Localisation of notch and delta in the human cornea [Abstract]

Purpose: To investigate if Notch and its ligand Delta, which play important roles in cell fate decisions in various epithelia, are present in the human cornea. Methods: Cryosections of human corneas, retrieved within 24 hours cadaver time, were immunofluorescently labelled using antibodies directed against Notch 1 and Delta (AutogenBioclear) followed by incubation in appropriate Alexor Fluor (Molecular Probes) secondary antibodies. SDS-PAGE and Western blotting of isolated corneal epithelial cells, keratocytes and endothelial cells were performed to confirm the localisation pattern. Results: Immunofluorescent localisation of Notch was demonstrated throughout the corneal epithelium in all suprabasal cells i.e. in the borders of the wing and superficial epithelial cells, in central, peripheral and limbal regions. Labelling was not detected in the epithelial basal cell layer in any region. In contrast, delta appeared to be expressed in cell borders throughout all regions, and all cell layers of the corneal epithelium, with an increased intensity underlying peripheral basal cells. Immunolabelling for notch was absent in both corneal keratocytes and the endothelium, however localisation of delta was observed in a small number of keratocytes within the corneal stroma. Immunoblotting confirmed the aforementioned cellular localisation: the 80kDa Delta protein was present in the Triton-soluble corneal epithelial cell fraction. Conclusions: The presence of Notch and Delta in the corneal epithelium suggests that it plays a pivotal role in corneal homeostasis by regulating cell fate and differentiation