O-Glycans on Mucins at the Ocular Surface

项目摘要/摘要 粘膜表面，包括眼表的粘膜表面，被上皮所覆盖，而上皮对 防止损坏和感染。细胞表面相关粘蛋白是细胞表面 该上皮的顶端糖萼，在质膜上方延伸至500 nm，远高于质膜 所有其他与膜相关的蛋白质和脂质。它们展示了大量的O-糖链 与蛋白质骨架中氨基酸的串联重复相关联，从而产生局部高 碳水化合物配体的浓度。细胞表面糖基化的改变与 与眼表上皮病(如干眼)的粘膜屏障的破坏有关，但 导致这种混乱的机制大多仍未确定。在上一次资助期间 期间，我们确定Galectin-3是人眼表达的主要碳水化合物结合蛋白。 表面上皮细胞。Galectin-3是一种多聚体凝集素，可以使蛋白质交联和寡聚 显示多个低聚糖链。这一发现使我们假设多价O- 葡聚糖结合Galectin-3低聚物形成坚固且高度有序的凝集素-碳水化合物晶格 在角膜和结膜细胞的糖萼中的起伏的膜脊上方。在这 应用，我们建议确定糖萼中的粘蛋白型O-糖链是否与 Galectin-3在眼表形成顶端细胞表面屏障，而这一屏障是否 防止外来物质通过质膜流入和进入 微生物和炎症介质对根尖细胞表面受体的影响。最后，我们建议 确定在患有眼表上皮病的患者中是否存在分子水平的破坏 O-葡聚糖：半乳糖屏障。了解上皮防御机制，包括 细胞表面糖基化的成分可以帮助开发新的治疗方法 旨在修复眼表患者上皮屏障成分的方法 疾病。

Project Summary/Abstract Mucosal surfaces, including those of the ocular surface, are covered by epithelium that is critical to preventing damage and infection. Cell surface-associated mucins are major components on the apical glycocalyx of this epithelium, extending up to 500 nm above the plasma membrane, far above all other membrane-associated proteins and lipids. They exhibit large numbers of O-glycan chains tethered to tandem repeats of amino acids in their protein backbone that create a locally high concentration of carbohydrate ligands. Alteration of cell surface glycosylation has been associated with the disruption of the mucosal barrier in ocular surface epithelial disease (e.g., dry eye), but the mechanisms that lead to this disruption remain mostly uncharacterized. During the last funding period, we identified galectin-3 as the major carbohydrate-binding protein expressed by human ocular surface epithelia. Galectin-3 is a multimeric lectin that can cross-link and oligomerize proteins displaying multiple oligosaccharide chains. This finding led us to hypothesize that multivalent O- glycans bind galectin-3 oligomers to form strong and highly organized lectin-carbohydrate lattices above the undulating membrane ridges in corneal and conjunctival cells' glycocalyx. In this application, we propose to determine whether mucin-type O-glycans in the glycocalyx associate with galectin-3 to form an apical cell surface barrier on the ocular surface, and whether this barrier prevents the influx of foreign substances through the plasma membrane and the access of microorganisms and inflammatory mediators to apical cell surface receptors. Finally, we propose to determine whether in patients with ocular surface epithelial disease there is a molecular breach of the O-glycan:galectin barrier. Understanding the mechanism of epithelial defense that includes components of the cell surface glycocalyx could help with the development of new therapeutic approaches aimed to restore components of the epithelial barrier in patients with ocular surface disease.