An integrated experimental and theoretical approach to understanding corneal epithelial maintenance

This multidisciplinary systems project studies maintenance and regeneration of the ocular surface. We have developed an in silico model of corneal epithelial maintenance incorporating cell proliferation, loss and centripetal migration, which stably recapitulates observed patterns of clonal cellular arrangement in adult life. In this project we will refine the model by fitting to experimental data, obtain new empirical data on epithelial cell loss, and hence make predictions about the response of the corneal to damage or ageing.

The in silico model will be validated in vivo using mosaic reporter transgenic mice (LacZ and GFP mosaics) and mutant mice to track the long-term patterns of cell migration and orientation in normal, wounded and ageing situations. Corneas of mouse genetic mosaics have radial striped patterns (meeting at a central spiral), consistent with centrifugal movement of cells from the peripheral limbus (putative stem cell niche). Predictions about how ocular surface wounding or ageing causes disruption to epithelial migration patterns will be tested using these reporters. Genetic disruption will be effected by crossing the mosaic reporter mice onto Pax6 and Gli3 mutant backgrounds. The location of stem cells in the ocular surface will be determined using lineage tracing with tamoxifen-inducible CAGG-CreER;loxP-reporter mice (R26R-LacZ, R26R-YFP). By applying low doses of tamoxifen we will label individual cells and, through longitudinal studies, determine whether active stem cells, producing long-term clones during normal homeostasis (without wounding), are ever found in the central cornea rather than limbus.

The cornea is an excellent model of how stem cell activity, cell movement and loss are balanced in vivo. This project is important as there are major gaps in knowledge of the basic science of corneal maintenance that are essential to resolve, both as a model for epithelial homeostasis and in relevance to corneal degenerative problems.