Composite membrane based on gelatin and amino acid, and method for cultivating limbal stem cells on membrane

An amino acid and composite membrane technology, applied in biochemical equipment and methods, animal cells, tissue culture, etc., can solve the problems of poor transplantation effect, exogenous cells that easily cause immune reactions, thick thickness, etc., and achieve high biophase Capacitance, promoting cell differentiation, and facilitating fine-tuning effects

Active Publication Date: 2017-09-01
ZHONGSHAN OPHTHALMIC CENT SUN YAT SEN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Due to limited technical level, limbal stem cells cultured in vitro are too thick, exogenous cells are likely to cause immune reactions and other shortcomings, making the transplantation effect not good

Method used

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  • Composite membrane based on gelatin and amino acid, and method for cultivating limbal stem cells on membrane
  • Composite membrane based on gelatin and amino acid, and method for cultivating limbal stem cells on membrane
  • Composite membrane based on gelatin and amino acid, and method for cultivating limbal stem cells on membrane

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0056] Embodiment 1 prepares composite film

[0057] After dissolving 10% (W / V) gelatin PBS solution in a water bath at 10°C, add methacrylic anhydride with a volume ratio of 5% at a rate of 0.5 mL / min, stir and react in a water bath at 50°C for 24 hours, and then add 5 Double the amount of DPBS to stop the reaction. The obtained liquid is placed in a 8-14KD dialysis bag, dialyzed in distilled water for 3 days, and then freeze-dried to obtain a white porous foam product, which is the Gelma material, which is stored in a dry place at low temperature.

[0058] Mix PEG and benzene at a ratio of 20% (W / V), distill in an oil bath to remove water, cool to room temperature, add triethylamine and acryloyl chloride to react in an ice bath for 48 hours in the dark (PEG: triethylamine: acryloyl chloride) The molar ratio is 1:4:4). After the reaction, the insoluble salts were removed by suction filtration, and the filtrate was post-treated with glacial ether. The resulting solid product ...

Embodiment 2

[0061] Embodiment 2 prepares composite film

[0062] After dissolving 10% (W / V) gelatin PBS solution in a water bath at 10°C, add methacrylic anhydride with a volume ratio of 5% at a rate of 0.5 mL / min, stir and react in a water bath at 50°C for 24 hours, and then add 5 Double the amount of DPBS to stop the reaction. The obtained liquid is placed in a 8-14KD dialysis bag, dialyzed in distilled water for 3 days, and then freeze-dried to obtain a white porous foam product, which is the Gelma material, which is stored in a dry place at low temperature.

[0063] Mix PEG and benzene at a ratio of 20% (W / V), distill in an oil bath to remove water and cool to room temperature, add triethylamine and acryloyl chloride to react in an ice bath for 48 hours in the dark (PEG: triethylamine: acryloyl chloride) The molar ratio is 1:4:4). After the reaction, the insoluble salts were removed by suction filtration, and the filtrate was post-treated with glacial ether. The resulting solid produ...

Embodiment 3

[0066] Embodiment 3 prepares composite film

[0067] After dissolving 10% (W / V) gelatin PBS solution in a water bath at 10°C, add methacrylic anhydride with a volume ratio of 5% at a rate of 0.5 mL / min, stir and react in a water bath at 50°C for 24 hours, and then add 5 Double the amount of DPBS to stop the reaction. The obtained liquid is placed in a 8-14KD dialysis bag, dialyzed in distilled water for 3 days, and then freeze-dried to obtain a white porous foam product, which is the Gelma material, which is stored in a dry place at low temperature.

[0068] Mix PEG and benzene at a ratio of 20% (W / V), distill in an oil bath to remove water and cool to room temperature, add triethylamine and acryloyl chloride to react in an ice bath for 48 hours in the dark (PEG: triethylamine: acryloyl chloride) The molar ratio is 1:4:4). After the reaction, the insoluble salts were removed by suction filtration, and the filtrate was post-treated with glacial ether. The resulting solid produ...

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Abstract

The invention discloses a composite membrane based on gelatin and amino acid, and a method for cultivating limbal stem cells (LSCs) on the membrane. A preparation method of the composite membrane comprises the steps of preparing methacrylic anhydride gelatin (Gelma) by utilizing the gelatin, methacrylic anhydride, Dulbecco phosphate buffered saline (DPBS) and distilled water under a controlled condition, preparing poly(ethylene glycol) diacrylate (PEGDA) by utilizing polyethylene glycol (PEG), benzene, triethylamine, acryloyl chloride and glacial ether under a controlled condition, obtaining an unsaturated polymer (U-PEA) by utilizing polymerization of amino acid, diol and diacid under a controlled condition or modification of methacrylic anhydride, and finally preparing the composite membrane by utilizing a reaction product by the action of a photoinitiator. The composite membrane can serve as a matrix layer to support growth of the LSCs on the membrane. The method is very simple and convenient to operate, simple in condition and good in repeatability; a prepared product is reliable in performance; and industrialization is very easy to realize.

Description

technical field [0001] The invention belongs to the technical field of tissue engineering. More specifically, it relates to a composite film based on gelatin and amino acids and a method for culturing limbal stem cells on the film. Background technique [0002] The cornea is a layer of transparent membrane that exists at the front of the eyeball and has a certain radius of curvature in space. From a physiological point of view, the outer layer of the cornea contains continuous smooth epithelial cells in the middle and limbal stem cells (LSCs) in the edge. There is a layer of stroma in the middle, which accounts for 90% of the corneal thickness. Collagen type, type five, adhesives and keratinocytes. The endothelial layer is formed by a layer of hexagonal endothelial cells and is directly connected to the aqueous humor. Limbal stem cells can continuously differentiate corneal epithelial cells to the center of the cornea, and play an important role in maintaining corneal tra...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C08L77/08C08L89/00C08L71/02C08J5/18C08J3/28C08J3/24C12N5/0775
CPCC08J3/24C08J3/28C08J5/18C08J2377/08C08J2471/02C08J2489/00C08L77/08C08L2203/02C08L2203/16C08L2205/03C12N5/0662C12N2501/01C12N2501/11C12N2501/30C12N2501/33C12N2501/39C08L89/00C08L71/02
Inventor 欧阳宏吴钧
Owner ZHONGSHAN OPHTHALMIC CENT SUN YAT SEN UNIV
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