Human corneal limbal stem cell tissue engineering product nourished by fibroblast and preparing process thereof

A technology of limbal stem cells and human fibroblasts, applied in the field of human limbal stem cells-amnion tissue engineering complex and its preparation, can solve the problems of clinical application threats and achieve the effect of promoting cell differentiation

Inactive Publication Date: 2005-07-06
天津医科大学眼科中心
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Problems solved by technology

In addition, 3T3 fibroblasts need to be added to the epithelial cell culture system as a trophoblast, but because 3T3 fibroblasts are mouse-derived immortal cell lines, they contain mouse-derived xenogeneic genes and retroviral genes, which bring great potential for clinical application. potential threat

Method used

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  • Human corneal limbal stem cell tissue engineering product nourished by fibroblast and preparing process thereof
  • Human corneal limbal stem cell tissue engineering product nourished by fibroblast and preparing process thereof
  • Human corneal limbal stem cell tissue engineering product nourished by fibroblast and preparing process thereof

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Embodiment Construction

[0027]A method for culturing human limbal stem cells-amnion tissue engineering complex for treating limbal stem cell-deficient ocular surface diseases is completed according to the following steps: 1. Preparation of amnion: take cesarean section fetal membranes of healthy pregnant women, and place them along the physiological gap Separate the amnion and chorion, spread the amniotic epithelium on the nitrocellulose membrane, put it in a preservation solution consisting of 50% DMEM (Gibco, USA) and 50% glycerin, and store it at -80°C for three more than a month. When in use, take out the frozen amnion from the refrigerator and put it in a 37°C water bath to thaw. Act with 0.1% EDTA for 30 minutes, then act with 0.1% trypsin for 30 seconds, and gently scrape off the amnion epithelial cells with a cell scraper. Lay the amniotic epithelium face up in the culture insert (Millipore, USA) for later use. 2. Preparation of human corneal limbal fibroblast trophoblast: Fetal corneal lim...

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Abstract

A human corneal limbal stem cell-amnion tissue engineering complex using human corneal limbal fibroblasts as the trophoblast is characterized in that: it is close to the human corneal epithelium form in histology form, and is analogous to the human corneal epithelium in immunology phenotype, its ultrastructure shows that the connection between the cells grows mature and the connection between the epithelia and the amnion carrier is a close conglutination. The culture method includes the following steps: (1) preparing the amnion for culturing the human corneal limbal stem cell in vitro; (2) preparing the human corneal limbal fibroblast trophoblast; (3) preparing the human corneal epithelia suspension; (4) culturing the human corneal limbal stem cell-amnion complex. The human corneal limbal stem cell-amnion complex has analogous phenotype to the human corneal epithelium, and provides a safe and effective biology tissue engineering product to treat corneal limbal stem cell defective eye diseases.

Description

[0001] Field: [0002] The present invention relates to a product for treating ocular surface diseases in the field of bioengineering and a preparation method thereof, in particular to a human limbal stem cell-amnion tissue engineering complex capable of repairing limbal stem cell damage and used for ocular surface reconstruction and its preparation method. Background technique: [0003] In the treatment of ocular surface diseases in recent years, the most important breakthrough is the understanding of the function and localization of limbal stem cells. A large number of research results have shown that some cells in the basal layer of the limbus maintain undifferentiated characteristics and are "stem cells" of corneal epithelial cells. These cells have the following characteristics: (1) have proliferation potential, (2) can self-renew, (3) ) can produce a large number of terminally differentiated functional cells, and (4) can repair tissue damage. Ocular surface diseases ch...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N5/074
Inventor 张晓敏孙慧敏李筱荣
Owner 天津医科大学眼科中心
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