EXTRACELLULAR MATRIX ABNORMALITIES IN CORNEAL EDEMA

描述(改编自申请人的摘要)：假晶状体(PBK)和 无晶状体眼(ABK)大泡性角膜病变是角膜最常见的适应症 移植。研究人员表明，在PBK/ABK角膜中， An的特定亚型的表达和沉积增加 细胞外基质蛋白TN-C(TN-C)，在 正常的角膜。TN-C可影响细胞的黏附、迁移和增殖 对伤口愈合和组织重塑很重要。他们还拥有 显示PBK/ABK角膜有相应的表达增加 TN-C整合素受体和某些生长因子/细胞因子，它们可以 诱导TN-C表达。提出了以下假设：(1) PBK/ABK角膜呈现一个持续的“损伤-反应循环”，其中整个 角膜处于持续重塑状态，(2)生长因子和/或 细胞因子在这个周期中起着重要的作用；(3)细胞因子的表达和 TN-C的沉积影响粘附性、迁移性、增殖性和 角膜细胞的功能特性。了解这些方面可以 导致未来的治疗干预。 而TN-C、生长因子和细胞因子被认为在 在PBK/ABK发病机制中的作用，其他尚未确定的异常有 可能也牵涉其中。因此，强大的新型微敏技术 已经被应用于对个体基因的差异筛选 眼角膜。假设有了这些技术，脑部的异常 其他对PBK/ABK发病重要的基因的表达将是 已确认身份。具体目标有三个：(1)确定功能 PBK/ABK角膜细胞在不同载体上生长对TN-C亚型的影响 TN-C的亚型，并测定细胞的黏附，迁移， 增殖和功能；(2)测定生长因子的影响 和细胞因子对TN-C和TN-C结合整合素表达的影响： (A)鉴定PBK/ABK角膜中的异常生长因子/细胞因子， (B)确定这些因素和地塞米松对 TN-C及其结合整合素在体外培养的角膜细胞中的表达 (C)确定TN-C的各种异构体是否会影响 TN-C结合整合素的体外实验；(3)检测差异基因 正常和PBK/ABK角膜中的表达及鉴定独特的基因表达 模式：(A)筛选正常与PBK/ABK角膜和 使用差异显示、核酸阵列和差减的细胞层 文库；(B)确定差异表达的基因是否具有特异性 用于PBK/ABK；以及(C)确定PBK/ABK中改变的基因表达 反映在蛋白质水平上。

DESCRIPTION (Adapted from applicant's abstract): Pseudophakic (PBK) and aphakic (ABK) bullous keratopathy is the most common indication for corneal transplantation. The investigators have showed that in PBK/ABK corneas, there is an increased expression and deposition of specific isoforms of an extracellular matrix protein, tenascin-C (TN-C), that is not expressed in normal corneas. TN-C can affect cell adhesion, migration and proliferation that are important in wound healing and tissue remodeling. They have also shown that PBK/ABK corneas have a corresponding increase in the expression of TN-C integrin receptors and certain growth factors/cytokines, which could induce TN-C expression. The following hypotheses are proposed: (1) that PBK/ABK corneas present an ongoing "injury-response cycle" where the entire cornea stays in a continuous state of remodeling, (2) growth factors and/or cytokines play an important role in this cycle, and (3) the expression and deposition of TN-C affects the adhesive, migratory, proliferative and functional properties of corneal cells. Understanding these facets could lead to future therapeutic interventions. While TN-C, growth factors and cytokines are thought to play a significant role in PBK/ABK pathogenesis, other as yet unidentified abnormalities are probably also involved. Therefore, powerful new micro-sensitive techniques have been adapted for differential screening of genes from individual corneas. The hypothesis is that with these techniques, abnormalities in the expression of other genes important for PBK/ABK pathogenesis will be identified. There are three specific aims: (1) to determine the function of TN-C isoforms in PBK/ABK corneas by growing corneal cells on various isoforms of TN-C and determine rates of cell adhesion, migration, proliferation and function; (2) to determine the influence of growth factors and cytokines upon the expression of TN-C and TN-C binding integrins by: (a) identifying the abnormal growth factors/cytokines in PBK/ABK corneas, (b) determining the effects that these factors and dexamethasone have on the expression of TN-C and its binding integrins in corneal cells in vitro, and (c) determining if various isoforms of TN-C can affect the expression of TN-C binding integrins in vitro; and (3) to examine the differential gene expression in normal and PBK/ABK corneas and identify unique gene expression patterns by: (a) screening cellular mRNAs in normal vs. PBK/ABK corneas and cell layers using differential display, nucleic acid array and subtraction libraries; (b) determining if differentially-expressed genes are specific for PBK/ABK; and (c) determining whether altered gene expression in PBK/ABK is reflected at the protein level.