem cell chondrogenic differentiation medium, including HG-MEM medium, and the following components and their concentrations:

[0047] Fetal bovine serum (FBS)

10% (volume percentage)

sodium pyruvate

100mg / ml

ascorbic acid

50μg / ml

proline

20μg / ml

Dexamethasone

100nM

ITS

1% (volume percentage)

TGF-β 3

5ng / ml

Insulin Growth Factor-2 (IGF-2)

20nM

[0048] Preparation method: Add FBS, sodium pyruvate mother solution, ascorbic acid mother solution, proline mother solution, dexamethasone mother solution, ITS mother solution, TGF-β to HG-MEM medium according to the above concentrations 3 Mix the mother liquor and the IGF-2 mother liquor evenly, and filter and sterilize with a 0.22 μm filter membrane.

[0049] Preparation of ascorbic acid mother solution: Take 50 mg...

Embodiment 3

[0050] Example 3: Human mesenchymal stem cell chondrogenic differentiation medium

[0051] Human mesenchymal stem cell chondrogenic differentiation medium, including HG-MEM medium, and the following components and their concentrations:

[0052]

[0053]

[0054] Preparation method: similar to the preparation method provided in Example 2.

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Abstract

The invention discloses a human mesenchymal stem cell cartilage-formation induced differentiation culture medium. The human mesenchymal stem cell cartilage-formation induced differentiation culture medium is characterized by being prepared from the following components: an alpha-MEM / HG-DMEM culture medium, fetal bovine serum (FBS) with the volume percent of 5 to 50 percent, sodium pyruvate with the concentration of 50 to 200mg / ml, ascorbic acid with the concentration of 20 to 200mu g / ml, proline with the concentration of 10 to 100mu g / ml, dexamethasone with the volume of 50 to 500nM, ITS withthe volume percent of 0.5 to 20 percent, TGF (Transforming Growth Factor)-beta with the concentration of 0.1 to 20ng / ml and an insulin growth factor-2 with the volume of 0.5 to 50nM. The human mesenchymal stem cell cartilage-formation induced differentiation culture medium is prepared through the following steps: disinfecting a culture dish; adding a source culture medium; mixing the components; adding FBS, a sodium pyruvate mother solution, an ascorbic acid mother solution, a proline mother solution, a dexamethasone mother solution, an ITS mother solution, a TGF-beta3 mother solution and an insulin growth factor-2 mother solution according to the concentration, and uniformly mixing; filtering and sterilizing; filtering and sterilizing a mixed culture solution by utilizing a 0.22mu m filtering membrane. The human mesenchymal stem cell cartilage-formation induced differentiation culture medium has a rapid induction speed, and has a better induced differentiation effect on differentiation raw materials with different sources, such as human bone mesenchymal stem cells, human umbilical cord mesenchymal stem cells and human adipose mesenchymal stem cells; a preparation method of the culture medium is simple, the culture medium is convenient to use and a process is stable.

Description

technical field [0001] The invention relates to the technical field of medium, in particular to a human mesenchymal stem cell chondrogenic differentiation medium and a preparation method thereof. Background technique [0002] Mesenchymal stem cells exist in various tissues such as bone marrow, umbilical cord, fat, etc., and can be collected by separation and expanded and cultured in large quantities in vitro. Studies have proved that mesenchymal stem cells, as a type of pluripotent stem cells, have good self-renewal and Multidirectional differentiation potential, induced by specific conditions, can differentiate into various types of tissue cells such as fat, bone, cartilage, muscle, liver, nerve, skin, etc. Based on this characteristic, mesenchymal stem cells have been tried to be applied to various tissues and organ function damage repair research, among them, cartilage tissue reconstruction and repair research based on the chondrogenic differentiation characteristics of m...

Claims

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Application Information

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IPC IPC(8): C12N5/077C12N5/0775
CPCC12N5/0655C12N2500/25C12N2500/30C12N2500/32C12N2500/38C12N2501/105C12N2501/15C12N2501/39C12N2506/1346C12N2506/1353C12N2506/1384C12N2506/1392
Inventor 罗乐朱灏
Owner 安徽瑞杰赛尔生物科技有限公司
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