Method and application for preparing tissue engineering corneal carrier stent by utilizing fresh porcine cornea

A technology of tissue engineering and porcine cornea, applied in prosthetics, medical science, etc., can solve the problems of unsatisfactory biocompatibility, insufficient structure density, large degree of structural damage, etc., and achieve easy attachment and growth, strong mechanical properties , the effect of low production cost

Active Publication Date: 2014-12-10
青岛彩晖生物科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

For example, the source of human-derived natural biological materials is limited, which cannot meet the needs of large-scale reconstruction of tissue-engineered corneas in vitro, and the tissue-engineered corneal carrier scaffolds made of natural biological materials such as heterogeneous (pig and other animal sources) corneal stroma after decellularization treatment have also been reported. There are many defects such as large structural damage, low transparency, and poor mechanical properties.
And some use detergents such as sodium lauryl sulfate, triton X-100 (Triton X-100) in the process of preparing acellular keratoma sheet, and some use

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0033]First get fresh pig eyeballs, rinse them with 0.9% (w / v) normal saline, and cut out a corneal piece with a thickness of 100 microns with the Bowman's membrane with a microkeratome;

[0034] Then take 800 milliliters of ultrapure water, add 1 gram of KCl, and dilute to 1000 milliliters with ultrapure water after completely dissolving to prepare a hypotonic solution; rinse the corneal piece with the hypotonic solution, immerse it in the hypotonic solution, and dry it at 37 Swell in a shaker at ℃ for 2 hours; after that, carefully peel off the epithelial layer of the corneal sheet with tweezers to obtain the corneal stroma sheet;

[0035] Then take 800 ml of ultrapure water, add 15.764 g of Tris-HCl, after completely dissolving, adjust the pH value to 7.5, and dilute to 1000 ml with ultrapure water to prepare a rinsing buffer; In a 50ml centrifuge tube of the buffer solution, freeze at -80°C for 1 hour, then thaw at 37°C for 10 minutes, repeat the freeze-thaw 3 times; then ...

Embodiment 2

[0040] First get fresh pig eyeball, after washing with 0.9% (w / v) normal saline, use microkeratome to cut out 300 micron thick corneal slices with Bowman's membrane;

[0041] Then take 800 milliliters of ultrapure water, add 2 grams of KCl, and dilute to 1000 milliliters with ultrapure water after completely dissolving to prepare a hypotonic solution; rinse the corneal piece with the hypotonic solution, immerse it in the hypotonic solution, and put it at 37 Swell in a shaker at ℃ for 2.5 hours; after that, carefully peel off the epithelial layer of the corneal sheet with tweezers to obtain the corneal stroma sheet;

[0042] Then take 800 ml of ultrapure water, add 15.764 g of Tris-HCl, after completely dissolving, adjust the pH value to 7.5, and dilute to 1000 ml with ultrapure water to prepare a rinsing buffer; In a 50ml centrifuge tube of buffer solution, freeze at -80°C for 1.5 hours, then thaw at 37°C for 15 minutes, repeat the freeze-thaw 4 times; then rinse twice with wa...

Embodiment 3

[0047] First take fresh pig eyeballs, wash them with 0.9% (w / v) normal saline, and cut out a corneal sheet with a thickness of 500 microns with Descemet's membrane with a microkeratome;

[0048] Then take 800 milliliters of ultrapure water, add 3 grams of KCl, and dilute to 1000 milliliters with ultrapure water after completely dissolving to prepare a hypotonic solution; rinse the corneal piece with the hypotonic solution, immerse it in the hypotonic solution, and dry it at 37 Swell in a shaker at ℃ for 3 hours; after that, carefully peel off the endothelial layer of the cornea with tweezers to obtain a corneal stroma sheet;

[0049] Then take 800 ml of ultrapure water, add 15.764 g of Tris-HCl, after completely dissolving, adjust the pH value to 7.5, and dilute to 1000 ml with ultrapure water to prepare a rinsing buffer; In a 50ml centrifuge tube of buffer solution, freeze at -80°C for 2 hours, then thaw at 37°C for 30 minutes, repeat the freeze-thaw 5 times; then rinse twice...

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Abstract

The invention discloses a method and application for preparing a tissue engineering corneal carrier stent by utilizing fresh porcine cornea. The method comprises the following steps: cutting a fresh porcine cornea for swelling, repeatedly freeze thawing and breaking cells, removing residual nucleic acid substances through DNA-RNA enzyme treatment, cross-linking, airing, and finally performing irradiation sterilization through ionization rays, thereby obtaining the product. The method is applied to carrier stents of tissue engineering corneal epithelium, stroma, endothelium, front board layer half cornea, rear board layer half cornea and full-layer board layer half cornea, so that the requirement on batch production of tissue engineering corneas is met. The carrier stent serves as a substitute of human corneal stroma to be used for clinical transplant and treatment of un-accumulated whole layer keratohelcosis. The method is applied to large-scale production and clinical popularization and application. The prepared carrier stent has the characteristics of high transparency, dense structure, high mechanical property, high biocompatibility with human corneal seed cells and the like. Therefore, the method is suitable for batch production, so that the requirements on lots of carrier stents for tissue engineering corneas are met.

Description

technical field [0001] The invention relates to a method and application for preparing a tissue engineering cornea carrier bracket by using fresh pig cornea, and belongs to the preparation technology of the cornea carrier bracket in the field of biological tissue engineering. Background technique [0002] The cornea is a transparent membrane located at the very front of the eyeball, covering the iris, pupil, and anterior chamber, and provides most of the refractive power for the eye. Because the cornea is in direct contact with the external environment, it is extremely vulnerable to mechanical trauma, thermal burns, and microbial infections, which can cause corneal lesions. In mild cases, the transparency of the cornea will decrease and affect the patient's vision. In severe cases, it will cause corneal opacity and blindness. According to WHO statistics, there are currently nearly 60 million blind patients with corneal disease in the world, and nearly 5 million people in my ...

Claims

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Application Information

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IPC IPC(8): A61L27/36
Inventor 樊廷俊于苗苗徐彬庞鑫邱月
Owner 青岛彩晖生物科技有限公司
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