Human limbal niche cells are a powerful regenerative source for the prevention of limbal stem cell deficiency in a rabbit model

Guigang Li,Xinyu Li,Sean Tighe,Subo Cai,Shen Li,Juan Wang,Shuangling Chen,Yingting Zhu,Huatao Xie,Ming Sun,Yuan Zhang

doi:10.1038/s41598-018-24862-6

Guigang Li, Xinyu Li + Show 9 more

Open Access

https://doi.org/10.1038/s41598-018-24862-6

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Abstract

In this article, human limbal niche cells (LNC) or bone marrow derived mesenchymal stem cells (BMMSC) were used to prevent limbal stem cell deficiency (LSCD) in an alkali burn rabbit model and their results were compared. The epithelial cell defect area, corneal neovascularization, and the print cell cytometry were quantified to grade the severity of LSCD. Three months after the alkali burn, a partial LSCD was observed in the control group (no treatment) indicated by chronic corneal epithelial defects, positive corneal fluorescein staining, neovascularization and goblet cell migration. In contrast, the severity of LSCD in both the LNC and BMMSC transplantation groups was dramatically reduced as shown by smaller epithelial cell defects, decreased fluorescein sodium staining, decreased neovascularization and decreased goblet cell density. Interestingly, the LNC group was shown to more effectively prevent LSCD than the BMMSC group. Further analysis indicated subconjunctivally transplanted LNCs were more powerful than BMMSCs to prevent LSCD, at least partially, due to increased activation of SCF-c-Kit signal. We conclude that LNCs are a more powerful resource than BMMSCs to prevent LSCD in an alkali burn rabbit model, at least partially due to increased activation of SCF signaling.

Highlights

Corneal transplantation is a standard treatment for serious cornea diseases, many patients are not able to recover from blindness due to limbal stem cell deficiency (LSCD)
To characterize limbal niche cells (LNC) and bone marrow derived mesenchymal stem cells (BMMSC), we double immunostained cornea-limbus sections with pan cytokeratin (PCK), vimentin (Vim) to delineate the epithelium and the stroma in the limbal and cornea region and double immunostained C-kit/SCF (Fig. 1A), PCK/P63α (Fig. 1B), PCK/C-kit (Fig. 1C), and C-kit/Vim (Fig. 1D) to show SCF and c-kit were expressed much higher in the limbus compared to other regions of the cornea in which most SCF was expressed in the basal layer (Fig. 1A)
The results indicated surface antigens that are characteristic to mesenchymal stem cells (MSC), including CD73, CD90, CD105, were expressed in both LNCs and MSCs

Summary

Introduction

Corneal transplantation is a standard treatment for serious cornea diseases, many patients are not able to recover from blindness due to limbal stem cell deficiency (LSCD). Niche cells (LNC) are progenitor cells isolated from the corneal limbal niche using collagenase digestion and cultured in modified embryonic stem cell medium (MESCM)[13] on Matrigel coated plastic surface. LNCs have been shown to more effectively prevent limbal epithelial progenitors from aging compared to BMMSCs14–17. It is unclear whether LNCs can prevent LSCD, and if so, whether LNCs are better than BMMSCs. In this study we compare the efficiencies between human LNCs and BMMSCs to prevent LSCD, and elucidate their potential mechanism. Our results suggest for the first time that subconjunctivally transplanted LNC are more powerful than BMMSC to prevent LSCD in an alkali burn rabbit model, at least partially, due to activation of SCF-c-Kit signaling

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Results

Conclusion