Culture medium for obtaining mesenchymal stem cells and exosomes thereof and preparation method of culture medium

A mesenchymal stem cell and culture medium technology, applied in the field of culture medium and preparation of mesenchymal stem cells and their exosomes, can solve the problem of expensive serum-free medium, poor cell proliferation effect, short cell growth cycle, etc. problem, achieve the effects of shortening the culture time, promoting division and proliferation, and increasing secretion

Pending Publication Date: 2022-05-13
杭州荣泽生物科技集团有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0003] First of all, in the traditional method, the culture system of stem cells mainly uses the medium containing animal serum, but the heterogeneous pollution brought by animal serum and the pollution of known or unknown pathogens such as viruses and mycoplasma, the growth in this environment Stem cells, what kind of changes will happen to their internal structure is not yet known. Secondly, the serum-free medium on the market has disadvantages such as high price, short cell growth cycle, poor cell proliferation effect and fast cell aging.

Method used

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  • Culture medium for obtaining mesenchymal stem cells and exosomes thereof and preparation method of culture medium
  • Culture medium for obtaining mesenchymal stem cells and exosomes thereof and preparation method of culture medium
  • Culture medium for obtaining mesenchymal stem cells and exosomes thereof and preparation method of culture medium

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Experimental program
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Effect test

Embodiment 1

[0039] The present invention provides a technical solution: a medium for obtaining mesenchymal stem cells and their exosomes, with DMEM / F12 as the base medium adding the following components: 10-22g / mL serum substitute, 20-55ng / mL Growth-promoting factor, 5-10ng / mL muscle cell repair factor HCGF, 5-16mM anti-aging factor, buffer, 0.5-3mg / L antioxidant, 1-20mg / L vitamin, antibiotic, 5-65mg / L trace element , 1-16 μg / mL protective agent, the volume ratio of the buffer agent to the DMEM / F12 basal medium is 1%, and the volume ratio of the antibiotic to the DMEM / F12 basal medium is 2%;

[0040] Serum substitutes include 10-20 g / L human albumin, 5-20 mg / L human transferrin, 1-10 mg / L human insulin, 1-5 mg / L fibronectin and 5-30 μg / mL cholesterol;

[0041] Growth-promoting factors include 5-10 ng / mL epidermal growth factor EGF, 5-10 ng / mL fibroblast growth factor FGF, 5-10 ng / mL platelet-derived mass proliferating factor PDGF, 5-10 ng / mL growth hormone-releasing inhibitory factor SRIH...

Embodiment 1-6

[0061] Table 2 Example 1-6 cell morphology conclusion

[0062]

[0063] It can be seen from Table 1 and Table 2 that: (1) growth-promoting factors are important constituent factors for obtaining mesenchymal stem cells, and the medium lacking growth-promoting factors cannot obtain mesenchymal stem cells; (2) muscle layer cell repair factors are beneficial to The cell outline is clearer; (3) anti-aging factors have the effect of delaying cell aging; (4) antioxidants are beneficial to promote cell proliferation; (5) antibiotics have the effect of preventing cell culture from being contaminated by fungi or bacteria;

[0064] The composition of the mesenchymal stem cell culture medium of the present invention is determined, does not contain serum components, the quality is controllable, and it can be produced on a large scale. It can be used for in vitro culture of stem cells, and the stem cells can grow normally. Moreover, the mesenchymal stem cell culture medium of the present ...

Embodiment 116-119

[0081] The amount of exosome protein per unit volume in the medium of table 8 embodiment 116-119

[0082] Example Exosome protein amount (mg) Example 116 6.88 Example 117 3.45 Example 118 4.24 Example 119 5.06

[0083] It can be seen from Table 7 and Table 8 that the addition of recombinant interferon γ and cytochalasin D in the medium can significantly increase the secretion of exosomes, and cooperate with the damaged tissue environment provided by inorganic salts to the medium, so that The production of exosomes has been improved, realizing the purpose of using mesenchymal stem cells to obtain large-scale production of exosomes.

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Abstract

The invention discloses a culture medium for obtaining mesenchymal stem cells and exosomes thereof and a preparation method of the culture medium. DMEM/F12 is used as a basic culture medium, and the following components are added: 10-22 g/mL of a serum substitute, 20-55 ng/mL of a growth promoting factor, 5-10 ng/mL of a muscle cell repair factor HCGF, 5-16 mM of an anti-aging factor, a buffer agent, 0.5-3 mg/L of an antioxidant, 1-20 mg/L of vitamins, antibiotics, 5-65 mg/L of trace elements and 1-16 [mu] g/mL of a protective agent. The mesenchymal stem cell culture medium is definite in component, free of serum components, controllable in quality and capable of being produced on a large scale, stem cells can normally grow when the mesenchymal stem cell culture medium is used for in-vitro culture of the stem cells, moreover, the mesenchymal stem cell culture medium is beneficial to division and proliferation of the stem cells, the stem cell amplification rate is greatly increased, and the mesenchymal stem cell culture medium is suitable for large-scale production. Compared with the prior art, the stem cell culture medium has the advantages that stem cell culture time is shortened, and stem cells still can keep original biological characteristics and multidirectional differentiation potential after being subjected to multiple passage times by the aid of the stem cell culture medium.

Description

technical field [0001] The invention relates to the technical field of mesenchymal stem cell culture, in particular to a culture medium for obtaining mesenchymal stem cells and exosomes and a preparation method thereof. Background technique [0002] Mesenchymal stem cells are a kind of pluripotent stem cells derived from mesoderm, which widely exist in bone marrow, umbilical cord, placenta, fat and other tissues and blood, capable of unlimited self-renewal and multi-directional differentiation. Directed differentiation into adipocytes, chondrocytes, muscle cells, etc. in vitro or in vivo. As seed cells for tissue repair, mesenchymal stem cells can be cultured and expanded in vivo or in vitro, and can secrete a large number of cytokines. Inhibition and other functions are used to treat immune-related diseases, and mesenchymal stem cells basically do not have immunogenicity and can be transplanted allogeneically. Generally speaking, mesenchymal stem cells have great clinical m...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N5/0775
CPCC12N5/0668C12N5/0665C12N5/0667C12N5/0663C12N2500/90C12N2501/11C12N2501/113C12N2501/115C12N2501/119C12N2501/135C12N2501/10C12N2501/117C12N2501/998C12N2500/24C12N2501/33C12N2500/30C12N2501/727C12N2501/72C12N2500/38C12N2500/05C12N2500/46C12N2501/24C12N2509/00C12N2509/10
Inventor 陈相波雷鸣田朋飞应荣
Owner 杭州荣泽生物科技集团有限公司
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