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Cell culture method using plant epidermis as tissue engineering scaffold material

A technology of tissue engineering scaffold and plant epidermis, which is applied in the field of cell culture, can solve the problems of spreading diseases, causing inflammatory reactions, accumulation of local acid products, improper handling, etc., and achieves good transparency and is conducive to the growth and arrangement of stratification tight effect

Active Publication Date: 2017-08-11
XIAMEN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The most mature material is amniotic membrane, but the use of amniotic membrane has the risk of improper handling and disease transmission; collagen is a natural degradable material for ocular surface reconstruction, but its poor stability and rapid degradation limit its development; chitosan is Another natural scaffold material, because chitosan macromolecules have a stable ring structure, strong hydrogen bond interaction, and poor solubility, so it is less used in tissue engineering scaffold materials
Synthetic polymer materials such as polylactic acid (PLA) and polyglycolic acid (PGA) have good biocompatibility and can undergo non-enzymatic hydrolysis, but their hydrophilicity is poor, and the metabolic process in the body is not completely clear, which can cause Inflammatory response, accumulation of local acid products, etc., need to be further studied if they are to be used as scaffold materials for tissue engineering corneas
In short, the current biomaterials have good histocompatibility, but poor plasticity; synthetic materials have good mechanical properties, but are cytotoxic; composite materials combine the advantages of biomaterials and synthetic materials, but their new properties further research is needed

Method used

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  • Cell culture method using plant epidermis as tissue engineering scaffold material
  • Cell culture method using plant epidermis as tissue engineering scaffold material
  • Cell culture method using plant epidermis as tissue engineering scaffold material

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0031] Example 1: Rabbit Primary Corneal Epithelial Cell Culture

[0032] Among the present embodiment, the medium and reagents used are as follows:

[0033] SHEM medium: 5% FBS (fetal bovine serum), 0.5g / mL hydrocortisone, 1% ITS (insulin, transferrin and selenium solution), 0.5% DMSO (di methyl sulfoxide), 10 ng / mL human EGF (human epidermal growth factor), 1% PS (P means Penicillin penicillin, S means Streptomycin streptomycin, and the respective concentrations of penicillin and streptomycin are 1%).

[0034] Dispase (neutral protease) II digestion solution: commercially available, working concentration 2U / mL.

[0035] 10×PBS: NaCl 20g, KCl 2g, NaCl 2 HPO 4 12H 2 O 18.2g, KH 2 PO 4 2.4g, stir evenly with a stirrer, adjust the pH to 7.2-7.4, dilute it into 1×PBS and sterilize before use.

[0036] Among the present embodiment, the onion bulb inner epidermis is selected, and the rabbit primary corneal epithelial cell culture method using the onion bulb inner epidermis ...

Embodiment 2

[0047] Example 2: Rabbit Primary Corneal Stromal Cell Culture

[0048] Among the present embodiment, the culture medium and culture conditions adopted are as follows:

[0049] Medium: DMEM (basic), 10% FBS, 1% PS; culture conditions: 37°C, 5% CO 2 incubator.

[0050] In this example, the preparation method of the onion epidermal scaffold is the same as step 1) in Example 1.

[0051] Such as Figure 6 Shown on the left side of the middle is a schematic diagram of the comparison of cell culture of rabbit primary corneal stromal cells (Rabbit Corneal StromaCells P0) on onion epidermal scaffolds and ordinary culture dishes in this embodiment. After 4-6 days of culture, the primary rabbit keratocytes grown on the onion epidermis could well maintain the shape of the keratocytes, and there was no obvious difference in shape compared with the cells on the culture dish, but the cell fusion rate was higher.

Embodiment 3

[0052] Embodiment 3: breast cancer cell culture

[0053] Among the present embodiment, the culture medium and culture conditions adopted are as follows:

[0054] Medium: DMEM (basic), 10% FBS, 1% PS; culture conditions: 37°C, 5% CO 2 incubator.

[0055] In this example, the preparation method of the onion epidermal scaffold is the same as step 1) in Example 1.

[0056] Breast cancer cell MDA-MB-231 was cultured and digested by conventional cell culture methods according to the above conditions to obtain a cell suspension, and then 5-8×10 4 piece / cm 2 Inoculated on the onion epidermis support, cultivated according to the method of step 6) in Example 1. At the same time, cells cultured in ordinary culture dishes were used as a control.

[0057] After culturing for 1 to 2 days, the results are as follows: Figure 6 As shown in the middle right, the cell morphology of the onion epidermal scaffold group has not changed compared with that of the ordinary culture dish group, in...

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Abstract

The invention discloses a cell culture method using a plant epidermis as a tissue engineering scaffold material. The scaffold material selecting onion bulb endepidermis and other plant epidermises has the advantages of non-toxicity, small immunogenicity, good biocompatibility, high transparency, certain mechanical strength, good plasticity, low price and easiness in material drawing. The proliferation rate of the cells in the plant epidermis is similar to that of existing scaffold materials, and the corneal epithelial cells can form an epithelium-like structure on the plant epidermis and are closely arranged, so the plant epidermis provides a good basis for cell proliferation and differentiation, and the plant epidermis is better than existing tissue engineering scaffold materials.

Description

technical field [0001] The invention belongs to the technical field of cell culture, and in particular relates to a cell culture method using plant epidermis as a tissue engineering support material. Background technique [0002] Tissue engineering materials are the basis of tissue engineering. Scaffold materials for tissue engineering can provide the basis for cell proliferation and differentiation. The characteristics of scaffold materials determine the shape of the constructed tissue. In recent years, people have been looking for scaffold materials with good biocompatibility, good mechanical properties, and strong light transmission that are suitable for constructing tissue-engineered corneas. The currently used scaffold materials mainly include biological, synthetic, and composite materials, and there are also scaffold-free culture methods. The most mature material is amniotic membrane, but the use of amniotic membrane has the risk of improper handling and disease trans...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N5/071C12N5/077C12N5/079C12N5/09
CPCC12N5/0621C12N5/0625C12N5/0656C12N5/069C12N5/0693C12N2533/90
Inventor 李程陈佩王国良薛玉花刘祖国
Owner XIAMEN UNIV
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