ALPHA-2,3 SIALYLATION DIFFERENTIATE THE LIMBAL AND CORNEAL EPITHELIAL-CELL PHENOTYPES

Purpose. The initial differentiation event for the corneal epithelial cell lineage occurs as the limbally localized stem cells yield, throug h mitosis, the highly proliferative, transiently amplifying corneal pe ripheral cells. This differentiation is characterized by the expressio n of tissue-specific cytokeratins, as well as the loss of alpha-enolas e and pigmentation. All these are intracellular events. The aim of thi s study was to identify and characterize, through lectin analysis, cha nges in cell surface properties associated with differentiation. Metho ds. Cryostat sections of the limbo-corneal area from freshly dissected pigmented rabbit corneas were stained with fluorescent lectins. Resul ts. Peanut lectin (PNA; binds to Ser/Threo-GalNAc-beta-1,3Gal, if the Gal residue is not sialylated) stained the plasma membrane of all laye rs of the conjunctiva and limbus but was excluded from corneal cell me mbranes. Maakia amurensis agglutinin (MAA; binds to sialic acid attach ed to galactose through alpha-2,3 bonds in either N-glycans or O-glyca ns) stained exclusively corneal cell plasma membrane. After complete t issue desialylation, all corneal plasma membranes became PNA positive with equal stain intensity across both sides of the limbo-corneal marg in. The binding of the agglutinins from Limax flavus (binds unselectiv ely to sialic acid) and Sambucus nigra (binds to sialic acid attached through alpha-2,6 bonds) to the basement membrane displayed a large in crease at the corneal side of limbo-corneal demarcation. Conclusions. Limbal (stem) cells express on the cell surface unsialylated galactose residues that are recognized by PNA and that lack any sialic acid bou nd through alpha-2,3 bonds. The initial differentiation involves sialy lation of these residues and the concurrent appearance of alpha-2,3 si alic acid residues, suggesting expression or activation of alpha-2-3 s ialytransferase. Changes in basement membrane composition, charge, or both may underpin this expression.