Preparation method and application of autologous mesenchymal stem cell-loaded human amniotic membrane cornea paster

Abstract

The invention discloses a preparation method of an autologous mesenchymal stem cell-loaded human amniotic membrane cornea paster, wherein autologous sourced mesenchymal stem cells are planted onto an amniotic membrane to be taken as bearing carriers to treat the corneal injury. The method comprises the following steps of: inoculating monocyte separated from bone marrow into an alpha-MEM culture medium, amplifying to a third generation to inoculate 2.5*10<5> cells onto the 1.2cm*1.2cm amniotic membrane, and tightly arranging the cells after 48 hours, wherein the cover area of the mesenchymal stem cell under an inverted microscope is greater than 80%, and the immunofluorescent staining shows that the positive rate of cells CD44 and CD90 is greater than 90%. The human amniotic membrane-loaded mesenchymal stem cell paster is free from dissolving and falling after being transplanted within one week, and new vessels can not be generated within eight months. The method is free from immunological rejection, an enough cell source can be provided, and the method can be used for the clinic treatment of the corneal injury.

Description

technical field [0001] The present invention relates to the field of regenerative medicine, in particular to a preparation method for planting autologous bone marrow mesenchymal stem cells on human amniotic membrane to form a patch for treating corneal damage. Background technique [0002] The treatment of corneal injury is a complicated clinical problem. The cornea is divided into five layers, namely the corneal epithelium, the anterior limiting layer, the corneal stroma, the posterior limiting layer, and the corneal endothelium. Clinical corneal diseases include physical and chemical burns, mechanical trauma, infection, recurrent pterygium, Steven-Johnson syndrome, pekoidoid, trachoma, etc. Severe conjunctival and corneal diseases are still a major part of ophthalmologists’ eyesight treatment. problem. After corneal burns, a large number of limbal stem cells are lost, the corneal epithelium cannot be repaired normally, the cornea is cloudy, new blood vessels increase and...

AI Technical Summary

Problems solved by technology

[0004] 1. The short-term corneal transparency after traditional keratoplasty, its main disadvantage is that only the central φ7mm optically transparent area is transplanted. Due to the obvious rejection, long-term use of immunosuppressants is required to cooperate with the treatment

[0005] 2. Although there is no immune rejection in autologous corneal limbus transplantation, the source of materials is limited. The main disadvantage is that it increases the risk of disease in healthy eyes. It cannot be transplanted in a 360° circle, and the effect is limited.

[0006] 3. Donor materials for allogeneic limbal transplantation are extremely limited, treatment opportunities are very few, and immune rejection is obvious, long-term medication is required, and the cost is high

Stem cells are the source of cell proliferation and differentiation in self-renewing tissues. Clinically, stem cells solve the problem of organ source caused by organ transplantation. Recently, corneal limbal stem cell transplantation has become a hot spot in the treatment of corneal diseases. The main method is to take autologous or allogeneic Limbal stem cells are cultured from the limbus of the cornea and planted on the amniotic membrane for transplantation to treat corneal diseases. However, due to the limited source of the limbus, the rejection of allogeneic limbal stem cells is obvious after transplantation, so although it has a significant therapeutic effect, it has not been popularized in clinic. application

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