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Method for constructing composite tissue engineering skins

A technology for engineering skin and composite tissue, which is applied in the field of construction of composite tissue engineering skin, can solve the problems of long time and incompatibility with clinical application, and achieve the effect of promoting cell growth, huge potential, and short construction time

Active Publication Date: 2017-05-10
SHANXI MEDICAL UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

For example, the separation and cultivation of seed cells used to construct artificial skin takes a long time and cannot be adapted to clinical applications; the biocompatibility and three-dimensional pore structure of the currently constructed dermal scaffolds are still far behind those of normal dermal matrix ; Although the composite skin used in clinical practice can cover skin defect wounds at one time, the artificial skin with skin appendages is still under study

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0030] Example 1: Isolation, cultivation and identification of epidermal stem cells.

[0031] Seed cells were derived from the remaining skin grafts from pediatric circumcision or burn plastic skin grafting.

[0032]Under aseptic conditions, after the fat and subcutaneous tissue were removed as thoroughly as possible, the skin slices were washed repeatedly with saline, cut into 2mm×2mm sizes, immersed in 5g / L Disapse II, and digested at 4°C for 16-18h. The epidermis was separated from the dermis, and the epidermal slices were picked out and placed in 0.125wt% trypsin digestion solution containing 0.01wt% EDTA, digested at room temperature for 15min, and the serum terminated the digestion. Filter, centrifuge (1000rpm, 5min) and discard the supernatant, and use K-SFM medium containing 10wt% fetal bovine serum as the epidermal stem cell medium to suspend the cells. Dilute the resuspended cells in 4×10 5 The concentration of each / ml was inoculated in the culture dish pre-coated ...

Embodiment 2

[0036] Example 2: Preparation of sheep acellular dermal matrix scaffold.

[0037] The sheep skin of healthy adult white sheep is selected, the subcutaneous tissue is removed, the epidermis and dermis are reserved, and the hair components in the skin are removed to make a 0.4mm thick skin piece. The skin slices were cut into squares with a size of 3cm×3cm, and the small squares were immersed in 1mmol / L NaCl solution, shaken at a constant temperature at 37°C for 24 hours to remove the epidermis, and washed repeatedly with PBS buffer. Then add it to 0.1wt% NP-40, and shake continuously for 24 hours on a shaker at room temperature. After removing the residual cells and their fragments, they are washed repeatedly with PBS buffer. Then immerse in 0.25wt% trypsin-EDTA digestion solution for 20 minutes, and rinse with PBS buffer solution repeatedly to obtain sheep acellular dermal matrix scaffold.

[0038] The obtained sheep acellular dermal matrix scaffold is stored in sterile PBS b...

Embodiment 3

[0041] Example 3: Construction of composite tissue engineered skin.

[0042] Spread the sheep acellular dermal matrix scaffold prepared in Example 2 with the dermis facing upwards in a 35mm culture dish, take the second-generation epidermal stem cells with strong proliferative power amplified in Example 1, digest with trypsin for 20min, and stop the serum , to 1 x 10 6 / cm 2 Inoculate the above-mentioned sheep acellular dermal matrix scaffold at a density of 10%, add enough K-SFM medium containing 10% fetal bovine serum to completely immerse the dermal matrix scaffold, and store at 37°C, 5% CO 2 Submerged culture in the incubator for 1 week, during which the medium was changed every 2 days. Afterwards, the tissue-engineered skin obtained by immersion culture was transferred to a stainless steel grid, and fresh K-SFM medium containing 10% fetal bovine serum was added to make the liquid level of the medium flush with the bottom of the dermal matrix scaffold. In this way, the ...

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Abstract

The invention discloses a method for constructing composite tissue engineering skins. The method includes that human epidermal stem cells are used as seed cells, sheep accellular dermal matrixes are used as scaffold carriers, and the composite tissue engineering skins can be constructed by means of in-vitro culture by the aid of gas-liquid interface separation processes. The method has the advantages that the composite tissue engineering skins constructed by the aid of the method have perfect three-dimensional netted structures, and accordingly growth of cells and formation of new collagen fibers can be promoted; the composite tissue engineering skins are short in construction time and complete in histological structure and are similar to natural skins, and morphology requirements on tissue engineering skins can be met.

Description

technical field [0001] The invention belongs to the technical field of tissue engineering skin and relates to a construction method of composite tissue engineering skin. Background technique [0002] Skin is the largest organ of the human body, covering almost the whole body, with complex structure and function. However, as the first barrier between the human body and the environment, the skin is extremely vulnerable. Various diseases such as extensive burns, trauma, chronic skin ulcers, and scar excision can cause skin defects, which require different methods of repair. The human body itself has a certain repair ability, which can realize the self-recovery of damaged skin structure and function to varying degrees. However, when the damaged area of ​​the skin is large and the damage is severe, surgery must be used to ensure the reconstruction of the skin structure and function. [0003] Skin transplantation requires sufficient skin sources, and autografting and allografti...

Claims

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Application Information

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IPC IPC(8): A61L27/38A61L27/36A61L27/60
CPCA61L27/3633A61L27/3687A61L27/3804A61L27/3895A61L27/60
Inventor 解军张琪孟凡秀于保锋曹睿温晓薇杨巧艳加三三温丽敏白欣艳姚志坚曾思衡袁洋洋张紫燕张丽娜王萱田九博马培元刘志贞傅松涛杨丽红郭睿王惠珍徐钧刁海鹏陈显久胡晓年弓韬王海龙赵虹
Owner SHANXI MEDICAL UNIV
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