Tracing the fate of limbal epithelial progenitor cells and their progeny in the murine cornea

Abstract

Purpose: Stem cell (SC) division, deployment and differentiation are processes which contribute to corneal epithelial renewal. Until now studying the destiny of these cells in a living organism has not been possible. However, the advent of inducible multicolor genetic tracing and powerful imaging technologies has rendered this achievable in the transparent and readily accessible cornea.

Methods: K14CreERT2-Confetti mice which harbor two copies of the Brainbow 2.1 cassette, yielding up to ten possible colors from the stochastic recombination of fluorescent proteins, were used to monitor K-14+ progenitor cell dynamics in the corneal epithelium of live animals.

Results: Multicolored columns of cells emerged from the basal limbal epithelium as they expanded and migrated linearly at a rate of 10 µm/day towards the superficial layers of the central cornea. The permanent expression of fluorescent proteins, passed from progenitor to progeny, assisted in discriminating individual clones as spectrally distinct streaks containing over 1000 cells. Our data are in agreement with the limbus as the SC repository, as opposed to SCs being evenly distributed throughout the central cornea. Furthermore, the centripetal clonal expansion is suggestive that a single progenitor is responsible for the long-term maintenance of a narrow corridor of corneal epithelial cells.

Conclusions: This is the first report describing progenitor cell fate determination in the murine cornea using multicolor genetic tagging. This model represents a powerful resource to monitor SC kinetics and fate choice under homeostatic conditions, and may assist in assessing clonal evolution within the corneal epithelium during development, aging, wound-healing and disease.

Keywords: 721 stem cells • 482 cornea: epithelium