N-Cadherin is Expressed by Putative Stem/Progenitor Cells and Melanocytes in the Human Limbal Epithelial Stem Cell Niche

Abstract

Purpose:: N-cadherin is a member of the classical cadherin family and has previously been demonstrated to be expressed by hematopoietic stem cell niche. The purpose of this study was to investigate N-cadherin expression in human corneal epithelial stem cell niche.

Methods:: Human corneoscleral rims from USA eye bank eyes were used. The expression patterns of N-cadherin in limbal and corneal epithelium were investigated by immunofluorescence staining. Epithelial cells were isolated from the limbus and central cornea, and flow cytometric analysis for N-cadherin was performed. Epithelial cells were separated to N-cadherin⁺ and N-cadherin^- cells by cell sorting, and then subjected to colony-forming assay (CFA) and real-time RT-PCR.

Results:: Immunofluorescence staining revealed that N-cadherin was expressed in limbal basal epithelium, but not in any epithelial layers of the central cornea. Flow cytometric analysis revealed that a small portion of cells (9.9 ± 0.3% N=7) expressed N-cadherin in limbal epithelium. CFA demonstrated that N-cadherin⁺ cells exhibited significantly higher colony-forming efficiency than N-cadherin^- and all sorted cells. Real-time RT-PCR showed that N-cadherin⁺ cells expressed significantly higher stem cell-related markers and lower differentiated markers than N-cadherin^- cells. Additionally, double staining for Melan-A and N-cadherin showed that limbal melanocytes appeared to adhere to epithelial cells in the basal layer, via N-cadherin⁺ regions.

Conclusions:: N-cadherin may be a critical cell-to-cell adhesion molecule between corneal epithelial stem/progenitor cells and their corresponding niche cells in the limbal epithelium.

Keywords: cornea: epithelium