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A method for constructing a recombinant epidermal model for in vitro skin irritation detection

A skin irritation and construction method technology, applied in biochemical equipment and methods, epidermal cells/skin cells, tissue culture, etc., can solve misjudgment, insignificant improvement of lipid lamellar fine structure, and accuracy impact and other problems, to achieve the effect of simple process, high product uniformity and stability, and improved accuracy

Active Publication Date: 2020-11-20
GUANGDONG BOXI BIO TECH CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The difference in lipid composition is the reason for the abnormal secretion of lamellar body and lipid lamellar in the model stratum corneum, and the imperfect barrier function. These defects lead to the high permeability of the epidermal model to the test substance, thus affecting the in vitro safety The accuracy of the test results is affected, and there is a certain misjudgment of the judgment of the irritant standard substance. Although lipid components (such as palmitic acid, oleic acid, linoleic acid, and arachidonic acid) can be added to the culture medium, the Improve the abnormal state of lipid composition to a certain extent, but the improvement effect on the fine structure of lipid lamellar is not obvious

Method used

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  • A method for constructing a recombinant epidermal model for in vitro skin irritation detection
  • A method for constructing a recombinant epidermal model for in vitro skin irritation detection
  • A method for constructing a recombinant epidermal model for in vitro skin irritation detection

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Embodiment 1

[0027] The embodiment of the present invention provides a method for constructing a recombinant epidermal model for in vitro skin irritation detection, using the following steps:

[0028] 1. Isolation and large-scale culture of epidermal stem cells:

[0029] (1) Isolation of epidermal stem cells: put human skin tissue in a petri dish, add 10mL of 75% alcohol, rinse quickly, transfer to PBS solution, peel off the subcutaneous loose connective tissue with scissors, soak in PBS, and remove the tissue block Cut into a size of 0.3cm×0.5cm, add 15ml of Dispase digestion solution with a mass ratio of 1%, digest overnight at 4°C, separate the dermis and epidermis, collect epidermal slices, cut them into pieces, and then add 10ml with a mass ratio of 0.025% The EDTA-trypsin digestion solution was digested at 37°C for 10 minutes to obtain a solution of epidermal cells separated into single cells. The epidermal fragments were filtered with a 200-mesh stainless steel screen, centrifuged at ...

Embodiment 2

[0041] The embodiment of the present invention provides a method for constructing a recombinant epidermal model for in vitro skin irritation detection, using the following steps:

[0042] 1. Isolation and large-scale culture of epidermal stem cells:

[0043] (1) Isolation of epidermal stem cells: put human skin tissue in a petri dish, add 10mL of 75% alcohol, rinse quickly, transfer to PBS solution, peel off the subcutaneous loose connective tissue with scissors, soak in PBS, and remove the tissue block Cut into a size of 0.3cm×0.5cm, add 15ml of Dispase digestion solution with a mass ratio of 1%, digest overnight at 4°C, separate the dermis and epidermis, collect epidermal slices, cut them into pieces, and then add 10ml with a mass ratio of 0.025% The EDTA-trypsin digestion solution was digested at 37°C for 10 minutes to obtain a solution of epidermal cells separated into single cells. The epidermal fragments were filtered with a 200-mesh stainless steel screen, centrifuged a...

Embodiment 3

[0055] The embodiment of the present invention provides a method for constructing a recombinant epidermal model for in vitro skin irritation detection, using the following steps:

[0056] 1. Isolation and large-scale culture of epidermal stem cells:

[0057] (1) Isolation of epidermal stem cells: put human skin tissue in a petri dish, add 10mL of 75% alcohol, rinse quickly, transfer to PBS solution, peel off the subcutaneous loose connective tissue with scissors, soak in PBS, and remove the tissue block Cut into a size of 0.3cm×0.5cm, add 15ml of Dispase digestion solution with a mass ratio of 1%, digest overnight at 4°C, separate the dermis and epidermis, collect epidermal slices, cut them into pieces, and then add 10ml with a mass ratio of 0.025% The EDTA-trypsin digestion solution was digested at 37°C for 10 minutes to obtain a solution of epidermal cells separated into single cells, and the epidermal fragments were filtered with a 200-mesh stainless steel screen, centrifug...

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Abstract

The invention provides a recombinant epidermis model construction method used for in-vitro skin irritation detection, which relates to the technical field of tissue engineering. The method has the advantages of simple operation, stable technology and large scale production, the product lipid composition is analogous to natural human skin, the barrier function is complete, and the method is used for detecting irritation of the cosmetic. The method comprises the following steps: 1) separating epidermal stem cells and culturing the epidermal stem cells with large scale; and 2) performing tissue engineering epidermis construction.

Description

technical field [0001] The invention relates to the technical field of tissue engineering, in particular to a method for constructing a recombinant epidermal model for in vitro skin irritation detection. Background technique [0002] Cosmetics are usually in direct contact with human skin, and their safety issues have attracted much attention, so it is necessary to conduct skin irritation tests. Traditional skin irritation tests are mostly carried out on animals, which consume large batches of animals, take a long time, are expensive, and have disadvantages such as species differences, and some experiments will cause great pain to animals, which does not meet the requirements of animal protection and welfare . With the introduction of the "3R" principle, foreign countries first began to vigorously carry out research on alternative methods of animal experiments on this basis. After nearly 20 years of research on skin irritation test in vitro alternative methods, the method ...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N5/074
CPCC12N5/0625C12N2500/14C12N2500/25C12N2500/30C12N2500/32C12N2500/36C12N2500/38C12N2500/40C12N2501/11C12N2501/385C12N2501/39C12N2533/30C12N2533/54
Inventor 李润芝卢永波
Owner GUANGDONG BOXI BIO TECH CO LTD