Method for preparing cornea lamina material

Abstract

The invention discloses a method for preparing a cornea lamina material. The method comprises the following steps of: carrying out accellular processing on a cornea-sclera complex of a mammal eyeball, culturing an amnion epithelial stem cell, inducing and secreting TSP (Thrombospondin)-1 and co-culturing accellular cornea stroma and the amnion epithelial stem cell, sizing, packaging and sterilizing, and the like. According to an obtained cornea lamina transplanting material, the promotion and the balance among the biocompatibility, the stability, the transparency, the mechanical strength and the bioactivity are realized; the integrity of cornea collagen molecules and a lamina structure can be kept to the largest extent, and compared with that prepared with the traditional method, the accellular cornea stroma prepared according to the method disclosed by the invention is larger in tensile strength and has the moisture content and the transparency close to those of normal cornea; and various cell growth factors and the TSP-1 are gathered, and the cornea lamina material has special effects of inflammatory resistance and vascularization resistance, has the capability of greatly enhancing the bioactivity of the cornea lamina material and can be widely used for repairing complicated ocular surface coloboma such as inflammation and corneal ulcer grown in a vascularization way.

Description

technical field [0001] The invention belongs to the technical field of tissue engineering medical biomaterials, and in particular relates to a preparation method of a corneal lamellar material for transplantation. Background technique [0002] The cornea is divided into epithelial cell layer, Bowman's layer, stroma layer, Bowman's layer and endothelial cell layer according to the structure. Trauma, microbial infection, lesions and other factors will cause corneal function loss and vision loss, and severe corneal blindness will be formed. For corneal endothelial damage, penetrating keratoplasty is required; for corneal epithelium and stroma damage with intact endothelial cell layer, corneal lamellar transplantation can be used to repair. Corneal stroma injury is usually accompanied by corneal ulceration, complex inflammatory response, vascularization, leading to continuous degradation of corneal stroma. In the case of complex inflammation, it is difficult to obtain long-ter...

AI Technical Summary

Problems solved by technology

[0006] The current preparation methods of acellular corneal stroma all have certain safety risks and shortcomings of insufficient biological activity.

For example, 1) Chinese patent application (200410018107.5) discloses a cell-free heterogeneous corneal stroma and its preparation method and use. It uses ionic or non-ionic detergent combined with nuclease to remove cell components, and then dehydrates the stroma. Dry; due to the introduction of immunogenic nucleases, it is difficult to ensure complete elution in subsequent operations, and the remaining residues will increase immunogenicity; the bioactivity of the prepared corneal lamellar material is low

2) The decellularized corneal stroma and its preparation method disclosed in the Chinese patent application (200710027698.6) use ionic or non-ionic detergents to remove the cells to a certain extent, but the decellularization is not complete, and the collagen telopeptide The epitope structure still exists, and the resulting decellularized corneal stroma has high immunogenicity and safety risks

Similarly, the method for preparing the decellularized matrix disclosed in the Chinese patent application (200810026972.2) uses a combination of phospholipase and non-ionic detergent for decellularization, which improves the biological activity, but does not fundamentally solve the problem of safety. The problem

3) The bioartificial cornea disclosed in Chinese patent application (200510120794.6) uses freeze-thaw-detergent-trypsin-ultrasound to decellularize the heterogeneous cornea, cross-links with epoxy, and uses acid anhydride or methylating reagent Active groups such as hydroxyl, amino, and sulfhydryl groups are blocked; although this method can improve the mechanical properties of the corneal stroma, the freezing treatment in the decellularization method will damage the molecular structure of collagen, resulting in disordered arrangement of collagen molecules and lamellar structures; The degree of cross-linking and sealing is too high, which reduces the biological activity of the material and the ability to integrate with the implant bed

[0007] The bioactivity of the corneal laminar material prepared by using the acellular corneal stroma can be enhanced by adding one or more cytokine compositions, such as epidermal growth factor (EGF), fibroblast growth factor (FGF), etc.; However, this addition method generally has a high preparation cost, and also lacks targeted therapeutic effects on pathological microenvironments such as corneal ulcers, inflammation, and vascularization.

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