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Anti-aging stem cell culture medium and culture method of mesenchymal stem cells

A technology of stem cells and culture methods, applied in the field of anti-aging stem cell culture medium and mesenchymal stem cell culture, can solve problems such as the decline of BMSC function, and achieve the effect of delaying cell aging, simple formula and large quantity

Inactive Publication Date: 2018-10-02
广东芙金干细胞再生医学有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Then, how to solve the decline of BMSC function has become an urgent problem to be solved at present

Method used

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  • Anti-aging stem cell culture medium and culture method of mesenchymal stem cells
  • Anti-aging stem cell culture medium and culture method of mesenchymal stem cells
  • Anti-aging stem cell culture medium and culture method of mesenchymal stem cells

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0038] Embodiment 1 Culture medium configuration

[0039] Step 1: Basal medium configuration:

[0040] Configure 880ml of α-MEM medium according to the product instructions, add 100ml of fetal bovine serum, 10ml of penicillin, and 10ml of streptomycin in order, mix well, configure the basic medium, filter and sterilize through a 0.22μm microporous membrane, and put in 4 Store at ℃.

[0041] Step 2: Anti-aging Stem Cell Medium Configuration:

[0042] Add 10 μg of miR-705 antagomir, 10 mg of vitamin C and 2 mmol of glutamine in sequence to 1 L of basal medium, mix well and prepare an anti-aging medium, filter and sterilize through a 0.22 μm microporous membrane, and store at 4°C.

[0043] Optionally, the anti-aging stem cell culture also includes: amphotericin. Among them, amphotericin is a kind of polyene antifungal drug isolated from the culture fluid of Streptomyces nodosus.

Embodiment 2

[0045] The method for cultivating mesenchymal stem cells using the basal medium provided in Example 1 and the anti-aging stem cell medium comprises the following steps:

[0046] Step 1: Extract 5ml of bone marrow through the iliac crest puncture, add 10,000U heparin for anticoagulation; add 5ml of phosphate buffer solution to dilute the bone marrow, add slowly to two centrifuge tubes filled with 10ml of mononuclear cell separation solution, Make sure that the bone marrow is distributed on top of the mononuclear cell separation medium. Gently put it into a centrifuge and centrifuge at 2000 rpm for 20 minutes until the bone marrow is layered in the mononuclear cell separation medium. Then use a pipette to carefully absorb the mononuclear cell layer cells, add 10ml PBS to resuspend, wash and centrifuge at 800 rpm for 5 minutes, suck up the supernatant, and collect the mononuclear cells.

[0047] Step 2: Resuspend the bone marrow mononuclear cells collected in step 1 in 10ml of b...

Embodiment 2

[0050] The identification method of the 10th generation mesenchymal stem cells in Example 2 is as follows:

[0051] 1 Observation of cell morphology:

[0052] Cell morphology was observed under 40-fold and 100-fold magnifications using an inverted microscope. Such as figure 1 As shown, the morphology of normal mesenchymal stem cells should be long spindle, and the morphology of cells is uniform.

[0053] 2 Detection of cell surface markers

[0054] The 10th passage mesenchymal stem cells were digested and collected, incubated with CD90, CD105, CD45, and CD31 fluorescent antibodies for 1 hr, washed with PBS three times, and the proportion of positive cells for different markers was detected by flow cytometry. Such as figure 2 As shown, from left to right is a schematic diagram of surface labeling with CD90, CD105, CD45, and CD31 fluorescent antibodies.

[0055] 3 Detection of cell clone formation ability

[0056] The 10th passage mesenchymal stem cells were inoculated in...

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Abstract

The invention discloses an anti-aging stem cell culture medium and a culture method of mesenchymal stem cells. The anti-aging stem cell culture medium is prepared from a low-sugar alpha-MEM culture medium, fetal bovine serum, miR-705 antagomir, vitamin C, glutamine, penicillin and streptomycin. The anti-aging stem cell culture medium can delay the aging and functional decline of the mesenchymal stem cells caused by long-term in vitro passage and amplification.

Description

technical field [0001] The embodiments of the present invention relate to the technical field of bioengineering, in particular to an anti-aging stem cell culture medium and a method for culturing mesenchymal stem cells. Background technique [0002] Bone marrow-derived mesenchymal stem cells (BMSCs) are a group of mesenchymal stem cells that exist in the bone marrow and have strong clone-forming ability and self-renewal ability. High expression of mesenchymal stem cell cell surface markers CD29, CD73 and CD105, with multi-directional differentiation potential, can differentiate into osteoblasts, adipocytes, chondrocytes, myoblasts and other cell types. It is generally believed that BMSCs play a key role in the development, repair and regeneration of bone tissue. BMSCs are the source of osteogenic precursor cells and are directly involved in bone development, remodeling, repair and regeneration. During the bone remodeling process, BMSCs migrate to the site of bone resorptio...

Claims

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Application Information

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IPC IPC(8): C12N5/0775
CPCC12N5/0663C12N2500/32C12N2500/38C12N2500/40C12N2500/84
Inventor 廖立帅逸赵宇邓志宏
Owner 广东芙金干细胞再生医学有限公司
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