Preparation method and application of acellular matrix biological material

An acellular matrix and biomaterial technology, which is applied in the field of the preparation of acellular matrix biomaterials and can solve problems such as uncertain clinical effects

Inactive Publication Date: 2015-11-04
武汉蓝普医品有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

There are many problems in the cell therapy of stem cells for tissue and organ repair. Although there are clinical application reports, the clinical effect is uncertain

Method used

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  • Preparation method and application of acellular matrix biological material

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0046] Example 1: Preparation of dermis-based acellular matrix material:

[0047] 1) The skin is taken from the skin of the above-mentioned animals, which can be taken from dead bodies and excess skin from surgical procedures;

[0048] 2) Separate the epidermis and dermis: soak the skin in 0.25% trypsin and 1mM EDTA solution for 3 hours to digest;

[0049] 3) The epidermis is separated from the dermis by mechanical means;

[0050] 4) decellularization treatment;

[0051] 5) Decellularization solution preparation: 0.125% trypsin, 1mM EDTA, 0.25% Triton X-100;

[0052] 6) Soak the corium obtained above in the decellularized solution and digest it with shaking at 37 degrees for 3-4 hours;

[0053] 7) Rinse with PBS, and then soak in phosphate-buffered saline (PBS) containing 0.1% sodium azide for several minutes to several days;

[0054] 8) Sterilization treatment: the obtained dermal matrix is ​​irradiated with gamma rays to achieve the purpose of sterility.

Embodiment 2

[0055]Example 2: Preparation of bladder-based acellular matrix material:

[0056] 1) Place the bladder tissue in a petri dish, add 50ml of PBS containing 0.1% sodium azide, and scrape off the mucosal layer with a glass slide;

[0057] 2) The remaining muscle layer and submucosa continued to remain in the above solution for 5 hours to dissolve the cells;

[0058] 3) Rinse the bladder tissue with 50ml PBS;

[0059] 4) DNase digestion: digest the bladder with 50 ml of 1 M sodium chloride solution containing 2000 units of DNase (Sigma Company) for 12 hours. Then remove the solution, add new digestion solution and repeat 3 times;

[0060] 5) Then treat with 50 ml of 4% sodium deoxycholate (sodium deoxycholatel) containing 0.1% sodium azide for 6 hours and stir, and repeat this process a total of 3 times;

[0061] 6) The obtained acellular matrix is ​​kept at 4°C in a solution containing 0.1% sodium azide and 10% neomycin sulfate;

[0062] 7) Sterilization treatment: the obtaine...

Embodiment 3

[0063] Embodiment 3: adipose stem cell is free:

[0064] 1) Adipose tissue or fat obtained by liposuction;

[0065] 2) Rinse with PBS containing 1% penicillin and streptomycin;

[0066] 3) Cut the tissue into small pieces of 1 mm square with a blade;

[0067] 4) Add 0.075% collagenase type 1A (collagenase type IA, Sigma company) at the ratio of solution:tissue 5:1;

[0068] 5) Incubate with a shaker at 37 degrees for 1 hour;

[0069] 6) Centrifuge at 250 g for 10 minutes at room temperature. At this time, the solution is divided into three layers, and the lower layer is the digested cell pellet;

[0070] 7) Remove the upper two layers and keep the cell layer;

[0071] 8) Resuspend the cells in adipose tissue maintenance solution containing 200mg / ml KH2PO4, 200mg / L KCl, 2.16g / L Na 2 HPO 4 .7H 2 O, 8g / L NaCl, 30,000 units / L SOD, 5g / L human serum albumin.

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Abstract

The invention relates to a preparation method of an acellular matrix biological material. The method comprises the following steps: obtaining tissues; carrying out acellular treatment; carrying out bactericidal treatment; harvesting an acellular matrix material; obtaining adipose tissues; dissociating adipose-derived stem cells; harvesting the adipose-derived stem cells; planting the adipose-derived stem cells; and harvesting the acellular matrix biological material. The invention further provides an application of the acellular matrix biological material. The acellular matrix biological material can be applied to regeneration and repairing of damaged tissues, covering and repairing of burn wounds, repairing of bladders and urethra substitution.

Description

technical field [0001] The invention belongs to the field of biotechnology, and in particular relates to a preparation method of an acellular matrix biological material. Background technique [0002] Stem cells are primitive cells with self-renewal ability and multilineage differentiation potential, and are ideal cells for tissue regeneration and repair after cell transplantation. Although embryonic stem cells are the most primitive stem cells with the most differentiation potential, their sources are currently very limited. Stem cells induced by high expression of stem cell factors, also known as iPS cells, have many characteristics of embryonic stem cells, but due to the complicated preparation methods and safety issues such as the use of viral vectors, there are still many bottlenecks in clinical application. Mesenchymal stem cells (Mesenchymal stem cells, MSC) is a sub-totipotent stem cell population with multilineage differentiation potential existing in human tissues,...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A61L27/36A61L27/38A61L27/50A61L27/54
Inventor 李龙承沈乔李辉李先华沈传宝
Owner 武汉蓝普医品有限公司
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