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Method for simultaneously preparing autologous epidermal cells and fibroblasts, and biological beauty product comprising autologous epidermal cells and fibroblasts

A technology of fibroblasts and epidermal cells, applied in animal cells, cosmetic preparations, cosmetic preparations, etc., can solve the problems of increased usage, insufficient skin beauty efficacy, poor effect, etc., and achieves regeneration and proliferation. Effect

Inactive Publication Date: 2015-05-13
GUANGZHOU SAIYIDE BIOTECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The application of these two factors tends to produce obvious dependence. As the number of injections increases, people's sensitivity to them decreases, which leads to patients using this method more frequently, and the amount of each use will gradually increase.
[0005] To sum up, due to various reasons and problems, biological beauty products currently on the market have various ineffective effects, and the curative effect on people’s skin beauty is far from enough, even if various nutrients are added to them

Method used

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  • Method for simultaneously preparing autologous epidermal cells and fibroblasts, and biological beauty product comprising autologous epidermal cells and fibroblasts
  • Method for simultaneously preparing autologous epidermal cells and fibroblasts, and biological beauty product comprising autologous epidermal cells and fibroblasts
  • Method for simultaneously preparing autologous epidermal cells and fibroblasts, and biological beauty product comprising autologous epidermal cells and fibroblasts

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0049] Example 1 Preparation of human autologous epidermal cells

[0050] By taking people 0.1-2cm 2 For the skin, infiltrate 5ml of 1×phosphate buffered saline (PBS, pH 7.4, containing 100IU / ml penicillin and 0.1mg / ml streptomycin) for 15 minutes, suck it up with a 1ml pipette gun, and then rinse with 1×PBS (pH Value 7.4, containing 100IU / ml penicillin and 0.1mg / ml streptomycin) were also washed repeatedly, digested with disperse II after washing, placed in a refrigerator at 4°C overnight, and the skin was taken out the next day and the epidermis and dermis were torn apart with tweezers.

[0051] Cut the epidermis to 1mm with ophthalmic scissors 3 After washing with 1×PBS, add it to a 15ml centrifuge tube, centrifuge at 1000 rpm for 5 minutes, discard the supernatant, add 3ml 0.25% (mass volume ratio) trypsin to digest for 30 minutes, wherein trypsin contains mass volume ratio 0.01% EDTA; pass through a 40 μm cell sieve after digestion, add 10ml 1×PBS after filtration, cent...

Embodiment 2

[0052] Example 2 Preparation of human autologous fibroblasts

[0053] Get the dermis obtained in Preparation Example 1 and cut it to 1mm with ophthalmic scissors 3 After washing with 1×PBS, add it to a 15ml centrifuge tube, centrifuge at 1000 rpm for 5 minutes, discard the supernatant, add 3ml 0.25% trypsin to digest for 30 minutes, pass through a 40μm cell sieve after digestion, add 10ml 1 ×PBS, centrifuge at 1000 rpm for 10 minutes to obtain a single cell pellet, resuspend in 10ml 1×PBS again, and centrifuge at 1000 rpm for 10 minutes to obtain a single cell pellet, add α-MEM medium to resuspend, Counting, using α-MEM medium (containing 10% fetal bovine serum, 50ng / ml recombinant human stem cell growth factor, 25ng / ml recombinant human epidermal growth factor and 25ng / ml recombinant human basic fibroblast growth factor) according to 1- 10×10 6 Cells / ml add 15ml to 75cm 2 culture flask at 37°C, 5% CO 2 Cultivate in an incubator; replace with fresh culture medium every 2 d...

Embodiment 3

[0055] Example 3 Detection of specific protein expression in human autologous epidermal cells and fibroblasts

[0056] Take 1 part of 50 μL each of the autologous epidermal cells prepared in Example 1 and the fibroblast suspension prepared in Example 2 (2×10 6 1) Add 500 μL of pre-cooled ready-to-use total protein extraction and lysate (RIPA, Beijing Biotech Company) (containing 1% protease inhibitor by volume, Roche Company, Switzerland), incubate on ice for 20 minutes, 13000 rpm, 4 Centrifuge for 20 minutes. Take the supernatant, aliquot and store at -80°C for later use. The protein concentration was determined according to the instructions of the bicinchoninic acid (BCA) protein quantification kit (Wuhan Boster Company). Adjust the protein concentration with RIPA, the final concentration of the sample is 1.0 μg / μL, add 5× protein sample buffer, and keep at 95°C for 5 minutes.

[0057] According to the molecular weight of the target protein, a 10% separating gel was prepa...

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Abstract

The invention provides a method for simultaneously preparing autologous epidermal cells and fibroblasts. The method comprises the steps of obtaining the epidermal cells from autologous skin, and culturing the epidermal cells to obtain the fibroblasts. The invention further provides a biological beauty product comprising the autologous epidermal cells and / or the fibroblasts. One or two cells are added into autologous peripheral blood platelet-rich plasma, and meanwhile, nutritional factors can be added to prepare the biological beauty product comprising autologous living cells; the product has beautifying effects of removing vitiligo macules, repairing scars, removing skin wrinkles and spots, and the like.

Description

technical field [0001] The invention relates to a method for simultaneous preparation of autologous epidermal cells and fibroblasts and a biocosmetic product obtained by the method. In particular, the present invention relates to fibroblasts obtained by obtaining epidermal cells from autologous skin and culturing them, adding one or two types of cells to the platelet-rich plasma of autologous peripheral blood, and at the same time adding nutritional factors to produce The bio-cosmetic product containing autologous living cells can be used for cosmetic effects such as vitiligo freckle removal, scar repair, skin wrinkle removal and freckle removal. Background technique [0002] A few days ago, a variety of cells used in beauty products have appeared on the market, and have been endowed with various magical effects and hopes by merchants, especially the application of stem cells in beauty. In fact, living cells have achieved some good results in the application of medical cosm...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N5/071A61K8/98A61K8/97A61K8/64A61K8/66A61Q19/08A61P17/02A61P17/10
Inventor 陈镇洲寇蕊刘兵
Owner GUANGZHOU SAIYIDE BIOTECH
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