Use of human amniotic mesenchymal stem cell exosome

A technology of mesenchymal stem cells and human amniotic membrane, applied in the field of stem cells, can solve the problem of exosome application without human amniotic membrane mesenchymal stem cells

Inactive Publication Date: 2015-03-04
GUANGZHOU SALIAI STEMCELL SCI & TECH CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] At present, there is no application of human amniotic

Method used

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  • Use of human amniotic mesenchymal stem cell exosome
  • Use of human amniotic mesenchymal stem cell exosome
  • Use of human amniotic mesenchymal stem cell exosome

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0073] Example 1. Culture of human amniotic mesenchymal stem cells and collection of culture supernatant

[0074] 1. Culture of human amniotic mesenchymal stem cells:

[0075] Well-growing human amniotic mesenchymal stem cells were taken, cultured in serum-free medium, digested and passaged with 0.25% trypsin, passed to the P2 generation, and when they grew to a confluence of 80%, the culture supernatant was discarded and washed three times with PBS. Add 1640 basal medium, collect the culture supernatant every day, and replace it with fresh 1640 medium, collect and culture continuously for 3 to 5 days, then collect the culture supernatant for exosome extraction. Among them, the cell state used to collect the culture supernatant is as follows: figure 1 shown.

[0076] 2. Extraction of exosomes (unless otherwise specified, the whole extraction process is carried out at 4°C):

[0077] (1), centrifuge all collected culture supernatants at 300 g for 10 min, retain the supernatan...

Embodiment 2

[0082] Example 2. Preparation of human amniotic mesenchymal stem cell exosome freeze-dried powder:

[0083] Take the exosomes purified in Example 1, add trehalose to a final concentration of 10%, mix well, pass through a 0.22 μm filter membrane, divide and store in an ultra-low temperature refrigerator (-80°C) for 12 hours, and then transfer the sample to Freeze-dried in a vacuum freeze-drying chamber, the vacuum degree of freeze-drying was kept at 10 Pa, and the temperature was frozen at -50°C for 12 to 48 hours to obtain the human amniotic mesenchymal stem cell exosome freeze-dried powder.

Embodiment 3

[0084] Example 3. Anti-photoaging test of human amniotic mesenchymal stem cell exosomes in vitro

[0085] 1. MTT method to determine the effect of human amniotic mesenchymal stem cell exosomes on the growth of human dermal fibroblasts

[0086] Test sample: human amniotic mesenchymal stem cell exosome freeze-dried powder prepared in Example 3 (protein content: 50.0±0.5 μg / cartridge).

[0087] Test organism: Human fibroblasts (HDF cells).

[0088] The specific steps are as follows:

[0089] (1) Use 10% (v / v) FBS DMEM / F12 medium (penicillin 200U / ml, streptomycin 250U / ml, pH7.2) to cultivate HDF cells, when the cell density reaches 100% fusion, use the cell Digest the cells with the digestive solution and adjust the cell concentration to 1×10 5 cell / ml, inoculated into 96-well plate, 100 μL per well, set 4 duplicate wells, 37°C, 5% CO 2 , in a cell incubator with saturated humidity to continue culturing;

[0090] (2) After 24 hours, the medium was discarded, and the final con...

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Abstract

The invention relates to the field of stem cells, and discloses novel use of a stem cell exosome, and particularly discloses novel use of a human amniotic mesenchymal stem cell exosome. The experiment result shows that a human amniotic mesenchymal stem cell is capable of promoting growth of HDF cells, improving the activity of SOD (Superoxide Dismutase) enzyme in the cells, reducing attacks and damages to the cells caused by oxygen radicals generated by ultraviolet radiation induction, and reducing the death rate of the cells. The use shows that the human amniotic mesenchymal stem cell exosome has an anti-photoaging effect, and meanwhile, the human amniotic mesenchymal stem cell exosome is free of phototoxicity, so that application of the human amniotic mesenchymal stem cell exosome in preparation of an anti-ageing preparation is provided. The invention also provides an anti-ageing preparation of the human amniotic mesenchymal stem cell exosome.

Description

technical field [0001] The present invention relates to the field of stem cells, in particular to new uses of exosomes from stem cells, in particular to new uses of exosomes from human amniotic mesenchymal stem cells. Background technique [0002] Exosomes were first discovered in the supernatant of sheep erythrocytes cultured in vitro. They are vesicle-like bodies actively secreted by cells with a uniform size, a diameter of 40-100 nm, and a density of 1.10-1.18 g / ml. The application of exosomes mainly focuses on clinical diagnosis and anti-tumor. [0003] Mesenchymal stem cells (MSCs) are cells with self-renewal and multi-directional differentiation capabilities, produce and release nutrients, promote cell repair and angiogenesis through paracrine pathways, and play an important role in the field of regenerative medicine. Lai et al. analyzed by mass spectrometry and found that exosomes of MSCs contained 857 proteins including pyruvate kinase (pyruvate kinase, PK), milkfat...

Claims

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Application Information

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IPC IPC(8): A61K8/98A61K8/02A61K35/28A61K9/19A61K8/06A61Q19/08A61P39/06
Inventor 陈海佳王一飞葛啸虎麦锦连王小燕马岩岩舒辉萍
Owner GUANGZHOU SALIAI STEMCELL SCI & TECH CO LTD
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