Method for separating and culturing mesenchymal stem cells from umbilical cord and inducing and differentiating mesenchymal stem cells into cartilage cells

A technology of mesenchymal stem cells and chondrocytes, applied in animal cells, vertebrate cells, bone/connective tissue cells, etc., can solve the problems of low conversion rate and high cost, achieve great application prospects and facilitate large-scale production

Active Publication Date: 2015-10-21
SHENZHEN ZHONGJIHENGRUN INVESTMENT CO LTD +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0007] In order to overcome the current problems of high cost and low transformation rate of isolating and culturing mesenchymal stem cells from umbilical cord and inducing differentiation of mesenchymal stem

Method used

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  • Method for separating and culturing mesenchymal stem cells from umbilical cord and inducing and differentiating mesenchymal stem cells into cartilage cells
  • Method for separating and culturing mesenchymal stem cells from umbilical cord and inducing and differentiating mesenchymal stem cells into cartilage cells
  • Method for separating and culturing mesenchymal stem cells from umbilical cord and inducing and differentiating mesenchymal stem cells into cartilage cells

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0046] Example 1. Isolation and cultivation of mesenchymal stem from umbilical cord according to the method described in the present invention cell

[0047] Umbilical cord and umbilical cord blood source: 24 umbilical cords from healthy puerpera, which were effectively frozen.

[0048] Experimental group 1:

[0049] Isolate and culture mesenchymal stem cells from umbilical cord, the specific isolation and culture methods are as follows:

[0050] (1) Umbilical cord treatment:

[0051] Choose the umbilical cord of a healthy parturient, and wash the outer surface of the umbilical cord with double anti-saline to remove the clot on the surface. Replace the container, soak the surface-treated umbilical cord in double-antibody saline for 15 minutes; take out the umbilical cord, replace the container, and repeat the immersion in double-antibody saline twice, each time for 15 minutes. Replace the sterile container, cut a 5cm long section, replace the sterile container again, c...

Embodiment 2

[0120] Example 2: Differentiation of umbilical cord-derived mesenchymal stem cells to chondrocytes of the present invention

[0121] Differentiation of mesenchymal stem cells to chondrocytes obtained by using the method described in Experimental Group 1 in Example 1 in this example

[0122] Specifically, experimental group 1 of the present embodiment is as follows:

[0123] The 3rd generation (P3) cells were inoculated into a six-well culture plate at a certain cell density, and a sterilized cover glass was placed in the six-well plate. Plates were placed in 5% CO 2 , 80% saturated humidity, cultivated in a 37°C incubator, and change the medium 2-3 times a week as needed.

[0124] The components of the induced transformation medium include: DMEM / F12 medium containing 5% fetal bovine serum, 10ng / ml transforming growth factor βⅠ, 6.25μg / ml insulin, 100μmol / ml dexamethasone, 1μmol / ml sodium pyruvate and 6.25 μg / ml transferrin.

[0125] In fact, the parameters of experimenta...

Embodiment 3

[0181] Embodiment 3: adopt the experimental group 12,14 in the embodiment of the present invention 2 to prepare Obtained mesenchymal stem cells for osteoarthritis

[0182] The method described in the experimental group 12 and 14 in the embodiment 2 of the present invention will adopt the method of the experimental group 1 in the embodiment 1 of the present invention to separate and cultivate the third generation cells of the mesenchymal stem cells in the embodiment 2 In the induction differentiation medium in experimental groups 12 and 14, 1×10 8 / ml of mesenchymal stem cell-medium mixture. Take 1ml of cell suspension (containing 1×10 8 / ml of mesenchymal stem cells), and directly transfer them into the lyophilized sodium hyaluronate structure with a certain three-dimensional shape, so that it forms a homogeneous mixture.

[0183] (1) Patients with mild osteoarthritis: After local anesthesia, inject directly into the joint cavity with sterile injection.

[0184] (2) Pat...

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Abstract

The invention provides a method for separating and culturing mesenchymal stem cells from an umbilical cord and inducing and differentiating the mesenchymal stem cells into cartilage cells. The method comprises the steps that the umbilical cord is treated; the cells are separated from the umbilical cord and umbilical vessels and subjected to primary cell culture; cell subculture is conducted, and the mesenchymal stem cells are obtained; the mesenchymal stem cells are induced and differentiated into the cartilage cells, wherein in the step that the cells are separated from the umbilical cord and the umbilical vessels, the umbilical cord and the umbilical vessels are digested through digestive juice. By means of the method, the mesenchymal stem cells can be separated from the umbilical cord and cultured, and the obtained mesenchymal stem cells are subjected to further induced transformation; the method can be used for mass production of the mesenchymal stem cells and has importance significance in treatment of injured tissue which can hardly regenerate.

Description

【Technical field】 [0001] The invention relates to the technical field of mesenchymal stem cells, in particular to a method for isolating and culturing mesenchymal stem cells from umbilical cords and inducing differentiation to chondrocytes. 【Background technique】 [0002] Osteoarthritis (OA), known as the "undead cancer", is a chronic progressive bone and joint disease characterized by articular cartilage degeneration and periarticular bone hyperplasia, whose incidence increases with age , mainly infringes on articular cartilage, bone and synovial tissues, leading to joint pain, deformity and dysfunction. At present, the treatment of osteoarthritis mainly includes: exercise and life guidance, physical therapy, drug therapy (non-steroidal anti-inflammatory drugs, oral glucosamine, local injection of sodium hyaluronate and hormones, etc.), surgical joint replacement, etc. It has improved the quality of life of patients to a certain extent, but there is still no ideal treatmen...

Claims

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Application Information

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IPC IPC(8): C12N5/0775C12N5/077
Inventor 黄慈波
Owner SHENZHEN ZHONGJIHENGRUN INVESTMENT CO LTD
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