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Human placenta chorion mesenchymal stem cell culture system

A technology of stem cells and culture system, applied in the field of biomedicine, can solve the problems of high chance of virus infection, unacceptable, decreased differentiation ability and cell number, etc., to achieve the effect of benefiting clinical application and improving utilization rate

Active Publication Date: 2017-10-10
GUANGZHOU WOMEN AND CHILDRENS MEDICAL CENTER
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0003] At present, the mesenchymal stem cells used are mainly bone marrow mesenchymal stem cells, and their source is convenient and fast. As the age increases, the expansion, differentiation ability and cell number show an obvious downward trend; 2. There are ethical issues and the source is limited; 3. The donor feels uncomfortable and is not easy to accept; 4. The chance of virus infection is high; 5. Allogeneic transplantation strong immune rejection
my country started early in the basic research of mesenchymal stem cells, and some basic research is still at the international leading level, but the level of clinical transformation in my country is insufficient, and there are no relevant clinical cell products

Method used

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  • Human placenta chorion mesenchymal stem cell culture system
  • Human placenta chorion mesenchymal stem cell culture system
  • Human placenta chorion mesenchymal stem cell culture system

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0041] Example 1: Human placental chorionic mesenchymal stem cell culture

[0042]After the human placental chorionic tissue was treated, the primary culture was performed. After 7 days of static culture, the medium was completely replaced. The tissue blocks, culture medium and washing fluid in the primary culture flask were re-inoculated after centrifugation, and then cultured again. After 7 days of static culture, the full amount was replaced. culture medium; re-inoculate the tissue pieces, culture fluid and washing fluid in the culture flask after centrifugation, carry out the third culture, and replace the culture medium in full after 7 days of static culture; when the confluence of the three cultured cells reaches 80-90%, use Trypsin digestion and passage to obtain human placental chorionic mesenchymal stem cells. The number of cells obtained from the primary culture, the re-culture and the third culture and the harvest time are shown in Table 1.

[0043] cultur...

Embodiment 4

[0049] Example 4: Passaging, cryopreservation and recovery of human placental chorionic mesenchymal stem cells

[0050] When the cells in the flask reach 80-90% confluence, remove the medium, rinse the flask twice with normal saline, add 3ml of trypsin, place it in a 37°C incubator to digest for about 3-5 minutes, and add the medium 15ml, stop trypsin digestion, and take 100ul for counting cell viability and cell number. After centrifugation (1500r / min, 5min), the cells were subcultured at a ratio of 1:2 and placed in a saturated humidity, 37°C, and a volume fraction of 5% CO. 2 In the incubator, generally 80-90% confluent state can be reached in 2-3 days, and it needs to be passaged again. When the passage that needs to be cryopreserved is reached, according to the passage of digestion, according to the number of cells, add culture medium and cell cryopreservation solution to make the cell concentration 1 × 10 6 / ml, the cell cryopreservation solution is 10%. After mixing,...

Embodiment 5

[0051] Example 5: Identification of biological characteristics of human placental chorionic mesenchymal stem cells and comparison of three cultured cells

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Abstract

The invention relates to a human placenta chorion mesenchymal stem cell culture system which includes the steps: treating human placenta chorion tissues, performing primary culture and replacing a culture medium 7 days after; centrifuging tissue blocks, culture solution and flushing fluids in a primary culture bottle, performing renewal vaccination and secondary culture, and replacing the culture medium 7 days after; centrifuging tissue blocks, culture solution and flushing fluids in a secondary culture bottle, performing renewal vaccination and third culture, and replacing the culture medium 7 days after; performing digestive passage by pancreatin after the fusion degree of cells cultured for three times reaches 80-90% to obtain human placenta chorion mesenchymal stem cells. The system is simple and convenient to operate, and the number of the P0-generation human placenta chorion mesenchymal stem cells obtained by culture for three times doubles that of cells in primary culture.

Description

technical field [0001] The invention belongs to the technical field of biomedicine, and in particular relates to a human placental chorionic mesenchymal stem cell culture system Background technique [0002] Mesenchymal stem cells are pluripotent adult stem cells that commonly exist in different tissues. At present, mesenchymal stem cells have been isolated and cultured from bone marrow, adipose, dental pulp, amniotic membrane, chorion, decidua, umbilical cord, umbilical cord blood and other tissues. Studies have shown that mesenchymal stem cells have the characteristics of strong differentiation ability, short doubling time, immunomodulatory effect, low immunogenicity, and easy transfection. They are an ideal seed cell and gene therapy vector for regenerative medicine. Research hotspots in clinical translational application of stem cells. [0003] At present, the mesenchymal stem cells used are mainly bone marrow mesenchymal stem cells, which are convenient and quick to so...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N5/0775
CPCC12N5/0605C12N5/0668C12N2509/00
Inventor 吴韶清孙逊沙欧志英金宇林李发涛唐婕吴洁莹陈劲松谢闺娥陆琰梁绮华刘东
Owner GUANGZHOU WOMEN AND CHILDRENS MEDICAL CENTER
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