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Thuy T. Truong, Kyle Huynh, Martin N. Nakatsu, Sophie X. Deng; Use Of SSEA-4 To Enrich The Isolation Of Human Corneal Stem/Progenitor Cells. Invest. Ophthalmol. Vis. Sci. 2011;52(14):5126.
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To assess whether Stage-Specific Embryonic Antigen-4 (SSEA-4) can serve as a negative marker to enrich the isolation of human corneal stem cells.
The SSEA-4 expression in the human cornea and limbus was examined by RT-PCR and immunohistochemistry. The SSEA-4+ and SSEA-4- cells were then separated using DynaBeads®SSEA-4 kit. The phenotype of these two cell populations were evaluated using cell size, clonogenic assay, and expression of putative limbal stem cell and proliferation marker.
SSEA-4 was expressed in all layers of the corneal and anterior limbal epithelia. There were discrete clusters of SSEA-4+ cells in the central and posterior limbal epithelia. SSEA-4+ cells accounted for an average of 40% of the total limbal epithelial cells. SSEA-4- population contained 5-time more small cells (≤11 µm in diameter) than SSEA-4+ population. The expression levels of putative LSC markers, ABCG2, ΔNp63α, and K14 were significantly higher in the SSEA-4- population than those in the SSEA-4+ population. In contrast, the expression levels of K12 and proliferation marker Ki-67 were substantially higher in the SSEA-4+ population. Interestingly, expression of Ki-67 in the SSEA-4- population increased significantly upon culturing in vitro and was 39±11% higher than that in the SSEA-4+ cells. The CFE was 15±1.9% (p<0.001) higher in the SSEA-4- population compared to the unsorted and 60.±10% (p<0.05) higher than the SSEA-4+ population.
This is the first study to characterize SSEA-4 expression in the human ocular surface epithelium. SSEA-4 could be used as a negative marker to enrich the limbal epithelial stem/progenitor population.
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