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Culture method for inducing adipose tissue-derived stromal cells to differentiate to chondrocyte

A technology of mesenchymal stem cells and chondrocytes, applied in the direction of cell culture active agents, biochemical equipment and methods, culture process, etc., can solve the problem of difficult self-healing of injuries, achieve short induction time, good experimental repeatability, osteoblast The effect of high differentiation rate

Inactive Publication Date: 2017-01-25
中卫华医(北京)生物科技有限公司 +1
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Problems solved by technology

Because cartilage tissue has no blood supply and can only rely on joint fluid to provide nutrition, once cartilage tissue is damaged, it is difficult to activate progenitor cells in the blood or bone marrow to enter the defect to repair, and the damage is difficult to heal itself

Method used

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  • Culture method for inducing adipose tissue-derived stromal cells to differentiate to chondrocyte
  • Culture method for inducing adipose tissue-derived stromal cells to differentiate to chondrocyte
  • Culture method for inducing adipose tissue-derived stromal cells to differentiate to chondrocyte

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Embodiment Construction

[0024] The steps of the culture method for inducing the differentiation of adipose-derived mesenchymal stem cells into chondrocytes of the present invention include:

[0025] 1) Separation of primary cells of adipose-derived mesenchymal stem cells: the adipose tissue derived from liposuction was washed to remove blood cells and a small amount of mixed fibrous connective tissue, and at the same time, the fat was tested on a plate to ensure that the fat was not polluted before the experiment; after centrifugation The adipose tissue is in the lower layer. After removing the upper layer of blood, disperse the fat in the lower layer of the centrifuge tube, add 2 times the volume of type I collagenase and trypLETM digestive enzyme solution mixture, digest at 37°C for 30-45 minutes, stop the enzymatic hydrolysis, and discard the upper layer by centrifugation supernatant; and filter with a sieve to obtain adipose-derived mesenchymal stem cell suspension;

[0026]2), Adipose-derived me...

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Abstract

The invention relates to a culture method for inducing adipose tissue-derived stromal cells to differentiate to chondrocyte, and aims to solve the problem that the prior art is low in differentiation rate. The culture method comprises the following steps: 1) performing separation of primary cells of adipose tissue-derived stromal cells; 2) performing amplification and passage of the adipose tissue-derived stromal cells; and 3) performing differentiation culture on P5-generation adipose-derived stem cells to chondrocyte, namely, adding the adipose tissue-derived stromal cells into a condition culture medium, namely, an adipose tissue-derived stromal cell chondrocyte differentiation culture medium, after P5 generation of passage, performing chondrocyte differentiation culture, replacing the medium of the cells every 3 days, observing the morphological change of the cells, and after 16 days of induction, performing Alcian blue dyeing, and identifying the chondrocyte differentiation situation of the adipose tissue-derived stromal cells. The result of Alcian blue dyeing identification shows that the chondrocyte can be formed, and the Alcian blue dyeing is positive. The culture method has the characteristics of being simple and feasible, short in induction time as the induction culture medium is a serum-free culture system, good in test repeatability and high in osteoblast differentiation rate.

Description

technical field [0001] The invention relates to a method for inducing differentiation of stem cells, in particular to a culture method for inducing the differentiation of adipose mesenchymal stem cells into chondrocytes. Background technique [0002] Articular cartilage is a special form of connective tissue, which belongs to hyaline cartilage and is mainly composed of chondrocytes and extracellular matrix (ECM). . Since cartilage tissue has no blood supply and can only rely on joint fluid to provide nutrition, once cartilage tissue is damaged, it is difficult to activate progenitor cells in the blood or bone marrow to enter the defect to repair it, and the damage is difficult to heal itself. Traditional articular cartilage repair methods mainly promote cartilage healing by mobilizing the intrinsic healing potential of chondrocytes or repairing cartilage defects by promoting chondrocyte regeneration. Mesenchymal stem cells (mesenchymal stem cells, MSCs) MSCs are derived fr...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N5/077C12N5/0775
CPCC12N5/0655C12N5/0667C12N2500/24C12N2500/30C12N2500/34C12N2500/38C12N2501/15C12N2506/1384
Inventor 李亚平王晶谷广其邱丽媛房芳熊开宇何辉吴志宏赵进军何文丽
Owner 中卫华医(北京)生物科技有限公司
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