Immunohistochemical studies on the role of glycosaminoglycans in thepathogenesis of corneal disorders

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To clarify the surface topography of D-periodic collagen fibrils, we applied atomic force microscopy (AFM) to the collagen fibrils of mouse corneal stroma and sclera. AFM showed that the depth of the groove in the D-periodicity of corneal fibrils was shallow compared with that of the sclera. Ruthenium red (RR) method showed that proteoglycans (PGs) or glycosaminoglycans (GAGs) distributed on the surface of collagen fibrils in the cornea and sclera. The difference in ultrastructure between cornea and sclera could explain the differences in the number or the distribution of molecule in both tissues.In the other experiment, we compared the surface structure of human corneal collagen before and after the digestion with chondroitinase ABC or kertanase by using corneal samples obtained from autopsy. The surface structure did not changed after the chondroitinase ABC digestion where as the groove of the D-periodicity changed to be much deeper after keratanase.This shows that keratan sulphate associate PGs should depress the groove of the surface of human corneal collagen fibrils, and these PGs may act for the regular distribution of collagen fibrils, and thus, may contribute for maintaining the transparency of the cornea. We further investigated the distribution of PGs or GAGs during corneal wound healing by the immunohistochemical study by using the human corneal graft obtained during regrafting, and showed that chondroitin sulphate A had an important role during wound healing.