Method for constructing skin tissue

A construction method and skin technology, applied in the field of cell culture, can solve the problems of expensive patients and restrictions on the development and application of tissue engineered skin, and achieve the effect of convenient control

Inactive Publication Date: 2018-12-21
山东麦德克斯生物科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

But the high price is unacceptable to patients, especially in China
Each large area of ​​skin transplantation (the required cost is more than 500,000 US dollars), which limits the development and application of tissue engineered skin in China

Method used

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  • Method for constructing skin tissue
  • Method for constructing skin tissue
  • Method for constructing skin tissue

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0070] A kind of keratinocyte and fibroblast, adopt following method to prepare:

[0071] Configure the following media:

[0072] Optimized Green culture solution: DMEM medium and Ham's F12 nutrient solution were mixed at a volume ratio of 3:1, and then the following components were added based on the volume after the two were mixed: FCS with a volume percentage of 10%, EGF 10ng / ml, hydrocortisone 0.4μg / ml, adenine 0.18mM, insulin 5μg / ml, transferrin 5μg / ml, glutamine 2mM, iodothyronine 0.2μM, amphotericin B 0.625μg / ml, Penicillin 100U / ml, streptomycin 100μg / ml.

[0073] 1. Skin Biopsy Sampling

[0074] 1.1 Under sterile conditions, take 1.5cm of inner thigh skin tissue 2 The size was placed in DMEM containing 10% newborn calf serum (FBS) and double antibodies (100 IU / ml penicillin, 100 ug / ml streptomycin), and stored at 4°C.

[0075] 1.2 The skin tissue was cut into three pieces of 1x5 mm in size, and washed 10 times with phosphate-buffered saline (PBS) containing 50 U / m...

Embodiment 2

[0099] A method for constructing skin tissue, comprising the steps of:

[0100] Configure the following media:

[0101] Optimized Green culture solution: DMEM medium and Ham's F12 nutrient solution were mixed at a volume ratio of 3:1, and then the following components were added based on the volume after the two were mixed: FCS with a volume percentage of 10%, EGF 10ng / ml, hydrocortisone 0.4μg / ml, adenine 0.18mM, insulin 5μg / ml, transferrin 5μg / ml, glutamine 2mM, iodothyronine 0.2μM, amphotericin B 0.625μg / ml, Penicillin 100U / ml, streptomycin 100μg / ml.

[0102] The fibroblasts and keratinocytes prepared in Example 1 were used.

[0103] Seed fibroblasts in scaffolds at a density of 5x10 5 / cm 2 , add 1ml of optimized Green culture medium to the scaffold.

[0104] Add 2ml of RPMI1640 culture solution around the scaffold.

[0105] 37°C, 5% CO 2 Cultured for 3 days, changing the culture medium every 2 days.

[0106] On the fourth day, keratinocytes (1x10 6 cells / ml), adh...

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Abstract

The invention relates to the field of cell culture, in particular to a method for constructing skin tissue, comprising the following steps: planting fibroblasts in a scaffold of a three-dimensional flow bioreactor, and adding an optimized Green culture solution into the scaffold; adding an RPMI1640 culture solution around the scaffold, culturing for 90-110 h, and then adding keratinocytes to the surface of the scaffold for culture; regularly changing the culture solution after a gas-liquid interface is formed and culturing for another 21 plus or minus 1 days. In the method, the fibroblasts andthe keratinocytes are jointly and stereoscopically cultured in the three-dimensional flow bioreactor to obtain the desired skin tissue available for skin transplantation.

Description

technical field [0001] The invention relates to the field of cell culture, in particular to a method for constructing skin tissue. Background technique [0002] The skin is the largest organ of the human body. The leading cause of extensive skin loss is burns and scalds. According to the statistical survey in 1952, among people aged 15-44, when more than 60% of the total body surface area was lost, the mortality rate was 100% (Bulland Fischer, 1954); but the mortality rate dropped to 41.4% in 2003 (Chua et al 2003). The main reason for the reduction in mortality is the improvement of treatment technology. Currently, the treatments for large area of ​​skin loss are mainly autologous free skin grafts and synthetic skin grafts. [0003] Synthetic skin can help patients in time and save lives. Tissue-engineered skin has high cytocompatibility, has more obvious advantages, and is more suitable for patients. But the high price is unacceptable to patients, especially in China...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N5/071
CPCC12N5/0625C12N5/0629C12N5/0698C12N2500/25C12N2500/32C12N2500/40C12N2501/11C12N2501/395C12N2501/855C12N2502/09C12N2502/094C12N2509/00C12N2513/00
Inventor 张辉赵鸿李涛张玉光王旭左岩霞张兆蕾慈鹏
Owner 山东麦德克斯生物科技有限公司
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