Loss of Corneal Epithelial Heparan Sulfate Leads to Corneal Degeneration and Impaired Wound Healing

PURPOSE. Heparan sulfate (HS) is a highly modified glycosaminoglycan (GAG) bound to a core protein to form heparan sulfate proteoglycans (HSPGs) that are vital in many cellular processes ranging from development to adult physiology, as well as in disease, through interactions with various protein ligands. This study aimed to elucidate the role of HS in corneal epithelial homeostasis and wound healing. METHODS. An inducible quadruple transgenic mouse model was generated to excise Ext1 and Ndst1, which encode the critical HS chain elongation enzyme and N-deacetylase/N-sulfotransferase, respectively, in keratin 14-positive cells upon doxycycline induction. RESULTS. EXT Delta/Delta CEpi mice (deletion of Ext1 in corneal epithelium) induced at P20 presented progressive thinning of the corneal epithelium with a significant loss in the number of epithelial layers by P55. EXT Delta/Delta CEpi mice presented tight junction disruption, loss of cell-basement membrane adhesion complexes, and impaired wound healing. Interestingly, EXT Delta/Delta CEpi and NDST Delta/Delta CEpi mice presented an increase in cell proliferation, which was assayed by both Ki67 staining and 5-ethynyl-2'-deoxyuridine (EdU) incorporation. Moreover, EXT Delta/Delta CEpi mice presented compromised epithelial stratification 7 days after a debridement wound. The conditional knockout of HS from keratocytes using the keratocan promoter led to no corneal abnormalities or any disruption in wound healing. CONCLUSIONS. Corneal epithelial cells require HS for maintaining corneal homeostasis, and the loss of epithelial HS leads to both impaired wound healing and impaired corneal stratification.