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Title: Morphology and proteoglycan content of the sulfotransferase- and glycosyltransferase-deficient mouse cornea
Author: Littlechild, Stacy
ISNI:       0000 0004 5915 4135
Awarding Body: Cardiff University
Current Institution: Cardiff University
Date of Award: 2016
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Keratan sulfate (KS) is an elongated glycosaminoglycan (GAG) chain found throughout the cornea, the clear tissue at the front of the eye. It is thought that KS plays a specialized role in maintaining the ordered spatial arrangement of collagen fibrils that comprise the thickest layer of the cornea, the stroma. Repeating N-­‐acetylglucosamine (GlcNAc) and galactose monosaccharides make up the fine structure of KS, and sulfate groups frequently modify their C-­‐6 positions. Recent studies have linked improper sulfation of GlcNAc residues to macular corneal dystrophy, a vision condition characterized by a progressive loss of corneal transparency. Since then, in vitro experiments have implicated four enzymes in highly sulfated KS biosynthesis. It is currently believed that β-­‐1,3-­‐N-­‐ acetylglucosaminyltransferase 7 (β3GnT7), corneal GlcNAc 6-­‐O sulfotransferase (CGn6ST), and β-­‐1,4-­‐ lactosyltransferase 4 (β4GalT4) sequentially catalyze the addition of GlcNAc, transfer sulfate to its C-­‐6 position, and link galactose to the growing KS backbone, respectively. The fourth enzyme, KS galactose 6-­‐O sulfotransferase (KSGal6ST) is thought to act last, sulfating galactose residues. Mutant mice deficient in CGn6ST, KSGal6ST, or β3GnT7 were recently developed to investigate the consequences of dysfunctional sulfo-­‐ or glycosyltransferases on corneal morphology. Electron microscopy and immunohistochemistry data in this thesis showed that the systemic absence of CGn6ST or β3GnT7 resulted in a corneal stroma devoid of KS, but mutation of only KSGal6ST led to a KS phenotype similar to that of wild type controls. Western blot analysis conducted on β3GnT7-­‐null corneas indicated that KS assumed an unusually truncated form, as compared to wild type controls. A secondary result evident in electron micrographs was that in cases where KS levels dropped below the detectable threshold (i.e. in CGn6ST and β3GnT7 mutant corneas), a concurrent appearance of atypically elongated GAGs was visible, suggesting a compensatory mechanism to preserve corneal organization. Since the unusual GAGs were susceptible to chondroitinase ABC digestion, it is thought they are comprised of chondroitin/dermatan sulfate (CS/DS). Studies using high performance liquid chromatography were also undertaken to establish protocols in which future work could quantify the change in CS/DS expression observed in CGn6ST and β3GnT7 mutant corneas.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available
Keywords: RE Ophthalmology