Cornea metaphase preservation solution, and preparing and using methods thereof

A technology of preservation solution and cornea, which is applied in the preservation, application, and animal husbandry of human or animal bodies. It can solve the problems of new blood vessel formation, persistent wound healing, and corneal stroma turbidity, so as to improve cloning ability and enhance survival ability. , to avoid the effect of apoptosis

Inactive Publication Date: 2013-04-24
SHANDONG EYE INST
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  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The corneal epithelium is an important barrier of the ocular surface, and its integrity has an important impact on the success of corneal transplantation, especially corneal limbal tissue t

Method used

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  • Cornea metaphase preservation solution, and preparing and using methods thereof
  • Cornea metaphase preservation solution, and preparing and using methods thereof
  • Cornea metaphase preservation solution, and preparing and using methods thereof

Examples

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preparation example Construction

[0013] 1. Preparation of medium-term corneal preservation solution

[0014] The preservation solution is MEM cell culture medium (Gibco, USA) supplemented with chondroitin sulfate, low molecular weight dextran, L-glutamine, dexamethasone, tobramycin, Hepes and Y-27632. Concentrations are as follows: Chondroitin Sulfate 25g / L (purchased from Sigma, USA), Low Molecular Dextran 10g / L (Sigma, USA), L-Glutamine 0.375 mg / L (Gibco, USA), Dexamethasone 20mg / L L. Tobramycin 100 mg / L, Hepes 9.5 g / L (Sigma, USA), Y-27632 10 μM / L (Sigma, USA).

[0015] A kind of preparation method step is as follows:

[0016] 1) Take 9.4g of MEM powder and add it to 500ml of water for injection, dissolve it and sterilize it under high pressure.

[0017] 2) Take 25g of chondroitin sulfate, add 200ml of water for injection, heat, dissolve and autoclave to make a chondroitin sulfate solution with a concentration of 0.125g / ml; add 10g of low-molecular-weight dextran to 100ml of water for injection to dissol...

Embodiment 1

[0037] Example 1 Corneal Medium-term Preservation Solution Preserves Human Limbal Stem Cell Rings for Ring Stem Cell Transplantation

[0038] Fresh corneal rings taken after clinical transplantation are stored in the Rho kinase inhibitor corneal preservation solution of the present invention, and stored in a refrigerator at 4°C for 2 weeks. For patients who need to undergo stem cell ring transplantation in clinical practice, routine preoperative preparations include taking out the preserved corneal ring, rinsing with sterile saline, cutting off the diseased tissue, and performing ring stem cell transplantation. After the operation, the transplanted corneal ring healed well, without infection and hyperemia, and the corneal ring was transparent.

Embodiment 2

[0039] Example 2 Preservation of human corneal limbal stem cells in medium-term corneal preservation solution for transplantation of amniotic membrane cultured stem cells

[0040] Fresh corneal rings after clinical transplantation were stored in Rho Kinase Inhibitor Corneal Preservation Solution and refrigerated at 4°C for 2 weeks. When it is clinically necessary to do amnion cultured stem cell patients, take out the corneal ring, immerse it in 1.2U / ml Dispase II, take it out after overnight at 4°C, separate the corneal limbal epithelial cells under a dissecting microscope, add 0.25% trypsin-0.02% EDTA, Digest at 37°C for 15min, centrifuge at 1500r / min for 5min, add limbal stem cell culture medium to make a single cell suspension, inoculate on the pre-prepared amniotic membrane sleeve, 37°C, 5% CO 2 Culture for about 10 days, air-liquid culture for 3 days, and cell stratification for transplantation. The patient is prepared routinely before surgery, the original diseased tiss...

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Abstract

The invention provides a cornea metaphase preservation solution, and preparing and using methods thereof. The cornea metaphase preservation solution is a cell culture minimum essential medium (MDM) with chondroitin sulfate, low molecular dextran, L-glutamine, dexamethasone, tobramycin, 2-hydroxyethyl and Y-27632 added. The preservation solution not only can keep activity and normal morphology of cornea endothelial cells, but also can enhance viability of corneal limbus epithelial cells and improve clone ability of corneal limbus stem cells. And especially, a medium to long term preservation effect is obvious, phenomena of deformation, conjugation and the like of endothelial cells of a control group cornea do not appear in long term preservation, endothelial morphology of the control group cornea is consistent with endothelial morphology of a cornea preserved for 4 days, and the endothelial cells of the control group cornea are still regular and few in cell conjugation phenomena. The preservation solution can effectively prevent cell apoptosis phenomena during the process of preserving isolated cornea materials, increases activity and clone forming ability of the corneal limbus stem cells, and enables the cornea to maintain a transparent feature in a long preservation time.

Description

technical field [0001] The invention belongs to the technical field of preparation of corneal preservation solution, and specifically relates to a corneal active preservation solution containing a Rho kinase inhibitor and a preparation method thereof, that is, a method for treating fresh cornea by adding a small molecule compound Rho kinase inhibitor Y-27632 Preservation solution. Background technique [0002] In recent years, corneal disease has become the second leading cause of blindness in my country. But as a kind of curable blindness, patients can regain their light by performing lamellar or penetrating keratoplasty. Due to the limitations of my country's national conditions, corneal materials available for transplantation are relatively scarce and unevenly distributed, and a considerable part of corneal materials used in clinical practice needs to be preserved for different periods of time. At present, the evaluation of corneal materials after preservation is mostly...

Claims

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Application Information

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IPC IPC(8): A01N1/02
Inventor 周庆军王瑶段豪云谢立信
Owner SHANDONG EYE INST
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