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Method for obtaining artificial skin through stem cell in-vitro differentiation and application of method

An artificial skin and stem cell technology, applied in cell dissociation methods, epidermal cells/skin cells, artificial cell constructs, etc., can solve problems such as reducing the depth of wounds, and achieve accelerated inflammatory response subsidence, depth reduction, and satisfactory healing effect. Effect

Inactive Publication Date: 2020-11-06
北京臻溪谷医学研究中心(有限合伙)
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  • Abstract
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  • Claims
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Problems solved by technology

[0003] In order to solve the above technical problems, the present invention provides a method for obtaining artificial skin by differentiating stem cells into skin cells in vitro. The artificial skin prepared by this culture method can further reduce the depth of the wound surface, and the acellular dermal matrix is ​​supplemented by the in vitro differentiation technology. The missing cell components, differentiated skin cells and acellular allogeneic dermis constitute tissue engineered skin, which can promote wound healing and accelerate the regression of graft inflammatory reactions; this method can be used for trauma, burns, diabetic foot, tumor resection, etc. skin damage

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  • Method for obtaining artificial skin through stem cell in-vitro differentiation and application of method

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Embodiment 1

[0038] This embodiment provides a method for obtaining artificial skin by differentiating fat stem cells into skin cells in vitro, the culture method comprising the following steps:

[0039] S1: Subculture the mesenchymal stem cells, culture them with complete medium until the cells are 90% confluent, collect the P3 generation cells, and make a mesenchymal stem cell suspension. The preparation method of the mesenchymal stem cell suspension is to take 2×10 6Mesenchymal stem cells were prepared by digesting 0.25% trypsin for 3 minutes; the complete medium was: DMEM / F12 as the base medium, adding human fibronectin at a concentration of 25 μg / ml, and alkaline at a concentration of 10 ng / ml Fibroblast growth factor, human epidermal growth factor at a concentration of 15 ng / ml, ITS at a concentration of 1%, human albumin at a concentration of 5%, NEAA at a concentration of 1%, hydrogenation at a concentration of 0.1 μmol / L Cortisone, 0.1% beta-mercaptoethanol;

[0040] S2: Attachme...

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Abstract

The invention relates to a method for obtaining artificial skin through stem cell in-vitro differentiation and application of the method, and provides a method for differentiating stem cells to skin cells in vitro to obtain artificial skin and application. The culture method comprises the following steps: S1, subculturing mesenchymal stem cells until 60-90% of cells are merged, and collecting P3 generation cells to prepare a mesenchymal stem cell suspension; S2, attaching, specifically, placing acellular dermis and the mesenchymal stem cell suspension in an ultralow adsorption reactor, and carrying out culturing by using a complete culture medium; and S3, inducing the mesenchymal stem cells loaded by the acellular dermis to differentiate to skin cells in vitro, specifically, changing liquid by using an induction culture solution, wherein the induction culture solution takes a KFSM culture medium as a basic culture medium, astragalus injection with the concentration of 30-50 g / L and fetal calf serum with the volume fraction of 1-3% are added, and the induction culture time is 14 days. TM4SF1 and broad-spectrum cytokeratin expression are positive, the obtained artificial skin is moresimilar to cells forming skin tissues, and a better treatment effect can be achieved.

Description

technical field [0001] The invention belongs to the technical field of biological tissue and organ repair, and particularly relates to a method for obtaining artificial skin by differentiating stem cells into skin cells in vitro and its application. Background technique [0002] The skin is the largest organ of the human body, and it can be damaged by trauma, burns, diabetic foot, tumor resection, etc. Wound repair is generally divided into three overlapping stages: inflammatory reaction stage-proliferative stage-remodeling stage, any stage out of control will lead to pathological healing, such as keloid formation, delayed wound healing or incomplete healing; severe cases It can cause local function limitation, loss of skin barrier function, trigger sepsis and other infectious diseases, and threaten the lives of patients. Currently, autologous skin grafts or flaps are the clinically used repair methods for deep or large skin defects. However, autologous skin graft or skin ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N5/071C12N5/0775A61L27/38A61L27/60
CPCC12N5/0629C12N5/0662C12N5/0667C12N5/0668A61L27/3834A61L27/60C12N2506/1346C12N2506/1384C12N2506/1392C12N2533/92C12N2500/76C12N2533/52C12N2501/115C12N2501/11C12N2500/25C12N2501/998C12N2501/39C12N2500/44C12N2500/32C12N2509/00A61L2430/34
Inventor 曹毓琳滕睿頔张海林白志惠
Owner 北京臻溪谷医学研究中心(有限合伙)
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