Characterization of stem cells on the mouse ocular surface

The adult corneal epithelial stem cells reside at the limbus, the border between the cornea and the conjunctiva on the ocular surface. The properties of these cells as well as how and when they form during post-natal development are not fully understood. One of the characteristics of these cells is their slow-cycling property, which has been used in label-retaining studies to identify and investigate these cells. Neonatal mice are injected with a label (5Bromo-2-deoxy-uridine) early in development. After a chase period, cells that retain their label are considered to be slow-cycling. In this study a whole mount procedure was developed and utilized in order to visualize and characterize the label-retaining cells at the limbus based on their integrin expression profiles, using the slow-cycling property of these cells.; It is shown here that alpha9 integrin is regionally distributed around the circumference of the eye at the limbus; more alpha9-positive cells were present at the nasal region in the adult mouse ocular surface. Also, this pattern gradually develops over time from one to six weeks after birth. Using 3-D reconstruction of the limbal region at high resolution, it is shown that alpha9 integrin is more abundant in the apical-most cytoplasm of the limbal basal cells and it can occasionally colocalize with E-cadherin within the plasma membrane. Analyzing alpha3, alpha6, beta1 and beta4 integrin expression profiles, it is concluded that these integrins are highly expressed at the central cornea and their expressions decrease moving towards the limbus.; BrdU label-retaining cells and their integrin expression profiles on the mouse ocular surface are analyzed in this study. Using simultaneous staining for BrdU and integrins, it is demonstrated that BrdU retention is negatively correlated with alpha9 integrin expression but positively correlated with beta1 and beta4 integrin expression. It is also shown that there are more BrdU-positive cells present at the superior and inferior quadrants than the nasal and temporal quadrants and that 0.94 to 3.6 percent of cells on the mouse ocular surface are label-retaining or slow-cycling.; In conclusion, slow-cycling cells at the limbal region of the mouse ocular surface are enriched in beta1 and beta4 integrins and have little or no alpha9 integrin. These studies are important in providing a better understanding of the development and cell biology of the corneal epithelial stem cells, which may lead to developing diagnostic tools in identifying the causes of corneal epithelial stem cell deficiency diseases.