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A kind of fat stem cell culture medium and application thereof

A technology of adipose stem cells and culture medium, which is applied in the field of stem cell culture, can solve problems such as difficult clinical treatment, increase the ability of stem cells to resist adversity and the growth environment and the effect of transplantation treatment, and affect the stability of stem cell phenotype, so as to ensure continuous value-added and increase Effects on resilience to adversity, maintenance of phenotype and function

Active Publication Date: 2020-11-06
JINAN UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

However, the addition of these growth factors or small molecular components will affect the phenotype and post-transplantation stability of the stem cells used for transplantation therapy to a certain extent, and fail to significantly increase the ability of the transplanted stem cells to resist the adverse growth environment and the therapeutic effect of transplantation. , it is difficult to be directly applied to clinical treatment

Method used

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  • A kind of fat stem cell culture medium and application thereof
  • A kind of fat stem cell culture medium and application thereof
  • A kind of fat stem cell culture medium and application thereof

Examples

Experimental program
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Embodiment 1

[0033] The LPA / S1P medium of the present invention can be prepared by using dry powder or liquid DMEM, M199, MEM, HBSS, F12, BME, RPMI1640, MCDB104, MCDB153 or a mixed medium thereof.

[0034] Taking DMEM / F12 mixed medium as an example, the preparation method of the complete medium for in vitro culture of adipose stem cells with a standard volume of 1000ml:

[0035] (1) Dissolve 10g DMEM / F12 dry powder in 800ml ultrapure water, add 100ml FBS, then add either or both of 1~25μMLPA and 0.05~0.5μM S1P, and use ultrapure water to make up to volume after fully dissolving To 1000ml, filter with a 0.22-0.1μm membrane filter, fully mix and dissolve, and store at 4°C for later use.

[0036] (2) Add 1000ml of liquid DMEM / F12 medium with 10% FBS, then add either or both of 1-25μM LPA and 0.05-0.5μM S1P, mix well and dissolve, then store at 4°C for later use.

Embodiment 2

[0037] Embodiment 2 The preparation of the optimized LPA / S1P in vitro culture medium of the present invention

[0038] The LPA / S1P medium of the present invention can be prepared by using dry powder or liquid DMEM, M199, MEM, HBSS, F12, BME, RPMI1640, MCDB104, MCDB153 or a mixed medium thereof.

[0039] Taking DMEM / F12 mixed medium as an example, the preparation method of the complete medium for in vitro culture of adipose stem cells with a standard volume of 1000ml:

[0040] (1) Dissolve 10g of DMEM / F12 dry powder in 800ml of ultrapure water, and add 100ml of FBS, 5μM LPA and 0.25μM S1P accordingly. After fully dissolving, use ultrapure water to set the volume to 1000ml, and use a 0.22-0.1μm filter membrane Filter, mix well and dissolve and store at 4°C for later use.

[0041] (2) Add 10% FBS, 5 μM LPA and 0.25 μM S1P to 1000 ml of liquid DMEM / F12 medium in sequence, mix well and dissolve, then store at 4°C for later use.

Embodiment 3

[0042] Example 3 Using the LPA / S1P medium of the present invention to carry out the anti-cell injury experiment of adipose stem cells

[0043] Using the LPA / S1P adipose stem cell medium prepared in Example 1 (2) and Example 2 (2), the adipose stem cells were cultured in vitro, and the in vitro anti-cell damage growth experiments were carried out respectively.

[0044] 0.1 μg / ml LPS and 200 μM H 2 o 2 , and 400mM pure alcohol treatment, the treatment time is 24 hours. In the experiment, cell counting, flow cytometry to detect cell apoptosis, and ELISA kits to detect changes in cellular caspase-3 / 7 activity were used to detect the use of LPA, S1P alone and LPA and S1P at the same time. Adipose-derived stem cells within the recommended concentration For LPS, H 2 o 2 and changes in resistance to cell damage caused by ethanol. like figure 1 and figure 2 It was shown that adding LPA or S1P alone within the recommended concentration can enhance the ability of adipose-derived ...

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Abstract

The invention provides an adipose-derived stem cell culture medium which comprises a basic culture medium and an additive, wherein the additive is one or two of lysophosphatidic acid (LPA) or sphingosine 1-phosphate (S1P). A culture medium used in a conventional adipose-derived stem cell in-vitro culture method is optimized, and compared with a conventional product, the culture medium additionally comprises the special LPA and S1P components. In-vitro tests and in-vivo tests show that the adverse environment growth capability of stem cells can be improved if LPA and S1P are independently or simultaneously used in the adipose-derived stem cell culture process, meanwhile continuous value increase of adipose-derived stem cells can be ensured, and phenotype and function stability of the adipose-derived stem cells can be maintained. By adopting the culture medium, the value increase efficiency, the phenotype stability and the security of in-vitro culture of the adipose-derived stem cells can meet levels of conventional culture systems, meanwhile the efficiency in treating acute liver injury and alcoholic hepatitis after transplanting is greatly improved, and the cultured adipose-derived stem cells can be relatively applicable to stem cell transplanting clinical treatment on multiple diseases.

Description

technical field [0001] The invention relates to the technical field of stem cell culture, and more specifically, to a culture medium for adipose stem cells and its application. Background technique [0002] Acute or chronic liver injury caused by drugs, toxins or alcohol is a serious clinical problem worldwide, for example, about 10% of acute hepatitis cases each year are drug-induced liver injury. In the United States, 15.1 million adults are reported to suffer from alcohol abuse, including 9.8 million men and 5.3 million women. An estimated 88,000 people die each year from alcohol-related diseases. Liver injury caused by these drugs / toxins or alcohol may progress to liver failure requiring prompt liver transplantation. [0003] Due to the rapid development of regenerative medicine, stem cell transplantation has become a promising effective strategy for the treatment of severe liver damage caused by drugs, toxins, alcohol, and to solve many problems in the liver transplan...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N5/0775A61P1/16
CPCC12N5/0667C12N2500/42C12N2500/46
Inventor 肖佳朱江李绵欢刘映霞吕翼何留民
Owner JINAN UNIVERSITY
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