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Improved mesenchymal stem cell (MSC) culture medium, bone marrow mesenchymal stem cells (BMSCs) and culture method and application of BMSCs

A bone marrow mesenchymal and mesenchymal stem cell technology, applied in the field of improved mesenchymal stem cell culture medium, can solve the problems of mitochondrial DNA damage and high levels of reactive oxygen species

Active Publication Date: 2020-12-22
源生生物科技(青岛)有限责任公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] However, due to the high level of reactive oxygen species in the environment of mitochondria in mesenchymal stem cells, their mitochondrial DNA is easily attacked and damaged

Method used

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  • Improved mesenchymal stem cell (MSC) culture medium, bone marrow mesenchymal stem cells (BMSCs) and culture method and application of BMSCs
  • Improved mesenchymal stem cell (MSC) culture medium, bone marrow mesenchymal stem cells (BMSCs) and culture method and application of BMSCs
  • Improved mesenchymal stem cell (MSC) culture medium, bone marrow mesenchymal stem cells (BMSCs) and culture method and application of BMSCs

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0057] (1) Isolation of bone marrow mesenchymal stem cells: 15 mL of human bone marrow was extracted, and anticoagulated at 100U heparin per mL of bone marrow. Add the same amount of normal saline to the bone marrow and mix well, add 1 part of lymphocyte separation solution (Ficoll separation solution) to 2 parts of diluted bone marrow, slowly add the diluted bone marrow to the surface of Ficoll, and centrifuge at 2000 r / min in a horizontal centrifuge at 20 °C. After 30 minutes, the bone marrow mesenchymal stem cells taken out from the interface were washed three times with normal saline, and then an appropriate amount of culture medium was added for counting.

[0058] (2) Primary culture of bone marrow mesenchymal stem cells: according to 0.3×10 6 The bone marrow mesenchymal stem cells obtained in step (1) were inoculated in different mesenchymal stem cell culture media (composed of basal medium + medium additives, the specific composition is shown in Table 1) for 3 days at a...

Embodiment 2

[0064] (1) Isolation of bone marrow mesenchymal stem cells: 15 mL of human bone marrow was extracted, and anticoagulated at 100U heparin per mL of bone marrow. Add the same amount of normal saline to the bone marrow and mix well, add 1 part of lymphocyte separation solution (Ficoll separation solution) to 2 parts of diluted bone marrow, slowly add the diluted bone marrow to the surface of Ficoll, and centrifuge at 2000 r / min in a horizontal centrifuge at 20 °C. After 30 minutes, the bone marrow mesenchymal stem cells taken out from the interface were washed three times with normal saline, and then an appropriate amount of culture medium was added for counting.

[0065] (2) Primary culture of bone marrow mesenchymal stem cells: according to 5×10 6 The bone marrow mesenchymal stem cells obtained in step (1) were inoculated into different mesenchymal stem cell culture media at a density of / mL, wherein: the mesenchymal stem cell culture medium used in the first group was made of ...

Embodiment 3

[0097] The effect of the bone marrow mesenchymal stem cells prepared in Example 2 in the treatment of type 2 diabetes

[0098] The P3 generation bone marrow mesenchymal stem cells obtained by culturing the mesenchymal stem cell medium consisting of serum-free mesenchymal stem cell medium, 1 μM coenzyme Q10, 1 μM melatonin and 1 μM nicotinamide mononucleotide in Example 2 were used (We call it "enhanced bone marrow mesenchymal stem cells", hereinafter referred to as eBMSCs) in preclinical experiments in mice with type 2 diabetes. specifically:

[0099] (1) Eight-week-old male C57BL / 6 mice were divided into two groups. One group (12 mice) was fed a high-fat diet with a fat content of 60%, which was the HFD (high-fat diet) group; One group (6 mice) was fed a normal mouse maintenance diet, which was the ND (normal diet) group. The statistical results of the changes in body weight of the two groups of mice over time are shown in Table 7.

[0100] Table 7

[0101]

[0102] It...

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Abstract

The invention relates to an improved mesenchymal stem cell (MSC) culture medium, bone marrow mesenchymal stem cells (BMSCs) and a culture method and application of the BMSCs. The improved MSC culturemedium comprises a base culture medium, a first component and melatonin, wherein the first component is selected from at least one of coenzyme Q10 and mitoxantrone mesylate, the working concentrationof the first component is 1-20 [mu]M, the working concentration of the melatonin is 1-20 [mu]M, and the molar ratio of the first component to the melatonin is 1: (0.2-10). The improved MSC culture medium can improve mitochondrial activity and mitochondrial quantity in MSCs.

Description

technical field [0001] The invention relates to cell culture technology, in particular to an improved mesenchymal stem cell culture medium, bone marrow mesenchymal stem cells and a culture method and application thereof. Background technique [0002] Mitochondria are important semi-autonomous organelles in cells. They have mitochondrial DNA that can encode part of the genetic information of mitochondria. They are the main site of aerobic respiration in cells and are responsible for providing energy to cells. In addition, mitochondria play an important role in cell differentiation, cell secretion, cell proliferation and apoptosis. [0003] Mesenchymal stem cells (MSCs) are a type of pluripotent stem cells belonging to the mesoderm, which mainly exist in connective tissues and organ interstitium. They have strong proliferation and differentiation capabilities, and have low immunogenicity. Bone marrow mesenchymal stem cells (BMSCs) are one of the most widely used mesenchymal s...

Claims

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Application Information

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IPC IPC(8): C12N5/0775A61K35/28A61P3/10A61P1/16
CPCC12N5/0663A61K35/28A61P3/10A61P1/16C12N2501/825C12N2501/999C12N2500/40C12N2500/34C12N2501/11A61P1/18A61P3/08C12N2500/38C12N2501/30C12N2500/90
Inventor 杨智广姬广聚张梦琦
Owner 源生生物科技(青岛)有限责任公司
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