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Method for preparing full-thickness skin for toxicity test by stem cell raft type cultivation

A full-thickness skin and stem cell technology, applied in the field of toxicology tests, can solve the problems of short skin function maintenance time, inapplicability to chronic toxicity tests, long culture period, etc., to achieve small batch-to-batch differences, complete morphology and tissue structure, and maintain long time effect

Inactive Publication Date: 2012-03-07
程树军 +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, these skin substitutes are mainly used for the clinical treatment of patients with skin defects such as burns, ulcers, and wounds, and cannot replace the mature skin of live animals for skin toxicity testing.
As tissue engineered skin, there are also the following disadvantages: the artificial epidermis is not a real artificial skin because it does not contain a dermis; the existing artificial skin tissue structure with a double-layer structure needs to be improved; Short maintenance time in vitro, not suitable for chronic toxicity test

Method used

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  • Method for preparing full-thickness skin for toxicity test by stem cell raft type cultivation
  • Method for preparing full-thickness skin for toxicity test by stem cell raft type cultivation
  • Method for preparing full-thickness skin for toxicity test by stem cell raft type cultivation

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Experimental program
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Effect test

Embodiment 1

[0043] Include the following steps in turn:

[0044] 1. Preparation and modification of polymer dermal scaffolds

[0045] 1) Preparation of PHBV scaffold:

[0046] (a) Dissolving an appropriate amount of PHBV powder in chloroform to make a solution with a concentration of 120-130g / L, then adding 940g of sodium chloride (NaCl) to each liter of the solution as a porogen, fully mixing to obtain a thick slurry;

[0047] (b) Pour the thick slurry into a circular metal mold with a hole diameter of 0.8 to 1.0 cm. After about 80% of the chloroform solvent is volatilized, take out the forming bracket from the mold, and then place it in a fume hood at room temperature for 48 hours to dry. Vacuum drying to remove residual solvent;

[0048] (c) Immerse the formed scaffold in deionized water, change the water once every 6 hours, and change the water 8 times within 48 hours until the porogen is filtered out, dry at room temperature and vacuum dry for 24 hours to make a PHBV porous three-...

Embodiment 2

[0071] 1. Preparation and modification of polymer dermal scaffolds

[0072] 1) Preparation of PHBV scaffold:

[0073] (a) dissolving an appropriate amount of PHBV powder in chloroform to make a solution with a concentration of 130g / L, then adding 930g of sodium chloride (NaCl) to each liter of solution as a porogen, fully mixing to obtain a thick slurry;

[0074] (b) Pour the thick slurry into 0.15-0.2ml of a circular metal mold with a diameter of 1.0cm. After the solvent volatilizes, take out the forming bracket from the mold, place it in a fume hood at room temperature for 54 hours, and dry it in vacuum to remove the residual solvent. ;

[0075] (c) Immerse the formed scaffold in deionized water, change the water once every 7 hours, change the water 7 times within 49 hours until the porogen is filtered out, dry at room temperature and then vacuum dry for 24 hours to make a PHBV porous three-dimensional scaffold ;

[0076] (d) Soak the PHBV porous three-dimensional scaffol...

Embodiment 3

[0094] 1. Preparation and modification of polymer dermal scaffolds

[0095] 1) Preparation of PHBV scaffold:

[0096] (a) Dissolving an appropriate amount of PHBV powder in chloroform to make a solution with a concentration of 130g / L, then adding 940g of sodium chloride (NaCl) to each liter of solution as a porogen, fully mixing to obtain a thick slurry;

[0097] (b) Pour the thick slurry into 0.15-0.2ml of a circular metal mold with a diameter of 1.0cm. After the solvent volatilizes, take out the forming bracket from the mold, place it in a fume hood at room temperature for 54 hours, and dry it in vacuum to remove the residual solvent. ;

[0098] (c) Immerse the formed scaffold in deionized water, change the water once every 8 hours, and change the water 7 times within 56 hours until the porogen is filtered out, dry at room temperature and vacuum dry for 24 hours to make a PHBV porous three-dimensional scaffold ;

[0099] (d) Soak the PHBV porous three-dimensional scaffold...

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Abstract

The invention relates to a method for preparing full-thickness skin used in toxicity testing by adopting stem cell raft culture, which comprises: 1. preparing and modifying a polymeric dermal scaffold; 2. preparing epidermal stem cells from embryonic stem cells and skin tissues; 3. preparing pancreatic islet cells; 4. preparing a special medium for the full-thickness skin; 5. and constructing the full-thickness skin. By utilizing the characteristics of strong differentiation and proliferation capacity of the epidermal stem cell, the skin morphology, organizational structure and functional activity of the constructed full-thickness skin of the invention can meet the demands of sub-chronic toxicity testing; the synthetic scaffold material has high degree of standardization and small batch-to-batch variation; as a serum free medium is adopted through the skin construction process, the factors influencing tissue construction are reduced, thus providing a foundation for the future use in toxicity testing. The all-thickness skin prepared by the method of the invention is closer to a natural skin, the application of which in the mode of toxicity testing and detection index agrees with practical situation better, thus being able to replace animals to be directly applied to the skin toxicity testing of chemicals, cosmetics, medicines and other health-related products.

Description

technical field [0001] The invention relates to a method for constructing full-thickness replacement skin by using stem cell tissue engineering technology, which can be used to replace living animal (or human) skin in the field of toxicology tests. Background technique [0002] The skin is the largest organ of the human body, and it is the main organ for the toxic effects of exogenous chemical factors (chemicals, drugs, cosmetics, etc.) or physical and mechanical factors (ultraviolet rays, microwaves, etc.). Skin toxicity testing using animal or human skin is a routine testing item for health and safety evaluation of cosmetics, pharmaceuticals, food additives and raw materials, pesticides, biological products, and chemicals. Traditional skin safety evaluation experiments not only require a certain number of high-quality experimental animals, but also have a long test period and high cost, and some reactions will cause great pain to the animals. Countries around the world at...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N5/071
Inventor 程树军焦红黄亚东秦瑶
Owner 程树军
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