Culture and expansion of stem cells located at the corneoscleral limbus

Document Type : Research Paper

Authors

1 Bo Ali Research Institute, Mashhad University of Medical Sciences, Mashhad, Iran

2 Department of Clinical Biochemistry, Faculty of Medicine, Mashhad University of Medical Sciences, Mashhad, Iran

Abstract

Introduction:
Corneal epithelial cells are destroyed naturally and the limbal stem cells replace them by multiplying and differentiating. In case of defects in these cells, it is impossible to replace them, and in many cases, the patients lose their vision. One of the newest methods of treating limbal stem cell deficiency (LSCD) is to cultivate them in the laboratory and transfer them to the patient's eye. For this purpose, in this study, we aimed to culture limbal stem cells in vitro and optimized the culture quality.
Methods:
First, the limbus region of the donated eye samples was separated. Then the separated parts were cultured on the amniotic membrane by the explant method. The presence and proliferation of limbal stem cells were investigated by immunofluorescent method and antibody against ΔNP63α and ABCG2 markers on the 10th, 14th, and 21st days. Also, by using the flow cytometry method and antibody against ΔNP63α marker, the percentage of limbal cells on the best day of their culture based on immunofluorescent results was determined.
Results:
Findings showed that limbal stem cells could be cultured properly in vitro. All immunofluorescent assays confirmed limbal stem cell attendance, however, the result of day 14th of culture was more considerable. Therefore, the flow cytometry assay was performed on the 14th day and proved that there were 5.8% limbal stem cells using explant culture.
Conclusion:
The results indicated the successful cultivation of limbal cells in laboratory conditions, but for their clinical application, further optimization of cultivation is required.

Keywords

References

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medical journal of mashhad university of medical sciences
Volume 66, Issue 1
May and June 2023
Pages 1-10

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