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Preparation method of hair follicle stem cells

A technology of hair follicle stem cells and mesenchymal stem cells, applied in the field of preparation of hair follicle stem cells, can solve problems such as difficulty in obtaining raw materials and failure to meet ethical requirements

Active Publication Date: 2017-09-26
GUANGZHOU RAINHOME PHARM&TECH CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] In view of this, the present invention provides a method for preparing hair follicle stem cells, which is used to solve the technical defects of difficulty in obtaining raw materials and inability to meet ethical requirements in the prior art.

Method used

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  • Preparation method of hair follicle stem cells
  • Preparation method of hair follicle stem cells

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0035] Medium A: Mix REGM and MEF medium at a volume ratio of 1:1.

[0036] Medium B: 450mLDMEM basal medium, 50mLFBS, 80pM insulin, 5mML-glutamine, 85ug / LbFGF, 15ug / LSCF and 3×10 -8 mol / L dexamethasone.

[0037] Medium C: 450 mL DMEM basal medium, 50 mL FBS, 10 ng / mL hEGF, 10 ug / LbFGF, 3 ng / mL HGF, 5 ng / mL PDGF and 5 ng / mL TGF-β.

[0038] Medium D: 450 mL DMEM basal medium, 50 mL FBS, 15 ng / mL hEGF, 10 ng / mL IGF, 0.5 ug / L hydrocortisone, 100 ng / mL linoleic acid, 4 ug / mL HGF, 150 ng / mL Wnt3a and 13 ug / mL KGF-7.

[0039] Medium E: 450 mL DMEM basal medium, 50 mL FBS, 0.015% T-β, 10 ng / mL hEGF, 10 ng / ml VEGF, and 2.5 mmol / LL-glutamine.

[0040] Step 1. Collect urine cells

[0041] 1) Add 2mL penicillin / streptomycin double antibody to each collection cup.

[0042] 2) Collect the urine. If the follow-up operation is not performed immediately, store the urine in a 4°C refrigerator and complete the follow-up operation on the same day.

[0043] 3) One well of a six-well plate wa...

Embodiment 2

[0065] Medium A: Mix REGM and MEF medium at a volume ratio of 1:1.

[0066] Medium B: 450mL DMEM basal medium, 50mLFBS, 1pM insulin, 1mML-glutamine, 50ug / LbFGF, 5ug / LSCF and 2×10 -8 mol / L dexamethasone.

[0067] Medium C: 450 mL DMEM basal medium, 50 mL FBS, 1 ng / mL hEGF, 100 ug / LbFGF, 1 ng / mL HGF, 10 ng / mL PDGF, and 10 ng / mL TGF-β.

[0068] Medium D: 450 mL DMEM basal medium, 50 mL FBS, 10 ng / mL hEGF, 1 ng / mL IGF, 1 ug / L hydrocortisone, 1 ng / mL linoleic acid, 5 ug / mL HGF, 30 ng / mL Wnt3a and 10 ug / mL KGF-7.

[0069] Medium E: 450 mL DMEM basal medium, 50 mL FBS, 0.001% T-β, 1 ng / mL hEGF, 100 ng / ml VEGF, and 1 mmol / LL-glutamine.

[0070] Step 1. Collect urine cells

[0071] 1) Add 2mL penicillin / streptomycin double antibody to each collection cup.

[0072] 2) Collect the urine. If the follow-up operation is not performed immediately, store the urine in a 4°C refrigerator and complete the follow-up operation on the same day.

[0073] 3) One well of a six-well plate was prep...

Embodiment 3

[0095] Medium A: Mix REGM and MEF medium at a volume ratio of 1:1.

[0096] Medium B: 450mL DMEM basal medium, 50mL FBS, 100pM insulin, 10mML-glutamine, 200ug / LbFGF, 50ug / LSCF and 2.5×10-8mol / L dexamethasone.

[0097] Medium C: 450 mL DMEM basal medium, 50 mL FBS, 100 ng / mL hEGF, 1 ug / LbFGF, 50 ng / mL HGF, 20 ng / mL PDGF and 25 ng / mL TGF-β.

[0098] Medium D: 450 mL DMEM basal medium, 50 mL FBS, 115 ng / mL hEGF, 100 ng / mL IGF, 10 ug / L hydrocortisone, 150 ng / mL linoleic acid, 10 / mL HGF, 200 ng / mL Wnt3a and 65 ug / mL KGF-7.

[0099] Medium E: 450 mL DMEM basal medium, 50 mL FBS, 0.1% T-β, 100 ng / mL hEGF, 1 ng / ml VEGF, and 10 mmol / LL-glutamine.

[0100] Step 1. Collect urine cells

[0101] 1) Add 2mL penicillin / streptomycin double antibody to each collection cup.

[0102] 2) Collect the urine. If the follow-up operation is not performed immediately, store the urine in a 4°C refrigerator and complete the follow-up operation on the same day.

[0103] 3) One well of a six-well plate...

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Abstract

The invention belongs to the technical field of biochemistry, and particularly relates to a preparation method of hair follicle stem cells. The invention provides the preparation method of the hair follicle stem cell. The method comprises the following steps of 1, collecting urine cells; 2, reprogramming urine cells; 3, inducing the induced culture of multifunctional cells; 4, performing induced culture of mesenchymal stem cells to obtain the hair follicle stem cells. Through the technical scheme provided by the invention, the hair follicle stem cells are prepared; the positive mark CK15 of the hair follicle stem cells is in positive expression in most cobblestone cells as hair follicle resources; meanwhile, through the analysis and display by a flow cytometry, the CK15, CK19, CD200 and beta 1 expression in the hair follicle stem cells is positive; the condition conforms to the expression result of the surface mark of the hair follicle stem cells. The preparation method of the hair follicle stem cells provided by the invention solves the problems of raw material obtaining difficulty and ethics requirement meeting incapability of the preparation method of the hair follicle stem cells in the prior art.

Description

technical field [0001] The invention belongs to the technical field of biochemistry, and in particular relates to a preparation method of hair follicle stem cells. Background technique [0002] The skin is the organ with the largest surface area in the human body. It plays an important role in resisting microbial invasion, ultraviolet radiation, preventing water loss, and regulating body temperature. Limited skin volume and barriers to additional damage. [0003] The use of artificial skin cultured in vitro, that is, tissue engineered skin, as a source of transplantation can fundamentally solve this problem, but how to obtain seed cells has been puzzling people. With the development of medical biology and the deepening of skin tissue engineering, hair follicle stem cells have gradually attracted people's attention as a new seed cell. More and more studies have shown that hair follicle stem cells have the potential of multidirectional differentiation. However, in the prior ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N5/10
CPCC12N5/0628C12N5/0666C12N2500/32C12N2501/11C12N2501/115C12N2501/125C12N2501/33C12N2501/39C12N2506/1376C12N2506/45C12N2510/00
Inventor 黄燕飞车七石刘少辉
Owner GUANGZHOU RAINHOME PHARM&TECH CO LTD
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