Composition and application of stem cells and hyaluronic acid
A composition and technology of hyaluronic acid, applied in the field of cosmetic medicine, can solve problems such as not being able to get immediate results, and achieve the effects of long duration, rapid onset duration, significant volume and wet weight
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Embodiment 1
[0034] Example 1 Preparation of Adipose Stem Cells
[0035] Divide human adipose tissue into 50mL centrifuge tubes, 20mL per tube.
[0036] Add an equal volume of 0.5% type Ⅰ collagenase (final concentration: 0.25%) to each tube, mix thoroughly, seal, transfer to a constant temperature air shaker, and digest for 1 hour at 37°C and 100R.
[0037] Add 4mL of FBS to each tube to stop digestion and mix well.
[0038] Centrifuge at 1500rpm / min for 5min.
[0039] After centrifugation, take 2 mL of the upper layer of blood (middle layer), divide it into two EP tubes, and keep samples for inspection.
[0040] Discard the upper two layers of liquid, add 40mL PBS to each tube to resuspend, and resuspend the cells.
[0041]Centrifuge at 1500rpm / min for 5min, discard the supernatant, and obtain the adipose stem cells. Add DMEM-F12 medium containing 15% FBS to resuspend the cells at 1×10 5 individual / mL inoculation.
[0042] Gently shake the culture dish back and forth to distribute ...
Embodiment 2
[0050] Example 2 Quality Identification of Adipose Stem Cells
[0051] The adipose stem cells prepared in Example 1 were taken, and the surface antigen detection, adipogenic differentiation, and osteogenic differentiation were respectively performed to identify the quality of the adipose stem cells provided in Example 1.
[0052] 1. Surface antigen detection of adipose-derived stem cells
[0053] Detection adopts flow cytometry, respectively detects the expression of CD44, CD73, CD90, CD105, CD34, CD45 and HLA-DR in the fat stem cells provided by Example 1, specifically:
[0054] Adipose stem cells of different passages were taken, the medium was sucked off, and 0.25% trypsin was routinely digested to make 1×10 5 Take 5 μL of monoclonal antibodies against human CD44, CD73, CD90, CD105, CD34, CD45 and HLA-DR respectively, add 500 μL of cell suspension, and incubate at room temperature for 20 minutes in the dark, and set up a blank isotype control at the same time. Centrifuge ...
Embodiment 3
[0059] Embodiment 3 Composition function identification provided by the present invention
[0060] 1. Preparation of the composition provided by the present invention: the adipose stem cells prepared in Example 1 and hyaluronic acid were mixed according to Table 1.
[0061] Table 1 Contents of each component
[0062]
Volume fraction of hyaluronic acid (%)
Adipose stem cell concentration (unit / mL)
Composition a
80
1.0×10 4
85
1.0×10 5
90
1.0×10 6
composition d
85
1.0×10 7
[0063] Two, the effect identification of the composition provided by the invention
[0064] 1. Take the composition recorded in Table 1, hyaluronic acid and the adipose stem cells prepared in Example 1, and prepare two preparations with a volume of 0.5 mL / bottle, respectively.
[0065] 2. The nude mice were randomly divided into 6 groups, 15 in each group. Each group of mice was injected with equal do...
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