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A method for promoting osteogenic differentiation of bone marrow mesenchymal stem cells

A bone marrow mesenchymal and osteogenic differentiation technology, applied in the field of stem cells, can solve the problems of low induction differentiation efficiency, long time, and reduced safety, and achieve the effect of shortening the osteogenic differentiation time and improving efficiency

Inactive Publication Date: 2021-10-26
深圳市旷逸生物科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] In the process of induction towards osteogenesis, it is often necessary to add exogenous serum such as fetal bovine serum, which reduces the safety, and there are problems such as low induction differentiation efficiency and long time, which cause inconvenience to clinical application.

Method used

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  • A method for promoting osteogenic differentiation of bone marrow mesenchymal stem cells
  • A method for promoting osteogenic differentiation of bone marrow mesenchymal stem cells
  • A method for promoting osteogenic differentiation of bone marrow mesenchymal stem cells

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Experimental program
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Effect test

Embodiment 1

[0020] A method for promoting osteogenic differentiation of bone marrow mesenchymal stem cells, comprising the following steps:

[0021] (1) Human bone marrow mesenchymal stem cells were inoculated in the basal medium DMEM / F12 in a 12-well culture plate at a seeding density of 1×10 4 cells / mL, at 37°C, 5% CO 2 cultured in an incubator, and when the cell confluency reached 80%, it was digested and subcultured with 0.25% trypsin, and the ratio of subculture was 1:3;

[0022] (2) Inoculate the third-generation bone marrow mesenchymal stem cells obtained from the subculture in the above step (1) on the differentiation medium of a 6-well culture plate, at 37°C, 5% CO 2 Induced differentiation in an incubator with a seeding density of 2×10 5 cells / mL, change the solution every other day;

[0023] The above differentiation medium includes DMEM / F12 medium, insulin 20 μg / mL, dexamethasone 8 ng / mL, ascorbic acid 45 μg / mL, sulfobutyl-β-cyclodextrin 20 μg / mL, betaine 50 μg / mL, indium a...

Embodiment 2

[0025] A method for promoting osteogenic differentiation of bone marrow mesenchymal stem cells, comprising the following steps:

[0026] (1) Human bone marrow mesenchymal stem cells were inoculated in the basal medium DMEM / F12 in a 12-well culture plate at a seeding density of 1.5×10 4 cells / mL, at 37°C, 5% CO 2 cultured in an incubator, and when the cell confluency reached 85%, it was digested and subcultured with 0.25% trypsin, and the subculture ratio was 1:4;

[0027] (2) Inoculate the fourth-generation bone marrow mesenchymal stem cells obtained from the subculture in the above step (1) on the differentiation medium of a 6-well culture plate, and inoculate at 37°C, 5% CO 2 Induced differentiation in an incubator with a seeding density of 3×10 5 cells / mL, change the solution every other day;

[0028] The above differentiation medium includes DMEM / F12 medium, insulin 18 μg / mL, dexamethasone 5 ng / mL, ascorbic acid 40 μg / mL, sulfobutyl-β-cyclodextrin 15 μg / mL, betaine 45 μ...

Embodiment 3

[0030] A method for promoting osteogenic differentiation of bone marrow mesenchymal stem cells, comprising the following steps:

[0031] (1) Human bone marrow mesenchymal stem cells were inoculated in the basal medium DMEM / F12 in a 12-well culture plate at a seeding density of 2×10 4 cells / mL, at 37°C, 5% CO 2 cultured in an incubator, and when the cell confluency reached 90%, it was digested and subcultured with 0.25% trypsin, and the subculture ratio was 1:4;

[0032] (2) Inoculate the fifth-generation bone marrow mesenchymal stem cells obtained from the subculture in the above step (1) on the differentiation medium of a 6-well culture plate, and inoculate at 37°C, 5% CO 2 Differentiation was induced in an incubator with a seeding density of 4×10 5 cells / mL, change the solution every other day;

[0033] The above differentiation medium includes DMEM / F12 medium, insulin 22 μg / mL, dexamethasone 10 ng / mL, ascorbic acid 50 μg / mL, sulfobutyl-β-cyclodextrin 23 μg / mL, betaine 52...

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Abstract

The invention discloses a method for promoting osteogenic differentiation of bone marrow mesenchymal stem cells, comprising the following steps: (1) taking human bone marrow mesenchymal stem cells and culturing them in DMEM / F12 medium until the degree of cell fusion reaches 80-90 %, digested and passaged with 0.25% trypsin; (2) inoculated the bone marrow mesenchymal stem cells obtained from the subculture in the above step (1) on the differentiation medium to induce differentiation; the differentiation medium included DMEM / F12 medium, Insulin 18‑22 μg / mL, dexamethasone 5‑10 ng / mL, ascorbic acid 40‑50 μg / mL, sulfobutyl‑β‑cyclodextrin 15‑23 μg / mL, betaine 45‑52 μg / mL, indium acetate 10‑ 15ng / mL, glutathione 50‑60μg / mL. In the present invention, the bone marrow mesenchymal stem cells obtained by subculture are inoculated on the differentiation medium, and sulfobutyl-β-cyclodextrin, betaine, and indium acetate are added to the differentiation medium to act synergistically to effectively improve the growth rate of the bone marrow mesenchymal stem cells. The efficiency of osteogenic differentiation and the shortening of osteogenic differentiation time provide a guarantee for its clinical application in tissue engineering bone construction.

Description

technical field [0001] The invention relates to the field of stem cells, in particular to a method for promoting osteogenic differentiation of bone marrow mesenchymal stem cells. Background technique [0002] Stem cells are a type of cells with self-renewal ability and multi-directional differentiation potential. They are the first cells in the process of human newborn cells from proliferation, migration, differentiation to maturity. They are the source of traditional Chinese medicine for cell therapy and tissue engineering seed cells. The common Mesenchymal stem cells are derived from bone marrow, peripheral blood, umbilical cord, fat, etc. In stem cell biology, it is of great significance to explore the relationship between cell proliferation and differentiation. [0003] Bone marrow mesenchymal stem cells are a type of mesenchymal stem cells that exist in bone marrow tissue and have self-renewal, strong proliferation ability, and multilineage differentiation ability. Bo...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N5/077C12N5/0775
CPCC12N5/0654C12N5/0663C12N2500/38C12N2501/33C12N2501/39C12N2501/90C12N2501/999C12N2509/00
Inventor 沈艳于梦梦马超群
Owner 深圳市旷逸生物科技有限公司
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