Method for inducing differentiation from mesenchymal stem cells to cartilage cells and application of mesenchymal stem cells in osteoarthritis

A technology for stem cells and chondrocytes, which is applied in the field of stem cells to achieve uniform differentiation state, good induction effect and rich sources.

Inactive Publication Date: 2013-01-30
黑龙江和泽北方生物科技有限公司
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0003] There is no attempt to induce chondrogenic differentiation of umbilical cord-derived mesenchymal stem cells in the prior art. We use umbilical cord-derived mesenchymal stem cells as

Method used

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  • Method for inducing differentiation from mesenchymal stem cells to cartilage cells and application of mesenchymal stem cells in osteoarthritis
  • Method for inducing differentiation from mesenchymal stem cells to cartilage cells and application of mesenchymal stem cells in osteoarthritis

Examples

Experimental program
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Effect test

Embodiment 1

[0018] Example 1 Induction of umbilical cord-derived mesenchymal stem cells into chondrocytes

[0019] 1. Monolayer expansion of umbilical cord-derived mesenchymal stem cells in MSC growth medium:

[0020] MSC growth medium (13.5ml DF12, 10% human serum, 20 ng / ml EGF, 5ng / ml FGF2 per 75 cm2 culture flask) was used to culture P3 generation umbilical cord-derived mesenchymal stem cells (UC-MSCs), and the cells grew After reaching 70-80% confluence, wash twice with D-Hank's washing solution, add 1.5ml trypsin-EDTA (0.25% trypsin-0.02% EDTA) for digestion, observe under the microscope, when the cells are in a spherical state, use 1.5ml of human serum was used to stop the digestion, the cells were sucked into a 50ml centrifuge tube, after mixing thoroughly, 200 microliters were taken to count and determine the number of cells, and the rest of the cell suspension was centrifuged at 1000rpm for 10min. Pour off the supernatant after centrifugation, subculture at a ratio of 1:3, cultu...

Embodiment 2

[0023] Example 2 Application of UC-MSCs induced to cartilage in osteoarthritis

[0024] The chondrogenically induced cell spheres obtained in Example 1 were digested with 3ml of 2% (w / v) type II collagenase in a centrifuge tube for half an hour, centrifuged at 1000rpm for 5min, and induced with the same method as in Example 1. The differentiation medium was resuspended and inoculated on the bottom area of ​​25 cm 2 UC-MSCs cell suspension induced by cartilage induction was made by digesting with 0.5ml trypsin-EDTA after overgrown.

[0025] Sixty healthy male Wistar rats, weighing 200±50 g, were selected. Randomly divided into (1) intra-articular injection of UC-MSCs cell suspension induced into cartilage treatment group 1; (2) intra-articular injection of UC-MSCs cell suspension induced into cartilage treatment group 2; (3) tail vein injection into cartilage-induced UC - MSCs cell suspension treatment group 3; (4) tail vein injection of cartilage-induced UC-MSCs cell suspens...

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Abstract

The invention relates to a method for inducing differentiation from mesenchymal stem cells to cartilage cells and application of the mesenchymal stem cells in osteoarthritis, and provides a method for preparing umbilical cord mesenchymal stem cells differentiated to the cartilage cells, mesenchymal stem cells differentiated to the cartilage cells and application of the prepared drug prepared by the mesenchymal stem cells in treatment of the osteoarthritis. The method comprises the following steps of: inducing and culturing the mesenchymal stem cell in a 15ml taper-bottom centrifuge tube by using a globule method; and slightly shaking the centrifuge tube for one time every 24 hours to separate cell sphere from the bottom of the tube, wherein the differentiation inducing culture medium is prepared by adding 100mu g/ml of sodium pyruvate, 10ng/ml TGFbeta3, 100nM of dexamethasone, 25mu g/ml of vitamin C, 40mu g/ml of proline and 1*ITS and 1 of premix into a high-glucose dulbecco modified eagle medium (DMEM). A non-bracket cartilage built by the method can be used for repairing damaged articular cartilages.

Description

technical field [0001] The invention relates to the technical field of stem cells, in particular to a method for preparing cartilage-induced UC-MSCs and its application in osteoarthritis. Background technique [0002] Articular cartilage is a kind of load-bearing connective tissue, which is often damaged by tumors, sports, degeneration or senile diseases; however, the ability of articular cartilage to repair itself is limited, which makes it difficult to heal cartilage defects clinically. In recent years, a variety of methods for the treatment of cartilage defects have emerged, including autologous chondrocyte transplantation, microfractures, and mosaicplasty, but each of these methods has its limitations. In recent years, tissue engineered cartilage has become a new hotspot in cartilage repair research, and mesenchymal stem cells (MSCs) are currently the most promising seed cells. MSCs are a type of pluripotent stem cells derived from the mesoderm and ectoderm in early dev...

Claims

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Application Information

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IPC IPC(8): C12N5/077C12N5/0775A61K35/44A61P19/02A61P29/00A61K35/28
Inventor 刘拥军王立华许放董方
Owner 黑龙江和泽北方生物科技有限公司
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