Chondrogenic differentiation culture medium and application thereof

A differentiation medium and chondrogenic technology, which is applied in the field of cell culture, can solve the problems of long differentiation time, low biological safety, and low differentiation efficiency, and achieve the effect of high differentiation efficiency, clear ingredients, and increased dosage

Active Publication Date: 2020-11-27
苏州依科赛生物科技股份有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] The first purpose of the present invention is to provide safer and more efficient chondrogenic differentiation of stem cells in view of the deficiencies of the prior art and the existing chondrogenic differentiation medium with low differentiation efficiency, long differentiation time and low biological safety serum-free medium

Method used

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  • Chondrogenic differentiation culture medium and application thereof
  • Chondrogenic differentiation culture medium and application thereof
  • Chondrogenic differentiation culture medium and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0030] The serum-free medium for chondrogenic differentiation of mesenchymal stem cells provided in this example consists of the following components: basal medium and additional components.

[0031] (1) The basal medium is marked as M1 and prepared according to the following scheme:

[0032]

[0033]

[0034]

[0035] The various components mentioned above can be purchased from reagent companies such as Sigma and Aladdin. Each component was dissolved in 1 L of water for injection and filtered through a 0.22 micron filter membrane to obtain the basal medium M1.

[0036] (2) The added component is recorded as S2 component, and the material composition of S2 component is as follows:

[0037]

[0038] In this example, dexamethasone and linoleic acid can be dissolved in absolute ethanol, Calcitonin and recombinant human insulin can be dissolved in 0.05M hydrochloric acid solution, and other components can be dissolved in water for injection. After the above components ...

Embodiment 2

[0043] This example provides verification of the effect of the serum-free medium for chondrogenic differentiation of mesenchymal stem cells in Example 1. In the experiments of this example, human bone marrow mesenchymal stem cells were used. The cells were derived from the ATCC standard cell bank (PCS-500-012, Lot 70011720, ATCC). These cells were used in all the experiments below.

[0044] 1. Experimental method

[0045] 1. Culture of human bone marrow mesenchymal stem cells

[0046] Serum-free medium for mesenchymal stem cells (Excell Bio, ME000-N023), resuscitated human bone marrow mesenchymal stem cells (ATCC, PCS-500-012, Lot 70011720, Passage 5), seeded in 100mm cell culture at a density of 8000 / cm2 Dish.

[0047]Add mesenchymal stem cell serum-free medium (Excell Bio, ME000-N023) for culture, the amount added is about 15ml, and the medium is replaced every 48h, and the bone marrow mesenchymal stem cells grow to about 85% abundance (about 72~ Cells were harvested at 9...

Embodiment 3

[0063] This example provides the effect test of different ratios. According to the scheme of Example 1 of this example, the basal medium (M1) and the added components (S2) are prepared, and the basal medium and the added components are used for mesenchymal stem cell growth The preparation of adipogenic differentiation medium, and the effect of chondrogenic induction and differentiation of each ratio of medium was tested. Alcian blue staining was performed 14 days after induction of differentiation.

[0064] The mesenchymal stem cells and culture method used in this example are the same as those in Example 2.

[0065] 1) Medium group

[0066]

[0067]

[0068] Prepare M1-M5 medium according to the ratio in the above table,

[0069] 2) Cell culture

[0070] Bone marrow mesenchymal stem cells were cultured according to the same scheme as in Example 2, and the mesenchymal stem cells collected after passage were counted, centrifuged at 300g for 5 minutes, and the cells wer...

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PUM

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Abstract

The invention relates to a chondrogenic differentiation culture medium and application thereof. The chondrogenic differentiation culture medium is composed of a basic culture medium and additive components, and comprises the components of an inorganic salt component, an amino acid component, a vitamin component, a microelement component, carbohydrate, TGF-[beta]3, BMP-6, BMP-2, human serum albumin, dexamethasone, magnesium ascorbyl phosphate, recombinant human insulin, recombinant human transferrin, selenic acid and linoleic acid. The invention further provides application of the chondrogenicserum-free differentiation culture medium for mesenchymal stem cells. The chondrogenic differentiation culture medium is definite in component, free of animal-derived components and serum and safer, and the components can efficiently induce differentiation of the mesenchymal stem cells to chondrocytes under the synergistic effect.

Description

technical field [0001] The invention belongs to the technical field of cell culture, in particular to a chondrogenic differentiation medium. Background technique [0002] Stem cells are a class of cells with high self-renewal and multilineage differentiation potential. According to the stage of development, it can be divided into embryonic stem cells and adult stem cells. Stem cell research is currently one of the hot research directions in the biomedical industry. Since embryonic stem cells involve ethical and moral issues, the research and application of ready-made somatic stem cells has become the mainstream. As a type of adult stem cells, mesenchymal stem cells have the advantages of rich sources, high self-proliferation, multi-directional differentiation, and low immunogenicity, and have become ideal seed cells for cell therapy. [0003] Due to the particularity of cell therapy, more and more attention has been paid to the identification of mesenchymal stem cells used...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N5/077
CPCC12N5/0655C12N2500/14C12N2500/20C12N2500/16C12N2500/12C12N2500/22C12N2500/34C12N2500/30C12N2500/32C12N2500/46C12N2500/38C12N2500/40C12N2501/15C12N2501/155C12N2501/34C12N2500/25C12N2500/36C12N2501/998C12N2501/39C12N2506/1384C12N2506/1353C12N2506/1361C12N2506/1369C12N2506/1392C12N2500/90
Inventor 于宝利李其雷陈旭陈刚杨建国孙芳
Owner 苏州依科赛生物科技股份有限公司
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