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CFSE: A New Method for Identifying Human Limbal Stem Cells and Following Their Migration in Human Cornea
Chiara Bonzano1, Barbara Canciani2, Sara Olivari3
1Eye Clinic, DiNOGMI, University of Genoa and IRCCS San Martino Polyclinic Hospital, Genoa, Italy oculistabonzano@gmail.com.
Aim:
To develop a method capable of identifying human corneal limbal stem cells (LSCs) and follow their proliferation and migration in the epithelium.
Materials And Methods:
Ten fresh matched pairs of cadaveric normal human corneas were obtained from donors. Carboxyfluorescein diacetate succinimidyl ester (CFSE) was used to target LSCs. The distribution of CFSE-positive cell clusters was analyzed by fluorescence microscopy by counterstaining with 4',6-diamidino-2-phenylindole (DAPI). Fluorescence was digitally recorded for seven days, and the rate of cell movement was determined.
Results:
CFSE-labeled cells were tracked in corneas. Analysis of time sequences revealed that they moved centripetally. Daily average CFSE-labeled LSC movement was 0.073±0.01 cm (±SD).
Conclusion:
CFSE allowed us to identify LSCs and to track their centripetal migration from the limbal basal layer to the anterior ocular surface. This experimental system appears to be a valuable tool for further studies on corneal epithelial cell migration and proliferation.
